Skip to main content

Full text of "A text-book of bacteriology, including the etiology and prevention of infective diseases and a short account of yeasts, and moulds, haematazoa, and psorosperms"

QR 



CORNELL UNIVERSITY 



THE 

IFlnmet Urt^rmatg ffiibtatji 

FOUNDED BY 

ROSWELL P. FLOWER 

for the use of the 

N. Y. State Veterinary College 

1897 



Cornell University Library 
QR 46.C94 1897 

A text-book of bacteriology, including th 



3 1924 000 246 714 



DATE DUE 



















































































































































GAYLORD 






PRINTED INU.S.A. 




Cornell University 
Library 



The original of tiiis book is in 
tine Cornell University Library. 

There are no known copyright restrictions in 
the United States on the use of the text. 



http://www.archive.org/details/cu31924000246714 



A TEXT-BOOK 

LIBRARY. 



BACTEEIOLOG 



INCLUDING THE 



ETIOLOGY AJSD PREVENTION 



II^FEOTIVE DISEASES 



AND A SHOET ACCOUNT OF 

YEASTS AND MOULDS, H^MATOZOA, AND 
PSOROSPERMS 



EDGAR M. CROOKSHANK, M.B. 

I'KOFESriOK OF COMPAEATnE PATHOLOGY AND BACTERIOLOGY, AND FELLOW OF KING'S 
COLLEGE, LONDON 



FOURTH EDITION 
RECOKSTRUCTED, REVISED AND GREATLY ENLARGED 



PHILADELPHIA 

W. B. SAUNDERS 

925 Walnut Sthbbt 
1897 



\\ 












Ho Ui"^') 



%0 
SIR JOSEPH LISTER, BART., M.B., P.R.S., 

WHO HAS CEEATED A NEW EPOCH IN 

MEDICINE AND SUEGERY, 

BY APPLYING A KNOWLEDGE OP MICRO-OESANISMS 

TO THE TREATMENT OP DISEASE, 

Ihte WioxU is, biiih yamtssion, Jciitattb 
BY THE AUTHOR 
AS A 
TOKEN OP ADMIRATION AND RESPECT 



PREFACE 

TO THE 

FOURTH EDITION. 



This book, though nominally a fourth edition, is practically 
speaking a new work. The progress of Bacteriology has been 
very rapid, and many new inyestigations have been made in 
connection with the etiology, prevention and treatment of 
communicable diseases. It has been necessary to reconstruct, 
enlarge and thoroughly revise the text of the third edition, 
and I have added twenty-six chapters. 

The most important researches conducted in bacteriological 
laboratories are those relating to the contagia. In many 
diseases of man and animals it has not been possible to 
identify the contagium with a bacterium, or indeed with any 
micro-organism ; but when the virus is chemically examined, 
or investigated with a view to protective inoculation, or utilised 
for experiments in serum-therapeutics, such researches are 
within the province of the bacteriologist. 

The recognition of the fact that in so many diseases the 
nature of the contagium has not yet been determined will 
have the effect of encouraging continued activity in this 
important field of scientific investigation. 

I hope that this work will continue, to be of use as a text- 
book for the bacteriological laboratory, and that the chapters 
on the etiology and prevention of the communicable diseases 



VIH PREFACE TO THE FOURTH EDITION. 

of man and animals will be not only of scientific interest, 
but of practical value to Medical Officers of Health and 
Veterinary Inspectors. 

I have divided the book into three parts. Part I., is 
mainly technical, and includes the most recent methods 
employed in studying bacteria and investigating the etiology 
of disease. Part II. deals with infective diseases and the 
bacteria associated with them. Any clinical or pathological 
evidence which may help to throw light on the nature and 
origin of the contagia is taken into account. The most 
.eifectual measures for stamping out these diseases are 
referred to, as they are intimately connected with a 
knowledge of the life-history of micro-organisms. Part III. 
contains descriptions of about five hundred bacteria. Many 
are of no practical importance and of very little scientific 
interest, but a text-book for the laboratory cannot be con- 
sidered complete unless an account is given of all bacteria 
which have been more or less completely investigated. I 
have endeavoured to refer to the original descriptions and 
to verify them by comparison with actual cultivations, but 
in a very great number of instances this has been quite 
impossible,, and I desire to acknowledge the assistance I have 
received from the works of several authors, especially those 
of Fliigge, Frankel, Eisenberg, Baumgarten, Frankland, 
Sternberg, Lehmann and Neumann. 

I have rearranged the bibliography according to the 
chapters, and the names of authors are given in alphabetical 
order. With the aid of the current numbers of the Annates 
ffe rinstitut Pasteur, the Zeitschrift fiir Hygiene, the 
Centralblatt fur Bakteriologie und Parasitenkunde, and the 
Journal of Comparative Pathology and Bacteriology, it is 
possible to become acquainted with the most recent litera- 
ture of the subject. 

Many of the coloured plates illustrating the last edition 
have not been reproduced. Those Bubstituted for them have 
been drawn from my own preparations, and most of them 



PREFACE TO THE FOURTH EDITION. IX 

have already appeared in my Reports to the Board of 
Agriculture and iu other publications. 

One hundred and thirty-three woodcuts and photographs 
have been added in the text, and I have reverted to the plan 
which T adopted in the second edition, of having many of 
them printed in. colours. 

I take this opportunity of thanking Professor Frankel for 
kindly permitting me to reproduce some of the photographs 
in his excellent Atlas. 

I am particularly indebted to Professor Hamilton for 
the use of cliches of figures in his classical treatise on 
Pathology, and to the New Sydenham Society for several 
from the' English translation of Professor Flugge's well- 
known work on micro-organisms. 

To my Demonstrator, Dr. George Newman, D.P.H., I am 
indebted for much assistance in correcting the proof-sheets, 
and for the preparation of an index. 

EDGAR M. CROOKSHANK. 

Saint Hill, East Geinstbad, Sussex, 
August 1st, 1896. 

P.S, — Since this work was finally passed for press the con- 
clusions of the Royal Vaccination Commissioners have been 
published. I have at the last moment added extracts in the 
form of a supplementary Appendix. E. M. 0. 

September IStJi, 1896. 



CONTENTS 



PAET I. 

THEORETICAL AND TECHNICAL. 

PAGE 

CHAPTER I. 
HISTORICAL INTRODUCTION ... . . 1 

CHAPTER II. 

MORPHOLOGY AND PHYSIOLOGY OF BACTERIA . . .11 

CHAPTER III. 

EFFECT OF ANTISEPTICS AND DISINFECTANTS ON BACTERIA . 30 

CHAPTER IV. 
CHEMICAL PRODUCTS OF BACTERIA . ... 39 

CHAPTER V. 
IMMUNITY 49 

CHAPTER VI. 
ANTITOXINS AND SERUM THERAPY . . .56 

CHAPTER VII. 

THE BACTERIOLOGICAL MICROSCOPE . . .65 

CHAPTER VIII. 
MICROSCOPICAL EXAMINATION OF BACTERIA . . 83 

CHAPTER IX. 

PREPARATION OF NUTRIENT MEDIA AND METHODS OF CULTIVA- 
TION ... ... . 99 



Xll CONTENTS. 

PAGE 

CHAPTER X. 

EXPERIMENTS UPON THE LIVIiSG ANIMAL . . .134 

CHAPTER XI. 

EXAMINATION OF AIR, SOIL, AND WATER 140 

CHAPTER XII. 

PHOTOGRAPHY OF BACTERIA . ■ . . .150 



PAET II. 

ETIOLOGY AND PREVENTION OF INFECTIVE 
DISEASES. 

CHAPTER XIII. 

SUPPURATION, PYAEMIA, SEPTICAEMIA, ERYSIPELAS 173 

CHAPTER XIV. 
ANTHRAX . . . . .191 

CHAPTER XV. 

QUARTER-EVIL. — MALIGNANT (EDEMA.— RAG-PICKERS' SEPTI- 

GMUIA. — SEPTICEMIA OF GCINEA-PIGS. SEPTICEMIA OF 

MICE . . . . . 217 

CHAPTER XVI. 

SEPTICEMIA OF BUFFALOES.— -SEPTIC PLEURO-PNEUMONIA OF 
CALVES. — SWINE FEVER. SEPTICEMIA OF DEER. — SEPTI- 
CEMIA OF RABBITS. FOWL CHOLERA. FOWL ENTERITIS. 

DUCK CHOLERA. — GROUSE DISEASE .... 226 

CHAPTER XVII. 

PNEUMONIA. — INFECTIOUS PLEURO-PNEUMONIA OF CATTLE. — 

INFLUENZA 233 

CHAPTER XVIII. 

ORIENTAL PLAGUE. — RELAPSING FEVER. TYPHUS FEVER. — 

YELLOW FEVER . . . .250 



CONTENTS. Xlll 

PAGE 

CHAPTER XIX. 
SCARLET FEVER.. — MEASLES . . . . .261 

CHAPTER XX. 

SMALL-POX. — CATTLE PLAGUE . 284 

CHAPTER XXI. 

SHEEP-POX. — FOOT-AND-MOUTH DISEASE . 297 

CHAPTER XXII. 

HORSE-POX. — COW-POX . . . . 303 

CHAPTER XXIII. 
DIPHTHERIA . . . . 330 

CHAPTER XXIV. 
TYPHOID FEVER . . . 340 

CHAPTER XXV. 
SWINE FEVER . . . . . . 347 

CHAPTER XXVI. 

SWINE MEASLES. DISTEMPER IN DOGS. — EPIDEMIC DISEASE OF 

FERRETS. — EPIDEMIC DISEASE OF MICE . . . 355 

CHAPTER XXVII. 

ASIATIC CHOLERA. — ^^CHOLERA NOSTRAS. — CHOLERAIC DIARRHCEA 
FROM MEAT POISONING. — DYSENTERY. — CHOLERAIC DIAR- 
RHCEA IN FOWLS .... . . 360 

CHAPTER XXVIII. 
TUBERCULOSIS . . . . . . 375 

CHAPTER XXIX. 

LEPROSY. — SYPHILIS. — RHINOSCLEROMA. — TRACHOMA . 406 

CHAPTER XXX. 

ACTINOMYCOSIS. — MADURA DISEASE . 413 

CHAPTER XXXI. 
GLANDERS . . . . . 451 



xiv CONTENTS. 

I'AGE 

CHAPTEK XXXII. 

TETANUS. — RABIES. — LOUPING-ILL . • • • 457 

CHAPTER XXXIII. 
FOOT-EOT 4^'* 

CHAPTEB XXXIV. 

FOUL-BROOD.— INFECTIOUS DISEASE OF BEES IN ITALY. 

Pl^BRINE.—FLACHERIE.— INFECTIOUS DISEASE' OF CATER- 
PILLARS 469 



PAET III. 

SYSTEMATIC AND DESCRIPTIVE. 

CHAPTER XXXV. 

CLASSIFICATION AND DESCRIPTION OF SPECIES . . .475 



APPENDICES. 

APPENDIX I. 

YEASTS AND MOULDS 577 

APPENDIX II. 

H^MATOZOA IN MAN, BIRDS, AND TURTLES. — H^MATOZOA IN 

EQUINES, CAMELSi AND FISH. HiEMATOZOA IN FROGS . 589 

APPENDIX III. 

PSOROSPERMS OR COCCIDIA. — AMCEBA COLI . . . 609 

APPENDIX IV. 
APPARATUS, MATERIAL AND REAGENTS EMPLOYED IN A BAC- 
TERIOLOGICAL LABORATORY . . . . .612 

APPENDIX V. 
BIBLIOGRAPHY . . . . 639 

SUPPLEMENTARY APPENDIX. 
EXTRACTS FROM THE FINAL REPORT OF THE ROYAL VACCINA- 
TION COMMISSION .667 



LIST OP ILLUSTRATIONS. 



WOOD ENGKAVINGS AND PHOTOGRAPHS. 

FIG. PAGE 

1. Asoocoocns Billrothii, x 65 (Gohn) . . 14 

2. Spiroohseta from Sewage Water, x 1200 (E.M.O.) 15 
B. Flagella (Koch, Brefeld, Warming, Zopf) 16 
i. Bacillus Megatherium (De Bary) . . 17 

5. Clostridium Butyricum, x 1020 (Prazmowski) . . .IS 

6. Leuconostoc- Meseuteroides ; Cocci-chains with Arthrospores (Van 

Tieghem and Cienkowski) ..... 19 

7. Spore-bearing Threads of Bacillus Anthracis, double-stained with 

Fuchsine and Methylene Blue, x 1200 (E.M.C.) 20 

8. Bacilli of Tubercle in Sputum, x 2500 (E.M.C.) . 21 

9. Comma Bacilli in Sewage Water, stained with Gentian Violet, x 1200 

(E.M.C.) . 22 

10. Vibrios in Water contaminated with Sewage, x 1200 (E.M.C.) 22 

11. Refraction of Light (Carpenter) . . 66 

12. Spherical Aberration (Carpenter) . , 67 

13. Combination of Lenses in Abba's Homogeneous Immersion (Car- 

penter) . . . ' 67 

14. Chromatic Aberration (Carpenter) 68 

15. Objective with Collar Correction (Zeiss) . 69 

16. Microscope— English Model (Swift) 71 

17. Removable Mechanical Stage (Swift) 72 

18. Microscope — Continental Model (Zeiss) . 7.S 

19. Iris Diaphragm (Zeiss) , 74 

20. Abba's Condenser (Zeiss) . 75 

21 . Microscope Lamp (Baker) . 6 

22. Large Microscope Lamp (Swift) . , .77 

23. Arrangement of Powell and Lealand's Microscope in working directly 

on the Edge of the Flame, with Stand for Micrometer Eye-piece to 
secure Steadiness and Accuracy of' Measurement (Carpenter after 

Nelson) . ..... 79 

24. Ramsden Micrometer Eye-piece (Swift) . . 80 

25. Micrometer Eye-piece (Zeiss) 81 

26. Inoculating Needles (E.M.C.) . 84 

27. Freezing Microtome (Swift) 94 
,28. Microtome (Jung) . . 95 

29. Wire-cage for Test-tubes (Muencke) . 100 

30. Hot-air Steriliser (E.M.C.) . 101 



XVI LIST OF ILLUSTRATIONS. 

FIG. r'^O''- 

31. Hot-water Filtering Apparatus (Muencke) 102 

32. Method of making a Folded Filter (E.M.C.) 103 

33. Steam Steriliser (Baird and Tatlock) . 103 

34. Incubator (Muencke) . 104 

35. Method of Inoculating a Test-tube containing Sterile Nutrient Jelly 

(E.M.C.) . • • 105 

36. Levelling Apparatus (B.II.C) , 107 

37. Iron box for Glass Plates (Muencke) 108 

38. Method of Inoculating Test-tubes in the Preparation of Plate-cultiva- 

tions (E.M.C.) . . • 108 

39. Damp-chamber containing Plate-cultivations (E.M.C) 110 

40. Pasteur's Large Incubator (Becker) 111 

41. Petri's Dish (Becker) . . 112 

42. Glass Benches and Slides (Becker) 112 

43. Koch's Serum Steriliser (Muencke) . 114 

44. Hueppe's Serum Inspissator (Baird and Tatlock) 115 

45. Box for Sterilising Instruments (Becker) . 116 

46. Damp Chamber for Potato-cultivations (E.M.C.) . . 117 

47. Apparatus for Sterilisation by Steam under pressure (Baird and 

Tatlock) . . . . 119 

48. Drop Cultivation (Fliigge) . 121 

49. Simple Method of forming a Moist Cell (Schafer) . 122 

50. Warm Stage (Schafer) .... 123 

51. Warm Stage shown in Operation (Schafer) 123 

52. Warming Apparatus in Operation (Israel) . 124 

53. Section of Warming Apparatus and Drop-culture Slide (Israel) 125 

54. Israel's Warming Apparatus . 125 

55. Gas Chamber in use with Apparatus lor generating Carbonic Acid 

(Schafer) . 126 

56. Gas Chamber (Schafer) .... 126 

57. Moist Cell adapted for Transmission of Electricity (Schafer) . 1!^7 

58. Apparatus arranged for Transmitting- Electricity (Schafer) 127 

59. Slide with Gold-leaf Electrodes (Schafer) . 128 

60. Lister's Flask (Becker) . 128 

61. Sternberg's Bulb (Becker) 128 

62. Aitken's Tube (Becker) 129 

63. Miquel's Bulb (Becker) . 129 

64. Pasteur's Flask (Baird and Tatlock) 130 

65. Pasteur's Double Tube (Baird and Tatlock) 130 

66. Frankel'S Anaerobic Tube-culture (Frankland) . 131 

67. Anaerobic Culture Tube (Liborius) 132 

68. Apparatus for Anaerobic Cultures (Eoscoe and Lunt) . 133 

69. Koch's Syringe (Baird and Tatlock) . 135 

70. Syringe with Asbestos Plug (Baird and Tatlock) 135 

71. Hesse's Apparatus (Muencke) . 142 

72. Sedgwick and Tucker's Tube (Baird and Tatlock) 143 

73. Pouchet's Aeroscope (Hamilton) . 143 

74. Apparatus for Estimating the number -.£ Colonies in a Plate-cultiva- 

tion (Muencke) . 146 

75. Esmarch's Eoll-culture (Frankland) 147 

76. Apparatus for Counting Colonies in a Eoll-culture (Becker) 148 

77. Horizontal Micro-photographic Apparatus (Swift) 156 



LIST OF ILLUSTRATIONS. XVll 

FIG. , PAGE 

78. Eeversible Micro-photographic Apparatus (E.MX!.) . . . 157 

79. Eeversible Micro-photographic Apparatus arranged in the Vertical 

Position (E.M.C.) ....... 158 

80. Large Micro-photographic Apparatus (Swift) .... 160 

81. Photograph of an Impression Preparation (E.M.C.) . '. . 1B2 

82. Photograph of a Cultivation of Bacillus anthracis (E.M.C.) . . 168 

83. Suppuration of Subcutaneous Tissue (Cornil and Ranvier) . , 174 

84. Pus with Staphylococci, x 800 (Fliigge) . . . .177 

85. Sabcutaneous Tissue of a Rabbit forty-eight hours after an Injection 

of Staphylococci, x 950 (Baumgarten) . . . . 177 

86. Ulcerative Endocarditis : Section of Cardiac Muscle, x 700 (Koch) 183 

87. Pure-cultures of Streptococcus Pyogenes (E.M.C.) . . . 184 

88. Section of Skin in Erysipelas (Cornil and Ranvier) . . . 185 

89. Streptococcus Pyogenes Hominis ; Pure-cultures on Nutrient Gela- 

tine (E.M.C.) ........ 187 

90. Streptococcus Pyogenes Bovis ; Pure-cultures on Nutrient Gelatine, 

(E.M.C.) 188 

91. Gonococous, x 800 (Bumm) . . . . 190 

92. Bacillus Anthracis, x 1200. Blood Corpuscles and Bacilli unstained; 

from an Inoculated Mouse (Frankel and PfeifEer) . ; . 192 

93. Pure-cultivation of Baoillus anthracis in Nutrient Gelatine (E.M.C.) 193 

94. Colonies of Bacillits anthraeis, x 86 (Flttgge) . . . 194 

95. Impression-preparation of a Colony, x 70 (E.M.C.) . . . 194 

96. Margin of a Colony, x 250 (E.M.C.) . . . . .195 

97. Filaments with Oval and Irregular Elements, x 800 (E.M.C.) 195 

98. Spores of Baoillus anthracis stained with Gentian Violet, x 1500 

(E.M.C.) ........ 197 

99. Anthrax in Swine (E.M.C.) . . . . . .203 

100. Anthrax in Swine (E.M.C.) . . . . . .205 

• 101. Bacilli of Quarter-evil, x 1000 (Frankel and PfeifEer) . . 218 

102. Pure-culture of Bacilli of Quarter-evil in Grape-sugar Gelatine 

(Frankel and PfeifEer) . . . . . . .218 

103. Bacilli of Malignant .(Edema, x 950 (Baumgarten) . . .221 

104. Pure-culture of Baoillus of Malignant Oedema in Grape-Sngar 

Gelatine (Frankel and PfeifEer) .... 222 

105. Bacillipf Malignant (Edema, x 1000 (Frankel and Pfeiffer) . 223 

106. Pure-cultivation oE the Bacillus of Septicseraia of Mice in Nutrient 

Gelatine (E.M.C.) . . . . . . .225 

107. Bacterium of Rabbit SepticEemia ; Blood of Sparrow, x 700 (Koch) 228 

108. Bacterium of Fowl-cholera, x 1200 (E.M.C.) . , , . .228 

109. Bacterium of Fowl-cholera, x 2500 (E.M.C.) . . . .228 

110. Bacterium of Fowl-cholera; Section from Liver of Fowl, x 700 

(Flugge) 229 

111. Bacillus of Hemorrhagic Septioseinia, x 950 (Baumgarten) . 231 

112. Bacillus of Hsemorrhagio Septicaemia : Pure-culture in Gelatine 

(Baumgarten) ........ 231 

113. Bacterium Pneumonias Croupos^ from Pleural Cavity of a Mouse, 

x 1500 (Zopf) . 234 

114. Friedlander's Pneumoooccus ; Pure-culture in Nutrient Gelatine 

(Baumgarten) ........ 234 

115. Capsule Cocci from Pneumonia, x 1500 (Baumgarten) . . 235 

116. Micrococcus of Sputum Septiocemia, x lOCO (Frankel and PfeifEer) . 236 

b 



XVlii .LIST OF ILLUSTRATIONS. 

FiC. ^^°^ 

117. Colonies of Sternberg's Micrococcus, x 100 (Frankel and PfeifEer) . 237 

118. Aeate Catarriial Pneumonia, x 480 (Hamilton) . . .239 

119. Infectious Pleuro-pneumonia of Cattle (Hamilton) . . . 240 

120. Infectious Pleuro-pneumonia of Cattle (Hamilton) . . .241 

121. Bacillus of Influenza, x 1000 (Itzerott and Niemann) . 248 

122. Bacillus of Influenza, x 1200 (B.M.C) . . 249 

123. Bacilli of Plague and Phagocytes, x 800 (Aoyama) . . .253 

124. Spirillum Obermeieri in Blood of Monkey inoculated with Spirilla 

after Removal of the Spleen (Soudakewitch) . . 258 

125. Pure-cultivations of Streptococcus Pyogenes (E.M.C.) . 263 

126. Free Surface of Diphtheritic Larynx, x 350 (Hamilton) . . 831 

127. Bacillus of Diphtheria ; from a Cultivation on Blood Serum, x 1000 

. (Frankel and Pfeiifer) ...... 332 

128. Pure-cultures of Bacillus Diphtherise on Glycerine-Gelatine (E.M.C.) 333 

129. Typhoid Fever. Ileum of Adult, showing Sloughy and Infiltrated 

Patches (Hamilton) . . ... 341 

130. Typhoid Bacilli from a Colony on Nutrient Gelatine, x 1000 

(Frankel and PfeifEer) . . . . . . .342 

131. Typhoid Bacilli, x 950 (Baumgarten) . . 342 

132. Flagella of Typhoid Bacilli, x 1000 (Frankel and PfeifEer) . 343 

133. Colonies of the Typhoid Bacillus (Frankel and PfeifEer) . . 344 

134. Pure-culture of Typhoid Bacilli inoculated in the Depth of Nutrient 

Gelatine (Baumgarten) .... 344 

135. Typhoid Bacilli in a Section of Spleen, x 800 (Fliigge) . . 345 

136. Typhoid Bacilli in a Section of Intestine invading the Submucous 

and Muscular Layers, x 950 (Baumgarten) . . . .346 

137. Ulceration of the Intestine in a Typical Case of Swine-fever (E.M.C.) 348 

138. Klein's Bacillus of Swine-fever (No. 1) . . . . 349 
189. From a Preparation of Bronchial Mucus of Klein's Swine-fever 

Bacillus (No. 2) . . . . . . 349 

140. Bacilli from an Artificial Culture with Spores, Bacillus No. 2 (Klein). 349 

141. Blood of Fresh Spleen of a Mou.se after Inoculation with Swine-fever 

Bacillfls No. 2 (Klein) . ... . 350 

142. Bacilli of Swine Erysipelas (Baumgarten) . . .. 356 

143. Blood of Pigeon inoculated with Bacilli of Swine Erysipelas, x 600 

(Schiitz) . . . . .356 

144. Pure-culture in Nutrient Gelatine of Bacilli from Swine Erysipelas 

(Baumgarten) . . . . . 357 

145. Cover-glass Preparation of a Drop of Meat Infusion containing a 

Pure-cultivation of Comma-bacilli (Koch) . . 361 

146. Arthrospores of Comma-bacilU (Hueppe) . . . 361 

147. Flagella of Comma-bacilli ; stained by Loffler's Method (Frankel 

and PfeifEer) . . . . . 862 

148. Involution Forms of Comma-baciUi, x 700 (Van Ermengem) . 362 

149. Colonies of Comma-baoilli on Nutrient Gelatine ; natural size (Koch) 362 

150. Colonies of Koch's Comma-bacilli, x 60 (E.M.C.) . . 863 

151. Cover-glass Preparation from the Contents of a Cholera Intestine, 

x 600 (Koch) . . ... 363 

152. Cover-glass Preparation of Cholera Dejecta on Damp Linen, x 600 

(Koch) . . ..... 363 

153. Section of the Mucous Membrane of a Cholera Intestine, x 600 

(Koch) . . ... . . 364 



LIST OF ILLUSTRATIONS. XIX 

^^^' PAGE- 

154. Pure-cultivations in Nutrient Gelatine of Koch's and of Finkler's 

Comma-bacilli (B.M.C.) ... . . 365 

155. Comma-shaped Organisms with other Bacteria in Sewage-con- 

. taminated Water, x 1200 . . 366 

156. Commarbaoilli of the Mouth,. X 700 (Van Ermengem) . 367 

157. Finkler's Comma-bacilli, from Cholera nostras, x 700 (Fliigge) . 367 

158. Deneke's Comma-bacilli, from Cheese, x 700 (Fliigge) . . 367 

159. Pure-cultivation of the Spirillum Finkler-Prior, in Nutrient Gelatine 

(E.M.C.) 370 

1 60. Tropical Dysentery ; Mucous Membrane of Large Intestine (Hamilton) 372 

161. Tubercle of the Lung in a very Early Stage, x 400 (Hamilton) . 376 

162. Primary Tubercle of Lung two to three weeks old, x 50 ( Hamilton) 377 

163. Large Oval Giant Cell from Tubercle of Lung, x 300 (Hamilton) . 377 

164. Bacillus Tuberculosis, from Tubercular Sputum, x 2500 (E.M.C.) . 379 

165. Pure-cultivation of the Tubercle Bacillus on Glycerine Agar-agar 

(B.M.C.) 380 

166. Pure-cultivation in Glycerine Agar-agar after ten months' growth 

(E.M.C.) ........ 381 

167. Pure-cultivations of Tubercle Bacillus in Glycerine Agar-agar ; a sub- 

culture from a Pure-culture in Glycerine Milk (E.M.C.) . . 381 

168. Section through a Lupus Nodule of the Nose (Hamilton) . . 387 

169. Tubercular Ulceration. of Mucosa of Ileum (Hamilton) . . 388 

170. Section of Lupus of the Skin ; Giant Cell containing Tubercle 

Bacillus (Fliigge) . . .... 389 

171. Tuberculosis of Pleura; "Grape-disease" (E.M^.C.) . . . 390 

172. Tubercular Ulceration of the Intestine of a Cow (E.M.C.) . . 393 
178. Tubercular Ulceration of the Intestine of a Rabbit (E.M.C). . -395 

174. Tubercular Lungs of Rabbit (E.M.C.) . . . . .396 

175. Cover-glass Preparation of Pus from a Chancre, x 1050 (Lustgarten) 410 

176. Wandering Cell containing Bacilli (Lustgarten) . . . 410 

177. Section of Liver from a Case of Actinomycosis in Man (E.M.C.) . 417 

178. -Actinomycotic Tumour in the Throat of a Steer (E.M.C) . . 424 

179. Actinomycotic Tumour of the Cheek (B.M.C.) . . . .424 

180. Steer with Emaciation the Result of Actinomycosis (E.M.C.) 425 

181. Actinomycotic Growths from the- Pleura resembling " Grape- 

Disease" (E.M.C.) . '. 425 

182. Aotinomycosi.s of the Skin (E.M.C.) . . .430 

183. Part of Human Foot with Madura Disease (E.M.C.) . . . 448 

184. Bacilli of Glanders, x 700 (Fliigge) . . . .452 

185. Section of a Branch of the Pulmonary Artery showing Glanders 

Bacilli penetrating the Wall (Hamilton) .... 453 

186. Pure-culture of the Tetanus Bacillus in Grape-sugar Gelatine (Fraukel 

and Pfeiffier) ... .... 458 

187. Foot of Sheep showing Disease of Horn (Brown) . . 465 

188. Section through the Foot showing a Crack exteading through the 

Wall " 465 

189. Secreting Membrane covered with Fungoid Growths (Brown) . 466 

190. Advanced Form of Disease of Skin between the Claws (Brown) 466 

191. Distortion of Hoof in an Advanced Form of Foot-rot (Brown) . 467 

192. Diseased Comb (Con an) . . . . . .469 

193. Spores of Bacillus Alvei (B.M.C.) . . . . .470 
191. Pure-culture in Nutrient Gelatine (Cheshire and Cheyne) . . 470 



XX LIST OF ILLUSTRATIONS. 

F[G. PAGE 

195. Cultivation on the Surface of Gelatine (Cheshire and Cheyne) . 471 

196. Cladothrix Dichotoma (Zopf) . . . . . .479 

197. Friedlander's Pneumoooccus, x 1500 (Zopf) .... 483 

198. Ascococcus Billrothii (Cohn) .... . 498 

199. Clostridium Butyricum (Prazmowski) .... 503 

200. Bacillus Cyanogenus, x 650 (Neelsen) .... 507 

201. Pure-cultivation of Bacillus figurans on the Surface of Nutrient 

Agar-agar (B.M.C.) ...... 509 

202. Photograph of Part of an Impression Preparation of Bacillus 

figurans on Nutrient Gelatine, x 50 (E.M.C.) . . . 510 

203. Part of the same Specimen, x 200 . . . 510 

204. Bacillus Indicus : Colonies in Agar, x 60 . . 518 

205. Bacillus Neapolitanus, x 700 (fimmerioh) . . . .522 

206. Bacillus Megatherium (De Bary) ..... 524 

207. Pure-culture of Bacillus Miegatherium in Gelatine (E.M.C.) . . 524 

208. Bacillus Putrificus Coli, x 1000 (Bienstock) . . . .529 

209. Bacillus Pyogenes Foetidus, x 790 (Passet) . . . .530 

210. Bacillus Saprogenes, No. 1 (Eosenbach) . . 531 

211. Bacillus Subtilis with Spores (Banmgarten) .... 535 

212. Pure-culture of Bacillus Subtilis in Nutrient Gelatine (Banmgarten) . 535 

213. Pure-culture of Bacillus Subtilis on the Surface of Nutrient Agar 

(E.M.C) . . . . . . .536 

214. Bacterium Zopfii (Kurth) . . . . . .542 

215. Beggiatoa Alba (Zopf) ...... 543 

216. Phase-forms of Beggiatoa Persicina (Warming) . . . 544 

217. Cladothi-ix Dichotoma (Zopf) . . . . . .545 

218. Crenothrix Kiihniana (Zopf) ... . . 546 

219. Lenconostoc Mesenteroides (Van Tieghem and Cienkowski) . . 550 

220. Micrococcus in Pyaemia in Rabbits (Koch) .... 55H 

221. Proteus Mirabilis ; Swarming Islands on the Surface of Gelatine, 

X 285 (Hauser) ....... 561 

222. Proteus Mirabilis ; Involution Forms, x 524 (Hauser) . . 561 

223. Proteus Vulgaris, x 285 (Hauser^ .... 562 

224. Sarcina, x 600 (Flugge) . . . . . .563 

225. SpirochffiU Plioatile (E.M.C.) . . . 566 

226. Comma-bacilli in Water contaminated with Sewage . . . 567 

227. Comma-bacilli of the Mouth, x 700 (Van Ermengem) . . 568 

228. Deneke's Comma-bacilli, from Cheese, x 700 (Flugge) . . 568 

229. Streptococcus in Progressive Tissue Necrosis in Mice (Koch) . 571 

230. Vibrio Eugula, x 1020 (Prazmowski) . . . . .574 

231. Black Torula ; Pure-cultivation on Potato (E.M.C.) . . .581 

232. Head and Neck of Calf with Advanced Ringworm (Brown) . 585 

233. Non-pigmented Amoeboid Forms (Marchiafava and Celll) . 590 

234. Pigmented Amoeboid Forms (Golgi) . . . . 590 

235. Semi-lunar Bodies of Laveran (Golgi) . . 590 

236. Rosette Forms with Segmentation (Golgi) . . . 591 

237. Flagellated Fornas (Vandyke Carter) . . 591 

238. " Suira " Parasites, occuning Singly and Fu.sed, x 1200 . . 598 

239. Parasites in the Blood of Rats (Lewis) .... 699 

240. A Monad in Rat's Blood, x 3000 (E.M.C.) . . 601 

241. Monads in Rat's Blood, X 1200 (E.M.C.) . . . .602 

242. Monads in Bat's Blood stained with Methyl Violet, x 1200 (E.M.C.) 603 



LIST OF ILLUSTRATIONS. XXI 

yiO- PAGE 

243. Organisms in tlie Blood of Mud-fish (Mitrophanow) 604 

244. Organisms in the Blood of the Carp (Mitrophanow) . 605 

245. Amoeba cola in Intestinal Mucus (Losch) . . .611 

246. Warm Stage (Sohafer) . . . . . . .613 

247. Warm Stage (Strieker) .... . . 614 

248. Combined Gas Chamber and Warm Stage (Strieker) . . . 614 

249. Vertical Micro-photographic Apparatus (Leitz) . . 622 

250. Koch's Steam Sterihser (Muencke) . . . 623 

251. flot-air Steriliser (Muencke) . . . . . 623 

252. Section of Hot-air Steriliser (Muencke) . . . 623 

253. Hot-water Filtering Apparatus with Eing Burner (Eohrbeck) . 624 

254. Wire-cage for Test-tubes (Muencke) , . . 626 

255. Platinum-needles (E.M.C.) .... .626 

256. Damp Chalnber for Plate-cultivations (B.M.C.) . . 626 

257. Apparatus for Plate-cultivations (E.M.C.) . . . 627 

258. Box for Glass-plates (Muencke) . . . 627 

259. Glass Benches for Glass-plates (Becker) . 627 

260. Israel's Case (Becker) ... .628 

261. Damp Chamber for Potato Cultivations (E.M.C.) . 628 

262. Koch's SerUm Steriliser (Muencke) . . . . .629 

263. Serum Inspissator (Muencke) .... . 629 

264. D'Arsonval's Incubator (Muencke) . . , . . 631 

265. Hohlosing's Membrane Regulator (Muencke) . . . 632 

266. Gas Burner protected with Mica Cylinder (Muencke) . . 633 

267. Koch's Safety Burner (Muencke) . . 633 
. 268. Babes' Incubator (Muencke) ... .634 

269. Moitessier's Gas-pressure Regulator (Muencke) 634 

270. Reichert's Thermo-regulator (Muencke) . . 635 

271. Meyer's Thermo-regulator (Muencke) ..... 636 

272. Siphon Bottle with Flexible Tube, Glass Nozzle, and a Mohr's Pinch- 

cook (E.M.C.) .637 

273. Desiccator (E.M.C.) 638 



DESCRIPTION OP PLATES. 



DESCRIPTION OF PLATE I. 
Bacteria, Schizomycetes, or Fission Fungi. 

Following p. 14. 

1. Cocci siDgly and varying in size. 2. Cocci in chains or rosaries (strepto- 
coccus). 3. Cocci in a mass (staphylococcus). 4 and 5. Cocci in pairs 
(diplococcus). 6. Cocci in groups of four (merismopedia). 7. Cocci in packets 
(sarcina). 8. Bacterium, termo. 9. Baciervwm termo x 4000 (Dallinger and 
Drysdale). 10. Bacterium septic<smiee htemorrhagiecs. 11. Bacterium pneu- 
monice crouposce. 12. Bacillus suitilis. 13. Bacillus murisepticus. U. 
Bacillus diphtherieK. 15. Bacilliis typhosus (Eberth). 16. Spirillum, widula 
(Cohn). 17. Spirillum ■Kolutans (Cohn). 18. Spirillum ckolerce Asiatics. 
19. Spirillum Oiermeisri (Koch). 20. Spi/rncheeta plieatilis (Flugge). 21. 
Vibrio rugula (Prazm'owski). 22. Cladothrix Forsteri (Cohn). 23. Cladotltrix 
diclwtoma (Cohn). 24. Monas Okenii (Cobn). 25. Monas Warmi/iigii (Cohn). 
26. Mliabdomonas rosea (Cohn). 27. Spore-formation {Bamllus alvd'). 28. 
Spore-formation (Bacillus anthracis}. 29. Spore-formation in bacilli cultivated 
from a. rotten melon (Frankel and Pfeiffer). 30. Spore-formation in bacilli 
cultivated from earth (Frankel and Pfeiffer). 31. Involution-form of Crenothrix 
(Zopf). 32. Involution-forms of Vibrio serpens (Warming). 33. Involution- 
forms of Viirio rugula (Warming). 34. Involution-forrhs of Clostridium 
polymyxa (after Prazmowski). 35. Involution-forms of Spirillum cholera 
Asiaticie. 36. Involution-forms of Bacterium aceti (Zopf and Hansen). 
37. Spirulina-form of Beggiatoa alba (Zopf). 38. Various thread-forms of 
Bacterium merismopedioides (Zopf). 39. False-branchijig of Cladotliri(i;(Zapt). 



DESCRIPTION OP PLATE II. 

Pure-cultivations of Bacteria. 

Following p. 100. 
Fig. 1. — In the dejith of Kvtrient Gelatine. A pure-cultivation of Koch's 
comma-bacillus (Spirillum choleras Asiaticae) stowing in the track of 
the needle a funnel-shaped area of liquefaction enclosing an air-bu"bble, 
and a white thread. Similar appearances are produced in cultivations of 
the comma-bacillus of Metchnlkoff. 
Fig. 2. — On tlie surface of Nutrient Gelatine. A pure-cultivation of Bacillus 
typhosus on the surface of obliquely solidified nutrient gelatine. 

xxii 



DESCRIPTION OF PLATES. XXlll 

Fig. 3. — On the surface of Nutrient Agar-agwr. Pure-cultivation of Bacillus 
indicus on the surface of obliquely solidified nutrient agar-agar. The 
growth has the colour of red sealing-wax, and a peculiar crinkled 
appearance. After some days it loses its bright colour and becomes 
purplish, like an old cultivation of Micrococcus prodigiosus. 

Fig. 4. — On tlw smfaoe of Nutrient Agar-agar. A pure-cultivation obtained 
from an abscess (Staphylococcus pyogenes aureus). 

Fig. 5. — On tlie sv/rfaoe of Nutrient Agar-agar. A pure-cultivation obtained 
from green pus (Bacillus pyocyaneus). The growth forms a whitish, 
transparent layer, composed of slender bacilli, and the green pigment 
is difEased throughout the nutrient jelly. The growth appears green by 
transmitted light, owing to the colour of the jelly behind it. 

Fig. 6. — On tJie surface of Potato. A pure-cultivation of the bacillus of 
glanders on the surface of sterilised potato. 



DESCKIPTION OF PLATE III. 

Plate-cultivation. 

FoUomng p. 108. 

This represents the appearance of a plate-cultivation of the comma-bacillus 
of Cholera nostras, when it is examined over a slab of blackened plate-glass. 
The drawing was made from a, typical result of thinning out the colonies by 
the process of plate-cultivation. At this stage they were completely isolated 
one from the other ; but later they became confluent, and produced complete 
liquefaction of the gelatine. 

DESCRIPTION OF PLATE IV. 

Streptococcus Pyogenes. 

Following p. 178. 

Fig. 1. — From a cover-glass preparation of pus from a pygemio abscess. 
Stained with gentian-violet by the method of Gram, and contrast-stained 
with eosin. x 1200. Powell and Lealand's apochromatic ^^ Hom. imm. 
E. P. 10. 
Fig. 2. — From cover-glass preparations of artificial cultivations of the strepto- 
coccus in broth and in milk at different stages of growth, x 1200. Powell 
and Lealand's apochromatic -^ Hom. imm. E. P. 10. 
In these preparations there is a great diversity in size and form of the 
chains and their component elements. In the drawing examples are 
figured of the following: 
(ffi) Branched chains. 
(J) Simple chains composed of elements much smaller than the 

average size. 
(c) Chains with spherical and spindle-shaped elements at irregular 
intervals. These are conspicuous by their size, and are sometimes 
terminal, 
((i e) Chains in which the elements are more or less uniform in size. 
(/) Complex chains with elements dividing both longitudinally and 
transversely, and varying considerably in size in different lengths 
of the same chain. 



XX1\- DESCRIPTION OF PLATES. 

DESCRIPTION OF PLATE V. 

Bacillus Authracis. 

FoUowing p. 192. ^ 

KiG.' I. — From a cover-glass preparation of blood from the spleen of a guinea- 
pig inoculated with blood from a sow. x 1200. Powell and Lealand's 
apochromatio yV Horn. imm. E.. P. 10. 

Fig. 2. — From a section of a kidney of a mouse. Under a low power the 
preparation has exactly the appearance of an injected specimen. Under 
higher amplification the bacilli are seen to have threaded their way along 
the capillaries between the tubules, and to have collected in masses in 
the glomeruli. Stained with Gram's method (gentian-violet), and eosin. 
X 500. 

Fig. 3. — Sacillus anihracis and Microcoemis tetragemig. From a section from 
the lungs of a mouse which had been inoculated with anthrax three days 
after inoculation with Micrococcus tetragenus. A double or mixed infection 
resulted. Anthrax-bacilli occurred in vast numbers, completely filling the 
small vessels and capillaries, and in addition there were great numbers 
of tetrads. Stained by Gram's method (gentian-violet), and with eosin. 
X 500. 



DESCKIPTION OF PLATE VI. 
Bacillus Murisepticus. 

Follovnnff p. 224. 

Fig. 1. — From a section of a kidney of a mouse which had died after inocula 
tion with a pure-cultivation of the bacillus. With moderate amplification, 
the white blood-corpuscles have a granular appearance, and irregular 
granular masses are scattered between the kidney tubules. Stained by 
Gram's method with eosin. x 200. 

Fig. 2. — Part of the same preparation with high amplification. The granular 
appearances are found to be due to the presence of great numbers of 
extremely minute bacilli, x 1500. 



DESCEIPTIOX OF PLATE VII. 

Casual Cow-pox. 

FoUotcinff p. 278. 

Fig. 1. — Case of W. P , a milker, infected from the teats of a cow with 

natural cow-pox. There was a large depressed vesicle with a small 
central crust and a tumid margin, the whole being surrounded by a 
well-marked areola and considerable surrounding induration. 

Fig. 2. — The same case a week later, showing a reddish-brown crust on a 
reddened elevated and indurated base. 



DESCRIPTION OF PLATES. XXV 

UESCRH'TION OF PLATE VIII. 

Bacillus diphtherise and Bacillus typhosus. 

Follovniiff p. 382. 

i'm. 1. — Cover-glass preparation from a pure-cultivation of Bacillus diph- 
therias on blood serum ; obtained from the throat in a, typical case of 
diphtheria. Stained with gentian-violet, x 1200. 

Fig. 2. — Cover-glass preparation from a pure-cultivation of Bacillus typhosus 
on nutrient-agar ; from the spleen in a case of typhoid fever, Stained 
with-gentian-violet. x 1200. 



DESCRIPTION OF PLATES IX. AND X. 

Swine Fever. 

FollotDing p. &i8. 

Plate IX. — Part of intestine from a typical case of swine fever, showing 
scattered ulcers and ulceration of the ileo-caecal valve. 

Plate X. — From the same case of swine fever. The lungs were extensively 
inflamed and partly consolidated, and the lymphatic glands were enlarged 
and of a deep red or reddish-purple colour. ' 



DESCRIPTION OF PLATE XI. 

Bacillus tuberculosis. 

Following p. 378, 

The figures in this plate represent the bacilli of tuberculosis in 
different animals, examined under the same conditions of amplifica- 
tion and illumination, x 1200. Lamp-light illumination. 

Fig. 1. — Bacilli in pas from the wall of a human tubercular cavity. In 
this specimen the bacilli are shorter than those in tubercular sputum, 
and are very markedly beajJed. 

Fig. 2. — Bacilli in pus from a tubercular cavity from another case in man. 
They are present in the preparation in enormous numbers. The proto- 
plasm occupies almost the whole of the sheath, and the bacilli are 
strikingly thin and long. 

Fig. 3. — Bacilli in sputum from an advanced case of phthisis, showing 
the ordinary appearance of bacilli in sputum ; some beaded, others 
stained in their entirety ; occurring both singly and in pairs, and 
in groups resembling Chinese letters. 

Fig. 4. — Bacilli in a section from the lung in a case of tuberculosis in man. 
The bacilli iu human tuberculosis are found in, and between, the tissue 
cells ; and sometimes, as in equine and bovine tuberculosis, in the 
interior of giant cells, but not so commonly. 

Fig. 5. — B'rom a cover-glass preparation of the deposit in a sample of milk 
from a tubercular cow. The bacilli were longer than the average 
length of bacilli in bovine tissue sections, and many were markedly 
beaded. 



XXVI DESCRIPTION OF PLATES. 

Fig, B. — From a section of the brain in a case of tubercular meningitis in a 
calf, showing a giant cell containing bacilli with the characters usually 
found in sections of bovine tuberculosis. 

Fig. 7. — From a section of the liver of a pig with tubercle bacilli at the 
margin of a caseous nodule. 

Fig. 8. — From a cover-glass preparation of a crushed caseous mesenteric 
gland from a rabbit infected by ingestion of milk from a cow with 
tuberculosis of the adder. 

Fig. 9. — From a section of lung in a case of equine tuberculosis, showing a 
giant cell crowded with tubercle bacilli. 

Fig.. 10. — From a section of lung from a case of tuberculosis in the cat, with 
very numerous tubercle bacilli. 

Fig. 11. — From a cover-glass preparation of a crushed caseous nodule from 
the liver of a fowl, with masses of bacilli. These are for the most part 
short, straight rods ; but other forms, varying from long rods to mere 
granules, are also found. 

Fig. 12., — From sections of the liver and of the lung in a case of tubercu- 
losis of a Rhea. Isolated bacilli are found, as well as bacilli packed in 
large cells, colonies of sinuous bacilli, and very long forms with terminal 
spore-like bodies and free oval grains. 

The preparations from vyhicli these figures were drawn were all 
stained by the Ziehl-Neelsen method, with the exception of the first, 
which was stained by Ehrlich's method. ' 



DESCRIPTION OF PLATE XIL 

Tubercular Mammitis. 

Folloieing p. 394. 

Fig. 1'.— From a section of the udder of a milch cow. The tubercular deposit 
is seen to invade the lobules of the gland. Lobules comparatively healthy 
are marked ofE, more or less sharply, from the diseased ones in which the 
new growth in its progress compresses and obliterates the alveoli. Stained 
by the Ziehl-Neelsen method and with methylene-blue. x 50. 

Fig. 2.— Part of the same preparation. On the right of the section part of a 
healthy lobule is seen. On the left a lobule is invaded by tubercular new 
growth composed of round cells, epithelioid cells and typical giant cells. 
Tubercle bacilli can be seen both singly and collected in groups. They 
are found in and between the cells, and in the interior of giant cells. 
Bacilli may be seen between the cells lining an alveolus and projecting 
into its lumen, x 800. 



DESCRIPTION OF PLATE XIII. 

Tuberculosis in Swine. 

Follomnff p. 400. 

Section of liver of a pig with scattered tubercular nodules. Microscopical 
sections of the liver showed tiibercle bacilli in very small numbers. 



DESCRIPTION OF PLATES. XXVU 

DESCEIPTION OF PLATE XIV. 

Sacillus Leprae. 

Follomtng p. 408. 

Fig. 1. — From a section of the skin of a leper. The section is, alinost in 
its entirety, stained red, and, with moderate amplification, has a finely 
granular appearance. Stained by the Ziehl-Neelsen method (carbblised 
fuchsine and methylene-blne). x 200. 

Fig-. 2. — Part of the same preparation with higb amplification, showing that 
the appearances described above are due entirely to an invasion of the 
tissue by the bacilli of leprosy, x 1500. 

DESCRIPTION OF PLATES XV. AND XVI. 

Actinomyces. 

Fallowing p. 432. 

Plate XV. 
Fie. 1. — From a preparation of the grains from an actinomycotic abscess in 
a boy ; examined in glycerine. The drawing has been made of a com- 
plete rosette examined by focussing successively the central and peripheral 
portions. Towards the centre the extremities of the clubs are alone 
visible ; they vary in size, and if pressed upon by the cover-glass give the 
appearance of an irregular mosaic. Towards the periphery the cliibs are 
seen in profile, and their characteristic form recognised. At one part 
there are several elongated elements, composed of separate links, x 1200. 
Fig. 2. — DifEerent forms of clubs from preparations in which the rosettes have 
been flattened out by gentle pressure on the cover-glass, x 2500. 

(«) Single club. (V) Bifid club, (o) Club giving rise to four 
secondary clubs, (d) Four clubs connected together, recalling 
the form of a bunch of bananas, (e) Mature club with a lateral 
bud. (/) Apparently a further development of the condition 
represented at (e). {g) Club with a lateral bud and transverse 
segmentation. Qi) Single club with double tranverse segmenta- 
tion, (i) Club with oblique segmentation, (j) Collection of 
four clubs, one with lateral gemmation,' another with oblique 
segmentation. (K) Club with lateral buds on both sides, and 
cut ofE square at the extremity. (Q Club with a daughter club 
which bears at its extremity two still smaller clubs. (»j) Club 
divided by transverse segmentation into four distinct elements. 
(») Elongated club composed of several distinct elements, (o) and 
(_p) Clubs with terminal gemmation, (j) Palmate group of clubs. 
(«•) Trilobed club, (s) Club with apparently a central channel. 
(f) Filament bearing terminally a highly refractive oval body. 

Plate XVI. 
Fig. 1. — From a section of a portion of the growth removed from a boy 

during life. The tissue was hardened in alcohol, and cut in celloidin. 

The section was stained by Gram's method and with orange-rubin. x 50. 
Fig. 2. — From the same section. A mass of extremely fine filaments occupies 

the central part of the rosette. Many of the filaments have a terminal 

enlargement. The marginal part shows a palisade of clubs stained by the 

orange-rubin. x 500. 



XXVlll DESCRIPTION OF PLATES. 

Figs. 3 and 4.— From cover-glass preparations of the fungus teased out of the 
new growths produced by inoculation of a calf with pus from a boy 
suffering from pulmonary actinomycosis. Stained by Gram's method and 
orange-rubin. The threads are stained blue and the clubs crimson (a) 
In the younger clubs the thread can be traced into the interior of the 
club (*). In some of the older clubs the central portion takes a yellowish 
stain, and in others the protoplasm is not continued as a thread, but is 
collected into a spherical or ovoid or pear-shaped mass. In others, again, 
irregular grains stained blue are scattered throughout the central portion 
(Fig. 4). X 1200. 

Fig. 5. — From a pure-culture on glycerine-agar. (a) branching filaments, 
(ft) a mass of entangled filaments. Gram's method, x 1200. 

Fig. 6.— From a similar but older cultivation, (a) a filament with spores, 
(ft) chains of spores simulating streptococci. Gram's method, x 1200. 



DESCRIPTION OF PLATES XVII. AND XVIII. 
Actinomycosis Bovis. 

Following p. 434. 

Plate XVII. 
Section of an actinomycotic tongue stained by the method of 
Gram and with eosin. 

Fig. 1. — This illustrates the appearance which is usually seen under a low 
power, when a section is stained by Gram's method and with eosin. The 
central portion of a mass of the fungus is either unstained or tinged virith 
eosin, while the marginal portion is stained blue. The reverse is seen, as a 
rule, in sections from man ; although under a low power the general appear- 
ance of sections from these two sources is somewhat similar, x 50. 

Fig. 2. — a, ft, o, d, represent the earliest recognisable forms of the ray fungus 
in the interior of leucocytes. In e the club-forms can be recognised. In 
/ and g there are small stellate groups of clubs, x 500. 

Fig. 3. — A part of the section represented in Fig. 1, under a high power. The 
marginal line of blue observed under a low power is now recognised as the 
result of the stain being limited to the peripherally arranged clubs. At 
(a) part of a rosette has undergone calcification ; the clubs are granular, 
and have not retained the stain. At (ft) and close to it there are the 
remains of rosettes in which the process of calcification is almost complete. 
X 500. 

Plate XVIII. 

The figures in this plate are taken from sections of a case of 
so-called " osteosarcoma," in which the growth of the fungus was 
remarkably luxuriant. The specimens were stained by Plants' 
method. 

Fig. 1. — Different forms of clubs in different specimens : x 1200. 
(a) Very small club-shaped elements, 
(ft) A club with transverse segmentation, 
(c) A club with lateral daughter clubs. 



DESCRIPTION OF PLATES. XXIX 

(d and e) Clubs with terminal offshoots resembling teleutospores. 
(J) A club with developing daughter clubs on the left, and on the 

right a mature secondary club. 
(g) A segmental club with lateral offshoots. 
(A) Two clubs undergoing calcification. 
Fl&. 2. — A very remarkable stellate growth comprised of nine wedge-shaped 
collections of clubs radiating from a mass of finely granular material. 
X 500. 
Fig. 3. — A rosette undergoing central calcification, and consisting in part of 
extremely elongated clubs resembling paraphyses. Calcareous matter is 
also being deposited in the club-shaped structures, x 500. 
Fig. 4. — Part of a rosette with continuation of the club-shaped bodies 
into transversely segmented branching cells apparently representing short 
hyphae. * 500. 
Fig. 5. — A rosette from another section in which similar appearances are 
observed as in Fig. 4. x 500. 

DESCEIPTION OF PLATE XIX. 
Pure-cultivations of Actinomyces. 

FolUming p. 438. 

These tubes were selected from a great number of cultivations 
in which there were different appearances. In some instances the 
growths had a faint tinge of pink. 

Fig. 1. — Pure-cultivation on the surface of potato, showing a luxuriant 
sulphur-yellow growth entirely composed of entangled masses of fila- 
ments. After three months' growth. 

Fig. 2. — Pure-culture from the same series, on glycerine-agar. In this case 
the culture remained perfectly white. The jelly was coloured reddish- 
brown. After fifteen months' growth. 

FiG.'S. — Pure-culture on glycerine-agar in which the growth was dark- 
brown, in parts black, and the jelly stained dark-brown. After nearly 
two years' growth. 

DESCRIPTION OF PLATES XX. AND XXI. 
Actinomycosis Bovis. 

FoUoiBing p. 440. 

Plate XX. 

Fig. 1. — From a section of an actinomycotic tongue stained by the triple 
method (Ziehl-Neelsen, logwood and orange-rubin). In this section the 
separate centres of growth are clearly shown. Each neoplasm consists of 
a fungus system, in which the masses of the fungus, situated more or less 
centrally, are surrounded with round cells, epithelioid cells, sometimes 
giant cell's, and lastly fibrous tissue forming a more or less distinct 
capsule. In parts the fungi have fallen out of the section. x 50. 

Fig. 2. — From a section of a "tubercular" nodule from the lungs of a 
Norfolk heifer with pulmonary actinomycosis. The nodule is a multiple 
growth surrounding a bronchus, and is enclosed by a capsule, in the 



XXX DESCRirriON OF PLATES. 

vicinity of which the pulmonary alveoli are compressed. It is composed 
of a number of separate neoplasms, and each of the latter is composed of 
secondary centres of growth resembling the giant-cell systems of bacillary 
tuberculosis. The new growth is composed of ray-fungi, large multi- 
nucleated cells, sometimes distinct giant cells, round cells, epithelioid cells, 
and, surrounding them, fibrous tissue. On examination of the same 
specimen with a higher power the typical rosettes of clubs are sometimes 
surrounded by multinucleated cells, and sometimes small rosettes are 
found like tubercle bacilli, in the interior of giant cells. From a pre- 
paration stained by Ziehl-Neelsen, logwood, and orange-rubin. x .50. 

Plate XXI. 

Fie. 1. — (a) A leucocyte containing the fungus in its earliest recognisable 
form. (J) A large multinucleated cell containing the fungus in an early 
stage with the club-form already visible, (c) A leucocyte containing a 
small stellate fungus, (d) A large cell containing clubs arranged in a 
small rosette. («) A multinucleated cell with clubs arranged in a palmate 
form. All the above are drawn from sections of actinomycotic tongues 
stained by the triple method, x 500. 

FiC4. 2. — A giant cell with large vesicular nuclei at the periphery, and in the 
centre a fully formed rosette of actinomyces with a smaller growth within 
ci " daughter " cell. From a section of the tongue of an ox stained by 
the triple method, x 500. 

Fig. 3. — A very large circular giant cell, with its ring of niiclei at the 
periphery, enclosing several isolated tufts of actinomyces. From a section 
of a nodule in the lung. Stained by the triple method, x 500. 

Fig. 4. — Three rosettes of actinomyces surrounded by a row of large, some- 
what angular multinucleated cells. From a section, of the tongue of an 
ox stained by the triple method, x 430. 



DESCKIPTION OF PLATE XXII. 

Sacillus tetani. 

Folloialng p. 45S. 

Fig. 1. — From a cover-glass preparation of a pure-cultivation of the tetanus 
bacillus in broth ; stained with Neelsen's carbolised fuchsine. y 1200. 
Lamplight illumination, 

Fig. 2. — From a cover-glass preparation from the same source ; stained with ' 
Neelsen's solution and methylene blue, x 1200. Lamplight illumination. 



PART I. 

THEORETICAL AND TECHNICAL. 



BACTERIOLOGY 

AND 

INFECTIVE DISEASES. 



CHAPTEE I. 

HISTORICAL INTRODUCTION. 



The researches of Pasteur into the rtle played by bacteria in the 
processes of fermentation and putrefaction, and the investigations of 
the practical mind of Lister, with the resulting evolution of antiseptic 
surgery, demonstrated the necessity for a more intimate acquaint- 
ance with the life-history of these micro-organisms. Further re- 
searches in diseases such as anthrax, the silkworm malady, pysemia, 
septicsemia, and fowl-cholera, invested the science of Bacteriology 
with universal interest and vast importance; while the investiga- 
tions which established an intimate connection between bacteria 
and other infective diseases, and more especially the discovery by 
Koch of bacteria in tuberculosis and in Asiatic cholera, claimed the 
attention of the whole thinking world. 

Those bacteria which are connected with disease, and more 
especially those which have been proved to be the causa causans, are 
of predominant interest and importance. 

The first attempt to demonstrate the existence of a contagimn 
vivvm, dates back almost to the discovery of the microscope. 
Athanasius Kircher, nearly two and a half centuries ago, expressed 
his belief that there were definite micro-organisms to which diseases 
were attributable. The microscope had revealed that all decom- 
posing substances swarmed with countless micro-organisms which 
were invisible to the naked eye, and Kircher sought for similar 
organisms in diseases which he considered might be due to their 
agency. The microscope which he described obviously could not 

1 



^ BACTERIOLOGY. 

admit of the possibility of studying, or even detecting, the micro- 
organisms which are now known to be associated with certain 
diseases ; and it is not surprising that his teacliings did not at the 
time gain much attention. They were destined, however, to receive 
a great impetus from the discoveries which emanated from "the 
father of microscopy." 

Antony van Leeuwenhoek had learned as a youth to grind and 
polish lenses, and later in Uf e employed his spare time in constructing 
microscopes, and in conducting those researches which have made 
for him a name which is famihar to all microscopists. His researches 
were pubhshed in a series of letters to the Royal Society. In 1675 
he described^ extremely minute organisms in rain-water, well-water, 
infusions of pepper, hay, and other vegetable and animal substances, 
in saliva, and in scrapings from the teeth ; and, further, he was 
able to differentiate these minute living things by their size, their 
form, and the character of their movements. In 1683 these 
discoveries were illustrated by means of woodcuts, and there can be 
little doubt, from the drawings of these micro-organisms, that they 
are intended to represent leptothrix filaments, vibrios, and spirilla. 
Indeed, we can almost recognise these micro-organisms as bacteria 
from Leeuwenhoek's graphic descriptions, apart from his figures. 
They were described as moving in the most characteristic manner, 
progressing with great rapidi ty, or . spinning round like a top, and 
so excessively minute that they were only perceived with great 
difiiculty. The smallest forms could hardly be examined individually ; 
but, viewed en masse, they closely resembled a swarm of gnats or 
flies. In another communication, published in 1692, he gives 
some idea of the size of these animalcules by stating that they 
were a thousand times smaller than a grain of sand. Others 
which were, comparatively speaking, of considerable length, were 
characterised by their peculiar mode of progression, bending and 
rolling on themselves— movements which, he adds, created both 
delight and astonishment in the mind of the observer. Leeuwenhoek 
himself was not disposed to believe in the possibility of such 
organisms being found in the blood in disease; but as soon as he 
had proved the actual existence of such minute creatures, theoretical 
physicians were not wanting who at once attributed various maladies 
to their agency. Among these, Nicholas Andry is made conspicuous 
by his work published in 1701. Andry classed the minute organisms 
discovered by Leeuwenhoek as worms. 

In 1718 Lancisi believed that the deleterious effect of the air of 
malarial districts depended upon animalcules, and others considered 



HISTORICAL INTRODUCTION. A 

that the plague in Toulon and Marseilles in 1721 arose from a 
similar cause. In fact, by some, all diseases were attributed to 
vermicules, and this led to the theory being ridiculed arid discredited. 

In spite of adverse criticism, the theory of contagium vivum 
survived, and Linnseus acknowledged it by placing the micro- 
organisms discovered by Leeuwenhoek, the contagia of specific 
fevers, and the causes of putrefaction and fermentation, into one 
class — " chaos." The theory was further supported by the writings 
of Plenciz, who, in 1762, very ably discussed the nature of contagium, 
as well as the relation of animalcules to putrefaction and disease. 
However, no proofs in Support of these theories were forthcoming,, 
and gradually the idea of contagium vivum fell into obscurity, and 
indeed came to be regarded by some as an absurd hypothesis. 

Though a causal relation of animalcules to diseases was for a 
time discredited, the natural history of these micro-organisms was 
studied with increasing interest. In 1778 Baron Gleichen described 
and figured a great number of micro-organisms which he had 
discovered in various vegetable infusions. Joblot, Lesser, Reaumur, 
HUl, and many others worked at the same subject. Hill remarked 
that there was hardly the least portion of matter or the least drop 
of fluid of any kind naturally found in the earth, which was not 
inhabited by multitudes of animalcules. But these observers incUned 
rather to searching for new forms than to studying more thoroughly 
those which had been already discovered ; and, as a result, but little 
scientific progress \yas made until the time of Miiller, of Copen- 
hagen. Miiller, in 17&6, criticised the work of previous writers, 
and pointed out that they had been too much occupied witli merely 
finding new micro-organisms. Miiller took into account the form 
of the micro-organism, its mode of progression, and other biological 
characters, and on such data based a classification. Thus the 
scientific knowledge of these minute beings was considerably advanced 
by his writings and illustrations. 

The subject which now eclipsed all others in interest was the 
origin of these micro-organisms. Two rival theories were widely 
discussed — spontaneous generation, and development from pre-exist- 
ing germs; and the researches that were made in the course of 
this discussion, and the discoveries which resulted, indirectly yet 
materially advanced the germ theory of disease, and explain many 
of the phenomena in the life-history of the pathogenic microbes 
which have been brought to light in recent years. 

Spontaneous development of micro-organisms in putrescible 
infusions was believed in by many, but was supported by no one 



4 BACTERIOLOGY. 

with greater persistency than Needham. Needham found that 
animalcules readily developed when meat infusion was boiled an^ 
transferred to a well-stoppered flask, and he could only explain this 
by supposing that they originated spontaneously from the material of 
the infusion. In 1768 Bonnet strenuously opposed these conclusions 
on purely theoretical grounds, and maintained that it was far more 
probable that the ova of the animalcules were present in the infusions 
or were suspended in the air enclosed in the flask. 

Spallanzani was the first to demonstrate by experiment the 
correctness of Bonnet's arguments. It occurred to him to boil the 
infusion in flasks, and to seal the vessels during the process of boiling. 
As a result the flasks remained free from putrefaction, and 
animalcules only developed when the infusion was exposed to the air 
by making a hole in the flask. That Spallanzani's experiments 
were reliable, and his conclusions correct, was evidenced by the fact 
that his simple precaution led to great practical results, as i'ran9ois 
Appert introduced, on this principle, the method of preserving meats, 
vegetables, and other provisions. 

The disciples of Needham nevertheless brought forward counter 
objections. Treviranus urged that a certain quantity and quality of 
air was necessary for the spontaneous development of these infusoria, 
and that by sealing the flasks, too small a quantity of air was in 
■contact with the infusion, and, further, that this air had become 
changed in quahty by the process of boiling. 

Spallanzani argued against these objections, but did not support 
his opinions by further experiments, so that the question remained 
for a time undecided. 

In 1836 Francis Schulze devised an experiment which brought 
still further evidence against Needham's theory. Schulze filled a 
glass vessel half full with distilled water and different animal and 
vegetable substances. This was plugged with a doubly-bored cork, 
and through each perforation a glass tube was introduced, bent at 
a right angle. On boiling the flask, steam issued freely from each 
tube, and all parts were thoroughly sterilised. Each tube was then 
connected with a bulbed tube, one bulb containing concentrated 
sulphuric acid and the other a solution of potash. Fresh air was 
drawn into the flask by aspiration, and this was deprived of any 
germs which might be present by its passage through the siilphuric 
acid. The result was that the infusion remained without any 
development of micro-organisms. When, on the other hand, air was 
admitted without first being drawn through the sulphuric acid, the 
infusion in a short time teemed with animalcules. In other words, 



HISTORICAL INTRODUCTION. Cf 

Schulze demonstrated that in spite of free access to air, which Imd not 
been heated, the infusions remained free from germs. 

Schwann, in 1837, arrived at similar results. He found that 
putrescible substances remained sterile if exposed to an abundant 
supply of air which was heated by being passed through a melted mix- 
ture of metals. This convinced him that the cause of the decompo- 
sition which would otherwise have occurred must exist in the air. 

The objection remained that in the experiments of Schvilze and 
Schwann, the air which was admitted to the flasks had undergone 
either a chemical or a thermal change, and therefore the theory of 
Needham was not yet entirely disposed of. 

In 1854 the final blow was dealt by Schroder and Yan Dusch. 
These investigators demonstrated that decomposition could be obviated 
without resorting either to thermal or chemical treatment of the 
air, as simple filtration of the air through cotton-wool was shown to 
be efficacious in excluding germs. Finally, Hoffman in 1860, and 
independently, Ohevreuil and Pasteur in 1861, showed that even 
cotton-wool could be dispensed with, as a sterile solution would 
remain sterile when the neck of the vessel was bent into an 
S -shaped curve. Micro-organisms in the air entering the flask 
were deposited by gravitation in the bend of the tube. 

The advocates of spontaneous generation were ready with fresh 
objections. They now urged that the medium lost its power of 
undergoing decomposition by being boiled. This objection was at 
once set aside by the fact that when unfiltered air was admitted to 
the infusion, decomposition set in. Additional evidence was brought 
against spontaneous generation by the experiments of Pasteur, 
Burden Sanderson, Lister, and others, in which it was shown that 
blood, urine, and milk would remain without decomposition, when all 
precautions were adopted to avoid contamination in filling the 
sterilised flasks. 

Even at this stage of this great scientific controversy fresh 
difiiculties arose, for it was found that in certain solutions which had 
been boiled and hermetically sealed in flasks micro-organisms made 
their appearance. In 1872 Charlton Bastian published a research 
which was to prove that spontaneous generation actually took 
place. Decoctions of turnip and cheese which had been filtered, 
neutralised, and boiled for ten minutes, and hermetically sealed 
during the boiling, were found after a time to contain micro 
organisms. These results, however, were before long explained by 
the fact that in milk, infusions of hay, and certain other decoctions, 
the spores of bacilli are present, which are much more resistant 



6 BACTERIOLOGY. 

than tlie bacilli themselves. In such cases mere scalding or boiling 
for a few minutes wiU not sterilise the solution. The bacilli are 
destroyed, but not their spores; and if the latter remain unhurt, 
they will germinate, and rapidly multiply. But if, as Tyndall 
foiind, the boihng be repeated a second and a third time, all the 
spores will be destroyed ; for in the intervals between the boilings 
the spores sprout into bacilli, and the bacilli at the next boihng 
perish ; so that after three or four repeated boilings the infusion is 
rendered perfectly free from germs. 

While this discussion was occupying the attention of the whole 
scientific world, some investigators had been again following up the 
theory of a connection between micro-organisms and disease. 

In 1837 Cagniard Latour and Schwann independently made the 
•discovery that the yeast plant was a living organism, and the true 
cause of yeast fermentation. The close analogy between the pro- 
cesses of fermentation and of certain diseases had long been held; 
and, therefore, when it was proved that fermentation was due to a 
micro-organism, fresh advocates appeared in support of the theory 
that diseases were produced by similar agencies. Boehm, in 1838, 
described certain organisms in cholera, which was at that time 
raging in Europe; but the researches of Bassi, who a year 
previously had discovered the cause of a disease of silkworms, 
attracted much greater attention. 

Bassi discovered that in this disease extremely minute spores 
existed on the bodies of the worms, which were conveyed from the 
sick to the healthy. They destroyed the healthy worms by 
germinating in their skins and growing into their bodies. These 
discoveries may be said to have brought the theory of contagium 
vivum to Ufe again ; and Henle, in reviewing the facts of the case in 
1 840, came to the conclusion that the cause of all contagious diseases 
must be of a living nature, and this he maintained, although he 
had searched in vaecine and small-pox lymph, in the desquamation 
of scarlet fever, and in other diseases without success. 

Bassi's discovery and Henle's doctrine encouraged a number of 
investigators, and remarkable results followed. In favus, in herpes 
tonsurans, in pityriasis versicolor, fungus threads and spores were 
found, and were regarded as being of etiological importance, 
inasmuch as the morbid lesions corresponded vsdth the growth of the 
particular fungus. 

Cholera became especially a subject for research. Swaine, 
Brittan, and Budd found micro-organisms in choleraic dejecta. 
Davaine described certain monads in the intestinal contents, but no 



HISTOKICAL INTRODUCTION. 7 

causal connection was established between these organisms and the 
disease ; and when the cholera disappeared the interest in contagium 
vivum waned, and was eclipsed by the question of fermentation. 
The discoveries which followed in this subject had a very important 
bearing on the micro-parasitic origin of communicable diseases. 

Pasteur, following up the researches of Cagniard Latour and 
Schwann, demonstrated in 1857 that the lactic, acetic, and butyric 
fermentations were produced by micro-prganisms. 

Previously to this, in 1850, Davaine and Eayer had noted the 
existence of little rod-like or filamentous bodies about the size of a 
blood corpuscle in the blood of a sheep that had died of splenic fever. 
Pollender had seen similar bodies in the blood of cows. Davaine 
did not at first pay much heed to this discovery; but in 1863 he 
thoroughly reinvestigated the subject, and conducted a series of 
experiments which led him to the conclusion that the actual cause 
of splenic fever was an organised being whose presence and 
multiplication in the blood produced changes in that fluid of the 
nature of fermentation, resulting in the death of the animal. 

These conclusions were not accepted by all, and indeed, the 
evidence was so far incomplete that sceptics were justified in con- 
sidering that these experiments afforded only a working hypothesis. 
But Davaine's comparison between this disease and fermentation 
attracted the attention of Pasteur, whose mind had been fully trained 
for entering upon this investigation by the researches which he had 
been carrying on in the interval between Davaine's publications of 
1857 and 1863. 

Pasteur, as already mentioned, had been working at fermentation, 
and his attention was next directed to studying the so-called diseases 
of wines, and subsequently to a contagious disease which committed 
ravages among silkworms. By laborious researches Pasteur was 
able to confirm the belief that this disease of silkworms was due to 
the presence of micro-organisms discernible with the aid of the micro- 
scope. These oval shining bodies in the moth, worm, and eggs had 
been previously observed by Cornalia, and described by Nageli as 
Nosema bombycis, and by Lebert as Panhistophyton. But it was 
reserved for Pasteur to introduce a means of combating the disease. 
Pasteur showed that when a silkworm, whose body contained these 
micro-organisms, was pounded up with water in a mortar, and the 
mixture painted with a. brush on the leaves on which healthy worms 
were fed, they would all without fail succumb to the disease. 

As the contagious particles were transmitted to the eggs, a 
method for preventing the spread of the disease suggested itself. 



8 BACTERIOLOGY. 

Each female moth was kept separate from the others, and allowed to 
deposit her eggs on a small Unen cloth. The moth was then pinned 
to the corner of the cloth, and left for future examination. When 
the time for this arrived, the moth was crushed up with water in a 
mortar, and a drop examined under the microscope. When any 
trace of corpuscular matter was found to be present, the cloth with 
its collection of eggs was burnt ; and if not, the eggs were set aside 
for use. 

Complete as this appears to be as a demonstration of a causal 
connection between the micro-organisms and the disease, it could 
obviously be objected that there was no distinct proof that the 
corpuscular bodies constituted the actual contagium. There was no 
isolation of the organisms, no artificial cultivation of them apart 
from the diseased moth or worm, and svibsequent production of the 
disease by means of the isolated organisms. The same objection 
was applicable to Davaine's investigations. Davaine found rods 
in association with anthrax, and maintained that they were 
causally related ; but others stated that it was possible to inoculate 
animals with anthrax blood containing rods, and to produce the 
disease without being able to detect the rods again in the blood 
of the animal experimented upon. It was also urged that it was 
possible to infect with anthrax blood after the rods had disappeared, 
and to find a reappearance of the bacilli in the blood of the 
inoculated animal. 

The well-known fact that anthrax was especially prevalent in 
certain seasons and certain localities appeared to lend great support 
to these objections. The disease, in fact, was regarded by some 
as originating from peculiar conditions of climate and soil. The 
fallacies in these objections were, however, rapidly dispelled. 
Bollinger, in 1872, pointed out that the blood, from which the rods 
had disappeared, was still virulent owing to the presence pf the 
spores of the bacillus, and that it was owing to the soil being impreg- 
nated with these spores that the disease broke out in certain 
localities. Yet there still remained many who refused to regard 
these particles as Uving bodies, some looking upon them simply as 
crystals ; and the question of their importance remained undecided 
for several years. 

In 1877 Robert Koch published a memoir in which he fully 
described the life-history of the anthrax or splenic fever baciQus, 
and gave a complete demonstration of the hfe-history of the micro- 
organism, and the definite proofs of its pathogenic properties. He 
pointed out how the rods grew in the blood and tissues by lengthen- 



HISTORICAL INTRODUCTION. 9 

ing and by cross division. Further, that in the blood or in serum 
or in aqueous humour they not only grew into long leptothrix 
filaments, but they produced enormous numbers of seeds or spores. 
He traced, by continuous observation on the warm stage, the whole 
life cycle, from the fission of the rods to the formation of spores and 
the sprouting of the spores into fresh rods. Further, he cax-ried 
on the disease by inoculating from mouse to mouse for several 
generations, and observed that in the blood of the animal and in 
the swollen spleen the glass-like rods were always to be found. 

Pasteur also studied the microbe of splenic fever, and amply 
confirmed and extended the observations of Koch by his researches 
on the attenuation of the anthrax virus. 

Pasteur also met with adverse criticism. Paul Bert argued 
that the bacilli were of no importance, because he could destroy 
them by exposing material containing them to great pressvire, and 
yet the material produced the disease on inoculation. But such 
measures did not destroy the spai'es; and finally, Paul Bert was 
convinced of his error when Pasteur demonstrated cultures of the 
anthrax bacillus in urine, from which successive generations were 
started, and that with such cultivations the disease could always be 
produced. 

It was, however, principally the researches of Koch which 
placed the doctrine of contagium vivum on a scientific basis. 

Koch's improvements in the methods of cultivation, his recom- 
mendation of the necessary microscopical apparatus, his histological 
methods for examining these minute organisms, and his famous 
postulates for proving beyond controversy the existence of specific 
pathogenic micro-organisms, elevated the theory of contagium vivum 
to a demonstrated and established fact. The chain of evidence 
regarded by Koch as essential for proving the existence of a 
pathogenic organism was as follows : — 

1. The micro-organism must be found in the blood, lymph, or 
diseased tissue of man or animal suffering from or dead of the 
disease. 

2. The micro-organisms must be isolated from the blood, lymph, 
or tissues, and cultivated in suitable media — i.e., outside the animal 
body. These pure cultivations must be carried on through successive 
generations of the organism. 

3. A pure cultivation thus obtained must, when introduced into 
the body of a healthy animal, produce the disease in question. 

4. In the inoculated animal the same micro-oi'ganism must 
again be found. 



10 BACTERIOLOGY. 

The chain of evidence is still more complete if we can from 
artificial cultures obtain a chemical substance which is capable 
of producing the disease independently of living micro-organisms. 

It is of very little value merely to detect or artificially to 
cultivate a bacterium associated with disease. We must endeavour 
to establish the exact relationship of the bacteria to disease processes, 
and the determination of the true pathogenic microbe is beset 
with fallacies. In many diseases bacteria have been regarded as 
the actual contagia, until a searching inquiry by other investigators 
has shown that the evidence was most unsatisfactory or entirely 
misleading. For example, in diseases with lesions of the external 
or internal linings of the body, extraneous micro-organisms may 
get into the circulation and be swept into the internal organs, 
where they either perish in the battle with the healthy tissues 
which are opposed to their existence, or they may gain the upper 
hand, and set up destructive processes. Such organisms, when 
found in association with these diseases, may be discovered in the 
blood and internal organs ; and though only accidental epiphytes, 
often associated with septic complication, they may too readily be 
accepted by the enthusiast as the actual contagium of the disease 
in question. 

It is only when such fallacies are exposed that we are brought 
■once more face to face with the fact that the nature of the contagium 
in hydrophobia, variola, vaccinia, scarlet fever, measles, and many 
other diseases, is still undetermined. 



CHAPTER II. 

MORPHOLOGY AND PHYSIOLOGY OF BACTERIA. 

Bacteria may be considered as minute vegetable cells destitute of 
nuclei. They are distinguished from animal cells by being able to 
derive their nitrogen from ammonia compounds, and they differ 
from the . higher vegetable cells in being unable to split up carbonic 
acid into its elements, owing to the absence of chlorophyll. Von 
Engelmann and Van Tieghem include among the bacteria certain 
organisms, named by them Bacterium chlorinum, Bacterium viride, 
and Bacillus virens, which are coloured green by this substance, but 
it is quite possible that they may be Algse, and further researches 
are required before any conclusions are definitely arrived at as to 
the exact place these particular organisms occupy in the vegetable 
kingdom. 

Composition. — For our knowledge of the chemical composition 
of bacteria we are chiefly indebted to Nencki. Their constituents 
are found on analysis to vary slightly, according to whether the 
bacteria are in zoolgoea or in the active state. In the latter condition 
they are said to consist of 83'42 per cent, of water. In one hundred 
parts of the dried constituents there are the following : — 

A nitrogenous body ..... 84:"20 

Fat . 6-04 

Ash 4-73 

Undetermined substances .... 5'04 

This nitrogenous body is called myco-protein, and consists of 

Carbon 52-32 

Hydrogen ....... 7'55 

Nitrogen 14-75 

but no sulphur or phosphorus. 

The nitrogenous body appears to vary in different species, for 
in Bacillus anthracis a substance has been obtained which does not 

11 ' 



12 BACTERIOLOGY. 

give the reactions of myco-protein, and, therefore, is distinguished 
as anthrax-protein. 

Considering bacteria as cells, we may speak of the cell-wall and 
the cell-contents. The cell-wall consists of cellulose, oi-, according 
to Nencki, in the putrefactive bacteria of myco-protein. It may be 
demonstrated by the action of iodine, which contracts the proto- 
plasmic contents, and renders the cell-wall visible. By staining 
cover-glass preparations of the anthrax bacillus by the method of 
Gram, the rods are at first uniformly stained, by subjecting them to 
iodine solution the protoplasmic contents are contracted, and alcohol 
decolorises the sheath, which may be then stained in contrast, with 
eosin. 

The cell-wall may be either pliable or rigid. Phability is 
observed in the long filaments, which are endowed with a slow 
vermicular movement, while rigidity accounts for the maintenance 
of the characteristic form of several species, such as spirilla. 

The cell-protoplasm yields myco-protein. In some it is homogene- 
ous, and in others granular. The action of the aniline dyes indicates 
a close relation to nuclear protoplasm, though all nuclear stains are 
not suitable for bacteria. In some cases also the bacteria remain 
stained under the influence of a reagent, which removes the colour 
from nuclei. The power of fixing the stain is not always present, 
and indicates a difference in the protoplasm of different species. 
Thus in staining phthisical sputum, the nitric acid removes the 
stain from all bacteria and bacilli present, with the exception of the 
tubercle bacillus. This difference in the protoplasm of different 
species is also illustrated by the necessity, in many cases, of using 
special processes, owing to the ordinary methods being unsatisfactory 
or not producing any result. 

The protoplasm of some bacteria contains starch granules ; thus 
Clostridium butyricum gives the starch reaction with iodine. 
Sulphur granules are present in some species of Beggiatoa which 
thrive in sulphur springs. The colouring-matter of the pigmented 
bacteria is probably external to the cell as a rule : for example, in 
Micrococcus prodigiosus the pigment granules are distictly between 
the ceUs ; on the pther hand, in Beggiatoa roseo-persicina, or the 
peach-coloured bacterium, the special pigment hacterio-pu/rpwrin 
appears to be dissolved in the cell protoplasm. In Bacillus 
pyocyaneus the pigment is certainly not locahsed entirely in the 
cell, for it becomes rapidly diffused in the surrounding medium, 
considerably beyond the confines of the growth itself. 

In several species, either as a result of a secretion from the cell or 



MORPHOLOGY AND PHYSIOLOGY OF BACTERIA. 13 

of the absorption of moisture and consequent swelling of the outer 
layer of the cell-wall, a mucinous or gelatinous envelope develops 
around them. This envelope may form a capsule, such as we meet 
with in certain bacteria found in the rusty sputum of pneumonia, and 
in Micrococcus tetragenus ; or it may occur as a continuous sheath 
around a chain of bacteria, which by its disappearance sets free 
the individual links. ' The capsule is soluble in water, and under 
some circumstances is difficult to demonstrate. In the pneumo- 
cocous of Friedlander the capsule disappears on cultivation, but 
reappears in preparations made from an inoculated animal. In the 
pleuritic fluid of a mouse these cocci are often found with a parti- 
cularly well-marked capsule, and in other encapsuled cocci the extent 
of the envelope has been observed to vary considerably in the same 
species of bacterium. 

When this gelatinous material forms a matrix, in which 
numbers of bacteria are congregated in an irregular mass, we have 
what is termed a zooglcea. The zoogloean stage is a resting stage, 
often preceded or followed by a motile stage. Thus bacteria may 
be present in a solution in an active state, and after a time a scum 
or pellicle forms on the surface of the liquid, which consists of 
zooglcea. At the edges of the zoogloea, individuals may be seen to 
again become motile, and after detaching themselves to swim oflf in 
the surrounding fluid. 

The zooglcEan stage may be observed sometimes in cultivations in 
broth, and also in nutrient gelatine which has become liqviefied. 
The inoculated bacteria grow and multiply, and after a time a film 
appears on the surface of the liquefied layer. In cultivations on 
potato the appearances in this stage are varied, and sometimes 
extremely characteristic. In the case of a bacillus which readily 
develops on unsterilised potatoes, the zoogloea may spread over the 
cut surface, forming a pellicle which can be raised en masse like 
a delicate veil. Another bacillus forms a zooglcea, consisting of a 
tenacious layer which can be drawn out in long stringy threads. 
In Ascococcus Billrothii the gelatinous envelope develops to such 
an enormous extent that it forms the characteristic feature of the 
species. (Fig. 1.) 

Form. — The individual cells vary in form, and may either 
remain isolated or attached to each other. Round cells and egg- 
shaped cells are called cooci. The spherical form is the most 
common, but cocci are occasionally exclusively ovoid, as in Strepto- 
coccus bombycis. The giant cocci of some species are spoken of as 
megacocci, to distinguish them from the ordinary cocci, or micrococci. 



14 BACTERIOLOGY. 

The fission by which the cooci increase may take place in one 
chrection only, and if the two resulting cells remain attached to 
each other they form a dijdococcm. If these two cells again dhide, 
and the resulting cells remain hnked together, we get a chain or 
rosary, termed streptococcus. These chains may consist of a few 
individuals linked together, or of several hundreds, in which case 
the chains are generally curved or twisted. When the division 
occurs in two directions, so that four cocci result, a tetrad or 
mensmopedia is formed ; when in three directions, one coccus divides 
into eight, and the result is a packet form or sarcinacoccus. 
Immediately after division, the daughter cells are not perfectly 
circular, but are flattened or facetted where they are opposite to 




Fig. 1. — Ascococous Billkothh, x 65. [After Cohn.] 

each other. They gradually become rounded off, and each daughter 
cell is then ready to divide in its turn. In other cases the cocci 
after division only form irregular heaps or collections like bunches 
of grapes. This form is sometimes distinguished as staphylococcus, 
but it cannot be considered an important feature. When we find 
irregular masses of cocci united by intercellular svibstance and 
embedded in a tough gelatinous matrix, the form is described as 
ascococcus. 

Another type is the rod, characteristic of hacteriam and bacillus. 
The rods may vary considerably in length. The very short rods 
with rounded ends are difficult to distinguish from the oval cocci, 
but differ in that a rod, however short it may be, must have 
two sides parallel. The vibrio or bent rod may be considered as 
the connecting link between the rods and the corkscrew forms or 



DESCRIPTION OF PLAT^ I. 
Bacteria, Schizomycetes, or Fission Fungi. 

1. Cocci singly and varying in size. 2. Cocci in chains or rosaries (strepto- 
coccus). 3. Cocci in a mass (staphylococcus). 4 and 5. Cocci in pairs 
(diplococcus). 6. Cocci in groups of four (merismopedia). 7. Cocci in packets 
(sarcina). 8. Bactenum termo. 9. JBacierium termo x 4000 (Dallinger and 
Drysdale). 10. Bacterium septicceni'us hcemorrTiagioo!. 11. Baeterium pneu- 
vumice crouposce. 12. Bacillus suiiilis. 13. Bacillus murisepticus. 14. 
Bacillus diplttherite. 15. Bacillus typlwsus (Bberth). 16. Spirilhmi wiidvla 
(Cohn). 17. Spirillum volutans (Cohn). 18. Spirillum cholerte Asiaticte. 
19. Spirillum Obervieieri (Koch). 20. SpirocJiceta plicatilis (Fliigge). 21. 
Vibrio rugula (Prazmowski). 22. CladotJtnx Forsteri (Cohn). 23. Cladothrix 
dichotona (Cohn). 24. Manas OTtenii (Cohn). 25. Monas WarmingiA (Cohn). 
26. BJidbdomonas rosea (Cohn). 27. Spore-formation (^Bacillus alvci). 28. 
Spore-formation (Bacillus antltracis). 29. Spore-formation in bacilli cultivated 
from a rotten melon (Frankel and Pfeiffier). 30. Spore-formation in bacilli 
cultivated from earth (Frankel and PfeifEer). 31. Involution-form of Crenbtli/rias 
(Zopf). 32. Involution-forms of Vibris serpens (Warming). 33. Involution- 
forms of Vibrio rugula (Warming). 34. Involution-forms of Clostridium 
poLymyxa (after Prazmowski). 35. Involution-forms of Spirillum choleree 
Aslaticce. 36. Involution-forms of Bacterium aceti (Zopf and Hansen). 
37. Spirulina-form of Beggiatoa alba (Zopf). 38. Various thread-forms, of 
Bacterium merismopedioides (Zopf). 39. False-branching of Cladothrix (Zopf). 



J''3 1. Fig. «. 






f IJ * 



.2'7y .^ 









%■ 






' Fig « 






1^ 



^ 






rf*' 



Fig. II jr,g Ig, 






Fig IS 








Fuf Ze J^iff- ZJ 




Fh "-f 



Tic). ?/.' 






'^~ 



.-^1^^3-2- 



,t> 



-"ooo, 



'o 






Fif, •?.'<. 




/•■/>. .5/ 7p .;r 



Fly JJ. 



V 






ll\CTEliTA, S("'.F[[ZOMYaLTJ-:S, OR PJSSION l-fMOI. 



MORPHOLOGY AND PHYSIOLOGY OF BACTERIA. 15 

spirilla. Lastly, we have tlie filamentous forms, which may be 
straight, leptothrix, or wavy, spirochceta, or the wavy thread may 
be looped and entwined on itself, spirulina. 

The term involution form is applied to certain peculiar shapes, 
which result more especially in bacteria grown under abnormal 
conditions. They are round, oval, pear-shaped, or club-formed 
enlargements. 

Movement — Many bacteria are devoid of movement through- 
out the whole of their life history. Others, during certain stages of 
their life cycle, and possibly some forms always, are endowed with 
locomotive power. The character of the movement is very varied, 
and ranges from a slow undulatory motion to one of extreme 
rapidity. Many appear to progress in a definite direction. Others 
move continuously, first in one direction and then in another, and 
others again seem to hesitate before altering their course. They 
may either glide along smoothly or progress with a tremulous motion. 




FiK. 2. — Spikoch^ta feom Sewage Water, x 1200. 

They appear to be able to avoid obstacles, and to set themselves 
free from objects with which they have accidentally come into 
contact. Vibrios have a pecuUar serpentine movement, but other 
forms, such as the commonly known Bacterium termo and segments 
of spirilla, such as comma-bacilli, revolve around their long axis 
as well as make distinct progression. The complete spirilla are 
characterised by the famihar corkscrew movement. With regard 
to cocci there is soine doubt as to whether they are endowed with 
independent movement, any quivering or oscillation being generally 
regarded as only Brownian or molecular. In some straight thread- 
forms, which are motile, the movement is very slow and vermicular 
in character, but in wavy threads, such as the Spirochseta plicatilis, 
there is not only an undulatory motion, with rapid progression across 
the field of the microscope, but if they are confined by more or 
less debris, they give very pecuUar and characteristic spasmodic 
movements. (Fig. 2.) 

The rod-forms of Proteus vulgaris exhibit very extraordinary 



16 



BACTERIOLOGY. 



movements on the surface of solid nutrient gelatine. Groups of 
rods may be observed to pass each other in opposite directions. 
Single individuals meet and progress side by side, or one or more 
individuals may part from a group and glide away independently. 
Occasionally a number of rods progress in single file. It is, however, 
difficult to believe that these movements can occur on a solid surface. 




Fig. 3. — Flagella. 

1. Coccus with flagellum. 2. Similar coccus dividing, with two flagella. 3. Colony 
of flagellated macrococci of Beggiatoa roseo-persicina. 4. Short rod from the 
same Beggiatoa with flagella [all after Zopfj. 6. Bacillus with flagella [from 
a photograph by Koch]. 6. Bacillus subtilis [after Brefeld]. 7, 8. Short rod- 
forms of Beggiatoa roseo-persicina with one flagellum [after Zopf]. 9. Very 
long rod of the same, with flagellum at both ends [after Warming]. 10. Vibrio, 
with double flagellum at each end [after Warming]. 11. Vibrio, with flagella 
[from a photograph by the author]. 12. Spirillum with flagella [from a photo- 
graph by Koch]. 13. Spirillum with flagella [after Zopf]. 14. Spirillum with 
double flagella [after Zopf]. 15. Beggiatoa roseo-persicina, with a triple 
flagellum at one end ; and 16, with a double flagellum at both ends [after 
Warming]. 



The author is inclined to beheve that there is an almost inappreciable 
layer of liquid on the surface of the gelatine, which is expressed 
after the gelatine sets. In tubes of nutrient agar-agar gelatinised 
obliquely and then kept upright the liquid so expressed collects at 
the bottom of the sloping surface. 



MORPHOLOGY AND PHYSIOLOGY OF BACTERIA. 



17 



The means by which bacteria are endowed with the power of 
spontaneous movement and of progression may still be said, in 
some cases, to be unsettled. The author has watched the move- 
ment of long slender threads in sewage-contaminated water, which 
could only be explained by the inherent contractility of the proto- 
plasmic contents ; for if any drawing or propelKng organ existed 
in proportion to the length of the organism, it would probably have 
been visible. But in many cases the organism is provided with a 
vibratile lash or flagellv/m at one end, or with one or more at both 
ends, or with numerous lateral and terminal flagella. 

Some observers believe that the movement of cooci is due to the 




Fig. 4. — Bacillus megathehium. 

a. A chain of rods, x 250. The rest x 600. 
6. Two active rods. , 

A to /. Successive stages of spore-formation. 
h to m. Successive stages of germination. 

[After De Bary.] 

existence of a flagellum. In Bacterium termo the existence of a 
lash at either end was first determined by the researches of Dallinger 
and Drysdale. In motile bacilli, such as the hay bacillus and 
Bacillus ulna, and in vibrios and spirilla, the flagella can be readily 
recognised by expert microscopists with the employment of the best 
lenses, and, what is of equal importance, proper illumination. They 
are objects of extreme delicacy and tenuity, and in stained prepara- 
tions may be absent from retraction or injury. Koch succeeded 
in photographing them after staining with logwood, which turned 
them a brown colour. The author has observed them in vibrios in 
preparations stained with gentian violet, from which also they have 
been photographed", in spite of the violet colour, by the use of 



18 



BACTERIOLOGY. 



isochromatic dry plates, and more recently special methods have 
been introduced, by Lbffler and others, by which they can be stained 
and photographed with comparative facility. 

It is not certain whether the flagella are extensions of the cell- 
wall, or derived from the internal protoplasm. Van Tieghem holds 
the first view, and does not regard them as motile organs at all. 
Zopf , on the other hand, adheres to the second view, and moreover 
believes that they can be retracted within the cell-wall. 

Eeproduction.— Bacteria multiply by fission and by processes 
which may be considered as representing fructification. The 




^o ^o ^^ 



Fig. 5. — Clostbidium butybioum, x 1020. 

B. Stages of spore-formation. 

C. Stages of germination. 

■[After Prazmowski.] 



bacteria exhibiting the latter processes have been divided into two 
groups, distinguished by the formation of endospores in the one and 
of arth/rospoi'es in the other. In the process of fission the cell first 
increases in size, and a transverse septum forms from the cell-wall, 
dividing the internal protoplasm into two equal parts ; these may 
separate and lead an independent existence, or remain linked 
together. In chains of cocci the individual cells are easily visible 
and distinct, but in the thread-forms resulting from the linking 
together of rods, as in the anthrax bacillus, the composition of the 
thread is only demonstrated by the action of reagents. 

Endospore formation may be conveniently studied in Bacillus 
anthracis. Bacillus megatherium, or Bacillus subtilis. The proto- 



MORPHOLOGY AND PHYSIOLOGY OF BACTERIA. 19 

plasm becomes granular, and at certain points in the thread a speck 
appears, which gradually enlarges and develops into a circular 
or egg-shaped, sharply defined, highly refractive body. The spore 
grows at the expense of the protopJasm of the cell, which in time, 
together with the cell-wall, entirely disappears, and the spore is set 
free. These phenomena are best seen in an immotile bacillus in a 
■drop-cultivation on a warm stage ; the whole process may then be 
observed continuously from beginning to end. Spores may form in 
each link of the thread, so that a regular row results, or they may 
occur at irregular intervals. Spore-formation also occurs in bacilli 
which do not develop into leptothrix filaments. The spores may 
develop in the centre or at one end of the rod. In the tetanus 
bacillus a spore develops at the extreme end, producing the appear- 
ance of a drum-stick. The spore may be considerably wider, but 
is never longer than the parent cell. 




Fig. 6. — Lboconostoo mesenteboidbs ; Cocoi-ohains with Akthkospokes 
(after Van Tieghem and Oienkowski). 

Arthrospore formation is illustrated in Leuconostoc mesenteroides. 
■Certain elements in the chain of cocci, apparently not differing from 
the rest, become larger, with tougher walls, and more refractive. 
The remaining cells die, and these cells having acquired the pro- 
perties of spores are set free, and can reproduce a new growth in 
any fresh nourishing soU. That this occurs in all species which 
do not form endospores is at present only a supposition. 

Spores are invested by a thick membrane, which is beHeved to 
consist of two layers. To this they probably owe the property they 
possess of retaining vitality when desiccated, and of offering a, 
greater resistance to the action of chemical reagents and heat than 
the parent cells. > 

Spore-formation has been regarded by some as occurring when 
the nourishing soil is exhausted, thus providing for the perpetuation 



20 BACTERIOLOGY. 

of the species. In antKrax the bacilli do not form spores in the 
living body, but when the animal dies the development of spores 
takes place, and hence the danger of contaminating the soil if the 
body is disposed of by burial. Klein, however, has pointed out that 
if mice and guinea-pigs which have died of anthrax are kept un- 
opened, the bacilli simply degenerate and ultimately disappear. 
Thus there is good reason for believing that spore-formation is 
not due to exhaustion of the pabulum, but probably free access to 
oxygen constitutes an important factor in inducing this condition. 
If we inoculate a potato with anthrax, copious spore-formation 
occurs, though we cannot consider that the nourishing soil has been 
exhausted. But we have in this case the surface of the potato 
freely exposed to the air in the damp chamber. In the same way, 
in cultivations on agar-agar solidified obliquely (so as to get a large 
surface), spore-formation readily takes place. Contamination of 






Fiu. 7. — Spobe-beabing Theeads of Bacillus Antheacis, Double-stainbd- 

WITH FUOHSINE AND METHYLENE BlUE, X 1200. 

the ground results, therefore, from animals in which a post-mortem^ 
examination has been made and the blood and organs freely exposed 
to the air ; or from carcasses the hides of which have been soiled 
with excretions, and with blood which issues from the mouth and 
nostrils before death. 

When spores are introduced into a suitable medium at a favour- 
able temperature they develop again into rods. The spore loses its 
sharp contour, and, at one pole or on one side, a pale prdcess bursts 
through the membrane, gradually growing into a rod from which 
the empty capsule is thrown off. 

Spores differ from the parent cells in their behaviour to staining 
reagents. Like them, they can be stained with aniline dyes but 
not by the ordinary processes. They require to be specially treated. 
This is probably due to the tough capsule, which must be altered 
or softened by heat or strong acid, until it allows the stain to 
penetrate. 



MORPHOLOGY AND PHYSIOLOGY OF BACTERIA. 21 

Once stained, they again differ from the parent cells in resisting 
decolorisation ; this fact is taken advantage of to double-stain spore- 
bearing bacilli (Fig. 7). 

In staining micro-organisms, the protoplasm is sometimes broken 
lip into irregular segments or granules, as in many spirilla, and we 
may add the bacilli of tuberculosis and leprosy. The beaded 
appearance of the tubercle bacillus is well known. Some observers 
have regarded the beads, others the bright spaces between them, 
as spores. But spores in unstained preparations appear as glistening 
bodies with sharp contour. They do not stain at all, or very little, 
by the ordinary processes. These considerations led the author to 
Stain and examine tubercular sputum and pure-cultures under 
careful illumination, and with such lenses as Powell and Lealand's 
•jL in. hom. imm. The tubercle bacillus in sputum (Fig. 8), as a rule. 




N 



Fig. 8. — Bacilli oi' Tubekole in Sputum, x 2500 (from photographs). 

consists of a very delicate sheath, holding together a number of 
deeply stained granules, for the most part round or cylindrical, with 
irregular contour, and differing considerably in size, while the light 
interspaces are seen to vary in form according to the shape of the 
granules. On the other hand, particularly in old cultures, more or 
less spherical, sharply defined bodies are observed in the bacilK, and 
also set free. These are the true spores of the tubercle bacillus, 
and are quite distinct from the irregular granules. There can be 
no doubt that a tubercle bacillus consists of a very delicate sheath, 
with protoplasmic contents which have a great tendency to break 
up or coagulate into little segments or roundish granules, partly 
owing to their age and the conditions under which they are grown, 
and partly to the treatment they are subjected to in making a 
microscopical preparation. This does not always occur, for the 
bacilli at times are not beaded, but are stained in their entirety. 
In the leprosy bacilli a similar appearance occurs. In stained 



22 BACTERIOLOGY. 

sections the rods have a beaded appearance, but the intervals between 
the granules are sometimes very long, and occasionally the protoplasm 
appears to have collected only at the extreme ends of the rod. 

The appearances in the case of the bacillus of glanders and the 
bacillus of hsemorrhagic septicaemia may be similarly explained. 

The fact that tubercular sputum preserves its virulence for 
several months, even after desiccation, is to be attributed to the 
formation of spores. Babes claims to have succeeded in differen- 
tiating them by double staining. 

In his definition of spirilla, Zopf gives the spore-formation as 
absent or unknown. In comma-bacilli in sewage water the author 
has often noted appearances suggestive of refractive spores ; and 
the same also may be observed in vibrios, differing by their regular 
contour from the irregular spaces occasionally observed in stained 
preparations ; but they are only vacuoles. 






Fig. 9.— Comma-bacilli in Sew- Fig. 10. Vibmos in Water con- 

age Water, stained with taminated with Sewage, 

Gentian Violet, x 1200. x 1200. 

Respiration and Nutrition — Like all a-chlorophyllous vegeta- 
bles, bacteria require for their nutrition oxygen, nitrogen, carbon, 
water, and certain mineral salts. Many require free access to oxygen, 
others can derive it from the oxidised compounds in the medium 
in which they grow. Pasteur divided bacteria into two great classes 
—the aerobic and anaerobic, and considered that the latter net 
only had no need of oxygen, but that its presence was actuaUy 
deleterious. Though this view must be considerably modified, the 
terms are convenient, and are stUl retained. They are well illus- 
trated by the bacillus of anthrax, and the bacillus of malignant 
oedema ; and a simple plan of demonstration has been employed 
by the author. A fragment of tissue from the spleen, for example, 
kriown to contain anthrax bacilli, is deposited with a sterihsed 
inoculating needle, with the necessary precautions, on the surface of 
nutrient agar-agar in a test-tube ; another tube of nutrient agar- 
agar is Hquefied, and when cooled down almost to the point of 




MORPHOLOGY AND PHYSIOLOGY OF BACTERIA. 23 

gelatinisation, a part is poured into the first tube, so that when it 
sets the piece of tissue is completely embedded. A piece of tissue 
from an animal suffering from maKgnant oedema is treated in the 
same way, and the tubes are placed in the incubator. If we 
examine them after two or three days, we shall find no change in 
the anthrax tube ; the bacillus being eminently aerobic, no growth 
whatever has occurred. In the tube containing the bacilli of 
malignant oedema there will be a more or less characteristic 
cultivation. 

The nitrogen which is essential for building up their protoplasm 
can be obtained either from albumins, or from ammonia and its 
derivatives. That the albumins can be dispensed with was shown 
by Pasteur, who employed an artificial nourishing solution consti- 
tuted upon a formula representing the essential food constituents. 

Carbon is derived from such substances as cane sugar, milk 
sugar, and glycerine, and, in some cases, by the splitting up of 
complex proteid bodies. 

Water is essential for their growth, but deprivation of water 
does not kill all bacteria. Desiccation on potato is employed for 
preserving some micro-organisms, as a new growth can be started, 
when required, by transferring some of the dried potato to fresh 
nourishing ground. Comma-bacilli, on the other hand, are 
destroyed by drying. Sugar is used in making preserves, because 
by abstracting water it prevents the development of micro- 
organisms. 

Mineral or inorganic substances, such as compounds of sodium 
and potassium, and different phosphates and sulphates, are necessary 
in small propoi'tions. 

Circumstances affecting the Growth of Bacteria. 

Mature of tlie Soil. — Though we know the elements necessary, 
we are, nevertheless, as yet unable to provide a pabulum suitable for 
all kinds of bacteria. Thus we are quite unable to cultivate some 
species artificially. Others will only grow upon special media. 
Many grow vipon nutrient gelatine ; but some species only if it be 
acid or alkaline respectively. Whether in the latter case this is due 
purely to the reaction or to the presence of the particular ingredients 
is an unsettled point. Though the comma-bacillus of Koch, like the 
majority of organisms, grows best on an alkaline medium, yet it 
is well known to flourish at the temperature of the blood on the 
surface of potato, which is acid. 



24 BACTERIOLOGY. 

Temperatti/re.—'Ihe influence of temperature on bacteria will be 
found to vary according to the species, but still for the majority we 
may distinguish a maximum, optimwm, and minimwm temperature. 

Many grow best at the temperature of the blood, and hence the 
value of nutrient agar-agar, which is not liquefied at 37° 0. The 
tubercle bacillus will only grow satisfactorily at a temperature 
varying between 30° 0. and 41° C. On the other hand, many forms 
grow between the limits of 5° 0. and 45° C. At these temperatures 
their functional activity is paralysed, but they are not destroyed, 
for by removal to favourable conditions they spring again into Hfe. 
Bacteria seem to have a special power of resisting the effects of cold. 
It has been stated that comma-bacilli exposed to a temperature 
of— 10° C. for an hour, and bacilli of anthrax after exposure to a 
temperature of — 110° C, still retained their vitality. Temperatures 
over 50° to 60° C. destroy most bacteria, but not their spores ; spore.s 
of anthrax retain their vitality after immersion in boiling water; but 
are destroyed by prolonged boiling. Roughly speaking, all patho- 
genic bacteria grow best at the temperature of the blood, and 
non-pathogenic bacteria at the ordinary temperature of the room. 

Movement. — Bacteria probably grow best when left undisturbed. 
Yiolent agitation of a, vessel in which they are growing certainly 
retards their growth, but a steady movement is stated not to affect 
it ; at any rate, anthrax bacilli grow with enormous rapidity in the 
blood-vessels, in spite of the circulation. 

Compressed Air. — Paul Bert maintained that a pressure of 
twenty-three to twenty-four atmospheres stopped all development 
of putrefactive bacteria. Oxygen, under a pressure of five or six 
atmospheres, is stated to stop their growth. Other observers have, 
however, obtained different results. 

<9ases.— Hydrogen and carbonic acid are stated to stop the 
movements of the motile bacteria. Chloroform is beheved to arrest 
the changes brought about by the zymogenic species. 

-Electricity.— Gohn and Mendelssohn found that a constant 
galvanic current produced a deleterious effect owing to electrolysis. 
At the positive pole the Uquid became distinctly acid, and at the 
negative pole distinctly alkaline. With a weak current there 
appeared to be no effect, two powerful cells at the very least being 
necessary. 

LighL-^Bownes has shown that sunlight is fatal to putrefactive 
bacteria. This is believed to be due to a process of induced hyper- 
oxidation, from which living organisms ordinarily are shielded by 
protective developments of the cell-wall, or of colouring-matter 



MORPHOLOGY AND PHYSIOLOGY OF BACTERIA. 25 

-which cut off injurious rays. Duclaux has investigated the same 
subject, and observed that micrococci were more sensitive to sun- 
light than the spore-bearing bacilli. Engelmann has described a 
bacterium whose movements cease in the dark, and Zopf states that 
in his cultures of Beggiatoa roseo-persicina the growth was much 
more strongly developed on the side of the vessel facing the light. 
Arloing, Marshall Ward, and Dieudonne have studied the effect 
of the sun's rays on anthrax spores, and on chromogenic and 
other bacteria, and maintain that they are bactericidal. The 
effect is due chiefly, if not entirely, to the blue rays.- 

Chemiccd ReUgents. — Many substances, such as carbolic acid, 
■corrosive sublimate, chlorine, bromine, have a marked effect upon 
the growth of bacteria. This will be more fully described in 
another chapter. In several cases the bacteria themselves secrete 
a substance which is injurious to their future development. 

, Phoducts of Geowth. 

Bacteria may be grouped together according to the changes pro- 
duced in the media in which they grow. Thus wo have pigment- 
forming, phosphorescent, fermentative, putrefactive, nitrifying, and 
disease-producing bacteria. 

GhroTnogenic or pigment- forming bacteria elaborate during their 
growth definite colour stuffs. Such species are exemplified by Bacillus 
violaceus, which produces a striking purple growth ; Bacillus 
pyocyaneus, which secretes pyocyanin, a substance which has been 
isolated and obtained in a crystalline form ; Micrococcus prodigiosus, 
which produces a pigment allied to fuchsine ; Beggiatoa roseo-per- 
sicina, which is characterised by the presence of bacterio-purpurin ; 
Sarcina lutea. Bacillus cyanogenus, and many others. 

Photogenic, or light-producing, bacteria are found more especially 
in sea-water. There are several species of phosphorescent bacUli, 
and according to Beyrinck the best medium for their cultivation is 
fish-broth made with sea-water. Photographs can be obtained of 
cultures by their own light. 

Zymogenic or ferment bacteria produce their changes in non- 
nitrogenised media. Bacterium aceti, by its growth produces the 
acetic fermentation in wine by which alcohol taking up atmospheric 
oxygen is converted into vinegar : — 

C^ffO + 0^ = C^H^G^ + H^O. 

The fermentation of urine, by which urea is converted into carbonate 
of ammonia, can be brought about by several micro-organisms, but 



26 BACTERIOLOGY. 

notably by the Bacterium urese. The change produced is represented 
by the following formula : — 

CoH|^ + 2ffO = (NH*) 2C0». 

Clostridium butyricum converts the salts of lactic acid into' 
butyric acid, producing the butyric fermentation in solutions of 
starch, dextrine, and sugar. These bacteria are agents in the 
ripening of cheese, and the production of sauerkraut. Thus, in a 
solution neutralised with calcium carbonate : — 

2[Ca(C»H=0«y] + H'^O = CaCC'H'O')'' + CaCO» + 3C0= + ff. 

In the so-called viscous fermentation of wines. Streptococcus viscosus- 
produces a gummy substance. According to Pasteur, the change 
may be thus represented : — 

2.5(C"H'=0") + 25(H'=0) = 12(C"'H»0'») + 24(C''H"0'') + 
12(00'') + 12(H'=0). 

And as another example, the Bacillus acidi lactici may be mentioned, 
through the agency of which sugar of milk is converted into lactic 
acid : — 

cm^O^' _ 4(C3H«0''). 

Saprogenic or putrefactive bacteria play a most important part 
in the economy of nature. They produce changes allied to fermenta- 
tion in complex organic substances. Their action on proteids, 
according to Hoppe-Seyler, may be compared to digestion ; bodies 
like peptones are first produced, then leucin, tyrosin, and fatty 
acids ; lastly indol, phenol, sulphuretted hydrogen, ammonia, carbonic 
acid, and water. They abstract the elements they require, and the 
remainder enter into new combinations. Associated with the forma- 
tion of these substances are certain bodies which have a poisonous 
effect when introduced into animals. These poisonous alkaloids, 
ptomaines, produce a septic poisoning, which must be distinguished 
from septic infection. The effects of septic poisoning depend on the 
dose, whereas the effects of septic infection are, to a certain extent, 
independent of the dose. A small quantity of a septic poison may 
produce only transient effects, and a relatively large quantity may 
be necessary to produce vomiting, rigors, and death. Septic in- 
fection, on the other hand, may result equally from a small dose, 
because the poison introduced is a living organism which is capable 
of propagation and multiplication. Our knowledge of these 
alkaloids is largely attributable to the researches of Selmi, Gautier 
and Brieger, and the result of their work will he referred to again. 



MORPHOLOGY AND PHYSIOLOGY OF BACTERIA. 27 

NiVrifying bacteria play a very important part by providing 
plant life with a most necessary food. They occur in the soil, and 
two kinds have been described — the" one kind converting ammonia 
into nitrous acid, and the other changing nitrous into nitric acid. 
To Winogradsky and Frankland we are principally indebted for our 
knowledge of these bacteria. 

Pathogenic bacteria are those which are genetically related to 
disease. Many organisms have been supposed to be pathogenic, or 
have been described in connection with diseases, which are only 
saprophytic associates. By saprophytic we mean organisms which 
feed upon dead organic matter. They include many forms which 
are found on the skin, in the intestinal canal, and sometimes in the 
internal organs, especially the Uver and kidneys ; the tissues have 
lost their vitality, and the organisms, through some lesion, have 
been carried into the circulation. 

That many organisms are causally related to disease, there is 
strong evidence in proof. No organism can be considered to be pro- 
ductive of disease unless it fulfils the conditions which have been 
laid down by Koch. Great stress must be laid upon the importance 
of successive cultivation through many generations, as the objection 
that a chemical virus may be carried over from the original source 
is thus overcome. Any hypothetical chemical poison carried over 
from one tube to another would, after a great number of such 
cultivations, be diluted to such an extent as to be inappreciable 
and absolutely inert. 

Though we may accept as a fact the existence of pathogenic 
organisms, we are not in all cases in a position to assert the means 
by which they produce their deleterious or fatal effects. Many 
theories have been propounded. It has been suggested that the 
pathogenic organisms may be compared to an invading army. 
The cells or phagocytes arrayed against them endeavour to as- 
similate and destroy them, but perish themselves in the attempt. 
This might explain the breaking down of tissue, and the for- 
mation of local lesions, but does not assist us in understanding 
the fatal result in thirty-six to forty-eight hours produced by the 
inoculation of the bacilh of anthrax. Another view is that the 
invading army seizes upon the commissariat, appropriating the 
general pabulum, which is so essential to the life of the tissues. 
This would hardly account for so acute and fatal a result as in 
anthrax, but would lead one to expect symptoms of inanition and 
gi-adual exhaustion. Moreover, against this theory we have the 
fact that death may result, in some cases, with the presence 



28 BACTERIOLOGY. 

of comparatively few bacilli in the blood; and, again, the 
blood may teem with parasites such as the flagellated monads in 
well-nourished, healthy-looking rats, without, apparently causing 
a,ny symptoms whatever. In the same category may be placed the 
theory that eminently aerobic organisms seize upon the oxygen of 
the blood and produce death by asphyxia. Another explanation is 
afforded by the suggestion of interference with the functions of the 
lung and kidney by mechanical blocking of the capillaries. Here 
the same objection is met with in the case of anthrax, the same 
fatal result may occur with only a few bacilh, while other cases 
yield very beautiful sections, looking like injected preparations from 
the mapping out of the capillaries with the countless crowds of 
bacilli. 

Putrefactive bacteria derive their necessary elements from com- 
plex organic substances, and accompanying the residue we find the 
presence of poisonous substances. Pathogenic bacteria, in a similar 
way, give rise to virulent poisons. Anthrax bacilli produce poisonous 
principles in the blood which cause death, independently of the 
number of baoilH, provided there are suflficient present to develop a 
fatal dose. 

It has been also suggested that possibly a special ferment is 
secreted by some organisms, and that by the changes ultimately 
wrought by the action of this ferment the symptoms and phe- 
nomena of disease arise. We have an analogy with this theory 
in the alkaline fermentation of urine by means of the torula urese. 
By the researches of Musculus, and later of Sheridan Lea, it has 
been shown that a ferment is secreted by the cells which can be 
isolated in aqueous solution, and is capable of rapidly inducing an 
active fermentation of urea. 

We can now understand how it is that in anthrax or in tuber- 
culosis we may find the presence of only a few bacilli, or that in 
tetanus we can have such a violent disturbance of the system 
produced by the presence of very few micro-organisms. We may 
conceive that different species of bacilli may vary greatly in their 
power of producing a toxin or secreting a. ferment, just as the 
elaboration of pigment is much more marked in some species than 
in others ; thus it need not follow that the number of micro- 
organisms bears any relation to the virulence or activity of the 
substance they produce. There is, however, yet another factor in 
the production of disease. We know that in health we are proof 
against most of these micro-organisms ; if it were not so, we should 
all rapidly fall victims to the tubercle bacillus or others, which in 



MORPHOLOGY AND PHYSIOLOGY OF BACTERIA. 29 

health we inhale with impimity. We know that a microbe may only 
cause a local lesion in one animal, but death in another. It is still 
more striking that the same micro-organism, as is the case with 
anthrax, may have no effect whatever upon certain species of 
animals, though it is deadly to others. Again, an animal natiirally 
susceptible to the effect of a pathogenic organism may be rendered 
proof against it. These matters will be discussed in a future, 
chapter. 

Distribution of Bacteria. 

Bacteria are commonly described as ubiquitous. They are ever 
present in the air, though not in such exaggerated numbers as is 
commonly supposed. In nutrient media exposed to the air one is often 
astonished at times at the comparativelji few bacteria which develop 
in comparison to the amiount of floating matter,, such as mineral 
particles, scales, spores of fungi and debris known to be present. 
In water they are also present in considerable numbers, though of 
course varying according to the character of the water. Wherever 
there is putrefaction, they are present in vast numbers. In the soil, 
in sewage, in the intestines and, in uncleanly persons especially, 
on the skin and between the teeth, various species may always be 
found, but in the healthy blood and healthy tissues bacteria are 
never present. In a previous chapter the method of examining the 
blood of living persons has been described, and there is, by this 
means, ample opportunity for satisfying oneself that bacteria are 
never to be found in the blood in health. The organs removed from a 
perfectly healthy animal, with the necessary precautions, and placed 
in sterilised media, can be kept indefinitely without undergoing 
putrefaction, or giving any development of bacteria. This has been 
established by many observers, notably Oheyne and Hauser ; and 
the results of former observers to the contrary must be attributed 
to imperfect methods admitting of accidental contamination. 



CHAPTER III. 

EFFECT OF ANTISEPTICS AND DISINFECTANTS ON BACTERIA. 

In the previous cliapter several conditions were alluded to which 
affected the growth of bacteria, such as the nature of the 
nutrient soil, temperature, light, and electricity. The effect of 
certain chemical substances, and of excessive heat and cold, was 
also mentioned ; but this constitutes a subject of such practical 
importance that it must be considered more fully. v 5 T 

Agents which retard the growth of bacteria are generally spoken 
of as antiseptics, as distinguished from disinfectants which altogether 
destroy their vitality. 

Though chemical disinfectants, or germicides, when diluted, act 
as efficient antiseptics, the converse, that an antiseptic in a suffi- 
ciently concentrated form will act as a disinfectant, is not the case. 
The term " antiseptic,'' indeed, should be restricted to those sub- 
stances or agents which arrest the changes bacteria produce, but 
which do not prevent their springing into activity when removed 
to favourable conditions. Thus excessive heat, which destro3's 
bacteria and their spores, is a true disinfectant ; and excessive cold, 
which only benumbs them, retarding their development without 
killing them, is an antiseptic. 

Spores have a greater power of resisting the action of these 
various agents than the parent cells, and many species of micro- 
organisms differ from each other in their resisting power. An 
exact knowledge of the subject can, therefore, only be based upon 
investigations which will determine the effect of these agents upon 
pure cultivations of the different micro-organisms causally related to 
putrefaction and disease. In the latter case, especially, this is not 
possible in the present state of our knowledge. In some cases of 
communicable disease there is considerable doubt as to the etiological 
importance of the organisms which have been described ; in other 
cases no organisms have as yet been discovered, or the organisms 

30 



EFFECT OF ANTISEPTICS AND DISINFECTANTS ON BACTERIA. 31 

cannot be artificially cultivated, or the disease is not reproduced by 
inoculation, so that there is no means of testing whether the agents 
have had any efEect. One can, therefore, only draw general 
•conclusions by selecting some well-known pathogenic and non- 
pathogenic micro-organisms, and considering the influence of 
chemicals, of hot air and of steam upon them, as representing the 
■effect upon the various contagia of disease and the causes of 
putrefaction. 

Such knowledge must necessarily prove of the greatest im- 
portance : to the sanitarian, who is concerned in preventing the 
spreading of disease and in the disposal of putrefactive matter ; to 
the surgeon, who is anxious to exclude micro-organisms during 
surgical operations, and to arrest the development of bacteria which 
have already gained an entrance in wounds ; to the physician, in 
the treatment of micro-parasitic diseases. 

The sanitarian and the surgeon must profit directly by such 
experiments, for in the disinfection of clothes and the sick-room by 
the one, and in the application of antiseptic dressings and lotions 
by the other, the micro-organisms are encountered, as in the experi- 
ments, outside the living body. 

The physician, on the other hand, is principally concerned in 
dealing with micro- parasites when circulating in the blood, or 
carrying on their destructive processes in the internal tissues. So 
far as our knowledge at present goes, the physician can avail him- 
self but httle of the effect of the direct application of the substances 
which have been found to retard or destroy the growth of the 
organisms in artificial cultivations, for the concentrated form in 
which they would have to be administered would prove as deleteri- 
ous or as fatal to the host as to the parasites. Thus Koch has 
stated that to check the growth of the anthrax bacillus in man it 
would be necessary that there should be twelve grammes of iodine 
constantly in circulation, and that the dose of quinine necessary 
to destroy the spirilla of relapsing fever would be from twelve to 
sixteen grammes. The retarding iniluence, however, of certain 
iiubstances when diluted, and the fact that disinfectants are some- 
times equally efiicacious in a diluted form when their application is 
prolonged, seem to indicate measures which may be adopted, in some 
cases, with chances of success, such, as the inhalation of antiseptic 
vapours in phthisis. For the most part the physician must look 
rather to combating the effects of micro-organisms by restoring to 
its normal standard the lowered vitality which enabled the bacteria 
to get a footing. 



32 BACTERIOLOGY. 

There is no wider field for research than the determination of 
the real effect of disinfectants and antiseptics. Painstaking and 
laborious as the researches are which have been hitherto made, the 
subject is so beset with fallacies, leading, in some cases, to totally, 
erroneous conclusions, that it is not surprising that one meets on 
all sides with conflicting statements. The author has no intention 
of analysing these results, but a general idea will be given of the 
methods which have been employed, and for further details reference 
must be made to the original papers mentioned in the bibliography. 

Chemical Substances. — It was customary to judge of the power 
of a disinfectant or antiseptic by adding it to some putrescent liquid. 
A small portion of the latter was, after a time, transferred to 
some suitable nourishing medium, and the efficacy of the substance 
estimated by the absence of cloudiness, odour, or other sign of 
development of bacteria in the inoculated fluid. Koch pointed out 
the errors that might arise in these experiments from accidental 
contamination, or from there being no evidence of the destruction 
of spores, and we are indebted to him for a complete and careful 
series of observations with more exact methods. 

Instead of employing a mixture of bacteria, Koch's plan was to 
subject a pure cultivation of some well-known species with marked 
characteristics to the reagent to be tested. A small quantity was 
then transferred to fresh nourishing soil, under favourable con- 
ditions, side by side with nutrient material inoculated from a 
cultivation without ' treatment with the disinfectant. The latter 
constituted a control test, which . is most essential in all such 
experiments. To test the resistant power of bacteria which are 
easily destroyed, two species were selected. Micrococcus prodigiosus,. 
and the bacillus of blue pus. These were cultivated on potatoes, 
the surfaces of which were sliced off and dried. A fragment trans- 
ferred to freshly prepared potato gave rise to a growth of the 
particular micro-organism ; but if after treatment with some reagent 
no growth occurred, the conclusion was drawn that the reagent was 
efficacious in destroying the vitality of the bacteria. 

Anthrax bacilli in blood, withdrawn from an animal just killed, 
were taken to represent sporeless bacteria, while silk threads steeped 
in an artificial cultivation of the bacilli and dried, afforded a means 
of testing the vitality of spores. 

Even by employing pure cultivations on solid media, great 
precautions were necessary to avoid mistakes. When, for instance 
a large quantity of the growth which had been subjected to some 
chemical sokition w;is carried over to the fresh tube containing 



EFFECTT OF ANTISEPTICS AND DISINFECTANTS ON BACTERIA. 33 

the nutrient medium, or when a silk thread, which had been dipped 
in a sohition, was directly transferred to the new soil, enough of the 
supposed disinfectant might be mechanically carried over to retard 
the development of the bacteria, though it was ineffectual in 
destroying them. From a growtli not appearing, it was concluded 
that the spores or the bacteria had been affected, and so a 
mistake occurred. To avoid this, Koch made a point of transfer- 
ring a minimum of the disinfected growth to as large a cultivation 
area as possible, so that any chemical substance mechanically 
carried over would be so diluted as to be inert. For the same 
reason, threads, after withdrawal from the disinfecting solution, 
were rinsed in sterilised water, or weak alcohol, and then trans- 
planted; or, instead of judging from , the ' development on nutrient 
gelatine, the effect of inoculation in a healthy animal was made 
the test. 

A few examples may be quoted in illustration. Silk threads, 
impregnated with anthrax spores, were placed in bottles containing 
carbohc acid of various strengths. A thread was removed from each 
on successive days, and transferred to nutrient gelatine, and the 
result noted. It was found that immersion of the thread in a 5 per 
cent, solution of carbolic acid was sufficient in two days to effect 
complete sterilisation, and seven days in a 3 per cent, solution was 
equally efficacious. Since for practical purposes a strength should 
be selected which would be effectual in twenty-four hours, Koch 
recommended that for general use, allowing for deterioration by 
keeping, a solution containing not less than 5 per cent, should be 
employed, and for complex fluids probably a still higher percentage 
would be necessary. In the case of sporeless bacilli the results were 
very different. Blood containing the bacUli, from an animal just 
killed, was dried on threads, and after exposure for two minutes to 
a 1 per cent, solution, was completely sterilised; and fresh blood 
mixed with a 1 per cent, carbolic solution produced no effect when 
inoculated. On the other hand, when the blood was mixed with a 
■5 per cent, solution, the virulence was not destroyed. The facility 
with which the bacilK are destroyed, compared with their spores, 
illustrates how easily errors may occur, when mere arrest of growth 
or loss of motihty is regarded as a sign of the efficacy of disinfection. 

To test vapours, Koch exposed anthrax spores or the spores 
which occiir in garden earth by suspending them over solutions, 
such as bromine or chlorine, in a closed vessel. After a time they 
were transferred to a nutrient medium to test their vitality. To 
test the power of sulphurous acid gas, the spores were spread about 

3 



S4 BACTERIOLOGY. 

in a room in which the gas was generated by burning sulphur in 
the ordinary way for disinfecting a room. To test chemicals which 
might be recommended for disinfecting vans and railway carriages, 
spores were laid on boards, which were then washed or sprayed, and 
the spores then transferred to the nutrient gelatine. 

Sternberg has also made an elaborate series of experiments with 
regard to the action of germicides. In this case cultivations of 
well-known pathogenic organisms in liqiiid media were employed. 
The supposed germicide was added to the liquid cultivation, and 
after two hours a fresh flask of sterilised culture was inoculated from 
the disinfected cultivation, and placed in the incubator. In twenty- 
four to forty-eight hour.s, if the chemical proved inefficient, there 
was evidence of a growth of bacteria. Blyth has investigated the 
disinfection of cultivations of Bacterium termo, of sewage, and 
typhoid excreta, and, in conjunction with EHein, the effect of well- 
known disinfectant materials on anthrax spores. Miquel, Laws, 
and others have also contributed to our knowledge of the . effect 
of antiseptics and disinfectants upon micro-organisms. In spite of 
all that has been done there is room for many workers ; a great 
deal of ground must be gone over again to rectify discrepancies,, 
examine conflicting results, and thus determine what observations 
may be relied upon for practical application. 

This may be illustrated by referring in detail to some experiments 
made with corrosive sublimate. Koch investigated a long Hst of 
chemical reagents, and according to these experiments the salts of 
mercury, and the chloride especially, proved most valuable. Where 
heat is not admissible, these compounds were therefore highly 
recommended, though their poisonous nature is a drawback to their 
indiscriminate use. Koch stated that for disinfecting a ship's bilge, 
where a 5 per cent, solution of carbohc acid must be left forty-eight 
hours, a 1 in 1000 solution of mercuric chloride would only require 
a few minutes. 

But there was good reason for doubting the efficacy of very dilute 
solutions; for, though according to Koch's experiments anthrax 
spores subjected to a 1 in 20,000 solution of mercuric chloride for 
ten minutes,- and then washed in alcohol, gave no growth in nutrient 
gelatine, silk threads exposed for ten minutes to a 1 in 20,000 
solution, or even 1 in 10,000, still proved fatal to mice. 

Herroun cultivated ordinary septic bacteria in albuminous 
filtrates, containing . 1 in 2000, and concluded that the value of 
mercuric chloride as an antiseptic was much over-rated. It is pre- 
cipitated by albumins though, as Lister has shown, the precipitate 



EtTECT OF ANTISEPTICS AND DISINFECTANTS ON BACTERIA. 35 

of albuminate of mercury is redissolved when there is an excess of 
albumin present. 

Geppert, and later Behring, recognised that the methods employed 
for testing the eificacy of corrosive sublimate were um-eliable. Thej^ 
found, for example, that corrosive sublimate could not be removed 
from silk threads by washing ; and therefore to study the effect of 
this antiseptic acting for a given time, it was necessary to dip the 
threads in ammonium sulphide solution after the treatment with 
corrosive sublimate. 

The author confirmed the results of Geppert and Behring, and 
made a series of experiments to test the value respectively of carbolic 
acid and corrosive sublimate in antiseptic surgery. The method 
of dipping an infected thread into an antiseptic solution for a few 
minutes, and then transferring it to the surface of a nutrient medium 
to test its efficacy in a given time, was discarded as fallacious ; the 
thread being still wet with the solution when transferred to the 
medium, it was obvious that the action of the antiseptic continued 
for many, days. To wash infected silk threads with alcohol after 
exposure to the antiseptic to stop its further action, also proved to 
be a fallacious method, for the author found in control experiments 
that absolute alcohol will destroy Streptococcus pyogenes, erysipelatis, 
and Staphylococcus pyogenes aureus, acting for only one minute. 
Other methods were therefore resorted to, and cultures on the 
sloping surface of nutrient agar were at first used. The antiseptics 
was poured into the culture tube until the growth was covered, 
and when it had acted for a definite time (one minute, five minutes,, 
or fifteen minutes) a solution was added which immediately stopped 
further action. In the case of corrosive sublimate, ammonium sul- 
phide was employed, which is quite inert as an antiseptic. The liquid 
contents of the test tube were carefully poured off, and an inoculation 
was made into a fresh tube of broth or agar from the culture still 
adhering to the surface of the nutrient medium. As the results 
disproved the efficacy of corrosive subhmate, it was thought possible 
that the solution had not been able in the time to penetrate the 
film of growth. Another plan was accordingly adopted. Cultures 
were made in broth, and when fully developed the supernatant liquid 
was carefully poured off. Corrosive sublimate solution was added to 
the test, tube, and agitated until any flocculent masses were dis- 
integrated and the whole of the liquid became uniformly turbid. 
Ammonium sulphide was added when the time had expired, and 
tubes of fresh broth were inoculated with the mixture. In the case 
of carbolic acid the cultures, after its action, were thoroughly washed 



36 BACTERIOLOGY. 

with water, and its efficacy tested by making inoculations from the 
cultures in fresh media. The results were entirely in favour of 
carbolic acid. Staphylococcus pyogenes aureus and Streptococcus 
pyogenes were not destroyed, even when corrosive sublimate solution 
of 1 in 1000 was allowed to act for an hour. In the case of the 
cultures of streptococcus of erysipelas the results were different. 
A solution of 1 in 10,000 had no effect, but 1 in 4,000, acting for 
one minute, destroyed the culture. With carbolic acid the results 
were very striking. Cultures were exposed to solutions of 1 in 20, 
1 in 30, 1 in 40, 1 in 50, for one minute, five minutes, fifteen 
minutes. The attempts to make subcultures in every case failed. 
Carbolic acid 1 in 40, acting for only one minute, was sufficient to 
destroy Streptococcus pyogenes and Streptococcus erysipelatis and 
Staphylococcus pyogenes aureus. Further experiments were made 
with tubercular sputum, the test being subsequent inoculation of 
guinea-pigs. Corrosive sublimate solution as strong as 1 in 500 had 
no effect, but 1 in 20 carbolic acid, shaken up with the sputum for 
one minute, completely neutralised it. 

Koch's statements with reference to the germicidal power of 
corrosive sublimate in extremely weak solutions had led Lister to 
substitute it for carbolic acid as a detergent in surgery. The author's 
experiments, which were undertaken in 1892, encouraged Lister to 
revert to the use of carbolic acid, w-hich, indeed, had always proved 
efiicacious in surgical practice. Lister pointed out that carbolic acid 
has also the great advantage of combining eagerly with fats and 
epidermis, so that the seat of operation can be effectually cleansed. 

These experiments also point to the conclusion that carbolic acid 
should be used in hospital wards for the disinfection of tubercular 
.sputum instead of mercuric chloride and other less efficacious dis- 
infectants commonly in use. 

Hot Air and Steam. — Koch, in conjunction with WolfhUgel, also 
made exhaustive experiments to test the value of hot air. A similar 
plan was adopted to that employed in disinfection with chemicals. 
Bacteria and spores were subjected for a certain time to a known 
temperature in the hot-air chamber, and then were transferred to a 
nourishing soil or inoculated in animals. 

Paper parcels, blankets, bags, and pillows, containing samples of 
micro-organisms wrapped up inside, were also placed in the hot-air 
chamber, to test the power of penetration of heat. 

The conclusions from these experiments were as follows : 

Sporeless micro-organisms at a little over 100° C. are destroyed 
in one hour and a half. 



EFFECT OF ANTISEPTICS AND. DISINFECTANTS ON BACTERIA. 37 

Spores of bacilli require three hours at 140° 0. 

If enclosed in pillows and blankets, exposure from three to four 
hours to 140° 0. is necessary. 

Spores of fungi require one and a half hours at 110° 0. to 
115° 0. 

Further experiments showed that at the temperature necessary 
for the destruction of spores of bacilli almost all fabrics are more or 
less injured. 

Koch, in conjunction with Gaffky and Loffler, also investigated 
the effect of steam under pressure and at the atmospheric pressure. 

Rolls of flannel with anthrax spores or earth spores, and a 
thermometer wrapped up Inside, were subjected to steam, and 
the results compared with the effect obtained with hot air. 

Thus in hot air four hours' exposure to a temperature of 130° C. 
to 140° C. brought the temperature inside the roll to 85° C, and the 
spores were not injured ; on the other hand, exposure to steam. 
under pressure at 120° 0. for one and a half hours, raised the 
internal temperature to 117° C. and killed the spores. 

By such experiments the superior penetrative power of steam- 
heat was established. 

To test steam-heat at the atmospheric pressure, water was boiled 
in a glass flask with its neck prolonged by means of a glass tube, the 
temperature in which was found to be uniform throughout. Anthrax 
and earth spores placed in the tube were found to be unable to with- 
stand steam at 100° C. even for a few minutes. It was, therefore, 
concluded that disinfection by steam at atmospheric pressure was 
superior to hot air from its greater efficiency, and to steam under 
pressure from the simplicity of the necessary apparatus. 

Parsons and Klein made some experiments which were more 
in favour of dry heat than the above. These observers state that 
anthrax bacilli are destroyed by an exposure of five minutes at 
from 100° C. to 103° 0. and that anthrax spores are destroyed 
in four hours at 104° C, or in one hour at 118° C. Guinea-pigs 
inoculated with tuberculous pus which had been exposed for five 
'minvites to 104° C, remained unaffected. They concluded that as 
none of the infectious diseases, for which disinfecting measures 
are in practice commonly applied, are known to depend upon the 
presence of bacilli in a spore-bsaring condition, their contagia 
are not likely to retain their activity after being heated for an 
hour to 105° C. (220° Fahr.) 

In experiments with steam the results were in accordance 
with those already given, and complete penetration of an object 



38 BACTERIOLOGY. 

by steam-heat for more than five minvites was deemed sufficient. 
They also arrived at the same result as in Koch's experiments, 
viz., that steam-chambers are preferable to those in which dry heat 
is employed, though it must be borne in mind that some articles, 
.such as leather, are injured by exposure to steam. 

Practical Application. 

Nurses and others attending infectious oases should freely use 
1 in 40 cai-bolic for the hands and a weaker solution for the body 
generally. The skin of patients after recovery should be sponged 
with 1 in 40 carbolic. The dead should be wrapped up in a sheet 
soaked in 1 in 20 carbolic acid or a strong solution of chloride of 
lime. Infected clothing and bedding should be burnt unless in excep- 
tional cases, when they may be disinfected by boiling, or by exposure 
to dry heat at 105° C. to 110° 0. for three hours, or by steaming 
at 100° C. for fifteen minutes. Leather and other articles which 
would be destroyed by any of these processes should be sponged with 
1 in 40 carbolic. The walls of the sick-room and furniture should 
be exposed to the fumes of burning sulphur, and next day washed 
down with 1 in 40 carbolic, and the room freely ventilated by 
opening all windows and doors. Rags should be burnt, or dis- 
infected by boiling or exposure to steam when supplied to manu.- 
facturers. . The importation of rags from places where there are 
cases of cholera or small-pox should be prohibited. Infected ships 
must be fumigated with sulphur, and the bilge disinfected -with 
carbolic acid. Infected railway carriages should be disinfected in 
the same way as a sick-room. 

Tubercular sputum, cholera and typhoid evacuations and other 
excreta should be disinfected by I in 20 carbolic acid, or by a strong 
solution of chloride of lime. 



CHAPTER IV. 

CHEMICAL PRODUCTS OF BACTERIA. 

The products of the metabolism induced by bacteria may be divided 
into three classes : (1) ptomaines or alkaloids ; (2) albumoses or tox- 
albumins ; and (3) enzymes. Alkaloids and albumoses are directly 
poisonous ; enzymes or ferments are harmless except in the presence 
of proteids, which they are capable of transforming into poisonous 
albumoses. 

Ptomaines and Tox- Albumins. 

The study of these products may be said to date back to 1822, 
when Gaspard and Stick found an intensely poisonous principle 
in cadaverous extracts. In 1856 Panum discovered a poisonous 
substance in putrid ilesh; and in 1863 Bergmann and Smiedeberg 
found a nitrogenous crystallisable substance in putrid beer which 
they named sepsin. In 1872 Gautier found that the decomposition 
of fibrine led to the formation of various complex alkaloidal sub- 
stances, and in 1875 Richardson obtained in pyaemia an alkaloid, 
septin. This subject, however, received most attention from the 
classical researches of Selmi, the Italian toxicologist. Selmi, in a 
celebrated poisoning case, demonstrated the presence of an alkaloid 
as the result of post-mortem changes. Similar substances were 
found in alcohol in which morbid specimens had been preserved. 
Thus the researches of Gautier and Selmi established the fact that 
aJbuminoid material tindergoing decomposition leads to the forma- 
tion of cadaveric alkaloids. These animal alkaloids Selmi named 
ptomaines. Brieger, finding the bases derived from the products of 
putrefaction less poisonous than those obtained from the pathogenic 
bacteria, suggested the' term toaains for the latter. Ptomaines have 
been divided into two classes — those which are non-oxygenous, liquid, 
and volatile, and those which are oxygenous, solid, and crystallisable. 
They are, for the most part, precipitated by the ordinary reagents 

39 



40 BACTERIOLOGY. 

for alkaloids, such as chloride of gold, double iodide of mercury 
and potassium, picric acid, and tannin. Phospho-molybdic acid 
precipitates them without exception. They are powerful reducing 
agents. Ferro-cyanide of potassium is converted into ferri-cyanide in 
theii' presence, and the addition of ferric chloride gives the Prussian 
blue test. Selmi discovered this tsst, and Brouardel and Boutmy 
regarded it as absolutely characteristic of ptomaines; but this is 
not the case ; some vegetable alkaloids, for example, behave in the 
same way. 

As examples of the non-oxygenous ptomaines there are : — 

Parvolin (O^H^'N) an oily base of an amber colour prepared 
from putrid mackerel and horse-flesh. 

Hijfh-ocollidin (C^Hi^N), from the same source. It is highly toxic, 
being compared by Gautier to the venom of the cobra di capello. 

GoUidin (C^H^'N), from putrid gela'tine and the pancreas of a 
bullock, also highly toxic. 

Xeuridin (O^H"N^), from fish, flesh, and decaying cheese. 

Sa/prin (CHi^N^), isomeric with neuridin. 

Cadaverin (CH^^N"^), a third isomeride, from ordinary putrefac- 
tion and herring brine. 

Putrescin (C^H^^N^) from putrefaction. 

The oxygenous ptomaines are in some instances found also in 
healthy tissues. They include the following : — 

Newrin (C^H^'NO), found in cadaveric putrefaction. 

Cholin (O-'^H^NO^), in bile. 

Jfuscarin (CH^'NO*), in a poisonous mushroom, Agaricus mus- 
carius, and in putrid fish. These are all highly poisonous. 

Gadinin (O^Hi^NO^), in putrefying codfish. 

Mytilotoxin (C^ff'NO^), in poisonous mussels. 

Poisonovis alkaloids are of great importance in connection with 
those cases of meat poisoning produced by sausages, hams, poultry, 
and cheese. Tyrotoxicon is a poisonous alkaloid obtained from cheese. 

The toxic substances of most interest to the bacteriologist are 
those isolated from pure cultivations of pathogenic bacteria, such as 
typhotoxin, isolated by Brieger from cultivations of the bacillus of 
typhoid fever, and tetanin, from cultivations of the tetanus baciUus ; 
and the poisons known as albumoses or tox-albumins, which are 
allied to the albumose of snake poison. 

Pasteur, in 1885, suggested that in anti-rabic inoculations the 
immunity resulted from the action of a substance secreted by a 
microbe, though the microbe has not as yet been discovei-ed in 
rabies. Salmon produced immunity from hog cholera by the iniec- 



CHEMICAL PRODUCTS OF BACTERIA. 41 

tioii of the toxic products in filtered culture fluids. Wooldridge, 
Hankin, and Martin studied the products of Bacillus anthracis. 
Charrin, and later Woodhead, Wood, and Blagovestchensky, investi- 
gated on these lines Bacillus pyocyaneus, Eoux and Chamberland 
experimented with the bacillus of malignant cedema; Eoux with 
symptomatic anthrax; Chantemesse and Widal with the typhoid 
bacillus. Roux, Yersin, Brieger, Fraiikel, Mai-tin, and Behring 
worked on the same lines with diphtheria. Koch introduced 
tuberculin, Kalning mallein, while others have utilised the products 
of streptococci and pneumococci. Anrep found an albumose in the 
medulla of rabid animals, and Babfes claims to have found an 
albumose in both rabies and glanders. 

Cholera. — Brieger found several ptomaines, including putrescin 
and cadaverin, ip. pure cultures of the spirillum of Asiatic cholera, 
and Petri found in addition to poisonous bases a proteid body which 
produces in guinea-pigs muscular tremors, paralysis, and a rapidly 
fatal result. Roux and Yersin obtained from cultures a tox-albumin 
insoluble in water, which kills guinea-pigs in two or three days, 
but has no effect on rabbits. Pfeifler also investigated the toxic 
substances in cultures. Chloroform, thymol, and drjdng destroyed 
comma -bacilli, leaving their toxic products unaffected. Concentrated 
solutions of neutral salts and boiling produced secondary toxic 
substances, but the original toxic substances were ten or twenty 
times more virulent. 

Typhoid Fever.— Typhotoxin (CH^NO^), the alkaloid ob- 
tained by Brieger from cultures of the typhoid fever bacUlus, produces 
in mice and guinea-pigs salivation, rapid breathing, dilatation of tl^ 
pupil, diarrhoea, and 'death in twenty- four to forty-eight hours. At 
the post-mortem examination the heart is found in a state of systolic 
contraction, and the condition of the heart after death and the 
absence of convulsions during life serve to distinguish typhotoxin 
from an isomeric base obtained by Brieger from putrid horse-flesh. 
Roux and Yersin have obtained a tox-albumin. It is soluble with 
difficulty in water, and more toxic to rabbits than guinea-pigs. 

Tetanus Brieger obtained the alkaloid tetanin from impure 

cultures of the tetanus bacillus. It is a base having the formula 
Qi3jj22]v^2Q4_ - 'jijg hydrochloride is a veiy deliquescent salt, and 
soluble in alcohol. Tetanin injected into guinea-pigs produces 
rapid . breathing, followed by tetanic convulsions. Anothei' toxic 
product, tetanotoxin (C^H^^N), produces the same effects as tetanin. 
The foi-mula of a third base, spasmotoxin, has not been determined. 
Cadaverin and putrescin are also present in cultures. Kitasato and 



42 BACTERIOLOGY. 

Weyl analysed the products of pure-cviltures, and obtained the same 
substances, tetanin and tetanotoxin ; and subsequently Brieger and 
Frankel found that in pure-cultures a tox-albumin could be 
obtained which is soluble in water, and infinitely more active tlian 
the toxic ptomaines. 

Anthrax.— In 1887 Wooldridge succeeded in protecting rabbits 
from anthrax by a new method. A proteid body obtained from the 
testis and from the thymus gland was used as the culture fluid. 
This proteid substance was dissolved in dilute alkaH, and the solution 
sterilised by repeated boiling. This was inoculated with the anthrax 
bacillus, and kept at 37° C. for two or three days. A small quantity 
of the filtered culture fluid injected into the circulation in rabbits 
produced immunity from anthrax. Subcutaneous inoculation of 
extremely virulent anthrax blood, made simultaneously with the injec- 
tion of the protecting fluid, produced no effect. Wooldridge showed 
that the growth of the anthrax bacillus in special culture fluids 
gave rise to a substance which, when injected into the organism, 
protected not only against an immediate but also subsequent attacks. 
In 1889 Hankin worked under the guidance of Koch in the 
Hygienic Institute of Berlin. The acquired tolerance of the effect 
of ordinary albumoses, and the experiments of Sewall, who pro- 
duced immunity against lethal doses of the albumose of snake 
poison by the injection of minute doses, led Hankin to expect that 
an albumose developed in anthrax cultures, arid that the anthrax 
albumose would probably confer immunity from the disease. Hankhi 
succeeded in isolating it from culture fluids. It was precipitated by 
excess of absolute alcohol, well washed in alcohol to free it from 
addition of ptomaines, filtered, dried, then redissolved and filtered 
through a Chamberland filter. With this substance Hankin suc- 
ceeded in producing immunity in mice and rabbits. 

Sidney Martin, working quite independently, grew anthrax bacilli 
in a solution of pure alkali albumin made from serum proteids. After 
ten or fifteen days the organisms were removed by filtration throvxgh 
a Chamberland filter. The filtrate contained p)-oto-alhumose and 
deutero-alhuniose, a trace of peptone, an alkaloid, and small quantities 
of leuciii and tyrosin. The mixture of albumoses proved poisonous 
to mice. The anthrax alkaloid produced symptoms and lesions 
similar to the albumoses, but mvich more rapidly and severely. It 
is an amorphous yellow body, soluble in alcohol and alkaline in 
reaction. Martin concluded that the anthrax bacillus formed the 
albumoses and the alkaloid by digesting the alkaU albumin ; and 
.suggested that the alkalinity of the albumoses explained their toxiC 



CHEMICAL PRODUCTS OF BACTERIA. 43 

properties, the alkaloid probably being in a nascent condition in the 
albumose molecule. 

Tuberculosis. — Koch prepared a glycerine extract of the 
product of the tubercle bacillus in pure cultivations, and found 
that the injection of small doses produced a remarkable reaction, 
both local and general, in tubercular cases, and especially lupus. 
This extract, called tuh&culin, came to be extensively used as a 
therapeutic agent, but with disappointing results. Yerj- shortly 
after the first announcement of Koch's discovery, the author, in 
conjunction with Herroun, investigated the chemical properties and 
physiological effects of the products of the tubercle bacillus. 
Cultures in glycerine-broth were filtered through porcelain, and 
a clear amber-coloured liquid was obtained, which gave important 
and suggestive chemical reactions. As this filtrate contained the 
products of the growth of the bacillus most probably in minute 
quantities, it was evaporated at m, low temperature over sulphuric 
acid. The viscous residue was dissolved in distilled water and 
tested on the healthy guinea-pig. The result was a marked fall of 
temperature, staring coat, extreme irregularity of the heart's action, 
muscular spasmii, loss of control over the extremities, and death. 

A preliminary examination of glycerine-broth cultivations having 
fihown the presence of non-coagulable proteid bodies of the nature 
of alburhose and peptone, and a crystallisable precipitate of a 
remarkable character resulting on the addition of iodine, the idea 
Naturally suggested itself that the tubercle bacillus might form 
albumoses and an alkaloid or ptomaine similar to the substances 
isolated by Martin from pure cultivations of the Bacillus anthracis. 

Koch pointed out that the effective substance in his extract 
could be precipitated by absolute alcohol ; the author and Herroun 
detex'mined to investigate the properties and physiological effects 
of the separated products. They accordingly set to work to isolate 
the ptomaine, of the existence of which they had some quahtative 
indication, and at the same time to examine the properties of the 
albuminous bodies. 

In this endeavour the general method they found satisfactory was 
as follows. The clear filtrate from the culture was evaporated at 
40° 0. to a very small bulk, and the residue thus obtained was 
mixed with an excess ef absolute alcohol, which precipitated the 
albumoses and peptone. It was found that by adding the alcohol 
by degrees a partial separation of the albumose from the peptone 
could be effected, the latter being only precipitated when the alcohol 
was nearly absolute. The precipitated albumose was collected on 



44 BACTERIOLOGY. 

a filter and redissolved in distilled water. In another experiment 
the albumose underwent a second precipitation, and after washing 
was again dissolved. 

The alcoholic filtrate from the precipitated albuminous bodies was 
then concentrated at a very gentle heat until a viscous residue was 
left containing the glycerine originally present in the cultivating 
medium and the extractives and products of the bacillus soluble 
in alcohol. With this residue definite reactions of an alkaloidal 
substance or ptomaine were obtained. 

Careful experiments, however, led to the belief that the whole 
of the ptomaine was not separated from the albuminous precipitate 
by simple addition of alcohol, and the above method was therefore 
slightly modified. 

The ptomaine is soluble in water and alcohol, and sparinglj' 
soluble in amyl-alcohol, but insoluble in benzine, ether, or chloro- 
form, which liquids therefore fail to extract it from aqueous 
solutions. In its aqueous solutions it is distinctly but not strongly 
alkaline to test-paper. Phospho-tungstic acid gives with it a white 
flocculent precipitate. Phospho-molybdio acid gives a pale yellow 
precipitate, soluble in ammonia to a blue solution which becomes 
colourless on boiling. In this respect it resembles the vegetable alka- 
' loids, aconitin and a'tropin. It must be remembered, however, that 
albuminous bodies are precipitated by both this and the preceding 
reagents, and in the case of the former a reduction of the phospho- 
molybdate giving the blue solution with ammonia is obtained. 

The reducing power of the ptomaine is shown by the conversion 
after a short time of ferri-cyanide of potassium to ferro-cyanide, 
giving the Prussian blue test with ferric chloride, to which much 
undue importance was attached by Brouardel and Boutmy. The 
solution of albumose and sokition of peptone are both capable of 
giving this reaction as well as many vegetable alkaloids. A solution 
of the ptomaine is not precipitated by ferro-cyanide of potassium or 
potassic bichromate. 

In strong solutions it yields precipitates with platinic chloride 
(yellow), gold chloride (pale yellow), and mercuric chloride (white). 
That yielded by the first of these reagents is granular in character, 
and quite insoluble in alcohol, though apparently soluble in water. 
The precipitation by gold chloride excludes amides and ammonium 
salts. 

With iodine in hydriodic acid or potassic iodide a precipitate is 
obtained which is occasionally crystalline, more often granular or 
amorphous. 



CHEMICAT. PRODUCTS OF BACTERIA. 45 

This precipitate is soluble in alcohol, and is redeposited when the 
alcohol is evaporated. On heating it is redissolved into oily drops of 
a dark colour. With picric acid a granular precipitate is obtained, 
■which under the microscope is seen to consist of minutfe crystals. 
This precipitate, on standing, is converted into rounded crystalline 
masses with numerous small crystals admixed. 

The ptomaine appears to be easily broken up by heating, 
especially in the presence of mineral acids or of baryta. The actual 
quantity obtained from a considerable amount of culture fluid was 
very small, and as it was possible that when the bacilli were grown 
in a medium richer in albumin, such as the animal body, more of 
these products might be formed, the liquid obtained by extracting 
large masses of tubercular growths from cattle was examined in a 
similar manner. In this extract, after filtration through porce- 
lain, an albumose, and minute quantities of a ptomaine were 
obtained which in reactions was identical 'W'ith that obtained from 
the artificial cultivation of the bacillus, but present in even smaller 
amount. The probable explanation of this is, that, in the living 
animal the ptomaine is constantly being removed ; or it may indicate 
that it is only formed in minute quantity under those conditions. 

Having succeeded in obtaining the albumose and the ptomaine 
in separate solutions, we next proceeded to ascertain the effects of 
these substances upon healthy and tubercular guinea-pigs. 

The effect of the ptomaine isolated from different sei'ies of 
cultures was as follows. A rise of temperature occurred in tuber- 
cular animals, and distinct enlargement of tubercular glands. There 
was a slight indication of a depression of temperature or hypothermic 
effect on healthy animals. The albumose, whether obtained from 
pure cultivations of the bacillus or from tubercular tissue, pro- 
duced a marked rise of temperature in tubercular guinea-pigs. On 
the other hand, in a control experiment on a healthy guinea-pig 
there was an equally well-marked fall of temperature. The effect 
upon the tubercular glands in the cases associated with marked rise 
of temperature was to render them well-defined, indurated, and 
painful, rather than any considerable increase in volume. 

Hunter made a chemical examination of Koch's crude extract, 
and confirmed the presence of albumoses and alkaloidal substances. 
The albumoses consisted chiefly of proto-albumose and deutero-albu 
mose with hetero- albumose, and occasionally a trace of dys-albumose. 
Two alkaloidal substances were obtained in the form of platinum 
compounds of their hydrochlorate salts. In addition there were 
extractives, mucin, inorganic salts, glycerine, and colouring-matter. 



46 BACTERIOLOGY. 

Swine Fever. — Schweinitz applied Brieger's metliods in the 
investigation of the products of the swine fever, or hog cholera 
bacillus. Broth- cultures were neutralised with dilute hydrochloric 
acid, and 'evaporated in the water bath. The residue was treated 
with 96 per cent, alcohol, and the filtered solution with mercuric 
chloride. A heavy crystalline precipitate was separated by filtra- 
tion, treated with water, and decomposed with sulphuretted hydrogen, 
and cadaverin and methylamine were isolated. The filti'ate from 
the mercuric chloride pi-ecipitate was freed from excess of mercury 
by sulphuretted hydrogen, and the miercury sulphide filtered off. 
The residue, after concentration of the filtrate, was extracted with 
absolute alcohol, and the solution showed the presence of an alka- 
loidal salt. The double salt obtained with platinum chloride was 
submitted, after crystalhsation, to an analysis, and the results gave 
the formula (Oi^HS^NsPtOF). The hydi-o-chloride is soluble in abso- 
lute alcohol as well as in water, and produces needle-hke crystals. 

On treating the culture fluids with excess of absolute alpohol a 
white flocculent precipitate was obtained partly soluble in water, and 
re-precipitated by alcohol. It was obtained in the form of white 
crystalline plates. A watery solution gives almost insoluble needle- 
crystals on the addition of platinum chloride. These products were 
respectively termed sucholo-toxin and sucholo-albumin. Small doses 
of these substances produce in guinea-pigs a slight rise in tempera- 
ture, and ulcsration at the seat of injection. Large doses produce 
a fatal result in six to twenty-four hours. Schweinitz asserts that 
he has pi-oduced immunity in guinea-pigs. An attempt to produce 
immunity in swine by injection of the albumose gave unsatisfactory 
results. 

Diphtheria. — Eoux and Yersin finding that filtered cultures of 
the diphtheria bacillus produced paralysis, affecting chiefly the hind 
legs, and a fatal result in rabbits and guinea-pigs, proceeded to 
investigate the chemical products. They succeeded in obtaining a 
white amorphous substance which was extremely active when 
injected into guinea-pigs. It was precipitated by alcohol from an 
aqueous solution, and it was calculated that -0004 gram would 
destroy eight guinea-pigs of 400 grams, or two rabbits of 3 kilos, 
each. They concluded that the poison was an enzyme or ferment 
as it not only acted in extremely small doses, but it was attenuated 
by heat and destroyed by boiling. 

Brieger and Frankel confirmed these experiments, and asserted 
that the poison was a tox-albumin ; but according to Martin their 
chemical analysis and reactions were vitiated by the fact that they 



CHEMICAL PKODUCTS OF BACTERIA. 47 

bad peptone in their cultivating medium. Martin examined the 
products by using as a culture medium a 1 to 2 per cent, solution 
of alkaU-albumin in broth made from beef, omitting the peptone. 
After about thirty days the baciUus had converted the alkali- albumin 
into albumoses, which gave the reactions of proto- and deutero- 
albumose, with small quantities of an organic acid. A single dose 
of these albumoses produced weakness of the hind hmbs, which after 
a time passed off. The animal was killed, and the nerves which 
were examined showed degeneration. Repeated intravenous in- 
jection on successive days, amounting in all to a dose of 1-69 grams 
per kilo, of body weight, produced high fever, followed by depi-es- 
sion of temperature, severe watery diarrhoea, and emaciation. The 
tendon reflexes began to diminish after the ninth day, on the 
eleventh or twelfth day there was definite paralysis of the hind legs, 
and on the seventeenth day reflexes could scarcely be obtained. 

Martin ^hus gives his method of abstracting the poisonous pro- 
ducts either from cultvires or from diphtheritic tissues. In deahng 
with tissues, the spleen and other organs are first finely minced and 
placed in rectified spirit, and the blood is also placed in spirit, and 
allowed to stand till the proteids are coagulated ; they are then 
filtered, and the residue extracted with cold water, all the extracts 
are mixed together, and evaporated at 35° 0. to a small bulk, and 
thrown into absolute alcohol. Most of the albumoses are precipi- 
tated, the alcohol is poured off, evaporated to dryness at a low 
temperature, and extracted by absolute alcohol until nothing more 
dissolves. The residue is deutero-albumose and mineral salts. All 
the proteid is mixed together, dissolved in water, and precipitated by 
alcohol, the process being repeated to remove any traces of bodies 
soluble in alcohol and the excess of mineral salts. At the last 
precipitation the precipitate is allowed to stand under alcohol for 
about two months. The alcohol is then poured off, and the pre- 
cipitate dried in vacuo. 

The resulting product is a light yellowLsh-brown powder soluble 
in water, cold or boiling, giving a yellowish and faintly acid or 
nearly neutral reaction. It is composed of deutero-albumose with 
a slight amount of proto-albumose but no peptone. It gives the 
ordinary actions of proteids and a well-marked biuret reaction. It 
is precipitated from solution by ammonium sulphate, and slightly 
by nitric acid. The reactions are similar to those of peptic deutero- 
albumose. The alcoholic extract of the tissues is strongly acid, and 
con,tains free fatty acid and an organic acid insoluble in chloroform. 
The organic acid is readily soluble in water and absolute alcohol, 



48 BACTKRIOLOGY. 

and insoluble in ether, chloroform, and benzine. It is a yellowish 
amorphous body, becoming a deep brown when made alkaline. 

Martin concludes that whereas the Bacillus anthracis produces 
albumoses and an organic base, in diphtheria we find albumoses and 
an organic acid. 

Glanders.^ — Kalming has obtained from cultures of the glanders 
bacillus an extract similar to tuberculin. This crude extract is 
known as mallein, and is extensively used for the diagnosis of 
glanders. In a glandered horse it causes a rise of temperature 
and swelling at the seat of the injection, and the glandered nodules 
become swollen and painful. Finger claims to have produced 
immunity from glanders by inoculation of the products contained 
in sterilised cultures. Schweinitz extracted from cultures a non- 
poisonous albumose, and obtained onl)' traces of a ptomaine. 

Suppuration and Pneumonia. — Brieger obtained a ptomaine 
from cultures of Staphylococcus pyogenes aureus, and Eoux and 
Yersin a tox-albumin fatal to rabbits and guinea-pigs in a few 
days. There was pus-formation at the seat of inoculation, with 
redness and swelling of the surrounding parts. 

From pure-cultures of the micrococcus of pneumonia Klemperer 
obtained a tox-albumin, for which the name pneumo-toxin has been 
suggested. 

Enzymes or Ferments. 

Many bacteria liquefy the nutrient gelatine m which they are 
cultivated. This is due to the development of a ferment or enzyme, 
which dissolves the albumin and gelatine. 

Enzymes are products of the vital activity of living bacteria. 
Bitter, and independently Sternberg, showed that when a liquefying 
bacterium is removed by filtration or destroyed by heat, the culture 
fluid retains the power of liquefying gelatine. As this occurs 
usually when the reaction is alkaline, bacterial enzymes resemble 
trypsin and papain rather than pepsin. They can be extracted with 
glycerine, and are quite harmless. If injected into animals no effect 
is produced, and after a few hours no trace of them can be found. 
According to Fermi, the influence of temperature on the enzymes 
produced by different bacteria will be found to vary very consider- 
ably. The enzyme of Staphylococcus pyogenes aureus is destroyed 
at 55° C, while the enzyme of Bacillus anthracis succumbs at n 
temperature of 65° 0. to 70° C. 

Some bacteria produce both enzymes and toxins, hut many pro- 
duce enzymes and not toxins, and others toxins but not enzymes 



CHAPTEE V. 

IMMUNITY. 

The condition of being insusceptible to an infective disease may be 
either natural or acquired. In studying the pathogenic organisms 
several examples of natural immunity will be encountered. The 
bacillus of septicaemia, so fatal to house mice, has been shown to 
have no eifect upon field mice. The bacillus of anthrax is innocuous 
to cats and white rats. The bacterium of rabbit septicaemia is 
equally inert in dogs, rats, and guinea-pigs. The immunity may 
be as in these cases complete, or only partial. Ordinary sheep are 
very easily affected with anthrax, but Algerian sheep succumb only 
to large doses of the virus. Natural immunity may not only be 
characteristic of certain species, but it may occur in certain indi- 
viduals of a susceptible species. The same immunity occurs in man, 
for certain individuals, though equally exposed during an epidemic 
of small-pox, may escape, whereas others readily fall victims to the 
disease. 

Acquired immunity is illustrated by the protection afforded by 
one attack of the exanthemata against subsequent attacks. Thus 
one attack of measles or small-pox, as a rule, affords complete 
protection. A knowledge of the immiinity resulting in the latter 
case led to the introduction of inoculation of small-pox as a 
protection against natural small-pox. 

Immunity may be acquired by acclimatization, for the inhabit- 
ants of tropical climates are less susceptible to the diseases of the 
country, malarial fevers, for instance, than strangers. 

In civihsed communities also, there appears to be a degree of 
acquired immunity, for infectious diseases like measles introduced 
among savages or isolated communities have assumed the most 
malignant type. 

The immunity acquired by protective inoculation constitutes, in 
connection with the study of pathogenic micro-organisms, a subject 
of pre-eminent interest and importance. Pasteur, in his researches 

49 4 



50 BACTERIOLOGY. 

upon fowl-cholera, observed that after non-fatal cases the disease 
either did not recur, or the severity of a subsequent attack was in 
inverse proportion to the severity of the first attack. It occurred 
to him to endeavour to obtain the virus of this disease in a form 
which would provoke a mild attack of the disease, and thus give 
protection against the virulent form. This attenuation or miti- 
gation of the virus was successfully attained by allowing cultiva- 
tions of the microbe in chicken-broth to remain with a lapse 
of several months between the carrying on of successive cultiva- 
tions in fresh media. The new generations which were then 
obtained were found to have diminished in virulence, and ultimately 
a viriife was obtained which produced only a slight disorder ; on 
recovery the animal was found to be proof against inoculation with 
virulent matter. The explanation given by Pasteur of this change 
was, that prolonged contact with the oxygen of the air was the 
influence which diminished the virulence, and he endeavoured to 
prove this by showing that when broth was inoculated in tubes 
which .could be sealed up, so that only a small quantity of air 
was accessible to the microbe, the virulence of the cultures was 
retained. 

Toussaint investigated the possibility of attenuating the virus of 
anthrax. Sheep injected with 3 cc. of defibrinated blood, con- 
taining anthrax bacilli, which had been exposed to 55° C. for ten 
miiiutes, recovered, and were afterwards insusceptible. Pasteur 
subsequently argued that this method did not admit of practical 
application, because difficulties would arise in dealing with infective 
blood in quantity, and artificial cultivations started from this blood 
could not be relied upon, as they proved sometimes as virulent as 
ever. 

Pasteur endeavoured to apply the same method for • obtaining 
an attenuated virus of anthrax, as he had successfully employed 
in fowl-cholera. A difficulty was soon encountered, for in culti- 
vations of this bacillus, with free access of air, spore-formation 
readily takes place, and the spores are well known to have an 
extraordinary power of retaining their virulence. Pasteur found 
that the bacilh ceased to develop at 45° C, and- he believed that 
spore-formation ceased at 42° to 43° C, the bacilH continuing to 
develop by fission only. The cultivations were, therefore, kept at 
this temperature, and at the end of eight days the ^bacilli were 
found to have lost their virulence, and were quite inert when 
incJculated in guinea-pigs, sheep, or rabbits. This total destruction 
was,-' however; preceded by a gradual mitigation j so- that a virus 



IMMUNITY. 51 

could be obtained, by taking it at the right time, which gave only a 
mild disease, and afforded subsequent protection. 

At Melun, in 1881, the protective inoculation against anthrax 
was put to a practical test. Sheep and oxen were inocula,ted with 
the mitigated virus, and then with a virulent form ; at the same 
time other sheep and oxen were inoculated with the virulent form 
without previous vaccination, as a control experiment. The unpro- 
.tected sheep died without exception ; the unprotected oxen suffered 
from oedematous swellings at the seat of inoculation, and a rise of 
temperature ; but all the protected animals remained healthy. 

As a result of these experiments an idea arose that by preventive 
inoculation with attenuated virus all communicable diseases would 
in time be eradicated ; but this does not follow, for all communi- 
cable diseases do not confer immunity after a first attack ;, in 
influenza the very reverse is believed to occur, and erysipelas of the 
face leads to an increased liability to subsequent attacks. Even 
with regard to the prevention of anthrax, Pasteur's researches were 
opposed and criticised. Koch investigated the subject, and came to 
the conclusion that the process did not admit of practical applica- 
tion, chiefly on the ground that as immunity lasted only a year, the 
losses from the vaccination process would be as great or even 
greater than from the spontaneous disease ; further, there was 
danger in disseminating a vaccine of the strength required to be 
effectual. 

Chauveau proved that the attenuation was due to the tempera- 
ture, and not to the prolonged effect of oxygen. By keeping 
cultivations at 42° to 43° C in vacuo, the virulence was found 
to disappear in twenty-four hours, and by keeping cultivations 
at a low temperature with free access of air, the virulence was 
retained. Chauveau considered, therefore, not only that oxygen was 
not the agent, but that the mitigation was much more easily effected 
in its absence. In spite of these adverse criticisms, these researches 
nevertheless confirmed the principle of Pasteur's conclusion, that 
immunity could be induced by experimental measures, and further 
showed that he had considerably advanced the methods by which this 
could be effected. 

Chauveau succeeded also in attenuating the virus by a modifica- 
tion of Toussaint's method. Sterilised broth was inoculated with 
the bacilli, and placed in the incubator at 42° to 43° C. After the 
lapse of twenty hours it was removed to another incubator at 47° C. 
According to the time of exposure to this increased temperature, the 
mitigation varied in degree. Thus inoculation with the virus,, before 



52 BACTERIOLOGY. 

it was exposed to 47° C, was fatal to gviinea-pigs ; but after one 
hour at 47° 0. the virulence was diminished, and, though ultimately 
fatal, life was prolonged ; after two hours' exposure at 47° C. only- 
half the animals died ; and after three hours' exposure they 
recovered, and were rendered refractory to subsequent inoculation. 

Attenuation of the virus of anthrax has also been induced by 
chemical means. Ohamberland and Eoiix stated that a fresh growth 
started from a cultivation of bacUli which had been subjected for 
twenty-nine days to ^^ of carbolic acid was found to be inert in 
guinea-pigs and rabbits. Bichromate of potash added to a cultiva- 
tion in the proportion of y^^inj- to j^^nnr 8*^6, after three days, a 
new growth, which killed rabbits, guinea-pigs, and half the sheep 
inoculated ; after ten days, rabbits and guinea-pigs, but not sheep ; 
and after a longer time even guinea-pigs were unaffected. 

In other diseases similar results have been obtained. Arloing, 
Cornevin, and Thomas found that by inoculating a small quantity 
of the virus of symptomatic anthrax anywhere in the subcutaneous 
connective tissue, or a moderate quantity at the root of the tail, 
and even by intravenous injection, immunity was obtained from a 
virulent dose. 

In swine-erysipelas, Pasteur and Thuillier obtained attenuated 
virus upon quite another principle. They discovered that by 
passing the virus through pigeons the virulence was increased, but 
by passing it through rabbits it was progressively diminished. Thus 
a virus was obtained from the rabbit, which produced only a mild 
disease in pigs, and after recovery complete immunity. Similarly 
in rabies, Pasteur found that passage of the virus through various 
animals considerably modified its properties. By inoculating a 
monkey from a rabid dog, and then passing the virus through other 
monkeys, the virulence was diminished ; but by inoculating a rabbit 
from the dog, and passing the virus from rabbit to rabbit, the 
virulence increased. 

In rabies, Pasteur has employed another method of attenuating 
the virus. The spinal cord of inoculated rabbits is removed with 
all possible precautions, and portions a few centimetres in length 
are suspended in flasks in which the air is dried by fragments of 
potash. By this process the virulence is found to gradually diminish 
and finally disappear. Animals inoculated with portions of these 
cords, after suspension for a certain time, are rendered refractory to 
inoculation with virulent cords. Having rendered dogs, which had 
been previously bitten, free from the supervention of symptoms of 
hydrophobia by means of protective inoculation, Pasteur proceeded 



IMMUNITY. 53 

to apply the same treatment to persons bitten by rabid animals, with 
results which tend to the beKef that a real prophylactic for rabies 
has been discovered. 

Immunity may also be produced by injecting the toxic products 
existing in pure cultivations after removal of the bacilli. Salmon 
was the first to produce immunity in this way, by utilising the toxic 
products of the bacterium of hog-cholera, which were separated by 
filtration from the living micro-organisms ; and shortly afterwards 
Wooldridge demonstrated that filtered anthrax cultures contained 
a substance which conferred immunity. Behiing and Kitasato 
produced immunity by mixing cultures with terchloride of iodine. 
Vaillard filtered the cultures through porcelain, and attenuated the 
products by heating at different temperatures. 

Lastly, in the course of Behring's and Kitasato's experiments, it 
was found that the blood serum of animals rendered immune was 
capable of conferring immunity on other animals. The injection 
of the toxic products of pathogenic bacteria leads to the development 
of substances in the blood to which the term " antitoxin " has been 
applied. These protective substances neutralise or destroy the 
injected poison, and blood serum which has thus been rendered 
antitoxic can be utilised to confer immunity on other animals. 

Haffkine's system of vaccination as a protection against Asiatic 
cholera is supposed to be based upon the principle of inducing the 
formation of antitoxins or defensive proteids. 

Mechanism of Immunity. 

Eaulin has shown that Aspergillus niger develops a substance 
which is prejudicial to its own growth, in the absence of iron salts 
in the nutrient soil, and Pasteur suggested that in rabies, side by 
side with a living microbe, there is possibly some chemical product 
or anti-microbe which has, as in Eaulin's experiment, the power of 
arresting the growth of the microbe. If we accept the theory of 
arrest by some chemical product, we must suppose that in the 
acquired immunity afforded by one attack of an infectious disease 
this chemical substance is secreted, and, remaining in the system, 
opposes the onset of the micro-organism at a future time. In the 
natural immunity of certain species and individuals we must suppose 
that this chemical substance is normally present. 

Another theory is, that the micro-organisms assimilate the 
elements which they require for their nutrition from the blood and 
tissues, and render the soil impoverished or otherwise unsuitable for 



54 BACTERIOLOGY. 

the development of the same species of micro-organisms hereafter ; 
this condition may be permanent, or the chemical constitution of 
the tissues may be restored to normal, when immunity ceases. If, 
however, we explain acquired immunity by the result of the growth 
of a previous invasion of micro-organisms, we are stiU confronted , 
with the difficulty of explaining natural immunity. 

A third theory is that the tissues are endowed with some 
power of vital resistance to the development of micro-organisms, 
similar to the vital resistance to coagulation of the blood, which is 
supposed to ejdst in the ■ lining membrane of the healthy blood- 
vessel ; that in some species and individuals this exists to a high 
degree, and hence their natural immunity. But this does not 
explain how one attack confers immunity from a subsequent one. 
One would expect that the vital resistance would invariably be 
lowered by a previous attack, and increased liability be the constant 
result. 

A fourth theory was propounded by Metchnikoff, who maintaiins 
that immunity depends upon phagocytosis. If anthrax bacilli are 
inoculated in the frog, white blood-cells, or phagocytes, are observed 
to incorporate and destroy them until they entirely disappear, and 
the animal is not affected. But if the animal, after inoculation, 
is kept at a high temperature, the bacilli increase so rapidly that 
they gain the upper hand over the phagocytes, and the animal 
succumbs. 

It has also been suggested that bacteria may attract or repel 
the phagocytes, exercising either a positive or a negative chemio- 
taxis. This power, is supposed to depend upon some special product 
of the bacteria, or possibly upon their toxins, as suggested by Roux. 
We must suppose that the negative chemio-taxis has become changed 
to a positive chemio-taxis in an immunised animal, so that the 
phagocytes, instead of withdra-\ving and leaving the bacteria to 
multiply, are readily drawn into the contest and destroy the 
invaders. 

In septicaemia of mice, the white blood-cells are attacked and 
disintegrated by the bacUli in a remarkable way. It is difficult, 
however, to accept these observations as affording a complete ex- 
planation of immunity. It is difficult to conceive that the leucocytes 
in the blood and tissues in the field mouse are differently constituted 
from those in the house mouse, so that they form an effectual 
barrier to the onset of bacteria in the one, case, though so readily 
desti'oyed in the other, or that in acquired immunity the result is 
due to ediioating the phagocytes to respond to a positive chemio-taxis. 



IMMUNITY. 55 

Phagocytosis cannot explain the immunity which results from 
the injection of filtered cultures, or of antitoxins, but when 
blood serum of immunised animals was shown to possess antitoxic 
properties, a new explanation of immunity was at once forthcoming. 
In the light of these discoveries immunity, whether natural or 
acquired, was regarded as due to the accumulation in the blood 
and tissues of substances which have the property of counteracting 
partially or entirely the" products by which pathogenic bacteria 
produce their poisonous effects. These antitoxins, or protecting 
proteids, can be obtained not only from the blood but also from 
the spleen and the lymphatic and other glands. They result 
from the metabolism of the cells of the tissues of the body. 
Phagocytes in their conflict with bacteria may play a small 
part, but it is more than probable that immunity is altogether 
independent of phagocytosis, 



CHAPTER VI. 

ANTITOXINS AND SERUM THERAPY. 

It has been clearly shown by the experiments of Fodor and Nuttall 
that some species of bacteria are killed by a mixtiu-e with fresh 
blood. Fodor pointed this out in the case of the anthrax bacUlus, 
and Nuttall confirmed the experiments, and repeated them with 
a number of different species of bacteria. 

Behring and Nissen followed up this line of inquiry, and found 
that there was a great difference in the behaviour of freshly drawn 
blood to different bacteria. In some cases the bacteria were 
destroyed, in others their growth was only retarded, and in others 
again they were not affected at all. Bouchard pointed out that 
although the normal blood serum of a rabbit may be used for the 
cultivation of Bacillus pyocyaneus, the blood serum of a rabbit, which 
has been rendered immune, will attenuate or entirely nullify the 
pathogenic properties of the bacillus. 

Ogata and Jasuhara obtained similar res\ilts by cultivating 
anthrax bacilli in the blood of immune animals. Buchner demon- 
strated that this property of fresh blood belonged to the serum 
and not to the cellular elements, and strongly advocated the theory 
that the force opposed to invading bacteria was to be found in the 
serum rather than in phagocytes. 

Similar experiments were made with the bacteria of swine-fever, 
and Emmerich and Mastbaum discovered that the blood serum of 
immune rabbits could be used as a therapeutic agent to prevent 
the progress of the disease in animals already showing symptoms 
of infection. 

A new light was thrown upon this question by the experiments 
of Behring, Kitasato, Tizzoni and Cattani, and others in connection 
with tetanus and diphtheria. In these diseases the bacteria do not 
invade the body, but the poisonous principles elaborated at the 
seat of inoculation are absorbed into the system and produce 
deleterious effects. It was obvious that attention must be turned 
towards counteracting or destroying these poisonous products. 

56 



ANTITOXINS AND SERUM THERAPY. 



57 



It was in this direction that the experiments of Behring and 
Kitasato, in 1890, proved to be of profound importance. It was 
shown that the blood serum of a rabbit rendered immune against 
tetanus or diphtheria had no destructive or retarding effect on the 
growth of the bapilli, but it possessed the power of neutralising the 
poison developed by the agency of the bacilli. In short, the serum 
was shown to possess an antitoxic instead of a bactericidal power. 

Hankin conceived the idea that this property is due to siibstances 
of the nature of defensive proteids, and the blood serum of the 
naturally immune rat was found to contain a proteid body with well- 
marked alkaline reaction, possessing the power of destroying anthrax 
bacilli. Injection of this proteid into mice, together with fuUy 
virulent anthrax spores, prevented the development of the disease. 
Young rats are susceptible to anthrax, and, according to Hankin, 
they can be protected from anthrax by injection of the blood serum 
of the parent. Tizzoni and Oattani expressed the opinion that the 
antitoxic substance in the blood serum of animals rendered immune 
against tetanus is a globulin to which they gave the name tetanus 
antitoxin. Buchner proposed the term alexins (d\ef<o, I defend), 
to signify these substances. Hankin subdivided them into sozins 
and phylaadns. Sozins are defensive proteids occurring in normal 
animals ; phylaxins are only found in animals artificially immune ; 
and each of these are sub-classed by Hankin according to their power 
of attacking the bacteria themselves or the products they generate; 



Defensive proteids 

(Hankin) 
Alexins (Buchner) 



Sozins : 
Defensive proteids . 
present in the nor- 
mal animal. 



'Myco-sozins : 

Alkaline globulins from rat 
(Hankin), destroying an- 
thrax bacillus. 

Toxo-sozins : 
Of rabbit, destroying poison 
of Vibrio Metchnikovi 
(Gamaleia). 



Phylaxins : 
Defensive 
present 



proteids 
in the 



animal after it has 
artificially been 
made immune. 



I'Myco-phylaxins : 

Of rabbit, destroying pig 
typhoid bacillus (Em- 
merich). 

Toxo-phylaxins : 

Of rabbit, etc., destroying 
diphtheria and tetanus 
poisons (Behring and 
Kitasato, anti-toxin of 
Tizzoni and Cattani). 



Tizzoni and Oattani immunised dogs and other animals against 
tetanus, and employed the antitoxin as a therapeutic agent. Its 



58 . . BACTERIOLOGY. 

active substance was precipitated by alcohol. Behring, Kitasato, and 
Schiitz experimented with a view to conferring immunity upon 
horses. The cultures were mixed with terchloride of iodine, and 
injected at intervals of eight days, and the antitoxic power tested 
on mice. By using increasingly virulent cultures, the blood became 
increasingly antitoxic. 

Vaillard filtered tetanus cultures through porcelain, and heated 
the filtrate at gradually diminishing temperatures. The first in- 
jections were made with 10 cc, which had been raised to 60° C. for 
an hour, then a filtrate was used which had been heated to 55° C, 
and lastly, a filtrate which had been heated to 50° 0. The blood 
became antitoxic, and by injecting increasing quantities of wulent 
filtrates the antitoxic power was rapidly intensified, and animals 
which were injected with antitoxin of full strength possessed 
immunity many months afterwards. 

Roux and VaUlard introduced another method. Virulent cultures 
were filtered through porcelain, and the filtrate mixed with Gram's 
solution of iodine in iodide of potassium. To give immunity to a 
rabbit, 3 cc. of toxin with 1 cc. of Gram's solution were injected on 
the first day, and increasing doses of toxin mixed with increasing 
doses of Gram's solution on the following days. The same method 
was applied to horses, sheep, and cattle. The antitoxin was found 
not only in the blood, but in the urine and saliva, and in the 
milk in cows-. With cows and goats it is necessary to proceed 
with the utmost care; while horses, on the other hand, bear the 
injections well, and are therefore more suitable for this purpose. 
It is also very easy to obtain large quantities of blood from the 
horse by inserting a trocSir and cannula into the jugular vein. 

Frankel was the first to produce immunity against diphtheria 
by injecting guinea-pigs with toxin which ha,d been heated to 70° C. 
Behring mixed the toxin with terchloride of iodine, or employed small 
doses of pure toxin. Horses, sheep, goats, and dogs were rendered 
immune. 

Preparation of Diphtheria Antitoxin. 

For the preparation of diphtheria antitoxin Roux cultivates the 
diphtheria baciUus in alkaline broth with 2 per cent, of peptone, and 
by preference, in flasks in which the cultivating liquid can be exposed 
to a current of moist air at 37° 0. After about three weeks the 
culture is filtered through a Chamberland filter, arid if tested on 
a guinea-pig it will be found that Jg^ of a cc. will kill an animal 
weighing five hundred grammes in forty-eight hours. The diphtheria 



ANTITOXINS AND. SERUM THERAPY. 59 

toxin immediately before the injection is mixed with i of its 
volume of Gram's solution. This is used for several weeks, and 
afterwards only pvire toxin is injected. 

The horses employed for this purpose are animals no longer fit 
for work, and it is necessaiy to inject them first of all with mallein 
to be sure that they are not suffering from glanders. 

In a horse inoculated by Roux, the injection began with | cc. 
of iodised toxin, increased to 1 cc. by the thirteenth day, and the 
injection continued daily. On the seventeenth day \ cc. of pure 
toxin was injected, and this was increased by the forty-first day 
to 10 cc. ; and on the forty-third day 30 cc. of pure toxin were 
injected, causing pronounced oedema. The doses were still further 
increased, untU on the eightieth day 250 cc. were injected. In 
two months and twenty days the horse had received 800 cc. of 
toxin. 

On the eighty-seventh day the serum obtained had an immunising 
power of over 50,000. By this is meant that a guinea-pig resisted 
inoculation of j cc. of virulent diphtheria culture when injected 
twelve hours beforehand with serum in quantity equal to the -gxjwff 
part of its body weight. 

There are two tests which can be appKed to the serum. Pirst, 
the antitoxic serum added to diphtheria toxin rendei'S it inert ; and, 
secondly, if serum is injected into a guinea-pig and toxin injected 
several hours afterwards, no result follows. 

Several ways have been suggested for estimating the ' immunising 
power of the serum. , 

In Ehrlich's system, the unit is represented by '1 cc. of anti- 
toxic serum, which, added to '8 cc. toxin, will neutralise it so that 
the whole may be injected subcutaneously in a guinea-pig without 
producing cedema. The standard toxin is a toxin of which -3 cc. 
is fatal to 1 kilo, of guinea-pig. 

But the preventive power of the serum is best expressed by the 
result of a subsequent injection of toxin. The immunising power is 
estimated by the number of grammes of guinea-pig which can be 
protected against the minimum fatal dose of toxin by 1 cc. of anti- 
toxic serum. 

The antitoxic serum can be kept in sterilised flasks in the dark, 
with the addition of a small piece of camphor, or it may be dried 
in vacuo, powdered, and thus supplied in a convenient form for trans- 
port. It has n>prely to be dissolved in water before use. 

Klein employed a modified plan by which he claimed to have 
obtained antitoxin in afar shorter time i than is possible by Roux's 



60 BACTERIOLOGY. 

method. Unfiltered attenuated cultures were injected into the 
horse. Later, large quantities of living diphtheria bacilli from 
the surface of solid cultures, of gradually increasing virulence, were 
repeatedly injected so as to allow the bacilli to grow and multiply. 
In twenty-three days an antitoxic serum was obtained, one part 
of which was found capable of protecting 20,000 to 40,000 
grammes of guinea-pig against more than a fatal dose of both living 
bacilli and the resulting toxin. 

Serum Treatment of Diphtheria. —The results obtained by 
Behring, Ehrhoh, Kossel, and Wasserman, in the^ treatment of 
diphtheria in children in Germany by means of the curative serum, 
and by Eoux and others in France, led to the adoption of the treat- 
ment in this country. It is best to use an especially constructed 
hypodermic syringe, which can be easily taken to pieces, and placed 
in boiUng water to sterilise it. The skin surface of the flank is 
washed, and disinfected with 1 in 20 carbolic, and the antitoxin is 
then injected. The syringe is taken to pieces, placed again in boiling 
water, and thoroughly cleaned. 

The dose will depend upon the age of the patient and the 
strength of the serum. From 10 cc. to 20 cc. are injected in children 
under fifteen, and 30 cc. to 40 cc. in older patients,, and the 
injection may be repeated in 12 hours. The best results are said 
to be obtained by injecting every 12 hours, for the first 12, 36 or 
48 hours, according to the nature of the case, 1,000 Behring's units, 
this being the dose calculated according to the immunising power of 
the serum. The result of the injection is to lower the temperature 
and pulse, but frequently the reverse occurs, and in about half the 
cases an urticarial and sometimes a scarlatiniform rash is produced. 
Pains in the joints, in rare cases effusion, may also result from the 
injection. 

The beneficial results of the treatment are, according to the 
Report of the Medical Superintendents of the hospitals of the 
Metropolitan Asylums Board, as follows : — ■ 

(1) Diminution of the fauoial swelling and of the consequent 
distress ; 

(2) Lessening or entire cessation of the irritating and offensive 
discharge from the nose ; 

(3) Limitation of the extension of membrane ; 

(4) Earlier separation of the exudation ; 

(5) Limitation and earlier separation of membrane in laryngeal 
cases ; 

(6) Improvement in general condition and aspect of patients • 



ANTITOXINS AND SERUM THERAPY. 61 

(7) Prolongation of life, in cases which terminate fatally, to ah 
extent not obtained with former methods of treatment. 

Statistics have also been brought forward which show, assuming 
them to be reliable, a great reduction in the mortality after the 
antitoxin treatment. A few instances may be quoted to illustrate 
the statistical evidence. 

According to Behring, in the four years prior to the employment 
of antitoxin, there were in Berlin 15,958 cases of diphtheria, with a 
mortality of 35'2 per cent. In 1894-5 there was an epidemic of 
5,578 cases. Behring asserts that if the mortality had not been 
reduced by the antitoxin treatment 1,963 would have died instead 
of 1,056. Behring also states that in the Oharite Hospital there 
were 299 patients, with 53 deaths, or 16'7 per cent. In the Bethania 
Hospital, where antitoxin was not employed, there were 249 
patients, with 112 deaths, or 43 per cent. 

At Vienna, at the Anna Hospital for children, the mortalit}' 
in 760 cases was 50'65 per cent., but after the introduction of anti- 
toxin there were 40 deaths in 159 cases, giving a mortality of 
25'5 per cent. 

In New York, it is said that before the introduction of anti- 
toxin the mortality ranged from 30'67 to 37'34, while in 1895, under 
treatment with antitoxin, the mortality fell to 19 "43; but it was 
also pointed out that since the introduction of antitoxin many 
children with trifling attacks had been treated, and reported as 
suffering from actual diphtheria, and that they would have recovered 
without antitoxin, and therefore these cases hg,ve_ given the remedy 
some credit which it does not deserve. 

In London, according to the Report of the Medical Superin- 
tendents of the hospitals of the Metropolitan Asylums Board there 
were in 1894, before antitoxin was employed, 3,042 cases of 
diphtheria with 902 deaths or 29-6 per cent., and in 1895, when 
antitoxin was used, 3,529 cases with 796 deaths or 22"5 per cent. : 
a reduction of 7'1 per cent, below that of 1894. The conclusions 
drawn from the statistical and clinical observations are summed up 
in the Eeport thus : — 

The improved results in the diphtheria cases treated during the year 
1895, are :— 

(I.) A great reduction in the mortality of cases brought under treat- 
ment on the first and second day of iUness. 

(II.) The lowering of the combined general mortality to a point 
below that of any former year. 

(III.) The still more remarkable reduction in the mortality of the 
laryngeal cases. 



62 BACTEEIOLOGY. 

(IV.) The uniform improvement in the results of tracheotomy at 
each separate hospital. 

(V.) The beneficial effect produced on the clinical course of the 
disease. 

A consideration of the statistical tables and clinical observations, 
covering a period of 12 months and embracing a large number of cases, 
in our opinion sufficiently demonstrates the value of antitoxin in the 
treatment of diphtheria. 

It must be clearly understood, however, thaj; to obtain the largest 
measure of success with antitoxin it is essential that the patient be 
brought under its influence at a comparatively early date— if possible not 
later than the second day of disease. From this time onwards the chance 
of a successful issue will diminish in proportion to the length of time 
which has elapsed before treatment is commenced. This, though 
doubtless true of other methods, is of still greater moment in the case 
of treatment by antitoxin. 

Certain secondary effects not infrequently arise as a direct result of 
the injection of antitoxin in the form iii. which it has at present to be 
administered, and, even assuming that the incidence of the normal com- 
plications of diphtheria is greater than can be accounted for by the 
increased number of recoveries, we have no hesitation in expressing the 
opinion that these drawbacks are insignificant when taken in conjunction 
with the lessened fatality which has been associated with the use of this 
remedy. 

We are further of the opinion that in antitoxin serum we possess 
a remedy of distinctly greater value in the treatment of diphtheria than 
any other with which we are acquainted. 

On the other hand it has been urged that the decline in the 
mortality in 1895 in London, which has been attributed entirely 
to the antitoxin treatment, may possibly be partly due to the pre- 
valence of a mild type of the disease, and that the fall in the 
mortality during the seven pre^T-Ous years from 59 per cent, in 
1888 to 29 per cent, in 1894, continued in 1895. 

It is obvious that the whole subject requires to be very carefully 
considered, and before any final conclusion can be arrived at as to 
the therapeutic value of antitoxin, the evidence of others who have 
had great experience in the treatment of diphtheria by the old and 
the new methods must be taken into account, and reliable statistics 
allowed to speak for themselves. 

Prepaeation of Tetanus Antitoxin. 

Antitoxin for use in the serum treatment of tetanus is obtained 
from the horse. The tetanus bacillus is cultivated in an atmosphere 
of hydrogen, in flasks specially constructed for the purpose. In 



ANTITOXINS AND SERUM THERAPY. 63 

about a fortnight the cultures are extremely toxic. The toxin is 
obtained free from bacilli by filtration through porcelain. Injec- 
tions may be given daily, subcutaneously or intravenously, beginning 
with 1 cc. of iodised toxin, and gradually increasing the dose until 
the pure toxin may be injected without danger. 

. Roux and Vaillard produced immunity in about three months. 
When a few days have elapsed after the last injection, the blood is 
drawn, by means of a trocar and cannula, from the jugular vein into 
a sterilised glass vessel, and set aside to coagulate ; next day the 
serum is drawn off with a pipette, and used in the liquid state, or 
dried in a vacuum over sulphuric acid, and subsequently powdered. 
When required for use the powder is dissolved in cold water. About 
5 grammes are used for a dose. 

Serum Treatment of Tetanus.— The result, so far, of the 
employment of tetanus antitoxin in animals suffering from tetanus 
is disappointing, and the serum treatment is not likely to be of much 
value in veterinary practice. Nocard infected sheep with tetanus 
by inserting splinters of wood infected with spores into the muscles 
of the leg. Tetanus supervened in eleven days, and the splinters 
were removed, the tissues excised, and the wounds dressed with 
iodoform. About twelve hours after the symptoms had shown 
themselves, the sheep were inoculated with antitoxic serum at 
intervals of one hour, but they all succumbed to tetanus. In one 
case the total amount injected was 160 cc. of highly antitoxic serum. 

The antitoxin has been employed in tetanus in man. Kanthack 
has collected the history of a number of cases, and they indicate 
that the treatment is useless in acute cases in man with a short 
incubation period, while chronic cases with a long incubation period 
often recover after the treatment. At the same time it must be 
remembered that recovery often took place in chronic cases before 
the introduction of the antitoxin treatment. 

The question must still be considered to be sub judice, and a 
trustworthy conclusion can only be based upon a more extended use 
■of the antitoxin and impartial reports of every individual case. 

Antitoxin op Septic Infections. 

An anti-streptococcic serum has been prepared by Marmorek. A 
culture of streptococcus was intensified in virulence by inoculation 
from rabbit to rabbit, and highly virulent cultures gave rise to a 
powerful toxin. E-oget and Oharrin also, found that the serum of 
immunised rabbits and of a horse conferred immunity, A patient 



64 BACTERIOLOGY. 

with puerperal fever was injected with 8 cc, on the follow- 
ing day with 16 cc, and on the third day with 25 ce. On the 
fourth day the temperature had fallen, and the patient recovered. 
Favourable results are said to have followed the use of the serum 
in 46 cases of erysipelas. 

Bokenham, working independently, cultivated the streptococcus 
in a mixture of broth and serum. Horses and asses were inocu- 
lated, and a considerable degree of immunity established. The 
serum of an inoculated ass possessed antitoxic power. 

Ruffer and Bullock succeeded in immunising four horses against 
the toxin of Streptococcus pyogenes ; two had been previously immu- 
nised against the toxin of the diphtheria bacillus. The streptococcus 
was cultivated by Marmorek's methods in a mixture of two parts 
of blood-serum and one part of peptonised broth, and the virulence 
of cultures maintained by inoculation of rabbits. On testing the 
immunising power of the antitoxic serum on rabbits, the effect 
appeared to be shght in comparison with the antitoxins of the 
bacilU of diphtheria and tetanus. In treating cases of septic infection 
in the human subject, it has been recommended to commence with 
two injections of 10 cc, and;it is said that no unfavourable results 
have been met with which could be attributed to the effect of the 
serum. 

Antitoxin of Typhoid Fever and Other Diseases. 

An antitoxic serum has been obtained by Ohantemesse for use in 
cases of typhoid fever, and it is probable that attempts will be made 
to extend the principle of the antitoxic treatment to other infective 
diseases. 



CHAPTEE VII. 

THE BACTERIOLOGICAL MICROSCOPE. 

The instruments sometimes in use in biological and pathological 
laboratories are not sufficient for the study of bacteria. It is 
absolutely essential for the examination of such minute objects that 
the microscope should be equipped with an objective of sufficiently 
high magnifying power and with a special illuminating apparatus, 
while the mechanical arrangements of the stage must admit of the 
examination of plate-cultivations. It would not be within the scope 
of this work to give a detailed account of the mechanical arrange- 
ments and optical principles of the microscope. These matters are 
fully dealt with in special works on the subject,* but sufficient will 
be said to afford assistance in the selection of a suitable instrument, 
and to explain the improvements in the microscope which have been 
such an aid in bacteriological investigations. 

A magnified image of an object is the result of the change 
produced in the direction of rays of light which are made to pass 
through lenses. This alteration in the course of the rays is known 
as refraction. A ray of light passing from a larer into a denser 
medium is refracted towards a hne drawn perpendicularly to the 
surface of the latter. A ray of light passing through air and 
impinging on water will not pass on in the same direction, but will 
be refracted towards a line drawn perpendicularly towards the 
surface of the water. If the ray pass into glass instead of water 
a greater refraction will take place, and if it pass into diamond the 
bending in its course will be still greater (Fig. 11). 

The sines of the angle of incidence and refraction of different 
substances have a constant ratio to each other, which is known 
as the index of refraction, and this is determined for different 
substances by the refraction produced by the passage of rays from 
a vacuum. Thus the index of refraction for flint glass is about 1'6, 

* Carpenter ; TJie Microscope, Nageli and Schwenderer : Tlw Microscope 
in Theory and Practice. 

65 .5 



66 



BACTERIOLOGY. 



the sine of the angle of incidence of a ray passing from a vacuum 
into glass being to the sine of the index of refraction as 1-6 to 1. 

If we study the course of a j^encil of rays we find that some 
of the rays are reflected instead of entering the medium and bemg 
refracted. When, for example, a pencil of rays falls upon water 
or glass, after passing through air, some of the rays are lost by 
reflection, and the proportion of the lost rays will increase with 
their obliquity. The diminution of the brightness of the image 
when pencils of rays have to pass through lenses is thus accounted 
for, and this loss of light increases when the number of surfaces 




Fig. 11. — The Refeaction of Light. 



through which the rays pass are, as in high-power objectives,, 
increased. There is an additional loss when there is an increase in 
the difference between the refractive power of the different media 
through which light passes. When pencils of rays pass from glass 
into air, and then into glass again, the loss is much greater than 
when the air is I'eplaced by a medium with a refractive index more 
neai'ly approaching that of glass. This explains the value of the 
immersion system, which will be referi'eil to more fully later on, 
and also the advantage of cementing pairs of lenses with Canada 
balsam or glass paste. The lenses used in the optical arrangements 



THE BACTERIOLOGICAL MICROSCOPE. 



^7 



of a microscope are principally convex, and the imperfections whick 
result mnst, if possible, be entirely overcome. These imperfections 
are spherical and chromatic aberration. 

Spherical aberration results from the unequal refraction of 
rays passing through lenses with equal curvatures. The rays passing 
through an ordinary convex lens do not all come to the same focus.. 
The rays passing through the marginal portion come to a focus at a 



R 


N 


^ 






n^ 


^^ 


f'f 






A~~- 






1 




.-^H 




R^ 


/ 


^ 




Ml 


v 





Fig. 12.— Spheeical Aberbation. 

point much nearer to the lens than the focus of the rays passing 
through the more central portion of the lens (Fig. 12). If the whole 
aperture of the lens is used there must of necessity be bkirring, for 
at the point at which the marginal rays form a distinct image the 
central rays will be out of focus, and at the point at which the 
central rays form a distinct image the marginal rays will have 
diverged, causing indistinctness. 

This is partially remedied by using a diaphragm 
and shutting out the marginal rays ; but this is 
at the cost of loss of light .and diminution of the 
angle of aperture. The difficulty is approximately 
overcome in practice by using a combination of 
lenses. The aberration of a convex lens is the 
opposite of that of a concave lens (Fig. 13). The 
makers of the best lenses endeavour to obtain this 
correction as perfect as possible to get the sharpness 
of the image, so essential in studying the mor- 
phology of bacteria. 

Chromatic aberration is the result of tke 
unequal refrangibility of the coloured rays which 
compose white kght. If parallel rays of light pass through a 
convex lens the violet rays, which are the most refrangible, will 
come to a focus at a point much nearer to the lens than the 
focus of the red rays, which are the least refrangible ; and the 
intermediate rays of the spectrum will be focussed at points between 
the red and the violet. A screen held at either of these foci 
will show an image with prismatic fringes (Fig. 14). 




Fig. 13.— Com- 
bination o F 
Lenses in 
Abbe's Homo- 
geneods im - 

MEKSION. 



68 BACTERIOLOGY. 

The chromatic aberration may be reduced by stopping out the 
marginal rays; but as it is necessary to get the most perfect 
correction possible, advantage is taken of the different relations 
which the refractive and dispersive powers bear to each other in 
different glasses. By combining a double convex lens of crown 
glass with a plano-convex lens of flint glass, correction is obtained 
for the violet and red rays. An achromatie objective is constructed 
on this principle, but the result is not perfect, as the intermediate 
coloured rays remain uncorrected, and what is termed a seconda/ry 
specia-um gives rise to images with coloured fringes, especially at the 
margin of the field. Abbe and Schott, after a great number of 
experiments, succeeded in discovering a glass with optical properties 
which removed the secondary spectrum, and objectives made with 
the new glass are termed apo-chromatic. There is much more 



A i 


Ib' ^"^ 




-^' I 




A'' 1 


If 


■ ^^""^---^ 







Tis. 14. — Chkomatic Abeebation. 

perfect concentration of the component rays than in the ordinary 
achromatic objectives, and the advantages thus obtained are very 
great. The objectives can be made of higher angle and admit of 
higher eye-pieces being used without materially diminishing the 
brilliancy and definition of the image. There is a complete absence 
of coloured fringes, and the perfect definition is invaluable in 
micro-photography. 

Another fault which has to be corrected is the aberration caused 
by covering a microscopical preparation with a cover-glass. Ross 
was the first to point out -the difference in the image when the 
object was examined under a cover-glass, and that by altering the 
position of the front pair of lenses, in an objective corrected for an 
uncovered object, the objective could be corrected for the covered 
object (Fig. 15). 

Objectives are generally corrected for a standard thickness of 
cover-glass, but H. Lister devised a screw-collar adjustment by 
which the position of the front pair of lenses could be altered at 
will ; and as it is almost impossible to obtain cover-glasses which 



THE BACTERIOLOGICAL MICROSCOPE. 



69 



do not vary slightly in thickness, the most perfect definition 
can only be obtained by adjusting for each separate cover-glass 
preparation. 

Immersion system. — All objectives were formerly used dry — 
that is to say, with an air space between the objective and the 
specimen to be examined — but high-power objectives are now almost 
entirely made on the immersion S3'stem, a drop of liquid being 
interposed between the objective and the cover-glass. 

About fifty years ago Amici observed that if a drop of water 
intervened between the cover-glass oi' an uncovered oljject and the 
lens the image was more brilliant. The passage of raj's from the 
object or the cover-glass into air, 
and again from air into glass, caused 
considerable loss of light. With 
objectives of wide angle of aperture 
the advantages were counteracted 
by the reflection of rays falling ob- 
liquely upon the lens. Hj inter- 
posing water more rays are bent 
in or refrticted, and enter the lens 
instead of being reflected and lost. 

Hartnack, Nachet, and others 
adopted the immersion system, and 
high-power water immersion lenses 
were constructed with high angle 
of aperture.* It was found that there was less necessity for 
correcting for covers of different thickness, as the aberration from 
this cause was diminished. The lenses were corrected for an average 
thickness of cover, and slight deviations produced hardly any 
appreciable effect. 

Wenham, Stephenson, Abbe, and Zeiss carried the system to 
perfection. They argued that the advantages obtained by water 
immersion would be intensified if a liquid could be found of the 
same refractive and dispersive power as crown glass. The media 
would be optically uniform, and the residt a homogeneous immersion 
system. 




Fig. 1.5. — Objective with Collar 

GOERECTION (h). 



* The angle of aperture is " the angle made by the most diverging of the 
rays of the pencil issuing from any point of an object that can enter the lens, 
and take part in the formation of an image of it." 

The numerical aperture is defined by Abbe as equal to " the sine of the 
angle of aperture multiplied by the refractive index of the medium between 
the object and the objective." 



70 BACTERIOLOGY. 

After experimenting with different liquids — solutions of salts, 
and various essential oils — Abbe recommended cedar oil as most 
suitable for the purpose. In its optical properties it very closely 
resembles crown glass, and it is far more convenient for use than 
any watery solutions of salts, especially when it is necessary to 
make a more or less prolonged examination of an object. 

The difference between the dry, water, and oil immersion systems 
may be illustrated, as Frankel has pointed out, by a v6ry simple 
experiment. If a glass rod is inserted into an empty test-tube, it 
is easily visible owing to the difference in refraction between the 
glass and the surrounding air. If the tube is filled up with 
water the rod is seen with difficulty, and if, instead of water, cedar 
oil is used, the part of the rod immersed in the oil will entirely 
disappear from view. The rays of light pass through an optically 
uniform medium in the experiment with cedar oil, and no refraction 
or reflection of rays of light can occur. 

To use an oil immersion objective, a minute drop of cedar oil 
is placed on the centre of the cover-glass, and the lens lowered 
by means of the coarse adjustment until it touches the oil. The 
specimen is then carefully brought into focus with the fine adjust- 
ment. If the slide is held between the finger and thumb of one 
hand, and moved from side to side while the other hand is working 
the fine adjustment, there can be no danger of injuring either the 
objective or the specimen. 

Microscopes are made upon either the Ross or the Jackson 
model. In the Ross model the body of the microscope is fixed 
at its base to a transverse arm, which is raised or lowered with 
it by the rack and pinion. In the Jackson model the body is 
supported for a great part of its length on a solid " limb." 

In the Ross model, unless the body and transverse arm are very 
solid as in Powell and Lealand's microscopes (Fig. 23), there will be 
vibration at the ocular end ; but in the Jackson model vibration is 
practically prevented, and this is most essential, especially in working 
with very high powers. 

The steadiness of the microscope also largely depends upon the 
form of stand. There are four different types of stands. The 
tripod (Fig. 23) ; the plate, with double columns ; the single column, 
ending in a plate or a bent claw; and the horse-shoe (Fig. 18). 

The tripod stand with cork feet is the steadiest form of stand, 
but it is cumbrous and expensive, and these objections also apply to 
the model made by Ross. 

The single upright should be unquestionably condemned, as it 



THE BACTERIOLOGICAL MICROSCOPE. 



71 




Fig. 16. — Bnalish Model. 



72 



BACTERIOLOGY. 



freely admits of vibration, and is most inconvenient for laboratory 
work, The heavy horse-shoe form is compact and firm, and the 
weight of it can hardly be considered an objection. 

The tubular body is from eight to ten inches in length, and within 
it is a draw-tube with engraved scale. By extending the draw-tube 
greater magnification is obtained; but as this is at the cost of 
definition it should hardly ever be used in the examination of 

bacteria. 

A triple nose-piece is a great convenience, saving the time which 
is otherwise spent m replacing objectives of different magnifying 
power, and there is less risk of injuring them. 

Foats should be obtained by means of a rack and pinion coarse 




Fig. 17. — Removable Mech-inical Stage. 



adjustment. The sliding tube is not to be recommended, as the 
motion may be stiff, encouraging the use of force, which in turn may 
result in the objective being brought violently into contact with the 
specimen, injuring the lens or damaging the preparation ; or it may 
get too loose and readily slip out of focus. 

The stage should be flat and rigid, either rectangular or circular, 
so long as it is sufficiently large to accommodate a plate- cultivation. 
A removable mechanical stage is of great advantage for working 
with high powers, as a motile bacterium can be constantly kept in 
view while one hand is engaged in working the fine adjustment 
(Fig. 17). It may also be employed as a finder if it is engraved 
with a longitudinal and vertical scale, and provided with a stop. 
The mechanical stage must be removable, so that the stage proper 



THE BACTERIOLOGICAL MICROSCOPE. 



73 








Tig. 18. — Continental Model. 



74 



BACTERIOLOGY. 




may be free from any attachments when required for the examina- 
tion of cultures. 

Diaphragms are necessary for regulating the amount of Kght. 
The plan of using a series of discs, with apertures of different sizes, 
should be avoided, as they are easily lost, and bacteriological investi- 
gations may have to be made under conditions in which it is difficult 
to replace them. A better plan is a revolving plate with apertures 
of different sizes, but the most convenient form is the iris diaphragm 
(Fig. 19). 

The sub-stage condenser is quite as necessary in bacteriological 
work as a high-power objective. In fact, the condenser and the 
objective should be considered as forming one optical apparatus, and 

the microscope regarded quite as • 
incomplete without a condenser as 
it would be without an objective. 

By means of the sub- stage con- 
denser (Fig. 20) the rays of light 
are concentrated at one point or 
on one particular bacterium ; and 
for the best definition it is essential 
that there should be niechanic.iJ 
arrangements for accurately cen- 
tring and focussing the condenser. 
It may even with advantage be provided with a fine adjustment. 

To sum up, a microscope for bacteriological investigation should 
be provided with (1) a steady stand of either the tripod or horse- 
shoe form ; (2) a tubular body on the Jackson model ; (3) a -ivide- 
angled sub-stage condenser, such as Abbe's ; (4) objectives of an inch, 
■j-th of an inch, and a yVth homogeneous immersion ; (5) a removable 
mechanical stage ; and for the most accurate work there should be 
centring arrangements and a coarse and fine adjustment to an oil- 
immersion sub-stage condenser such as Powell and Lealand's, and 
a vgth homogeneous oil-immersion apo-chromatic objecti\'e. 

With regard to the choice of a microscope, it is chiefly a 
question of price. The most perfect instrument is the large model 
by Powell and Lealand, but it is most expensive, and C|uite unsuit- 
able for laboratory work. For general use excellent instruments 
are made by Zeiss, Leitz, Eeichert, or Swift. The bacteriological 
microscopes of these makers are in the necessary equipment 
practically identical. The Zeiss microscope is the most finished, and 
costs about twenty pounds. A similar microscope by Leitz and 
by Swift costs about eighteen, and both make an excellent students' 



FlU. 19. — Ims DiAPHR.VGM. 



THE BACTERIOLOGICAL MICROSCOPE. 



75 



bacteriological microscope, with a cheap form of adjustment to the 
sub-stage condenser, at a total cost of about fifteen pounds. 

Method of Illumination. — Good daylight is the best for 
general work. The microscope should be placed near a window 
with a northern aspect. Direct sunlight should never be utilised, and 
the best light is that reflected from a white cloud. When daylight 
is not available good results can be obtained with either gas or a 




Fig. 20.— Abbe's Condenser consteucted by Zeiss. 



paraifin lamp. In the author's laboratory the microscope lamps 
are fitted with Welsbach incandescent mantles. These have many 
advantages over an Argand burner or a paraffin lamp. A steady 
and beautifully white light is obtained, and the lamps are quickly 
lit, and require comparatively little attention. In using high powers 
and carefully focussing the sub-stage condenser, the image of the 
fabric of the mantle is embarrassing, and is an objection to this 
light for the most accurate observations, but in other resjjects, and 



i 



76 



BACTERIOLOGY. 



for general use, it is the best form of artificial illumination for the 
microscope. 

An ordinary parafiin lamp of the cheapest form may be used, 
but there are many objections to it, such as the shape of the 
chimney, and the strise and defects in the glass. The best form of 
paraffin lamp is constructed by Baker and by Swift from sug- 




FiG. 21. — MICBOSCOPE Lamp. 



gestions by Nelson and Dallinger (Fig. 21) ; there is also a similar 
but much larger pattern which is made by Swift (Fig. 22). This 
form of lamp has a large flat bowl for the oil. It is attached to 
a standard, and can be raised or lowered to the desired position. 
The chimney is of metal and blackened, so that there is no reflected 
light, and it may also with advantage be provided with a shade, 
so that no light reaches the eye except through the microscope. 

The burner may be made to revolve, so that either the edge or 



THE BACTERIOLOGICAL MICEOSCOPE. 



77 



the flat of the flame may be utiUsed. Great care should be taken 
to have the wick evenly trimmed. The best paraffin oil should 
be burnt, and it is as well to add a small lump of camphor. The 
metal chimney has an aperture in front, giving exit to the rays of 
light, which is closed in by a slip of glass. The glass is very liable 
to crack when exposed to the full force of the flame, and it is as well, 
therefore, to be provided with a stock of glass slips, which have 




Fig. 22. — Large Mickoscope Lamp. 



been annealed by being enveloped in a cloth and boiled for two or 
three hours. 

The flat of the flame is used with low powers. The image of 
the flame is reflected by a plane mirror, and a bull's-eye condenser 
interposed between the lamp and the mirror to give an equal 
illumination of the whole fleld. In working with high powers the 
lamp is turned with the flame edgewise, and the mirror is dispensed 
with. By working, as it is termed, directly on the edge of the 
flame, the illumination is greatly increased, and a band of light can 



78 BACTERIOLOGY. 

be concentrated on that part of the microscopical preparation which 
requires most careful study (Fig- 23). 

To obtain the best definition considerable time must be spent in 
the arrangement of the illumination. The lamp and microscope 
having been placed in position, a low power is first used and the 
smallest diaphragm. On looking through the microscope it will 
probably be observed that the image of the diaphragm is not in 
the centre of the field. By moving the centring screw of the con- 
denser this may be adjusted. The image of the edge of the flame 
may not be central, and this must be adjusted by moving the lamp 
into position. The low power is then replaced by a high power, 
the largest diaphragm used, and the bacteria brought into focus. 
The diaphragm must now be replaced by one of medium size, and 
by racking the condenser up and down, a point will be arrived at 
when the image of the edge of the flame appears as an intensely 
bright band of light. If this is not exactly in the centre of the 
field the centring screws of the condenser must again be adjusted. 
Lastly, by trying different sizes of diaphragms, and focussing with 
the fine adjustment, and using the correction collar, we arrive at the 
sharpest possible image of the bacteria. 

When the condenser has been accurately centred, it will still be 
necessary to focus it for each individual specimen, so as to correct 
for difference in the thickness of slides and the layers of mounting 
medium. Correction for different thickness of cover-glasses must 
in each case be made by means of the collar adjustment in the follow- 
ing way. A high-power eye-piece is substituted for the ordinary 
eye-piece, and the fault in the image will thereby be intensified. By 
moving the cqllar completely round, first in one direction and then 
the other, while carefully observing the effect on the image, it will 
be seen to become obviously worse whichever way the collar is turned. 
The collar must then be turned through gradually diminishing dis- 
tances imtil an intermediate point is reached at which the best image 
results with the high-power eye-piece, and on replacing this by the 
low-power eye-piece the sharpest possible image will be obtained. 

Effect of the sub-stage condenser. — The sub-stage condenser 
gives the most powerful illumination when it has been racked up 
until it almost touches the specimen. It produces a cone of rays of 
very short focus, and the apex of the cone should correspond with the 
particular bacterium or group of bacteria under observation. The 
effect of the condenser without a diaphragm is to obliterate what 
Koch has termed the structure picture. If the component parts of a 
tissue section were colourless and of the same refractive power as 



THE BACTERIOLOGICAL MICEOSCOrE. 



79 







3 < 

S q 



^^ a 



Ph o 



:3 ^ 

5I 



80 



BACTERIOLOGY . 



the medium in which the section is mounted, nothing would be 
visible under the microscope. As, however, the cells and their nuclei, 
and the tissue fibres do differ in this respect, the rays which pass 
through them are diffracted, and an image of lines and shadows is 
developed. If in such a tissue there were minute coloured objects, 
and if it were possible to mount the tissue in a medium of exactly 
the same refractive power, the tissue being then invisible, the 
detection of the coloured objects would be much more easy. This 
is exactly what is required in dealing with bacteria which have been 
stained with aniline dyes, and the desired result can be obtained 
by the use of the sub-stage condenser. 

If we use the full aperture of the condenser the greatly converged 
rays play on the component parts of the tissue, light enters from 




Fig. 24.— Ramsden Micbometee Eye-piece. 



all sides, the shadows disappear, and the structure picture is lost. 
If now a diaphragm is inserted, so that we are practically only 
dealing with parallel rays, the structure picture reappears. As the 
diaphragm is gradually increased in size the structure picture 
gradually becomes less and less distinct, while the colour jjicture, 
the image of the stained bacteria, becomes more and more intense. 
When, therefore, bacteria in the living condition and unstained tissues 
are examined a diaphragm must be vised, and when attention is 
to be concentrated upon the stained bacteria in a section or in a 
cover-glass preparation, the diaphragm must be removed and the 
field flooded ^vith light. 

Micrometer. — For the measurement of bacteria a stage micro- 
meter may be used with a camera lucida". The stage micrometer 
consLsts of a slip of thin glass ruled with a scale consisting of tenths 
and hundredths of a millimetre. The image of this can be projected 



THE BACTERIOLOGICAL MICROSCOPE. 



81 



on a piece of paper, and a drawing made, and the object to be 
measured can then be projected on the paper and compared with the 
scale. 

In the Eamsden micrometer eye-piece (Fig. 24) two fine wires 
are stretched across the field of an eye-piece, one of which can be 
moved by a micrometer screw. In the field there is also a scale 
with teeth, and the interval between them corresponds to that of the 
threads of the screw. The circumference of the brass head is usually 
chvided into one hundred parts, and a screw with one hundred threads 
to the inch is used. The bacterium to be measured is brought into a 




Fig. 25. — Micrometeh Eye-piece by Zeis 



position in which one edge appears to be iu contact with the fixed 
wire, and the micrometer screw is turned until the travelling wire 
appears to be in contact with tlie other edge. The scale in the 
field and the scale on the milled head together give the number of 
complete turns of the screw and the value of a fraction of a turn in 
separating the wires. 

In the micrometer eye-piece constructed by Zeiss, the eye-piece 
with a glass plate with crossed Hues is carried across the field by 
means of a mici'ometer screw (Fig. 25). Each divi.sion on the edge 
of the drum corresponds to '01 mm. Complete I'evolutions of the 
drum are counted by means of a figured scale in the visual field. 
Another method of measuring bacteria will be referred to in the 

C 



82 - BACTERIOLOGY. 

chapter on micro-photography. The unit of measurement is one 
thousandth of a milhmetre or a micro-millimetre or micron, and is 
expressed by the Greek letter ju. 

CARE OF THE MICEOSCOPE. 

After use the objectives, sub-stage condenser, and eye-piece 
should be carefully wiped with soft linen, an old silk handkerchief, 
or chamois leather, and the microscope covered with a bell-glass to 
protect it from dust. If a lens comes into contact with Canada 
balsam it must be very carefully wiped with a soft rag moistened 
with alcohol, and then cleaned with a soft leather. Microscopes 
should not be exposed to the fumes of sulphuretted hydrogen, 
chlorine, or volatile acids. 



CHAPTER VIII. 

microscopical examination of bacteria. 

(a) Bacteria in Liquids, Cultures, and Eeesh Tissues. 

In conducting bacteriological researches the importance of absolute 
cleanliness cannot be too strongly insisted upon. All instru- 
ments, glass vessels, slides, and cover-glasses should be thoroughly- 
cleansed before use. A wide-mouthed glass jar should always be 
close at hand, containing refuse alcohol for the reception of re- 
jected slide preparations or dirty cover-glasses. When required 
again for use, shdes can be easily wiped clean with a soft rag. Cover- 
glasses require further treatment, for, unless they are perfectly 
clean, it is difficult to avoid the presence of air biibbles when 
mounting specimens. They should be left in strong acid (hydro- 
chloric, sulphuric, or nitric) for some hours ; they are then washed, 
first with water and then with alcohol, and carefully wiped with a 
soft rag. The same principle applies in the preparation and 
employment of culture media ; any laxity in the processes of 
sterilisation, or insufficient attention to minute technical details, 
will surely be followed with disappointing results by contamination 
of the cultures, resvilting in the loss of much time. 

For the preparation of microscopical specimens it will be found 
convenient to use a platinum inoculating needle. This consists of 
two or three inches of platinum wire fused into the end of a glass 
rod about eight inches in length. Platinum is employed as it 
rapidly cools after being raised to a white heat in the flame of a 
Bunsen burner. It is thus completely sterilised, and in a few 
moments is cool enough not to destroy the bacteria with which it is 
brought into contact. 

When using platinum needles, either for inoculating fresh tubes 
in carrying on a series of pure cultures, or in transferring a small 
portion of a cultivation to a cover-glass for examination under the 
microscope, the careful sterilisation of the needle by heating the 

83 



g4 BACrERIOLOGY. 

platinum wire till it is white hot in every part, and heating also 
as much of the glass rod as is made to enter the test-tube, must 
be carried out with scrupulous care. Indeed it is a good plan to 



Fig. 26.— Inocolatins Needles. 

let it become a. force of habit to sterilise the needle before and after 
use on every occasion, whatever may be the purposes for which it 
is employed. 

Unstained Bacteeia. 

The bacteria in liquids, such as pus, blood, and culture- fluids, can 
be investigated in the unstained condition by transferring a drop with 
a looped platinum needle or a capillary pipette to a slide, covering 
it with a clean cover-glass, and examining without further treat- 
ment. If it is desirable to keep the specimen under prolonged 
observation, a drop of sterilised water or salt solution must be run in 
at the mai'gin of the cover-glass to counteract the tendency to dry. 

Cultures on solid media can be examined by transferring a small 
portion with a sterilised needle to a drop of sterilised water on a 
slide, thinning it out, and covering with a cover-glass as already 
described. 

Tissues in the fresh state may be teased out with needles in 
sterilised salt solution, and pressed out into a sufficiently thin layer 
between the slide and cover-glass. Glycerine may in many cases 
be substituted for salt solution, especially for the examination of 
micro-organisms such as Actinomyces and mould fungi. 

There is, as a rule, no difficulty in recognising the larger micro- 
organisms such as those just mentioned ; but when we have to 
deal with very small bacilli and micrococci, they may possibly be 
mistaken for granular detritus or fat-crystals, or vice versa. They 
are distinguished by the fact that fatty and albuminous granules 
are altered or dispersed by acetic acid, and changed by solution 
of potash; alcohol, chloroform, and ether dissolve out fat-crystals 



MICROSCOPICAL EXAMINATION OF BACTERIA. 85 

or fatty partwiles ; on the other hand, micro-organisms remain 
unaffected by these reagents. Baumgarten demonstrated tubercle 
bacilli in sections by treating them with potash, which clarified 
the tissues and brought the bacilli clearly into view. Actinomyces 
and other vegetable structures will not disappear when sections are 
immersed in weak hydrochloric acid and mounted in glycerine. 

In examining unstained bacteria, it is necessary, in order to 
obtain the structure picture, that the light entering the microscope 
should be reduced by employing a small diaphragm, and the sub-stage 
condenser carefully centred and focussed. To focus an unstained 
specimen in which only bacteria are present, is often difficult. The 
shde may be gently I'aised towards the objective, and the stage 
may be constructed to enable this to be done with the index finger 
(Fig. 16). If on tilting the slide the organisms come into focus it 
will serve as a guide in working the fine adjustment. Another plan 
when bacteria are examined in water, is to look for an air-bvibble, 
and then to focus its edge until the bacteria appear in view. 

The simple method of covering the liquid with a cover-glass will 
not answer for a prolonged examination, as the liquid evaporates and 
the specimen dries up. To keep living bacteria under observation 
for any length of time, in order to study their movements or spore- 
formation, a special slide must be employed (p. 120). 

Stained Bacteeia. 

Weigert first pointed out the vakie of the aniline dyes for 
staining bacteria, and we are principally indebted to Koch, Ehrlich, 
Gram, and Loffler for many valuable processes. 

The staining of fresh preparations, especially those with no 
coagulable albumen to fix them, miay be carried out by the method 
of His. A slide is prepared as ali'eady described for the exami- 
nation of micro-organisms in the fresh state. The reagents are 
then applied by placing them with a pipette drop by drop at 
one margin of the cover-glass, and causing them to fiow through 
the preparation by means of a strip of filter-paper placed at the 
opposite margin. 

Bab^s recommends another rapid means of examining cultivations. 
A little of the growth, removed by means of a sterilised platinum 
hook or small loop, is spread out on a cover-glass into as thin a 
film as possible : when almost dry, a drop or two of a weak aqueous 
solution of methyl violet is allowed to fall from a pipette upon the 
film. The cover-glass with the drop of stain is, after a minute. 



86 BACTERIOLOGY. 

carefully turned over on to a slide, and the excess of stain gently 
and gradually removed by pressure with a strip of filter-paper. 
This affords a rapid means of demonstration — for example, of a 
cultivation of Koch's comma bacilli in nutrient gelatine — enabling 
the microbes to be seen in some parts of the preparation both 
stained and in active movement. 

Cover-glass Peepaeations. 

Bacteria may be spread out into a thin layer on a cover-glass, 
and then treated with a dye, or sections of tissues containing bacteria 
can be stained and then mounted in the usual way. 

The method of making a cover-glass preparation is one which is 
very commonly employed. In addition to its value as a means of 
examining bacteria in liquids and solid culture media, it affords 
the additional advantage of enabling, if necessary, a lar^e number 
of preparations to be made, which, when dried, can be preserved, 
stained or unstained, in ordinary cover-glass boxes ; they are 
then in a convenient form for transport, and can be mounted 
permanently at leisure. 

The method is as follows : A cover-glass is smeared with the 
cut surface of an oi-gan or pathological growth, or with sputum ; 
or a di'op of blood, pus, or culture-fluid is conveyed to it with a 
looped platinum needle. It is absolutely necessary to spread out 
the micro-organisms into a sufficiently thin layer, so that the 
individual bacteiia may be as much as possible in the same plane, 
otherwise some in the field will be in focus and others out of 
focus, and it would be impossible to obtain a satisfactory photograph 
of such a specimen. To overcome this it will be necessary, in the 
case of cultures on solid media, to diffuse the bacteria in a httle 
sterilised water ; and even cultures in liquids may sometimes with 
advantage bs diluted in the same way. By means of another 
cover-glass the jiiice or fluid is squeezed out between them into a 
thin layer, and on sliding them apart each cover-glass bears on one 
side a thin film of the material to be examined ; or a culture is 
spread out into a thin film by means of a hooked platinum needle. 
The cover-glass is then placed with the prepared side upwards, and 
allowed to dry. After a few minutes, it is taken up with a pair of 
flat-bladed or spring forceps, with the prepared side uppermost, and 
passed rapidly from above downwards three times through the 
flame of a spirit lamp or Bunsen burner. Two or three drops of 
an aqueous solution of fuchsine or methyl violet will be suflicient to 
cover the film, and after a minute or two the surplus stain is washed 



MICROSCOPICAL EXAMINATION OF BACTERIA. 87 

off witli distilled water by means of a siphon apparatus or a wash- 
bottle. The cover-glass may be allowed to dry, and then mounted 
in Canada balsam, or it may, while still wet, be turned over on to a 
slide, the excess of water removed with filter-paper, and the exposed 
surface wiped dry. It may first be examined with a power of about 
250 diams. ; and if a high magnification is required, which is usually 
the case, a droplet of cedar oil is placed on the cover-glass, and the 
specimen examined with an immersion lens. 

If the specimen is to be made permanent, fix the cover-glass at 
one corner with the thumb, and with a soft rag carefully wipe off 
the cedar oil ; then float off the cover-glass by running in distilled 
water at its margin, and having made a little ledge with a strip of 
filter-paper, place the cover-glass up against it upon one of its 
edges and leave it to dry. When perfectly dry mount in Canada 
balsam, or put it away in a cover-glass box provided with a label of 
contents. 

In many cases it is necessary or preferable to apply the stain 
for a much longer period. This may best be effected by pouring 
some of the staining solution into a watch-glass, and allowing the 
cover-glasses to swim on the surface, with their prepared side, of 
course, downwards. Throughout all these manipulations it is 
necessary to bear in mind which is the prepared surface of the 
cover-glass. 

Instead of using the watery solutions of the aniline dyes the 
author prefers in many cases to use stronger solutions, and to reduce 
the staining by a momentary immersion in alcohol. Very beautiful 
preparations of streptococci, sarcinse and other bactei'ia can be 
obtained by this method, which is as follows : Cover-glass prepara- 
tions are stained with carbolised fuchsine (Neelsen's solution) for 
about two minutes, rinsed in alcohol for a few seconds, quickly 
washed in water, and either examined in water or dried and 
mounted in the usual way. The extent of decolorisation is a 
m.atter of practice : a momentary immersion in alcohol is sometimes 
sufficient ; too long immersion will remove too much of the colour ; 
too short immersion will leave the dehcate outlines indistinct. This 
method is especially valuable for sarcinse and streptococci, the 
divisions between the elements being sharply defijied, and as any 
albuminous particles or debris in the preparation are decolorised, 
much cleaner and sharper preparations are obtained than with the 
watery solutions. Lbffler's and other concentrated solutions may 
also be used, but Neelsen's solution may be regarded as the standard 
one for this method. 



88 BACTERIOLOGY. 

Aniline oil, carbolic acid, and some other chemicals, when added 
to the aniline dyes, have the property of acting in the manner 
of mordants, in some way fixing the colour in the bacteria, so that 
they are not so readily acted upon by decolorising agents. 

Loffler's Solution.— Potash intensifies the staining power, and 
Koch and Lbffler have both used it with methylene blue. Loffler's 
solution consists of 30 grammes of methylene blue in 100 grammes 
of 1 in 10,000 solution of potash. It may be used with advantage 
for almost all kinds of bacteria. 

Gram's Method. — With a solution of gentian -violet the whole 
film on the cover-glass is at first stained violet. By immersing the 
cover-glass in a solution of iodine in iodide of potassium the stain 
is fixed in the bacilli, but not in any debris, pus colls, or tissue 
elements present in the film. Consequently by transferring the 
cover-glass to alcohol the bacilli alone remain stained, the violet 
colour being merely changed to blue. By employing a contrast 
colour, such as eosin, a double staining is obtained. In some 
bacteria the sheath is by this method differentiated from the 
protoplasmic contents. 

The stock solution of gentian-violet is prepared by shaking up 
1 cc. of pure aniline with twenty parts of distilled water, and 
filtering the emulsion.. Half a gramme of the best finely powdered 
gentian- violet is dissolved in the clear filtrate, and the solution filtered 
before use. 

The details of the method will now be described. In the first 
place, it is much better to employ the aniline-gentian- violet solution 
quite freshly prepared, and the following useful method is invariably 
used by the author : Place four or five drops of pure aniline in 
a test-tube, fill it three-quarters full with distilled water, close the 
mouth of the tube with the thumb, and shake it up thoroughly. 
Filter the emulsion twice, and pour the filtrate into a watch-glass 
or glass capsule. To the perfectly clear aniline water thus obtained 
add drop by drop a concentrated alcoholic solution of gentian-violet 
till precipitation commences. Cover-glasses must be left in this 
solution about ten minutes, transferred to iodine-potassic-iodide 
solution until in itwo or three minutes the film becomes uniformly 
brown, and then rinsed in alcohol. The process of decolorisation may 
be hastened by dipping the cover-glass in clove-oil and returning it 
again to alcohol. The cover-glass is once more immersed in clove-oil, 
then dried by gently pressing between two layers of filter-paper, 
and finally mounted in Canada balsam. 



microscopical examination of bacteria. 89 

Double Staining op Cover-glass Preparations. 

To double stain cover-glass preparations they can be treated by 
Ehrlich's method for staining tubercular sputum, or by ISTeelsen's 
modification, or by staining mth eosin after treatment by the 
method of Gram. 

Ehrlich's method is as follows : Five parts of aniline oil are 
shaken up with one hundred parts of distilled water, and the 
emulsion filtered through moistened filter-paper. A saturated 
alcoholic solution of fuchsine, methyl-violet, or gentian-violet, is 
added to the filtrate in a watch-glass, drop by drop, until precipitation 
commences. Weigert recommended that exactly eleven parts of the 
dye should be used to one hundred parts of the aniline solution. 

Cover-glass preparations are floated in this mixture for fifteen 
minutes to half an hour, then washed for a few seconds in dilute 
nitric acid (one part nitric acid to two of water), and then rinsed 
in distilled water. The stain is removed from everything except 
the bacilli; but the ground substance can be after-stained brown 
if the bacilli are violet, or blue if they have been stained red. 

yeelsen's Solution and Methylene Blue. — Ziehl suggested the use of 
carbolic acid as a substitute for aniline oil, and IsTeelsen recommended 
a solution composed of 100 cc. of a 5 per cent, watery solution 
of carbolic acid, 10 cc. of absolute alcohol, and 1 gramme of 
fuchsine. This stain is commonly known as the Neelsen or Ziehl- 
Neelsen solution. Cover-glass preparations are floated on the hot 
dye for two minutes, they are then rinsed in dilute sulphuric acid 
25 per cent., washed in water, immersed in watery solution of 
methylene blue for three minutes, again washed in water, dried, 
and mounted in balsam. 

Ora/m's Solution and Eosin. — Double staining of cover-glasses can 
be obtained by combining Gram's method with eosin. The method 
is very useful for differentiating the sheath of Streptococcu.s 
pyogenes and Bacillus anthracis, from the protoplasmic contents, 
and for staining preparations of pneumonic sputum, or of micrococci 
and other micro-organisms in pus. After decolorising the prepara- 
tion in alcohol, the cover-glass is transferred to a weak solution 
of eosin for two or three minutes, then washed again in alcohol, 
immersed in clove-oil, dried between fllter-paper, and mounted in 
balsam. 

Staining of Spores. 

A shght modification of the ordinary process employed in making 
cover-glass preparations has to be adopted to stain the spores of 



90 BACTERIOLOGY. 

bacilli. Under ordinary circumstances the stain will not penetrate 
the sheath, but if it can be made to penetrate, it is not readily 
removed. The cover-glass preparation must be heated to a tem- 
perature of 210° 0., for half an hour, or passed as many as twelve 
times through the flame of a Bunsen burner, or exposed to the 
action of strong sulphuric acid for several seconds, and then a few 
drops of a watery solution, of an aniline dye may be applied in the 
usual way. 

To double stain spore-bearing bacilli the cover-glass preparations 
may be floated, for from twenty minutes to an hour, on Ehrlich's 
fuchsine-aniline-water, or on the Ziehl-JSTeelsen solution. The stain 
must be heated — by preference in a capsule placed in a sand-bath — 
until steam rises. The fuchsine is removed from the bacilli by 
rinsing in water and washing in weak hydrochloric acid, and then 
the preparations are washed again in water, and floated for a few 
minutes on a watery solution of methylene blue. They are again 
rinsed in water, dried, and mounted. Neisser's decolorising solution 
consists of 25 parts of hydrochloric acid to 75 parts of alcohol. 

Staining of Flagella. 

Koch first stained flagella by floating the cover-glasses on a 
watery solution of h8ematox}din. From this they were transferred 
to a 5 per cent, solution of chromic acid, or to Muller's fluid, by 
which the flagella obtain a brownish-black coloration. The author 
succeeded in demonstrating and photographing flagella in prepara- 
tions stained with a saturated solution of gentian violet in absolute 
alcohol ; but these methods are now superseded owing to the much 
more satisfactory method introduced by Lbffler. 

Lbffler's method depends upon the employment of a mordant. 
Lbfiler tried tannate of iron, and after a number of experiments 
the following method was introduced. An aqueous solution of 
ferrous sulphate is added to an aqueous solution of tannin (20 per 
cent.), until the mixture turns a violet-black colour, then 3 or 4 cc. 
of a 1 in 8 aqueous solution of logwood are added. This constitutes 
the mordant, and a few drops of carbolic acid may be added, and the 
solution kept in well-stoppered bottles. The dye consists of 1 cc. of 
a 1 per cent, solution of caustic-soda, added to 100 cc. of aniline 
water, in which 4 or 5 grammes of either methyl violet, methylene blue, 
or fu.chsine, are dissolved. A cover-glass preparation is made in 
the ordinary way, the bacteria being diffused in water, and then 
spread out in a very thin film. After drying and very carefully 
fixing, the film is covered with the mordant, and the cover-glass 



MICROSCOPICAL EXAMINATION OF BACTERIA. 91 

held over the flame until steam rises. The mordant is then washed 
off with distilled water, and all traces removed from the edge of 
the cover-glass with alcohol. The stain is filtered, and a few drops 
allowed to fall on the film, and after a few minutes the cover-glass 
is again very carefully warmed until steam rises. The stain is then 
washed off with distilled water, and is ready to be examined and 
subsequently mounted. For some bacteria it is necessary to modify 
the solutions, either by the addition of acetic or sulphuric acid, or 
by varying the quantity of soda solution. 

Trenkmann introduced a modification of Lbffler's system. Cover- 
glasses are floated for from two to twelve hours on a solution 
consisting of 1 per cent, tannin and g per cent, hydrochloric acid. 
After washing in water the preparation is stained with a saturated 
alcoholic solution of any of the aniline dyes diluted in the propor- 
tion of- 2 drops of the dye to 20 of water. The cover-glasses 
which remain in the solution for from two to four hours are then 
washed in water, and examined. The best results are obtained with 
carbolised fuchsine, diluted in the proportion of 2 drops to 20 drops 
of 1 per cent, carbolic. Trenkmann also recommended the use of 
catechu and logwood as mordants, with the addition of very dilute 
acid, and subsequent staining with fuchsine. 

Lutesch suggested the use of ferric acetate. To avoid any 
deposit on the surface of the preparation, freshly prepared saturated 
ferric acetate is used, and 5 to 10 drops of acetic acid are added to 
16 cc. of the mordant. After warming the solution the preparation 
is washed in water, followed by 20 per cent, acetic acid, again 
thoroughly washed, and then stained with hot sokition of fuchsine or 
gentian-violet in aniline water. 

Van Ermengem. used a mordant composed of 1 part of 
2 per cent, solution of osmic acid, 2 parts of 10 to 25 per cent, 
solution of tannin, with to every 100 cc. of this mixture 4 or 5 drops 
of acetic acid. A black ink is thus formed, and the solution is 
applied for from five to thirty minutes. After washing in water and 
alcohol the cover-glasses are placed in a sokttion of nitrate of silver 
and transferred to another solution composed of 5 grammes of gallic 
acid, 3 grammes of tannin, 10 grammes of acetate of soda, and 330 
grammes of distilled water. In a few moments they are again placed 
in nitrate of silver, and then washed and mounted in balsam. 

Sclavo's method answers well for certain micro-organisms. The 
preparations are left for one minute in, solution of tannin, washed in 
distilled water, transferred for a minute to 50 per cent, phospho- 
molybdic acid, again washed and stained from three to five minutes 



92 BACTERIOLOGY. 

in hot saturated solution of f uchsine in aniline water, washed in water, 
dried on filter paper, and mounted in balsam. The tannin solution 
consists of 1 part of tannin to 100 cc. of 50 per cent, alcohol. 

Mcolle and Morax also, have modified Lofiler's method. Per- 
fectly clean cover-glasses are used, and the film is dried without 
fixing in the flame. Cover-glasses are covered with the mordant, 
and heated for abont ten seconds, and when steam rises the mordant 
is shaken off and the film rinsed with water. The same process is 
repeated three or four times, and finally the cover-glass is stained 
with Neelsen's solution, holding it over the flame once or twice 
for a quarter of a minvite ; it is then washed and examined. 

Bunge prefers as a mordant a mixture of aqueous solution of 
tannin with 1 in 20 aqueous solution of sesquichloride of iron in the 
proportion of 3 parts of the tannin solution, 1 part of the iron 
solution, with the addition of 1 cc. of a saturated watery solution of 
f uchsine added to 10 cc. of the mixture. The mordant is kept before 
use, and ajiplied for five minutes. The preparation is then washed and 
stained with Neelsen's solution. In another plan the cover-glasses 
are immersed for one half to one minute in 5 per cent, solution of 
acetic acid, washed and dried. The mordant is then applied three or 
four times, and the cover-glasses washed, dried, and then stained with 
gentian-violet, dipped in 1 per cent, acetic acid, washed, dried, and 
mounted. Peroxide of hydrogen may he added to the mordant, 
drop by drop; it becomes reddish-brown in colour, and must be 
shaken up and filtered before use. Cover-glasses are exposed to its 
action for about a minute, and Neelsen's solution is used foi- 
staining. 

Hessert dispenses with the mordant. The film is fixed by 
treating cover-glasses with a saturated alcoholic solution of corrosive 
.sublimate. After washing, the cover-glass is stained for thirty to 
forty minutes in a hot dye, by preference a 10 per cent, watery 
solution of saturated alcoholic solution of fuchsine. 

Cover-glass Impeessions. 

One of the most instructive methods for examining micro- 
organisms is to make an ivipression-prejMt-ation. This enables us, 
in many cases, to study the relative position of individual micro- 
organisms one to another in their growth on solid cultivating media, 
and in some cases produces the most exquisite preparations for the 
microscope. A perfectly clean, usually small-sized, cover-glass is 
carefully deposited on a plate-cultivation, and gently and evenly 
pressed down. One edge is then carefully levered up, with a needle, 



MICROSCOPICAL i:XAMINATION OF BACTERIA. 93 

and the cover-glass lifted off by means of forceps. It is then 
allowed to dry, passed through the flame three times, and stained 
as already described. In some oases of plate-cultures, especially 
where no liquefaction has taken place, the growth is bodily trans- 
ferred to the cover-glass, and a vacant area left on the " gelatine 
or agar-agar, cori-esponding exactly with the form and size of the 
cover-glass employed. 

Presebvation of Preparations. 

After examining a cover-glass preparation with an oil immersion 
objective the cedar oil must be carefully wiped off, and the slide 
set aside for tbe Canada balsam to set. At a convenient time all 
preparations should be sealed with a ring of Hollis' glue ; the 
cedar oil used at subsequent examinations of the specimen will 
not be able to work its way under the cover-glass, and prevent 
the balsam from hardening. When it is ringed cedar oil can be 
readily wiped off, and the specimen cleaned without danger of 
moving the cover-glass and injuring the preparation. 

(b) Bacteria in Sections op Tissues. 

Methods of Hardening and Decalcifying Tissues. — To harden small 
organs, such as the viscera of a mouse, they should be placed on 
a piece of filter-paper at the bottom of a small wide-mouthed glass 
jar, and covered with about twenty times their volume of absolute 
alcohol. Larger organs, pathological growths, etc., are treated in 
the same way, but must first be cut into small pieces, or cubes, 
varying from a quarter of an inch to an inch in size. Miiller's 
fluid may also be employed, and methylated spirit may be sub- 
stituted for alcohol, from motives of economy. Tissues hardened in 
absolute alcohol are ready for cutting in two or three days, and 
those hardened in Miiller's fluid in as many weeks. 

Teeth, or osseous structures, must first be placed in a decalcifying 
solution, such as Kleinenberg's. When sufiiciently softened, they 
are allowed to soak in water, to wash out the picric acid, and then 
transferred through weak spirit to absolute alcohol. Ebner's solu- 
tion also gives excellent results, especially when the structures to 
be decalcified are placed in fresh solution from time to time. 

Methods of Embedding, Fixing, and Cutting. — The author 
finds that freezing with ether combined with the method of em- 
bedding in celloidin gives excellent results. The pieces of tissue 
to be embedded are placed, after the process of hardening is com- 



94 



BACTERIOLOGY. 



pleted, in a mixture of ether and alcohol for an hour or more. 
They are then transferred to a solution of celloidin in equal parts 
of ether and alcohol, and left there, usually for several hours. 

The piece of tissue is then placed in a glass capsule, and some 
of the celloidin solution poured over it. The capsule can be placed 



1 




Fig. 27.— Swift's Freezing Microtome. 



bodily in 60 to 80 per cent, alcohol, and left until the following' 
morning. The celloidin will then be of the consistency of wax. 
The piece of tissue is next cut out, and after trimming off superfluous 
celloidin is put in water until it sinks. It is then transferred to 
gum, and frozen and out with a freezing microtome. 

For cutting with Jung's mici'otome, the tissues are embedded 



MICROSCOPICAL EXAMINATION OF BACTERIA. 



95 



in paraffine or celloidin, and mounted on cork ; or, if firm enough, 
they may be fixed upon cork without any embedding material at 
all. Paraffine, dissolved in chloroform, will be found vei'y .service- 
able as an embedding material. 

Corks ready cut for the clamp of the microtome are smeai'ed 
over with the solution of celloidin. This can be applied with a 
glass rod to the surface which is to receive the piece of tissue- 
The corks are then set aside for the film of celloidin to harden. 
In the case of lung, or degenerated broken-down tis.sue, the 
specimen should be left for a much longer time than is found to 
be sufficient for firmer structures. When ready, it is removed 
from the celloidin solution with forceps and placed upon the pre- 




FiG. 28. — JL'^"G's Microtome. 



pared cork. Enough of the solution, which is of syrupy consistence, 
is allowed to fall on the piece of tissue to cover it completely, and 
the mounted specimen is placed in the alcohol to harden. The 
specimen will be ready for cutting next day. 

The specimen may be more neatly embedded by fixing it with 
a pin in a small paper tray, pouring the celloidin solution over it, 
and then placing the tray in alcohol to harden the celloidin. The 
embedded specimen is then fixed on a cork, which has been cut for 
the clamp of the microtome. The celloidin in the section disappears 
in the pi'ooess of clearing with clove-oil. 

In the case of specimens embedded in celloidin, or mounted 
directl)' on a cork, the tissue, as well as the blade of the knife, should 
be kept constantly bathed with alcohol, and the sections transferred 
from the blade with a camel's-hair brush, and floated in alcohol. 



96 BACTERIOLOGY. 

For fixing directly on cork, small organs and pieces of firm tissue 
such as the kidneys of a mouse, or liver, we may employ gelatine or 
glycerine gelatine, liquefied over a Bunsen burner in a porcelain 
capsule. Glycerine gelatine may be used with advantage for fixing 
irregular pieces of tissue, as it does not become of a consistency 
that would injure the edge of the knife. The cork, with specimen 
affixed, is placed in alcohol, and is ready for cutting sections next 
day. 

Material infiltrated with paraffine must be cut perfectly dry, 
■ and the sections prevented from rolling up by gentle manipulation 
with a camel's-hair brush. They must then be picked off the blade 
of the knife with a clean needle, and dropped into a watch-glass 
containing xylol. This dissolves out the paraffine. The sections are 
then transferred to alcohol to get rid of the xylol, and then to the 
staining solution. 

Staining Bacteria in 'Tissue Sections. — Sections of fresh tissues 
made with the. freezing microtome are to be floated in '8 per cent, 
salt solution, and then carefully transferred, well spread out on a 
platinum lifter, to a watch-glass containing absolute alcohol. Simi- 
larly, sections selected from those cut with Jung's microtome may 
be transferred from the spirit to absolute alcohol. The sections 
may be then stained by any of the methods to be described. 

It Ls often advisable to employ some method which will enable 
one to study the structure of the tissue itself ; and sections, however 
stained, should always be first examined with low powers, to enable 
one to recognise the tissue under examination, and to examine in 
many cases the topographical distribution of masses of bacteria. 
With a power of about 250 diams. (one-sixth), very many bacteria 
can be distinguished ; and with the oil immersion lenses the minutest 
bacilli and micrococci can be recognised, and the exact form of 
individual bacteria accurately determined. As most good modern 
instruments are provided with a triple nose-piece, there is no loss 
of time in examining a preparation successively, with these different 
powers. 

Weigert's Method.— A very useful method for staining both 
the tissue and the bacteria is as follows: Place the sections for 
from six to eighteen hours in a 1 per cent, watery solution of any of 
the basic aniline dyes (methyl violet, gentian violet, fuchsine, Bis- 
marck brown). To hasten the process, place the capsule containing 
the solution in the incubator, or heat it to 45° C. A stronger 
solution may also be employed, in which case the sections are far 
more rapidly stained, and are easily over-stained. In the latter case 



MICROSCOPICAL EXAMINATION OF BACTERIA. 97 

they must be treated with a half-saturated solution of carbonate of 
potash. In either case the sections are next washed with distilled 
water, and passed through 60 per cent, alcohol into absolute alcohol. 
When almost decolorised, spread out the section carefully on a 
platinum lifter and transfer it to clove-oil, or stain with picro-carmine 
solution (Weigert's) for half an hour, wash in water, alcohol, and 
then treat with clove-oil. After the final treatment with clove-oil, 
transfer with the platinum lifter to a clean glass slide. Dry the 
preparation by pressure with a piece of filter-paper folded several 
times, and preserve in Canada balsam, dissolved in xylol. 

Gram's Method. — In the method of Gram sections are stained 
for ten minutes in a capsule containing aniline-gentian- violet solution. 
Great care must be taken not to injure the sections. If there is 
any difficulty in finding them, it is best to carefully pour off the 
stain and fill up the capsule with water. The sections are then readily 
visible, and can be taken up on the end of a glass rod and placed 
in the iodine and iodide of potassium solution, where they remain foi- 
two or three minutes, until stained uniformly brown and resembling 
in appearance a boiled tea-leaf. They are then placed in absolute 
alcohol, and washed by carefully moving the sections in the liquid 
with a glass rod. When completely decolorised they are spread out 
on a lifter, and transferred to clove-oil until completely clarified. 
Each is transferred with a hfter to a slide, and the clove-oil is 
run off and then completely removed by gently pressing two or 
three layers of filter-paper upon the section. Finally, the section 
is mounted in Canada balsam. 

The process of decolorisation may be hastened by transferring the 
section from alcohol to clove-oil, and back agairi to alcohol, repeating 
this two or three times. 

On examination the tissue appears coloui^less, or slightly tinged 
yellow from too long immersion in the iodine solution, while the 
micro-organisms are stained blue or blue-black. 

Double staining is obtained by transferring the sections after 
decolorisation to eosin, Bismarck brown, or vesuvin. They are left 
in a watery solution for two or three minutes, then again washed in 
alcohol, before clarifj'ing in clove-oil and mounting in balsam. 

Another instructive method is to place the decolorised sections 
in picro-carminate of ammonia for three or four minutes, and then 
treat with alcohol and clove-oil. 

A similar result is obtained by placing the sections in Orth's 
solution (picro-lithium carmine), transferring to acidulated alcohol, 
and then passing through clove-oil and mounting in balsam. 

7 



98 BACTERIOLOGY. 

In EhrKch's method delicate sections are lia))le to be injured by- 
immersion in the nitric acid, and therefore Watson-Oheyne suggested 
the use of formic acid. 

The Ziehl-Neelsen method, in which sulphuric acid is used instead 
of nitric acid, is much to be preferred to Ehrlich's method. 

Ziehl-Neelsen Method-. — The solution is warmed, and sections 
left in it for ten minutes. The red colour, which disappears when 
the section is placed in weak sulphuric acid (25 per cent.), may 
partly return when the section is placed in water. In this case the 
section must be again immersed in acid and passed backwards and 
forwards from acid to water until the red colour has completely, or 
almost completely, disappeared. It must be thoroughly washed in 
water to remove all traces of the acid, and then placed in a watery 
solution of methylene blue for two or three minutes, washed again 
in water, immersed in alcohol, clarified in clove-oil, and mounted in 
the usual way. Sections are brilliantly stained, and the results are 
very permanent. 

Many special methods of staining have been introduced, and will 
be given in subsequent chapters with the description of the bacteria 
to which they apply. The methods already described are those 
which are more or less in constant use in studying bacteria and in 
conducting original researches. 



CHAPTER IX. 

PREPARATION OF NUTRIENT MEDIA AND METHODS OF 
CULTIVATION. 

To cultivate micro-organisms artificially, and, in the case of tlie 
pathogenic bacteria, to fulfil the second of Koch's postulates, they 
must be supplied with nutrient material free from pre-existing 
micro-organisms. Hitherto various lands of nutrient liquids have 
been employed, and in many cases they still continue to be 
used with advantage, but for general use they have been, in a 
great measure, supplanted by the methods of cultivation on sterile 
solid media about to be described. The advantages of the latter 
methods are numerous. In the first place, in the case of liquid 
media, in spite of elaborate precautions and the expenditure of much 
labour and time, it was almost impossible or extremely difficult to 
obtain a pure culture. When a drop of liquid containing several kinds 
of bacteria is introduced into a liquid medium, we have a mixed 
cultivation from the very first. If in the struggle for existence 
some bacteria were unable to develop in the presence of others, or 
a change of temperature and soil allowed one form to predominate 
over another, then we might be led to the conclusion that many 
bacteria were but developmental forms of one and the same micro- 
orgd,nism ; while possibly the contamination of such cultures might 
lead to the belief in the transformation of a harmless into a patho- 
genic bacterium. The secret of the success of Koch's methods greatly 
depends upon the possibility, in the case of starting with a mixture 
of micro-organisms, of being able to isolate them completely one 
from another, and to obtain an absolutely pure growth of each 
cultivable species. When sterile nutrient gelatine has been liquefied 
in a tube and inoculated with a mixture of bacteria in such a way 
that the individual micro-organisms are distributed throughout it, 
and the liquid is poured out on a plate of glass and allowed to solidify, 
the individual bacteria, instead of moving about freely as in a liquid 
medium, are fixed in one spot, where they develop individuals of 

99 



100 BACTERIOLOGY. 

their own species. In tlii.s way colonies are formed each possessing 
its own biological characteristics and morphological appearances. 
When an adventitious germ from the air falls npon the culture, it also 
gi'ows exactly upon the spot upon which it fell, and can be easily 
reco<^nised as a stranger. To maintain the individuals isolated from 
one another during their growth, and free from contamination, it is 
only necessary to thin out the cultivation, and to protect the plates 
from the air. The slower growth of the micro-organisms in solid 
media, affording much greater facility for examining them at various 
intervals and stages of development, is an additional point in favour 
of these methods ; and the characteristic macrosoopical appearances 
so frequently assumed are, more especially in the case of morpho- 
logical resemblance or identity, of the greatest importance. The 
colonies on nutrient gelatine (examined with a low power) of micro- 
organisms such as Bacillus anthracis and Proteus mirabilis, the 
naked eye appearances in test-tubes of the growth of the bacilli 
of anthrax and tubercle, and the brilliant growth of Micrococcus 
prodigiosus, may be quoted as examples in which the appearances are 
often very striking and sometimes quite characteristic. 

Solid Media. 

(a) Preparatiox^of Nutrient Gelatine and Nutrient 
Agar- A GAR. 

Nutrient Gelatine is prepared as follows : Take half a kilo- 
gramme of beef (one pound), as free as possible from fat. Chop it 
up finely, ti'ansfer it to a flask or cylindrical 
vessel, and shake it up well with a litre of 
distilled water. Place the vessel in an ice- 
pail, ice-cupboard, or in winter in a cold 
cellar, and leave for the night. Next morn- 
ing commenc3 with the pi'eparation of all 
requisite apparatus. Thoroughly wash and 
linse with alcohol about 100 test-tubes, and 
allow them to dry. Plug the mouths of the 
'"^'^^ ' " test-tubes witli cotton-wool, taking care that 

Fig. 2a-WiKE Cage ^.j^^ , ^^ g^.^^^j ,^^^^ ^^^^ ^^^ tightly. 

FOR Test-tubes. . . . . . 

Place them in their wire cages in the hot-air 

sterilisei-, to Ije heated for an houi' at a tempei-iture of 150° C. In 
the same manner cleanse and sterilise several flasks and a small 
glass funnel. In the meantime the meat infusion must be again 
well shaken, and the liquid iiortion separated b}- filtering and 



DESCRIPTION OF PLATE II. 
Pure-cultivations of Bacteria. 

Fig. 1. — In the depth of Nutrient Gelatine. A pure-cultivation of Kochs 
comma-bacillus (Spirillum cholerse AsiaticEe) showing in the track of 
the needle a funnel-shaped area of liquefaction enclosing an air-bubble, 
and a white thread. Similar appearances are produced in cultivations of 
the comma-bacillus of MetchnikofE. 

Fig. 2. — On the surface of Nutrient Gelatine. A pure-cultivation of Bacillus 
typhosus on the surface of obliquely solidified nutrient gelatine. 

Fig. 3. — On the surface of Nutrient Agar-agar. Pure-cultivation of Bacillus 
indicus on the surface of obliquely solidified nutrient agar-agar. The 
growth has the colour of red sealing-wax, and a peculiar crinkled 
appearance. After some days it loses its bright colour and becomes 
purplish, like an old cultivation of Micrococcus prodigiosus. 

Fig. 4. — On the surface of Nutrient Agar-agar. A pure-cultivation obtained 
from an abscess (Staphylococcus pyogenes aureus). 

Fig. 5. — On the surface of Nutrient Agar-agar. A pure-cultivation obtained 
from green pus (Bacillus pyocyaneus). Thfe growth forms a whitish, 
transparent layer, conlposed of slender bacilli, and the green pigment 
is diffused throughout the nutrient jelly. ' The growth appears green by 
transmitted light, owing to the colour of the jelly behind it. 

KiG. H.— On tJie surface of Potato. A pure-cultivation of the bacillus of 
glanders on the surface of sterilised potato. 



Plate IL 




pin 








en 




z 







-? 


1 — I 


.■op 


H 


Pn 


< 




C 




I-' 




K-H 




H 




^ 




Id 


o 


o 


■o^ 


1 


E 


w 




tf 




t3 




dl 






^ 

f^ 



f^MWnf ^-V^JcAiitry J^ •^O'l.iit'l - 



NUTRIENT MEDIA AND METHODS OF CULTIVATION. 



101 



squeezing through a linen cloth or a meat press. The red juice thus 
ohtained nmst be brought up to a litre by transferring it to a large 
measuring glass and adding distilled water. It is then poured 
into a sutfieiently large and strong beaker, and set aside after the 
addition of 10 grammes of peptone, 5 grammes of common salt and 
11)0 grammes of best gelatine. 

In about half an hour the gelatine is sufficiently softened, and 
subsequent heating iu a water-bath cairses it to ba completely 




Fig. 30. — Hot Am Stekiliser. 



dissoh'ed. The danger of breaking the beaker may be avoided by 
placing a cloth, several times folded, at the bottom of the water-bath. 

The next process requires the greatest care and attention. Some 
micro-organisms grow best in a sUghtly acid, others in a neutral 
or slightly alkaline, medium. For example, foi- the growth and 
characteristic appearances of the comma bacillus of Asiatic cholera 
a faintly alkaline soil is absolutely essential. This slightly alkaline 
medium will be found to answer best for most micro-organisms, and 
may be obtained as follows : — 

With a clean glass rod dipped in the mixture, the reaction 
upon litmus-paper may be ascertained, and a concentrated solution 
of carbonate of soda must be added drop by drop, until red litmus- 



102 



BACTERIOLOGY. 



paper becomes faiutly blue. If it has been made too alkaline, it can 
be neutralised by the addition of lactic acid. 

Finally, the mixture is heated for an hour in the water-bath. 
Ten niinvites before the boiling is completed, the white of an egg 
beaten up with the shell is added, and the liquid is then filtered 
while hot. For the filtration, the hot-water apparatus (Fig. 31) 
can be used with advantage, furnished with a filter of Swedish 

paper, which may be conveniently 
made in the follov/ing way : — 

About eighteen inches square 
of the best and stoutest filter paper 
is fii'st folded in the middle, and 
then creased into sixteen folds. The 
filter is made to fit the glass funnel 
by gathei'ing up the folds like a fan, 
and cutting off the superfluous part. 
The creasing of each fold should be 
made fii-mly to within half an inch 
of the apex of the filter, which part 
is to be gently inserted into the 
tube of the funnel. To avoid 
bursting the filter iit the point, the 
Ijrotli, when poured out from the 
flask, should be diieoted against 
the side of the filter with a glass 
rod. During filtivition the funnel 
should be covered over watli a 
circular plate of glass, and the pro- 
ce.ss of filtration must be repeated, 
if necessar}', until a pale, straw- 
coloured, pei'fectly transparent 
filtrate results. 
The sterilised test-tubes are filled to about a thu-d of then- depth 
by pouring in the gelatine carefully and steadily, oi' by employing a 
small sterilised glass funnel. The object of this care is to prevent 
the mixture touching the part of the tube with which the plug 
comes into contact ; otherwise, when the gelatine sets, the cotton- 
wool adheres to the tube and becomes a source of embarrassment in 
subsequent procedures. As the tubes are filled they are placed in 
the test-tube basket, and must then be sterili.sed. They are either 
lowered into the steam steriliser, when the thermometer inchcates 
100° C:, for twelve minuter for four or five successive days, or they 




Fig. 31.- 



-HOT-W.iTEE FiLTEKIXG 
App.iR.iTUS. 



NUTRIENT MEDIA AND METHODS OF CL'LTIVATION. 



103 



may be transferred to the test-tube water-bath, and heated for an 
hour a day for tliree successive days. 

If the gelatine shows any turbidity after these processes it must 




Fio. 32. — Metholi of .m.\king a Folded Filteu. 



be poui-ed back from the test-tubes into a flask, bailed up for ten 
minutes, and iiltered once more, and the processes of sterilisation just 
described must be repeated. 




Fig. 33.— ISteam Stehilisei;. 



Nutrient Agar-agar. — Agar-agar is a substance prepared 
from seaweed which grows on the coasts of Japan and India, 
and is supplied in long crinkled strips. It VjoiLs at 90° C, and 



104 



BACTERIOLOGY. 



i-emains solid up to a temperature of about 45° C. It is there- 
fore substituted for gelatine in the pi'ejjaration of a jell}- for the 
cultivation of those bacteria which will only grow, or grow best, in 
the inculiatoi' at the temperatui'e of the blood. It may also be 
employed at ordinary temperatures for bacteria which liquefy 
gelatine. The preparation is conducted on much the same principles 
as those already described. Instead, however, of 100 grammes of 
gelatine, only about 20 grammes of agai--agar are employed (1-.5 to 
2 per cent.), and to facilitate its solution it must be allowed to soak 
in salt water overnight. For the filtration, flannel is substituted 

for filter- paper, or may 
be used in combination 
■with the latter. The 
hot-water apparatus 
is invariably emploj-ed, 
unless, to accelerate 
the process, the glass 
funnel and receiver are 
bodily transferred to 
the steam sterihser. If 
the conical cap cannot 
be replaced, cloths laid 
over the mouth of the 
steriliser must be em- 
ployed instead. It may 
be necessary to repeat 
the process of filtra- 
tion, but it must not be 
expected that such a 
brilliant transparency 
can be obtained as with 
gelatine. The final 
hould be colourless and clear ; but if slightly 




Flu. 34. — Incubator 



result, when solid 

milky, it may still be employed. 

A little liquid gradually collects in the tubes, Ijeing expiressed by 
the contraction of the agar-agar. 

Wort-gelatine is used in studying the bacteria of fermentation. 
It is made by adding from 5 to 10 per cent, of gelatine to beer-wort. 

Glycerine Agar-agar, — This is prepared by adding 5 per 
cent, of glycerine to nutrient agar-agar, after the boiling and before 
the filtration, and other modifications can be made for special 
purposes by the addition of grape-sugar or of gelatine. 



I 



NrXRIENT MEDIA AND METHODS OF CULTIVATION. 



105 



After the final treatment in the steam steriKser some of the 
tubes of gelatine and agar-agar are placed upright and allowed to 
set, and others are placed on an inclined plane or in the blood-serum 
inspissator, and left to gelatinise with an oblique surface. 



(b) Methods of employing Nutrient Jelly in Test-tubes 
AND ON Glass Plates. 

Test-tube-cultivations. — To inoculate test-tubes containing 
nutiient jelly, the cotton-wool plug is first twisted round in case 
there are an}' adhesions between the plug and the test-tube. It is 
then removed with the thumb and 
index finger of the right hand, and 
placed between the fourth and fifth 
fingers of the left hand, instead of 
being put down on the laboratory table 
and thereby probably contaminated 
with bacteria oi' the spores of mould 
fungi. A steriHsed needle charged, 
for example, with blood or pus con- 
taining bacteria, oi' with a colony from 
a. plate-culture, is thrust once in the 
middle line into the nutrient jelly, 
and steadily withdrawn. The tube 
should be held horizontally or witli 
its mouth downward, to avoid, as far 
as possible, accidental contamination 
from the gravitation of gei'ms in the 
air; and the plug replaced as quickly 
as possible. The cotton-wool project- 
ing bej'ond the mouth of the tube is 

then thoroughly bui'nt in the flame of a Bunsen burner oi' blow- 
pipe, and an india-rubber cap fitted over the mouth of the tube 

The chances of error arising from contamination of the culti- 
vations are reduced by avoiding di'aughts at the time of inoculation, 
and it is best that these manipulations should be carried on in a 
quiet room in which the tables and floor are wiped with damp cloths, 
rather than in a laboratory in which the air becomes charged with 
germs through constant sweeping and dusting, and the entrance 
and exit of classes of students. In conducting any investigation 
a dozen or more tubes should be inoculated, and if by chance an 
adventitious germ, in spite of all pi-ecautions, gains an entrance. 




Fig. 35. — Method of Inoculat- 
ing A Test-tube containing 
Sterile Nutrient Jelly. 



106 BACTERIOLOGY. 

the contaminated tube can be rejected, and the experiments con- 
tinued with the remaining pure cultivations. 

When, however, one tube containing a liquid medium is in- 
oculated from another, as in the process of preparing plate-cultures, 
or when a cultiire is made from a tube in which the growth has 
liquefied the gelatine, it is obvious that the tubes cannot be inverted 
or held horizontally, and they must then be held and inoculated as 
in Fig. 38. To inoculate those tubes of nutrient media which have 
been solidified obliquely, the point of a straight sterilised needle 
charged with the material to be cultivated is traced over the surface 
of the jelly from below upwards, or the inoculated material may be 
spread out with a hooked or looped needle. 

Examination of Test-tube-cultivations. — The appearances pro- 
duced by the growths in test-tubes can be in most cases sufficiently 
examined with the naked eye. In some cases the jelly is partially 
or completely liquefied, while in others it remains solid. The 
growths may be abundant or scanty, coloured or colourless. The 
nutrient jelly may itself be tinged or stained with products resulting 
from the growth of the organisms. When liquefaction slowly takes 
place in the nesdle track, or the organism grows without producing 
this change, the appearances which result are often very delicate, 
and in some cases very characteristic. The appearance of a simple 
white thread, of a central thread with branching lateral filaments, 
of a cloudiness, or of a string of beads in the track of the needle, 
may be given as examples. 

In some cases much may be learnt by examining the growth with 
a magnifying glass. Here, however, a difficulty may be encountered, 
for the cylindrical form of the tube so distorts the appearance of its 
contents, that the examination is rendered somewhat difficult. To 
obviate this, a very simple contrivance may be employed with 
advantage. This consists of a rectangular vessel, about four inches 
in height and two inches in width, which may be easily constructed 
by cementing together two slips of glass to form the back and front, 
with three slips of stout glass with ground edges forming the sides 
and base (Cheshire). The front may be constructed of thin glass, 
and the base of the vessel made to slope so that the test-tube when 
placed in the vessel has a tendency to be near the front. The 
vessel is filled with a mixture of the same refractive index as the 
nutrient gelatine. The latter has a refractive index rather higher 
than water, which is about 1-333 ; alcohol has a refractive index of 
1-374. The vessel is filled with water, and alcohol is then added 
until the proper density is reached. The test-tube is placed in the 



NUTRIENT MEDIA AND METHODS OF CULTIVATION. 



107 



vessel, and held in position by means of a clip. Tlie vessel can be 
fixed on the inclined stage of the microscope, and the contents of the 
tube conveniently examined with low-power objectives. 

Plate-cultivations. — By this method, as already mentioned, a 
mixture of bacteria, whether in fluids, excreta, or in cultivations on 
solid media, can be so treated that the different species are isolated 
one from the other, and perfectly pure cultivations of each of the 
cultivable bacteria in the original mixture established in various 
nutrient media. We are enabled also to examine under a low 
power of the microscope the individual colonies of bacteria, and to 
distingui.sh hj their characteristic appearances, micro-organisms 
which, in their individual foi-m, closely resemble one another, or 
ai'e even identical. The same process, with slight modification, is 
also employed in the examination of air, soil, and water, which will 
be referred to later. 

The preparation 
of plate-cultivations, 
therefore, must be 
described in every de- 
tail : and to take an 
example, we will sup- 
pose that a series of 
plates is to be pre- 
pared from a te.st- 
cube - cultivation . 

Arrangement of 
Levelling Apparatus.- 




Fic. 36. — Levelling Appakatus, 



-In 



order to spread out the liquid jelly 
evenly on the surface of a glass plate, and hasten its sohdifica- 
tion, it is necessary to place the glass plate upon a level and 
cool surface. This is obtained in the following manner : Place a 
large shallow glass dish upon a tripod stand, and fill it to the brim 
with cold water ; carefully cover the dish -vvith a slab of plate-glass, 
or a pane of window-glass, and level it by placing the i^pirit-level in 
the centre and adjusting the screw.s of the tripod. Substitute for 
the spirit-level a piece of filter-paper the size of the glass plates to 
be employed, and cover it with a shallow bell-glass. 

Sterilisation of Glass Plates. — The glass plates are sterilised in 
an iron box placed in the hot-air sterilisei', at 1.50° C, from one to 
two hours. As these plates are used also for other purposes, a 
quantity ready sterilised should always be kept in the box. 

Preparation of Damp Chambers. — The damp chambers for the 
reception of the inoculated plates are prepared thus : Thoroughly 



108 



BACTERIOLOGi'. 



cleanse and wash out with Tin 20 carbolic acid a shallow glass dish 
and bell. Cut a piece of filter-paper to line the bottom of the glass 
dish, and moisten it with the same solution. 
Metliod of Inoculating the Test-tubes. — In 
a glass beaker or an ordinary glass tumbler, 
with a pad of cotton-wool at the bottom, 
place the tube containing the cultivation, 
the three tubes to be inoculated, three glass 
rods which have been sterilised by heating 
in the flame of a Bunsen burner, and a 
thermometer. Provide a strip of paper, a 
large label, a pencil, a pair of forceps, and 
inoculating needles. All is now ready at 
hand to commence the inoculation of the tubes. 
Lifpiefy the gelatine in the three tubes b}' placing them in a 
beaker containing watei' at 30° C, or by gently warming them in 
the flame of the Bunsen })urner. Keejp the tubes, both before and 
;ifter the inoculation, in the warm water, to maintain the gelatine 
in a state of liquefaction. Hold the tube containing the cultivation 




Fig. 37.— Iron Bo.\ 
Gl.'Iks Plates. 




Fig. 38.— Method of Inoculating Test-tubes in the Pkepakation 
OF Plate-cultivations. 



and a tube of the liquefied gelatine as nearly horizontal as possible 
between the thumb and index finger of the left hand. With the index 
finger and thumb of the right hand loosen the plugs of the tubes. 
Take the looped platinum needle in the right hand and hold it like 
a pen. Remove the plug from the culture-tube by using the fourth 
and fifth fingers of the right hand as forceps, and place it between 



DESCRIPTION OF PLATE III. 

Plate-cultivation. 

This represents the appearance of a plate-cultivation of the comma-bacillus 
of Cholera nostras, when it is examined over a slab of blackened plate-glass. 
The drawing was made from a typical result of thinning out the colonies by 
the process of plate-cultivation. At this stage they were completely isolated 
one from the other ; but later they became confluent, and produced complete 
liquefaction of the gelatine. 



Plate IIL 




o 

I— I 
H 

H 
O 

H 

H 
<i 

^A 



Jf. Crooltslia.jOcf'.ci 



Vi.n^£ni Brvohs.Dc^ SfSonMth. 



NUTRIENT MEDIA AND METHODS OF CULTIVATION. 109 

the fourth anci fifth fingers of the left. Remove the plug of the 
other tube in the same way, placing it between the third and fourth 
fingers of the left hand. With the needle take up a droplet of the 
cultivation and stir it round in the liquefied jelly. Replace the plugs, 
and set aside the cultivation. Hold the freshly inoculated tube be- 
tween the index finger and thumb of either hand, almost horizontally, 
then raise it to the vertical, so that the liquid gelatine gently flows 
back. By repeating this motion and rolling the tube between the 
fingers and thumbs the micro-organisms which have been introduced 
are distributed throughout the gelatine. Any violent shaking, and 
consequent formation of bubbles, must be carefully avoided. From 
the inoculated tube, in the same manner inoculate a fresh tube of 
liquefied gelatine, introducing into it three droplets with a sterilised 
needle. After tilting and rolling this tube, as in the previous case, 
the same process is repeated with a third tube, which is inoculated 
from the second tube. This last tube must be inoculated in different 
ways, according to experience, for different micro-organisms. Some- 
times a sufficient separation of the micro-organisms is attained by 
inoculating the last tube with a straight, instead of a looped, needle, 
dipping it from the one into the other from three to five times. 

The next process consists in pouring out the' gelatine on glass 
plates and allowing it to solidify. 

Preparation of the Gelatine- Plates. — The directions to be observed 
in pouring out the gelatine are as follows : — 

Place the box containing sterilised plates horizontally, and so 
that the cover projects beyond the edge of the table ; remove the 
cover, and withdraw a plate with sterilised forceps ; hold it between 
the finger and thumb by opposite margins, rapidly transfer it to 
the filter-paper under the bell-glass, and quickly replace the cover 
of the box. On removing the plug from the tube which was first 
inoculated, an assistant raises the bell-glass, and the contents of the 
tube are poured on to the plate ; with a glass rod the gelatine must 
be then i-apidly spread out in an even layer within about half an 
inch of the margin of the plate. The assistant replaces the bell- 
glass, and the gelatine is left to set. Meanwhile a glass bench or 
metallic shelf is placed in the damp chamber, ready for the reception 
of the plate-cultivation, and when the gelatine is quite solid the 
plate is quickly transferred from under the bell-glass to the damp 
chamber ; precisely the same process is repeated with the other 
tubes, and the damp chamber, labelled with the details of the 
experiment, is set aside for the colonies to develop. Not only plate- 
cultures should be carefully labelled with date and description, but 



110 BACTERIOLOGY. 

the same remark applies equally to all preparations — tube-cultures, 
potato-cultures, drop-cultures, etc. 

Corresponding with the fractional cultivation of the micro- 
organisms obtained in this manner, the colonies wUl be found to 
develop in the course of a day or two, the time varying with the 
temperature of the room. The lower plate will contain a countless 
number of colonies which, if the micro-organism liquefies gelatine, 
speedily commingle, and produce, in a very short time, a complete 
liquefaction of the whole of the gelatine. On the middle plate the 
colonies will also be very numerous, but retain their isolated position 
for a longer time ; while on the uppermost plate the colonies are 
completely isolated from one another, with an appreciable surface of 
gelatine intervening. 

Examination of Plate-cultivations: — The macroscopical appear- 
ances of the colonies are best studied by placing the plate on a 




Fig. 39. — Damp Chamber containing Plate-cultivations. 

slab of blackened glass, or on a porcelain slab if the colonies are 
coloured. 

To examine the microscopical appearances, a selected plate is 
placed upon the stage of the microscope. The smallest diaphragm 
is employed, and the appearances studied principally with a low 
power. These appearances should be carefully noted, and a 
sketch or photograph of the colony made. The morphological 
characteristics of the micro-organisms of which the colony is formed 
can be examined in the following way: A small looped or 
hooked platinum needle is held like a pen, and the hand steadied 
by resting the little finger on the stage of the microscope. The 
extremity of the needle is steadily directed to the space between 
the lens and the gelatine without touching the latter, until, on 
looking through the microscope, it can be seen in the field, above 
or by the side of the colony under examination. The needle 
is then dipped into the colony, steadily raised, and withdrawn. 
Without removing the eye from the microscope this manipulation 
can be seen to be successful by the colony being disorganised or 



NUTRIENT MEDIA AND METHODS OF CULTIVATION. 



Ill 



completely removed from the gelatine. It is, however, not ea.sy to 
be successful at first, but with practice this can Vje accomplished 
with rapidity and pi'ecision. A preparation is then made by rubbing 




Fig. 40.— Pasteur's Laege Incubatok. 

the extremity of the needle in a droplet of water on a slide, covering 
with a cover-glass, and examining in the fresh state, or by spreading 
out the droplet on a cover-glass, drying, passing three times through 
the flame, and staining with a drop of fuchsine or gentian violet. 
Inoculations should be made in test-tubes of nutrient gelatine 



112 llACTEKIOLOGY. 

and agar-agar, from the micixi-oi'gaui.sms transferred to the euver- 
u-lass before it is dried and stained, from any remnants of the colony 
which was examined, or from other colonies bearing exactly similar 
appearances. In this way pure cultivations are established, and 
the macroscopical appearances of the growth in test-tubes can be 
obtained. The plates should be replaced in the damp chamber 
as soon as possible ; drying of the gelatine, or contamination with 
micro-organisms giuvitating from the air dui'iug their exposure, 

may spoil them for subsequent examination. 
A much simpler method of plate -cultivation 

is to dispense with the levelling apparatus, and 

„ , pour the liquetied ielly into .shallow, flat dishes. 
Fig. 41.^Petri's i^ '^ ■' r ' 

j)jgjj -Lhey take up much less I'oom, and m many 

ways are more convenient (Fig. 41). 

Nutiient agar-agar can also be employed for the preparation 
of plate-cidtivatious, but it is much more difficult to obtain satis- 
factory results. The- test-tubes of nutrient agar-agar must be 
placed in a beaker with Avater and heated until the agar-agar 
is completelj- liquefied. The gas is then turned down, and the 
temperature of the water allowed to fall until the thermometer 
•stands just above 50° C. The water must be maintained at this 
tempei'rttui-e, and the test-tubes must be in tui-n rapidly inoculated 
and poured out upon the glass plates, or bettei' still, into glass 
dishes, as alieady described. 

A very much simpler plan is to liquefy the agar, pour it into 



I 





Fii;. 42.— Gl.\sm Benches and Sudes. 

a, shallow dish, and allow it to solidify. The culture material is 
thinned out in sterilised broth, and a few drops are spread out over 
the surface of the agar. The dishes are tlien i>laceil in the incubator 
at 37° 0. 

Glass plates may also l^e employed in a, much simpler way. 
The nutrient jelly is liquefied, poured out, and allowed to set. A 
needle charged with the material to be inoculated is then drawn 
in lines over the sui-face of the jelly. This method is of use for 
inoculating different organisms .side by side, and watching the effect 
of one >ipon the other, or a. micro-organism in this Avay may be 



NUTRIENT MEDIA AND METHODS OF CULTIVATION. 113 

sown upon the gelatine whicli has been already altered by the 
growth of another micro-organism ; the change produced in the 
gelatine, as in the case of the Bacillus pyocyaneus, extending far 
beyond the limits of the growth itself. 

Nutrient jelly may also be spread oiit on sterilised glass slides, 
which after inoculation are placed in dam.p chambers for the growths 
to develop. 

Esmarch's Roll-cultures Esmarch introduced a modification 

of the method of plate- cultivation which may sometimes be used with 
advantage. The ordinary test-tubes may be employed, or tubes 
considerably larger in size. 

After the liquid jelly has been inoculated in the tube, instead 
of pouring it out on to a glass plate or into a dish, the cotton-wool 
plug is replaced, and an india-rubber cap fitted over the mouth of 
the tube. 

The tube is then placed horizontally on a block of ice or 
in a vessel containing iced water. The neck of the tube is steadied 
with the left hand, and the tube turned round and round with the 
right hand. In a very short time the gelatine sets, and the tube 
is Kned inside with a thin coating. 'There is far less danger of 
contamination, and the cultures are in a much more convenient 
form when circumstances render it necessary to move them. 

(c) Prepaeation and Employment of Solidified Blood Sebum. 

Solid Blood Serum. — The tubercle-bacillus, the bacUH of 
glanders and of diphtheria, and many other micro-organisms, thrive 
well when cultivated on solid blood serum. This medium has the 
Additional advantage of remaining solid at all temperatures. The 
technique required for its preparation and sterilisation is as follows : 
.Several cyKndrical vessels, about 20 cm. high, are thoroughly washed 
with carbolic acid (1 in 20), and then with alcohol, and finally 
rinsed out with ether. The ether is allowed to evaporate, and the 
vessels are then ready for use. The skin of the animal selected — 
■calf, sheep, or horse — is washed with carbolic at the seat of operation, 
and the bleeding performed with a sterilised knife or a trocar and 
■cannula. The first jet of blood from the vein is rejected, and that 
which follows is allowed to flow into the vessels until they are almost 
full. The ground-glass stoppers, greased with vaseline, are replaced, 
and the vessels set aside in ice, as quickly as possible, for from 
twenty-four to thirty hours. By that time the separation of the 
clot is completed, and the clear serum can then be transferred to 

8 



]14 



BACTERIOLOGY. 



plugged sterilised test-tubes. These should be filled, with a sterilised 
pipette, to about one-third of their capacity. 

Formerly the tubes were sterilised by Tyndall's process of dis- 
continuous sterilisation. The tubes were placed in Koch's serum 
sterilisei-, with the temperature maintained for an hour or more at 
.56° C, and this was repeated for six successive days, the temperature 
on the last day being gradually raised to 60° C. This completed the 
stei-ilisation, and to solidify the serum the tubes were arranged in 
the inspissator at the angle required, and the temperature was kept 

between 65° 0. and 68° C. 
Directly solidification took 
place the tubes were removed. 
The new process is much 
less tedious, and consists in 
taking every possible pre- 
caution to obtain the blood 
without contiimination by bac- 
teria in the air or in the 
vessels employed. There is 
then no need to sterilise the 
serum, and it can be coagu- 
lated immediately. The tubes 
are tested by placing them 
in an incubator at .37° C. for 
a week, and if any show 
signs of contamination they 
are discarded, and the rest 
can be used or kept in stock. 
The serum should then 
pi'esent the chai-acter of being hard, solid, of a pale straw colour, 
a.ud transparent. A little liquid collects at the lowest point, and 
the serum is sometimes milky in appearance at its thickest part. 

Loffler's Blood Serum is prepared by mixing two-thirds of fresh 
serum with one-third of broth, prepared in the usual way but with 
the addition of 1 per cent, grape-sugar. The mixture is decanted 
into test-tubes, avoiding the formation of air-bubbles, and it is then 
coagulated in the usual wa^-. The sei'um may be employed not only 
in test-tubes, but also in small flasks, glass capsules, or other vessels, 
all of which must be cleansed and sterilised. 

Hydrocele fluid and other serous effusions may be prepared in 
the same manner. Gelatine may be added to the serum in the pro- 
portion of .5 per cent. 




Fig. 43. — Koch's Sebum Stekiliskh. 



'NUTRIENT MEDIA AND METHODS OF CULTIVATION. 115 

Inoculation of the, Tubes. — A small portion of a culture or of the 
material to be inoculated is taken up with a sterilised platinum 
needle, and traced over the sloping surface of the serum ; or a 
fragment of tissue, such as diphtheritic membrane or tubercle, may 
be introduced into the tube and rubbed gently over the serum so as 
not to Ijreak the sui'face. ' 




Fig. 44. — Hueppe's Sekuji Ikspissator. 



(d) PRErAKATION AND EMPLOYMENT OF STERILISED PoTATO. 

Potato-cultivations. — Sterilised jiotatoes form an excellent 
medium for the cultivation of many micro-organisms, more especially 
the chromogenic species. Potato-cultivations also give in some cases 
very characteristic appearances, which are of value in distinguishing 
bacteria which po.ssess morphological resemblances. 

Preparation of Sterilised Potatoes. — Potatoes, preferably smooth - 
skinned, which are free from " eyes " and rotten spots, should be 
selected. If they cannot be obtained without eyes and spots, these 
must be carefully picked out with the point of a knife. The potatoes 
are well scrubbed with a stiff brush, and allowed to soak in 1 in 20 
carbolic for a few minutes. The}' are then transfei'red to the j)otato- 



116 



BACTERIOLOGY. 



receiver, and steamed in the steam steriliser for twenty minutes to 
half an hour, the time varying according to the size of the potatoes. 
When cooked, the potato- receiver is withdrawn and left to cool, the 
potatoes being retained in it until required for use. 

Damp chambers are prepared ready for the potatoes, the vessels 
being cleansed and washed with carbolic as described for plate- 
cultivations. Small glass dishes of the same pattern as the large 
ones may be employed for single halves of potatoes. Potato-knives 
and scalpels, which have been sterilised in an iron box by heating 
them in the hot-air steriHser at 151° 0. for one hour, should 
be ready to hand. Knives steriHsed by heating them in the flame 
of a Bunsen burner should afterwards be placed upon a steriHsed 
glass plate and covered with a bell-glass. It miist not be forgotten. 




Fio. 45. — Box FOR Sterilising Instruments. 



however, that heating the blades in the flame destroys the temper 
of the steel, and therefore knives and other instruments should 
preferably be sterilised in the hot-air steriliser, enclosed in an iron 
box, or simply enveloped in cotton-wool. 

Inoculation of Potatoes. — The coat-sleeves should be turned back, 
and the hands, after thorough washing with good lathering soap, 
be dipped in 1 in 40 carbolic. An assistant opens the potato- 
receiver, and a potato is selected and held between the thumb and 
index linger of the left hand. With the knife held in the right 
hand, the potato is almost completely divided in the direction 
which will give the lai-gest surface. The assistant raises the cover 
of the damp chamber, and the potato is introduced, and while the 
knife is withdrawn, allowed to fall apart. The cover is quickly 
replaced, and another potato treated in the same way, is placed 
in the same damp chamber. The four halves are then quite ready 
for inoculation. As an extra precaution, the left hand is again 
dipped in carbolic, and one half of a potato is taken up between the 
tips of the thumb and index finger, care being taken to avoid 
touching the cut surface. Holding it with its cut surface vertical, 



NUTRIENT MEDIA AND METHODS OF CnLTIVATION. 



117 



a small portion of tlie substance to be inoculated is placed on the 
centre with a sterilised platinum needle. With a sterilised scalpel 
the inoculated substance is rapidly spread over the surface of the 
potato with the flat of the blade, to within a quarter of an inch 
of the margin, and the potato is then as quickly as possible replaced 
in the damp chamber. With another sterilised scalpel a small 
portion of the potato from the inoculated surface of the first half 
is in the same way sprend over the surface of the second half, thus 
thinning out the bacteria as in plate-cultivations. Exactly the 
same is repeated with a third potato, and even a fourth, so that 
a still further thinning out or fractional cultivation of the micro- 
organisms may be obtained. In some cases it is necessary to place 
the cultures in an incubator (Fig. 40) ; others grow very well at the 




Fig. 46.— Damp Chamber for Potato-cultivations. 

temperature of the room. As in plate-cultivations, the potato may 
also be inoculated by simply streaking it in lines with a needle 
charged with the material to be cultivated. 

Potato in Test-tubes. — Large surfaces of potato are employed when 
we wish to obtain cultures of micro-organisms in considerable quanti- 
ties, as in the examination of the products of chromogenic bacteria ; 
but under ordinary circumstances potato is employed in test-tubes. 
The central portions of raw potatoes are cut out in cylindrical pieces 
with a cork-borer. These are divided obliquely in their whole 
length, and each half is placed in, a test-tube. The test-tubes are 
plugged with cotton-wool, and then steam in the steam -steriliser 
for twenty minutes. The sloping surface is inoculated in the same 
way as obliquely solidified jelly, and the advantages are great. ^ The 
cultures are obtained in a more convenient form, and there is less 
danger of contamination. 

Potato- paste may be employed when it is desirable to obtain an 
extensive growth of certain bacteria. The potatoes are boiled for 
an hour, and the floury centre squeezed out of the skins. This 
is then mashed up with sufticient sterilised water to produce a thick 



118 BACTERIOLOGY. 

paste, and is heated in the steam steriliser for half an hour for three 
successive days. 

(e) Preparation and Employment of Bread-Paste, Vegetables, 
Fruit, White of Egg. 

Some micro-organisms, more especially mould fungi, grow very 
well on bread-paste. This is prepared by removing the crust from 
slices of bread and drying them in the oven. They are then 
broken up, and reduced to a fine powder with a pestle and mortar. 
Small, carefully cleansed, conical, or globe-shaped flasks are plugged 
with cotton-wool and sterilised in the oven. When cool, a small 
quantity of the powder is placed in them, and sterilised water added 
in the proportion of one part to every four of the powder. The 
paste is sterilised by steaming in the steriliser at 100° C. for half an 
hour for three successive days. The flasks can be reversed, and may 
be inoculated with a platinum needle. 

Boiled carrots and other vegetables, and various kinds of stewed 
fruit, are also occasionally employed for the cultivation of bacteria. 
The sterilisation of these media must be carried out on the principles 
already explained. 

White of egg may be solidified in shallow glass dishes, in the 
steam steriliser. After inoculation the dishes should be placed 
in a damp chamber. 

Liquid Media. 

(f) Preparation of Sterilised Broth, Liquid Blood Serum, 
Urine, Milk, Vegetable Infusions, and Artificial Nourish- 
ing Liquids. 

Nutrient liquids are still largely employed. Eor inoculation ex- 
periments when the presence of gelatine is undesirable, for studying 
the physiology and chemistry of bacteria, and when for any object 
a rapid growth of micro-organisms is necessary, the employment 
of liquid media is not only advisable, but absolutely necessary. 
Liquid media comprise two distinct groups — natural and artificial. 
Natural media include meat broth, blood, urine, milk, and vegetable 
infusions ; artificial media are solutions composed from a chemical 
formula representing essential food constituents. 

Broth may be made from beef, pork, chicken, or fish in the 
manner which has been described for the preparation of nutrient 
gelatine, simply with omission of the gelatine. After the process 



NUTRIENT MEDIA AND METHODS OF CULTIVATION. 



119 



of neutralisation with carbonate of soda solution, the flask of broth 
is placed in the steam steriliser for half an houi- at 100° C A 
clear liquid results on iiltration which is transferred to plugged 
sterihsed flasks or test-tubes, and sterilisation effected by exposing 
them in the steam steriUser for half an hour at 100° 0. for two 




Fig. 47.~Appaeatus foe Steeilisatiox by Steam unuer pkessuhe. 



or three successive days, or by I using the apparatus for sterilising 
b\' steam under pressure. For some bacteria a more suitable 
cultivating medium is obtained by the adchtion of glycerine or 
grape-sugar. 

Liquid Blood Serum. — The preparation of blood serum has 
already been described. It may be required for cultivations before 
the final treatmeiit -by;, which it is solidified, for example, in the 
method of drop-cultivation, and may be used -ivith the addition of 
glycerine or grape-sugar. Hydrocele fluid, peritoidtic and pleuritic 



120 BACTERIOLOGY. 

effusions, can also be employed after sterilisation in the steam 
steriliserj The fluid should be withdrawn with a sterilised trocar 
and cannula, and received into plugged sterilised flasks. 

Urine. — In order to obtain urine free from micro-organisms the 
following precautions must be observed : The orifice of the urethra 
must be thoroughly cleansed with weak carbolic. The first jet of 
urine should be rejected, and the rest received into sterilised vessels, 
which must be quickly closed with sterilised plugs. If these pre- 
cautions be not attended to, the urine must be rendered sterile by 
the means described for the sterilisation of broth. 

Milk. — If milk has been drawn into sterile flasks, after thoroughly 
cleansing and disinfecting the teats and hands, it may be kept 
without change. If procured without these precautions, it must 
be steamed in the steriliser for half an hour for five successive 
days. 

Vegetable and other Infusions Infusions of hay, cucumber, 

and turnip are used for special purposes, and more rarely decoctions 
of plums, raisins, malt, and horse-dung. They are mostly prepared 
by boiling with distilled water, after maceration for several hours. 
The filtrate is received into sterile flasks and sterilised in the usual 
way in the steam sterihser. 

Artificial Fluids. — Pasteur's solution is prepared by mixing the 
ingredients in the following proportions : — 

Distilled water . 100 

Pure cane-sugar 10 

Ammonium tartrate . 1 

Ash of yeast . . -075 

Mayer's modification of the nourishing fluid employed by Cohn is 
as follows : — 

Distilled water . 20 

Ammonium tartrate . -2 

Phosphate of potassium . . -1 

Sulphate of magnesium -1 

Tribasic calcium phosphate . -01 

Drop-cultures — This method of cultivation is a particularly 
instructive one. It enables us to study many of the changes which 
take place during the life history of micro-organisms. This is 
illustrated, for example, in a drop-culture of the anthrax bacillus, 
in which we can watch the gradual growth of a single bacillus into 



NUTRIENT MEDIA AND METHODS OF CULTIVATION. 



121 



a long filament, and the subsequent development of brigiit oval 
spores. It is necessary carefully to observe the minutest details 
in order to maintain the cultivation pure. An excavated shde is 
thoroughly cleaned, and then sterilised by being held with the 
cupped side downwards in the flame of the Bunsen burner. A ring 
of vaseline is painted round the excavation, and the slide is then 
placed under a glass bell. Meanwhile a carefully cleansed cover- 
glass is also sterilised by passing it through the flame, and should 
be deposited on a sterilised glass plate. With a sterihsed looped 
needle, a drop of sterile broth is transferred to the cover-glass, 
and this is inoculated by touching it with another sterilised needle 
charged with the material to be examined, without disturbing 
the form of the drop. It is quite suflicient just to touch the drop 




.-3 



Fig. 48. — Deop Cultivation. 
(a) Drop of broth ; (b) layer of vaseline. 



instead of transferring a \'isible quantity of blood, juice, or growth, 
as the case may be. The slide is then inverted and placed over the 
cover-glass, so that the drop will come exactly in the centre of the 
excavation, and is gently pressed down. On turning the shde over 
again the cover-glass adheres, and an additional layer of vaseline 
is painted round the edges of the cover-glass itself. The slide must 
be labelled, and if necessary, placed in the incubator, and the results 
watched from time to time. Instead of broth, liquid blood serum 
may be employed in this form of cultivation. If it is required 
to preserve the drop-cultivation as a microscopic preparation, the 
cover-glass is gently lifted off and allowed to dry. Any vaseline 
adhering to the cover-glass should be wiped off, and the cover-glass 
can then be passed through the flame and stained in the usual manner. 
3foist Cells.— Unlens drop-cultures are very carefully prepared 



122 BACTERIOLOGY. 

they are liable to dry up, if kept for examination for several days. 
Many therefore prefer employing a moist cell, of which there are 
several different forms in use. 

The drop-culture slide may be converted into a moiist cell 
by having a deep groove cut round the circumference of the con- 
cavity. This groove is filled with sterilised water by means of a 
pipette. A ring of vaseline is painted with the camel's-hair brush 
outside the groove, and the cover-glass, with the drop-cultivation, 
is inverted and placed over the concavity. This form is very useful, 
as the slide can be easily cleansed and effectually steriUsed by 
holding it in the flame of the Bunsen burner. 

A very simple form of moist cell recommended by Schafer 




Fig. 4tl.— Simple Method of Forjiing a Moist Cell. 

may be used in some cases, but posse.sses the disadvantage of not 
admitting of sterilisation by heat. A small piece of putty or 
modelling wax is rolled into a cord about two inches long and g inch 
thick. By uniting the ends a ring is formed, which is placed on the 
middle of a clean glass slide. A drop of water is placed in the 
centre of the ring, and the cell roofed in by applying a cover-glass. 

A cell somewhat similar in form, which has the advantage of 
permitting of thorough cleansing, may be constructed by cementing 
a glass ring with flat surfaces to an ordinary slide. Vaseline is 
applied with a camel's-hair brush to the upper surface of the ring, 
and one or two di'ops of water placed with a pipette at the bottom 
of the cell. The cover-glass, with the preparation, is then inverted 
over the cell and gently pressed down upon the glass ring. The 
vaseline renders the cell aii'-tight, and, to a certain extent, fixes 
the cover-glass to the ring. 



NUTRIENT MEDIA AND METHODS OF CULTIVATION. 



123 



Warm Stages. — To apply warmth while a preparation is under 
continuovis observation, we must either place the microscope Vjodily 




Fig. 50. — Wakm Stage. 



within a special incubator, with the eye-piece protruding through an 
opening, or we must employ some means of applying heat directly 
to the prepai-ation. 

A simple warm stage may V)e made of an oblong copper plate, 




Fig. .51.— Warm Stage showx in Opekation. 

two inches long by one inch wide, from one side of which a rod of 
the same material projects. The plate has a round aperture in the 



124 



BACTERIOLOGY. 



middle, half an inch in diameter, and is fastened to an ordinary 
slide with seahng-wax. The drop to be examined is placed on a 
large-sized cover-glass and covered with a smaller one. Ohve oil 
or vaseline is painted round the edge of the smaller cover-glass to 
prevent evaporation, and the preparation is placed over the aperture 
in the plate. 

The slide bearing the copper plate is clamped to the stage of 




Fig. 52. — Israel's Waeming Appakatus in Opkkation. 



the mici'oscope. The flame of a spirit-lamp is apphed to the 
extremity of the rod, and the heat is conducted to the plate and 
thence transmitted to the specimen. In order that the temperature 
of the copper plate may be approximately that of the body, the lamp 
is so adjusted that a fragment of cacao butter and wax, placed close 
to the preparation, is melted. 

Israel's Warming Apparatus. — It is obvious that in employing 
very high powers a difficult}' will be presented by the warm stages 



NUTRIENT MEDIA AND METHODS OF CULTIVATION. 125 

commonly used for accurate observations, such as Sohafer's or 
Strieker's, owing to their interference with the ilhimination. To 




Fig. 53.— Section of Iskael's Warming Appahatuk and Dkop-cultuee 

Slide. 

overcome this an apparatus has been constructed by wdiich the 
sHde is warmed from above (Figs. 52, 53). 

The drop-culture sHdes are provided with a shallow groove, -1 mm. 
deep and 1 mm. broad, cut round the concavity. Into this the 
cover-glass fits, so that its upper .surface is level with that of 




Fig. 54.— Ishael's War.ming Apparatus. 

the slide. The heating apparatus consists of a flat disk-shaped box 
with a central conical aperture. 

The entrance and exit pipes are fixed on at a right angle to the 



126 



BACTERIOLOGY. 



side (Fig. 54). The former, s, is of metal, andihe latter, a, of glass 
fitted with a thermometer, the bnlb of which, k, is contained within 
the box. A partition, s, keeps up a current between the openings 




Fig. 



55. — Gas Ghambek in use with Appaeatus fou Generating 
Caebonic Acid. 



of the pipes, which are supported on a stand and connected by 
tubing with the hot-water supply. 

A mixture of paraffine and vaseline is recommended for indicating 
the temperature of the chamber, and experience has shown that if 
a temperature of 37° 0. is required the temperature of the water 
in the box must range between 42° and 47° C. 




Fig. 56.— Gas Chambek. 

Gas Ghamhers. — To investigate the action of gases or vapours 
upon micro-organisms, a modification of the moist cell may be 
emplo3'ed. 

A jjiece of glass tubing is first fixed to the slide by means of 



NUTRIENT MEDIA AND METHODS OF CULTIVATION. 



127 



sealing-wax, and the ring of putty is so placed as to include one end 
of it, leaving a small interval at the side, or a little notch is made 
in the putty opposite, so as to afford an exit for the gas or vapour. 




Fig. 57.— Moist Cell adapted fok Thansmission of Electricity. 

ApiMcation of Electricitij. — To study the effect of electricity 
we may prepare a drop-culture in the moist cell. The cover- 
glass to be used is provided with two strips of tinfoil, which are 




Via. 58. — Apparaths akkanged for Transmitting Electricity. 



isolated from the bi'ass of the microscope, and so arranged that a 
current of electricity may be pa.ssed through them. 

A much simpler plan, which may also be emidoyed, is to take 
an ordinary gla.ss slide and coat the surface with gold-size. The 



128 



BACTEHIOLOGY. 



slide is tlien pressed firmly down on gold-leaf or tinfoil and allowed 
to dry. When dry, the metal is scraped away, leaving two triangles 
with a small interval between tliem. 




Fig. 59.— Slide with Gold-le.\f Electrodes. 

The liquid containing the micro-organisms is placed between the 
electrodes, covered with a cover-glass, and then subjected to the 
electric current. 

(g) Methods of Employixg and Stoking Liquid Media. 

Cultivations in liquid media can be carried on in test-tubes, but 
it is more satisfactory to employ special forms of flasks, bulbs, and 
(J tubes, such as those employed by Pasteur and his school, and by 
Lister, Sternberg, and Aitken. 

Lister s Flasks. — These flasks were especially introduced by Lister 
as a means of storing liquid nutrient media. 
They are so constructed that after removal 
of a portion of the contents, on restoring 
the vessel to the vertical position, a drop of 
liquid always remains in the extremity of 
the nozzle, which jjrevents I'egurgitation of 
unfiltered air. 

Sternberg's Bulbs. — The method of intro- 
ducing liquid into the bulb employed by 
Sternberg, and of .stei-ilising and inoculating 
it, is as follows : The bulb is heated slightly 
over the flame, and the extremity of the neck, after the sealed point 
has been broken off, is plunged beneath the surface of the liquid. 
As the air cools the liquid is drawn into the bulb, 
usually filling it to about one-third of its capacity. 
The neck of the flask is again sealed up, and the 




Fig. 60.— Lister's 
Flask. 



Fig. 61.— Stekn- 
bekg's Bdlb. 



hquid which lias been introduced is sterilised by 
repeatedly boiling the flasks in the water-bath. 
They should then be placed in the incubator for two or three days ; 



NUTRIENT MEDIA AND METHODS OF CULTIVATION. 



129 




Tig. 62.— Aitken's 
Tube. 



and if the contents remain transparent and free from film, they 
may be set aside as stock-bulbs, to be used when required. 

To inoculate the liquid in the bulb the end of the neck is heated 
to sterilise the exterior, the bulb is gently warmed, and the extremity 
of the neck nipped off with a pair of sterilised forceps. The open 
extremity is plunged into the liquid containing the micro-organisms, 
and a minute quantity enters the tube and mingles with the fluid in 
the bulb without fear of contamination by atmospheric germs. The 
extremity of the neck is once more sealed up in the flame of a 
Bunsen burner. 

Aitken's Tubes. — These tubes are plugged and sterilised, and the 
nutrient medium introduced as into ordinary test-tubes. Instead 
of withdrawing the cotton-wool plug, they are 
inoculated through a lateral arm. The sealed 
extremity of the arm is nipped oflf with 
sterilised forceps, and the inoculating needle is 
carefully introduced through the opening thus 
made. It is directed along the arm until it 
touches the opposite side of the test-tube, where 
it deposits the material with which it was charged. 
The needle is withdrawn, and the end of the 
lateral arm again sealed up in the flame ; the test-tube is then 
tilted until the liquid touches the deposited material ; on restoring 
the tube to the vertical, the material is washed down with the 
nutrient liquid. 

Miguel's Bulbs. — The tube ct 
boule of Miquel is also a very 
useful form. It consists of a 
bulb of 50 cc. capacity, blown 
in the middle of a glass tube. 
The part of the tube above the 
bulb is contracted in two places, 
and can either be left quite 
straight or made to curve 
slightly. Between the contrac- 
tions the tube is plugged with 
asbestos. The portion of the 
tube below the bulb is S shaped, 
and drawn out at its extremity 

into a fine point. The bulb is charged with nutrient liquid and 
inoculated by aspiration, and the point of the S tube sealed in the 
flame of a Bunsen burner. 

9 




Fig. 63.— Miquel's Bulb. 



130 



BACTERIOLOGY. 




Fig. 64. — Pasteur's Flask. 



Pastern's Apparatios. — Special forms of tubes, bulbs, and pipettes 
are employed by the school of Pasteur. The tubes are provided 

with lateral or with 
curved arms drawn 
out to a fine point, 
and with slender 
necks plugged with 
cotton-wool. A 
double form (Pig. 65) 
shaped like a tuning- 
fork, each limb with 
a bent arm, is con- 
venient for storing 
sterihsed broth. The 
sealed end of an arm is nipped off with sterilised forceps, the sterile 
broth aspirated into each limb, and the arm again sealed in the 
flame ; a series of such tubes can be 
arranged upon a rack on the working 
table. 

Bulbs with a vertical neck drawn out 
to a fine point, others with a neck bent 
at an obtuse angle, plugged with cotton- 
wool, and a lateral curved arm drawn 
out to a fine point, are also employed. 
For a description of these various vessels 
and their special advantages, the woi-ks 

of PasteiTr and Duclaiix must be con- „ , ^ 

Fig. 65.— Pasteue's Double 
suited. Tpbj,. 




(h) Cultivation of Anaerobic Bacteria. 

To cultivate anaerobic organisms the same media are employed 
as for aerobic oi-ganisms, but the methods must be modified, or special 
apparatus used, so that the oxygen in the air may be excluded. 

In the preparation of plate-cultivations, before the film of gelatine 
has completely hardened it is covered with a sheet of mica, and the 
edges are sealed with melted paraiiine. By this process the air is not 
completely excluded, so that only those organisms which are not 
strictly anaerobic can be grown by this method. Liborius recom- 
mends boiling a considerable volume of gelatine in a tube, cooling it^ 
and after thoroughly distributing the organisms in the still liquid jelly, 
rapidly solidifying it by placing the tube in iced water. By this 



NUTRIENT MEDIA -AND METHODS OF CULTIVATION. 



131 




process very little air re-enters the jelly, and colonies of even strictly 
anaerobic bacteria will develop in the lower part of the tube. The 
drawback is the difficulty encountered in examining the colonies, 
and in preparing sub-cultures. For this purpose the tube 
must be broken, or carefully warmed until the jelly can be 
shaken out. 

Esmarch first prepares a roll culture, and when the gelatine film, 
has set, the tube is completely filled with liquefied gelatine which has 
been cooled down almost to the temperature at which it solidifies. The 
same diflficulty arises £ 
in the previous method, in 
the examination of the 
colonies. 

Buchner places the 
culture tube inside a much 
larger tube containing a 
small quantity of pyro- 
gallic acid and closed with 
a gutta-percha cap. The 
pyrogallic acid absorbs the 
oxygen, but the method 
is not altogether success- 
ful. 

The most satisfactory 
plan is to exhavist the air 
with an air pump, or to 
substitute an atmosphere 
of hydrogen which does 
not affect the growth of 
the bacteria. 

"Various forms of flasks 
and tubes for cultivating ^^^ ee.-FBANKEL's Anaekoeio Tube-culture" 
bacteria have been devised, jj_„^ gjass tube through which hydrogen is 
which can be easily con- passed; b, exit tube; e, india-rubber 

nected with an exhausting ^^X^LANnr "*""''" "^'^ '""*'' 

apparatus, and • readily 

sealed by the flame of the blowpipe when the air has been removed. 

If hydrogen is employed the most convenient plan is to use 
a Zipp's apparatus, from which the hydrogen is passed through 
two bottles, one containing a solution of lead, to remove any 
sulphuretted hydrogen, and the other pyrogallic acid, to intercept 
any oxygen. 



132 



BACTERIOLOGY. 



In the method recommended by Frankel a tube of gelatine is 
Hquefied, and inoculated. A gutta-percha stopper is substituted 
for the cotton-wool plug (Fig. 66). It is perforated by two holes, 
through which two tabes pass which are bent at a right angle. 
One tube only just passes through the stopper, the other reaches 
down to the bottom of the test-tube. The 
horizontal part of each tube has a narrow 
neck. The long tube has a plug of steril- 
ised cotton-wool, and is connected with 
a short piece of india-rubber tubing by 
which it can be connected with Kipp's 
apparatus. The hydrogen di'ives the air 
out of the liqviefied jellj^ and out of the 
test-tube, and after about half an hour ■ 
the horizontal tubes are sealed up, and 
the test-tube is made into a roll culture. 

Liborius employs a tube with a narrow 
neck and a lateral arm (Fig. 67). The 
tube is filled up to the height of the arm 
with either nutrient agar or a mixture of 
nutrient agar with 2 per cent, of grape- 
sugar. The hquefied jelly is inoculated 
in the usual way, and hydrogen passed 
through the lateral arm. When the air 
has been completely driven out, the tube 
is sealed up. 

To cultivate anaerobic organisms in 
broth, such as the tetanus bacillus, a flask 
is inoculated with the bacillus, and a 
stream of hydrogen is passed through the 
broth by means of a tube passing down to the bottom of the 
flask. The air in the flask escapes by a lateral arm which is bent 
■down at a right angle, and immersed in a capsule of mercury. 
When the air has been completely expelled the entrance tube is 
hermetically sealed, and the mercury in the capsule prevents any 
air from re-entering the flask by the lateral arm (Fig. 68). 




Fig. 67. — Anaeeobic Cul- 

TUEE-TUBE (LiBOEIUS). 



Method of Fixing Cultures. 



The colonies in plate-cultivations and the growths of bacteria 
in test-tubes may be stopped at any stage of their growth, and 
permanently fixed by exposing the culture to the fumes of formic 



NUTRIENT MEDIA AND METHODS OF CULTIVATION. 



133 



aldehyde. The test-tubes, dishes, or capsules are placed in a cylin- 
drical glass vessel containing a pledget of cotton- wool moistened with 




Fig. 68. — Apparatus foe Anaeeobic Cultures, 
(roscoe and lunt.) 



formic aldehyde. The vessel is fitted with a ground glass stojjper 
and set aside. The growth almost immediately ceases. Any 
liquefied gelatine is hardened, so that the exact appearances of 
cultures are obtained in a permanent form. 



CHAPTER X. 

EXPERIMENTS UPON THE LIVING ANIMAL. 

To carry out the last of Koch's postulates, and so complete the 
chain of evidence in favour of the causal relation of micro-organisms 
to disease, and to study the mode of action, of a pathogenic bacterium, 
it is necessary to introduce into a living animal a pure cultivation 
of the micro-organism or its chemical products. For this purpose 
various animals are employed, such as mice, rabbits, guinea-pigs, 
pigeons, and fowls. 

Inhalation. — The animals may be made to inhale an atmosphere 
impregnated with micro-organisms by means of a spray. In this 
way Friedlander succeeded in administering the bacteria of pneu- 
monia to mice ; and the production of tuberculosis by experimental 
inhalation has thrown light upon the clinical records of cases 
reported as instances of the infectiousness of phthisis. 

Ingestion. — A sheep fed upon potatoes which have been the 
medium for the cultivation of the anthrax bacillus dies in a few 
days. Babbits fed on cabbage sprinkled with a culture of the 
bacillus of fowl cholera, rapidly succumb to the disease. Animals 
fed upon the nodules of bovine tuberculosis or upon tubercular 
flesh and milk will be readily infected. 

Milk, or bread soaked in milk, is a very convenient medium, 
and from a public health point of view, a most instructive way of 
administering and testing the effect of pathogenic bacteria. 

Vaccination and Subcutaneous Inoculation. — Vaccination may be 
performed by making a few superficial scratches and inoculating the 
wound with a steriHsed platinum needle charged with the micro- 
organisms. Another simple method is to take a sterilised scalpel, 
infect the point with the material to be inoculated, and then make 
a mmute puncture or incision. In either case a situation should be 
selected, such as the root of the ear, which cannot be licked by the 
animal after the opejation. 



EXPERIMENTS UPON THE LIVING ANIMAL. 



135 



Subcutaneous inoculation is very simple and effectual, and con- 
sequently the method most frequently employed. The animal 
selected — for example, a guinea-pig — is held by an assistant, who 
covers it with a towel, leaving only the hind extremities exposed. 
By so doing, and gently laying it upon its back, with its head low, 
a guinea-pig passes apparently into a state of hypnotism, and the 




^SfW*' 



if Itr^fMI IwSaiBMfifaSlillS 



Fig. 09. — Koch's Syhinoe. 



trivial operation can be performed with little or no movement on the 
part of the animal. From a spot on the inner side of the thigh the 
hair is cut close with a small pair of scissors curved on the flat, and 
the skin must be thoroughly purified with 1 in 20 carboHc acid. A 
small fold of skin is then pinched up with a pair of sterilised forceps, 
and with a pair of sharp sterilised scissors, or with a tenotomy knife, 
a minute incision is made. A sterilised platinum loop is charged 
with the material to be inoculated, and the loop is gently inserted 
under the skin, forming a small pocket in the subcutaneous tissue. 
The needle is then withdrawn, and the sides of the wound gently 
pressed into apposition and painted over with collodion. 




Fig. 70. — Syeinge with Asbestos Plug. 

In inoculating a mouse the same process is adopted, with the 
exception that the root of the tail is the usual site of the 
operation. 

In some cases it may be necessary to inoculate cultures diffused 
in sterilised salt solution, or blood or lymph containing bacteria, 
or a culture in broth, or a filtrate containing the toxic products. 



136 BACTERIOLOGY. 

and then a hypodermic syringe may be required. One o£ the ordinary 
pattern may be \ised, but it is very much better to employ a syringe 
which has been especially constructed to admit of thorough dis- 
infection. Koch's syringe is a convenient form, the liquid being 
expressed by pressure on a rubber ball. 

The author has generally preferred to improvise a substitute for 
the hypodermic syringe which can be quickly made, and is destroyed 
after use, so that there can be no possible risk of accidentall}' 
infecting other .animals. A short length of ordinary glass-tubing is 
sterilised, and plugged at one end with sterilised cotton- wool ; about 
three inches from the plug a bulb is blown about the size of a 
marble, and two inches below this the glass is drawn out into a long 
capillary tube. A sufficient quantity of the liquid to be injected rises 
up into the tube by capillary attraction, or can be drawn up by 
means of an india-rubber ball, until the bulb is full. The point of 
the capillary tube is inserted through the opening in the skin, and 
gently pushed into the subcutaneous tissue, and then withdrawn for 
a short distance. By pressure on the bulb the contents of the tube 
are injected. In dealing with chemical products there is no risk in 
applying the lips and blowing out the contents of the tube, or indeed 
of fiUing it by suction, for if too much force were applied the liquid 
which might enter the mouth would be stopped by the cotton- wool 
plug. 

A number of these capillary tubes can be placed in a small case, 
and when it is necessary to go to a distance to investigate an outbreak, 
they will be fovind most convenient to bring back lymph or blood to 
the laboratory for further study. 

Sternberg takes a piece of glass-tubing, blows a bulb at one end, 
and draws oiit the other end into a thin tube. By heating the bulb 
and then dipping the tube into the liquid to be inoculated the latter 
rises in the tube as the bulb cools. After inserting the point of 
the tube subcutaneously the bulb is again heated, and the liquid is 
forced out into the tissues. 

Inl/i'avenous Injection. — A cultivation of micro-ox-ganisms may be 
mixed with sterilised water, and then injected with a syringe directly 
into the circulation. This may be performed without much difficulty 
by injecting, with a hypodermic syringe, the large vein at the base 
of the ear in rabbits, or the jugular vein in large animals. 

Special Operations. — In many cases it is absolutely necessary to 
perform an operation of greater severity. After the administration 
of an anaesthetic, infective mateiial may be inserted, or injected, into 
the peritoneal cavity, or injected into the duodenum in the manner 



EXPERIMENTS UPON THE LIVING ANIMAL. 137 

employed in the case of Koch's comma bacilli by Nicati and Rietsch. 
In such cases antiseptic precautions must be rigidly followed, and 
use made of iodoform and other antiseptic dressings. The disinfec- 
tion of the skin of the animal, of the instruments employed, and of 
the hands of the operator, are details essential to secure success. 

To inoculate tubercular matter, sputum may be rubbed up with 
distilled water, and some, of the mixture injected into a tracheal 
fistula ; or the first steps of the operation of iridectomy may be 
performed and tubercular material inserted into the anterior chamber 
of the eye, but this method is only justifiable when it is absolutely 
necessary for the results and changes to be observed from day 
to day. 

To inoculate rabbits or other animals with the virus of rabies, 
the skull is trephined, and an emulsion prepared from the spinal cord 
of a rabid animal is injected beneath the dura mater. 

Before every inoculation the instruments must be sterilised in a 
hot-air sterihser or by immersion in boiling water in a flat dish oi' 
enamel tray heated by a spirit-lamp, and after each operation all 
instruments should be placed in carbolic acid (1 in 20) or in boiling 
water, wiped dry, and again sterilised in the hot-air steriliser, before 
they are put away. If these precautions are not observed, instances 
of accidental infection are sure to occur. 

After the inoculation is completed a careful record must be made 
of the date and details of the experiment. The form in which the 
virus was used, the quantity employed, and the seat of inoculation, 
must be taken into account. The animals must be kept under close 
observation, the temperature taken, and any signs of illness, such as 
ceasing to feed, difiiculty in breathing, staring coat, and any local 
signs, such as the development of a tumour or an enlargement of the 
lymphatic glands, must be carefully noted. 

It is perhaps hardly necessary to add that in this country no 
experiments of any kind may be performed on living animals without 
a license. 

Method op Dissection and Examination. 

All animals that die after an experimental inoculation should 
be examined immediately after death. Every precaution must be 
taken in conducting the dissection, to exclude extraneous micro- 
organisms, and all instruments employed must have been sterilised 
in the hot-air steriliser, or by inlmersion in boiling water. If a mouse, 
for example, has died after inoculation with anthrax, it should be at 



\^8 BACTERIOLOGY. 

once pinned out by its feet on a slab of wood or in a gutta-percha 
tray, and bathed with 1 in 40 carbolic. In the same way, before 
examining a dead rabbit, a stream of carbolic should be directed 
over it to lay the fur, which otherwise interferes with the dissection. 
The hair should be cut away with sterilised scissors from the seat 
of inoculation, which is the first part to be examined, and any 
suppuration, haemorrhage, oedema, or other pathological change should 
be carefully noted. From any pus or exudation that may be 
present, material for inoculations should at once be taken, and 
cover-glass-preparations made for microscopical examination. 

To examine the internal organs and to make inoculations from 
the blood of the heart or spleen, the skin is cut through from below 
upwards in the median hne of the abdominal and thoracic regions. 
The abdominal cavity is then opened, and the walls pinned back, 
on either side of the animal. Any abnormal appearances in the 
peritoneum should be noted, and the state of the spleen should be 
carefully examined by turning the intestines aside. After noting 
its appearances, it should be removed with sterilised forceps and 
scissors, and deposited upon a sterilised glass slide, and incised with 
sterilised scissors. The cut surface is then touched with the point 
of a sterilised inoculating needle, and cultures are made in test-tubes 
of nutrient gelatine and agar-agar, and also on potato, and in broth 
in the form of drop -cultivations. Precisely the same care must be 
taken in examining lymphatic glands, tubercles, or pathological 
nodules ; any chance putrefactive micro-organisms on the surface 
should be destroyed by carbolic acid or the actual cautery; an 
incision is then made, and a minute fragment snipped out of the 
centre of the nodule, which can be inoculated in the living animal or 
transferred to a cultivating medium. 

The examination of the thorax is made by cutting through the 
ribs on either side of the sternum with sterilised scissors, and 
turning the sternum up where it will be out of the way. The 
pericardium is then opened, and the right auricle or ventricle pierced 
with the point of a sterilised scalpel, and inoculations and cover- 
glass-preparations are made from the blood which escapes. 

The lungs also require to be especially studied. They should be 
incised with a sterihsed scalpel, and inoculations and cover-glass- 
preparations made from the ciit surface. It may be necessary to 
embed a piece of lung or fragment of spleen, so that it shall be free 
from air. This may be done by isolating a fragment with the 
precautions just described, and depositing it upon the surface of a 
test-tube of nutrient agar-agar. The contents of another tube. 



EXPERIMENTS UPON THE LIVING ANIMAL. 139 

which have been, liquefied, and allowed to cool almost to the point 
of gelatinisation, must then be poured over it. From a potato a 
little cube must be cut, the tissue deposited in the trough thus 
formed, and the cube replaced, or cultures may be prepared by any 
of the methods which have been described for dealing with anaerobic 
bacteria. Blood may also be taken directly from a vein by laying it 
bare by dissection, making a small opening with sterilised scissors, 
and inserting a looped platinum needle, the needle of a hypodermic 
syringe, a capillary tube, or the extremity of the capillary neck of 
a Sternberg's bulb. If the cultivation, in spite of these precautions, 
is contaminated, or if there was more than one organism present 
in the blood or tissues under examination, it will be necessary to 
separate the different kinds by plate-cultivation. 

Having completed the dissection, the organs of suchi a small 
animal as a mouse may be removed en masse, and transferred to 
absolute alcohol for subsequent examination. In other cases it may 
be only necessary to reserve portions of each organ. In experimenting 
with a virulent micro-organism like anthrax, any remaining part of 
the animal should be cremated, and the hands and all instruments 
should be thoroughly disinfected. 

Isolation of Micro-organisms during Life. — Micro-organisms in 
the Jiving subject may be isolated from the pus of abscesses, or 
other discharges, and from the blood and tissues. Abscesses should 
be opened, and other operations performed, when practicable, with 
Listerian precautions, and a drop of the discharge taken up with 
.1 looped needle or capillary pipette, as already explained." 

To make a cultivation from the blood of a living person, the tip 
of a finger must be well washed with soap and water and sponged 
with 1 in 20 carbolic. Venous congestion is produced by applying 
an elastic band or ligature to the finger, which is pricked with a 
.sterilised sewing needle. From the drop of blood which exudes the 
necessary inoculations and examinations can be made. Another 
way of extracting blood from the living patient is to apply a leech. 
This method has -been found of considerable value in experimenting 
upon the blood of patients suffering from malai'ia, and may be 
useful in other diseases, if the blood is reqxiired for further 
examination, or in quantity. 



CHAPTER XI. 

EXAMINATION OF AIR, SOIL, AND WATER. 

AlE. 

The air, as is well known, contains in suspension, mineral, animal, 
and vegetable substances. . The mineral world is represented 
by such substances as silica, silicate of aluminium, carbbnate and 
phospate of calcium, which may be raised from the soil by the 
wind, and particles of carbon, etc., which gain access from acci- 
dental sources. Belonging to the animal kingdom we find the 
debris of perished creatures, as well as, sometimes, living animals. 
The vegetable world supplies micrococci, bacilli, and other forms 
of the great family of bacteria, spores of other fungi, pollen seeds, 
parts of flowers, and so forth. The air of hospitals and sick rooms 
has been found" to be especially rich in vegetable forms ; fungi 
and spores have been stated to be present in particularly large 
numbers in cholera wards; spores of tricophyton have been dis- 
covered in the air of hospitals for diseases of the skin, and of achorion 
in wards with cases of favus. The tubercle bacillus is said to have 
been detected in the breath of patients suffering from phthisis. 

These points indicate that, in addition to the interest for the 
micro-biologist, considerable importance, from a hygienic point of 
view, must be attached to the systematic examination of the air, 
A knowledge of the microbes which are found in the air of marshy 
and other unhealthy districts, and in the air of towns, dwellings, 
hospitals, -njorkshops, factories, and mines, will be of practical value. 

Miquel, who has particularly studied the bacteria in the air, 
has found that their number varies considerably. The average 
immber per cubic metre of air for the autumn quarter at Mont- 
souris is given as 142, winter quarter 49, spring quarter 85, and 
summer quarter 105. In air collected 2,000 to 4,000 metres above 
the sea-level,'>not a single bacterium or fungus spore was found, 
while in 10 cubic metres of air from the Rue de Rivoli (Paris) the 
number was computed at 55,000. 

140 



EXAMINATION OF AIR, SOIL, AND WATER. 141 

The simplest method for examining the organisms in air consists 
in exposing plates of glass or microscopic slides coated with 
glycerine, or with a mixture of glycerine and grape sugar, which is 
stable, colourless, and transparent. Nutrient gelatine spread out on 
glass plates may be exposed to the air for a certain time, and then 
put aside in damp chambers for the colonies to develop. Sterilised 
potatoes, prepared in the usual way, may be similarly exposed. In 
both the last- mentioned methods separate colonies develop, which 
may be isolated, and pure cultivations carried on in various other 
nutrient media. Nutrient gelatine has also been employed in the 
special methods of Koch and Hesse. 

Koch's Appouratus. — This consists of a glass jar, about six inches 
high, the neck of which is plugged with cotton-wool. In the 
interior is a shallow glass capsule, which can be removed by means 
of a brass Kfter. The whole is sterilised by exposure to 150° C. 
for an hour in the hot-air steriliser. The nutrient gelatine in a 
stock- tube is liquefied, and the contents emptied into the glass 
capsule. The jarSs exposed to the air to be examined for a definite 
time, the cotton -wool plug replaced, and the apparatus set aside for 
the colonies to develop. 

Hesse's Apparatus. — The advantage of this apparatus is that it 
enables the experimenter to examine a known volume of air. A 
glass cylinder, 70 cm. long and 3"5 cm. in diameter, is closed at one 
end by an india-rubber cap, perforated in the centre. Over this fits 
another cap, which is not perforated. The opposite end of the 
cylinder is closed with a caoutchouc stopper, perforated to admit 
a glass tube plugged with cotton-wool. The tube can be connected 
by means of india-rubber tubing with an aspirating apparatus, 
which consists of a couple of litre-flasks, suspended by hooks from 
the tripod-stand which supports the whole apparatus. The cylinder, 
caps, and plug are washed with solution of carbolic acid, and 
then with alcohol. After being thus cleansed, 50 cc. of nutrient 
gelatine are introduced, and the whole sterilised by steaming for 
half an hour for three successive days. After the final sterilisation, 
the cylinder is rotated on its long axis, so that the nutrient medium 
solidifies in the form of a coating over the whole of the interior. 
When required for use, the cotton-wool plug is removed from the 
small glass tube, and the latter connected with the upper flask by 
means of the india-rubber tubing. 

The apparatus is placed in the air which is to be examined, the 
outer india-rubber cap removed from the glass cylinder, and the 
upper flask tilted until the water begins to flow into the lower one. 



142 



BACTERIOLOGY. 



The emptying continues by siphon action, and air is drawn in along 
the cylinder to replace the water. When the upper flask is empty, 
the position of the two is reversed, arid the flow again started. 
When a sufficient volume has been drawn through the cylinder, the 
outer cap and the cotton-wool plug are replaced, and it is set aside 
for the colonies to develop. As an example, twenty-five litres of air 
from an open square in Berlin gave rise to three colonies of bacteria 
and sixteen moulds ; on the other hand, two litres from, a school- 




FiG. 71. — Hesse's Apparatus. 



room just vacated by the scholars gave thirty-seven colonies of 
bacteria and thirty-three moulds. 

Porous substances, such as sand, powdered glass, or sugar, may 
be used for the filtration of samples of air ; and an apparatus is 
employed in a convenient form to be conveyed to the laboratory for 
the subsequent examination. 

Petri's Appa/ratus consists of a glass-tube 9 cm. long, containing 
two sand-filters separated from each other. A known volume of air 
is aspirated through the tube. The bacteria are arrested and can 



EXAMINATION OF AIR, SOIL, AND WATER. 



143 



be examined by spreading the sand out in a dish and covering it 
with nutrient gelatine ; or it may be shaken up with sterilised water 
and plate-cultivations prepared. The sand-filter nearest to the 
aspirator should remain free from bacteria. 

Sedgwick and Tucker employ a glass cylinder which is drawn out 
at one end into a narrow tube to contain sterilised powdered cane 
sugar. Both ends of the apparatus are plugged with sterilised 
cotton-wool. By means of an exhausting apparatus a known volume 
of air is drawn through the tube. The cotton-wool plug is re- 



> 



Fig. 72. — Sedgwick and Tuckek's Tube. 

moved, and liquid gelatine is introduced into the cylinder, the 
plug is replaced, and the sugar is shaken into and quickly dissolves 
in the jelly. The cylinder is then treated in the same way as a 
roll-culture, and set aside for the colonies to develop (Fig. 72). 

Various forms of " aeroscopes " and " aeroniscopes " have from 




Fig. 73. — Pouohet's Aekosoopk. 

time to time been employed. Pou chefs aeroscope consists of a small 
funnel, drawn out to a point below which is a glass slip coated with 



144 BACTERIOLOGY. 

glycerine. The end of the funnel and the glass slip are enclosed 
in an air-tight chamber, from which a small glass tube passes out 
and is connected by india-rubber tubing with an aspirator (Fig. 73). 
The air passing down the funnel strikes upon the glycerine, which 
arrests any solid particles. For a full description of the apparatus 
employed by Maddox, Cunningham, and Miquel, reference should 
be made to the writings of these authors, and particularly to the 
treatise published by the last-named. 

Soil. 

Surface soil is exceedingly rich in bacteria. Miquel has com- 
puted that there exists in a gramme of soil an average of 750,000 
germs at Montsouris, 1,300,000 in the Rue de Rennes, and 2,100,000 
in the Rue de Monge. As agents in putrefaction and fermenta- 
tion they play a very important rdle in the economy of nature ; 
but there exist in addition, barfteria in the soil which are patho- 
genic in character. Pasteur has succeeded in isolating the bacillus 
of anthrax from the earth. Sheep, sojourning upon a plot of 
ground where animals with anthrax have been buried, may succumb 
to the disease. Pasteur considered that the spores were conveyed 
by worms from buried carcasses to the surface soil. The bacilli 
of malignant oedema and tetanus are also present in soil. Nicolaier 
produced tetanus in mice and rabbits by inoculating a little garden 
earth under the skin. 

To obtain a cultivation of the microbes in soil a sample of the 
latter must be first dried and then triturated. It may then be 
.shaken up with distilled water, and from this a drop ti'ansferred to 
sterilised broth. The employment of solid media is, however, 
much more satisfactory: a sample of earth is collected, dried, and 
triturated, and a small quantity sprinkled over the surface of 
nutrient gelatine prepared for a plate-cultivation. In another 
method the gelatine is hquefied in a test-tube, the powder added, 
and distributed, in the usual way, throvighout the medium, which 
is then poured out upon a glass plate or made into a roll-culture. 
In the same way the dust which settles from the air in houses and 
hospitals, or food substances in powder, may be distributed in 
nutrient gelatine, and examined both for aerobic and anaerobic 
bacteria. The different kinds which develop, must be thoroughly 
investigated as regards their morphological and biological charac- 
ters, and pathogenic properties. 



examination of aie, soil, and water. 145 

Water. 

In the case of water, as in that of air, a knowledge of the 
micro-organisms which may be present is not only of interest 
to the bacteriologist, but of the greatest importance in practical 
hygiene. Common putrefactive bacteria and vibrios may not be 
hurtful in themselves, but they indicate the probability of the 
presence of organic matter in which there may be danger. The 
detection of Bacillus coli communis may be taken to indicate a 
probable contamination with human excreta. 

The Microzyme Test which was introduced for the detection 
of putrefactive bacteria, consisted in adding three or four drops of 
the sample of water to 1 or 2 cc. of Pasteur's fluid, the nourishing 
fluid having been previously boiled in a steriHsed test-tube. If the 
microzymes or their germs existed in the water, the liquid in a few 
days became turbid from the presence of countless bacteria. This 
test is of no real value, for it does little more than indicate that 
bacteria are present, which we know to be the case in all ordinary 
water, and even in ice. On the other hand, the bacteriological test 
of Koch is a most valuable addition to the usual methods of water 
analysis. It enables us not only to detect the presence of bacteria, 
but to ascertain approximately their number, and to study very 
minutely their morphological and biological characteristics. The 
importance of a thorough acqtiaintance with the life-history of the 
individual micro-organisms cannot be too strongly insisted upon. 
For example, by such means the spirillum of Asiatic cholera can 
be distinguished from most other comma-shaped organisms, and 
inasmuch as its presence may be an indication of contamination 
with choleraic discharges, such water should be condemned for 
drinking purposes, even though we are not yet in a position to 
affirm that the microbe is the cause of the disease. The detection 
of the bacillus of typhoid fever or of the Bacillus coli communis 
in suspected water or milk would be evidence of considerable 
importance. 

Koch's test, in short, consists in making plate- cultivations of a 
known volume of water, counting the colonies which develop, 
isolating the micro-organisms, and stvidying the characters of each 
individual form. 

Collection and TroMsport of Water Samples. — Sternberg's bulbs, 
or Erlenmeyer's conical flasks of about 100 cc. capacity, may bo 
employed with advantage for collecting the samples of water. The 
latter are cleansed, plugged, and sterilised in the hot-air steriliser. 

10 



146 BACTERIOLOGY. 

When required for use, the plug is removed and held between the 
fingers, which must not touch the part which enters the neck of 
the flask. About 30 cc. of the water to be examined are intro- 
duced into the flask, and the plug must be quickly replaced and 
covered with a caoutchouc cap. If collected from a tap, the water 
should first be allowed to run for a few minutes, and the sample 
should be received into the flask without the neck coming into contact 
with the tap. From a reservoir or stream, the flasks may be filled 
by employing a sterilised pipette. During transport contact between 
the water and cotton-wool plug must be avoided, and if likely to 
occur the sample must be collected and forwarded in a Sternberg's 
bulb. 

ExaTnination hy Plate-cultivation. — The apparatus for plate- 
cultivation should be arranged as already described. Crushed ice 



Fig. 74. — Apparatus foe Estimating the Number of Colonies in a 
Plate-cultivation. 

may be added to the water in the glass dish to expedite the setting 
of the gelatine, so that the plate may be transferred as quickly 
as possible to the damp chamber. The caoutchouc cap is removed 
from the flask, and the cotton-wool plug singed in the flame to 
prevent contamination from adventitious germs on the outside of 
the plug. The flask is then held slantingly in the hand, and the 
plug twisted out and retained between the fingers. "With a 
graduated pipette a measured quantity (J^ or Jg- cc.) of the sample is 
transferred to a tube of liquefied nutrient gelatine, and the plugs of 
the flask and of the tube quickly replaced. If the water is very 
impure, it may be necessary to first dilute the sample with sterihsed 
water. The inoculated tube must be gently incHned backwards and 
forwards, and rolled as already explained, to distribute the germs 
throughout the gelatine, and the gelatine finally poured on a plate. 
When the gelatine has set, the plate is transferred to a damp 
chamber, which should be carefully labelled and set aside in a place 



EXAMINATION OF AIR, SOIL, AND WATER. . 147 

of moderate temperature. In about two or three days the cultivation 
may be examined. In some oases the colonies may be counted at 
once ; more frequently they are so numerous that the plate must 
be placed on a dark background, and a special process resorted to. 
A glass plate, ruled by horizontal and vertical lines into centimetre 
squares, some of which are again subdivided into ninths, is so 
arranged on a wooden frame that it can cover the nutrient-gelatine 
plate without touching it (Fig. 74). A lens is used to assist in dis- 
covering minute colonies. If then the colonies are very numerous, 
the number in some small division is counted, if less in some large 
one, and an average is obtained from which the number of colonies 
on the entire surface is calculated. A separate calculation of the 
Uquefled colonies should be also made, and their number, as well 
as the total number of colonies present in 1 cc. of the sample, 
recorded. Any peculiar macroscopical appearances, colours, etc., 
should be noted, and then the microscopical appearances of the 
colonies studied. Lastly, examination of the individual organisms 
should be made by cover-glass preparations, and by inoculation of 
nutrient gelatine, potatoes, and other media. 

Instead of plates, Petri's dishes may be used both for gelatine 
.and agar-agar cultivations. 




Fig. 75. — Bsmabch's Eoll-Cultube. 

■a, India-rubber caps; bhh, longitudinal line drawn on the tube ; ccc, transverse 

lines for counting colonies (Frankland). 

Another plan is to take a measured quantity of the sample of 
water and prepare a roll-culture, using a large-sized test-tube 
(Fig. 75). The colonies can be counted with the aid of a lens (Fig. 76). 
Microscopical preparations and sub-cultures can be made from the 
.colonies, and the anaerobic bacteria can be examined by Frankel's 
modification of this method (p. 131). 

A drop of the sample of water may also be added to liquefied 
nutrient gelatine in a test-tube, the organisms distributed, and the 
.gelatine allowed to solidify in the tube. A rough comparison of 
water samples may be made in this way. 

Microscopic Examination. — A drop of the water may be mounted 
^nd examined without staining ; or allowed to evaporate on a cover- 



148 BACTERIOLOGY. 

glass, which is then passed through the flame, and stained in the- 
usual manner. 

Parietti's Method. — As tj^phoid fever bacilli are apt to be 
crowded out by more rapidly growing micro-organisms, some method 
had to be devised for restraining the growth of the latter, and 
Ohantemesse and Widal suggested the use of carbolic acid. Parietti 
put this into practice by the method he introduced. This consists in 
adding to tubes of broth about five drops of a mixture composed of 
sterilised water (100 parts), hydrochloric, acid (4 parts), and carbolic 
acid (5 parts). The tubes are first tested by incubation, and are 
then readv for use. A few drops of the suspected water are added 



Fig. 76.— Apparatus for Counting Coloniks in a Roll Culture. 

to the broth, and if it becomes turbid in a day or two the typhoid 
fever bacillus is present in the form of a pure-culture. 

An excess of bacteria in a fresh sample indicates an excess of 
organic matter, and points to possible contamination with sewage. 
Where there is such contamination we are very likely to find 
pathogenic bacteria ; and moreover impure water is a constant 
source of danger, for if the contagia of infectious diseases are 
introduced they will retain their vitality in such water for a long 
period, and will in some cases even multiply, whereas the same 
organisms inti'oduced into pure water wovild in a short time perish. 

The actual number of bacteria in water is not of very great 
importance, and it must be remembered that if a sample is set aside 
for a few days there will be an enormous increase in the number 
of bacteria present ; but in dealing with perfectly fresh samples it 



EXAMINATION OF AIR, SOIL, AND WATER. 149 

"may be said that water containing less than 100 bacteria to the 
<!ubic centimetre is very pure water. Water containing 1,000 or 
more should be filtered. Water containing 100,0.00 to 1,000,000 
is contaminated with surface water or sewage. It is necessary to 
bear in mind that in typhoid fever and Asiatic cholera the excreta 
contain the bacteria in great numbers, and wells and streams 
receiving surface water may be contaminated in various ways. 
The cholera bacillus dies as a rule quickly in distilled water, while 
it preserves its vitality for a long time in water of a bad quality. 

It is necessary to lay stress upon the fact that a bacterio- 
logical analysis may show the presence of pathogenic bacteria when 
their detection is not possible by any other means. They may 
be present in water in such small numbers that no chemical 
analysis would detect any contamination, but as they are living 
organisms capable of increasing in a suitable environment, they can 
readUy be discovered by bacteriological methods. 

The examination of rain water, drinking water, tap water, sea 
water, various liquids and infusions, by these methods, opens up 
a wide field for research. Pettenkofer has shown that impregna- 
tion of water containing many bacteria with carbonic acid diminishes 
the number of the latter. The examination of waters before and 
after filtration, or after addition of chemical substances, are matters 
which require further investigation, though a great deal of work has 
already been accomplished. The reader will find in Micro-organisms 
in Water by P. and G. Frankland, a very complete account of this 
subject with valuable analytical tables. 



CHAPTEE XII. 

PHOTOGRAPHY OF BACTERIA. 

The production of pictures of microscopic objects by photographic 
means was attempted at an early date. Some authorities regard 
the very earliest recorded experiments as being the first experi- 
ments alike in photography and micro-photography. The experi- 
ments of Wedgwood and Sir Humphry Davy were embodied in 
a paper read before the Royal Institution in 1802. They obtained 
with the solar microscope impressions upon paper, and with greater 
success upon white leather, though the results were transitory when 
exposed to daylight. 

In 1816 Mcephore Niepce described his experiments in con- 
nection with fixing the image obtained by the camera. He was 
at first only able to obtain negatives, and these were transitory. 
But, after joining with Daguerre, who had been experimenting 
in the same direction, a process was invented which was published 
in 1839 under the name of daguerreotype. 

This invention, and the rapid improvements which followed, 
were taken advantage of by Reade, Donne, Hodgson, Kingsley, and 
Talbot, who were early workers in the field of micro-photography. 

So early as 1845 it is stated that Donne produced a work 
illustrated with engravings copied from daguerreotypes. 

Subsequently this interesting branch of photography was taken 
up by many in France and Germany, in America, and in England. 
Of those to whom we are indebted for the literature of the subject, 
and for many improvements, the names of Wenham, Dancer, 
Draper, Maddox, Shadbolt, Redmayne, Woodward, Highley, Deecke, 
Moitessier, Gerlach, Koch, Sternberg, Frankel, PfeifEer, and Pringle 
may especially be mentioned. 

Of these workers the name of Woodward stands pre-eminently 
foremost. His skill in microscopical manipulations, combined 
with access to the very best apparatus and objectives, placed at 

150 



PHOTOGRAPHY OF BACTERIA. 151 

his disposal in the Museum at Washington, enabled him to obtain 
photographs of diatoms which probably have never been surpassed. 

To Koch belongs the credit of being the first to extend the 
application of micro-photography to the delineation of bacteria. 
A series of instructive photographs was first published by him 
in 1877. These were photographs of cover-glass preparations, 
and all admirably illustrated the subjects from which they were 
taken ; while two, showing the flagella of bacilli and spirilla, 
were triumphs in this new departure. 

Lewis, in India, was one of the first to illustrate his writings on 
the subject of micro-organisms by means of photographs. 

About the same tim.e Sternberg, in America, took some excellent 
photographs of bacteria. Heliotyps reproductions of these were 
published in 1884. 

Hauser and "Van Ermengem and many other bacteriologists 
successfully resorted to photography for illustrating their researches, 
and Frankel and Pfeiffer's, and Itzerott and Niemann's atlases of 
photographs of bacteria, in microscopical specimens and cultivations, 
are especially worthy of mention. 

Opinions have differed widely as to the merits of photographic 
illustrations. Many, taking the standpoint solely of a comparison 
with drawings, have decried their use. By judging from such a 
comparison alone the real value of photographs may be lost sight 
of. On the other hand, many who have looked at the question 
from all sides, have been led to value even a defective photograph 
more than an ordinary drawing. 

In his first publication on this subject, Koch strongly advocated 
photography on the ground that illustrations would then be as true 
to nature as possible. The photographs which accompanied his 
paper were all taken from preparations of bacteria which had 
been made from blood, cultivations, or infusions, by drying a 
thiri layer on a cover-glass and staining, or from specimens prepared 
in the same way but left unstained. But when, having committed 
himself to this opinion, Koch attempted, later, to photograph the 
bacteria in animal tissues, he was led to modify his previous 
conclusion. For though no trouble was spared, yet disappointing 
results were met with. This was owing, he explains, to the fact 
that the smallest and most interesting bacteria can only be made 
visible in animal tissues by staining them, and thus obtaining the 
advantage of colour. 

This introdviced the same diificulties which are met with in 
photographing coloured objects, such as tapestry and oil paintings. 



1 52 BACTERIOLOGY. 

As these difficulties had been to a certain extent obviated by the 
use of eosin-collodion, Koch adopted the same method for photo- 
graphing stained bacteria. By the use of eosin-collodion, and by 
shutting off portions of the spectrum by coloured glasses, he 
succeeded in obtaining photographs of bacteria which had been 
stained with blue and red aniline dyes. But, owing to the long 
exposure which was necessary, and the unavoidable vibrations of 
the apparatus, the results were so wanting in definition that they 
not only proved unsatisfactory as substitutes for drawings, but did 
not in some cases give any evidence of what was to be seen in the 
preparations. 

Koch, in consequence, stated that he would abstain from 
publishing photographic illustrations until he had the advantage 
of improved methods. 

We find, however, in spite of this, that in 1881 Koch published 
a series of reproductions from his negatives in illustration of what 
could be accomplished by photography. 

Here again we find that many of the photographs of cover- 
glass preparations were admirable, but those of tissue-sections gave 
evidence of the difficulties Koch encountered, and were undoubtedly 
unsatisfactory from the want of flatness of field, some of tlie 
illusti'ations recalling rather a map of a mountainous country than 
a microscopical preparation. 

In consequence of the difficulties met with in attempting to 
photograph bacteria stained with the aniline dyes most commonly 
used, Koch recommended that the preparations should be stained 
brown, pointing out as his reason that, though the bright and 
concentrated colour of the red and blue aniline dyes catches the 
eye far more readily than the somewhat sombre brown colours, 
yet no one up to the time of his publication had succeeded in 
obtaining good photographs of bacteria which had been stained 
either blue or red, and mounted in Canada balsam, while there was 
no difficulty in obtaining photographic representations of prepara- 
tions stained yellow or brown. 

Though this stain could be easily employed in most cover-glass 
preparations, it was by no means easy to obtain a good differential 
stain of bacteria in the tissues by employing Bismarck brown. 
An attempt was, therefore, made to photograph preparations 
stained blue and red by the aid of the dry-plate process, and by 
interposing glasses of suitable tints. After many fruitless experi- 
ments this method had to be abandoned, and the method of staining 
the object brown was adopted. In many cases this gave excellent 



PHOTOGRAPHY OF BACTERIA. 15^ 

results ; in others again, compared with the results of staining 
with blue or red stains, there was much to be desired, and further 
improvement was needed. 

That a stain, such as yellow or brown, must be employed which 
absorbs the blue rays, and acts on the sensitive plate like black, 
which absorbs all the light, constituted the first condition laid 
down by Koch as an essential for success. It was further pointed 
out that the suitability of the stain could be ascertained by first 
passing the light, to illuminate the preparation, through a solution 
of ammonio-sulphate of copper, under which condition the bacteria 
would appear black on a blue ground. 

The second condition was, that sunlight must be employed, but 
that direct projection upon the object was disadvantageous, and it 
must, therefore, be diffused by the interposition of one or more plates 
of ground glass. 

Lastly, an illuminating condenser was recommended, of such 
construction that the diffused sunlight brightly illuminates the object 
from all sides. 

Sternberg encountered the same difficulty in photographing red, 
blue or violet preparations, while he produced excellent pictures of 
preparations stained with aniline brown, or a weak solution of iodine 
(iodine grs. iij, potassic iodide grs. v, distilled water grs. 200). Thus 
the results of a large number of attempts to photograph the tubercle 
bacillus in sputum, only ended in producing such extremely faint 
impressions, that any one unacquainted with the object as seen under 
the microscope could form scarcely any idea of its form or minute 
structure with even an accompanying explanation and the closest 
inspection of the photograph. 

Dufrenne, in attempting to photograph the same object by the 
ordinary method, found the plates were uniformly acted on, or the 
image was so faint, or so lacking in contrast, that they were useless 
for obtaining proofs on paper or glass. By interposing green glass 
between the objective and the sensitive plate, so that the red rays 
were absorbed, while the green rays passed through and acted on 
the plate, he states that better results were obtained. 

The work of Hauser illustrated the great value of photography 
in the production of pictures of impression-preparations and colonies 
in nutrient gelatine. To give the general effect, as well as faithfully 
reproduce the minute details in these difficult subjects would in most 
cases create insurmountable difficulties, except to the most accom- 
plished draughtsman. 

Hauser employed Gerlach's apparatus and Schleussner's dry 



154 BACTERIOLOGY. 

plates, and obtained the illumination by means of a small incan- 
descent lamp, which gave a strong, white light, with three or 
four Bunsen elements. In another respect Hauser's results were 
of practical value. The preparations to be photographed were 
stained brown as recommended by Koch, but they were mounted in 
the ordinary way in Canada balsam. The objection to the mounting 
medium most commonly employed was thus set aside. The prevalent 
idea, however, that the preparations must be stained brown was still 
a formidable obstacle, and the way out of this difficulty was clearly 
shown by Van Ermengem's photographs. These were pictures of 
comma-bacilli which had been stained with fuchsine and methyl 
violet. These photographs afforded the first practical illustration 
of the value of isochromatic plates in micro-photography which had 
been previously noted by Van Ermengem in 1884, and their intro- 
duction marks a distinct era in the progress of micro-photography. 

A .short explanation may be given of what is meant by isochro- 
matic, or what have been more properly termed orthochromatic dry 
platss. The difficulties encountered in photographing certain stained 
preparations have been mentioned. It is a familiar fact that in 
portraits, blue or violet comes out almost or quite white, while 
other colours, such as yellow, are represented by a sombre shade 
or perhaps black. This failure in correctly translating colours is 
explained by the want of equality between the strength of the . 
chemical and luminous rays. If the rays of the spectrum are pro- 
jected upon a photographically sensitive surface, the greatest effect 
is found to take place at the violet end. In other words, the violet 
and blue rays are more actinic or chemically powerful, while the 
yellow and orange have scarcely any effect. The dyes employed in 
staining give corresponding results : blue and violet give but a faint 
impression, yellow and orange a black picture. These results are 
most clearly demonstrated in a photograph of an oil painting taken 
in the ordinary way ; and they led to experiments being made which 
have resulted in orthochromatic photography. 

The effect of interposing coloured glasses has already been 
referred to. It was found later that, if plates were coloured yellow, 
e.g., with turmeric, the blue and violet rays were intercepted, and 
their actinism reduced. In 1881, Tailfer and Clayton produced the 
so-called isochi'omatic plates. The emulsion of bromide of silver 
and gelatine was stained with eosin, and it was claimed that colours 
would be represented with their true relative intensity. Chlorophyll 
and other stains have been tried, and by such methods the ordinary 
gelatine dry plates can be so treated that they will reproduce 



PHOTOGRAPHY OF BACTERIA. 155 

various colours, according to their relative light intensity, and thus 
be rendered iso- or, what is now more commonly known as, ortho- 
chromatic. 

Apparatcs and Material. 

Micro -photographic Apparatus. — As is well known, various forms 
of apparatus have from time to time been recommended and em- 
ployed by different workers. 

Many use the microscope in a vertical position, with the camera 
superposed or fitted to the eye-piece end of the microscope tube ; 
or the microscope tube may be screwed off from the body of the 
microscope, and a pyramidal camera, adjusted in its place, the base of 
the pyramid being represented by the ground glass screen. 

Others again prefer that the microscope and camera should be 
arranged horizontally. 

In another form the. ordinary microscope is dispensed with, the 
objective, stage, and mirror are adapted to the front of the camera, 
and provided with suitable arrangements for holding the object, 
supporting the mirror, and adjusting the different parts. 

Lastly, the camera may be dispensed with, the operating-room, 
wliich must be rendered impervious to light, taking its place, while 
the image is projected and focussed upon a ground glass screen, 
which has a separate support.* 

The horizontal position affords greater stability than the vertical, 
so that the former is to be preferred. The vertical model with the 
camera fixed to the microscope is particularly to be avoided, as the 
weight of the camera bears directly upon the microscope, and must 
affect the fine adjustment, and any vibration in one part of the 
apparatus is communicated throughout. 

The simplest apparatus consists of a camera fixed upon a base- 
board four or five feet in length, upon which the microscope can be 
clamped, and which cai'ries also a lamp and a bull's-eye condenser 
(Fig. 77). 

Simplicity and economy must always be borne in mind in 
recommending any apparatus of this kind, for to insist upon the 
necessity of a very elaborate apparatus, or a specially fitted- up room, 
or that a special room should be built with windows facing in a 
definite direction, will in most cases at once place photography beyond 
the reach of those who might otherwise employ it. Yet to fulfil 

* For an excellent account of the forms of apparatus which have been 
employed by different workers the reader is referred to the section on Micro- 
photography in Beale's Htm to imrk with the Microscope. 



156 



BACTERIOLOGY. 



all the purposes for which the apparatus may be required, inchidiug 
the employment of the highest powers, and also that one may be 
enabled to work for long intervals of time with due comfort, an 
accurate and complete apparatus will be found to be most desirable. 
Though most preparations will admit of being photographed 
when the stage of the microscope is vertical, yet if we require to 
photograph micro-organisms in liquids, or colonies upon partially 
liquefied gelatine, the apparatus must admit of being placed so that 
the stage of the microscope becomes horizontal. In addition, the 
apparatus is rendered somewhat complex if we employ powerful 



I 




Fig. 77. — Hohizoxtal Micko-photogkaphic Appaeatus. 



artificial light. Sunlight, no doubt, is the best and cheapest, but 
it is not always available, especially in a city like London ; and, 
moreover, evenings and dull days will probably be the very time 
which can be best spared for this work. We must, therefore, fall 
back upon the paraffine lamp, or the magnesium, oxyhydrogen, or 
electric light. 

To fulfil all these conditions Swift has constructed an apparatus 
under the author's directions (Fig. 78). It is merely a modification 
of the ordinary horizontal model, which admits of being readily placed 



PHOTOGRAPHY OF BACTERIA. 



157 



in the vertical position, while the illumination is supplied from an 
oxyhydrogeu lantern. 

To place the apparatus in the vertical position two small hinged 




brackets, at the end distant from the camera, are forced up with 
a smart blow of the hand. The corresponding ends of the stretcher 
bars are dislodged from their fittings, and allowed to descend ; when 



158 



BACTEHIOLOGY. 



horizontal, the opposite extremities of the bars are easily released 
from their sockets. The leg or support at this end cau then be 




Fig. 79. — Reversible Micro-photographic Apparatus arranged in the 
Vertical Position. 



turned up and fixed underneath the apparatus by a button, and 
the end of the apparatus itself gently lowered to the ground. 



PHOTOGRAPHY OF BACTERIA. 159 

A hinged end- piece is also to be turned out to increase the base upon 
which the whole apparatus will stand when raised to the vertical. 
The two-legged support at the opposite end of the apparatus is 
next worked down by a quick thread screw, and on raising the 
apparatus to the vertical, the two-legged support drops to the 
ground, and assists in maintaining the stability of the whole. If 
it is thought necessary, a simple means can be readily devised for 
clamping the apparatus, in either position, to the wall of the room, 
so as to eliminate as much as possible all chances of vibration. A 
second quick thread screw moves the base -board upon which the 
, camera and central shding-board are uiounted, so that the camera, 
iiiicroscope and lantern can be raised to a convenient height from 
the ground. 

The various parts of this apparatus, may be described in 
detail. 

The Microscope and its Attachments. — It is most essential that 
the micx'oscope should be perfectly steady. The microscope was made 
by Zeiss, and to ensure steadiness, the horse-shoe footpiece fits under 
a projecting ledge, and is then clamped by a cross-piece, so that 
it is- firmly fixed. 

The microscope with the means for clamping it and the oxy- 
hydrogen lantern are carried upon an independent sliding-board, 
which admits of movement to or from the camera. The sUding- board 
also moves upon a centre, which enables the microscope to be turned 
out from the median line ; in fact, to be turned at a right angle to the 
position it occupies when ready for the exposure. The object of this 
contrivance is to enable the operator to sit down by the side of the 
apparatus, and with comfort to arrange the object in the field of the 
microscope. On turning the microscope back into the median line, it 
is fixed in the optical axis of the apparatus by means of a suitable 
stop. The sUding-board is provided with a small grooved wheel 
receiving an endless cord, made of silk or fishing-line, which passes 
round the grooved, milled head of the fine adjustment of the 
microscope. When the slidmg-board is returned to the median 
line of the apparatus, the milled wheel connected with the fine 
adjustment impinges upon the wheel of the long focussing rod. 
The latter is provided with an india-rubber tire, which grips the 
teeth of the milled wheel, and thus the long focussing rod is placed 
in connection with the fine adjustment of the microscope. 

Illwmination. — The oxy-hydrogen lamp has been more frequently 
employed by the author than the paraiiine lamp, partly on account 
of the diminished time in exposure, especially when em.ploying very 



160 



BACTERIOLOGY. 




PHOTOGRAPHY OF BACTERIA. 161 

high powers ; this is of great importance where there is likely to be 
vibration from passing traffic. With rapid plates and the highest 
powers, the exposure has only been two or three seconds, whereas 
with the paraffine lamp it may vary from three to ten minutes, or 
even longer. 

Walmsley gives the following table for exposures with the 
paraffine lamp : — 



1| inch objective 3 to 45 seconds. 

TW ^' " ■ ■ i " ^ minutes. 



9 



I 



XT 



4 „ 10 



The illuminating apparatus represented in the accompanying 
engraving (Fig. 78) consists of a lantern which not only moves 
together with the microscope on the central sUding-board, but 
can be moved independently to or from the microscope, and be 
clamped with screws at the requisite distance for obtaining the best 
illumination. It is provided with two 3-inch condensing lenses of 
long focus, constructed of optical glass, which is much whiter than 
that used for ordinary lantern condensers. The lime-cylinders should 
be of the hardest and best quality, as they give a more actinic light 
than those made of soft lime. The " Excelsior " lime-cylinders are 
strongly recommended. They are . supplied in hermetically sealed 
tins which can be easily opened and re-sealed, so that a cylinder can 
be taken ovit and used, and the rest preserved for a future occasion. 
The hydrogen can be obtained by using the coal-gas supplied to the 
house, and the oxygen should be supplied preferably in a compressed 
state in iron bottles. Not only are the bottles much less cumbrous 
than the bags, but a small quantity of gas can be used, and the 
residue left for an indefinite time ; moreover, the gas is always 
at hand to be turned on when required. On the other hand, the 
retention of unused gas in bags is liable to cause their corrosion, 
owing, it is believed, to impurities which are carried over in the 
manufacture of the oxygen. If gas is not laid on in the house, then 
it also must be procured in a compressed state in bottles. As the 
blow-over jet is recommended on account of its safety, the bottles 
should be supplied in this case with a supplementary valve. It is 
then just as easy and free from danger to employ the compressed 
gas as it is to make use of the house-supply. 

I'he Camera. — A long-focus, half -plate camera is mounted upon 
a sliding platform. This admits of the camera being pushed up to 

11 



162 



BACTERIOLOGY. 



the microscope when it is in the long axis of the apparatus, so as to 
make a hglit-tight combination. The opening which is filled in an 
ordinary camera by the lens can be shut off by means of an internal 
shutter, which is opened and closed by turning a screw at the side 
of the camera. The dark-back is provided with plate-carriers, so 
that either half, quarter, or lantern-size plates can be emiiloyed. It 
wiU be found convenient to have two or more dark-backs, so that 
several plates may be exposed without rearranging the hght for 
each exposure. 

Much more elaborate and expensive micro-photographic cameras 
have been constructed by Zeiss, and also by Swift. The latter has 




Fiu. 81.— Photogkaph of an Isipkession PEEP.\KATIO^^ 



carried out a suggestion made by Pringle foi- a support at the 
ocular end (Fig. 80). 

The Bark-room. — In every bacteriological laboratory there should 
be a developing room provided with shelves, gas, water-tap, and sink, 
but these arrangements are not absokitely indispensable. All that 
is essential is a room impervious to light ; and a closet or cupboard, 
if it can be ventilated, will answer perfectly well, with a jug and 
basin instead of the tap and sink. The steam-steriliser employed 
in the preparation of nutrient media for cultivating bacteria, if not 
I'equired at the time for sucli purposes, may be filled to the brim 
with water, and will form an excellent cistern and tap, while a pail, 
or small sanitary bin, may be utilised as a sink. 

Various kinds of lamps are made for the dark-room, burning 



PHOTOGRAPHY OF BACTERIA. 163 

either candles, oil, or gas. In any case, the light must pass through 
two thicknesses of ruby glass. 

Dry Plates. — A small supply of any of the ordinary plates in 
the market may be procured for preliminary trials in acquiring a 
knowledge of the processes ; but to overcome the diflaculties of certain 
stained preparations, the isochromatic or orthochromatic plates should 
be used. The | plate will be found to be the most suitable size- 
There are numerous formulae for the requisite solutions for 
developing and fixing the negatives, and instructions are usually 
enclosed, in the boxes of dry plates, but it is best to abstain from 
trying a number of different formulse, as it, leads to a great expendi- 
ture of time. There is a temptation to do this, it being supposed 
that there is probably some great advantage in one formula over 
another. It is much better to get accustomed to the behaviour 
under different exposures of one, or perhaps two methods. 

In France the iron developer is much in vogue, and is recom- 
mended by Tailfer and Clayton for use with their isochromatic 
plates. It has the advantage of great simplicity in the mode of 
employment, and, therefore, is very suitable for a beginner. In 
England, on the other hand, the alkaline developer is very 
much used, as it gives more command over the plate, enabling the 
photographer more fully to compensate for incorrect exposure. 

It is very desirable before attempting to take photographs with 
the microscope to learn how to take photographs with an ordinary 
landscape camera, and to get thoroughly accustomed to the use of 
some good developer, so that mistakes may be corrected and the 
clearest and sharpest negatives obtained. 

Practical Manipulation. 

Arrangement of Apparatus. — For working with the parafiine 
lamp, the mode of procedure is, as regards the illumination, briefly 
as follows. The sub-stage condenser is dispensed with when a 
low power is employed, as well as the mirror, and the lamp is 
so placed that the image of the flat of the flame appears accurately 
in the centre of the field of the microscope. A bull's-eye condenser 
is then interposed, so that the image of the flame disappears, and 
the whole field is equally illuminated. With high powers the 
sub-stage achromatic condenser is necessary, and a more intense 
illumination is obtained by using the flame edgewise. In using a 
low power with the oxyhydrogen light, the lantern is withdrawn 
some little distance from the microscope, and the top combination 
of the achromatic condenser removed. 



1 64 BACTERIOLOGY. 

It is best to begin with the use of a low power, and a trial 
object, such as the blow-fly's tongue, spine of Echinus, or trachea 
of silkworm. 

In order to explain the management of the apparatus (as 
represented in Fig. 78) the steps in the arrangement of the 
apparatus and exposure of the plate will be described in detail 
for the employment of a high power and the oxyhydrogen light. 
The solutions being ready for use, it is proposed to take a photograph 
of tubercle bacilli in sputum, with a Jg- apochromatic oil-immersion 
objective. The first point to claim attention is the arrangement 
of the light. Having lighted the gas at the hydrogen jet, the 
lime-cylinder should be revolved until heated equally all round. 
The oxygen is then carefully turned on until only a small spot 
of incandescence is produced. The central sliding-board is turned 
out, a low power screwed on to the microscope, and the image of 
the bright spot focussed and accurately centered. To protect the 
sight, an eye-piece provided with a smoksd glass shade is used. 
The immersion objective is then substituted for the low power, 
and the oxygen turned on until the right admixture of gas is 
obtained to produce a brilliant illumination. It is well at this 
stage to sit down to focus the selected object, and to spend some 
httle time in searching for the most characteristic part of the 
specimen to be photographed. This being decided upon, the eye- 
piece is carefully withdrawn, and the central sHding-board rotated 
back into the median line. To make a light-tight connection 
between the camera and the microscope, the camera is pushed up 
until a velvet-lined tube, which occupies the position of the lenses 
of an ordinary camera, is enclosed within a short wide tube which 
is adapted to the eye-pieca end of the microscope. 

On opening the camera-shutter the image will be projected upon 
the ground glass screen of the camera. It is necessary, however, 
to obtain the exact focus, and to effect this the ground-glass 
screen is turned away, and the dark-back with a piece of plain 
glass is substituted. Here agam time may be well spent in 
getting the sharpest image, with the aid of a focussing glass of 
proper focal length. 

The greatest delicacy in manipulation is necessary, as in working 
with such high powers a turn too much of the fine adjustment will 
cause the image to vanish. Having determined the best visual 
focus, which will be found with the high-power objectives of most 
makers to correspond with the chemical focus, the dark-back must 
be cautiously removed, to prevent any vibration, and the plain 



PHOTOGRAPHY OF BACTERIA. 165 

glass replaced by a sensitive plate. To effect this change, the 
operator retires to the dark-room, and opens a box of plates with 
as little exposure to the red light as possible. Having removed 
a plate, it is necessary to ascertain which is the .sensitive side. 
This may be done by momentarily exposing it to the red light, 
and seeing which is the sensitive side by the dull appearance of 
the film. A less satisfactory way is to moisten the tip of a 
finger, and press it at one corner of the plate. The film side will 
be recognised by imparting a sticky sensation. The film must be 
dusted with a camel's-hair brush, as well as the dark-baick, and 
the plate is placed film-side downwards in the dark-back, which 
is then securely closed. 

Care should be taken that the plates then remaining in the box 
ai'e packed away before light is admitted to the dark-room. 

Exposure of the Plate. — On returning to the apparatus, the 
camera-shutter is closed. Then the dark-back is gently shd into 
its place, and its slide -vvithdrawn. A few moments are allowed 
to elapse, so that the least possible vibration, which might be 
caused by inserting the dark-back, has had time to cease, and all 
is ready for the exposure. 

In the case of the object we have selected, three seconds will 
probably be the exposure required. This is done by opening and 
closing the camera-shutter with one hand, while a watch can be 
held in the other. The slide of the dark-back is then carefully 
closed, and the plate is ready to be carried off to the developing 
room. 

As the hght will not be again required until the next exposure, 
the oxygen must be turned off, while the coal-gas may be allowed 
to play over the lime. 

Development and Fixation of the Image. — It is well to be 
systematic, and therefore, before the plate is taken out of the 
dark-back, light is admitted to the dark-room, and everything 
arranged so that the position of the trays and bottles may be 
remembered in the dark. First, let the ruby lamp be lit, place 
two dishes or trays close by, and a row of four dishes within easy 
reach. Pour out some fixing solution in the first porcelain dish, 
alum in No. 2, and water in Nos. 3 and 4. Put the necessary 
quantity of " pyro " solution into the glass measure, and place 
it with the ammonia drop-bottle in front of the ruby hght. 
Then, when all light except that from the ruby lantern has been 
excluded, everything is ready to commence the development of the 
plate. 



166 BACTERIOLOGY. 

Opening the dark-back, the plate may be turned out on to the 
palm of the hand. The film side is then uppermost, and the plate is 
to be transferred in the same position to a tray, and covered with 
water. This is to soak the film and obtain an equal action of 
the developer ; or the solution of fresh pyro may be poured on to 
the plate without previous soaking, if the flow is uniform, and the 
formation of bubbles avoided. In the first case the water is run o£E 
and the pyro allowed to flow evenly over the plate. To protect the 
plate from prolonged exposure to the ruby light, a second tray may 
be inverted over it, or the developing tray covered with a piece 
of card-board. Gently rock the tray for a minute or so, then 
to a few drops of ammonia in a measuring glass add the pyro 
from the developing tray, and pour the mixture back again 
over the plate. After again gently rocking the tray for a few 
minutes, more ammonia is added by drops in the same way. 
If the exposure has been properly timed, — and the time necessary 
must be ascertained by trial for each preparation, — the image will 
gradually begin to appear, and the action must be allowed to 
continue until sufiicient density has been obtained. To determine 
this requires some experience. It is generally recommended to take 
the plate out of the tray and hold it for a moment, film-side towards 
the operator, in front of the ruby light. Though the plate is not 
nearly so sensitive when the image has commenced to develop, and 
there is, therefore, not the same danger of fogging, a safer plan Ls 
to occasionally turn the plate film downwards in the tray, and when 
the image appears on the back the development vnll be found to be 
completed. 

With such a preparation as tubercle bacilli in sputum it is 
not easy to trace the gradual formation of the image, and hence the 
advantage of commencing with a well-marked object such as the 
blow-fly's tongue. It is then easy to watch the gradual progress of 
the image. The bright parts or high-lights appear first, then 
gradually the half-tones, or less brightly-lighted parts, and lastly 
every shade except the deepest shadows is represented. When, 
however, all action seems to have ceased, we nlust still wait until 
we have judged, in the manner already described, that the density 
is sufficient. This being determined, we pour off the developing 
solution and thoroughly wash the plate with water. It is then 
ready to be placed in dish No. 1, containing " hypo," and here it must 
be left for some minutes after all appearance of creaminess has 
disappeared from the back. White light may now be admitted, the 
plate removed from the hypo and thoroughly washed under the tap, 



PHOTOGRAPHY OF BACTERIA. 167 

and then placed in dish No. 2. When another plate is ready to 
take its place, transfer it to dish No. 3, and then to No. 4, and, 
after a good final washing under the tap, place it upon a rack 
to dry. If there is any tendency for the film to detach itself from 
the plate, " to frill," the alum bath must be used before fixing, as 
well as after. 

Frilling or blisteriiig may be due to an error in manufacture, 
and is liable to occur in hot weather, or when using a developer too 
strong in ammonia. If it occurs during washing or fixing, the alum 
bath must be employed before the hypo. Fogging, or the appear- 
ance of a veil over the plate, may arise from error in the manu- 
facture, from admission of extraneous light, from over-exposure, 
or from prolonged exposure to the ruby light during development. 
Care must be taken that the camera and dark-room are light-tight. 
GrystalUsation, or powdery deposit, upon the negative when dry, is 
due to insufficient washing out of the hyposulphite of soda. Thin- 
ness of the image, or want of density, may be due to insufiicient 
development, too weak a developer, or too short or too long an 
exposure. Too great density results from too long immersion in the 
developer. 

Spots may sometimes occur upon the negatives. They may be 
caused by dust upon the plate or by air bubbles in the developer. 

In the tesxt-books of photography full accounts of failures will be 
found, their causes and prevention ; but it will be advantageous 
when these difficulties are encountered to take the negatives to a 
skilled photographer and get advice upon them. It is necessary 
to persevere, and not be disheartened if several negatives have to 
he made of a preparation before a successful result is obtained. 

It may here be remarked that the beginner will far more 
rapidly learn the technique if he avail himself of a practical demon- 
stration from a photographer. When he has learnt to obtain suc- 
cessful negatives, if he prefer silver prints, and time is an object, it 
will be found to be true economy to get the printing and mounting 
done by a professional photographer. The credit of a successful 
photograph of bacteria is due to the bacteriologist who prepares 
the microscopical specimen and obtains the negative. 

Determination of the Amplification. — The amplification varies 
not only with the objective employed, but with the distance of 
the focussing screen from the object. In order to ascertain the 
amplification afforded by a certain objective at a certain distance, a 
photograph should be taken, iinder the same conditions, of the lines of 
a micrometer slide. It is easy then to calculate the amphfication 



If 



BACTERIOLOGY. 




obtained in the micro-photograph ; sujjposing, for example, in 
the micro- pliotograph the lines which are i^jjjj inch apart are 
delineated 1 inch apart, the magnifying power mnst be 1,000 
diameters. Moreover, ha\ing tlins ascertained the amplification, 
we can accurately compute from the photograph the size of the 
objects taken. 

Vahie of Photographs. — It is not necessary to compare the 
relative merits of diagrams and photographs. Diagrams which do 
not purport to be accurate representations, but are 
intentionallj' the means for simplifying instruction, 
will always be valuable, even if we have the 
original preparations under the microscope before 
us. We must consider the relative merits of 
photographs and of drawings which purport to be 
exact representations of what is seen under the 
microscope. Thus in the case of micro-organisms, 
when their biological characters are studied under 
low powers of the microscojae, photograjjhs are 
preferable because they give a more faithful re- 
presentation. At the same time, apart from this 
comparative value, we must not lose sight of the 
actual value of photograph}' in placing within the 
reach of the student or investigator, who may not 
be a draughtsman, a most valuable means for 
illusti'ating all kinds of preparations. 

For double-stained or triple-stained tissue pre- 
parations an accurately coloured drawing leaves 
little to be desired ; but if we reproduce the same 
by a wood engraving, and so lose the advantage 
of the coloured picture, which is instructive in 
indicating the method of staining, a photograph will, in many cases, 
be far more satisfactory. 

When we have to deal with the growth of bacteria en masse, 
as in test-tube and plate-cultivations, with colonies as seen under 
a low power of the microscope, and with impression-prepara- 
tions both under low and high powers, unless the bacteriologist 
is a most accompli-shed draughtsman as well as an accurate and 
reliable observer, photography undoubtedly affords the best mode 
of illustration. The apparatus being ready and at hand, a negative 
can be produced in a few minute's of a preparation which, from the 
amount of detail it contains, would take perhaps several hours to 
draw and colour. From that negative any number of facsimiles can 




I'iG. 82. — Photo- 
geaph of a 
Cultivation op 
Bacillus An- 
thkaois. 



PHOTOGRAPHY OF BACTERIA. 169 

be obtained, whereas an original dra-^ving, even in the best hands, 
if cut on wood or lithographed, is almost certain to fall short of 
being an exact copy. 

With regard to individual bacteria, the result is more satisfactory 
in many cases than a drawing ; for there is the advantage of being 
absolutely certain that any particular structure, form, or shape 
which may be represented is actually what exists, and not what 
may have been evolved by unconscious bias in the mind of the 
observer. Many illustrations might be given of this. Thus Lewis, 
who was a most conscientious observer, published an account of 
organisms in the blood of rats in India, and illustrated it with a 
wood engraving and with micro-photographs. The identity of the 
organisms which were found in the common brown rat of this 
country was established much more readily from these photographs 
than from the wood engraving or the description in the letterpress. 

A micro-organism, even under the highest powers, appears as 
so minute an object that to represent it in a drawing requires a 
very delicate touch, and it is only too easy to make a picture which 
gives an erroneous impression to those who have not seen the 
original. If, on the other hand, to represent the object more 
clearly we draw an enlarged picture, we can only do so by repre- 
senting what we think the object would be like if it could be 
amplified to the size represented. In such cases a photographic 
enlargement is certainly more valuable. 

Photography enables us also to record rapid changes, and it 
is possible that as the art advances we may find that the film is 
more sensitive than the human retina, and brings out details in 
bacteria which would be otherwise unseen. 

Photographs can be readily transmitted by post, and when we 
can neither make a great number of preparations to illustrate 
some object, nor perhaps be able to go to the expense of having a 
drawing reproduced, this method will be of value in enabling others 
to benefit by our observations. 

The author is convinced that if the employment of photography 
is encouraged in bacteriological and other research laboratories for 
depicting microscopical preparations and cultivations of bacteria, the 
results of increasing experience and practice will lead to its being 
made more general use of as a faithful and graphic method, valuable 
alike for class demonstrations and for illustrating publications. 



PAET II. 

ETIOLOGY AND PREVENTION OF INFECTIVE 
DISEASES. 



171 



CHAPTER XIII. 

suppuration, pyemia, septiggmia, erysipelas. 

Abscess. 

When inflammation is followed by an accumulation of leucocytes 
and of plasma which does not coagulate, the result is a white or 
creamy liquid called pus, and when the surrounding tissues are 
involved so that a cavity develops containing pus, we have what 
is termed an abscess. The almost constant association of bacteria 
with the production of pus has created a behef that they are 
the direct cause of suppuration. Ogston found micrococci present 
in all acute abscesses, and concluded that acute inflammation was 
invariably due to their presence. The fact that inflammation 
occurs more frequently in the external tissues of the body is 
accordingly explained by the ready entrance of bacteria which 
are in the air ; and suppuration following pericarditis, pleurisy, 
and other conditions in which air is excluded is attributed to the 
presence of pyogenic cocci, which have gained access by the blood 
stream. There is no pyogenic organism constantly present, but 
several different species of bacteria have been isolated from pus 
and carefully studied, and the antiseptic treatment is based upon the 
principle of excluding bacteria in surgical operations, and destroying 
any which may have previously obtained access to wounds and 
broken surfaces. Inflamed tissue and pus form a most suitable 
medium for the growth of bacteria, which in some cases are 
unquestionably only accidental epiphytes. 

In tuberculosis, actinomycosis, and glanders, pus formation may 
take place without the presence of pyogenic cocci ; and it is generally 
believed that chemical irritants, such as croton oil, turpentine, iodine, 
cadaverin, and tuberculin, will excite the formation of pus in the 
absence of bacteria, although Klemperer, after a number of very 
careful experiments, maintains that no genuine pus will be produced 
if the chemical irritants are first carefully sterilised. 

173 



174 



INFECTIVE DISEASES. 



The bacteria which have been isolated from pus include : — 
Staphylococcus pyogenes aureus, albus, and citreus, Staphylo- 
coccus cereus flavus and albus, Streptococcus pyogenes, Micrococcus 
pyogenes tenuis. Micrococcus pneumoniae crouposse, Bacillus pyo- 
cyaneus, Bacillus pyogenes f oetidus. Micrococcus tetragenus, Bacillus 
intracellularis meningitidis, Gonococcus, Bacillus septicus vesicae, 
Urobacillus liquefaciens septicus. Bacillus typhosus. Bacillus coh 
communis. Bacillus anthracis. Bacillus tuberculosis, Bacillus mallei, 
and Actinomyces. 




Jig. 83. — Suppubation of Subcutaneous Tissue. 

d, Leucocyte containing micrococci ; d', leucocyte with pale nucleus showing 
necrosis ; c, fixed connective tissue cells, much enlarged, containing several 
nuclei, of which some (n') are pale and necrotic ; numerous cocci, diplocoooi, 
and short chains. (Cobnil and Kanvier. ) 

Some idea of the distribution of the bacteria most commonly 
occurring in pus may be gathered from the records made by Passet 
and by Karlinski. 

Passet examined acute abscesses, and found Staphylococcus 
pyogenes aureus and albus in 11 cases. Staphylococcus pyogenes 
albus alone in 4, Staphylococcus pyogenes albus and citreus in 2, 
Streptococcus pyogenes alone in 8, Staphylococcus pyogenes albus 
and Streptococcus pyogenes in 1, and Staphylococcus pyogenes albus 
and citreus, and Streptococcus pyogenes in 1. i 



SUPPURATION, PYjEMIA, SEPTICEMIA, ERYSIPELAS. 

Karlinski tabulated his cases thus : — 



175 



Disease. 


1 


1 


si 

gb 


1^ 




is 
II 




^ 

a'S 


J 
1 






OljSj 


•"£■ 


="£■ 






I& 


(5^ 


» 


Mastitis .... 


3fi 


'/.l 


4 


4 


6 




_ 


_ 





Subcutaneous Abscess 


30 


10 


2 


8 


6 


2 


2 





— 


Phlegmon . 


24 









24 





— 





— 


Furuncle 


20 


9 





10 





1 


— 





— 


Bubo .... 


17 


8 


1 


1 


7 





— 





— 


Subperiosteal Abscess 


16 


6 





10 


— 




— 





— 


Panaritium Cutaneum 


Ifi 


7 





9 








— 





— 


Panaritium Osseum 


10 


7 





3 








— 





— 


Dental Abscess . 


10 


1 


— 4 


1 


3 


1 





— 


Hordeolum 


10 


fi 


— 4 













— 


Abscess of the Middle Bar 


4 


2 













2 


— 


Carbuncle . 


4 


2 


— 1 


1 





— 


— 


4 


Osteomyelitis 


3 1 2 


— 1 


— 


— 


— 




Total . 


200 


82 


7 


m 


45 


6 


3 


2 


4 



Pyemia and SsPTicaiMiA. 

When pyogenic micrococci get access to the blood stream they 
may be carried into distant parts, and by multiplying produce meta- 
static abscesses in the lymphatic glands, bones, joints, and internal 
organs, a condition which is recognised as pysemia. 

If there is a gelieral invasion of the blood stream by micrococci, 
and absorption of their poisonous products, septicaemia results, and 
death may occur before the development of any secondary lesions. 
When septic micro-organisms multiply locally, and their chemical 
products are absorbed, or their products are separated from putrid 
material and injected into the circulation, the result may be called 
sa/proemia. The blood in septicaemia contains living organisms, and is 
infective. The blood in saprsemia contains only the toxic chemical 
products, and is not infective. The one is septic infection and the 
other septic intoxication. Pysemia may follow accidental wounds, 
surgical operations, parturition, acute suppuration of bones, scarlet 
fever, typhoid fever, and other diseases. 

To avoid pysemia in surgery and midwifery, the greatest care 
must be taken to prevent micro-organisms from being conveyed by 
instruments, sponges, bandages, and by the hands of the surgeon 
or the obstetric physician. By the use of antiseptics and absohite 
cleanliness the chances df infection are reduced to a minimum. 



176 INFECTIVE DISEASES. 

Eosenbach examined six cases of metastatic pyaemia : Strepto- 
coccus pyogenes was found five times, partly in tlie blood and partly 
in the metastatic deposits, and twice in company with Staphylococcus 
pyogenes aureus. \ 

Baumgarten, also, found Streptococcus pyogenes in the internal 
organs in pysemic cases, and Eiselsberg found Streptococcus pyogenes 
in company with Staphylococcus pyogenes aiireus in the blood of 
cases of septicaemia. 

Frankel isolated a streptococcus from puerperal fever, which 
he at first called Streptococcus puerperalis, but subsequently 
identified with Streptococcus pyogenes. These researches have 
been confirmed by others. Winkel obtained a pure cultivation of 
a streptococcus from the blood of the heart in a case of puerperal 
peritonitis. It produced erysipelatous redness when inoculated 
in the rabbit's ear, and in form and in cultivation was similar to 
the streptococcus in erysipelas. Gushing also found Streptococcus 
pyogenes associated with puerperal infection. The cocci were 
^ found in endometritis diphtheiitica as well as in secondary puerperal 
inflammation. These observations were still fvirther confirmed by 
Baumgarten, and Bumm isolated the same organism in puerperal 
mastitis. 



Desceiption of Bacteria in Pus. 

A description may now be given of the cocci most frequently 
found. Staphylococcus pyogenes aureus and albus and Strepto- 
' coccus pyogenes and Gonococcus are the most important of these. 
Staphylococcus pyogenes citreus, cereus albus and flavus, are pro- 
bably merely epiphytic. Micrococcus tetragenus. Micrococcus 
pyogenes tenuis. Bacillus pyogenes foetidus. Bacillus pyocyaneus, 
Bacillus coli communis. Bacillus septicus vesicae, Urobacillus lique- 
faciens septicus, and Bacillus intracellularis meningitidis will be 
described fully in Part III. The description of Actinomyces, of 
Micrococcus pneumoniae crouposae and of the bacilli of anthrax, 
tuberculosis, glanders, and typhoid fever, will be found in other 
chapters in Part JI. 

Staphylococcus pyogenes aureus (Rosenbach). — Yellow 
coccus in pus (Ogston). Cocci singly, in pairs, very short chains, and 
irregular masses. Cultivated on nutrient agar-agar, an orange- 
yellow culture develops, looking like a streak made with oil paint. 
One variety grows on nutrient gelatine without liquefying it ; 
another produces rapid liquefaction, and the growth subsides as 



SUPPURATION, PYEMIA, SEPTICEMIA, ERYSIPELAS. 



177 






iui orange-yellow sediment. On potutoes and blood serum a similar 
orange-yellow culture grows luxuriantly. They may also form 
colourless growths in sub-cultures, and are then indistinguishable 
from Staphylococcus pyogenes albns. The cocci do not cause any 
septic odour in pus, nor does any gas develop. Albumin is con- 
verted b}- their action into peptones. 
They produce rapid ammoniacal fer- 
mentation in urine (Shattock). 

The micro-organisms injected into 
the pleura or knee of a rabbit produce, airait/ \.^ 
as a rule, a fatal result on the following " V"^ 

da}' ; but if it survives longer, it eventu- 
ally dies of severe phlegmon. If injected 
into the knee of a dog, suppuration 
occurs, followed by diusintegration of the 
joint. Injected into the peritoneal Fig, 84.— Pus with Staphylo- 
cavit}' of animals, they set up peiito- cocci, x 800 (Flijgge). 

nitis, and introduced into the jugular 

vein they produce septicaemia and death. When a small quantity of 
a cultivation is introduced into the jugular vein after previous fracture 
or contusion of the bones of the leg, the animal dies in about ten days, 
and abscesses aie found in and around the bones, and in some cases in 

the lungs and kidneys, 
'-, ^ -, - -)'^^/ '^'i^L^fiii^ — 



( 



'/^^ 



•*) 




-i* 



n 



]\- 



J. 



/ 



and the cocc-i are found 
in the blood and pus. 

Garre caused sup- 
puration by inoculating 
a pure-culture in a 
wound near his finger 
nail. Bockhart suffered 
from several pustules 
after vaccinating his 
arm with a pure-culture 
suspended in salt solu- 
tion, and Bumm gave 
himself a hypodermic 
injection of a pure- 
culture and produced 
an abscess. This micro- 
organism is practically ubiquitous. It has been cultivated fi'om 
the skin and mucous membranes and secretions of healthy persons, 
and it occurs in the air, in soil, in dust, and in water, and in 

12 



Fig. 85. — Subcutaneous Tissue of a Rabbit 48 

HOUHS AFTER AN In.IECTIOX OF STAPHYLOCOCCI, X 

050 (Bau5igakten)_. 



178 INFECTIVE DISEASES. 

association with suppuration, pyaemia, puerperal fever, and acute 
osteomyelitis. 

Staphylococcus pyogenes albus (Rosenbach). — Cocci micro- 
scopically indistinguishable from the above. In cultivations also 
they resemble Staphylococcus pyogenes aureus, but the growth 
consists of opaque white masses. They, as a rule, liquefy nutrient 
gelatine very rapidly, and subside to the bottom as a white sediment ; 
more rarely they liquefy very slowly ; and a variety has also been 
described which does not produce any liquefaction. They are similar 
to the above-mentioned in their pathogenic action. Pure-cultivations 
of the organism were obtained from a case of acute suppui-ation 
of the knefe-joint. 

Staphylococcus pyogenes citreus (Passet). — Cocci singly, in 
pairs, very short chains, and irregular masses. If cultivated on 
nutrient gelatine or nutrient agar-agar, a sulphur or lemon-yellow 
growth develops. When inoculated under the skin of mice, guinea- 
pigs, or rabbits, an abscess forms after a few days, from which a 
fresh cultivation of the micro-organism can be obtained. 

Staphylococcus cereus albus (Passet).. — Cocci, morphologic- 
ally similar to the above, but distinguished by forming on nutrient 
gelatine a white, slightly shining layer, like drops of stearine or wax, 
with somewhat thickened, irregular edges. In the depth of gelatine 
they form a greyish-white, granular thread. In plate-cultiva- 
tions, on the first day, white points are observed, which spread 
themselves out on the surface to spots of 1 to 2 mm. When culti- 
vated on blood serum a greyish-white, slightly shining streak 
develops, and on potatoes the cocci form a layer which is similarly 
coloured. 

Staphylococcus cereus flavus (Passet).— Cocci which produce 
in nutrient jelly a growth which, at first white, becomes lemon- 
yellow, somewhat darker in colour than Staphylococcus pyogenes 
citreus. Microscopically Staphylococcus cereus flavus corresponds 
with Staphylococcus cereus albus. Inoculation experiments with 
both kinds give negative results. 

Streptococcus pyogenes (Rosenbach). — Cocci occurring singly, 
in masses, and in chains. The individual cocci are small spherical 
cells, with a special tendency after fission for the resulting elements 
to remain attached to each other, forming chains or rosaries. In 
cultures on solid media they often occur in the form of staphylococci," 
but in liquid cultures there may be a few, three or more elements, 
linked together ; or a great number, forming long chains which may 
be straight, serpentine, or twisted. . ' 



DESCRIPTION OF PLATE IV. 
Streptococcus Pyogenes. 

Fig. 1. — From a cover-glass preparation of pus from a pyaemic abscess. 
Stained with gentian-violet by the method of Gram, and contrast-stained 
with eosin. x 1200. Powell and Lealand's apochromatic ^\ Hom. imm. 
E. P. 10. 

Fig. 2. — From cover-glass preparations of artificial cultivations of the strepto- 
coccus in broth and in milk at different stages of growth, x 1200. Powell 
and Lealand's apochromatio -f-^ Horn. imm. E. P. 10. 

In these preparations there is a great diversity in size and form of the 
chains and their component elements. In the drawing examples are 
figured of the following: 
(o) Branched chains. 

(b) Simple chains composed of elements much smaller than the 
average size. 

(c) Chains with spherical and spindle-shaped elements at irregular 
intervals. These are conspicuous by their size, and are sometimes 
terminal. 

((Z c) Chains in which the elements are more or less uniform in size. 
(/) Complex chains with elements dividing both longitudinally and 

transversely, and varying considerably in size in different lengths 

of the same chain. 



Plate IV 



\ '•'St*?'! 



'••»,! -. 



Fig 1. 

.,.•. 



'••' „' V 



% •* ""..• 


•■• 


^ 8 




^ ••" 








./ . ,.• ".. 


/ 


, e 

IllfO „«• 


..-•• 


"X. 





"v 

,:■' 






d 






x-"!!^'-.., '4.i"f- 



.* - •.«. .1... -f ■•> 

'• A*i f"."'Via.>".f.Xf 









'•.. 



ng2 
STREPTOCOCCUS PYOGENES 

E.M.CnijkeTuaiU.fea.b.. nrtcenb 'Brooha.Da^ 4 Son, lOk . 



SUPPURATION, PYEMIA, SEPTICEMIA, ERYSIPELAS. 179 

The individual elements composing the chains will be found to 
vary considerably in size : here and there in a preparation will 
be found a chain composed of excessively small cocci, in another 
part the elements will be all on a larger scale, and again in 
another pai-t they will be peculiarly conspicuous on account of their 
size. So great is the diversity in the size of the cocci in some of the 
chains, that one might imagine that there was more than one kind 
of streptococcus present in a preparation, until on examining some 
of the longest chains it is observed that various sizes are repre- 
sented in different lengths of the same chain. Very characteristic 
appearances result from the fact that the cocci enlarge and divide 
both longitudinally and transversely ; and, indeed, the largest, for 
the most part, clearly show a division in two directions, resulting 
in the formation of tetrads. In addition to the forms resulting 
from the fission of the cocci, there are here and there in a chain, 
and sometimes termina,Jly, lai-ger elements, which are spherical, 
spindle-shaped, or in the form of a lemon. In the length of the 
chains, as in the size of the individual cocci, there is usually great 
diversity, In some cases they are composed of only a few, three 
or four cocci; in others eight, ten, or twenty. Here and there 
an exquisite rosary vdll extend in a straight line across the field 
of the microscope, or be twisted, curved, or serpentine ; in some 
preparations twisted or entangled strands are observed which are 
.composed of several hundred elements. Such chains will be found 
to be much thicker in one part than another. Another char- 
acteristic appearance is produced by separation of the elements 
resulting from fission in the long direction of the chain, by which 
lateral twigs or branches are formed. Another character, which 
is very striking, may be seen when the individuals in a chain 
have become separated ; an unstained or faintly stained membrane 
may be found bridging across the interval. This will become still 
more visible in preparations contrast-stained with eosin. 

In plate-cultivations the appearances of the colonies ai-e not very 
striking. They appear to the naked eye after three or four days 
as extremely minute, greyish-white, translucent dots, which under 
the microscope have a slightly yellowish-brown colour. They are 
finely granular and well defined. They do not liquefy the gelatine, 
and after several weeks may not exceed the size of a pin's head. 

If the surface of nutrient gelatine solidified obliquely be traced 
over once or- twice with a platinum needle bent at the extremity 
into a little hook charged with the cocci, a ribbon-shaped film 
develops in two or three days. This film is composed of minute, 



180 INFECTIVE DISEASES. 

greyish-white, translucent dots or droplets, which can be more easily 
recognised with the aid of a pocket lens (Fig. 87). According to 
the number of organisms sown on the jelly, the dots or colonies 
may be completely isolated, or form a more or less continuous 
film. The film by reflected light has an iridescent appearance like 
mother-of-pearl, but has a bluish or bluish-grey tint by transmitted 
light, and with a pocket lens appears distinctly brownish. The 
gelatine is not liquefied, and even after several weeks the cultivation 
is limited to the inoculated area, and the individual colonies are, as 
a rule, not larger than pins' heads. In gelatine-cultivations of the 
same age, but kept in the incubator at 18° C, the colonies get 
ii^regular in form, especially at the margin of the film, and give the 
growth an ai'borescent, fringed, or serrated appearance. Cultivated 
on the oblique surface of nutrient agar-agar at 37° 0. the growth is 
very similar, forming a film composed of minute white colonies 
hke grains of sand ; but the film appears less transparent, is 
• whiter, and the colonies have a greater tendency to geit irregular 
in form. If inoculated with one tracing of the needle the growth 
is scanty, but tends to get thicker in the centre than towards the 
margins, which may have a terraced appearance. Inoculated in 
the depth of gelatine, there appears after a day or two at 18° 0. a 
thread-hke growth along the track of the inoculating needle. This 
delicate growth is found on examination with a pocket lens to consist 
of a linear series of extremely minute granules. In a few days' 
more, the beads or granules become more marked, but even after 
weeks, the cultivation only appears like a string of minute, white, 
compact, globular masses or grains. In broth at 37° C. the cocci in 
twenty-four hours create a turbidity, and gradually develop beauti- 
ful chains varying in length according to the age of the cultivations. 
Even in forty-eight hours there may be chains of eight, ten, twenty, 
or a hundred elements. After a few days the growth settles down 
at the bottom of the tube in the form of a white deposit, while the 
supernatant liquid becomes again clear. 

Inoculated subcutaneously in the ear of rabbits, they produce in 
two days an infiammatory thickening with erysipelatous redness, 
or sometimes suppuration. 

They may occur in vaccine lymph, as the result of con- 
tamination, and Pfeiffer suggested that before calf lymph is 
employed for vaccination it should be tested on a rabbit's ear. 
If in two days no rash has been produced, the possibility of the 
presence in the lymph of Streptococcus pyogenes or erysipelatis 
is excluded. 



SUPPUKATION, PYEMIA, SEPTICAEMIA, ERYSIPELAS. 181 

According to Fliigge and others, after subcutaneous inoculation 
of mice with a small quantity of a cultivation, there is no result in 
80 per cent, of the animals experimented upon. Sometimes there 
is limited pus formation at the seat of inoculation, sometimes the 
animals die without any very strildng pathological appearances. 

They occur in abscesses, pysemia, and septicaemia, and are often 
found in diseases such as scarlet fever and typhoid, associated with 
septic complications. They have been isolated from air, soil, and 
water. 

The streptococcus found in erysipelas agrees in description, and 
is merely a variety of Streptococcus pyogenes. It has been definitely 
estabUshed by the researches of Frankel and Freudenberg, and later 
by those of the author, Easkin, Prudden, and Bayard Holmes, 
that Streptococcus pyogenes is frequently found in scarlet fever and 
diphtheria, and in other diseases associated with septic complica- 
tions. The author has isolated Streptococcus pyogenes from acute 
abscesses, from suppuration after surgical operations, from pyasmia, 
from pyaemia after scarlet fever, and from purulent peritonitis. 
Some of these cultures have been kept up for very long periods, 
extending over some years, so that opportunities occurred for a 
complete investigation into the life history of this micro-organism. 
Variations in the appearances of cultures have been observed when 
obtained from the same source. A number of cultures from pus 
were prepared on gelatine and agar, made according to the usual 
formula, but at different dates, and, therefore, varying slightly iti 
composition and quality. Sub-cultures were also started in nutrient 
gelatine of precisely the same composition, but from primary cul- 
tures of the same micro-organism in different media — agar-agar, 
milk, and broth. The descriptions of the streptococcus hitherto 
published were then found to be inadequate. The different cultures 
and sub-cultures presented striking variations in the microscopical and 
macroscopical appearances. Some sub-cultures on gelatine, for exam- 
ple, exhibited a finely dotted appearance, others showed every variety 
in the size, and degree of opacity of the colonies (Fig. 89). Cultures 
in broth also, varied in appearance, owing to sKght variation in the 
composition of the medium, to slight differences of temperature, and 
other conditions difficult to determine. The addition of glycerine to 
broth materially alters the appearance of the culture. It was con- 
clusively proved that minute differences arise from different conditions 
of the cultivating media. The author was led to study exhaustively 
the streptococcus of acute suppuration in bovines. Primary cultures 
of Streptococcus pyogenes from man, and primary cultures from 



182 INFECTIVE DISEASES. 

a case of purulent peritonitis in a cow, were carried through 
sub-cultnres under exactly similar conditions. -Cultivations of the 
Streptococcus pyogenes bovis exhibited variations in microscopical 
and cultural characters which were even more marker! than in the 
case of the Streptococcus pyogenes hominis. By selecting certaia 
cultures from both sources there was a striking similarity if not 
identity between them, but, when compared under exactly identical 
conditions, there was more difference in cultural characters between 
the Streptococcus pyogenes bovis and the Streptococcus pyogenes 
hominis than between the Streptococcus pyogenes hominis and the 
Streptococcus erysipelatis, and they may therefore be regarded as 
distinct varieties (Kgs. 89, 90). 

Some of the diseases and conditions in which Streptococcus 
pyogenes has been found may be alluded to more in detail. 

SpreadiiKj Gangrene. — From a case of spreading gangrene, which was 
identical with Ogston's erysipelatoid wound gangrene, and regarded by 
him as the most intense and dangerous form of erysipelas, Eosenbach 
obtained pure-cultivations of a streptococcus by incising the skin of the 
limb, and inoculating tubes from the turbid reddish fluid which escaped. 
That the streptococcus was identical with Streptococcus pyogenes was 
ascertained by comparison with a cultivation derived from pus, of the 
mode of growth, and of the effect on animals. 

Surgiml Fever. — Eiselsberg proved the presence of a streptococcus 
in the blood of several cases of surgical fever in Billroth's clinic. The 
organism was identified by cultivation with Streptococcus pyogenes. 

Diphtheria.— la three cases of typical diphtheria, Loffler found a 
streptococcus. He isolated it by cultivation, found that it was similar 
in form, characters on cultivation, and effects after inoculation, to 
Fehleisen's streptococcus of erysipelas. Loffler was not inclined to 
regard them as identical, because Fehleisen never found his cocci in the 
blood-vessels. Fliigge named the organism Streptococcus articulorum, 
and states, that after subcutaneous inoculation or injection of a cultiva- 
tion in mice, a large proportion of the animals die, and in the sections 
of the spleen and other organs the streptococci are again seen. Baum- 
garten investigated the same subject, and decided that the streptococcus 
was identical with Streptococcus pyogenes. 

Small-pox.—'H.la.Ya. has established the presence of Streptococcus 
pyogenes in the pustules of variola, and Garre found streptococci in the 
internal organs in a case of variola hsemorrhagica. In a fatal case of 
variola complicated with pemphigus, Garr(5 found a streptococcus in 
the pemphigus vesicles. Whether it was identical with Streptococcus 
erysipelatis Garre left an open question. 

Yellow Fern-.—Bahhs observed the presence of streptococci in the 
vessels of the kidney and liver in yellow fever. Cultivation experiments 
are wanting. It was probably a case of secondary infection with Strepto- 
coccus pyogenes. 



SUPPURATION, PYAEMIA, SEPTICAEMIA, ERYSIPELAS. 



183 



Bilious Fever.— Bahhs, in a case of fifevre bilieuse typhoide, found 
masses of streptococci filling the vessels of the liver, kidney, and spleen. 
This was probably another instance of secondary infection with Strepto- 
coccus pyogenes. 

Measles.— From, the blood and inflammatory post-products in measles, 
Babes isolated a streptococcus, which he describes as closely resembling 
the Streptococcus pyogenes. 

Ulcerative Endocarditis. — Wyssokowitsch found cocci in the internal 
organs in ulcerative endocarditis, and produced the disease in animals, 
after injury to the valves, by injection of Streptococcus pyogenes and 
other organisms. Weiohselbaum, by microscopical research and by 
cultivation experiments, proved the presence of Streptococcus- pyogenes 
in acute verrucous endocarditis. Ban mgarteu confirmed this. He found 




SSfiiii^ttMA' s^ 




Tig. 86. — Ulcerative Endocakditis : Section of Cakdiac Muscle, ■" 700 (KocH). 



Streptococcus pyogenes alone in one case and accompanied by Staphylo- 
coccus aureus in another. 

Bronclw-pneumonia. — Thaon found a streptococcus in the lungs of 
children in fatal cases of broncho-pneumonia, complicating measles, 
diphtheria, and whooping cough. It was regarded as identical with the 
streptococcus isolated by Loffler from diphtheria. Frankel discovered 
a streptococcus in the lungs of a case of true croup complicated with 
broncho-pneumonia, and by cultivation established its identity with 
Streptococcus pyogenes. 

Anthrax. — Oharrin found cocci in rabbits, examined some hours after 
death from anthrax. These, when isolated, produced death in rabbits 
from septicaemia, without suppuration. Chains composed of from fifteen 
to twenty elements were found in all the organs. This was probably 
another instance of Streptococcus pyogenes. 

Syphilis. — Kassowitz and Hoohsinger found the presence of a strepto- 



184 



INFECTIVE DISEASES. 



coccus iu the tissues and internal organs, and especially in the blood- 
vessels, in fatal cases of congenital syphilis. These observers regarded 
their discovery as having an important bearing on the etiology of syphilis, 
but Kolisko pointed out that it was only the result of septic infection 
with presence of Streptococcus pyogenes, as had already been established 
in scarlet fever. 

Cerebru-.yi'nal Jfriunf/iti-^.— From the meningeal exudation of a case 
of apparently idiopathic cerebro-meningitis, Banti found Streptococcus 
pyogenes and Staphylococcus aureus and albus. The cocci probably 
entered through an abscess of the jejunum. 




Tig. 87. — Puke-Cultures of Streptococcus Pvogenes. 

a, On the .surface of nutrient gelatine ; h, in the depth of nutrient gelatine ; 
c, on the surface of nutrient agar. 



Blephai-(idi>nitin <inil Ducri/nryst/s. — Widmark isolated by cultivations 
Streptococcus pyogenes and other organisms from cases of blepharadenitis 
and phlegmonous dacryocystis. In phlegmonous dacryocystis Widmark 
found Streptococcus pyogenes almost exclusively. 

Leulxeiiiia, — FHigge cultivated a streptococcus from necrotic patches 
in the spleen of a fatal case of leukaemia. Cultures corresponded very 
closely with Streptococcus pyogenes. Inoculation in the ears of rabbits 
produced similar results to Streptococcus pyogenes or erysipelatis. 
Fliigge calls it Streptococcus pyogenes malignus, but concludes that it 
is probably dentical with the streptococcus from pus. 



SUPPURATION, PYjEMIA, SEPTICEMIA, ERYSIPELAS. 



185 



Erysipelas. 

Erysipelas is an acute inflammation of the skin, occurring in 
connection with wounds, when it is traumatic, and on surfaces 
apparently sound, when it is idiopathic, as in erysipelas of the 
face. It is highly contagious in surgical wards, and it gives rise 
to rapidly fatal puerperal fever in lying-in hospitals. In such cases 
the virus is obviously conveyed from sick to healthy persons by 
direct contact, or by instruments and sponges, or by the hand 
of the siirgeon, physician, or nurse, and possibly by the air. 




fet:n ^^K^g0^^-^ 










Fie. 88. — Section of Skin in Eeysipelas. 

v,v, two lymphatic vessels containing leucocytes and m,m, streptococci ; t, connective 
tissue ; a, connective tissue and wandering cells, x 600 (Cornil and !R.\nvier). 



Streptococcus of Erysipelas. — In 1882, Fehleisen isolated a 
streptococcus in erysipelas, described the appearances on cultivation, 
and maintained that it could be distinguished from the streptococcus 
of suppuration. Rosenbach agreed that the two micro-organisms 
could be distinguished by parallel experiments, and named the one 
Streptococcus pyogenes and the other Streptococcus erysipelatis. 
(Fehleisen). Rosenbach asserted that the colonies of tlie latter 
were more opaque and whiter than those of Streptococcus pyogenes 
and the growth more marked in the depth of nutrient gelatine, 
while microscopically the chains were better marked and larger, and 
the individual cocci larger than in Streptococcus pyogenes. Others 
who investigated this subject could not distinguish them with 
certainty, either by their morphological or cultural characters or 
effects on inoculation. Passet found that inoculation of Strepto- 
coccus pyogenes induced a condition very similar to that produced by 
inoculation of Streptococcus erysipelatis. Hoffa and Hajek described 
minute differences, but Biondi and Eiselsbei'g failed to confirm these. 



186 INFECTIVE DISEASES. 

Baumgarten failed to prove any essential difference. Mitchell 
Prudden found that Streptococcus pyogenes injected into the sub- 
cutaneous tissue of the ears of rabbits, produced in one no effect ; 
in four, slight transient redness ; in five, local redness followed by 
abscess; in twelve, well-marked erysipelatous redness, followed by 
complete resolution in seven, abscess in three, and death in two. 
Passet, Biondi, Eiselsberg, Baumgarten, and Mitchell Prudden 
concluded that, in their morphological, biological, and pathogenic 
characters, so far as animals are concerned, the two organisms are 
practically identical. 

The author investigated the morphology and cultural characters 
of the Streptococcus erysipelatis, which he had isolated from a 
typical case. This result cleared up the conflicting statements 
which had been made by different observers. By carrying out 
absolutely parallel experiments, the. Streptococcus pyogenes and 
Streptococcus erysipelatis were unquestionably distinguishable, as 
Fehleisen and Rosenbach had asserted. In both cases, however, 
inoculation of a trace of a culture from a solid medium produced 
only transient redness. Injection hypodermically of a broth-cultm-e 
produced in both cases a .spreading erysipelatous redness, followed 
by suppiu'ation. It was found that primary cultures of ~the two 
micro-organisms, cultivated under precisely the same conditions, 
differed in the size and character of their chains, in the size of the 
individual elements, in the greater opacity of the colonies of Strepto- 
coccus erysipelatis, in a greater tendency to confluence, and in 
a more rapid growth. The author found that the difference was 
most marked in broth-cultures. Abundant flocculi were formed by 
Streptococcus pyogenes ; a powdery deposit with special tendency to 
form a granular adhesive film at the bottom of the culture flask, 
in the case of the streptococcus of erysipelas. Lastly, they differed 
in their power of resisting germicides. 

Fehleisen inoculated patients in hospital suffering with malignant 
growths, and produced a typical erysipelas with sub-cultures after 
an incubation of from sixteen to twenty hours. The disease was 
marked by rigors, fever, and general disturbance. Patients who had 
recently suffered from erysipelas had an immunity. 

Emmerich succeeded in proving the presence of streptococci in 
the air of a hospital where erysipelas had broken out. These cocci 
in their form, their characters on cultivation, and their inoculation 
results, were identified with the Streptococcus erysipelatis. It is not 
therefore exclusively parasitic. 

Streptococci identical or agreeing very closely in their description 



SUPPURATION, PYEMIA, SEPTICEMIA, ERYSIPELAS. 



li. 



with Streptococcus pyogenes, ha\-e been found in cattle plague, foot 
and mouth disease, strangles, contagious mammitis in cows, and 
progressive tissue necrosis in mice, and thej' will be i-eferred to fully 
in subsequent chapters. 

EXAMIXATIOX AND C'ULTIA'ATIOX OF STREPTOCOCCI, 

Cover-glass pi'eparations can be stained witli the watery solutions 
of the aniline dyes. In some cases veiy beaiitiful preparations can 
he obtained by using Neelseii's solution, and I'emoving excess of 







Fig. 89.— Streptococcus Pyogenes Hominis. Pure-cultures on nutrient 

gelatine. 



«, Sub-culture from agar, 
c, Sub-culture from milk. 



b. Sub-culture from broth. 
d. Sub-culture from milk. 



stain by rinsing in alcohol. To examine pus, milk, or broth, take 
an ordinary platinum needle bent at the extremity into a booklet. 
Dip it into the liquid to be examined, and spread it on a cover- 
glass into as thin a film as possilile ; the preparation is treated 
in the ordinary way, that is to say, the film is allowed to dry, 
and the cover is taken up with forceps, and passed three times 
through the flame with its prepared side uppermost. 

Gram's Method toith Eosin.^ln this way the streptococci are 
stained blue, and stand out in marked contrast to the rest of the 
preparation. Use freshly prepai'ed solution. Float the cover-glasses 



INFECTIVE DISEASES. 



on the solution for ten minutes to h;ilf an hour, then tiULsfer them 
to ioaine-potassic-iodide solution, until they assume the colour of a 
tea leaf; then immerse them in alcohol until they are decolorised ; 
dip them in an alcoholic solution of eosin for a few moments, and 
then transfer them to clove oil to clarify the film ; to remove the 
clove oil gently press the cover between tw,j layers of clean filter 
paper, then mount in xylol Ijalsain. 

A good method for cultivating streptococci is to employ a steril- 
ised looped platinum wive, and to spread a droplet, for example, of 
pus or blood, over the surface of nutrient agar-agar solidified obliquely. 





h. c. 'I. 

Fig. 90. — STKErTOcoccus Pyogenks Bovis. Pure-cultures on nutrient 

gelatine. 
a, Sub-cnlture from agar. h. Sub-culture from broth. 

c, Sub-culture from milk. d. Sub-culture from milk. 

The tubes ai'e then placed in the incubator at 37° C. ; the .strepto- 
cocci will appear in the course of two or three days in the form 
of minute dotted colonies. If present alone, and in consiilerable 
(juantities, the inoculated surface will exhibit a pure cultivation 
consLsting of a number of such colonies, whilst a flocculent mass is 
observed in the liquid which collects at the bottom, of agar-agar 
tubes ; this florculeut mass will be found to be composed of chains. 
From such a tube inoculate a number of the small flasks employed 
in Pasteur's laboi'atory for cultivations in hqiiids. In this way 
a number of pure- cultivations in milk and broth are estaljlished, 
which can le re.idily examined fi'om time to time. From a pure- 



SUPPURATION, PYjEMIA, SEPTICEMIA, ERYSIPELAS. 189 

cultivation in broth or agar-agar tubes of nutrient gelatine can 
be inoculated. Cover-glass-preparations from the growths on sohd 
media can be made in the iisual way, and stained with either a 
watery solution of fuchsine or gentian violet : but to stain prepa- 
rations made from milk or broth, or from the liquid in agar-agar 
tubes, use the method of Gram ; the stain will then be removed, 
except fi'om the streptococci, and very beautiful pj'eparations 
result. 

GONOKEHOEA. 

Gonorrhoea is the result of a catarrhal inflammation of the 
mucous membrane of the urethra, vagina, or conjunctiva caused by 
a characteristic pyogenic organism discovered by Neisser in 1879. 

GonococGUs of Neisser. — Cocci, usually in pairs 1-6 /i, in 
length, -8 ft, in width, and tetrads, with those surfaces of the com- 
ponent elements which are in contact, flattened. The elements 
are more or less kidney-shaped, and are separated by a clear 
unstained interval. They are found free in the pus and also in 
the interior of the pus cells. They stain with the aniline dyes, 
but are decolorised by Gram's solution. They do not grow on 
the ordinary media, such as gelatine, agar, and potato, in mai'ked 
contrast to the common pyogeiiic cocci ; but Bumm succeeded in 
obtaining a cultivation by using human blood serum, which was 
procured for the purpose from the placenta. They give rise to 
a very delicate growth in the form of an almost invisible film, 
with a moist appearance, which attains its full development in 
a few days. Steinschneider used human blood serum and agar 
incubated at 35° C. 

Krall recommended either agar with grape-sugar and blood serum, 
or the same mixture with the addition of 5 per cent, glycerine. 
Others have employed nutrient agar with the surface moistened with 
sterilised human blood. More I'ecently Keifer has been successful 
with a medium which is prepared in the following way : ascitic 
fluid is filtered and sterilised by Tyndall's pi'ocess, to this is added 
an equal quantity of the following mixture, agar 3-5, peptone 5, 
glycerine 2, salt '5 (per cent.). The ascitic agar is solidified in a 
Petri's dish, and the culture incubated at 36° C. 

They have also been cultivated in albumin from plovers' eggs, 
and in the fluid obtained from a case of synovitis of the knee joint. 

Inoculation of rabbits, dogs, horses, and monkeys, has been 
invariably unsuccessful, but sub-cultures produce the disease in 
the healthy urethra. 



190 INFECTIVE DISEASES. 

The cocci are found in pus from the urethra and other mucous 
membranes affected by the disease. They have also been found in 
urethral and inguinal abscesses in association with Staphylococcus 
pyogenes aureus. 

Method op Staining. 

I 

CovQr-glass preparations are made in the usual way, and 
double stained with LofHer's methylene blue, and eosin. 

Schiitz recommends floating the cover-glasses for five or ten 
minutes in a saturated solution of methylene blue in 5 per cent, 
solution of carbolic acid. They are washed in water, rinsed in very 
weak acetic acid, and again washed in water. Safranin may be 
used as a contrast stain. 




7, 

Fig. 91.— Gonocoocus x 800 (Bumm). 
a, free cocci ; b, cocci in pus cells ; c, epithelial cell containing cocci. 

Egyptian Ophthalmia. 

There are two forms of ophthalmia in Egypt, one associated 
with Gonococcus and the other with a bacillus closely resembling 
the bacillus of mouse-septicsemia, but there are minute differences. 

Bacillus of Ophthalmia (Koch and Kartulis). Minute rods 
which do not grow on gelatine but readily on blood serum and 
nutrient agar, forming a plainly visible, whitish-grey shining growth. 
Animals are insusceptible, but cultures produced the disease in the 
human conjunctiva in two out of six cases. 



CHAPTER XIV. 

ANTHRAX. 

Anthrax is a very fatal malady, and most irregular in its be- 
haviour. At one time it attacks only one or two animals, and 
at another time it will destroy nearly all the stock on a farm. 
Farmers formerly regarded the disease as non-communicable, and 
possibly the result of excessive or improper feeding, or faulty sani- 
tation, or of climatic conditions over which no control could be 
exercised. It is obvious that so long as the disease was regarded as 
the result of unknown conditions, no explanation could be given of 
its recurrence from time to time, or of certain animals contracting 
the disease and others not, and no measures of any use could be 
suggested to cope with an outbreak. 

Anthrax has always been more prevalent on the Continent than 
in England, and this to some extent accounts for the fact that it has 
received greater attention abroad. In France, Germany, Hungary, 
Russia, and in India and Persia, anthrax at times produces wide- 
spread losses. In Siberia it is still known on this account as the 
Siberian Plague. 

On the Continent there are certain localities known as anthrax 
districts on -account of their reputation for anthrax — for example, in 
the Upper Bavarian Alps in Germany and in Auvergne in France. 

In 1849, Pollender happened to examine the blood of a cow 
after death from anthrax, and discovered peculiar rod-like bodies 
among the blood cells. The same observation was made independ- 
ently by Brauell and Davaine about the same time, but the greatest 
importance must be attached to the publication of Davaine's further 
researches in 1863. Many ridiculed the discovery of bacilli, and 
stoutly maintained that they were only blood crystals or accidental 
structures of no importance. 

For many years very little progress was made, and the statements 
of other observers who were able to verify and add to Pollender's 
and Davaine's discoveries, were still received with scepticism. 

191 



192 INFECTIVE DISEASES. 

Within the last few years u great change of ojainion has taken 
place. Bacteriologists have investigated the whole subject, so that 
at the present day we know exactly the cause of anthrax. 

Bacillus anthracis {Bacterldie du. charbon, Bacillus of splenic 
fever, Wool-sorters' disease, or malignant pustule). — Rods 5 to 20 /i 
long and 1 to 1-25 ;u. broad, and threads; spore-formation present. 
As a thorough knowledge of the life-history of this bacillus is of the 
greatest importance, the various steps to be followed in a practical 
study of it will be successively treated in detail. Its morphological 




'^''^r^ 



Fig. 92.— Bacillus Anthkacis, /. 1200. Blood corpuscles and bacilli 
unstained ; from an inoculated mouse (Fkankel and Pfeiffee). 

and biological characteiistics liave been very completely worked out, 
and it serves as an excellent subject foi' gaining an acquaintance 
• with most of the methods employed in studying micro-organisms. 

A mouse inoculated with the bacillus or its sjDores will die in 
from twenty-four to forty-eight houi's, oi' more rarely in from 
forty-eight to about sixty hours. 

Examination after Deatli. — The spleen is found to be considerably 
enlarged, and may be removed, and examined by making cover-glass 
preparations, inoculations in nutrient media, and subsequently 
sections. 

Vorer-ijlass Preparations. — In cover-glass preparations of the 
blood of the spleen the liacilli are found in enormous numbers. 
Pi'eparations should also be made with blood from the heart and 
witli the exudation from the lungs and other organs ; it will be 



DESCEIPTION OF PLATE Y. 
Bacillus Anthracis. 

Fig. 1. — From a cover-glass preparation of blood from the spleen of a guinea- 
pig inoculated with blood from a sow. x 1200. Powell and Lealand's 
apochromatic x\ Horn. imm. E. P. 10. 

Fig. 2. — From a section of a kidney of a mouse. Under a low power the 
preparation has exactly the appearance of an injected specimen. Under 
higher amplification the bacilli are seen to have threaded theirway along 
the capillaries between the tubules, and to have ' collected in masses in 
the glomeruli. Stained with Gram's method (gentian-violet), and eosin. 
X 500. 

Fig. 3. — Bacillus anthraeis and Micrococcus tetragenus. From a section from 
the lungs of a mouse which had been inoculated with anthrax three days 
after inoculation with Micrococcus tetragenus. A double or mixed infection 
resulted. Anthrax-bacilli occurred in vast numbers, completely filling the 
small vessels and capillaries, and in addition there were great numbers 
of tetrads. Stained by Gram's method (gentian- violet), and with eosin. 
X 500. 



Plate T. 













/• 



J 




^m 



/ 

Fig 2. 



/ 



.^^ 







Fig 3, 



BACILLUS ANTHRACIS 



ANTHRAX. 



193 



noted that in these the bacilli are present in very small numbers, or 
altogether absent. The bacilli should be examined both unstained 
and stained. The rods are straight or sometimes curved ; rigid and 
motionless. They can be stained with a watery solution of any of 
the aniline dyes, and are then seen to be composed of segments with 
their extremities truncated at right angles ; between the segments 
a clear linear space exists, which gives them a characteristic appear- 
ance (Plate v., Fig. 1). By double staining, with Gram's method and 
eosin, the rods are seen to consist of a membrane or hyaline sheath 
with protoplasmic contents. 

Drop-cultures. — A little of the blood from the spleen or heart 
may be employed to inoculate sterilised broth or blood serum. 
Several of these cultures should be prepared, and some of them 
placed in the incubator, and examined at intervals of a few hours. 
It will be observed that the rods grow into long homogeneous fila- 
ments, which are twisted up in strands, and partly untwisted in long 
and graceful curves. The filaments begin to swell, 
become faintly granular,, and bright, oval spores 
develop (Plate 1). The cultures in the incubator 
develop rapidly. A temperature of 30° to 37° 0. 
is the most favourable for spore-formation. The 
spores are eventually set free, and by making 
a fresh cultivation, or by injecting them into a 
mouse or guinea-pig, they germinate again into 
the characteristic bacilli, which in their turn 
grow into filaments and spores. When the spore 
germinates it swells, the envelope becomes jelly- 
like, and gives way at one or other pole, and the 
contents escape and gi-ow into a rod. 

Test-tube Cultivations in Ahitrient Gelatine. — 
Typically characteristic appearances are obtained 
by inoculating a 5 to 8 per cent, nutrient gelatine. 
A whitish line develops in the track of the inocu- 
lating needle, and from it fine filaments spread out 
in the surrounding medium (Fig. 93). The fila- 
ments are more easily observed with a magnifjdng 
glass. In a more soUd nutrient gelatine the gi-owth 
appears -only as a thick white thread. As lique- 
faction of the gelatine progresses, these appearances 
gradually alter, and the growth subsides to the bottom of the 
tube as a white flocculent mass. In exhausted culture-media, and 
sometimes in the blood, filaments are seen in a state of degeneration. 

13 





Pig. 93.— Puke Cul- 
tivation OF Ba- 
cillus ANTHKACIS 
IN Nutrient Ge- 
latine. 



194 



INFECTIVE DISEASES. 



This has also been observed in sections of the internal organs of a 
rabbit which had been inoculated with the anthrax bacillus and had 
died of septicemia the following morning. 

Test-tuhe Cultivations in Ntotrient Agar-agar. — Cultivated upon a 





Fig. 94. — Colonies of Bacillus akthracis, x 80 (Flugge). 
a, after 24 hours ; 6, after 48 hours. 

sloping surface of nutrient agar-agar a viscous snow-white layer is 
developed, but without access of air no cultivation can be obtained, 
the bacilli being aerobic. This can be demonstrated hj completely- 
embedding a piece of lung or spleen 
pulp containing bacilli, in nutrient 
agar-agar (p. 22). 

Potato - cultivations. — In about 
thirty-six to forty-eight hours a creamy- 
white or very faintly j^ello-wish layer 
forms over the inoculated surface, 
usually with a translucent edge, and 
sometimes a strong, penetrating odour 
of sour milk. 

Plate-cultivations. — From the spleen 
or blood of the heart cultivations may 
be made in nvitrient gelatine on plates. 
The colonies develop in about two days, according to the temperature 
of the room. They appear to the naked eye as little white spots 




Fig. 95.— Impeession-pkepaea- 



TION of a colony, 



70. 



ANTHRAX. 



195 



or specks, which, on examination with a low power of the microscope 
and small diaphragm, exhibit two distinct forms. One form, on 
careful focussing, has the appearance of a little compact ball of 

1 




Fig. 96.— Margin of a Colony, x 2.50 

twisted threads ; in the other, liquefaction of the gelatine has 
commenced, and the threads spread out like locks oi' plaits of 
hair in the neighbonring gelatine. These appearances are perfectly 
characteristic (Figs. 94, 96). 

Cover-glass Impressio7is. — The plate-cultivations should be also 
examined as soon as the colonies 
appear, by making cover-glass im- 
pressions (Fig. 95). The filaments, 
examined with a high power, will be 
.seen to consist of a number of rods 
or segments which are perfectly 
regular in form. On the other 
hand, filaments from a tube-cultiva- 
tion in a solid medium will often be 
found to be composed, not only of 
rods, but here and there of the so- 
called involution-forms (Fig. 97). ria, 
From cultures in gelatine and 
glycerine agar, very sti-iking preparations are sometimes obtained, 
with numerous large spherical and lemon-shaped elements. In' a 




97.— Filaments with oval and 

lEBEUULAli ELEMENTS, X 800. 



196 INFECTIVE DISEASES. 

cover-glass preparation from a potato-culture the individual, segments 
will be found to have a great tendency to be isolated one from the 
other, and there is copious spore-formation. 

Preservation of Spores.Spores may be preserved simply by allow- 
ing anthrax blood to dry and then seahng it in a tube. The spores 
from a potato-cultivation are treated as follows : — The inoculated 
surface bearing the creamy cultivation is sliced off in a thin 
layer, and is mashed up with distUled water in a glass capsule. 
Sterilised silk-thread is cut up into lengths of about a quarter of 
an inch, and allowed to soak in the paste for some hours, under a 
beU-glass. The threads are then picked out with a pair of forceps, 
and laid upon a sterilised glass plate, covered with a bell-glass, 
and allowed to dry. From the plate, when" perfectly dry, they 
are transferred to a small test-tube, which can be plugged with 
cotton-wool, or sealed in the Bunsen burner. 

Examination of the Tissues. — The organs should be hardened 
in absolute alcohol, and sections prepared and stained by the 
ordinary methods. The method of Gram is the most instructive, 
and eosin a very satisfactory contrast stain. The capillaries in 
the lungs, liver, kidney, spleen, skin, mucous membrane, etc., will 
be found to contain bacilli. In some cases the bacilK are so 
numerous that a section under a low power has the appearance 
of an injected specimen. 

Inoculation of Animals. — A thread containing spores, a drop of 
blood from an infected animal, or a minute portion of a cultivation, 
introduced under the skin of a mouse or guinea-pig, causes a fatal 
result, as a rule, in from twenty-four to forty-eight hours. Sheep 
fed upon potatoes which have been the medium for cultivating the 
bacillus, die in a few days. Goats, hedgehogs, sparrows, cows, horses, 
swine, and dogs are all susceptible. Rats are infected with diffi- 
culty. Frogs and fish have been rendered susceptible by raising the 
temperature of the water in which they hved. Oats, white rats, 
and Algerian sheep have an immunity from the disease. 

Attenuation of the Virv^.- — Toussaint attenuated cultures by 
exposing them for ten minutes to 55° 0. Pasteur obtained a similar 
result by resorting to lower degrees of temperature; and Koch, 
Gaffky, and Loffler concluded from their experiments, that from 
42° to 43° C. the bacillus was most easily deprived of its poisonous 
properties. By cultivating , the bacillus in neutralised broth at 
42° to 43° C. for about twenty days, the infecting power is weakened, 
and animals inoculated with it (jrremier vaccin) are protected against 
the disease. To obtain a still more perfect immunity, they are 



ANTHRAX. 197 

inoculated a second time witli material (deuxieme vaccin) which has 
been less weakened. The alnimals are then protected against the 
most virulent anthrax, but only for a time. From a weakened 
cultvire, according to Klein, new cultures of virulent bacilh can be 
started, and a c\ilture that can be used as a vaccine for sheep kills a 
guinea-pig, and then yields bacilli that are fatal to sheep. 

The virulence of the bacillus is also altered by passing the 
bacillus through different species of animals. The bacillus of sheep 
or cattle is fatal when re-inoculated into sheep or cattle ; but if 
inoculated in mice, the bacilli then obtained lose their virulence 
for sheep or cattle, only a transitory illness results, and the animals 
are pi'otected for a time against virulent anthrax. 

Exposure to a temperature of 55° C, or treatment with "5 to 
1 per cent, carbolic acid, deprives the bacilli of their virulence. 

Chauveau obtained a similar result by cultivating the bacillus at 
38° or 39° 0. under a pressure of eight atmospheres. The possibility 
of mitigating the virus depends upon the species of animal ; rodents 
cannot be rendered immune by any known anthrax vaccine. The 
nature of the toxic products has been described in a previous chapter 
(p. 42). 

Methods op Staining the BACiiiLus Antheacis. 

Cover-glass preparations of blood, etc., can be stained with a 
watery solution of any of the aniline dyes, or with Neelsen's solution 
and subsequent treatment with alcohol (p. 87). The preparations 
may be dried and mounted permanently in Canada balsam, but the 
typical appearances are best observed in freshly stained specimens 
examined in water. 

. The sheath and protoplasmic contents can be demonstrated in 
cover-glass preparations from the 
blood or spleen which have been 
stained with eosin after the method 
of Gram. 

Spores must be stained by the 
special methods already described. 
The most satisfactory preparations 
are obtained by double-staining with Fig. 98.— Spoees op Bacillus 
Ziehl-Neelsen solution and methy- anthkacis stained with 

, , _ ,_. _. Gentian Violet, x 1500. 

lene blue (Fig. 7). 

Tissue sections are best stained by the method of Gram, and 
afier-stained with eosin, picrobarniinate of ammonia, or picro-lithium- 
carmine. 




198 INFECTIVE DISEASES. 



Origin and Mode of Speead. 

As every outbreak of anthrax is the result of the introduction 
into the system of the bacilli, the question naturally arises, how- 
are they introduced on the farm ? Where do they come from ? and 
what are the channels of infection ? 

The spores of the bacilli may get into the soil, and may remain 
there in a dormant state for many years. The spores were believed 
by Pasteur to be taken up by earth-worms, carried to the surface 
and deposited in their castings. Animals grazing are thus liable 
to be infected ; but Koch's experiments tended to disprove this 
theory. Anthrax has been known to break out among cattle 
grazing on a field where several years previously some Russian 
hides from infected animals had been buried. By some means or 
other the spores may contaminate the grass, and hay imported 
from an anthrax district may start the disease on a farm on which 
it had never been known to occur. The spores may in a similar 
way be introduced with blood manure and bone manure, and with 
refuse used as manure. The skin, hair, wool, hoofs, and horns of 
infected animals, if soiled with blood, are contaminated by the 
bacUlus. 

Another way in which the disease can be communicated may 
be illustrated by the transmission of the disease to man. Those 
who handle carcasses, wool or hides of infected animals are liable to 
contract the disease. Slight scratches, cuts, bites, and pimples, may 
readily be inoculated with the bacilli or their spores. Veterinary 
surgeons, butchers, herdsmen, cattle drovers — in fact, all those whose 
occupation leads them to cut open or skin cattle, sheep, or horses, or 
to handle hides and wool — are liable to fall victims to this disease. 

In one case which was brought to the author's notice, a veteri- 
nary surgeon had been called to see a bullock which had died 
suddenly in a meadow. A post-mortem examination was made, and 
the veterinary surgeon wiped his hands, which were soiled with 
blood, on some rough grass, and then washed them in a stream. 
The sedgy grass made some small cuts on his fingers, and the 
result was that he was simultaneously inoculated with the blood 
of the bullock. Local anthrax followed, two of his fingers were 
amputated, and he fortunately recovered. In another case a butcher 
dressed the carcass of a beast which had died suddenly, and while 
doing so scratched a pimple on his neck. An anthrax pustule 
developed, and after a very serious illness he also recovered ; but 
in many cases the attack is ' fatal. " Wool-sorters' disease " is 



ANTHRAX. 199 

anthrax of the lungs. Bales of foreign wool contain not only wool 
from living sheep, but wool which has been clipped from skins of 
dead sheep. If any of the sheep died from anthrax the wool is 
sure to be contaminated with blood containing the bacilli, and then 
wool-sorters engaged in picking the wool readily inoculate them- 
selves through a scratch or pimple, or by inhaling the spores. In 
many cases Wool-sorters' disease is fatal. 

A farm may become extensively infected by the living animal. 
Blood containing the bacilli may be discharged from the mouth and 
nostrils, or be passed with the contents of the intestinal canal 
and bladder. The droppings contaminate the pasture or byre, and 
spore formation, especially in warm weather, quickly takes place. 
From this cause the disease may not only be conveyed to healthy 
cattle grazing with infected animals, but fresh cases may occur, year 
after year, on the same farm, and if hay is cut and sold off the farm, 
other cattle at a distance are similarly infected. If the flooring of 
cattle sheds is once soiled by infected animals it is easy to account 
for those otherwise mysterious outbreaks which occiir when the 
cattle are taken in for the winter. 

Another source of danger arises when blood from a diseased 
animal is washed into brooks or streams, for thus the disease may 
be carried to farms in which it was previously unknown. 

Preventive Measures. 

Early recognition and prompt action are essential to prevent the 
spread of any communicable disease. 

Unfortunately in the case of anthrax only too often the very 
first indication of the existence of the disease is the sudden death 
in the pasture or byre of an apparently healthy beast, or possibly 
of one or more sheep. Nevertheless, the importance of being able 
to recognise any early indications is very great, because an im- 
mediate and careful examination should at once be made of the 
stock on the farm, and suspicious cases isolated from the rest. The 
stock-man may notice that one or two animals tend to keep away 
from the others. They look dull and cease feeding, and possibly 
shivering may be observed. In horses swelling of the throat may 
occur, and in some places there is discharge of blood from the 
orifices. Death follows the appearance of these symptoms in a 
few hours, and often with startling suddenness. Cattle die rapidly, 
but sheep, though rapidly contracting the disease, do not as a rule 
die so suddenly. 



200 INFECTIVE DISEASES. 

The characteristic sign after death is enlargemeat of the spleen 
to three or four times its natural size. It is not only enlarged, 
but extremely soft and dark in colour. Blood spots are visible on 
the internal organs generally, and' the intestine often contains a 
quantity of blood. The examination of a drop of blood will show 
\inder the microscope the characteristic bacilli. It is, however, 
quite unnecessary to make an elaborate post-mortem examination 
in order to satisfy oneself whether the disease is really anthrax or 
not. If an animal has died suddenly and has created a suspicion 
of anthrax, all that it is necessar}' to do is to cut cff an ear — or a 
foot in the case of a sheep — and make a cover-glass preparation at 
the first opportunity. 

A farmer with a case of anthrax must be made to realise the 
fact that an enormous quantity of poisonous material has to be 
dealt with. In fact, an infected animal is more dangerous when 
dead than alive. The owner or person in charge must immediately 
notify to a police constable the existence, or even a suspicion of 
the existence, of the disease. Prompt measures must be taken 
to destroy the carcass and all traces of the blood, and thus to 
reduce to a minimum the chance of the disease spreading to the 
rest of the stock, and of creating fresh outbreaks in the future. 
Every possible precaution must be taken to prevent the blood of 
the dead animal from contaminating the pasture, byre, or water 
supply. The rest of the stock should be removed from the pasture 
or cowshed where the disease has broken out. It is desirable to 
give a complete change of food and water, and the whole of the 
stock should be examined every day for a week, and any animals 
showing a rise of temperature should at once be isolated from the 
rest. Preventive inoculation has been recommended to protect 
the rest of the stock, but there is not sufficient evidence of the 
safety . of the process to lead to the adoption of this treatment. 
Animals ready for the butcher may be removed from the risk of 
infection by immediate slaughter. To disinfect the pasture the 
best plan is a heavy top-dressing of lime, and after six weeks 
stock may be readmitted, though not without some risk. If 
year after year cases of anthrax occur on a particular pasture, 
the most obvious precaution is to keep stock from it altogether 
and convert it into arable land. As roots grown on anthrax- 
infected soil have been known to convey the disease, the wisest 
course if we have to deal with a small field or comparatively 
small tract of land is to throw it out of cultivation or to plant 
it with trees. 



ANTHRAX. 201 



Disposal of the Carcass. 

The surest method to render harmless all the bacilli which 
exist in the carcass is burning, but cremation offers practical 
difficulties, especially if several carcasses have to be destroyed. In 
the case of an animal dying in a tow^, the local conditions may 
render it best to adopt destruction by burning or by means 
of chemicals. In such a case the carcass should be covered 
with quicklime, and then taken, in charge of an officer of the 
Local Authority, to a horse -slaughterer's or knacker 's-yard, and 
destroyed by exposure to a high temperature, or by chemical agents 
especially in the vicinity of chemical works. Under the usual 
circumstances of death occurring on a farm, fortunately the simple 
plan of burial, with the addition of lime or other chemical agents, 
is perfectly efficacious, and even without the use of chemicals if the 
carcass has been left unopened, as the bacilli die ra/pidly if air is 
excluded. 

Some experiments carried out by M'Fadyean clearly indicate the 
importance of leaving the carcass unopened. 

On July 16th a sheep was infected with anthrax by feeding it with a 
virulent culture. Five days later it died, and a microscopic examina- 
tion of blood from the ear, immediately after death, showed very many 
anthrax bacilli. The carcass was left unskinned and unopened until 
July 27th, when the various organs were cut out of the chest and 
abdomen and placed in a tin box. The box was then buried at a depth 
of about two feet in garden earth, and left there undisturbed until 
February 16th, when it was exhumed. The organs had become con- 
verted into adipooere, and this was thoroughly mixed up with water and 
administered to a sheep. The sheep remained perfectly healthy. In 
another experiment a rabbit was inoculated with anthrax on June 1st. 
It died on June 3rd, and blood from the ear contained the bacilli. The 
rabbit was left unopened for three days, and then placed in a flower pot 
and buried in garden earth at a depth of two feet. It was exhumed on 
February 15th. The tissues were aU destroyed by putrefaction, and the 
earth in contact with the bones was administered to a sheep without 
conveying the disease or producing any ill effects. 

Thus, in the first experiment, the lungs and the intestines, in 
which spore formation was most likely to occur, were used as a 
test, and in the second experiment the entire carcass. In both cases 
there was destruction or disappearance of the bacilli, and these tests, 
therefore, confirm in a very marked way the opinion that prompt 
burial of the unopened carcass is a perfectly safe plan to adopt. 



202 INFECTIVE DISEASES. 

If an animal has died in a meadow, a pit six feet deep should 
be dug close to the carcass, and if quickhme can be procured with- 
out delay the carcass should be buried with a layer about a foot in 
depth beneath it and with about the same quantity to cover it, and 
the pit filled up with the excavated soil. 

If there are any traces of blood where the animal lay, the 
contaminated ground should be covered with quicklime or drenched 
with strong carbolic acid, and the whole of the site of burial fenced 
off for six months. If an animal dies near a brook or stream then 
the carcass must be removed for burial to a sufficient distance to 
prevent any reasonable probability of contamination of the water. 

If death has occurred in the byre, the carcass must be removed 
to the nearest and most convenient spot for burial, any fodder or 
litter which may have been in contact with the deceased animal must 
be destroyed, and the shed and cart and any utensils, hurdles, etc., 
disinfected. For the latter purpose thorough scouring with water 
and then washing with limewash is recommended. The limewash 
should be prepared immediately before use, and four ounces of 
chloride of lime, or half a pint of commercial carbolic acid, be added 
to each gallon of limewash. 

The following is an illustration of the value of prefventive 
measures based upon a knowledge of the exact nature of the disease. 
A farm on the banks of the Yeo was repeatedly attacked by 
anthrax. One morning two sheep died, and other cases followed. 
The farmer learnt that his predecessor had buried cattle which had 
died of anthrax on the very spot where the sheep were folded. He 
removed his flock, and had no further losses among the sheep, but 
he continued to lose cattle grazing in the pastures by the river. 
These pastures were occasionally flooded by the Yeo. Another 
farmer in the same locality heavily manured a field, and shortly 
afterwards anthrax broke out in a most deadly form on his farm. 

What was the cause of these mysterious outbreaks ? The 
explanation was forthcoming, and prevention an easy matter. The 
river Yeo received the washings from the wool factories at Yeovil, 
and the pastures were contaminated by anthrax spores in the 
deposit which was left behind when the flood subsided. In the 
second instance, it was found that the manure used for dressing 
the pasture consisted of a quantity of refuse from the wool factories. 

Infected wool from foreign countries is one of the principal 
sources of the disease in this coiintry, and the remedy is to insist 
upon the factories destroying their refuse instead of its being 
allowed to contaminate the rivers or to be sold as manure. 



ANTHRAX. 203 

So long as this source of the disease was unknown anthrax 
continued to be spread through the agency of the wool factories. 

Anthrax spores may also be introduced with foreign oats, hay, 
and manure, so that it is almost impossible absolutely to prevent 
the importation of the disease ; but the danger of its unlimited 
extension and disastrous losses can be minimised, and the com- 
munication of the disease to man and to swine entirely avoided by 
simple preoavitions. 

Anthrax in Swine. 

The occurrence of anthrax in swine is a siibject upon which 
there has long been considerable diversity of opinion. Some of the 




Fig. 99. — Antheax in Swine. From a photograph taken during lite, showing a 
swollen condition of the neck and throat six days after ingestion of part of the 
viscera of a bullock which had died from anthrax. 

earliest writers on the diseases of animals speak of outbreaks of 
anthrax among swine, but whether any or all of these outbreaks 
were examples of true anthrax has long been a matter of un- 
certainty; for it is well known that diseases quite distinct were 
included under the name anthrax. 

Menschel states that in an outbreak in which twenty-four 
persons were attacked with malignant pustule, many of them from 
eating the flesh of beasts suffering from anthrax, pigs which were 
fed on the same flesh also became affected, and a woman who ate 
some of the diseased pork was subsequently ill. 

Eoche-Lubin, while apparently accepting the occarrence of 
anthrax in swine, taught that the pig resisted inoculation with the 
blood of a different species. 



204 INFECTIVE DISEASES. 

In this country accounts have been published from time to time 
of a fatal disease in pigs induced by eating the flesh of animals 
which had died of what was described as " blood-poisoning." 

Some very striking cases occurred in the practice of Mr. "Wilson, 
of Berkhampstead, and were reported in the Veterinarian. A 
farmer consulted Mr. Wilson respecting an illness with which his 
pigs were affected, stating that two or three were dead and many 
others seriously ill. They were strong hogs, ra,nging from six to 
nine months old. On inquiry it was ascertained that the farmer 
had lost a beast suddenly about a week previously, that the carcass 
had been opened in the yard, and the viscera thrown to the pigs. 
Mr. Wilson expressed the belief that the disease was anthrax, and 
.stated that he found the pigs exhibiting many of the symptoms 
observable in cattle, with the additional one of enlargement round 
the throat from infiltration of a yellow fluid causing discoloration 
of the skin. 

Also, in the reports of the Agricultural Department of the Privy 
Council thirteen pigs were reported as suffering from anthrax in 
1886, and one hundred and fifty-nine in 1887. 

But the question arose whether the disease in the pigs was 
genuine anthrax or septic poisoning. 

Williams says : " The flesh of animals which have died or have 
been killed whilst suffering from the disease [anthrax] should not 
be used as food either for men, pigs, or dogs, as it is apt to cause 
death by blood poisoning " ; and Steel writes : " Pigs, dogs, and 
poultry should not be allowed to feed on blood, flesh, and ejecta 
of anthrax victims," birt no statement is made as to the nature of 
the illness produced. No doubt these writers have been greatly 
influenced by the opinion of many bacteriologists, for Toussaint 
maintained that pigs could not be infected with anthrax, and a 
similar view was at one time upheld in this country by Klein, 
who stated that pigs were very insusceptible. In Germany also, 
pigs have been credited with an immunity from this disease. 

In the face of these conflicting statements the author carried 
out a series of experiments in order to ascertain the nature of the 
disease in swine resulting from the ingestion of the offal of animals 
which had died of anthrax ; and the result of inoculation with blood 
of animals which had died of anthrax, and with pure cultivations 
of the Bacillus anthracis. 

As a result of these experiments genuine anthrax was produced 
in swine (a) by feeding them with anthrax offal ; (i) by injection 
of blood of a bullock which had died of anthrax; (c) by passing 



ANTHRAX. 



205 



bacilli through the guinea-pig, and transmitting them to swine by 
injection of blood from the spleen ; (d) by injecting a pure cultiva- 
tion of the anthrax bacillus ; (e) and lastly, the anthrax bacillus 
was isolated from swine in which the disease was accidentally induced 
on a farm, and the disease reproduced by inoculation of guinea-jjigs 
and mice with blood from the spleen. 

The Author's Conclusions. — Swine of all ages can be affected with 
anthrax. If the disease is induced by ingestion of anthrax offal, 
the tonsils are ulcerated, and constitute the point of access of the 
bacilli to the blood. In such cases the characteristic symptom is 




Fig. 100. — Anthrax in Swine. From a jihotogi-aph tsXaan post-mortem. Death 
occurred four days after the ingestion of offal from a bulloclc which had died of 
anthrax, and there was well-marked cedema of the throat, cheeks, and eyelids. 

enormous swelling around the throat. If the disease is induced by 
hypodermic injection, the same oedematous infiltration of the tissues 
occurs at the place selected for inoculation. Death may occur 
in twenty-four hours, or not until after five or six days. There 
is a rapid rise of temperature, usually a rash-like discoloration 
of the skin, sometimes loss of power over the hmbs, and general 
weakness and disinclination to move ; the a-nimal may lie helplessly 
on its belly, and utter plaintive cries when disturbed. At the post- 
mortem the most characteristic feature is the gelatinous redema 
which, in the case of ingestion of offal, is found around the throat. 
There is usually congestion of all the organs and engorgement of 



206 INFECTIVE DISEASES. 

the heart and large vessels, fluid in the cavities of the chest and 
abdomen, and enlargement and haemorrhage into the lymphatic 
glands. There is in some cases inflammation of the intestines with 
submucous and subserous haemorrhages. The spleen may be normal 
in size, pale and flabby, and the liver only slightly congested and 
friable; in other cases the condition is characteristic, the spleen 
is the seat of hsemorrhage, causing more or less local enlargement, 
which is superficially of a deep purple colour ; the liver may also 
bo greatly congested, very friable, and marked with purple patches. 
The examination of the blood of the heart and spleen for anthrax 
bacilli must be carried out with great perseverance and discrimi- 
nation, as they are present only in small numbers, and in some 
cases have given place entirely to septic organisms, Inoculation 
with the blood will produce either typical anthrax, or malignant 
cedema or some other form of septicaemia. Possibly in the cases 
arising from ingestion of offal the ulcerated condition of the throat 
affords a nidus and a means of access for septic organisms. It 
is also well known that blood in a state of putrefaction may 
contain the bacillus of malignant oedema. In the presence of 
putrefactive organisms the anthrax bacillus rapidly disappears. If, 
therefore, inoculation of guinea-pigs or mice is used as a test for 
ascertaining the nature of an outbreak in swine, it must not be 
concluded, if Pasteur's or some other form of septicaemia result, 
that the disease was not anthrax, while, on the other hand, the 
discovery of the anthrax bacillus in the blood of the pig, or the 
production of anthrax in guinea-pigs or mice, is positive evidence 
as to the nature of the original disease. 

Peuch, in France, had obtained similar results by injecting pigs 
with anthrax blood and anthrax cultures. He also carried out 
some interesting experiments bearing on public health. The leg of 
a pig which had died of anthrax was covered with pounded sea-salt. 
PrevioiTsly to the curing, a slice of the flesh was squeezed in a meat- 
press, and the liquid thus obtained was employed for inoculation. 
The animals inoculated died of typical anthrax. In six weeks the 
curing was considered to be completed, and a slice was cut from the 
ham and soaked in filtered water. The juice was extracted in the 
meat-press, and employed for the inoculation of four guinea-pigs 
and three rabbits. Slight swelling and a certain amount of redness 
at the seat of inoculation were the only results. A few drops of 
the muscle-juice were added to sterilised broth, and produced a 
mixed cultivation of micrococci and motile bacilK. A rabbit and two 
guinea-pigs inoculated with the cultivation remained quite healthj^ 



ANTHRAX. 207 

These experiments demonstrated that salting destroys the viru- 
lence of the flesh of pigs which have died of anthrax, but in order 
to obtain this result the salting must be thoroughly carried out. If 
the process be incomplete the flesh is still virulent. Thus the leg 
of a pig salted for only fourteen days furnished a juice which 
possessed a certain amount of virulence. Out of three inoculated 
rabbits, one died in ninety-seven hours of anthrax, and the others 
recovered. Three guinea-pigs all succumbed, and a fourth guinea- 
pig inoculated with a cultivation from the muscle- juice also died. 
Peuch considers that there is danger in consuming flesh which has 
not been thoroughly cured. 

As it has been clearly shown that pigs may become infected with 
anthrax, these anim.als come under the Anthrax Order of 1886. 
This provides for the disposal of the carcass ; and although Peuch 
has shown that salting destroys the virulence of the flesh of pigs 
which have died of anthrax, there can be no doubt that it is quite 
right that such animals should be condemned as unfit for food. 

Further, the recognition of the occurrence of true anthrax in 
swine is an additional reason for condemning the Continental practice 
of eating hams, sausages, etc., in the raw state. Indeed, the viru- 
lence of anthrax flesh suggests one possible explanation of some 
of those obscure cases of meat poisoning which have occurred in 
this country. It is possible that the flesh of animals which had 
died of anthrax was used in the preparation of sausages, pork- pies, 
etc., and that the cooking was not sufficient to deprive the meat 
of its poisonous properties. 

Equine Antheax. 

Veterinary authorities have described " Anthrax in the Horse," 
but it remains to be seen whether there are not two or more affec- 
tions included under this heading. Fleming says : " The most 
acute form of anthrax, the apoplectic, is somewhat rare in the 
horse, and has perhaps been most frequently observed on the 
Continent. Though cases are recorded, but through an error in 
diagnosis, under other names in the veterinary literature of this 
country, I have only witnessed two cases in England ; though during 
the intense summer heat in the north of China I had several." 

The question to which the author is in a position to give a 
definite answer is, whether the disease produced by the Bacillus 
anthracis ever occurs in the horse. Whether that has been pre- 
viously determined, at any rate in this country, it is diflScult to say. 



208 INFECTIVE DISEASES. 

Fleming in. describing the pathological anatomy of anthrax in the 
horse, says : " The spleen is double and treble its ordinary volume ; 
its surface is sometimes bosselated by tumours ; its texture is 
softened and transformed into a viscid reddish-brown or violet mass, 
and the mesenteric glands are infiltrated. The blood in it has 
been found to contain bacteridia when examined soon after death." 
Williams, who says that "anthrax in the horse rarely occurs 
in this country," adds, that it is prevalent in India, and is 
there termed " Loodiana disease," and in Africa " Horse-sick- 
ness." But " Horse-sickness," from recent researches, is certainly 
not anthrax. Williams described a case which occurred in 1879 as 
one of anthrax. A carriage-horse died suddenly while in harness ; 
" a large black tumour was found in the lungs, and the pulmonary 
arteries were engorged with black tarry blood, which, when micro- 
scopically examined, was found to contain the bacilli in a most 
perfect form, and very numerous indeed." In 1884, an outbreak 
of charbonous fever occurred in Liverpool. Williams proceeded 
to investigate the outbreak, and found two horses dead on his 
arrival, one having died only a few hours previously. The bacilli 
from the blood in this case are figured, and the following statement 
made : " These bacilli seem to differ from those of splenic fever, being 
rather smaller in diameter, and so far as my observations go, 
multiply by fission only, not developing spores." 

On the other hand, the author investigated the blood of a mare 
which was supposed to have died of anthrax, and on examining 
cover-glass preparations of the blood, it was found to contain large 
numbers of bacilli with the characteristic microscopical appearances 
of anthrax bacilli. To place the question beyOnd any possible doubt 
a number of tubes of agar-agar were inoculated. These, after three 
days in the incubator, produced typical cultivations, and on examina- 
tion by the ordinary methods and by double-staining, yielded very 
beautiful preparations of filaments and spores. 

At the same time that the cultivations were prepared, two mice 
were inoculated at the root of the tail with a trace of the blood. 
Two days afterwards they were both found dead, and with the 
characteristic post-mortem appearances, spleen much enlarged, and 
anthrax bacilli in enormous numbers. 

There can be no doubt that true anthrax occurs in the horse ; 
and the author, in 1887, recommended that it should be scheduled 
under the Contagious Diseases (Animals) Act, and equine anthrax 
has been included in the Anthrax Order of 1895. 

More recently Pemberthy has described cases' of equine anthrax 



ANTHRAX. 



209 



which he believes to have been the result of infection, from feeding 
on foreign oats or imported hay. 

Preventive Inoculation. — The prevention of anthrax by- 
means of protective inoculation or vaccination has been attempted 
on a very large scale in France, and it is claimed that the results 
have been very beneficial to agriculture in that country : — 







Total 
No. of 
Animals 
Vacci- 
nated. 


No. of 
Veteri- 
nary 
Be- 
portB. 


Animals 
Vacci- 
nated 
after 

E«C8)pt 

of 
Reports. 


Mortality. 


Total. 


Total 
loss 
per 
cent. 

1-08 


Average 
loss 
before 
Vacci- 
nation. 

10% 


Year. 


After 

1st 
Vacci- 
nation. 


After 
2nd 
Vacci- 
nation. 


During 
rest of 
Year. 




1882 


270,040 


112 


243,199 


766 


847 


1,037 


2,640 




1888 


268,506 


108 


193,119 


436 


272 


784 


1,492 


0-77 






1884 


316,563 


109 


231,693 


770 


444 


1,088 


2,247 


0-97 






1886 


342,040 


144 


280,107 


884 


735 


990 


2,609 


0-93 






1886 


■ 813,288 


88 


202,064 


662 


803 


614 


1,469 


0'72 




Sheepi 


1887 


293,572 


107 


■ 167,811 


718 


737 


968 


2,423 


1-29 




1888 


269,574 


60 


101,834 


149 


181 


800 


630 


0-62 






1889 


,239,974 


48 


88,483 


238 


286 


501 


1,024 


1-16 






1890 


223,611 


69 


69,866 


331 


261 


244 


836 


1-20 






1891 


218,629 


66 


63,640 


181 


102 


77 


360 


0-67 






1892 


259,696 


70 


63,125 


319 


183 


126 


628 


0-99 






>1893 


281,338 


SO 


73,939 


234 


66 


224 


614 


0-69 




Total 
/1882 


8,296,816 


990 


1,788,677 


6,668 
(0-32%) 


4,406 
(0'24%) 


6,798 
(0-38%) 


16,872 


0-94 


s% 


36,654 


127 


22,916 


22 


12 


48 


82 


0-36 




1883 


26,468 


130 


20,501 


17 


1 


46 


64 


0'31 






1884 


33,900 


139 


22,616 


20 


13 


62 


85 


0-37 






1885 


34,000 


192 


21,078 


32 


8 


67 


107 


0'.60 




Oxen 

or 
Cows 


1886 
1887 
1888 
1889 


89,154 
48,484 
34,464 
82,261 


136 

148 

61 

68 


22,113 
28,083 
10,920 
11,610 


18 
23 
8 

14 


7 

18 

4 

7 


39 
68 
85 
31 


64 

109 

47 

52 


0-29 
0'39 
0'43 
0-45 






1890 


83,965 


71 


11,067 


5 


4 


14 


28 


0-21 






1891 


40,786 


68 


10,476 


6 


4 


4 


14 


0-18 






1892 


41,609 


71 


9,767 


8 


3 


16 


26 


0-26 






^1898 


88,154 


45 


9,840 


4 


1 


13 


18 


0-18 








438,824 


1,266 


200,962 


177 
(0-09%) 


82 
(0-04%) 


432 
(0'21%) 


601 


0-84 



14 



210 



INFECTIVE DISEASES. 



The vaccine is supplied, by a company in Paris, in two strengths. 
Reports are supplied by veterinary surgeons, and the results have 
been tabulated by Chamberland and published, and commented upon 
by Cope in a report to the Board of Agriculture (1894). The column 
of deaths, in the above table, includes the animals which died from 
the vaccination, and those which died from natural infection. 

It is claimed that the percentage of losses has been reduced from 
10 per cent, to '94 per cent, in sheep, and from 5 per cent, to '34 per 
cent, in cattle. Cope, in the report just referred to, regards these 
conclusions as somewhat fallacious, because in order to prove that 
the animals inoculated received immunity, it should be shown that 
they were subsequently exposed to the risks of natural infection. 
This was not the case. But a report obtained from the Bureau in 
Paris gives the actual number of animals on each of the infected 
farms, and the number which have died of the disease ; and when 
compared with Ohamberland's statistics it is evident that nine-tenths 
were not on farms where the disease appeared — at least, during 
1889-92 — and that the deaths from anthrax on those farms where 
it was reported to exist were, if anything, higher than they were 
supposed to be prior to the introduction of the system of vaccination ; 
and in spite of the immense number of animals vaccinated the 
official returns obtained from Paris, by Cope, indicate that the 
mortality from anthrax, calculated in the ordinary way, remains as 
high as ever. 



Anthrax in France. 



Year. 


No. of 
Outbieaks re- 


No. of Animals 


No. of which 


Percentage of 




ported. 


in Premises. 


Died. 


Loss. 


1889 


618 


22,599 


1,458 


6-5 


1890 


536 


24,073 


1,123 


4-7 


1891 


570 


21,356 


1,444 


6-8 


1892 


607 


28,199 
Cattle. 


1,581 


5-6 


1889 


— 


6,059 


700 


11-6 


1890 


— 


5,865 


771 


14-4 


1891 


— 


7,299 


849 


11-6 


1892 




5,058 
, Sheep. 


804 


15-9 


1889 


— 


16,540 


755 


4-e 


1890 


— 


18,708 


352 


1-9 


1891 


— 


14,057 


545 


4-2 


1892 





23,141 


777 


3-4 



ANTHRAX. 211 

In Germany, veterinary and agricultural authorities agree that 
the results have not met with the success which has been claimed 
for vaccination in France. Experiments were undertaken for the 
German Government, and in one set of experiments twenty-five sheep 
were vaccinated with the first vaccine without an accident, but three 
died five days after the second vaccine. In another experiment two 
hundred and fifty-one sheep were vaccinated with only one death, 
and subsequent inoculation with virulent anthrax proved that they 
had immunity. 

Six head of cattle were vaccinated without any loss, and six more 
were used for a control experiment. Inoculation with virulent virus 
proved fatal to the control animals, but the vaccinated were pro- 
tected. These, with other animals similarly vaccinated, amounting 
in all to two hundred and sixty-six sheep and eighty-three head of 
cattle, were then turned out to graze on infected pastures with two 
hundred and sixteen unvaccinated sheep as a control experiment. 
Within five months four of the vaccinated and eight of the un- 
vaccinated sheep died of anthrax, and one of the vaccinated and one 
of the unvaccinated cattle. 

The result of these experiments led to the following conclusions : — 

(1) That the first vaccine is- mild and harmless. 

(2) That, the second vaccine, even in the hands of experts, is 
dangerous and often fatal. 

(3) That sheep are more affected than cattle by the injections, 
exhibiting fever and other indications of illness. 

(4) That cattle and sheep which recover from the vaccination 
have an immunity against anthrax when tested by experimental 
inoculation. 

(5) That vaccinated cattle' and sheep tested by exposure to 
natural infection by grazing on infected pastures contract the 
disease in the ordinary way. 

(6) That the time for which immunity is conferred has not been 
determined. 

In England, Klein tested the vaccine, with the result that animals 
either succumbed to the vaccine, or to virulent anthrax after recovery 
from the vaccine. Protective inoculation has also been employed in 
a few instances by leading agriculturists, but with very unsatis- 
factory results. 

Stamping-out System Iii Germany the conclusion is that 

the safest measures are destruction of carcasses and disinfection, and 
that inoculation will have no effect in lessening the loss caused by 
this disease. 



212 INFECTIVE DISEASES, 

In England the stamping- out system has been advocated for 
many years, and is still regarded as the only reliable means for 
suppressing the disease ; and the possible introduction of the disease 
among healthy stock by vaccination, and especially in localities in 
which anthrax is unknown, would be contrary to the principles upon 
which the system is based. These principles are illustrated by the 
following extracts from the Anthrax Order of 1895 : — 

Notification. 

2. — (1) Every person having or having had in his possession or under 
his charge, an animal affected with or suspected of anthrax, shall, with all 
practicable speed, give notice of the fact of the animal being so affected or 
suspected, to a constable of the police force for the police area wherein 
the animal so affected or suspected is or was. 

(2) The constable shall forthwith give information of the receipt by 
him of the notice to an Inspector of the Local Authority, who shall forth- 
with report the same to the Local Authority. 

(3) The Inspector of the Local Authority shall forthwith give 
information of the receipt by him of the notice to the Medical Officer 
of Health of the Sanitary District in which the affected or suspected 
animal is or was. 

Duty of Inspector to act immediately. 

3. An Inspector of a Local Authority on receiving in any manner 
whatsoever information of the supposed existence of anthrax, or having 
reasonable ground to suspect the existence of anthrax, shall proceed with 
all practicable speed to the place where such disease, according to the 
information received by him, exists, or is suspected to exist, and shall 
there and elsewhere put in force and discharge the powers and duties 
conferred and imposed on him as Inspector, by or under the Act of 1894 
and this Order. 

Public Warning as to Existence of Disease. 

4.— (1) The Local Authority may, if they think fit, give public 
warnmg by placards, advertisement, or otherwise, of the existence of 
anthrax in any shed, stable, building, field, or other place, with or without 
any particular description thereof, as they think fit, and may continue to 
do so durmg the existence of the disease, and, in case of a shed, stable, 
buildmg, or other like place, until the same has been cleansed and dis- 
infected in accordance with this Order. 

(2) It shall not be lawful for any person (without authority or 
excuse) to remove or deface any such placard. 

J/!lk of Diseased or Suspected Cow not to be Removed. 
5. Where anthrax exists or has existed in any shed, stable, building, 
or other place, it shall not be lawful to remove from such shed stable 



ANTHRAX. 213 

building, or other place the milk of any cow which is affected with or 
suspected of anthrax. 

Removal of Dung or other Things. 

6. It shall not bei lawful for any person to send or carry, or cause to 
be sent or carried, on a railway, canal, river, or inland navigation, or in 
a coasting vessel, or on a highway or thoroughfare, any dung, fodder, or 
litter that has been in any place in contact with or used about a diseased 
or suspected animal, except with a Licence of the Local Authority for 
the District in which such place is situate, on a certificate of an Inspector 
of the Local Authority certifying that the thing moved has been, so far 
as practicable, disinfected. 

Disposal of Carcasses. 

7. — (1) The carcass of an animal which at the time of its death was 
affected with or suspected of anthrax shall be disposed of by the Local 
Authority as follows : — 

(i.) Either the Local Authority shall cause the carcass to be buried 
as soon as possible in its skin in some convenient or suitable place 
removed from any dwelling house and at such a distance from any 
well or watercourse as will preclude any risk of the contamination 
of the water therein, and at a depth of not less than six feet 
below the surface of the earth, having a layer of lime not less 
than one foot deep beneath, and a similar layer of lime above, 
the carcass ; 
(ii.) Or the Local Authority may, if authorised by Licence of the 
Board, cause the carcass to be destroyed, under the inspection of 
the Local Authority, in the mode following: The carcass shall 
be disinfected, and shall then be taken, in charge of an officer of 
the Local Authority, to a horse-slaughterer's or knacker's-yard 
approved for the purpose by the Board, or other place so approved, 
and shall be there destroyed by exposure to a high temperature, 
or by chemical agents. 

(2) With the view to the execution of the foregoing provisions of this 
Article the Local Authority may make such Regulations as they think fit 
for prohibiting or regulating the removal of carcasses, or for securing the 
burial or destruction of the same. 

(3) Before a carcass is removed for burial or destruction under this 
Article it shall be covered with quicklime. In no case shall the skin of 
the carcass be cut, nor shall anything be done to cause the effusion of 
blood. 

(4) A Local Authority may cause or allow a carcass to be taken into 
the District of another Local Authority to be buried or destroyed, with 
the previous consent of that Local Authority, but not otherwise. 

Digging Up. 

8. It shall not be lawful for any person, except with the Licence of 
the Board or permission in writing of an Inspector of the Board, to dig 



214 INI-ECTIVE DISEASES. 

up. or cause to be dug up, the carcass of any animal that has been 
buried. 

Disinfection in Case of Anthrax. 

9. — (1) The Local Authority shall at their own expense cause to be 
cleansed and disinfected in the mode provided by this Article— 

(a) Air those parts of any shed, stable, building, or other place in 

•which a diseased or suspected animal has been kept or has died or 

been slaughtered ; 
(5) Every utensil, pen, hurdle, or other thing used for or about any 

diseased or suspected animal ; 
(c) Every van, cart, or other vehicle used for carrying any diseased 

or suspected animal on land otherwise than on a railway. 

(2) The mode of the cleansing and disinfection of such shed, stable, 
building, or other place, or the part thereof, shall be as follows : — 

(i.) All those parts aforesaid of the shed, stable, building, or other 
place shall be swept out, and all litter, dung, or other thing that 
has been in contact with, or used about, any diseased or suspected 
animal shall be effectually removed therefrom ; then 

(ii.) The floor and all other parts of the shed, stable, building, or other 
place with which the diseased or suspected animal or its droppings 
or any discharge from the mouth or nostrils of the animal has come 
in contact, shall be, so far as practicable, thoroughly washed or 
scrubbed or scoured with water ; then 

(iii.) The same parts of the shed, stable, building, or other place shall 
be washed over with limewash made of freshly burnt lime and 
water, and containing in each gallon of limewash four ounces of 
chloride of lime or half a pint of commercial carbolic acid, the 
limewash being prepared immediately before use ; 

(iv:) Except that where any place as aforesaid is not capable of being 
so cleansed and disinfected, it shall be sufiScient if such place be 
cleansed and disinfected so far as practicable. 

(3) The mode of the cleansing and disinfection of such utensil, pen, 
hurdle, or other thing, and such van, cart, or other vehicle aforesaid, shall 
be as follows : — 

(i.) Each utensil, pen, hurdle, or other thing, van, cart, or other 
vehicle, shall be thoroughly scraped, and all litter, dung, sawdust, 
or other thing shall be effectually removed therefrom ; then 

(ii.) It shall be thoroughly washed or scrubbed or scoured with water ; 
then 

(iii.) It shall be washed over with limewash made of freshly burnt 
lime and water, and containing in each gallon of limewash four 
ounces of chloride of lime or half a pint of commercial carbolic 
acid, the limewash being prepared immediately before use. 

(4) All litter, dung, or other thing that has been removed from any 
such shed, stable, building, place, van, cart, or vehicle as aforesaid, shall 
be forthwith burnt or otherwise destroyed or disinfected to the satisfac- 
tion of an Inspector of the Local Authority. 



ANTHRAX. 215 

(6) The Local Authority may make such Regulations as they think 
fit for the purpose of carrying out the provisions of this Article. 

Occupiers to give Facilities for Cleansing. 

10. — (1) Where the power of causing any place, thing, or vehic'e to 
be cleansed and disinfected under this Order is exercised by a Local 
Authority, the owner and occupier and person in charge of the place, 
thing, or vehicle shall give all reasonable facilities for that purpose. 

(2) Any person failing to comply with the provisions of this Article 
shall be deemed guilty of an offence against the Act of 1894. 

Regulatimis of Local Authority as to Movement of Animals, 
Fodder, etc. 

11. A Local Authority may make such Regulations as they think fit 
for the following purposes, or any of them : — 

(a) For prohibiting or regulating the movement of any diseased or 

suspected animal into or out of any shed, stable, building, field, or 

other place, or any part thereof ; 
(6) For prohibiting or regulating the movement of any animal into 

or out of any shed, stable, "building, field, or other place, or any 

part thereof, in which there is or has been any diseased or 

suspected animal ; and 
(c) For regulating the removal out of any shed, stable, building, 

field, or other place of any fodder, litter, or other thing that has 

been in contact with or used for or about any diseased or suspected 

animal ; 
but nothing in any such Regulation shall authorise movement in 
contravention of any provision of any Order of the Board for the time 
being in force ; and a Regulation under paragraph (6) of this Article 
shall operate so long only as any animal which in the judgment of the 
Local Authority is disease'd or suspsoted remains in the shed, stable, 
building, field, or other place to which the Regulation refers, and, in case 
of a shed, stable, building, or other like place, until the same has been 
cleansed and disinfected in accordance with this Order. 

Slaughter in Anthrax and Compensation. 

12. — (1) A Local Authority may if they thmk fit cause to be 
slaughtered — 

(a) Any animal affected with anthrax or suspected of being so 
affected ; and 

(6) Any animal being or having been in the same field, shed, or other 
place, or in the same herd or flock or otherwise in contact with 
animals affected with anthrax, or being or having been in the 
opinion of the Local Authority in any way exposed to the infection 
of anthrax. 

(2) The slaughter of animals under this Article shall be conducted 
in such mode as will so far as possible prevent effusion of blood. 



216 INFECTIVE DISEASES. 

■ (3) The Local Authority shall out of the local rate pay compensation 
as follows for animals slaughtered under this Article :-,- 

(a) Where the animal slaughtered was affected with anthrax the 
compensation, shall be one-half of the value of the animal 
immediately before it became so affected ; and 

(b) In every other case the compensation shall be the value of the 
animal immediately before it was slaughtered. ' 

' (4) Provided, that if the owner of th6 animal gives notice in writing 
tp the. Local Authority, or their Inspector or other officer, that he objects 
to' the animal being slaughtered, it shall not be lawful for the Local 
Authority to cauSse that animal to be slaughtered except with the further 
special authority of the Board first obtained. 

Keeping of Swine in Slaughter-houses. 

16. It shall, not be lawfulfor any person, in any case in which the 
slaughter of any animal is authorised or required by this Order, to use 
for such slaughter any slaughter-house in which swine are kept. 

Whether an anthrax virus can be obtained which is absolutely 
incapable of creating centres of infection, and can therefore be 
recommended with safety for vaccination as an axixiliary and 
voluntary measure, is a matter for further investigation. 



CHAPTEE XV. 

QUAETER-EVIL.- — MALIGNANT (EDEMA. BAG-PICKERS' SEPTICAEMIA.- — 

SEPTICEMIA OE GUINEA-PIGS. — SbPTIC^MIA OF MICE. 

QuAKTER-EViL in cattle, maKgnant csdema, and rag-pickers' septiesemia 
in man, septiesemia in guinea-pigs, and septicaemia in mice, are all 
varieties of septicaemia produced by bacilli. 

An account of quarter-evil, malignant cedema, and rag-pickers' 
septiesemia may appropriately follow the chapter on anthrax, as 
they have cei-tain similarities to that disease. They are, however, 
not only distinct from anthrax, but must be carefully distingiiished 
from each other. In connection with these forms of bacillary 
septiesemia in man and cattle we may study bacillary septiesemia 
in small animals. 

QUARTEK-EVIL. 

The disease known in this country as quarter-evil or black-leg 
is identical with the French Charhon symptonuUique and the 
German Rauschhrand. Symptomatic anthrax in a very slight degree 
resembles anthrax. The disease occurs usually in young cattle from 
a few weeks to about twelve months old, and attacks sheep and 
horses,, but not swine or poultry. It is characterised by the develop- 
ment of ah emphysematous swelHng of the subcutaneous tissue and 
muscles, generally over the hind quarter. Infected animals cease 
feeding, the temperature rises, lameness supervenes, and death 
occurs in about forty-eight hours. The tumour on incision is found 
to contain a quantity of dark sanguineous fluid, with characteristic 
bacilli. 

Bacillus of Quarter-evil [Bacille du chctrbon symptomatique, 
Rauschhrand bacillus). — Motile rods with rounded ends, 3 to 5 yu. in 
length, -5 to '6 /a in breadth. Spore-formation present. The 
spores are oval, generally situated near the extremity of the rods, 
and when fully developed considerably exceed the rods in diameter. 

217 



218 



INFECTIVE DISEASES. 



Involution forms are freely developed 
cultures made in unsuitable media. The 






\ 






0' 



7*/o 



V 



<l,r' 



in old cultures, and in 
bacilli possess numerous 
flagella, and theii' 
power of movement at 
once distinguishes them 
fi'om anthrax bacilli. 
They can be cultivated 
in the ordinary media 
in the absence of oxy- 
gen, but more readily 
■with the addition of 
grape-sugar or glyce- 
rine. E-adiating fila- 
ments grow out from 
the more or less spher- 
ical colonies directlv 



i"iG. 101. B.iciLLi OF Quarter-evil x 1000. From 
an agar culture (Frankel and Peeiffek). 

liquefaction commences. In the depth of 
nutrient gelatine the growth occurs in two 
or three days at 20° to 25° 0. towards the 
lower part of the track of the inoculating 
needle. The gelatine slowly liquefies, and 
there is considerable formation of gas with 
the development of a peculiar odour. Spore- 
foivmation occurs freely in cultures, but not 
in the blood of infected animals until after 
death. 

Guinea-pigs inoculated with a pure- 
culture, or with spore-bearing threads, die 
in twenty-four to thirty-six hours. An em- 
physematous infiltration with sanguineous 
serum is produced at the seat of inoculation, 
and the surrovmding muscles are of a dark 
colour. The internal organs are more or less 
congested. The bacilli are found in the 
local exudation and in the surrounding 
tissue, and some hours after death in 



Fig. 102. Pure-Cui.tuhe op 
Bacilli of Quaeter-evil 
IN Grape-sugar Gela- 
tine (Frankel and 
Pfeiffek). 



QUARTER-EVIL. 219 

increasing numbers in the blood of the heart and in the internal 
organs. 

Quarter-evil and malignant oedema, though possessing points of 
resemblance, are distinct diseases. Not only do the bacilh in the 
two cases differ in minute morphological and biological details, but 
Eatasato showed that guinea-pigs rendered immune against virulent 
quarter-evil had no immunity against malignant oedema. 

Protective Inoculation — Arloing, Oomevin and Thomas have 
produced immunity by inoculating healthy cattle with a small 
quantity of the fluid from the tumour of an infected animal. 
Eeoovery takes place, and subsequent inoculation with a strong dose 
is without effect. Similar results may be obtained by intravenous 
injection of a few drops of the exudation. For general application 
of the system of protective inoculation, the virulent liquid and 
afiFected muscles are dried at 32° to 35° C, and the dried mass 
triturated with water and heated to 100° 0. This is used as the 
first vaccine. 

An infusion similarly prepared, but only heated to 80° 0., forms 
the second vaccine. The dxy powder is a convenient form for 
general distribution, and ^-^ of a gramme is triturated with 5 cc. 
of water, and ^ cc. is injected into each animal. In about ten days 
the second vaccine is employed, and cattle so treated are said to 
have a complete immunity from fatal doses. 

The place chosen for the injection is the under surface of the 
tail, a short distance from the extremity. The hair is chpped at 
this spot, and the point of a syringe is pushed in between the skin 
and the bone, and the vaccine slowly injected. 

Eoux and Chamberland produced immunity by inoculation of 
filtered cultures. Cultures in broth were deprived of bacilli by 
heating to 115° C, or by filtration through porcelain. Guinea-pigs 
were inoculated with three doses of 30 cc. at intervals of two days, 
and subsequently injected with a solution of virulent black-leg 
powder and lactic acid, which killed control animals in twenty-four 
hours. Kitt employed cultures on agar a fortnight old, or fresh 
cultures sterilised by steam for thirty minutes. It was found 
possible to confer immunity in oxen, sheep, and guinea-pigs against 
the most virulent extract. Kitt's method has the advantage over 
others of only necessitating one single injection. Whether these 
experiments are of scientific interest rather than of practical value 
may be regarded as an open question. 

On the Continent, and especially in France, vaccination against 
quarter-evil has been carried out extensively ; and by comparing the 



220 INFECTIVE DISEASES. 

mortality among the vaccinated and unvacoinated in localities where 
the disease commonly occurs, it has been said that the results are 
extremely favourable. The matter was investigated in this country 
by a committee : of , the Midland Veterinary Medical Association, 
and in the course of the experiments some surprising results were 
obtained. Six calves and four sheep were vaccinated, and five 
calves and two sheep were left unvacoinated as a control experiment. 
The seventeen animals were subsequently inoculated with virulent 
virus in the form of dried and powdered muscle. In forty-eight 
hours all the sheep died, and all the calves exhibited a swelling at 
the seat of inoculation. In another set of experiments, healthy 
calves inoculated with fresh juice from the tumour in a case of 
quarter-evil were not materially affected. The possibility of those 
calves which possess a natural immunity being classed as protected 
by the inoculation must be admitted, and the efficacy and safety 
of the process is by no means established. 

Malignant CEdema. 

The disease known by surgeons as progressive gangrene, gan- 
grenous emphysema, or surgical gangrene, has been shown by the 
researches of Chauveau, Arloing, Eosenbach and Bab^s, to be 
due to a bacillus identical with the microbe septique of Pasteur and 
the bacillus of malignant cedema of Koch. The bacillus or its spores 
may be spread by the neglect of antiseptics. The disease occurs 
especially after compound fractures and gun-shot wounds. 

If a guinea-pig is subcutaneously inoculated with earth, putrid 
fluid, or hay dust, death frequently occurs in from twenty-four to 
forty-eight hours. At the autopsy the most characteristic symptom 
is a widespread subcutaneous CEdema accompanied by air-bubbles. 
This originates from the point of inoculation, and contains a 
clear reddish liquid full of motile and non-motile bacilli. The 
internal oi'gans are little changed, the spleen is enlarged and of a 
dark colour, and the lungs are hypersemic, and have hsemorrhagic 
spots. Examined immediately after death, few or no bacilU are 
detected in the blood of the heart, but in that of the spleen, liver, 
lungs, and other organs, in the peritoneal exudation, and in and 
upon the serous coating of the abdominal organs, they are present in 
large numbers. If, on the other hand, the animal is not examined 
until some time after death, the bacilli are found in the blood of 
the heart, and distributed all over the body. 

Bacillus CEdematis Maligni, Koch (Pasteur's Septicjemia). — 



MALIGNANT CEDEMA. 



221 



Rods from 3 to 3' 5 /x long and 1 to 1"1 fj, wide ; they mostly lie 
in pairs, and then appear to be double this length. The rods are 
rounded at their ends, and form threads which are sometimes straight, 
but more commonly curved. In stained preparations they have a 
somewhat granular appearance. They are motile, possessing flagella, 
and form spores. The bacilli are distinguished from anthrax bacilli 
by their being somewhat thinner, by their rounded ends, and by their 
motility. Moreover, anthrax bacilli never appear as threads in fresh 
blood, and are differently distributed throughout the body. They 
are anaerobic, and can be cultivated on blood serum and on neutral 
solution of Liebig's meat extract in an atmosphere of carbonic acid. 
By embedding material containing bacilli in nutrient agar-agar 



\v 




Fig. 103. Bacilli of Malignant CEdema x 950. From the subcutaneous tissue 
of a guinea-pig. (Baumgakten. ) 

and nutrient gelatine, characteristic cultivations are obtained. The 
following process may be adopted to obtain a pure cultivation. A 
mouse inoculated subcutaneously with dust, as a rule, dies in one 
to two days. It is then pinned out, back uppermost, on a slab of 
wood, and the hail- singed with a Paquelin's cautery from one hind 
leg up to the neck, across the latter, and down again to the opposite 
hind leg. Following the cauterised line, the skin is cut through with 
sterilised scissors, and the flap turned back and pinned out of the 
way. With curved scissors little pieces of the subcutaneous 
(Edematous tissue, in the neighbourhood of the inoculated spot, are 
cut out, and sunk with a platinum needle in a 1 per cent, nutrient 
agar-agar, or 5 per cent, nutrient gelatine. Fragments of tissue may 
also be embedded by the method already described for anaerobic 
bacteria. 



222 



INFECTIVE DISEASES. 



The inoculated tubes are placed in the incubator. In a few hours 
a whitish turbidity spreads out from the piece of tissue, and upwards 
in the needle track. Examined microscopically, the turbidity is found 
to be due solely to the development of the baciUi of oedema. The 
surface expo.sed to the air exhibits no trace of the bacilli. To 
investigate the tubes microscopically, a sterilised glass tube with a 
capillary end may be used, with its neck 
plugged with sterilised cotton-wool, and 
provided at the mouth with a suction ball. 
The capillary end is thrust into the cultiva- 
tion, and a small fragment removed by 
aspiration. 

In the course of the first day the bacilli 
spread throughout a great part of the 
agar-agar in such a way that a more or less 
equally diffused cloudiness of the medium en- 
sues, with subsequent appearance of strongly 
marked clouds or lines of turbidity. At the 
same time gas- bubbles develop along the 
needle ti'ack, and a collection of liquid takes 
place, while spore-formation also commences. 
The following day these appearances are more 
marked, the opacity is moi-e pronounced, the 
development of gas increases, and the liquid 
contains more spore-forming bacilli and nu- 
merous free-spores. 

The nutrient-gelatine cultures during the 
iirst day show no macioscopic change, but 
Fir;. 104. Puhe-cultuhe J^-fter a few days the piece of tissue is sur- 
Of B.iciLi.us OF Malig- rounded with a white halo. This gradually 
spreads in all directions, and is apparently 
beset with hairs. The gelatine liquefies, and 
the fragment of tissue, degenerated bacilli, 
and spores, sink to the bottom. The cultivation is also very 
characteristic in a i per cent, nutrient agar-agar. If placed in 
the incubator, in a few hours a cloudiness forms around the piece of 
embedded tissue, which is caused by bacilli gradvially spreading in all 
directions in the nutrient medium. Mice inocvilated from these 
cultivations die more quickly than from the oi'iginal infection from 
dust. On potatoes they are cultivated by introducing a piece of liver 
or other tissue containing the bacilli, into the interior of a sterilised 
potato, and incubated at 38° C. The bacillus is not deprived of its 
virulence by cultivation. 



X.VNT (JiDEMA IN GhaPE- 

SuGAK Gelatine (Fhan- 
KEL and Pfeiffee). 




MALIGNANT (EDEMA. 223 

The spores of the asdema-baciUi appeal' to be veiy widely dis- 
tributed. They are found in the upper cultivated layers of the soil, 
in hay dust, in decomposing liquids, and especially in the bodies of 
suffocated animals, which are left to decompose at a high temperature. 
From any of these sources animals can be successfully inoculated. 
The bacillus is not only pathogenic in guinea-pigs, rabbits, and 
mice, but also in man and in farm animals, including calves but not 
cattle. Pure-cultures inoculated in animals produce cedema at the 
seat of inoculation 
without appreciable 
gas formation and / ■ 

without any putrefac- / " 

tive odour. The odoui- .; 

and frothy effusion / . 

resulting from the in- | ,■ '■•';. ^ 

oculation of earth are / * ^ , » ■^■ 

due to other bactei-ia, -■■ <• . 

which are introduced |: r 

simultaneously with ^" . ■? ' ^^^ 

the bacilli of malig- -^V. ' \' v 

nant cedema. The ^ ' ^^^ 

spleen is sometimes . ' 

slightly enlarged. By "'^*^^£,' 

touching with a cover- ^''aB-. 

glass the capsule of -yig. 105. Bacilli op Malignant (Edema x 1000. 
the spleen, or by ex- From an agar culture (Feankel and Pfeiffek). 
amining the serous 

effusion, the bacilli are found in abundance ; but if a preparation 
is made from the interior of the spleen or from the blood of 
the heart, no bacilli will be found until several hours after 
death. In this respect there is a marked difference from 
anthrax. Another difference is shown in spore-formation, which - 
occurs in the living body in malignant oedema, but never in 
anthrax. Animals which recover from the disease are said to 
be protected. 

Protective Inoculation Roux and Ghamberland produced 

immunity by injecting the chemical products in the filtrate obtained 
from cultures in broth. The serum from fatal cases will, it is said, 
confer immunity on other animals. 

There is a variety of this bacillus in soil according to Fliigge, 
agreeing in morphological and cultural but not in pathogenic 
characters. 



224 INFECTIVE DISEASES. 

Rag-pickers' Septicemia. 

llag-picjcers' disease has a resemblance to anthrax or wool-sorters' 
disease. After death the spleen is found to be enlarged, the in- 
ternal organs are congested, and there are haemorrhages on the 
.serous membranes. Bordoni -Uffreduzzi isolated bacilli which are 
quite easily distinguished from anthrax bacilli. They were found 
in the blood and in sections of the internal organs. 

Proteus hominis capsulatus. — Rods with rounded ends, singly, 
in pairs, and in filaments, somewhat smaller than anthrax bacUli, 
and often irregular in form. Spore-formation not described ; they 
have a well-marked capsule. Colonies are circular, appearing at 
first granular, and later possessing a filamentous structure. In 
the depth of gelatine they grow in the shape of a round-headed 
nail, like a culture of Friedlander's pneumococcus. On the surface of 
gelatine they form a shining white layer. On agar the growth is 
.somewhat transparent. On potato a moist, glistening film gradually 
spreads over the surface. They do not liquefy blood serum, and 
the growth is similar to that obtained on agar. They prove fatal 
to mice and dogs, but rabbits and guinea-pigs are not very sus^ 
ceptible.^ Dogs die usually on the second day after intravenous 
injection, and after death there is congestion of the internal organs 
and of the intestinal mucous membrane. Oedema is produced at 
the seat of inoculation in mice. There are haemorrhages in the 
lymphatic glands, and congestion of the hver and kidneys. Similar 
organisms have been described by Kolb and by Babeis in purpura 
hsemorrhagica. 

Septicemia of Guinea-pigs. 

Guinea-pigs and mice sometimes die of septicaemia, characterised 
by congestion of the lungs, liver, and kidneys, inflamed peritoneum, 
pleural and pericardial exudation, congested spleen, and congestion 
of the mucous and serous coats of the intestine. Klein isolated 
a bacillus from the blood and the internal organs in these cases. 

Bacillus of Septicaemia in Guinea-pigs Rods with rounded 

ends, motile, with pleomorphic forms, cocci, short rods and filaments. 
Colonies appear as small, circular, white dots, which enlarge and 
become irregular in outline. In the depth of gelatine a white 
filament develops, and on the surface the growth rapidly spreads 
with a crenated outHne. Broth becomes turbid, and after the second 
day a copious white sediment is deposited. Spore-formation not 
observed. 



DESOEIPTION OF PLATE VI. 
Bacillus Murisepticus. 

Fig. 1. — From a section of a kidney of a mouse wMcli had died after inocula- 
tion with a pure-cultivation of the bacillus. With moderate amplification, 
the white blood-corpuscles have a granular appearance, and irregular 
granular masses are scattered between the kidney tubules. Stained by 
Gram's method with eosin. x 200. 

Fig. 2. — Part of the same preparation with high amplification. The granular 
appearances are found to be due to the presence of great numbers of 
extremely minute bacilli, x 1500. 







\ '• ..■''■ 


■ ■ . 




•* "■•*• K, .- 


^5 ■ 




C 1?*' 




V 




''■' -I 






i- '-v ■ 


.1 i 


-^ ^„ 


■.--i- ;^ ,,^-^ 




^ ^ ^< 


* 




/' * 


>. 


©• , ■' 


•si%-j 


./ 5f. 






.' 




''V' . 


\ 




1 ^ ~ 


?'' 


-.o,>-.r;,. 


■>'' -^V'y. 




.Ml' '^ ' u 


(-?;'*,.- 




V oT-c-'C""-- 








': '^', 


1 


^ 


"i '~ 


- \ 


^ 


' ' ~ a^. 


' 


X. ^ ' 


■ 






^^ ^0$ 




If 


^M. '^''m 


-■', 








-'S''-^ 1 ■'(' 1 


,j\, , / 


-~-- ^\_, 




"vV- -"■' '■■ 




"V* "'1' 


■~' '^V -''■■' 






';(y,f,\^ J 


'!' 


~i,'-~ - _ -_^"\Mf-V-- 




'"\\\ 


Fig 2, 


, 


BACILLUS MURLSEPTICUS 






Km^nt Sroofc5,Dcy/ *Joi,iit?L. 


E.Jf.Croofc6TianJc/Mit. 







SEPTICAEMIA OF MICE. 



225 



I 



[1 



■I'll. 

i 



M 



41 



According to Wooldridge, the chemical products of this bacillus, 
separated by filtration, produce on inoculation immunity against 
virulent bacilli. 

Septicemia of Mice. 

Mice inoculated with a minimum quantity of putrid fluid often 
die of septicaemia. They rapidlj' sicken, their 
eyes inflame, their eyelids stick together, they 
become soporific, and death occurs in forty to 
sixty hours. There is slight oedema at the seat 
of inoculation, and enlargement of the spleen ; 
the bacilli are found free and in the interior 
of white corpuscles, both in the cedematous 
tissue and in the blood capillaries. 

Bacillus of Septicaemia of Mice (Koch). 
— Extremely minute bacilli, -8 to I /x long, and 
•1 to '2 fjL broad, and filaments. In cultivations 
in gelatine they do not appear to make threads, 
but the bacilli lie together in masses. Spores 
have been observed. The bacilli are probably 
non-motile. They are most commonly in the 
interior of white blood corpuscles. In these 
they increase, and in many cases a white blood 
cell is represented only hj a mass of bacilli. 

A minimal quantity of blood containing the 
bacilli produces the disease if inoculated in 
house-mice or sparrows. Field-mice have an 
immunity. Rabbits and guinea-pigs inoculated 
in the ear suffer only from a local erythema, 
which disappears after five or six days, and 
renders them for a time immune. Rabbits 
inoculated in the cornea suffer from an intense 
inflammation of the eyes. The bacilli form in 
plate- cultivations scarcely perceptible cloud-like 
specks, and in a test-tube of nutrient gelatine they form a delicately 
clouded cultivation along tlie needle track. 

Ah identical bacillus has been isolated in swine measles. 



I 




il' 



Fig. 106.— Puke Ci'l- 
tivatiok of the 
Bacillus of Septi- 
cemia OF Mice in 
NuTBiEN'T Gelatine. 
After two days. 



15 



ICHAPTEE XVT. 

SEPTICiEMIA OF BUFFALOES. SEPTIC PLEURO-PNEDMOJJIA OF 

CALVES.— SWINE p-EVER. — SEPTICAEMIA OF DEER. — SEPTICAEMIA 

OF RABBITS. FOWL CHOLERA. FOWL ENTERITIS. DUCK 

CHOLERA. — GROUSE DISEASE. 

There are several varieties of septicsemia occurring naturally in 
buffaloes, deer, calves, and birds, and artificially induced by inocula- 
tion of rabbits with septic material. They are associated with 
bacteria which agree in their morphological and cultural characters, 
though in some cases differing in their pathogenic properties. As 
the differences between the bacteria cultivated from these different 
sources is not greater than the differences which exist between 
the morphological, biological, and pathogenic effects of varieties of 
the tubercle bacillus, it will be convenient and fully justifiable to 
follow Hueppe and Baumgarten, and regard them as varieties of 
the bacillus of hcemorrhagic septiccemia. 

Epidemic Disease of Buffaloes. 

Oreste and Armanni investigated an epidemic among herds of 
young buffaloes in Italy {Buffel-seuche). The disease was extremely 
acvite, death occurring in from twelve to twenty-four hours. It was 
probably identical with an epidemic disease described by Bollinger in 
deer. The symptoms were fever, rapid pulse, discharge of mucus 
from the nose and mouth, and a local swelling of the head and face 
leading to suffocation. The only marked feature after death was 
hsemorrhagic inflammation of the small intestine. 

The bacilli were identical with those found by Schiitz in swine 
fever. Cultures inoculated in young buffaloes produced the disease. 
The bacilli were pathogenic to mice, guinea-pigs, rabbits, pigeons, 
and fowls, death taking place in from one to three days. 

226 



EPIDEMIC DISEASE OF DEER AND BOARS. 227 

Septic Pleuro-pnbumonia in Calves. 

Septic pleuro-pneumonia is a disease which attacks young calves 
within the first two months after their birth. Percussion and 
auscultation reveal lung mischief. The disease is very rapid and 
fatal, death occurring on the second or third day. In the less 
acute cases one or more lobes of the lungs ate found after death 
in a state of lobular and inter-lobular pneumonia. The inter-lobular 
connective tissue is distended with exudation, giving rise to white 
or yellowish bands between the inflamed lobules, which produce a 
marbled appearance, recalling the condition of the lungs in infectious 
pleuro-pneumonia. The internal organs are congested, and there 
are very often hsemorrhagic spots on the mucous and serous coats 
of the small intestine. All the organs contain rods identical with 
those of septicsemia of rabbits. Rabbits, guinea-pigs, and mice were 
infected. A calf was injected in the pleural cavity with a broth- 
culture, and died in twenty hours. 

Swine Fever. 

This disease vriW be described in a separate chapter. Several 
bacteria have been isolated by different investigators. In swine 
fever in Germany (Schwein-seuche) Lbffler and Schiitz isolated a 
bacillus which has been identified with the bacillus isolated by 
Salmon and Smith from hog-cholera in America, and with the 
bacillus of rabbit septicsemia and of fowl cholera. 

Epidemic Disease of Debe and Boaes. 

A very fatal epizootic (Wildseuche) occurred in the royal game 
preserves near Munich, destroying one hundred and fifty-three deer 
and two hundred and thirty-four boars (Bollinger). The disease 
lasted from twelve hours to six days. In the less acute cases pneu- 
monia and pericarditis supervened. In cattle there was also severe 
hsemorrhagic inflammation of the small intestine. In another form 
it produced swelling of the head, face, neck, and tongue. The 
virus proved fatal to rabbits in six to eight hours, and to sheep and 
goats in about thirty hours. A pig inoculated with a few drops of 
blood died in twenty-two hours. Kitt also investigated this malady. 
The bacteria were found to be identical, in their appearance and 
pathogenic properties, with extremely virulent bacteria from swine 
fever. Schiitz distinguished them from the bacteria obtained from 
swine fever by their pathogenic effect on pigeons, but cultures 
obtained from swine fever do not act uniformly in this respect. 



228 INFECTIVE DISEASES. 

Septicemia in Rabbits. 

Koch minutely investigated a disease of rabbits produced 
by inoculation with impure river water and with putrid meat 
infusion. Bacteria are found in the blood in abundance, and may 
be readily cultivated. 

The smallest quantity inoculated subcutaneously or in the cornea 

of a rabbit produces a rise of tempera- 

^ » '.^ ^ ture and laboured breathing after ten 

Ma I '^'' W to twelve hours, and death in sixteen 

, ^fi^ ' ''^ to twenty hours. The spleen and 

lymphatic glands are found to be 

Pio. 107.— Baotekium of Rabbit enlarged, and the lungs congested, 

SEPTicaiMiA; Blood 01' Spae- , , ,, , ,. , 

BOW X 700 (Koch). ^^* there are no extravasations, and no 

peritonitis. Mice and birds ai'e very 

susceptible ; guinea-pigs and white rats have an immunity. 

DaVAINe's SEPTICiEMIA. 

A disease was produced by Davaine by injecting rabbits with 
putrid blood. Rabbits, mice, fowls, pigeons, and .sparrows are sus- 
ceptible, and guinea-pigs and rats are insusceptible to the bacteria 
found in this disease. Rabbits inoculated with a trace of blood con- 
taining the bacteria, or' with a culture, died in from twenty-four to 
thirty-six hours. The spleen, liver, lungs, and intestines are highly 
congested, and sometimes extravasations and peritonitis are found. 

Fowl Cholera. 

Fowl cholera is an epidemic disease of the poultry-yard much 
dreaded in France, and well known through the researches of 
Perroneito, Toussaint, Pasteur, and Eatt. 



Si' 



^ ,„„ ^ „ „ Fig. 109.— Bactebium of Fowt 

"i-onf-;: If '!J''.°^ ^T"- Choleba, Choleba, X 2500. Muscle juice 

X 1200. rrom blood of inoculated Fowl. ^j j> j 

Fowls suffering from the disease usually die in from twenty-four 
to forty-eight hours. The disease shows itself by the fowls becoming 




FOWL CHOLERA. 229 

somnolent. They suffer from weakness of the legs, and their wings 
trail. There is freqviently diarrhoea, with slimy greenish evacuations, 
and death usually ensues after a slight convulsive attack. On 
making a post-mortem examination the viscera will be found to be 
congested, and there is intense inflammation of the mucous membrane 
of the intestine, with haemorrhages. 

The blood from the heart, and the intestinal contents, contain 
the bacilli which were at one time believed to be pecuHar to 
this disease. Inoculation subcutaneously, or administration with 
food, of a small quantity of a broth cultivation will produce death 
in twenty-four to thirty-six hours. Pigeons, pheasants, sparrows, 

■:■■■■ f" ■ ■■fjSj'C- 









'ii 



kt 



J' 






Fig. 110. — BACTEKinii OJ? Fowl Choleea. Section from liver of Fowl x 700 

(Flugge). 

rabbits, and mice are svisceptible. In guinea-pigs, sheep, and horse.s, 
an abscess develops at the seat of inoculation. Rabbits are readily 
infected by sprinkling a broth-cultivatiou on cabbage leaves or any 
suitable food. It was with this microbe that Pasteur proposed to 
eradicate the plague of rabbits in Australia. 

Fowl cholera has an additional interest, as it was with this 
disease that Pasteur first investigated the attenuation of virus. 
Broth-cultures which were several months old were found, when 
injected, to produce apparently only a local effect. This weakening 
of the virus was attributed by Pasteur to exposure to oxygen. 
After recovery the fowls were protected against the action of 
virulent cultures, while fowls not immunised died the following day. 
Kitt, by working with pure-cultures on solid media, showed that the 



230 INFECTIVE DISEASES. 

weakening was not due to prolonged exposure to oxygen, but that 
old contaminated broth- cultures after a time completely lost their 
power, owing to the antagonism of the bacteria accidentally present. 
Filtered broth-cultures contain the toxic products of the bacillus, 
and produce slight illness and subsequent immunity. 

Fowl Enteeitis. 

Fowl enteritis is an acute infectious disease of fowls, the course 
and symptoms of which are regarded by Klein as distinct from fowl 
cholera. The fowls sviffer from diarrhoea, with liquid greenish 
evacuations, bvit are never somnolent, and death occurs in one or 
two days. After death the-mucous membrane of the intestine is 
found to be congested, and coated with grey or yellowish mucus; 
the liver is congested, spleen enlarged, and lungs normal. There are 
a few bacilli in the blood of the heart, very many in the spleen and 
Hver, and they are in the form of a pure-culture in the mucus of 
the intestine. Klein says that the bacilli are a little longer and 
thicker than those found in fowl cholera, which they only slightly 
resemble, and that the course of the disease, the symptoms and 
pathological appearances, .definitely distinguish it from fowl cholera, 
but that nevertheless it belongs to the same family of bacilh. 
Pigeons are said to be insusceptible, rabbits only slightly susceptible. 
By feeding and by subcutaneous inoculation the disease can be 
communicated to healthy fowls, but there is no sign of illness until 
the fourth day. As regards attenuation, the bacilli behave like 
those from cases of fowl cholera. 

Duck Cholera. 

Duck cholera is an epidemic disease of ducks which was investi- 
gated by Oornil. The symptoms are similar to those of fowl cholera. 
They suffer from diarrhoea and weakness, followed by death in two 
or three days. 

The bacillus cultivated from the blood of ducks is pathogenic in 
ducks but not in fowls or pigeons, and large doses are required to 
kUl rabbits. 

GrEousE Disease. 
Grouse disease is an acute infectious disease of red grouse. 
According to Klein the chief pathological feature is severe pneu- 
monia ; there is also patchy redness of the serous and mucous linings 



GROUSE DISEASE. 



2n 



of the intestine, and the liver is congested and dark, hut the spleen 
is not enlarged. The bacilli are found in the heart, lungs, and liver, 
and in the extravasated blood. 



Cultures inoculated in mice and 
guinea-pigs produce pneumonia 
and death. Sparrows are sus- 
ceptible, and other small birds. 
Fowls, pigeons, and rabbits are 
insusceptible. 

Bacillus of Haemor- 

rhagic Septicsemia.— Very 

short rods, with rounded ends, 

"6 to '7 fjL in width and 1 "4 yu, in 

length. 






Fig. 111.- Bacillus op H.emokrhagic 
Septicaemia. Blood of a Rabbit after 
death from Septicemia x 950 (Baum- 
garten). 



In stained preparations the rods 
are observed to be deeply stained at the 
ends and to have a clear interval in the 
middle ; they were on this account mis- 
taken by earlier observers for dumb-bell 
micrococci or diplococci. They are non- 
motile, and spore-formation is unknown. 
They grow readily in the ordinarjr media. 
The colonies in nutrient gelatine appear 
about the third day. They are circular 
in form, ^vith a sharp dark outline, and 
of a yellow colour, lighter at the peri- 
phery. Later, the central zone is iinely 
granular, and of a dark yellowish-brown 
colour, with the lighter peripheral zone 
more clearly defined. In the depth of 
gelatine a delicate filament develops in 
the track of the needle, composed of 
minute spherical colonies, somewhat trans- 
parent, and yellowish-white in colour. 
At the point of puncture there may be no 
growth visible, or a flat and very limited 
growth. Inoculated on the surface of 
nutrient media a thin layer develops, 
with an iriegular serrated and thickened 
border. On potato different results have 
been obtained by different observers. Some maintain that a 
greyish-white or yellowish film will develop at the temperature of 
the blood ; but aecording to Caneva, the bacilli, whatever their source. 




Fig. 113. — Bacillus op 
H/E,1I0ERHAGIC Septic/Emia 
(Rabbit Septicaemia). Pure- 
culture in Gelatine after 
four days (Baumgarten. ) 



232 INFECTIVE DISEASES. 

will not grow on potato, while Bunzl-Federn maintains that the 
bacilli from fowl cholera and rabbit septicsemia do grow upon potato, 
but those from septicemia in deer, buffaloes, and swine do not. 
Opinions differ with regard to their action on milk. The reaction 
for phenol and indol is given in all cases, except with cultures 
obtained from septicaemia of buffaloes. The virulence of the bacilli 
may be diminished and attenuated, but it may subsequently be 
restored by successive inoculation in aniiftals. The pathological 
lesions vary in different animals. The most common result is con- 
gestion of the internal organs and haemorrhage. The bacilli culti- 
vated from cattle or deer produce fatal results -when inoculated 
in swine. The bacilli from any of these sources inoculated in 
pigeons will produce fowl cholera, but the bacilli isolated by Schiitz 
from swine, and those from deer, are not fatal to fowls. Further, 
the bacilli cultivated from swine fever are fatal to guinea-pigs, 
while the bacilli from rabbit septicaemia have very little effect upon 
them. The bacilli have been found in association with diseases of 
cattle, swine, deer, birds, rabbits, and mice, and have been cultivated 
from healthy mucous membrane. Veranus Moore found the bacilli 
in the m^ucus from the upper air passages, of 71 per cent, of cattle, 
85 per cent, of cats, and 33 per cent, of dogs. From these sources 
inoculations were made in rabbits, and rapidly fatal septicaemia 
was produced, associated in less acute cases with peritonitis, pleurisy, 
and pericarditis. 



CHAPTER XVII. 

pneumonia. infectious pleuro-pneumonia of cattle. — 

influenza. 

Acute Croupous Pneumonia. 

Pneumonia is an acute inflammation of the lungs with fibrinous 
infiltration of the air vesicles and interstitial tissue. There are 
varieties of pneumonia, and one form is commonly believed to be 
infectious. 

The lung passes through three stages — engorgement, red hepati- 
sation, and grey hepatisation. In the first stage -the lung is of 
a deep red colour, but still vesicular ; in the second stage the 
affected part is" more or less solid, and has the consistency of Uver, 
owing to the fibrinous lymph which is poured out into the alveolar 
cavities. In the grey hepatisation, the exudation contains more 
leucocytes and less fibrin, and this is followed by the stage of 
suppurative softening and final absorption. The sputum at the 
commencement of the disease is rusty, from the presence of blood, 
and later on has the appearance of prune juice. Examination of 
the sputum by Gram's method will reveal numerous micro-organisms, 
and two of these are deserving of special study — the pneumococcus 
of Friedlander, which is present in a considerable proportion of 
cases, and Sternberg's micrococcus, which was found in sputum by 
Talamon. ■• 

In 1888, there was considerable prevalence of pneumonia in 
Middlesbrough, with strong tendency to occur in groups of cases; 
but there was admittedly room for doubt whether the clinical 
and post-mortem appearances were not identical with ordinary 
pneumonia. Dr. Ballard maintained that there were facts and 
considerations which appeared to show that the disease was com-" 
municable from the sick to the healthy, and that it was a specific 
febrile disease, and Klein isolated and described the micrococcus 
present in these cases. 

Bacterium Pneumoniae Crouposse (Pneumococcus, Pried- 

233 



234 



INFECTIVE DISEASES. 




Vm. 113. — Bacterium Pneumoni.e 
Ckoupos.e, from Pleural Cavity 
OF a Mouse, x 1500. A, B. 
Thread-forms. C, D, E. Short 
rod-forms. G. Diplococoi. H. 
Cocci. I. Streptococci. (Zopf.) 



lander). — Cocci ellipsoidal and round, singly, or in pairs (diplococoi), 

rods and thread-forms. The cell- 
membrane thickens, and develojJS 
into a gelatinous capsule, which is 
round if the coccus is single, and 
ellipsoidal if the cocci occur in pairs 
or in rod-forms. Cultivated in a 
test-tube of nutrient gelatine they 
grow in the form of a round- 
headed nail, without Kquefaotion 
of the gelatine (Fig. 114). The 
cocci when artificially cultivated 
have no capsule, but it again 
appears after their injection into 
animals, 
The cocci 
can also be 
cultivated 

on blood serum and on boiled potatoes. 

They occur in pneumonic exudation. In- 
oculation of dogs with a cultivation of the 

cocci occasionally gave positive results ; but 

in rabbits no results followed. Guinea- 
pigs proved to be susceptible in some cases ; 

but thirty-two mice, after injection of a 

cultivation diffused in sterilised water, into 

the lungs, died without exception. The 

lungs were red and soHd, and contained the 

cocci, which were also present in the blood, 

and in enormous numbers in the pleural 

exudation. Inhalation experiments by sjDray- 

ing the cocci diffused in water into mouse 

cages produced pneumonia and pleurisy in 

three out of ten mice. 

The nail-shaped cultivation is not always 

produced, nor are these conclusions acceoted 

^' Fig. 



by all investigators. 

METHODS OF STAINING FEIEDLANDEE'S 
PNEDMOCOCCUS. 

Cover-glass prepar 
treated as follows : — 



114. — Feiedlander's 
Pneumococcus. Pm-e- 
oulture in nutrient- 
gelatine four days old 
(Baumoarten). 



rations of pneumonic sputum or exudation may be 



PNEUMONIA. 



235 



(a) Stain by the method of Gram, and after-stain with eosin. ' , 
(6) Treat with acetic acid, then stain with gentian-violet or Bismarck- 
brown. Examine in distilled water, or dry and preserve in Canada 
balsam. 

(c) Float them on weak solutions of the aniline dyes twenty-four 
hours ; differentiation between coccus and capsule is thus obtained. 

(d) Stain with osmic acid ; the contour of the capsules is brought 
out. 




i^ t 



Fig. 115.— Capsule-coooi from Pneumonia, x 1500 (Baumgabtbn). 

Sections of pneumonic lung should be stained by — 

(a) Method of Gram. 

(J) Method of Friedlander. This method is employed to demonstrate 
the capsules in tissue sections. It consists in placing the sections twenty- 
four hours in the following solution : — 

Fuchsine . . . 1 

Distilled water . . 100 

Alcohol ... . . 5 

Glacial acetic acid . ... 2 

They are then rinsed with alcohol, transferred for a couple of minutes 
to a 2 per cent, solution of acetic acid, and treated with alcohol and oil 
of cloves in the usual way, and preserved in Canada balsam. 

Sternberg's micrococcus was first found in the blood of rabbits 
inoculated with saliva. Three months afterwards, Pasteur encoun- 
tered the same organism in rabbits inoculated with the blood of a 
child suffering from rabies. The same orgariism in 1883 was found 
by Talamon in pneumonic sputum. It was identified by Sternberg. 
Two years afterwards further observations were made by Frankel, 
Gamaleia, and others. It has also been found in purulent meningitis 
by Netter, and by Monti in cerebro-spinal meningitis, by Weichsel- 
baum in ulcerative endocarditis, and by others in acute abscess of 
the middle ear, and in purulent inflammation of the joints following 
pneumonia. 



236 INFECTIVE DISEASES. 

Sternberg's Micrococcus. [Microbe de salive, Pasteur ; Micro- 
coccus Fasteuri, Sterubei'g ; Lancet-shctped micrococcus, Talamon ; 
Strejitococcus lanceolatus Pasteuri, Gamaleia ; Bijdoaoccus pnevr 
monice, Weichselbaum ; Bacillus septicus sputigenus, Fliigge ; 
Micrococcus of sputum septiccemia, Frankel.) Spherical or oval 
cocci, siiigl}', ill pairs or in chains, often lanceolate and capsuled. 
Stain readily with the anUiiie colours and by Gram's method ; 
non-motile. They flourish in alkaline media in the incubator. In 
broth they produce in twelve hours a cloudiness due to the develop- 

# ##< A 

Fig. 116. — MicKOcoccus of Sputum SEPTicEMi.i. From the blood of a 
Rabbit, x 1000 (Feankel and Pfeippee). 

ment of cocci and short chains. After a time these subside to 
the bottom of the tube, and the liquid above becomes clear. In 
plate -cultivations the colonies are small, circular, white, and granular. 
In the depth of gelatine, minute white colonies develop along the 
track of the needle without hquefaction of the gelatine ; and on 
the sloping surface of nutrient agar or blood serum minute trans- 
parent drops appear along the line of inoculation. They grow in 
milk, coagulating casein ; but they do not grow on potato. Sub- 
cultures quickly lose their virulence, but regain it by inoculation. 




PNEUMONIA. 



237 



The injection of a minute quantity (-2 cc.) of a virulent culture 
subcutaneously proves fatal to mice and rabbits in from twenty- 
four to forty-eight hours. Immediately afterwards there is a rise of 
temperature of 2° or 3° C, later it falls, and just before death 
it is several degrees below normal. After death, the post-mortem 
appearances of septicaemia are observed, in addition to diffuse 
inflammatory oedema extending in all directions from the point of 
injection. The subcutaneous connective tissue contains sanguineous 
serum and micrococci in abundance. The liver and spleen are some- 




FiG. 117.— Colonies of Steknbehg's Miceococcus. Agar plate-cultivation, 
after 24 hours, x 100 (Feankel and Pfeiffee). 



times dark and engorged, and blood from the heart and internal 
organs teems with mici'ococci. 

There is no indication of pneumonia after subcutaneous inocula- 
tion, but intra-pulmonary injections produce fibrinous pneumonia, 
often fatal (Talamon, Gamaleia). The result is usually fatal in 
rabbits .ind sheep, but dogs, as a rule, recover. Injection of cultures 
into the trachea of rabbits is said to ind uce typjca l pneumoiik. 

(Monti). ;flHli 

Sternberg concludes that this micrococcus is the cause of acute 
infectious pneumonia, but the micrococcus is undoubtedly associated 
with widely different pathological processes, and the possibility of its 
being a saprophyte, which finds in pneumonia a suitable soil for its 
development, must not be overlooked. 



238 INFECTIVE DISEASES. 

Klein's Micrococcus. — Klein found in pneumonic sputum a 
diplococcus which does not appear to differ from Sternberg's micro- 
coccus. In cover-glass preparations the bacilli are surrounded with 
a halo, but no definite capsule, as in Friedlander's coccus. They 
appear as short rods constricted in the centrfe, or dumb-bell forms, 
and forms intermediate between cocci and bacilli. In gelatine, after 
two or three days, greyish-white spots appear, which enlarge in the 
next two or three days into flat, translucent, greyish-white plaques, 
with irregular serrated outline. Colonies beneath the surface are 
spherical, and of a brownish-yellow colour. In test-tubes in the 
depth of the gelatine a whitish-brown filament develops on incuba- 
tion, composed of minute spherical colonies, and on the surface the 
growth spreads out into a greyish-white film with serrated margin. 
On the surface of obliquely solidified gelatine the growth forms a 
thin whitish film, which enlarges in breadth with irregular outline, 
reaching its maximum in about a fortnight. The growth on agar 
is very similar. Broth becomes uniformly turbid in twenty-four 
hours, then a powdery precipitate makes its appearance. On potato 
there is a thin, moist, faintly yellowish- brown film. Cultures examined 
in the fresh state show many rods in a resting stage, and others 
actively motile. In addition to the dumb-bell forms there are others 
of greater length, and in old cultures involuted and degenerated 
forms. Spore- formation has not been observed. A broth-culture 
Btispulated into two rabbits produced^^ocal tumour which subsided 
in^a week. Death ensued in one ca™in eight days, and in the 
other in three weekl. There was purulent matter at the seat of 
peculation in one ; in the other, pericardial exudation and hypersemia 
oKthe lungs. Broth- cultures inoculated intravenously produced no 
effect. In guinea-pigs there was swelling at the seat of inoculation, 
or slight^ indication of disease and recovery. Cultures inoculated 
in mice produced rapid breathing, drowsiness, and death in from 
twenty-four to ninety-si^ hours. The internal organs ■wferacon- 
gested, the lungs inflamed, and the blood and organs in the inoculated 
animals contained the diplococci in considerable numbers. 

Foa isolated a coccus which he named the Micrococcus lanceolatus 
i, capsulatus. It produced in small animals either rapid septicaemia 
' and death, or local oedema and death at a later period. 

Protective Inoculation. — Immunity has been produced in 
rabbits by the intravenous inje ction, of the virus in a diluted form. 
Blood obtained from immunised raTOts was kept at 10° 0. for twelve 
hours, and then flltered, and animals injected with it acquired 
immunity against virulent cultures (Emmerich). 



INFECTIOUS PLEURO-PNEOMONIA. 



239 



Filtered cultures are said to confer immunity for six months, and 
raising the temperature of filtered cultures increases the strength of 
the substance which gives immunity (Klemperer). The blood serum 
of immune animals can confer immunity on other animals, and, it is 
said, will arrest the progress of the disease produced by injection 
of healthy animals with virulent cultures. The cultures contain a 
proteid body, for which the name pneumo-toxin has been suggested, 
and anti-pneumo-toxin has been isolated from immunised blood 
serum. 

Inff.ctious Pleuro-pxeumonia. 

Infectious pleuro-pneumonia is a highly infectious disease 
peculiar to cattle ; it is characterised by rise of temperature and 
exudation into the lungs. It is often fatal, and sometimes exi.sts in 
an extremely chronic form. It is believed to have been unknown in 
England previously to 1840, and is supposed to have been introduced 
from Holland, where in one year it destroyed seven thousand cattle. 











Fig. 118. — Acute Cataekhal Pneumonia (Ox). 
<i, Coagulated mucus with catarrhal cells (c) embedded in it; b, catarrhal cellSiMVC ^'^ 
sprouting from alveolar walL, ^'^80. (Hamilton.) '" ^^ 

The disease cannot be conveyed 'artificially. A living, diseased 
animal must be the medium of infection. The disease is apparently 
only communicated by cohabitation. Brown injected large quanti- 
ties of lymph from diseased lungs into the jugular vein, into the 



240 



INFECTIVE DISEASES. 



tissue of the lungs, and into the trachea, without any result except 
a small abscess at the seat of puncture. Administration of the 
virus by the mouth gave equally negative results. The lungs from 
a recently killed animal infected with pleuro-pneumonia were placed 
in a shed occupied by healthy heifers, and left there for several days. 
Fodder, litter, and manure were taken from places in which there 
were diseased cattle, and placed in contact with healthy cattle, and sub- 
sequently all the animals used in these experiments were slaughtered 
and carefully examined, and the results were absolutely negative. 
Similarly negative results followed experiments made by )Sander- 







L'^s'V' r.^ssitw...'.'. ™ 



Fig. 119.— Infectious Plbuko-Pneumonia of Cattle, x 480. 

a,a^a, Exudation in air-vesicles, composed of a network of fibrinous lymph with 
entangled leucocytes; 6,&, the same caseating ; c, the air- vesicle filled with 
leucocytes only. In the centre is a blood-vessel filled with a fibrinous plug. 
(Hamilton.) 



son and Duguid, and thus coniirmed the conclusion arrived at bj' 
Brown, that the disease could only be communicated by actual contact 
of a living, diseased animal with a healthy one. 

The symptoms of the disease in cattle are a rise of temperature 
to 105° or 107°, and a peculiar dry cough, and later the usual 
indications of pneumonia, difficulty in breathing, and dulness on 
percussion. As a rule, death follows from exhaustion ; but the disease 
may also assume a chronic form, if the animal escapes slavighter, 
and the lung may become gangrenous or tubercular. The period of 
incubation is about thirty days, but it is uncertain. The lesions are 




Fig. 120.— Infectious Pleuko-pneumonia of Cattle, x 50. 
a,a,a. Spaces in deep layer of pleura and interlobular septa iilled with fibrinous 
lymph; h, deep layer of pleura running down to an interlobular septum ; c,c, 
air-vesicles filled with fibrinous lymph ; d, blood-vessels of alveolar walls, 
much congested ; e, large congested blood-vessels ; /,/, interlobular septa 
infiltrated with fibrinous lymph ;i a, blood-vessel in interlobular septum 
(Logwood, Eosin and Farrant's solution).— Hamilton. 

16 



242 INFECTIVE DISEASES. 

limited almost entirely to the lungs ; congestion is quickly followed 
by inflammation and effusion into the air vesicles and the intra- 
lobular iibrous tissue which is so well marked in the lungs of cattle. 
Leucocytes are entangled in the fibrinous lymph, and the intra- 
lobular septa are enormously enlarged, so that the red lobules are 
mapped out by the paler septa, and produce on section of the diseased 
parts a very striking marbled appearance. A somewhat similar 
appearance is sometimes observed in septic pleuro-pneumonia in 
•calves. The effusion occurs also in the air vesicles. The stages 
of grey hepatisation and suppurative softening have not,, as a rule, 
time to develop. Hsemorrhagic infarctions are sometimes produced, 
which in turn become gangrenous or cheesy, and a capsule may form 
round the diseased part. Eoy found micro-organisms in the lymph, 
but attached no importance to them. Bruylants and Verriet also 
•described a micro-organism in the lymph. Later, Poels and Nolen 
isolated a micrococcus resembling Friedlander's pneumococcus. 
Inoculation in the lungs produced a condition in cattle which they 
•considered indicative of pleuro-pneumonia. 

Lustig was unable to confirm these observations, but succeeded 
in isolating from lymph a bacillus and three species of micrococci. 
One of the micrococci formed an orange growth when cultivated, 
and was regai'ded as the specific micro-organism, as it caused sub- 
cutaneous tumefaction, and, it is said, some degree of immunity. 

Brown cultivated a number of oi-ganisms which on inoculation 
only produced local irritation. Intravenous injection px'oduced death 
from septicsemia in one case in thirty-six hours. 

Arloihg isolated four different organisms, including a bacillus 
which was named Pneumo-baoillus liquefaciens bovis. Later, he 
prepared a fluid from broth-cultures, pnewmo-hacillin, which pro- 
•duced a more marked rise in temperature in animals suffering from 
pleuro-pneumonia than in healthy animals, and its use was 
suggested as an aid in diagnosis. Arloing named the micro- 
organisms provisionally Pneumo-bacillus liquefaciens bovis, Pneumo- 
coccus gutta cerei, Pneumococcus hchenoides, and Pneumococcus 
flavescens. 

Pneumo-bacillus liquefaciens bovis Short rods, non- 
motile ; spore-formation not observed. They rapidly liquefy gelatine, 
and form on potato a white layer, which becomes brownish and 
sometimes greenish. According to Arloing pure-cultures produce 
in the ox, when injected subcutaneously or in the lung, the same 
lesions which are produced by virulent lymph. Guinea-pigs and 
rabbits are slightly susceptible, dogs are immune. 



INFECTIOUS PLEURO-PNEUMONIA. 243 

Nocard does not accept Arloing's conclusions, and expresses the 
opinion that the virus is particulate, but is not due to any micro- 
organism which can , be detected or cultivated by the methods 
at present adopted. In the opinion of the author, who has 
also examined the micro-organisms in pleuro-pneumonia, it is 
fuUy justifiable to regard the nature of the contagium as 
unknown. 

Preventive Inoculation. — In 1852 Willems introduced inocula- 
tion. The liquid from the lungs of an animal with pleuro-pneumonia, 
which had recently died, was inoculated in the extremity of the tail 
by a puncture with a lancet. Swelling occurred at the seat of inocu- 
lation, and on recovery the animals were believed to be protected. 
A Dutch Commission reported that the inoculation gave a temporary 
protection. A Belgian Commission in the following year reported 
that the phenomena of inoculation could be produced several times 
in succession in the same animal, and that it was not a certain 
preventive. A French Commission in 1854 concluded that a power 
of resisting infection was given, but the period was undetermined. • 
Protective inoculation continued to be employed, and, various modi- 
fications of the method were introduced. Threads soaked in lymph 
were inoculated, or the lymph subcutaneously or intravenously 
injected. 

The usual result of the inoculation is swelling and, in about ten 
or fourteen days, effusion of straw-coloured fluid, which is occasion- 
ally blood-stained. Gangrene may follow, involving amputation of 
the tail. Germont and Loire in Queensland adopted the method — 
which was suggested by Pasteur — of inoculating calves ,in the loose 
cellular tissue behind the shoulder. This produces intense oedema 
and a quantity of lymph. There has been much controversy with 
regard to the value of protective inoculation. 

Stamping-out System. — Brown maintains" that pleuro- 
pneumonia can be exterminated only by slaughter of the diseased 
animals, and quotes the results experienced in the Netherlands in 
support of his views. 

In 1871 slaughter for pleuro-pneumonia was commenced in the 
Netherlands. There were 6,000 cattle attacked by the disease. In 
1872 owners were compelled to slaughter not only diseased cattle, 
but those which had been in contact with them, unless inoculated, 
and the attacks were, in consequence, reduced to 4,000. In 1873 it 
was forbidden to move cattle out of infected districts, and the attacks 
were reduced to 2,479. In 1876 slaughter of the whole herd was 
decreed, and during the first year of this heroic system the cases fell 



244 



INFECTIVE DISEASES. 



from 2,227 in 1875, to 1,723 in 1876, to 951 in 1877, to 698 in 
1878, to 157 in 1879, and to 48 in 1880. 

In England the Pleuro-pneumonia Act came into force on 
September 1st, 1890. Notification was to be given by the owner 
to a police constable of the district, who was required to transmit 
the information to the Local Authority and also to the Board of 
Agriculture. An inspector, with the aid of the veterinary surgeon, 
arranged for the slaughter of the suspected animal, and, if the 
disease proved to be pleuro-pneumonia, of the rest of the herd. The 
results are shown in the following table : — 



Tears, 


Numlier 

of 
Infected 
Counties. 


Number 

of 

Fresh 

Outbreaks. 


Number 

of 

Cattle 

Attacked. 


Diseased Cattle. 


Healthy 

Cattle in 

contact 

slaughtered. 


Cattle 
slaughtered 
as suspected, 
but found 
free from 
Pleuro- 
pneumonia. 


Killed. 


Died. 


1890 
1891 
1892 
1893 
1894 


36 

27 

10 

4 

2 


465 

192 

35 

9 

2 


2,057 

778 

134 

30 

15 


2,022 

778 

134 

30 

15 


37 

_ 


11,301 

9,491 

3,477 

1,157 

.391 


232 

188 
86 
41 



Thus the number of cases was reduced from 2,057 in 1890 to 
15 in 1894. 

A departmental committee appointed in 1892 to inquire into 
pleuro-pneumonia and tuberculosis, came to the following conclusions 
with regard to pleuro-pneumonia : — 

(1) That the system of eompulsory slaughter be applied not only to 
all diseased cattle, but also to all cattle which have been in association 
with them, or otherwise in any manner exposed to the infection of the 
disease. 

(2) Compulsory slaughter should be accompanied by supplementary 
measures, such as restrictions on the movement and sale of cattle within ,^ 
or coming from, infected districts. 

(3) Any exception to, or modification of, the system of compulsory 
slaughter, as provided in the Slaughter Order, 1888, should only be 
applicable to cattle in the dairy yards, byres, and cowsheds of large 
towns, the owners or occupiers of which may claim in writing the privi- 
lege of exemption for their cattle from immediate slaughter, on the 
following conditions :^ 



INFECTIOUS PLEURO-PNEDMONIA. 245 

(a) No head of cattle that has been brought into such dairy premises 
shall be removed therefrom, except for the purpose of immediate 
slaughter. 

(6) In the event of an outbreak of pleuro-pneumonia, all the diseased 
cattle shall be slaughtered. 

(c) All the remaining cattle on such premises where an outbreak has 
occurred shall be branded, and regularly subjected to the ther- 
mometer test ; and whenever a continuous increase of temperature, 
rising above 104°, is shown, they shall be slaughtered. 

(d) No fresh cattle shall be admitted into such premises while any 
of the cattle thus branded remain alive. 

(4) Inoculation cannot be recopimended as a means of eradicating 
pleuro-pneumonia, nor as practicable under existing conditions. Although 
it is open to owners to inoculate their cattle, it should be distinctly 
understood that that operation shall not give them any immunity from 
the regulations above suggested. 

The order at present in force is the Pleuro-Pneumonia Order 
of 1895. In addition to regulations for the movement of cattle, 
for disposal of carcasses, for markets, and for compensation for 
slaughter, the Order contains the following provisions : — 



Notification. 
Notice of Disease. 

(1) Every person having or having had in his possession or under his 
charge a head of cattle affected with or suspected of pleuro-pneumonia 
shall with all practicable speed give notice of the fact of the head of cattle 
being so affected or suspected to a, constable of the police force for the 
police area wherein the head of cattle so affected or suspected is or was. 

(2) The constable receiving such notice shall immediately transmit 
the information by telegraph to the Board of Agriculture. 

(3) The constable shall also forthwith give information of the receipt 
by him of the notice to an Inspector of the Local Authority, who shall 
forthwith report the same to the Local Authority. 

Duty of Inspector to act immediately. 

(1) An Inspector of a Local Authority on receiving in any manner 
whatsoever information of the supposed existence of pleuro-pneumonia, 
or having reasonable ground to suspect the existence of pleuro-pneumonia, 
shall proceed with all practicable speed to the place where such disease, 
according to the information received by him, exists, or is suspected to 
exist, and shall there and elsewhere put in force and discharge the powers 
and duties conferred and imposed on him as Inspector by or under the 
Act of 1894 and this Order. 

(2) The Inspector shall forthwith report to the Board of Agriculture. 



246 INFECTIVE DISEASES. 

No Movement into or out of Plewro-pneumonia Infected Place without 

Licence. 

Cattle shall not be moved into or out of an Infected Place except with 
a Movement Licence of an Inspector or officer of the Board, and such 
cattle shall not be moved except in accordance with the conditions 
contained in such Licence., 



in ama 



PleUro-pneumonia found in amtirket, Railway Station, Grazing Park, 
01' other like Place, or during Transit. 

The Inspector of the Local Authority shall cause to be seized all the 
cattle affected with pleuro-pneumonia, and also all cattle being in or on 
the market, fair, sale-yard, place of exhibition, lair, landing-place, wharf, 
railway station, common, uninclosed land, farm, field, yard, shed, park, or 
other such place as aforesaid, and shall forthwith transmit the information 
by telegraph to the Board of Agriculture. 

The Inspector of the Local Authority shall cause all such cattle so 
seized to be detained at the place where they are seized, or to be moved 
to some convenient and isolated place and there detained. 

Removal of Dung or other Things. 

It shall not be lawful for any person to send or carry, or cause to be 
sent or carried, on a railway, canal, river, or inland navigation, or in a 
coasting vessel, or on a highway or thoroughfare, any dung, fodder, or 
litter that has been in an Infected Place, or that has been in any place in 
contact with or used about a diseased or suspected head of cattle, except 
with a Licence of an Inspector or officer of the Board or of an Inspector 
of the Local Authority. 

Report to Board of Cattle that have heen in Contact with Cattle affected 
with Pleuro-pneumonia. 

Where it appears to a Local Authority that there is within their District 
any head of cattle which has been in the same field, shed, or other place, 
or in the same, herd, or otherwise in contact, with any head of cattle 
affected with pleuro-pneumonia, or otherwise exposed to the infection 
thereof, the Local Authority shall forthwith report the facts of the case 
to the Board of Agriculture. 

Disinfection. 

An Inspector or officer of the Board may cause or require any shed or 
other place which has been used for a head of cattle while affected with 
or suspected of pleuro-pneumonia, and any utensil, pen, hurdle, or other 
thing used for or about such head of cattle, to be cleansed and disinfected 
to his satisfaction. 

Occiipiers to give Facilities for Cleansing. 

(1) The owiler and occupier and person in charge of any shed or other 
place which has been used for any head of cittle while affected with or 
suspected of pleuro-pneumonia shall give all reasonable facilities to an 



INFLUENZA. 247 

Inspector or officer of the Board for the cleansing and disinfection of such 
place, and of any utensils, pens, hurdles, or other things used for or about 
such cattle. 

(2) Any person failing to comply with the provisions of this Article 
shall be deemed guilty of an offence against the Act of 1894. 

Prohibition to Expose or Move Diseased or Suspected Cattle. 
(1) It shall not be lawful for any person — 
(tt) To expose a diseased or suspected head of cattle in a market or 

fair, or in a sale-yard, or other public or private place where cattle 

are commonly exposed for sale ; or 
(J) To place a diseased or suspected head of cattle in a lair or other 

place adjacent to or connected with a market or a fair, or where 

cattle are commonly placed before exposure for sale ; or 

(c) To send or carry, or cause to be sent or carried, a diseased or 
suspected head of cattle on a railway, canal, river, or inland 
navigation, or in a coasting vessel ; or 

(d) To carry, lead, or drive, or cause to be carried, led, or driven, 
a diseased or suspected head of cattle on a highway or thorough- 
fare : or 

(e) To place or keep a diseased or suspected head of cattle on common 
or uninclosed land, or in a field or place insufficiently fenced, or 
in a field adjoining a highway unless that field is so fenced or 
situate that cattle therein cannot in any manner come in contact 
with cattle passing along that highway or grazing on the sides 
thereof ; or 

(/) To graze a diseased or suspected head of cattle on pasture being 
on the sides of a highway ; or 

((/) To allow a diseased or suspected head of cattle to stray on a 
highway or thoroughfare or on the sides thereof or on common 
or uninclosed land, or in a field or place insufficiently fenced. 

Influenza. 

Iniluenza is an infectious disease characterised by a catarrh of 
the respiratory or the gastric mucous membrane, accompanied by 
great prostration and mental depression, and frequently ending 
fatally by pneumonic complication. One attack is not protective. 
The disease has occurred in the form of great epidemics, like the 
pandemic of 1890, which is said to have started from Bokhara, and 
travelled to St. Petersburg, Berlin, Paris, and London, whence it 
spread all over this countrj'. The incubation period is extremely 
short, only a few hoiirs, so tliat numbers are attacked almost 
simultaneously. The occurrence of cases in succession in a family, 
the importation of the disease by an infected person, and the escape 
of persons in completely isolated localities, point to the existence 
of a living contagiiim. Pfeiffer claims to have identified it with a 



248 INFECTIVE DISEASES. 

bacillus which was found by him in the purulent bronchial secretion, 
and, by Canon, in the blood. 

Bacillus of Influenza. — Very small rods, singly or in leptothrix 
filaments. They stain with the aniline dyes, but not by Gram's 
method. They are non-motile and aerobic ; they do not grow in 
gelatine at the temperature of the room. On glycerine-agar very 
small transparent drop-like colonies develop in about twenty-four 
hours. In broth there is only a very scanty growth of whitish 
particles on the surface, which subside and form a woolly deposit. 
They are found especially in the bronchial secretion, and only in 
cases of influenza. Canon obtained them by puncturing the finger, 



% 



/ '3 












Fig. 121. — B.vcillus oj;' Iki'LUENZ.\. 
From a culture on gelatine, x 1000. (Itzeeott and Niemann. ) 

and allowing a few drops of the blood to fall upon the surface of 
glycerine-agar in a Petri's dish. The organism will retain its 
vitality for fourteen days in sputum, but is quickly detroyed by 
drj'ing. It is said that by applying the bacillus to the nasal 
mucous membrane in monkeys, symptoms similar to influenza were 
produced. 

Method of Staining. 

To stain the bacilli use Neelsen's solution or Lofiler's methylene- 
blue ; or the following method : — 

Canon's method. — Spread the blood on cover- glasses, allow them 
to dry, immerse for five minutes in absolute alcohol, and stain in the 
following solution : — 



INFLUENZA. . 249 

Aqueous solution of methylene- blue^^rong, 40 parts; ^ per 
cent, solution of eosin in 70 per cent, aldohol, 20 parts ; distilled 
water, 40 parts. 

Float the cover-glasses from three to six hours in a capsule 
placed in the incubator at 37° C, wash with water, and dry and 
mount in balsam. The red corpuscles will be stained pink, and 
the leucocytes, with the bacilli in them, blue. 




Fig. 122. — Bacillus of Influenza. 

From a cultivation showing filaments composed of long and short rods, 

oooci-f orms and irregular elements, x 1200. 

Equine Influenza. 

Equine influenza, or " pink-eye," has been noticed to be prevalent 
at the same time as epidemics of influenza in man, but there does 
not appear to be any evidence of intercommunicability or of any 
relation between the two diseases. 



CHAPTER XVIII. 

ORIENTAL PLAGUE. — EELAPSIN^ FEVER. — TYPHUS FEVER. — YELLOW 

FEVER. 

The Plague. 

The plague is a highly infectious disease, having its origin in 
putrefaction and filth, in tropical climates. The virus in its effects 
resembles that of typhus. The period of incubation varies from a 
few hours to a week. The disease produces high temperature and 
decomposition of the blood, and dark hiemorrhagic patches appear 
on the skin, but there is no eruption. Lymphatic inflammation and 
buboes almost invariably occur. The virus is intensified by warmth 
and overcrowding in houses, and dissipated by exposure to fresh air. 

When the plague occurred in this country it was recognised as a 
foreign pestilence from the East, and once imported it was fostered 
and intensified in virulence wherever there was filth, putrefaction, 
and overcrowding. The disease, like the small-pox, was communicated 
from one person to another. If a case occurred in a house other 
inmates were liable to suffer from the disease, while visitors to the 
house ran a similar but less risk. There was a good deal of variation 
both in the infectivity of the virus and in the susceptibility of 
individuals, so that one contemporary writer remarked that " no 
one can account for how it comes to pass that some persons shall 
receive the infection and others not." 

Medical men were credited with enjoying an extraordinary degree 
of immunity, though there were members of the medical profession 
who undoubtedly died of the plague. This tradition has been 
supported, to a certain extent, by the experience of the plague 
in modern times. In the epidemic in Egypt, in 1835, of the ten 
French physicians engaged there, only one died ; and while those 
who buried the victims of the plague were liable to suffer from 
it, and many did so, yet the medical men made more than one 
hundred post-mortem examinations without any death resulting. 

250 



THE PLAGUE. 251 

The clothes and coverings of the infected often spread the disease, 
and yet there are numerous examples of persons who without having 
adopted any method of protection occupied the beds of plague patients 
without contracting the malady. 

The plague is transmissible from one country to another 
by sea. An infected ship becomes an infective centre as readily 
as an infected house. Once imported, whether by land or sea, the 
virus from infected persons or merchandise spreads wherever the 
environment is favourable for its development and extension. 

Old London afforded in every way a suitable environment for 
the plague. The situation of the city was unhealthy, and the old 
town ditch was a receptacle for all kinds of filth. The houses 
projected over the roadway, and the streets were saturated with 
constant contributions of slops and of excrement from animals and 
human beings. The houses were often filthy and unventilated, and 
the floors strewn with rushes, which were seldom changed. Erasmus 
goes so far as to say that the rushes were piled the new upon the 
old for twenty years, and were fouled with spillings of beer, 
fragments of fish, expectoration, vomit, excrement, and urine. 
Another very striking insanitary feature of Old London was the 
overcrowded state of the graveyards, which was well calculated to 
predispose to pestilence, if not actually to produce it. The burials 
were so frequent in St. Paul's Churchyard that a new grave could 
scarcely be dug without bodies being exposed in all stages of 
putrefaction. 

In 1894 the plague broke out in China, with all the symptoms 
of the fatal bubonic pest of Old London. The disease was confined 
to the poorest classes and the most overcrowded and most filthy 
localities. In Canton the deaths exceeded one hundred thousand, 
and in Hong-Kong numbered about ten thousand. The disease 
was contagious, and mainly diffused by personal contact. Death 
occurred, as a rule, in from twenty-four hours to five days. The 
English and European community escaped, with the exception of a 
very few out of a large number, mostly soldiers, employed in cleansing 
the houses. The disease was a specific fever, intensely fatal, accom- 
panied by high temperature, cerebral congestion, delirium, and the 
formation of buboes. The buboes consisted of exquisitely painful and 
swollen lymphatic glands. All the glands, in some cases, were affected. 

According to Cantlie the glandular swelling when first recognised 
was almond-shaped in the inguinal region, and globular in other 
regions, with peri-glandular oedema. The swelling rapidly increased 
in size, becoming softer, less definite in outline, and less tender, until 



252 INFECTIVE DISEASES. 

by the end of five or six days it consisted of an elevated mass, doughy 
to the touch, almost circular, with a diameter of six inches. The 
skin over the swelUng was Uvid and dimpled. The swelKng was in 
some cases due to purulent effusion, but more frequently on incision 
there was only an escape of sero-sanguineous fluid. The cervical 
glands in very severe cases sometimes attained an enormous size. 

Three out of seven Japanese medical men were attacked and one 
died, but none out of eleven English doctors, though they were equally 
exposed to infection. Of eight Englishmen attacked seven were among 
the soldiers employed, and only two died. No nurses or attendants 
on the sick were attacked. The virus appeared to be intimately 
connected with filth in the soil. According to the Chinese, rats, 
poultry, goats, sheep, cows, and buffaloes are susceptible. In the 
houses and hotels dead rats were found in great numbers : it was said 
that they emerged from their haunts in sewers and drains, appeared 
to be dazed, and limped about, owing to the formation of buboes in 
their hind legs. Rats, mice, and guinea-pigs inoculated with virus 
from a human lymphatic gland died with development of buboes. 
It appears to be clearly proved that rats suffer from the plague in 
common with man, and it has also been suggested that they may 
serve to spread the disease. Bacilli were found in human blood and 
in the swollen lymphatic glands by Kitasato, and independently by 
Yersin. 

Bacillus of Plague. — Short rods with rounded ends. They 
stain with aniline dyes, but not by Gram's method. The stain 
collects at the ends of the rods, leaving a clear space in the middle. 
Sometimes the rods are surrounded by a capsule. They are found 
in abundance in the buboes, and in small numbers in the blood in 
very serious and rapidly fatal cases. 

Material from the buboes inoculated on agar gives rise to white 
transparent colonies, which have an iridescent edge when examined 
by reflected light. 

The bacilli grow more readily on glycerine-agar and on soUdified 
serum. In broth cultures the liquid remains clear, and a fiocculent 
deposit forms on the sides and at the bottom of the vessel. 

An alkahne solution of peptone 2 per cent., with from 1 to 2 per 
cent, of gelatine, is the best nutrient medium. In cultures the 
bacilli develop chains of short rods and well-marked involution 
forms. Swollen and degenerated forms are found most abundantly 
in old cultures, and stain with difficulty. 

Mice, rats, and guinea-pigs, inoculated with bubonic tissue, die in 
a few days, numerous bacilU being found in the lymphatic glands, 



THE PLAGUE. 253 

spleen, and blood. Guinea-pigs die in from two to five days, and 
mice in one to three days. 

In guinea-pigs after some hours there is oedema at the seat of 
inoculation, and the lymphatic glands are swollen. After twenty - 
four hours the animal refuses to eat, has a staring coat, and after 
a time suddenly falls on its side, and is attacked by convulsions, 
which become more and more frequent until death occurs. 

After death the seat of inoculation is found to be extensively 
cedematous, and the neighbouring lymphatic glands enlarged and 
filled with bacilli. The intestine is often congested, and the liver is 




Fig. 123. — Bacilli or Pl.^gue and Phagocytes, x 800. 
From human lymphatic gland. (Aoyama.) 

congested and enlarged. In less acute cases an abscess of the 
abdominal wall occasionally resvilts. 

The bacilli are sometimes found in the pleural and peritoneal 
exudation. The liver and spleen also contain many bacilli. Those 
in the blood are a little longer than those in the lymphatic glands. 

Inoculations can readily be made from guinea-pig to guinea- 
pig by using the pulp of the spleen, or the blood. Cultures lose then- 
virulence gradually, but the virus can be intensified by successive 
inoculations in animals. The disease is infectious to mice as 
well as inoculable. Pigeons are insusceptible. Rats and flies may 
convey the bacilli. 

According to Aoyama, the bacilli found in the blood of plague 



254 INFECTIVE DISEASES. 

patients and in the buboes are not identical. The bacilli in the 
buboes are different in form, and they stain by Gram's method. 

There is no doubt that the micro-organism which was found in 
blood is very similar to the bacillus of fowl cholera, and it is quite 
possible that the so-called plague bacillus is really identical with the 
bacillus of haemorrhagic septicaemia, and that the real nature of the 
contagium in bubonic plague is unknown. 

Protective Inoculation. — Yersin, Calmette, and Borrell claim 
not only to have produced immunity, but to have cured animals 
after infection. Cultures on agar heated to 58.° 0. for an hour 
were attenuated, and rabbits after intravenous or subcutaneous 
inoculation were protected against virulent cultures. The serum 
of immunised rabbits was capable of protecting from subsequent 
virulent cultures, and neutralising the effect of a previous inoculation 
of a virulent culture. A horse was inoculated with cultures which 
killed mice in two days, and after six weeks a serum was obtained 
which produced immunity in mice and guinea-pigs. 

Stamping-out System. — It is not until the sixteenth century 
that we hear of preventive measures being attempted in England, 
and then they appear to have been adopted only when an outbreak 
threatened to be very serious. 

Early in the sixteenth century all those who had the plague in their 
houses were ordered to put up wisps, and to carry white rods in their 
hands. 

In 1643 the Plague Order of Henry VIII. was issued. In place of 
wisps the sign of the cross was to be made on every infected house, and 
to remain there for forty days. Persons afHicted with the disease were 
to refrain, if possible, from going out of doors, or for :^orty days to carry 
a white rod in the hand. All straw from the infected houses was to be 
carried into the fields and burnt. Churchwardens were directed to keep 
beggars out of churches on holy days, and all streets and lanes were to 
be cleansed. 

In 1547 the means of notification was a blue cross with the addition 
of the inscription Lord have mtrcy wpon us. Later on the colour of the 
cross was changed to red. 

With the outburst of the plague in 1563, came an attempt to enforce 
a terrible system of compulsorily shutting up infected families. The 
doors and windows in such houses were to be closed, and no inmates were 
to leave the premises and no visitors to be allowed for forty days. No 
better incubator on a large scale could possibly have been devised for 
both breeding and intensifying the virulence of the plague bacillus, or 
whatever may be the contagium vivum of this disease. 

This compulsory shutting up of the sick with the healthy amounted 
to a compulsory infection of many of the unfortunate inmates who might 



THE PLAGUE. 255 

■otherwise have escaped, and very naturally the order was frequently 
infringed. 

In 1568 the Lord Mayor of London drew up instructions for the 
Aldermen for dealing with the plague. It was enacted that constables 
and officers should search out infected houses and report to the authorities. 
In other words, that there should be notification by the police. All infected 
houses were to be shut up, and no person to be allowed to come out for 
twenty days. All bedding and clothes used by the victims were to be 
destroyed. 

At Westminster these instructions were to be enforced under a penalty 
■of seven days in the stocks, with imprisonment to follow, making in all 
a punishment of forty days. 

In 1681 the Lord Mayor transferred notification from the constables 
to searchers. Two honest and discreet matrons in every parish were to 
■search the body of every such person that happened to die in the parish. 
They were ordered to make a true report to the clerk of the parish, and 
the said clerk had to report to the wardens of the parish. For failing 
to notify, the penalty was an exemplary term of imprisonment. The 
searchers were of course likely to be offered heavy bribes by the people 
to suppress reports, owing to their anxiety to avoid the shutting up of 
infected houses. 

The continued prevalence of the plague led to the publication, in 
1593, of a book by Simon KeUwaye. One chapter "teacheth what 
orders magistrates and rulers of citties and towns should cause to be 
observed," which included among other regulations that no dunghills 
were to be allowed near the city, and the streets were to be watered 
and cleansed. 

No surgeons or barbers who let blood were to cast the same into the 
streets. All those visiting and attending the sick to carry something in 
■their hand to be known from other people ; and if the infection were in 
few places, all the people were to be kept in their houses during the time 
of their visitation, and when this was over, aU clothes, bedding, and 
other such things used upon the sick, were to be burnt. 

In 1603 Thomas Lodge recommended that discreet and skilful men 
should be appointed in every parish to notify sipkness to the authorities, 
and so cause them to be visited by expert physicians, and that such as 
were sick should be separated from the whole, and that isolation hospitals 
should be built outside the City in separate and unfrequented places. 

In 1666 the Great Plague of London occurred, and was attributed by 
some to the importation of an infected bale of silks from the Levant. 

According to Hodges the disease stayed among the common people, and 
hence was called The Poor's Plague. He criticised the system of shutting 
up infected houses, and strongly recommended that those who were 
untouched in infected houses should receive " accommodation outside the 
city." The sick were to be removed to convenient apartments provided 
on purpose for them. To quote his own words, " Timely separation of 
the infected from the well is absolutely necessary to be done." 

For the purification of houses his directions were to place " a chafing 



256 INFECTIVE DISEASES. 

dish in the middle of the room, where proper things were burnt and 
exhaled all around." The use of sulphur and quicklime was mentioned.* 

Preyentive measures were drawn up and published by the Lord Mayor 
and Aldermen. Examiners in Health, watchmen, and searchers were 
appointed. Surgeons were selected to assist the searchers in making their 
reports, and a fee of twelve pence was allowed for every case. The disease 
was immediately notified to the Examiner of Health. Rules for disinfec- 
tion were made, and every infected house was shut up, and no one removed 
except to a pest-house or tent. Orders were issued for cleaning and sweep- 
ing the streets. Hackney coaches were not to be used after conveying 
patients to the pest-house until they had been well aired. Regulations 
were also made dealing with loose persons, assemblies, and drinking taverns. 

The plague was scarcely over before the whole city was in flames. A 
new city speedily rose upon the ashes of Old London. A few sporadic 
cases of plague are given in the London Bills of Mortality down to 1679, 
when they finally ceased. London was sterilised by the great fire. "Great 
as this calamity was," wrote Thomas Pennant, " yet it proved the provi- 
dential cause of putting a stop to one of far more tremendous nature. 
The plague, which, for a series of ages, had, with very short intervals, 
visited our capital in its most dreadful forms, never appeared there again 
after the rebuilding of the city in a more open and airy manner ; which 
removed several nuisances, which if not the origin of a plague, was 
assuredly one great pabulum, when it had seized our streets." 

In the years 1720-22 there was a terrible outburst of plague in France. 
It was attributed at Marseilles to importation by a ship from Syria. This 
caused a panic in England, and the Lords Justices considered it necessary 
for the public safety that measures should be taken to defend the country 
from a fresh invasion of this disease. Dr. Richard Mead was entrusted 
with drawing up the required recommendations. Mead laid it down as 
an essential doctrine that the plague was not native to this country, and 
therefore the first thing was to prevent importation, and if such a misfor- 
tune occurred, it was to be prevented from sp'reading. How was this to 
be accomplished ? Briefly stated, his system was as follows : Lazarettoes 
were to be provided for the reception of infected men and merchandise. 
The healthy were to change their clothes and to be kept in quarantine, and 
the sick were to be kept remote from the healthy and their clothes 
destroyed. 

If, through a miscarriage in the public care, by the neglect of officers 
or otherwise, the disease was imported, then " the civil magistrates were 
to make it as much for the interest of the afiiicted families to discover 

* During outbreaks of the plague amulets were extremely popular. Walnuts 
filled with mercury, pieces of cloth coated with arsenic, and arsenical cakes,, 
were very generally worn. The College of Physicians recommended issues on 
the arms and legs. Dr. Hodges wrote, that the more of the ulcers that were 
made the better, although their largeness answered as well as more in number. 
If two issues were preferred, it was recommended to make one on the left arm 
and the other on the opposite leg. A somewhat similar plan was adopted in. 
Ciroassia by small-pox inoculators. 



RELAPSING FEVER. 257 

their misfortune, as it was when a house was on fire, to call in the 
assistance of the neighbourhood." The shutting up of infected houses 
was condemned in the strongest terms, and a system of notification and 
isolation was proposed on the lines originally suggested by Dr. Hodges. 

1. A Council of Health was to be established, and entrusted with such 
powers as might enable them to see all their orders executed with im- 
partial justice. 

2. Notification. — The ignorant old women employed as searchers were 
to be replaced by understanding and diligent men, who were to report 
cases immediately to the Council of Health. 

3. Isolation. — Physicians were at once to be despatched to visit the 
suspected cases, and when the suspicion of plague was confirmed, all the 
families in which the sickness occurred were to be isolated. The sick 
were to be separated from the sound, and isolation houses to be provided 
three or four miles out of the town. 

The removal of the sick was to be made at night, so as to avoid the 
danger of spreading infection, and all possible care was to be taken to 
provide such means of conveyance for the sick that they might receive no 
injury. The poor were to be isolated in houses provided for the purpose, 
but the rich were to be allowed to be in their own homes provided that 
care was taken to separate the healthy from the sick, and no pains were 
to be spared to provide clean and airy apartments. All expenses were to 
be paid by the public, and a reward was to be given to the person who 
made the first discovery of infection in any place. 

Mead further pointed out that general sanitation must be carefully 
attended to. OfiBcers were to see that the streets were washed and kept 
clean from filth, carrion, and all manner of nuisances. Beggars and idle 
persons were to be taken up, and such miserable objects as were fit 
neither for the hospitals nor for the workhouses, were to be provided for 
in an establishment for incurables. Houses also were to be kept clean, 
and sulphur was to be used as a disinfectant. 

After centuries of experience we have learnt that the necessary 
conditions for avoiding the plague are more accurate knowledge on 
the part of the profession and the public of the way in which the 
disease spreads, and the adoption of sanitary precautions, which must 
include personal cleanliness, sanitary dwelKngs, absence of overcrowrd- 
ing, immediate notification, prompt separation of the sick from the 
healthy, disinfection of infected dwellings, destruction of infected 
clothing, and extra-mural burial or, better still, cremation. It was 
because the very reverse of these sanitary conditions existed that 
the virus of the plague found a suitable environment in Old London 
and in recent times in Hong-Kong. 

Relapsing Fevbe. 
Relapsing or famine fever is a contagious disease producing a 
state of high fever lasting about seven days, followed by apparent 

17 



258 



INFECTIVE DISEASES. 



recovevy, and in about fourteen days by another attack of fever, 
which may be repeated after another week. 

Starvation, in association with overcrowding and filth, is intimately 
connected with the causation of the disease. The subjec^ts of the 
disease contaminate the air around them, and the virus is principally 
conveyed by tramps raid dirty people. 

Obermeier discovered spirilla in the blood during the paroxysms 
of fever. The constant occurrence of the spirillum in relajjsing 
fever, and the fact of its not being found in any other conditions, 
render it ^ery probable that it is the cause of the disease. 



#^# "^^ ^ .\y] 







""^IK '^ 





Fig. 124. — SpiKn.LUM Obekmeieki ik Blood op Monkey Inoculated with 
Spirilla aftek Removal of the Spleen (Soudakewitch). 



Spirillum Obermeieri (Sjm-ochceta Obermeieri, Cohn). — 
Threads similar to the Spirillum plicatile. In length they are mostly 
16 to 40 fjL, with regular screw-curves. They move very rapidly, 
and exhibit peculiai- wave-Uke undulations. They are absent from 
the blood during the non-febrile intervals, but are found in the 
interior of leucocj'tes in the spleen. In blood serum and 50 per 
cent, salt solution, they preserve their movements. In cover-glass 
preparations they are readily stained by any of the aniline dyes, 
and in sections, by preference, -with Bismarck brown. They are not 



TYPHUS FEVER. — YELLOW FEVER. 259 

found in the urine, sweat, or saliva. They have not been cultivated 
artificially on any nutrient media. Monkeys have been successfully 
inoculated with blood containing the spirilla by Koch, Carter and 
Soudakewitch ; and Koch found the spirilla in the vessels of the 
brain, liver, and kidneys, after death. According to Soudakewitch 
a fatal result is produced in monkeys if the spleen is removed, 
and the spirilla are found in great numbers in the blood ; but if 
the spleen is not excised the spirilla rapidly disappear, and recovery 
follows. Munch and Motschutkowsky transferred blood containing 
the spirilla to healthy persons, and produced typical relapsing fever. 

Typhus Fever. 

Typhus fever is a highly contagious disease, which lasts for two 
or three weeks, and produces a measly eruption. Like the plague, 
it is intimately associated with overcrowding and filth, and is liable to 
occur where these conditions exist in cities, in armies, and in prisons. 
The virus produces profound changes in the blood, and after death 
the internal organs are found to be congested, especially the lungs, 
which are very friable. The spleen is softened and often enlarged, 
and the' blood is dark and imperfectly coagulated. 

The virus is dissipated by fresh air. It is given off by the 
breath of patients, and possibly from the skin. It clings to the 
clothes of patients, and the disease may be conveyed by their agency. 
One attack, as a rule, confers immunity. Some persons are naturally 
insusceptible, failing to contx'act the disease though daily exposed to 
it. Hlava has described a bacterium which he believes to be the 
specific micro-organism. Thoinot and Calmette found the same 
bacterium with others, but there was no particular micro-organism 
constantly present. There can be little' doubt that the nature of 
the contagium is unknown. 

Stamping-out System. — Sanitary precautions, and especially 
the operation of the Public Health Acts in relation to lodging- 
houses, prisons, and the better housing of the working classes, have 
been instrumental in almost completely stamping out the disease in 
this country. 

Yellow Fever. 

Yellow fever is a disease of tropical climates, characterised by 
abdominal tenderness, hemorrhagic vomiting (black- vomit), and 
jaundice. The disease may end fatally, or recovery occur in about 
two or three weeks. It is especially prevalent in the West Indies 
and in parts of North and South America. 



260 INFECTIVE DISEASES. 

The virus may be conveyed by infected ships, and has in this 
way made its appearance at British and French seaport towns. 
The disease is generally believed to be contagious, but the source 
■of the virus is not known. According to Sternberg the virus is not 
conveyed by water, but spreads where there is overcrowding and 
filth. 

Bacteria in Yellow Fever. — Freire asserts that there is a 
specific micrococcus in yellow fever which can be grown on all 
ordinary nutrient media, and that cultures can be used for protective 
inoculation with satisfactory results. Carmonay Valle also claims to 
have discovered the contagium ; but Sternberg, who has carried on 
investigations extending over several years, maintains that there is no 
characteristic micro-organism present in the blood or in the tissues 
after death. Aerobic and anaerobic cultures were made from the 
blood, Kver, kidney, urine, stomach, and intestines. The liver was 
found to contain after death a number of bacilli, most frequently 
Bacillus coh communis and Bacillus cadaveris. Blood or fresh 
liver does not produce any disease in rabbits or guinea-pigs, but 
liver tissue kept for forty-eight hours and inoculated subcutaneously 
in guinea-pigs is extremely pathogenic. Similar results occur after 
inoculation of healthy liver which has been kept in the same way. 
We may conclude from these experiments that the nature of the 
■contagium is unknown. 

Stamping-out System. — Sternberg states that there are many 
facts relating to the origin and extension of yellow fever epidemics 
which support the theory that the virus is present in the evacua- 
tions, and that accumulations of fsecal matter and of organic 
material of animal origin furnish in certain climates a suitable soil 
for the development of the contagium. According to this view the 
■evacuations should be thoroughly disinfected, and with other sanitary 
precautions and efiicient quarantine at seaports, the disease may 
be stamped out, and the danger of importation from tlie natural 
iome of the disease reduced to a minimum. 



CHAPTER XIX. 

SCARLET FEVER. MEASLES. 

Scarlet Fever. 

Scarlet Fever is a highly contagious disease peculiar to man. It 
produces inflammation of the tonsils and adjoining parts, fever, and 
a general punctiform eruption. The period of incubation is about, 
a week, and the rash usually appears on the second day. In some 
cases the disease manifests itself in an extremely mild form, known 
as latent scarlet fever, in which there is only a slight febrile attack, 
or a, mild sore throat, with very little or no rash. Many cases 
would not be recognisable as such if they were not capable of 
conveying scarlet fever, or unless other cases followed or occurred 
simultaneously which were undoubtedly typical cases of the disease. 
The occurrence of such cases in the early history of an epidemic 
often causes the greatest difficulty in tracing the origin of the 
outbreak, and indeed in some cases renders it quite impossible to 
do so. 

The virus is given off by the skin, in desquamation, and possibly 
by the urine. It maintains its vitahty in clothing for months, and 
sometimes longer. It may also be conveyed by the hands of the 
physician to women during parturition. The disease may be 
transferred by subcutaneously inoculating persons, who have not 
previously contracted scarlet fever, with virus obtained by puncturing 
the eruption on the skin. 

After death the internal organs appear to the naked eye more or 
less healthy. The liver is soft, the kidneys are congested, the ileum 
is inflamed, and Peyer's patches enlarged and congested ; but these 
conditions are also prodiiced by other causes. There are inflam- 
matory changes in the lymphatic follicles of the tonsils, and the 
larynx and trachea. Other morbid lesions, especially in the kidneys, 
are associated with the sequelae and comphcations, and though 
commonly occurring in scarlet fever are also found in other diseases. 

261 



262 INFECTIVE DISEASES. 

These changes appear to be due to the poison which is in the blood, 
and is excreted by the kidneys. The epithelium is in a state of 
cloudy swelhng, a condition found in other febrile diseases and in 
septic poisoning. 

Bacteria in Scarlet Fever. — The occurrence of micro-organ- 
isms in cases of scarlet fever has been observed by several investi- 
gators — Coze and Feltz, Orooke, Loffler, Babfes, Heubner and Bahrdt, 
and notably by Frankel and Freudenberg, and more recently by 
Klein, the author, Raskin, and others. 

Ooze and Feltz found cocci in the blood, and Orooke, in cases of 
scarlet fever with severely affected throat, found bacilli, cocci, and 
streptococci in the organs of the throat, and cocci in the internal 
organs. Orooke left it an open question whether these cocci were 
the specific organisms of scarlet fever, or were to be regarded as 
diphtheritic or septic associates. He inclined, for clinical reasons, 
to the latter view. 

Loffler, in cases of scarlatinal diphtheria, found the same chain - 
forming micrococcus which he had found in typical diphtheria. 

Babes was able constantly to prove the presence of a strepto- 
coccus in inflammatory jjroducts secondary to scarlatina. 

Heubner and Bahrdt, in a fatal case of scarlet fever in a boy, 
complicated with suppuration of the finger and knee-joints, and 
with pericarditis, found a streptococcus identical in form with Strepto- 
coccus pyogenes, but cultivations were not made. The secondary 
infection started from diphtheritically affected tonsils, which were 
followed by retro-pharyngeal abscesses. 

Frankel and Freudenberg examined, for micro-organisms, three 
cases of scarlatina with well-marked affection of the throat. In all 
three cases they obtained cultivations of cocci from the submaxillary 
lymphatic glands, spleen, liver, and kidney. These cocci could not 
be distinguished from Streptococcus pyogenes derived from pus, nor 
from the undoubtedly identical streptococcus which one of them 
(A. Frankel) had repeatedly cultivated in large numbers from 
puerperal affections. In two of the cases Streptococcus pyogenes 
was the only organism present, and in all three cases it was far in 
excess of other colonies which developed. The organisms were also 
found in sections of the organs by microscopical examination. 
Frankel and Freudenberg could in no way distinguish the strepto- 
coccus in scarlatina from the streptococcus in pyaemia and septi- 
caemia. The identity of this streptococcus with Streptococcus pyogenes 
and Streptococcus puerperahs was established by comparison of their 
macroscopical and microscopical appearances in cultivations on 



SCARLET FEVER. 



263 



nutrient agar-agar, nutrient gelatine, and in brotli, botli at the 
ordinary and at higher tempei-atures, and also by experiments on 
animals. They concluded that it conld be stated with certainty 
that the organi-sms in question did not stand in causal relation to 
scarlet fever. They considered that special methods of microscopical 
and biological research were apparently needed for demonstrating 
the true scarlet fever contagium, which probably was especially 
present in the skin. They considered that the presence of the 





Fig. 125. — Pure-Cultivatioxs of Streptococcus Pvogenes. 

(a) On the surface of nutrient gelatine ; (i) In the dej^th of nutrient gelatine : 
{n] On the surface of nutrient agar. 



streptococcus was due to a secondary infection, to which the dooi' 
was opened by the lesions of the throat — a view which was sup- 
ported by the fact that the organisms were found in submaxillary 
l3rmphatic glands. They pi-eferred to use the term " secondary " 
to " complicated " oi' " combined " infection, because this expresses 
the fact that by the effect of the scarlatinal virus the soil is 
rendered suitable for this ubiquitous microbe when it has once 
gained an entrance. 

This streptococcus was found by Klein in five out of eleven cases 



264 



INFECTIVE DISEASES. 



of scarlet fever in man, twice in association with certain other 
micro-organisms, and three times alone. The micro-organisms were 
isolated by inoculating tubes of nutrient gelatine, solidified obliquely, 
by streaking the surface with blood taken from the finger, the arm, 
or the heart after death. Those cases from which the organism was 
obtained were all oases with ulcerated throat, and the culture 
experiments, from the living patient, were made on or about the day 
at which the temperature was at its maximum. 



Name of Patient. 



L F 

aged 5 

K -F — 

aged 2 



H L , 

aged 8 
(a woman), 

aged 40 
(a girl), 

aged 19 
B M , 

aged 15 
E W , 

aged 22 
R H , 



F G- — , 

aged 2^ 

E— F , 

aged 3 

E B , 

aged 20 months 



Condition of 
Tonsils. 



Severely- 
ulcerated 

Much 
ulcerated 



Ulcerated 

Much 
ulcerated 



Ulcerated 

Much 
ulcerated 
Ulcerated 



Advanced 
ulceration 



Source 

of 
Blood. 



Micio-Organisms 
Isolated. 



Finger 



Arm 



Finger 



Heart 



Streptococcus 

(Staphylococcus 
pyogenes aureus 
Liquefying micro- 
coccus 
Streptococcus 
f Staphylococcus 
\ Streptococcus 
None 



Streptococcus 
None 

Streptococcus 
None 



Death or 
Recovery. 



Ultimately 
recovered. 



. Died of 
pyaemia. 

>- Recovered. 
[Not stated.] 



Recovered. 
[Not stated.] 

?» 
Died. 



Klein regards this streptococcus as the actual cause of scarlet 
fever in man. 

The author, Raskin, Holmes, and others who have investigated this 
subject agree with the conclusions of Frankel and Freudenberg. The 
author is convinced that the streptococci in suppuration, puerperal 
septicaemia, pysemia, and septicaemia, and in certain cases of measles, 
scarlatina, and diphtheria, are identical; and from overwhelming 
evidence we are justified in concluding that — (1) The nature of 
the contagium of scarlet fever is unknown. (2) The streptococcus 
regarded by Klein as the contagium is the Streptococcus pyogenes. 



MILK-SCARLATINA. 265 

(3) This streptococcus is found, sometimes in company with Staphylo- 
coccus pyogenes aureus, as a secondary result in scarlet fever and 
many other diseases, and its exact relation to scarlet fever and its 
identity with the streptococcus from pus and puerperal fever, were 
definitely established in 1885 by Frankel and Freudenberg. 

MiLK-SCABLATINA. 

It would not be necessary to say anything further on the etiology 
of scarlet fever if the generally accepted belief, that scarlet fever is 
a disease peculiar to man, were accepted by the Medical Department 
of the Local Government Board; but the theory is officially held 
that scarlet fever is in its origin a disease of cows. Bovine scarlatina 
is supposed to be an eruptive disease of the teats, and it is maintained 
that the virus, by contaminating the milk, produces scarlet fever in 
the human subject. As this theory is very naturally accepted by 
many medical officers of health, and is mentioned in EngUsh medical 
text-books, it will be necessary to discuss this question in considerable 
detail, and especially as these opinions were promulgated in this 
country with official svipport, and have since been proved to be 
erroneous. 

The theory of the origin of the exanthemata in diseases of the 
lower animals is a very old one. The Arabians imagined that small- 
pox arose from the camel. Jenner adopted a similar theory, and 
expressed his belief that small-pox originated in the horse, being 
generated by horses suffering with " greasy " hocks. Thus Jenner 
wrote : "May not accidental circumstances have again and again 
arisen, still working new changes ujDon it, until it has acquired 
the contagious and malignant form 'imder which we now com- 
monly see it, making its devastations among vis ? and from a 
consideration of the change which the infectious matter undergoes 
from producing a disease in the cow, may we not conceive that 
many contagious diseases now prevalent amongst us may owe their 
present appearance, not to a simple, but a compound origin ? For 
example, is it difficult to imagine that measles, scarlet fever, ulcerated 
sore throats, and spotted skin, all spring from the same source, 
assuming some variety in their forms according to the nature of 
their new combinations ? " Baron informs us that this idea was 
prevalent in Jenner's mind as early as 1787. It is related that in 
that year he accompanied his nephew, George Jenner, into a stable 
to look at a horse with diseased heels, and, pointing to them, he 
remarked : " There is the source of small-pox. I have much to say 



266 INFECTIVE DISEASES. 

on that subject whicli I hope in due time to give to the world." 
And again in 1794, when writing in connection with this subject, 
he adds : " Domestication of animals has certainly provided a prolific 
source of diseases among man." 

Jenner's views were found to be incorrect, and it was shown by 
Loy and others that the grease bears no relation to cow-pox, and 
it is now known that Jenner mistook horse-pox for the disease 
known as the grease. No one at the present day supports Jenner's 
theory of small-pox in man arising from any disease of the horse. 
Indeed, the origin of small-pox from a disease of the horse was not 
upheld even by Jenner's pupil and nephew, Henry Jenner. The 
latter promulgated the idea that small-pox originated from the cow. 
He believed that small-pox, in fact, was cow-jjox intensified in its 
virulence by being passed through man. He thus expressed himself • 
" Nor may it, perhaps, be too hypothetical to suppose that the 
cow-pox may possibly be the small-pox in its original iinadulterated 
state, before it became contaminated by passing thi'ough the impure 
and scrofulous habits of human constitutions." The theory of the 
origin, in animals, of human febrile diseases was, later, advocated by 
Copland, who stated, firstly, that scarlet fever in man was originally 
a disease of the horse, and that it formerly occurred, and had recently 
occurred, epidemically as an epizootic among horses ; secondly, that 
it was communicated in comparatively modern times from horses to 
man ; thirdly, that it might be, and had been, communicated to the 
dog. But this opinion has not been accepted, for the disease called 
scarlatina in the horse is a non-infectious disease, generally attack- 
ing but one or two horses in a large stud. It neither spreads by 
contagion nor infection ; and Williams states that it is impossible to 
transmit it from the horse to -any other animal, and that many cases 
of the so-called scarlatina of the horse are in reality identical with 
purpura. 

The theory was again revived, but in another form, and has 
been adopted by the Mechcal Department of the Local Government 
Board. Owing to failure in tracing, in some oases of milk scarla- 
tink, the contamination of the milk from a human source, the 
theory was started that in such oases the disease is derived from 
the cow — that, in other words, there is a disease, scarlet fever 
in the cow, which is responsible for outbreaks of scarlet fever in 
man. 

In 1882 an epidemic of scarlatina in St. Giles and St. Pancras 
was investigated by Mr. W. H. Power for the Board. The disease 
was distributed with a milk supply from a Surrey farm. In this case 



MILK-SCARLATINA. 267 

two facts were ascertained : the one, that a cow recently come into 
milk had been suffering from some ailment from the time of her 
parturition, of • which loss of hair in patches was the most con- 
spicuous manifestation ; the other, that there existed no discoverable 
means by which the milk could have received infective quality from 
the hviman subject. 

In 1885 an outbreak of scarlet fever occurred in Marylebone in 
connection with milk from a farm at Hendon, and again Power 
failed to establish infection from any human source in any commonly 
accepted way — such, for example, as handUng of milk, or milk utensils, 
by persons carrying scarlatina infection. But on examining the 
cows with a view to ascertain any new condition pertaining to 
them, it came to hght during the inquiry that some of them, 
which had recently been introduced from Derbyshire, were suffering 
from a vesicular disease of the teats. 

At this stage Klein became associated with Power in the 
inquiry; and their belief in the existence of a disease among the 
cows on the farm capable of producing scarlatina among human 
consumers of the cow's milk, became unreserved. Klein took away 
with him samples of milk, contents of vesicles, and discharges 
from ulcers, and afterwards two of the cows were purchased and 
kept under observation. 

Dr. Cameron of Hendon has given a detailed description of the 
clinical history of this disease. He expressed his belief that it was 
a specific disease capable of being communicated to healthy cows 
by direct inoculation of the teats with virus conveyed by the milker 
from a diseased animal. 

The condition of the teats is described as follows : The teats 
became enlarged, swollen to nearly twice their natural size, and 
oedematous. On handling them there was no feeling of induration. 
Vesicles appeared on the swollen teats and upon the udder between 
or near the teats. These varied in number from two to four on a 
teat, and in size from a pea to a horsebean. The vesicle contained 
a clear fluid. The vesicles were rubbed and broken in milking, and 
left raw sores, sometimes red, in other cases pale in colour, with 
raised, ulcerated edges. Sometimes a few accessory vesicles formed 
around the margin of these ulcerated sores. After the rupture of 
the vesicle a brown scab formed, which might remain attached for 
five or six weeks, or fall oft' in ten days or a fortnight, a smaller 
one forming afterwards. A thin, watery fluid exuded from vmder 
the scab, and the sore ultimately healed. 

Cameron examined the teats of several cows five or six weeks 



268 INFECTIVE DISEASES. 

after they were attacked. The scabs then varied in size from a 
shilling to a florin ; they were about one-eighth of an inch thick in 
the centre, thinning off towards the edges. 

Some of the cows were also suffering from an eruption on the 
rump and hind quarter, consisting of patches of eczematous crusts. 
When a crust was picked off, the hair came off with it, exposing a 
raw, moist sore, the crusts and sores looking exactly like eczematous 
scabs and sores ; but this condition corresponds in description with 
eczema, the result of lingworm which is very common in young 
stock. 

In addition to his own observations, Cameron obtained infor- 
mation from the farmers, and others familiar with cows, who 
thought they recognised in the disease at the farm one stage of a 
disease which they were able to describe. Cameron thus gives an 
account of what he and his informants together would regard as 
a connected clinical history of the disease. 

He did not see the earlier symptoms, and hence these were 
of necessity learnt from other persons. The account, therefore, 
of these symptoms was to be held liable to future correction or 
modification. 

Cameron stated that he learnt this disease was capable of 
being communicated to milkers by inoculation with virus from the 
vesicles on the teats, though the milkers on the Hendon farm 
escaped. " A trusty informant received the virus into a recent 
scratch on the forefinger while milking a diseased cow. General 
weakness, malaise, and loss of appetite resulted, and after about 
four or five days a vesicle or small blister appeared on the finger. 
This broke, and several others formed on the back of the hand. 
The whole hand and fingers became swollen and inflamed, the 
inflammation extending in broad lines as far as the elbow. The 
general disturbance lasted a fortnight." 

In the course of the inquiry, Cameron adds that it was 
strongly asserted by several people, who examined the cows, that 
they were suffering from cow-pox. He, however, dismissed the 
diagnosis of cow-pox on the ground that no papule had been 
observed or subsequent formation of pustule, areola, or pitting, and 
because the vesicles were not umbilicated. These reasons given 
for dismissing the diagnosis of cow-pox at Hendon were totally 
inadequate ; a comparison having been made between the characters 
of the eruption of vaccinia as it appears on an infant's arm, instead 
of the eruption of the natural or so-called spontaneous disease on 
the teats of the cow. 



MILK-SCAELATINA. '2(^9 

Klein stated that on the teats and udders of two cows which 
he investigated there were several flat irregular ulcers, varying in 
diameter from one-quarter to three-qiiarters of an inch. Some 
ulcers were more or less circular, others extended in a longitudinal 
direction on the teat. The ulcers were covered with a brownish or 
reddish-brown scab. The animals looked thin, but not strikingly so. 
In feeding capacity, milking power, and body temperature there 
was nothing abnormal. 

Four calves were inoculated in the corium of the groin and the 
inside of the ear, with scrapings from the ulcers after removal of 
the crusts. In one, which may h& taken as an example of the result 
obtained, there was vesiculation at the margin of the spot inoculated, 
and in the centre the commencement of the formation of a crust. 
On the seventh day each sore on the ear had enlarged to about 
half an inch in breadth, and was covered in its whole extent by 
a brownish crust. On the eighteenth day they had all healed up 
and become converted into flat scars. 

To search for micro-organisms, Klein removed the crust 
from an ulcer on the teat, scraped off the most superficial layer, 
squeezed the ulcer, and made cover-glass preparations. Tubes of 
nutrient gelatine and nutrient agar-agar were also inoculated, and 
a streptococcus was isolated which in m.orphological and cultural 
characters agreed with those of Streptococcus pyogenes. 

Two calves were inoculated in the groin with the cultivated 
micro-organism. One calf died in twenty-seven days. At the 
necropsy there were found peritonitis, and hsemorrhagic spots on 
the omentum ; the liver, kidneys, and lungs were congested, and 
there were petechise under the pleura, and pericarditis. The second 
calf was killed, and at the necropsy the lungs and kidneys were 
congested, and there were hsemorrhagic patches on the spleen. 

In these cases, the post-mortem appearances and anatomical 
features recalled to Klein the lesions of scarlatina. In the 
kidney, for example, the cortex was congested, and there were 
haemorrhages, glomerulo-nephritis, and granular or opaque swelling 
of the epithelial cells and infiltration with round cells. From the 
blood of the heart the streptococcus, which had been used in the 
inoculation, was recovered. In view of this evidence it was con- 
cluded that the streptococcus was the virus of the cow disease, and 
that it produced in calves a disease very closely resembling that of 
scarlatina in man. 

Two of the cows selected from the Hendon farm were killed, 
and it was observed in one that the lungs were congested, and 



270 INFECTIVE DISEASES. 

that there were numerous adhesions by recent soft lymph between 
the lower lobes of the king and the costal pleura. In the 
liver there were several reddish streaks and patches. The spleen 
and kidneys, with the exception of slight congestion, appeared 
normal. 

Sections of the kidney showed well-marked glomerulo-nephritis 
and infiltration of the sheath of the cortical arterioles with numerous 
round cells. The epithelium of the convoluted tubules was swollen, 
opaque, and in many places dLsintegrating. 

In the other cow there was great congestion of the lungs and 
pleural adhesions ; the cortex of the kidney was congested, but its 
medulla was pale. 

On microscopic examination there was a good deal of round- 
celled infiltration in the walls of the infundibula and bronchi in the 
lung, and round the arterioles in the Iddney. 

In sections of the ulcers on the teats, the corium was found to be 
infiltrated throughout the whole extent of the ulcer with round 
cells. In the superficial layers of the stratum Malpighi, close to 
the stratum lucidum, as also in the stratum lucidum itself, there 
were numerous cavities of different sizes. These cavities lay side by 
side, the most superficial ones being covered by the stratum lucidum, 
or extending between the layers of this stratum. 

At the marginal parts the cavities, although placed side by side, 
were well separated from one another by thicker or thinner 
trabeculse, composed of epithelium, while at or near the centre of 
the ulcer these trabeculse were destroyed, the cavities had become 
confluent, and the covering layers of the cuticle having here also 
given way, their contents extended on to the free surface of the 
ulcer. Ill short, Klein states that all the anatomical details of 
the distribution and arrangement of these cavities recalled vividly 
the conditions observed in the vesicles of cow-pox. Yet as a result 
of this investigation he concluded that the cow disease at Hendon 
was bovine scarlatina, and that towards its study and supervision 
every effort ought to be directed in order to check the spread of 
scarlet fever in man. 

As a result of this conclusion, the Board of Agriculture resolved 
to have the whole subject fully investigated, and the author was 
directed to study the bacteriology and micro-pathology of this disease 
and to rejjort thereon. Professor Axe investigated the origin of 
the outbreak of the disease in the cows, and Professor M'Fadyean 
carried out an investigation into the possibility of inoculating cows 
with the virus from cases of scarlet fever in man. 



MILK-SCARLATINA. 271 



The Authoe's Investigation. 



An outbreak of an eruptive disease of the teats, alleged to be 
identical with the so-called Hendon cow disease, was raging in some 
farms in Wiltshire. In this case every facility was given by the 
owner of the estate for a thorough investigation into the disease. 
Not only were animals sent from the farm to London, but the 
author was allowed to visit the farms, to inspect all the infected 
animals, and to make every investigation, with the hearty co- 
operation of the bailiff of the farms, and the voluntary assistance 
of the head cowmen and those under them. Some of these cowmen 
were unusually intelligent, while two had had experience of cows 
for more than half a century. Thus, there was not only every 
opportunity for studying the disease on the lines indicated by 
Klein, but it was possible by repeated visits to the farms to enter 
into the clinical history of the disease in the cow, to study very fully 
the nature of the disease on the hands of the milkers, and to trace 
the probable mode of its introduction on the estate, and the way in 
which it spread from one part of the herd to another. 

Two cows were sent to London vsdth disease of the teats and of 
the udder between the teats. On the right teats of one there were 
numerous sores, covered with crusts varying in size and in thickness, 
and generally fissured. In some they were flat, in others conical ; 
some were with difficulty removed with forceps, others were readily 
detached. The crusts varied in colour from reddish-brown to veiy 
dark brown or almost black. On detaching or scraping ii. crust 
there was a granulating and somewhat indurated base. On the 
right anterior teat there were several ulcers, from which appa- 
rently the thick crvists had been detached, and new scabs were 
forming. On the left posterior teat there were unusually large, 
dark brown, or blackish crusts, covering a very extensive area 
of ulceration, extending over the whole of the lower third of the 
teat. 

In the other cow from Wiltshire there was the same disease on 
the teats, but not in such a severe form. The sores were covered 
with thick crusts, but though varying in size they were moi-e 
regular in form, and more circumscribed. 

Having entirely removed the crusts from some of the ulcers, a 
number of inoculations in nutrient gelatine and nutrient agar-agar 
were made from the discharge, and cover-glass preparations were 
made and stained in the ordinary way. Cultures were obtained of 
the organisms commonly found in pus. 



272 INFECTIVE DISEASES. 

With the discharge and with scrapings from the ulcers two 
calves were inoculated. 

Of the two calves, one was inoculated by scarification in both 
ears ; the other, a small calf, was scarified in the left ear. Scrapings 
from the ulcers were rubbed into the places thus prepared. In 
addition, in the small calf an incision was made through the corium 
in the left groin, scrapings from different ulcers on the teats were 
well rubbed in with the blade of the scalpel, and a portion of crust 
inserted into a small pocket in the subcutaneous tissue. In the ears 
and the groin there were positive results. In the large brown calf 
one of two places inoculated in the right ear passed through the 
following changes : On the third day there was apparent vesiculation 
and commencing formation of crust. From day to day the crust 
thickened, and on the eighth day the crust was at its height and 
detached at its edges. By removing the scab an Tilcer was exposed ; 
there was slight inflammatory thickening. About the thirteenth 
day the ulcer had quite healed. 

"Very similar appearances resulted in the ear of the smaller calf. 
The result of inoculation in the groin was of a very much severer 
character. In the course of two or three days the incision had 
apparently commenced to heal by scabbing, but there was a surround- 
ing area which was inflamed, and painful on manipulation. The 
inflammatory thickening which resulted continued to increase around 
the seat of inoculation, and the thickening could be felt to extend 
deeply into the groin. Suppuration followed, and on firm pressure 
pus welled up through the wound. The wound then showed very 
little disposition to heal, and the calf began to exhibit marked 
constitutional symptoms. During the second week after inoculation 
the animal became very dull, and was reported by the attendant as 
refusing to feed. Diarrhoea supervened, and lasted for several days, 
and bloody urine was passed. The calf was also noticed to cough, 
and the cough gradually increased in severit}-. Thirty-six days after 
the date of inoculation it was decided to kill the calf and examine 
the condition of the viscera. The appearances which were found 
at the post-mortem examination were as follows : — 

The upper and middle lobes of each lung were adherent to the walls 
of the chest ; there was congestion, especially of the middle lobe, and 
patches of recent adherent lymph. Posterior parts of the upper lobes of 
both lungs were completely consolidated, and on section varied in colour 
from brick-red to greyish-white. The interlobular tissue was infiltrated 
with inflammatory products, which mapped out the tissue of the lung in 
small indurated areas, in which the tissue was granular-looking and friable. 



THE author's investigation. 273 

These appearances in the upper lobes were due to septic pleuro-pneumonia. 
They closely resembled, and were supposed to be due to, infectious 
pleuro-pneumonia. They were, however, found identical with the con- 
dition observed in septic pleuro-pneumonia in calves, and the disease was 
not conveyed by infection to other animals in the same stall. Scattered 
through the other lobes of both lungs were white, mostly firm, nodules 
raised above the level of the surface of the lung. They were surrounded 
by a, zone of congestion, and in some cases sections were composed of 
indurated, in others of friable, lung tissue. In the posterior part of the 
right upper lobe there was a recent infarct. The bronchial glands at the 
roots of each lung were enlarged to two or three times their 'natural size, 
and were firm and hard on section. The parietal surface of the pericardium 
was covered with recent adherent lymph. The visceral surface of the 
pericardium was normal. Along the external surface of the aorta were 
chains of enlarged lymphatic glands connected by dilated lymphatic 
vessels. These glands were dark red or purplish in colour, from haemor- 
rhage into their substance. The heart was normal, and the endocardium 
not stained. There were chains of red glands on the oesophagus similar 
to those along the aorta. The appearance of the mesenteric glands was 
very striking. The mesentery, along the lymphatic vessels, was dotted 
with glands, varying in size from a large shot to a pea, which were deep 
red or prune-coloured. In addition, there were here and there enlarged 
glands without hsemorrhage into their substance, and greyish in colour. 
There were scattered petechiae on the spleen. The kidneys were firm 
on section, and there was marked congestion in both, while it was more 
pronounced in one kidney than the other. The liver was congested, the 
congestion being more marked in patches. 

Sections from the consolidated upper lobes showed under the micro- 
scope thickening of the pleura and infiltration with round cells. The 
exudation filled the alveoli, and was breaking down in some cases in the 
centre. The vessels were injected, and there were hsemorrhages into the 
alveoli. The periphery of the lobules was infiltrated with round cells. 
In sections of the kidney there was slight infiltration around glomeruli 
and arterioles with round cells ; the epithelium in the convoluted tubules 
was granular and disintegrating ; there was haemorrhage in the straight 
tubules, and engorgement of vessels. In sections of liver the inter- and 
intra-lobular vessels were engorged ; there were interlobular collections 
of round cells displacing the liver cells, and the interlobular connective 
tissue was infiltrated with round cells ; the liver cells were granular and 
cloudy. 

There can be no doubt from the symptoms and post-mortem 
appearances that this calf had been suffering from septicaemia as 
the result of introducing the septic virus and crust subcutaneously 
in the groin. 

The two Wiltshire cows were killed, and there was nothing of 
importance to note in one, but in the other an incision into the 
udder revealed an enormous abscess. 

18 



274 INFECTIVE DISEASES. 

Though the naked-eye appearances of the kidney in this case 
were practically healthy, the results of examining sections of the 
kidney under the microscope were extremely instructive and interest- 
ing, as they showed that marked changes had taken place which 
were indicative of septic complication. 

The sections showed glomerulo-nephritis ; there was infiltration 
of the capsule of Bowman with round cells ; there was infiltration 
also of the sheaths of the vessels with round cells, especially in 
the cortex. The blood-vessels in the boundary zone of the medulla 
were engorged, the arterioles of the glomeruli were also engorged, 
and there were slight hsemorrhages into the capsule. The epithelium 
of the convoluted tubules was granular, opaque, and in some parts 
breaking down. 

Sections of the ulcers of the teats of these cows were also 
carefully examined, and the appearances corresponded exactly with 
the description given by Klein. 

On visiting the farms it was found that there were altogether 
about a hundred and sixty cows. Only a few had proved refractory, 
and had not taken the disease at all. The rest had contracted the 
disease in varying degrees of severity. About fifty at a time were 
dry, and they escaped until they were in milk again. The milk 
was drunk on the farms and in the village, and a quantity was 
supplied to a large town. Most careful inquiries were instituted to 
ascertain the existence of scarlatina among consumers of the milk. 
So far the research was completely analogous to the Hendon 
investigation; but, in spite of the contamination of the milk, no 
cases of scarlatina were found either on the farms or in the village, 
and there was no epidemic in the town in which the milk was 
distributed. 

The disease, in fact, was cow-pox, and in no way connected with 
scarlet fever; and to assist others who may undertake a similar 
inquiry the details will now be given of the author's investigation 
into the nature of the outbreak in Wiltshire. 

The Disease proved to be Cow-pox. 

Locality of the Wiltshire Outbreak.— There is considerable interest 
attached to the fact that the farms were situated a few miles from 
Cricklade. They are close to the borders of Gloucestershire, and about 
twenty-five mUes from Berkeley. They are, therefore, within that 
district in which in Jenner's time cow-pox was particularly prevalent. 

Time of Year.~The outbreak commenced about the end of September 
1886, and lasted until about the middle of December. In an outbreak 



AN OUTBREAK OF COW-POX. 275 

in 1885, a few miles from these farm.s, but on a separate estate, the 
disease appeared in June and July. 

Origin of the Outbreak. — The author made careful inquiries as to the 
origin of the outbreak, but beyond ascertaining with certainty that the 
disease appeared first at one farm, and was conveyed from this to the 
other farms, all evidence was negative. The milkers were unable to 
say whether it commenced in one particular cow or whether it broke 
out in several simultaneously. 

The only information which could be obtained, which was very 
suggestive, was that the milkers were in the habit of receiving their 
friends from neighbouring farms on Sundays. The friends would assist 
in the milking, to get the work done as quickly as possible on these 
occasions. As it was reported that the same disease had occurred 
that summer on a neighbouring farm, it is quite possible that it was 
introduced by one of the milkers' friends. 

Mode of Dissemination. — When the disease made its first appearance, 
the bailiff, attributing it to the farm being, for some reason, unhealthy, 
decided to remove the cows to other farms. The herd was therefore 
divided and sent to two other farms. From these cows the disease was 
communicated to healthy cows, and, as this interchange was repeated, 
not only of the cows, but of the milkers, the disease was communicated 
to four separate farms. 

In all cases the disease was limited to the teats, and was conveyed 
from the teats of a diseased cow to the teats of a healthy cow by the 
hand of the milker. In no case was there any evidence of the disease 
being produced in healthy cows by other means than contact. 

Bulls and dry cows remained free from the disease, while the cows 
in mUk, numbering about a hundred and twenty, were all attacked, with 
the exception of about a dozen, which proved to be entirely refractory. 

These facts explain how it is that the disease has been known from 
time immemorial as the " cow-pox.'' We never hear of cattle-pox or 
bull-pox. We have not, in other words, to deal with an infectious 
disease like cattle-plague or pleuro-pneumonia, but with a disease which 
is communicated solely by contact. 

The disease was only observed in the cows in milk, and was limited 
to the parts which come in contact with the hand of the milker. The 
virus was mechanically transferred from diseased to healthy cows, being 
communicated to all, or nearly all, the animals in the same shed, 
whether the milker had vesicles on his hand or not. 

Character of the Eruption on the Cow. — In a recent case which was 
carefully examined the teats were visibly infiamed, partly red and partly 
livid in colour. On each teat there were vesicles, some broken, and 
others which appeared to be just forming. In other cases there was 
nothing more than the remains of broken and dried vesicles, and more 
or less characteristic crusts on the teats. 

On visiting a byre at the time that the cows were brought in to 
be milked, it was a striking sight to look along the line and see one 
animal after another affected with the eruption ; and thus one character 



276 INFECTIVE DISEASES. 

of the disease was clearly shown— the tendency to spread through a 
whole herd. 

On examining the eruption carefully, the degree of severity was 
found to differ very much in different animals. In a few cases the 
condition was most distressing, both to the cow and to the observer. In 
such oases the teats were encrusted with huge, dark brown or black 
crusts, which, when handled in milking, were broken and detached, 
exposing a bleeding, suppurating, ulcerated base. Such ulcers varied 
in size from a shilling to a florin, and in form were circular, ovoid, or 
irregular. "Weeks afterwards, when the animals had recovered, the 
site of these ulcers was marked by irregular scars. 

All the milkers agreed as to the general characters of the malady, 
laying particular stress on the teats being red, swollen, and painful when 
handled. Vesicles would then appear on the teats — two, three, four, or 
more on each teat. They were soon broken in milking, and irritated into 
sores, which became covered with thick crusts. From four to six weeks 
elapsed before they had entirely healed. Other more observant milkers 
insisted that before the teats were red and swollen, spots or pimples first 
appeared which came to a head. This head increased if it was not broken, 
which might be the case if it was situated between the bases of the teats, 
until it formed a greyish vesicle of the size of a threepenny-piece or 
even larger. 

General Sym2)toiiis in the Cow. — As to the general condition of the cows 
nothing abnormal was observed. They appeared in the best of health, 
and in only one particular was any difference from their condition in 
health stated to exist. This was, that in the majority of the cases there 
could be no doubt that the milk had diminished. This might escape 
notice by inexperienced milkers in any particular animal, but the total 
amount of milk supplied by the herd was considerably below the average. 

History of the Eriqjtion communicuted to the Milkers. — The most striking 
characteristic of this outbreak was the communicability of the disease 
to the milkers. A milker, with vesicles which presented typically the 
characters of casual cow-pox, was taken to London and kept under 
observation. The various cases will be described in the order in which 
they first presented themselves, their history being given as much as 
possible in their own words. 

Case I. — J. R., milker, informed the author that he was the first to catch 
the eruption from the cows. He stated that it came as a hard, painful spot, 
which formed " matter " and then a " big scab." He had been inoculated 
about seven weeks previously. He pointed to the scar which remained 
on his right hand. This scar presented the characters of an irregular 
cicatrix, indicating considerable loss of substance. He stated that he had 
also two places on his back, where he supposes he had inoculated himself 
by scratching. He had continued milking ever since, but had had no 
fresh places. 

Case II. — W. H., milker. He stated that he was inoculated from the 
cows about the same time as J. R. They were the two milkers of 
the herd in which the cow-pox first made its appearance. The eruption 



AN OUTBREAK OF COW-POX. 277 

appeared in one place on each hand. He pointed to two irregular scars 
as the remaifis of the eruption. 

Case III. — J. L., milker, stated that he also caught the disease from 
the cows. On his right hand a spot appeared which formed a hlister, 
then discharged matter and produced a bad sore. Lumps formed at the 
bend of his elbow and in his armpit. He lost his appetite, felt very 
poorly, and was obliged to leave ofE work for two or three days. 

Case IV. — W. K., a labourer on the farm, was put on as a milker to 
take the place of one of the others with bad hands. After his fifth or 
sixth milking — that is to say, about three days after first milking the cows, 
— pimples appeared on his hands, which became blistered and then ran on 
to bad sores. He pointed to three irregular scars on the first and third 
fingers and palm of the right hand. Lumps appeared in his elbow and in 
his armpit, but he did not feel very poorly in consequence. 

Case V. — J. F., milker, stated that about a month ago he noticed 
spots which appeared on both hands. His fingers swelled and were pain- 
ful. He said it came first like a pimple, and felt hard. Then it " weeped 
out " water in four or five days. There were red marks creeping up to 
his arm. There was a sort of throbbing pain, and he could not sleep at 
night. On the right hand there was a scar, but on the left hand there 
was an ulcer about the size of a shilling covered with a thick black crust. 
The crust was partially detached, and exposed a granulating ulcer. It 
was in this stage the exact counterpart of the ulcers on the cow's teats. 

Case VI. — W. H., junior, milker, stated that he had both hands bad 
about a month previously : fii-st on the index finger of the left hand, 
and then on the right hand on his knuckle and between the first and 
second fingers. He said that it came up like a hard pimple, and the 
finger became swollen and red. After a few days it "weeped out" 
water, and then matter came away. Both his arms were swollen, but 
his left arm was the worst. About a fortnight after, he noticed kernels 
in his armpits, which were painful and kept him awake at night. His 
arms became worse, he could not raise them, and he had to give up 
milking. He also had had a " bad place ' on the lower lip. On examina- 
tion, I found that the axillary glands were stiU enlarged and tender. 
He volunteered the statement that the places were just like the sore 
teats. 

Case VII. — J. H., the bailiff's son, also milked the cows. He had 
a sore on the upper lid of his right eye and on his left hand. In both 
cases he had been previously scratched by a cat, and the scratches were 
inoculated from the cow's teats. The right hand also had been inoculated. 
The eruption broke out a fortnight previously. His hands were swollen, 
red and hot. He felt very poorly and went to bed. Little spots like 
white blisters appeared on the back of his right hand. His mother 
remarked that they "rose up exactly as in vaccination." Thick dark 
brown scabs formed. He was very ill for two or three days, but did not 
send for a doctor. He had painful lumps at the bend of his arm and in 
the armpit. He gave up milking, and had not taken to it since. 

On examining him, the thick crusts on his right hand were identical 



278 INFECTIVE DISEASES. 

with the stage of scabbing in vaccinia. The scabs fell ofE in about three 
■weeks to a month, and left permanent, depressed scars. 

Case VIII.— W. P., milker. This case was pointed out on the occasion 
of another visit, and is the only one in which the eruption was seen in 
its earlier stages. 

The history of this boy is as follows. He had taken the place of one 
of the other milkers who had vesicles on his fingers, and had been obliged 
to give up milking. After the seventh time of milking he noticed a 
small pimple on his right cheek. This became larger and vesicular. 
On examination it presented a depressed vesicle with a small central 
yellowish crust and a tumid margin, the whole being surrounded by a 
well-marked areola and considerable surrounding induration. On raising 
the central incrustation a crater-like excavation was seen, in which 
lymph welled up and trickled down the boy's cheek. On the following 
day the crust had re-formed, and was studded with coagulated lymph. 
The areola became more marked, and on pricking the margin of the 
vesicle, the exuding contents were slightly turbid. 

From this day the surrounding infiltration increased enormously, 
the whole cheek was inflamed, and the eyelids so oedematous that the 
eye was almost closed. There was enlargement of the neighbouring 
lymphatic glands. The crust which had re-formed thickened day by day. 
It retained the character of central depression, and was situated on a 
reddened, raised, and indurated base (Plate VII.). 

From this date the surrounding induration gradually diminished. 
The crust changed in colour from dark brown to black, and finally 
fell off, leaving an irregular, depressed scar. This scar, when seen several 
months afterwards, was found to be a permanent disfigurement. The 
eruption appeared on the fourth day after exposure to infection, and 
allowing two days for incubation, the vesicle was at 'its height on the 
seventh or eighth day, and a typical tamarind-stone crust fell ofE on 
the twenty-first day after infection, leaving a depressed, irregular 
cicatrix. 

A vesicle also formed on the thumb of the left hand. Two days 
after the pimple appeared on his cheek, the lad said that he first noticed 
a pimple on his thumb, and this, on examination, presented a greyish 
flattened vesicle, about the size of a sixpence. Later, its vesicular 
character was much more marked, and a little central crust had com- 
menced to form. The margins became very tumid, giving it a marked 
appearance of central depression. The vesicle was punctured at its 
margin with a clean needle, and from the beads of lymph which exuded 
a number of capillary tubes were filled. 

Two days afterwards suppuration had commenced, the vesicle con- 
tained a turbid fluid, and the areola was well marked. Later, the crust 
had assumed a peculiar slate-coloured hue, and, on pressing it, pus 
welled up through a central fissure. The areola had increased, and 
there was considerable inflammatory thickening. The lymphatic glands 
in the armpit were enlarged and painful. Though there was deep 
ulceration, which left a permanent scar, the ulceration did not assume 



DESCRIPTION OF PLATE VII. 
Casual Cow-pox. 

Fig. 1. — Case of W. P , a milker, infected from the teats of a cow with 

natural cow-pox. There' was a large depressed vesicle with a small 
central crust and a tumid margin, the whole being surrounded by a 
well-marked areola and considerable surrounding induration. 

Fig. 2. — The same case a week later, showing a reddish-brown crust on a 
reddened elevated and indurated base. 



Hate'Wr. 




Pis^i 




Jig 2, 



CASUAL COW-POX. 



Hn.^^n.t Er'^ohs.Day i.Jon.Ll 



M Craoks'ha.niltfe.ciZ. 



AN Ol'TBREAK OF COW-POX. 279 

quite so severe a character aa in some of the other milkers. Possibly 
this may be accounted for to some extent by the fact that the pock was 
covered with a simple dressing instead of being subjected to the irritation 
and injury incidental to working on the farm. 

Revaccination of the Milkers. — There were in all eight milkers, varying 
in age from seventeen to flfty-five, who had vesicles on their hands 
from milking the cows. Seven had been vaccinated in infancy, but not 
since ; one had been revaccinated on entering the navy at fifteen. They 
were all revaccinated by a public vaccinator after complete recovery 
from the casual cow-pox (that is to say, from three to four months 
afterwards), and were all completely protected. On the other hand, 
,two of the three milkers who had escaped infection from the casual 
cow-pox were also vaccinated, with the result in one of typical 
revaccination, in the other of very considerable local irritation. 

Retro-vaccbmtion of Calves. — The result of retro-vaccinating calves with 
the humanised lymph was strictly in accordance with the experience of 
Ceely, who has pointed out that in retro- vaccination from the milker's 
hands the results are doubtful, and depend greatly on the animals selected. 
"Those of a light colour and with thin skins were generally preferred, 
but often without avail, scarcely one-half of the operations succeeding." 
" Yaccine lymph, in passing from the cow to man, undergoes a change 
which renders it less acceptable and less energetic on being returned to 
many individuals of the class producing it ; some refuse it altogether." 
Two cases out of four succeeded, and an eruption was produced with 
all the typical characters of vaccinia, but running rather a rapid course, 
and the protection passing ofE after a few weeks, while the result 
obtained in calves inoculated with pus or scrapers from ulcers was in 
accordance with what is well known to occur if pus instead of lymph 
is taken for carrying on calf to calf vaccination. 

That the cow-pox in Wiltshire was identical vrith the so-called 
Hendon cow-disease there can bo little x-oom for doubt, for in both 
cases we find that — 

1. The disease spread through a whole herd of milch cows. 

2. The disease was characterised by the appearance of vesicles, 
which were broken by the hand of the milker, and irritated into 
deep ulcerations. 

3. The disease was conveyed from one cow to another by the 
hand of the milker. 

4. The vesicular eruption was communicable to the hand of 
the milker. 

5. The disease was not fatal, and in cows which were killed and 
examined the post-mortem appearances could not be distinguished 
from accidental complications. 

6. The naked-eye appearances and the duration of the ulcers of 
the teats were the same. 



280 INFECTIVE DISEASES. 

7. Sections of the ulcers showed vinder the microscope identical 
appearances of a cellular character, and the purulent discharge of 
the ulcers contained pyogenic cocci. 

8. The results produced by inoculation of calves with the septic 
virus were identical. 

If we examine the chain of argument which has been brought 
forward to maintain the existence of cow-scarlatina at Hendon, wo 
find that it was urged : — 

1. That the Hendon cow disease was a disease in which the 
post-mortem appearances resembled scarlatina. 

2. That this disease was associated with a streptococcus, which 
produced, by inoculation in calves, a disease with post-mortem 
appearances similar to those of the Hendon cows. 

3. That a streptococcus regarded as identical with the one 
above mentioned was found in certain cases of scarlatina in man, 
which when inoculated in calves produced post-mortem appearances 
similar to the post-mortem appearances in the original Hendon cows 
and in certain cases of scarlatina in man. 

But the microscopical appearances of the kidney of a Wiltshire 
cow were identical with those which were. regarded as indicating 
scarlatina in a Hendon cow ; and, indeed, the statements as to the 
post-mortem appearances in the Hendon cows, when studied, not 
only do not necessarily indicate scarlatina, but they cannot even be 
considered of j)rimary importance, or as throwing much light on the 
question of scarlatina at all. The description of the naked-eye 
appearances in both cows only suggests coincident pleurisy or 
pleurisy with pne^imonia. The microscopical appearances in both 
were suggestive of septic complication. 

A careful examination of the post-mortem appearances of calves 
inoculated with scraping of an ulcer of a Hendon cow, or with 
cultivations of the streptococcus from certain cases of scarlatina, 
brings to light much more striking changes. These appearances, 
however, cannot be regarded as indicative of scarlatina. They are 
in reality the post-mortem appearances of septic poisoning, and 
occur commonly in many diseases. This is clearly shown by com- 
paring the post-mortem appearances in the calf which was killed 
while suffering from septicsemia as the result of inoculation from 
the ulcers of a Wiltshire cow. These visceral changes are not to 
be distinguished from the post-mortem appearances described in 
the calves inoculated by Klein. Consequently, that the strepto- 



AJSf ODTBEEAK OF COM'-POX. 281 

COCCUS found in certain cases of scarlet fever should produce on 
inoculation in calves certain post-mortem appearances whicli are 
found in many diseases, and should fail to produce fever, ulceration 
of the tonsils, or Scarlatinal rash, or any condition in the least 
resembling, clinically, the disease in man, and yet that the result 
should be regarded as scarlatina in the calf, is a conclusion quite 
untenable. 

It is true that visceral lesions similar in character were produced 
in calves whether inoculated with scrapings or with streptococci from 
ulcers of the Hendon cows or with streptococci from certain cases of 
scarlet fever. In both cases the streptococcus is pathogenic, and 
inoculation of Streptococcus pyogenes or the inocvilation of septic 
virus, is liable to produce septicaemia. These facts constitute a mass 
of evidence which justifies the conviction that the pathological data 
which appeared to support the theory that the vesicular disease of 
the teats of cows at Hendon was scarlatina in the cow, admit of an 
entirely different interpretation, and there can be no longer any 
doubt that the milk was not infected by the cows but with the virus 
of scarlet fever from some human source which Mr. Power failed to 
discolor. 

All the other evidence reported to the Board of Agriculture pointed 
to the same conclusion. The disease at Hendon was admittedly 
introduced from Derbyshire ; and from Professor Axe's report it 
appears that only a part of the herd was sold to the farmer at 
Hendon; other cows with the same eruption were transferred to 
other dairy farms, and the disease communicated to healthy cows as 
at Hendon, but in no instance did scarlet fever occur among the 
consumers of the milk. At the farm of the brother of the dealer the 
disease was communicated to three of the milkers, and the eruption 
diagnosed by Dr. Bates as vaccinia. 

All this evidence must be regarded as conclusive. The con- 
tamination of the milk at Hendon with scarlet fever must neces- 
sarily have been a mere coincidence ; and the conclusion that 
the milk could not possibly have become infected from any 
human source is untenable. Professor Axe even ascertained 
that scarlet fever existed at Hendon during several months of 
1885, and that the dwellings where cases occurred stood within six 
hundred yards of the cowsheds which contained the incriminated 
cows, and that out of fourteen men on the farm six lived in 
a district where cases occurred. Professor Axe has also stated 
that the father and brother of a girl with scarlet ,fever, visited the 
dairy during her illness. Whether any of those engaged on the 



282 INFECTIVE DISEASES. 

farm suffered from latent scarlet fever does not appear to have 
been ascertained. 

There is, it is true, no evidence to show that any one daily carried 
infection to the milk, but the exact path of infection is not always 
easy to trace ; and because it was not actually traced it was hardly 
reasonable to assume that the possibility of contamination from a 
human source could be altogether eliminated. 

In attempting to communicate scarlet fever to cows Professor 
M'Fadyean confirmed the negative results which had been experi- 
enced in some earlier experiments by Klein In 1882 Klein 
inoculated and fed cows and yearling heifers with diseased products 
from human patients, using desquamated cuticle and the discharges 
from the throat ; but the experiments all 'failed. MTadyean's 
failures were still more marked. Cows and calves were inoculated 
with blood from scarlet fever patients, and they were made to drink 
water thickened with desquamated cuticle, but all the experiments 
proved unsuccessful. 

The author believes that the outbreak at Hendon was one 
of cow-pox, which was prevalent in this cotintry in 1886. The 
outbreak in Wiltshire could not be distinguished bacteriologically 
or clinically or in its micropathology, from the disease at Hendon, 
and the Wiltshire outbreak proved on investigation to be true cow- 
pox. This conclusion was questioned at the time, as cow-pox was 
generally believed to be extinct in England ; but that view is 
entirely fallacious, and the author's conclusions have since been fully 
confirmed by independent observers, whose work will be referred to 
in another chapter (p. 321). 

Stamping-out System. — The Notification Act of 1890 may be 
voluntarily adopted in sanitary districts, but it would be a great 
advantage if notification were carried out uniformly all over the 
country. Prompt information may lead to detecting the origin of 
cases of scarlet fever, and isolation and disiofection will assist in pre- 
venting its spread. Epidemics have occurred on a large scale owing 
to scarlet fever existing among those engaged in dairy work, and 
the precaution not being taken of stopping the milk supplied to the 
consumers. Scarlet fever cannot be so readily controlled as small- 
pox, for it may be spread by mild cases before the nature of the 
disease is suspected, and small-pox cannot be conveyed in milk. 



MEASLES. 283 



Measles. 



Measles is a contagious disease peculiar to man. It lasts for 
one or two weeks, and produces fever, catarrh of the respiratory 
mucous membrane, and a characteristic rash. It is highly contagious, 
especially before the nature of the disease is revealed; there is 
consequently great difficulty in preventing its spread in schools 
and households. The contagium appears to be given off from the 
body, principally if not entirely, by the breath. One attack is pro- 
tective against future attacks. ~"The whole population of a, country 
may acquire a certain degree of immunity. Measles introduced into 
countries where it was previously unknown assumes a most malig- 
nant form. There are no characteristic post-mortem appearances. 

Bacteria in Measles. — Micrococci have been found in the 
blood, catarrhal exudation, and skin, by Keating, Babfes, and others, 
but they are accidental epiphytes of no importance, or associated 
with secondary complications, as in scarlet fever. 

Canon and Pielicke have found in the blood small bacilh varying 
in form. They do not grow on nutrient agar or blood serum, but 
cultures were obtained by pricking the finger of a patient suffering 
from measles, and allowing the blood to drop into sterilised broth. 
After a few days the broth became cloudy, and later, a flocculent 
deposit formed. The bacilli were also obtained from the nasal and 
conjunctival secretions. The nature of the contagium of measles 
is unknown. 

Stamping-out System Measles is not easily controlled by the 

stamping-out system ; it is, in fact, extremely difficult, almost impos- 
sible, to prevent its spread, as it is especially infectious during the 
period of incubation. Notification, isolation, and disinfection assist in 
controlling an epidemic, but the value of the system does not apply 
to the-same extent in measles as in other infectious diseases. 



CHAPTEE XX. 

SMALL-POX. — CATTLE PLAGUE. 

Small-pox. 

Small-pox is an infectious and inoculable disease of man, charac- 
terised by sudden and severe fever, followed in forty-eight hours by a 
characteristic papular eruption which gradually becomes vesicular 
and then pustular. The virus is contained in the vesicles, and in a 
concentrated form in mature pustules. It also passes into the air 
from the breath and skin. Infection may occur from the dead body, 
and clothes and bedding may retain the coutagium for months. 
One attack, as a rule, gives immunity against future attacks. 

Small-pox is undoubtedly a disease foreign to this country. Its 
home is in the East. Some of the old writers held that it spread to 
Europe from Alexandria about the year 640 a.d., following in the 
wake of the Arab conquests in Egypt, Palestine, Persia, along the 
Asiatic coast, through Lycia, Gallicia, along the coast of Africa, 
and across the Mediterranean to Spain ; others maintained that it 
was not introduced until the end of the eleventh or beginning of the 
twelfth century, by the returning Crusaders. At any rate, small- 
pox was imported from the East, and probably from Egypt. Hero- 
dotus, who visited Egypt, leads us to infer that epidemics were 
unknown there during the rule of the Pharaohs ; but Egypt 
undoubtedly became a hotbed of pestilence during the Mohammedan 
occupation. Prosper Alpinus imagined that both the plague and the 
small-pox were concocted in the putrid waters of the Nile, but he 
would probably have been more correct if he had suggested that 
they arose from the insanitary condition of the Arab conquerors 
and their filthy camp followers, who did their best to destroy all 
that remained of that magnificent civilisation which had existed in 
the days of the ancient Egyptians. 

We do not know the exact period at which small-pox was first 
imported into England, and the records of the disease are very 
meagre until the sixteenth century. 

284 



SMALL-POX. 285 

In 1593 Simon Kellwaye appended to his work on the Plague 
a short treatise on the small-pox. " Oftentimes," he wrote, " those 
that are infected with the plague are in the end of the disease 
sometimes troubled with the small pocks or measels, as also by good 
observation it hath been seen that they are fore-runners or warnings of 
the plague to come." According to Kellwaye the disease arose from 
the " excrements of all the foul humours in our bodies, which striving 
with the purest doth cause a supernatural heat and ebullition of our 
blood, always beginning with a feaver in the most part." 

Small-pox steadily increased in the seventeenth century until it was 
a formidable scourge, for no advantage was taken of all the experi- 
ence which had been gained in dealing with the plague. No public 
measures were adopted to cope with the disease, and the people came 
to regard the new pestilence as a visitation which was unavoidable. 
Early in the eighteenth century, small-pox inoculation was introduced, 
and this was superseded in the nineteenth century by vaccination. 

Examination of small-pox cases after death does not reveal any 
characteristic lesions in the internal organs, but sections of small- 
pox vesicles show an important structure. A vesicle is formed by 
the exudation raising up the outer layer of epidermis, and the chief 
feature is the formation of a vacuolated structure in which, especially 
in the later stages, bacteria are found iji abundance. 

Bacteria in Small-pox. — Cohn and Weigert found cocci in 
variolous lymph. Hlava found Streptococcus pyogenes in the pustules, 
and Garre streptococci in the internal organs in a case of variola 
haemorrhagica. In a fatal case of variola complicated with pemphigus 
Garr6 found a streptococcus in the pemphigus vesicles. Klein 
and Copeman have found a small bacillus which they regard as 
characteristic, but its biological characters are unknown, as it will 
not grow on any nutrient media. The bacteria commonly found in 
variolous pus are the usual pyogenic organisms. The nature of 
the contagium of small-pox is unknown. 

Protective Inoculation. — Experience had taught that a person 
was not, as a rule, attacked with small-pox a second time ; but when 
and how the method of artificially inducing a mild form of the 
disease was discovered, or when this preventive treatment was first 
employed, is unknown. Avicenna of Bokhara was credited with the 
discovery, and it was supposed that the practice was carried by 
Tartar and Chinese traders to Surat, Bengal, and China, and by 
the Mahommedan pilgrims to Mecca. In Constantinople it was 
supposed by some to have been introduced from thiC Morea by 
an old woman, and by others by the women of Oircassia. The 



286 INFECTIVE DISEASES. 

Circassian women fastened three needles together, and pricked the 
skin over the pit of the stomach and heart, the navel, the right 
wrist, and the left ankle. The variolous matter was applied to the 
bleeding points, and the eruption came out in five or six days. In 
Constantinople scarifications were made on the forehead, wrists, 
and legs, and carefully selected virus appUed to the incisions. The 
needle used was a three-edged surgeon's needle, or the operation was 
performed with a lancet. The virus was obtained by pricking the 
vesicles, and pressing out the matter into a clean glass vessel. The 
Armenians preferred to be inoculated in both thighs. In Barbary 
a slight wound was made between the thumb and forefinger, and 
the virus obtained from a mild form of small-pox. In Hindustan 
the operation was performed at certain seasons of the year, and a 
preparatory regimen enforced. The inoctilators were very careful 
in the selection of the virus, as they had learnt its varying intensity, 
and they were credited with being able to control the amount of 
the eruption. They preferred to inoculate the outside of the arm, 
midway between the wrist and the elbow in males, and between the 
elbow and the shoulder in females. The skin over the part to be 
inoculated was first well rubbed with a piece of cloth ; then, with 
slight touches of a small instrument, little wounds were made over 
an area which might be covered by a small coin, and sufficient to 
cause just an appearance of blood. A pledget of cotton-wool 
charged with the variolous matter, and moistened with water, was 
applied to the wound. This virus was obtained from inoculated 
pustules of the preceding year. 

In China the contents of the variolous pustules were dried and 
kept for several years. If the virus was to be used from fresh 
pustules the " acrimony " of the matter was corrected by steaming. 
The dried powder was made into a paste, which was wrapped up in 
cotton-wool and introduced into the nostril. 

The Greeks were more cautious in their procedure, and were 
said to inoculate tens of thousands without an accident. They 
operated only upon those in perfect health, punctures were made 
with needles, and the virus was used in the crude state, freshly 
obtained from the " kindly " pustules of a young child. They were 
particularly careful in the choice of the " ferment." 

Dr. Perrot Williams, in 1722, wrote that the practice of com- 
municating small-pox had long been employed in South Wales. The 
oldest inhabitants said that it had been a common practice with 
them " time out of mind," but Lady Mary Wortley Montagu was 
responsible for the general adoption of small-pox inoculation in 



SMALL-POX. 287 

England by persuading physicians in London to employ it. Lady 
Mary had her child inoculated in Turkey. An old Greek woman inocu- 
lated one arm, and Mr. Maitland, surgeon to the Embassy, the other. 
The disease ensued in due course with an eruption of a hundred 
pustviles. This was the first time that the Byzantine method of 
inoculation was performed upon an English subject. In 1721 Dr. 
Harris delivered a lectiire before the College of Physicians,, and 
described the successful inoculation of fovir children of the French 
consul at Aleppo, by means of a thread imbued with variolous pus. 
A daughter of Lady Mary was inoculated in England by Maitland in 
1721, and subsequently a number of criminals were inoculated by 
him. Incisions were made through the cutis, and pledgets which had 
been steeped in variolous pus from ripe pustules, were apphed to the 
wound. This was known as Maitland's or the reformed operation, 
but it was soon miodified, as troublesome ulcers resulted. Shortly 
afterwards Maitland encountered another obstacle. The child of a 
Mr. Batt was inoculated, had plenty of pustules, and soon recovered, 
but six of Mr. Batt's domestic servants, " who all in turn were 
wont to hug this child while under this operation, and whilst the 
pustules were out, never suspecting them to be infectious, were all 
seized at once with the right natural small-pox of several and very 
different kinds." 

Dr. Jurin in 1729 reverted to the Eastern method, and recom- 
mended virus from a mild case of small-pox, but the virus was still 
taken from perfectly maturated pustules, and the operation continued 
to be followed by bad results. In order to diminish the risks, Burgess 
in 1766 advocated certain improvements. An incision about an inch 
long was made on each arm through the cuticle, but not so deep as 
to wound the csUular tissue. A variolous thread was laid along the 
whole length of the wound and fixed with plaster. Ulcerations and 
other accidents continued to take place, and a new epoch in the 
history of inoculation was the introduction of the Suttonian method, 
in 1764-6. 

It was said that Mr. Sutton, with his assistants, inoculated 
one hundred thousand persons. The method was kept secret at first, 
but the essential points were all discovered and published by Dr. 
Dimsdale. Dimsdale recommended a very slight puncture with a 
lancet wet with variolous matter. Subsequently, Sutton published 
an account of his method, and the result of his operation may be 
given in his own words. 

" The lancet being charged with the smallest perceivahle quantity 
(and the smaller the better) of unripe, crude, or watery matter, immediately 



288 INFECTIVE DISEASES. 

introduce it by puncture, obliquely, between the scarf and true skin, 
barely sufficient to draw blood, and not deeper than the sixteenth part of 
an inch. Neither patting, nor daubing of the matter, in or over the 
punctured part, is at all necessary to its efficacy. This practice, indeed, 
is rather prejudicial than otherwise, as it may afEect the form of the 
incision, and thus be apt to confound our judgment upon it. 

" Indications of the Incision. — In the incipient state of variolous 
increase in the incision, a small florid spot appears on the part of access, 
resembling a flea-bite in size ; and on passing the finger lightly over it a 
hardness is felt not larger than a small pin's head. This florid appearance 
and hardness denote that the variolous principle is effectually imbibed, 
and their indications point no farther, unless the progress to vesication 
be very slow, in which case an uncomfortable number of pustules may be 
expected to follow. The florid spot iii most instances of inoculation is 
somewhat larger, or more extended on the second, than on the third day 
after the insertion. 

"About the fourth day from inoculation, should the incision begin 
to vesicate, an itching sensation will be complained of on the place of 
insertion — the occurrence of which symptom is the first indication of a 
favourable event, yet not of sufficient importance to justify any present 
relaxation in the preparatory proceedings. 

" The vesication of the incision in most instances will begin to be visible 
on the fourth or fifth day after the insertion of the matter ; the sooner 
it becomes so, the more favourable may be expected to be the event. The 
extent or diameter of the vesication at this stage does not usually exceed 
that of a large pin's head, and it has invariably a dint or small depression." 

Adams obtained still more striking results by inoculating with 
variolous lymph from pearl-pox, a mild variety of small-pox. Starting 
with lymph obtained from this benign form of small-pox, and 
selecting the cases, and carrying on arm' to arm variolation, the 
results obtained vrere practically identical with the phenomena 
obtained by inoculation of the arm with cow-pox lymph. Similar 
results were obtained by Guillou, but more rapidly. In 1827 there 
was an epidemic of variola, and Guillou, having no vaccine virus, 
took variolous lymph from a girl fifteen years of age on the fifth day 
of the eruption. The case was one of varioloid or mild small-pox, attri- 
buted to previous vaccination. The variolous lymph was inserted in 
ten places on the arm of a healthy infant still at the breast. This 
inoculation produced ten beautiful "vaccine'' vesicles, from which, on 
the ninth day, forty -two infants were inoculated under the eyes of 
two of the local authorities. These furnished virus for the inoculation 
of one hundred, who were inoculated in the presence of magistrates 
and many medical men. This experiment was repeated with success. 
Variolous lymph was taken from two lads at school, and in ten 



SMALL-POX. 289 

cases produced appearances with a perfect similarity to ordinary- 
vaccination. 

Thiele produced a benign vesicle in the following manner. 
Variolous lymph was diluted with warm cow's milk, and inocailated 
like ordinary vaccine lymph. Large vesicles resulted. There were 
febrile symptoms from the third to the fourth day, and a secondary 
onset of fever much more pronounced between the eleventh and 
fourteenth days. The areola was strongly marked, and not con- 
fined to the inoculated place, which was occasionally surrounded 
by minute secondary vesicles. After watching through ten removes, 
the vesicles finally assumed the characters of an ordinary vaccination 
with cow-pox lymph. As soon as the secondary fever ceased to 
occur inoculation was practised from arm to arm without diluting 
the lymph with cow's milk. The lymph was designated lacto-varioline, 
and the result was variolation in its mildest form. The result of 
variolating the cow will be discussed in another chapter. 

Small-pox inoculation, or variolation, protected the individual 
when genuine small-pox was produced, and .endangered the com- 
munity. Persons inoculated became centres of infection, and con- 
veyed the disease to others. Haygarth, although in favour of 
inoculation, strongly condemned its use without precautions to 
prevent the spread of the disease. " The most serious and solid 
objection," he wrote, " that has been advanced against inoculation 
is deduced from a comparison of the Bills of Mortality for a series 
of years in various places. They show that a larger proportion 
of inhabitants have died of the small-pox in towns where it is prac- 
tised than in the same before it was known, or in others where it is 
prohibited." 

Even Dr. Dimsdale, an ardent inoculator, admitted that more 
lives were lost in London than before inoculation commenced, and the 
practice was more detrimental than beneficial to society ; and he 
added : " The disease by general inoculation throughout London 
spreads by visitors, strangers, servants, washerwomen, doctors, and 
inoculators, by means of hackney coaches in which the sick are sent 
out to take the air, or by sound persons approaching them in the 
streets. The poor in London are miserably lodged ; their habitations 
are in close alleys, courts, lanes, and old dirty houses ; they are often 
in want of necessaries, even of bedding. The fathers and mothers 
are employed constantly in laborious occupation abroad, and cannot 
attend the inoculated sick." In 1798 Jenner, who had practised 
small-pox inoculation, proposed the use of a benign non-infectious 
lymph obtained from a disease of the cow or horse as a substitute 

19 



290 INFECTIVE DISEASES. 

for vEwiolous lymph, and in 1840 small -pox inoculation was prohibited 
by Act of Parliament. 

Stamping-out System. — The disappointing and dangerous 
results of small-pox inoculation led to a widespread demand for 
some new method for dealing with small-pox. This induced Haygarth 
to turn his attention to the subject, and towards the end of the 
eighteenth century to bring before the medical profession and the 
public a plan for stamping out the disease. Haygarth, who was a 
close observer and an able physician, studied the question of the 
communicability of the disease from one person to another, and its 
conveyance by infected clothing and other means, and ultimately 
drew up rules and regulations for its prevention, the importance of 
which we are only now beginning to fully acknowledge. Haygarth's 
essential doctrine was " that mankind was not necessarily subject to 
the small-pox, and that it was always caught by infection from a 
patient or the poisonous matter," and might be avoided by observing 
his Rides of Prevention. 

These rules comprised a regular system of notification and isolation. 
Inspectors were to be provided to report cases of small-pox, and people 
were to be rewarded for carrying out the instructions. Several examples 
were given of the results at Chester, where the plan was adopted. 

Haygarth met with considerable encouragement from some of the 
leaders of the profession. Dr. Fothergill wrote to him in 1778, saying, 
" I have mentioned the intention of freeing this country from the 
small-pox to divers of the faculty, and shall continue to do so as it falls 
in my way. The proposal is variously received, but in exact proportion 
to their humanity." 

In 1793 Haygarth made considerable addition to his rules, and urged 
that legislation should follow to make them compulsory. Provision was 
to be made to reward the poor for observing the rules, and public thanks 
to the wealthy for their support were to be published in the parish church 
and newspapers. Transgression of the rules was to be punished by a fine of 
from £10 to £50, one half to go to the informer and the other half to the 
fund which supplied the expense of rewards to the poor, and all details were 
to be supplied to the press. It was further suggested that Great Britain 
should be divided into districts, including a certain number of parishes 
or townships, and that to each of them a surgeon or apothecary should 
be appointed as inspector to see that the regulations were exactly 
observed. In addition, there were to be directors of inspectors, superin- 
tended by a commission of Physicians in London and in Edinburgh. All 
salaries were to be paid by the county rates, and the rewards for observing 
the rules of prevention were to be guaranteed out of the parish funds. 
On the requisition of the director and inspector of a circuit, power 
was to be given to two or more justices of the peace to appoint a separate 
house for the reception of patients with the small-pox. In conclusion, 



SMALL-POX. 291 

Haygarth maintained that the plague had been completely exterminated 
from this country, for above a century, by civil regulations, and that 
there seemed to be little doubt that the small-pox was propagated on 
principles similar to the plague, and that it also might certainly be 
exterminated from this island. 

Haygarth's teachings had a profound influence upon both the profession 
and the educated public, but his system of compulsory notification was 
never carried out, for no legislation followed to enforce his recommenda- 
tions. This is a matter deeply to be regretted, for towards the end of the 
eighteenth century srpiall-pox was declining in London, general sanitation 
was making rapid advances, small-pox inoculation, which created fresh 
centres of infection, was falling into disfavour, small-pox hospitals were 
built, which served to limit centres of infection, and the profession and 
the public were influenced by the teaching of Haygarth with regard to 
the various ways of avoiding the spread of the disease. 

It only required the compulsory adoption of Haygarth's system 
uniformly all over the country to have kept the disease in control, 
if not to have entirely extirpated it from Great Britain. That a 
similar conviction existed at the time is evidenced by an article which 
appeared in 1779 in the Medical and Ghirurgical Review, in which 
the following statements were made : — 

" Plans for the extirpation of the small-pox have been suggested. . . . 
To do this, however, the exertions of the physician are incompetent unless 
they be aided by the powerful hand of Governments, but this has hitherto 
been withheld. The grand means, however, of extirpating this destructive 
malady is an early and strict separation of the infected from those that 
are sound." 

Small-pox in the present century has been largely controlled by 
legislation, especially in recent years, by the Public Health Acts for 
England and Wales, for Scotland, and for Ireland ; the Epidemic and 
other Diseases Prevention Act ; the Public Health Amendment Act ; 
the Labouring Classes' Dwellings Acts ; the Housing of the Working 
Classes Act ; the Public Health (Ships) Act ; the Local Government 
Board Act — and various orders and memoranda of the Local Govern- 
ment Board ; the Infectious Diseases Notification Act ; the Infectious 
Diseases Prevention Act ; and the Public Health (London) Act. 

By the Public Health Act of 1875 England was divided into 
Urban and Rural Sanitary Districts, and powers were given to 
enforce regulations of the Local Government Board for guarding 
against the spreading of infectious diseases ; to provide medical aid 
and accommodation for infected persons, to promote cleansing, 
ventilation, and disinfection, to provide hospitals, to provide for 



292 INFECTIVE DISEASES. 

destruction or disinfection of infected bedding, clothing, and other 
articles, and to appoint Medical Oificers of Health. 

As to the value of notification and isolation in cities such as 
London we have the evidence of the Metropolitan Asylums Board. 
In their Eeport for 1889 we read in reference to the diminution of 
smaU-pox : " These very satisfactory results confirm the view taken 
by the Committee two years ago to the effect that the rapid and 
systematic removal from crowded districts of infected persons, each 
of whom inight have become a centre of contagion, is an important 
factor in stamping out small-pox from the metropolitan population. 
The notification of cases will also greatly facilitate the action of the 
managers in this direction." 

More recently there has been a most striking confirmation of 
these statements. An outbreak of small-pox occurred in Maryle- 
bone, and by the energy of the ofiicials of the Board this outbreak 
was suppressed in a few days by means of notification and 
immediate isolation. 

The Isolation Hospitals Act of 1893 gives power to County 
Councils to provide, or cause to be provided, an isolation hospital 
in any district within their county. An application to a County 
Council for the establishment of an isolation hospital may be made 
by any one or more of the authorities defined as local authorities 
having jurisdiction in the county or any part of the county. 
Further, the County Council may direct an inquiry to be made by 
two medical ofScers of health in the county as to the necessity of au 
isolation hospital being established for the use of the inhabitants 
of any particular district in the county, and in the event of such 
medical officers reporting that such a hospital ovight to be 
established for the use of the inhabitants of a district, may take the 
same proceedings in all respects for the establishment of such 
hospital, as if a petition had been presented by a local authority for 
the establishment of an isolation hospital for the district named in 
the report of such medical officers of health. 

Lastly, the Local Government Act of 1894 provides for the 
formation of District Councils ; and the powers, duties, and 
liabilities are principally those which were conferred by the Public 
Health Act of 1875. 

In the opinion of the author the Government of this country should 
enter into friendly negotiations with the Governments of other countries, 
so that there, might be concerted action to prevent an avoidable 
disease like small-pox. Much good might result from the formation of 
a permanent International Board of Health. If civilisation is not yet 



, CATTLE PLAGUE. 293 

sufficiently advanced to admit of a system of international notification, 
our Consular authorities should he instructed to give immediate notifica 
tion of the existence of small-pox in other countries, and every measure 
should be enforced to diminish the possibilities of importation. The 
duties of a Central Health Office, presided over by a Minister of Health, 
should include the collection of information as to the existence of small- 
pox in other countries, and details should be published in the Annual 
Reports of the Department. Regulations, for example, for dealing with 
the importation of rags from smaU-pox stricken places should be enforced, 
as in the case of cholera ; and if, in spite of these precautions, isolated 
cases occurred in this country, they should be dealt with promptly. 

Notification should be enforced uniformly all over the country, and 
there is not the slightest reason why the authorities and the public should 
not immediately receive information of the existence of small-pox, whilst 
to procure immediate isolation we have only to imitate the excellent 
ambulance system of the Metropolitan Asylums Board. To procure 
prompt notification there must be no loophole for evading the Act, and 
there should be a heavy penalty for failure to notify not only small-pox, 
but any case which may reasonably be supposed to be one of small-pox. 

The police should be required to report any case of small-pox in 
common lodging-houses or shelters ; they should have power to require 
any tramp suffering from small-pox, or from any disease which may 
reasonably be supposed to be small-pox, to be examined by the medical 
officer of the Union, and kept under observation, or transferred at once 
to the isolation hospital ; and inmates of the workhouse should be daily 
inspected, and no case allowed to leave when there is the least suspicion 
of small-pox infection. 

Objections no doubt wiU be raised to this proposal, but the frequency 
with which small-pox is spread by tramps fully justifies these measures. 
All these measures should be carried out as a, matter of routine, and 
without the semblance of panic. 

Isolation should be uniformly enforced all over the country, and 
vaccination should be relegated to the position of a voluntary auxiliary 
measure, which should never be allowed to take the place of sanitary 
regulations to stamp out the disease. 

Cattle Plague. 

Cattle plague is a highly contagious disease of bovines producing 
high fever, and characterised by an eruption with a resemblance to 
human small-pox. The disease is transmissible to other ruminants, 
and is inoculable in man. One attack gjves immunity against 
future attacks. Cattle plague and small-pox are not intercom- 
municable, and are specifically distinct diseases, but the resemblance 
between them was recognised from early times. Ramazzini published 
an account of the cattle pest in Italy in 1711, and described the 
pustules which broke out over the body as similar to those of variola in 



294 INFECTIVE DISEASES. 

kind and appearance. Dr. Layard, in 1780, described this disease of 
horned cattle as an eruptive fever of the variolous kind, with the 
appearance and stages of small-pox. This resemblance vras endorsed 
by Murchison, one of the Commissioners appointed in 1866 to 
inquire into the origin and nature of cattle plague. 

Murchison pointed out that in both diseases the eruption con- 
sisted of pustules and scabs, and that in both it extended from 
the skin to the interior of the mouth and nostrils; in both, the 
pustules and scabs were preceded or accompanied by patches of 
roseola ; in both, they were occasionally interspersed with petechise ; 
and in both, they sometimes left behind pitted scars and discolora- 
tions on the cutis. The other prominent symptoms of rinderpest 
were also those of small-pox — viz., pyrexia, lumbar pain, saUvation, 
and running from the nostrils, alvine flux, albuminuria, hsematuria, 
and " the typhoid st^ate.'' The anatomical lesions of the internal 
organs in rinderpest and immodified small-pox were identical — viz., 
congestion or inflammation of the mucous membranes of the air 
passages and digestive canal, patches of ecchymosis and even 
gangrene of the stomach and other mucous surfaces, and darkly 
coloured blood. In both rinderpest and small-pox the duration 
of the pyrexial stage was on an average about eight days. In 
both diseases a peculiarly offensive odour was exhaled from the 
body before and after death. The two diseases resembled one 
another in their extreme contagiousness, and in the facility with 
which the poison was transmitted by fomites. Both diseases were 
easily propagated by inoculation, and in both cases the inoculated 
disease was milder and less fatal than that resulting from infection. 
In both diseases there was a period of incubation, which is shorter 
when the poison has been introduced by inoculation than when it 
has been received by infection. 

Oeely described the result of an accidental inoculation of cattle- 
plague virus in the human subject. A vesicle was produced which 
so closely corresponded with the result of inoculated cow-pox that 
Oeely inclined to the belief that cattle plague was a malignant form 
of cow-pox. The following is the account of this case as reported 
by Ceely. Mr. Hancock, a veterinary inspector at Uxbridge, was 
engaged in superintending the autopsy of a bullock recently dead of 
cattle plague. His assistant, who was performing the operation, 
while occupied in removing the skin from the scrotum, accidentally 
punctured the back of Mr. Hancock's hand with the point of the 
knife. The puncture being slight was disregarded at the time, but 
was washed as soon as piacticable, and thought of no more. Five 



CATTLE PLAGUE. 295 

days afterwards, a small, slightly elevated, hard pimple was felt 
and seen on the site of the puncture. This gradually advanced 
till the ninth day of the puncture, the fourth from papulation, 
when the enlargement became distinctly vesicular. At that time 
there were but slight constitutional symptoms. On the next day, 
the tenth from the receipt of the puncture, the fifth from papulation, 
and the second from vesiculation, Mr. Hancock consulted Mr. Rayner, 
of TJxbridge, who, on seeing the hand, inquired if the patient had 
been handling the udder of a cow, as he thought he covild recognise 
a cow-pox vesicle of the ninth day. The vesicle was distended 
with thin lymph, its margin elevated and slightly brown, its centre 
depressed and brownish, and the whole surrounded with a large 
bright red areola. There was then considerable tumefaction extend- 
ing from the knuckles to above the wrist. The absorbent vessels 
were considerably inflamed, and, like the axillary glands, were tender 
and painful ; the pulse, naturally slow, was accelerated ; there was 
much pain in the back and limbs, severe distracting headache, etc, ; 
all of which symptoms continued to increase during the two following 
days. At the end of that time the diffused areola had extended as 
far as the elbow. Fifteen days after the puncture, and ten days after 
papulation, the local inflammation and constitutional symptoms had 
partially subsided. The vesicle contained a rather turbid brownish 
fluid, and there were present all the indications of a declining vaccine 
vesicle. 

Murchison also saw and described the case, and gave practically 
the same account of it. He pointed out that the appearances and 
the entire history were very different from the results of a poisoned 
wound, but coincided with the appearances seen after vaccination. 

In 1832 Macpherson, in Bengal, inoculated eleven native children 
with cattle-plague crusts. There was no result in six, others 
suffered from local inflammation, and in one a vesicle formed. 
With lymph from this vesicle other children were inoculated. The 
results in all were similar in appearance to those of vaccination. 
Two children were subsequently inoculated with human variola, and 
were said to be protected. 

In 1834 Macpherson's example was followed by Mr. Eurrell 
in Assam. Furnell inoculated four children with cattle-plague 
crusts without result, but his assistant succeeded with crusts taken 
from the back and abdomen of the diseased cattle, and carried on 
the lymph from child to child. In one case there was a general 
eruption. Furnell inoculated his own child from one of the native 
children : a copious eruption followed, and the child died. Furnell 



296 INFECTIVE DISEASES. 

after this misfortune issued a strong' warning against taking the 
virus from the cow. The experiments were made in the belief that 
cattle plague was really small-pox in cattle, and that the virus 
would protect against human variola. 

Similar results were obtained by Mr. Wood at- Gowalpara 
in 1838. 

Bacteria in Cattle Plague. — Semner cultivated streptococsi 
from the blood and lymphatic glands of a sheep suffering from 
cattle plague. A calf inoculated with a cultivation died in seven 
days. The cocci were stated to lose their virulence by cultivation, 
and the weakened cultivation to protect against the virulent disease. 

The micro-organism was very probably Streptococcus pyogenes, 
and the calf may have died of septic infection. There can be no 
doubt that the nature of the contagium of cattle plague is unknown. 

Protective Inoculation. — In the great epidemic of cattle 
plague in England in 1866, owing to a belief that the analogy 
between cattle plague and small-pox was closer than it really is, 
vaccination with cow-pox was attempted as a preventive measure, 
but was proved to be absolutely useless. 

Stamping-out System. — When cattle plague was imported in 
1865 into London, dairymen and stock-owners made no attempts 
to prevent the extension of the disease, so that it spread rapidly 
all over the country through disposal of infected cattle. The losses 
were enormous, and an Order in Council was passed in July 1865, 
directing dairymen and others to notify outbreaks of any contagious . 
or infectious disease among the animals under their charge. A 
Yeterinary Department of the State was formed, and inspectors 
appointed in various parts of the country. A short Act was passed 
in February 1866. A stamping-out system, consisting of compulsory 
notification and the slaughter of diseased animals, was soon brought 
to the notice of the public. There was violent opposition, but 
nevertheless, after some delay, the system was carried out. The 
number of cases of cattle plague had reached 18,000 weekly, and 
on the introduction of the stamping-out system the disease rapidly 
declined. The disease was again imported into Great Britain in 
1872, and thei-e were outbreaks in 1877. In each instance the 
disease was promptly stamped out, and ever since that year the 
disease has been kept out of this country. 



CHAPTER XXI. 

SHEEP-POX. FOOT-AND-MOUTH DISEASE. 

Sheep-pox. • 

Sheep-pox, or variola ovina, is an acute febrile disease accompanied 
by a general vesiculo-pustular eruption, highly infectious, and 
capable of being propagated by inoculation or clavelisation. It is 
a common disease in some parts of Europe. In France the disease 
is called la clavelee, and in Italy vaociwlo. It has been introduced 
oji several occasions into this country, but has been effectually 
stamped out. As in human small-pox, there are varieties — the 
benign and the malignant, the discrete and the confluent ; and one 
attack is protective against the disease in future. 

It is very closely analogous to human small-pox. Vaccination 
with cow-pox lymph has been employed to protect sheep from sheep- 
pox, but unsuccessfully, and lymph for vaccination has been raised 
from sheep-pox to protect human beings from small-pox. These 
experiments were first performed in Italy. 

Marchelli, in 1802, took lymph from the vesicles of sheep-pox, and 
inoculated children. Sacco repeated these experiments, and found 
there was no appreciable difference from the results obtained with 
cow-pox lymph. Dr. Legni carried on the inoculations with ovine 
virus from arm to arm for several years, and when small-pox 
occurred in Pesaro, it was said that all those who were inoculated 
with the sheep virus were protected. 

Inoculation of children with ovine virus, direct from sheep, was 
repeated by Sacco and Magnani in 1806. 

Marson in England succeeded in producing on the human 
subject a vesicle with the physical characters of the vaccine vesicle. 
The vesicle had a bluer tinge, and subsequent inoculation of the 
patient with human variola was ineffectual. Other experimenters 
were unsuccessful, but their failures, as in the case of variolation of 
the cow, do not invalidate the results of those who were successful. 

297 



298 INFECTIVE DISEASES. 

Sheep-pox and cow-pox are quite distinct diseases. Sheep-pox 
is highly infectious, whereas cow-pox is only conveyed by direct 
inoculation, and is never infectious, and further, cow-pox inoculated 
in sheep does not produce sheep-pox. 

Bacteria in Sheep-pox. — Hallier and Zurn, Klein, and others, 
have found micrococci and bacteria in the lymph of the vesicles of 
sheep-pox, but they are only accidental epiphytes. The nature of 
the contagium is unknown. 

Protective Inoculation. — Extensive experiments were carried 
out in England to test the protective power of vaccination against 
sheep-pox. According to Marson and Simmonds, it was very difficult 
to get cow-pox to take on sheep, and when an effect was produced, 
the resulting affection, even when developed to its fullest extent, was 
veiy unlike the same disease in the human subject. In the sheep 
it seldom produced anything more than a small papule, which occa- 
sionally resialted in the formation of a minute vesicle, or more 
commonly, a pustule, which was sometimes, although very rarely, 
surrounded by a slight areola. Generally, however, neither vesica- 
tion nor pustulation followed, but a small scab was produced, which 
soon fell from the site of the puncture, leaving no trace behind. The 
disease passed quickly and irregularly through its several stages, 
and terminated by the eighth or ninth day, and not unfrequently 
even before that time. Lymph was but rarely obtainable, and then 
only in the smallest quantity, and this on the fifth or sixth day suc- 
ceeding the vaccination. The effects were only local, and the animal's 
health was not impaired. 

Sheep were found to be just as susceptible of the cow-pox virus 
on subsequent repetition of the inoculation as they were in the 
first instance, and hence the conclusion that cow-pox was utterly 
worthless as a protective against sheep-pox. According to Depaul, 
however, cow-pox takes characteristically on sheep, and sheep-pox 
lymph inoculated on cows produces a result indistinguishable from 
the appearances obtained vnth the inoculation of cow-pox lymph. 
It is impossible to say whether these conflicting results depended 
upon the employment in the experiments of different breeds of sheep 
or different stocks of vaccine lymph. 

The objection to clavelisation or ovination is that the disease 
may be introduced in localities where it was previously unknown. 
By ovination, as in the analogous case of variolation, fresh centres 
of infection are created, whereas every precaution should be taken 
to prevent the introduction of the disease. 

Stamping-out System — Sheep-pox has been imported into this 



FOOT-AND-MOUTH DISEASE. 299 

country on several occasions. It was introduced in 1847, and again 
in 1862; in 1865 it was introduced again, and active measures of 
repression were at once taken. The diseased flocks were carefully 
isolated, and day by day as fresh cases occurred the diseased animals 
were killed and buried. Owing to the adoption of these precaution- 
ary measures, the affection did not extend beyond the flock among 
which it first appeared. It was introduced again in 1866 at Long 
Buckby, in Northamptonshire. In this case the disease was exter- 
minated by the slaughter and burial of the whole flock, and imme- 
diate application of disinfectants to the hurdles and other things with 
which the sheep had been in contact. Then it was introduced 
again in Cheshire, and strict isolation being enforced the infection 
died ovit. Since 1866 we have had no outbreak of sheep-pox in 
this kingdom, but foreign sheep have been landed with sheep-pox 
in 1868, 1869j 1870, 1871, 1875, 1876, 1878, and 1880, but the 
disease has been prevented from spreading. 

The Sheep-pox Order of 1895 provides for the notification of 
the disease, for disinfection and for compulsory slaughter of infected 
sheep, and prohibits the movement of diseased or suspected sheep, 
and the local authority may, if they think fit, order the slaughter 
of suspected sheep and of sheep which have been in contact with 
diseased sheep. 

Foot-and-mouth Disease. 

Foot-and-mouth disease is a highly contagious and infectious 
febrile disease, characterised by a vesicular eruption affecting the 
lips, tongue, roof of the mouth, and feet of sheep, cattle, and pigs, 
and according to some observers it also attacks horses, poultry, 
hares, and rabbits. Sometimes the mouth only is affected, in other 
cases the principal seat of the eruption is in the feet. The vesicles 
soon break and give rise to ulcers. When these occur in the mouth 
they cause pain and difliculty in taking food. Extensive ulceration 
may occur on the feet, causing great pain and lameness. In milch 
cows it sometimes happens that the eruption occurs on the udder and 
teats, and it is this manifestation of the disease which has received 
so much attention from Eayer. The milk is contaminated by the 
discharge of the vesicles, and is unfit for use, either as food for the 
human being or for the lower animals. It induces a vesicular 
eruption in the mouth, larynx, pharynx, and intestinal canal. It 
acts most vigorously when administered warm to young animals, 
and calves occasionally die quite suddenly after sucking cows 



300 INFECTIVE DISEASES. 

affected with the eruption on the teats. Fatal effects also result 
when the milk is administered to young pigs. 

It has been stated that no injurious consequences arise from the 
consumption of the milk by human beings, but there is abundant 
evidence to the contrary, and the conflicting opinions probably arise 
from the fact that milk is seldom drunk direct from the cow, and 
rarely in an undiluted form. Hertwig experimented upon himself 
with milk freshly drawn from a cow with the eruption. He drank 
a pint, and two days afterwards experienced sHght fever, restless- 
ness, and headache. The mouth was dry and hot, and there was 
tingling in the skin of the hands and fingers. These symptoms 
continued for seven days after taking the milk. On the ninth day 
vesicles had formed on the tongue, principally on the edges, and on 
the mucous membrane of the cheeks and lips (the largest being 
about the size of a lentil). They were yellowish-white in colour, and 
contained a whitish turbid liquid, which flowed when the vesicles 
were pricked with a needle. At the same time a number of vesicles 
developed on the hands and fingers ; and most of them at the 
time of their first appearance were the size of a millet seed. They 
were firm to the touch, yellowish -white, and occasioned a slight 
tingling. The vesicles of the mouth increased in size and eventually 
broke, and the epithelium detached itself completely from the affected 
parts, leaving dark red spots, which disappeared gradually. The 
slight fever present during the first days ceased after the appearance 
of the eruption ; but from this time, until the disappearance of 
the red spots, Hertwig felt a continual burning pain in the mouth, 
and speaking and deglutition caused considerable uneasiness. On 
the lips the vesicles dried up, and were covered with thin brownish 
crusts, which fell off ten days after the appearance of the first 
vesicles. The vesicles which developed on the hands ran a slower 
course. From the tenth to the thirteenth day they filled with a 
liquid, like turbid lymph. They were large and confluent, and 
finally broke and dried up. 

Bacteria in Foot-and-mouth Disease Klein in 1885 isolated 

from the vesicles a streptococcus which in its microscopical and its 
cultural characters on gelatine, agar and blood serum resembled 
Streptococcus pyogenes. Minute differences in the size of the 
colonies and in their rate of growth, and in the character of the 
chains, were /observed on making comparative cultures with 
Streptococcus pyogenes from a human source, but no comparison 
was made with Streptococcus pyogenes from acute suj)puration in 
cattle.- Baumgarten regarded this micro-organism as Streptococcus 



FOOT-AND-MOUTH DISEASE. 301 

pyogenes, and not as the contagium of the disease. The author has 
pointed out the variation which exists in the size of the chains and 
of the colonies, and the difference which is found in the rate of 
growth of cultures of Streptococcus pyogenes, and these variations are 
especially marked in Streptococcus pyogenes bovis. Klein believes 
that the administrations of broth cultures produced the disease in 
sheep, but the results were very probably due to accidental infection. 
It is well known how very readily foot-and-mouth disease is spread. 
The appearance of a case in a flock of sheep or a herd of cattle will 
be almost certain to be followed by all or nearly all of the other 
animals being infected with great rapidity. The virus clings to the 
clothes of shepherds and others who have been in ^contact with 
infected sheep,' and may be readily conveyed to healthy animals by 
those who have been visiting infected premises. 

SchotteUus described chains composed of rounded elements, some 
of which resembled an amoeba or plasmodivim. The chains were said 
to be motile, and deHcate growths were obtained in blood serum and 
agar, and in broth and on potato. Inoculation in sheep and pigs 
and numerovis small animals gave negative results. These organisms 
were described as streptocytes, to distinguish them from bacteria. 

Piani and Fiorentini investigated the contents of the vesicles, and 
also described coi'puscular elements exhibiting amoeboid movements. 
They regarded these bodies as protozoa, and concluded that foot-and- 
mouth disease is due to their presence. 

Until a micro-organism is cultivated which will produce sheep- 
pox in sheep on a farm or on premises where the disease does not 
exist, and where there can be no possibiHty of accidental infection, 
we are fully justified in concluding that the nature of the contagium 
of this disease is unknown. 

Stamping-out System. — Foot-and-mouth disease was imported 
into this country in 1839. It has been successfully dealt with by the 
stamping-out system, which in this case is very difficult to apply 
because of the very short period of incubation, and the value of 
the stamping-out method very greatly depends upon the length 
of the incubation period. Foot-and-mouth disease very often, 
from infection to recovery, does not exceed ten days ; yet according 
to the reports of the Board of Agriculture, when foot-and-mouth 
disease exists in a manageable state, perfect isolation and effectual 
disinfection have proved equal to the complete control of the 
spreading of the infection, and the final extinction of the disease. 
Nothing more is necessary in any case than to close up all the 
channels through which infected matter can be conveyed ; but in 



302 INFECTIVE DISEASES. 

order that this may be done close supervision by conscientious and 
responsible officers is required ; without it the case is hopeless. 

The Foot-and-mouth Disease Order of 1895 enforces notification, 
isolation, and disinfection, and the question of slaughter is left to 
the local authority. 

(1) A local authority may, if they think lit, cause to be slaughtered — 
(a) Any cattle, sheep, or swine afEected with foot-and-mouth disease 

or suspected of being so afEected ; and 
(J) Any cattle, sheep, or swine being or having been in the same field, 
shed, or other place, or in the same herd or flock or otherwise in 
contact with animals affected with foot-and-mouth disease, or being 
or having been, in the opinion of the local authority, in any way 
exposed to the infection of foot-and-mouth disease. 



CHAPTER XXII. 

hoese-pox. — cow-pox. 

Constitutional Geease or Horse-pox. 

Horse-pox is- a vesicular disease of the horse communicable from 
animal to animal by inoculation, but never infectious. It is 
communicable by inoculation to man, and the attenuated virus 
produces phenomena indistinguishable from the results of vaccina- 
tion with cow-pox lymph. 

The existence of this disease of the horse had long been known 
to farmers and fai-riers, but Jenner was the first to draw attention 
to it in writing. " There is a disease to which the horse from his 
state of dome.stication is frequently subject. The farriers have 
termed it the grease ; it is an inflammation and swelling in the heel 
accompanied at its commencement with small cracks and fissures, 
from which issues matter possessing properties of a very peculiar 
kind." Jenner gave several instances in which this disease was 
communicated to man and to cows. 

Thus, a man named Merret attended to some horses with sore 
heels and also milked the cows. The cows were infected, and the 
man had several sores upon his hands. 

William Smith, on another farm, attended to hoi'ses with sore 
heels and milked the cows also. The cows were infected, and on one 
of Smith's hands there were several ulcerated sores. 

Simon NichoUs applied dressings to the sore heels of one of his 
master's horses and at the same time milked the cows, and the cows 
were infected in consequence. 

A mare, the property of a dairy farmer, had sore heels, and 
was attended to by the men of the farm, Thomas Virgoe, William 
Wherret, and William Haynes. They contracted " sores on their 
hands, followed by inflamed lymphatic glands in the arms and 
axiUse, shiverings succeeded by heat, lassitude, and general pains in 
the limbs," and the disease was also communicated to the cows. 

303 



304 INFECTIVE DISEASES. 

But Jenner's experience of this disease was not limited to cases 
in which the eruption occurred in the heel. 

He mentions a case in which — 

" An extensive inflammation of the erysipelatous kind appeared 
without any cause upon the upper part of the thigh of a sucking 
colt. The inflammation continued several weeks, and at length 
terminated in the formation of three or fovir small abscesses." Those 
who dressed the colt also milked the cows on the farm, and communi- 
cated the disease to them. 

Subsequently, Jenner gave a more comprehensive description of 
this disease. 

" The skin of the horse is subject to an eruptive disease of a 
vesicular character, which vesicle contains a limpid fluid, showing 
itself more commonly in the heels. The legs first become (Edematous, 
and then fissures are observed. The skin contiguous to these fissures, 
when accurately examined, is seen studded vnth small vesicles sur- 
rounded by an areola. These vesicles contain the specific fluid. It 
is the ill-management of the horse in the stable that occasions the 
malady to appear more frequently in the heel than in other parts. 
I have detected it connected with a sore on the neck of the horse, 
and on the thigh of a colt." 

Mr. Moore, of Chalford Hill, described a case in 1797, and re- 
garded the disease as virulent grease. His horse was attacked with 
what was supposed to be ordinary " grease.'' A cow was subse- 
quently infected, and the disease communicated to the servant, who 
had " eruptions on his hands, face, and many other parts of the 
body, the pustules appearing large, and not much unlike the small- 
pox, for which he had been inoculated a year and a half before, and 
had then a very heavy burden." 

In 1798, Mr. Fewster, of Thornbury, met with a case of this 
equine malady, and wrote a very full account to Jenner of its 
transmission to the human subject. 

"William Morris, aged thirty-two, servant to Mr. Cox of 
Almonsbury in this county, applied to me the 2nd of April, 1798. 
He told me that four days befor&he found a stiffness and swelling 
in both his hands, which were so painful it was with difficulty he 
continued his work ; that he had been seized with pain in his head, 
small of the back, and Hmbs, and with frequent chilly fits succeeded 
by fever. On examination I found him still affected with these 
symptoms, and there was great prostration of strength. Many 
parts of his hands on the inside were chapped, and on the middle 
joint of the thumb of the right hand there was a small phagedsenic 



CONSTITUTIONAL GREASE OR HORSE-POX. 305 

ulcer, about the size of a large pea, discharging an ichorous fluid. 
On the middle finger of the same hand there was another ulcer of a 
similar kind. These sores were of a circvilar form, and he described 
their first appearance as being somewhat like blisters arising from a 
burn. He complained of excessive pain, which extended up his arm 
into the axilla. On the 5th of April I again saw him, and found 
him still complaining of pain in both his hands, nor were his febrile 
symptoms at all relieved. The ulcers had now spread to the size of 
a seven-shilling gold coin, and another ulcer, which I had not noticed 
before, appeared on the first joint of the forefinger of the left hand, 
equally painful with that on the right. I ordered him to bathe his 
hands in warm bran and water, apply escharotics to the ulcers, and 
wrapped his hands up in a soft cataplasm. The next day he was 
much relieved, and in something more than a fortnight got well. 
He lost his nails from the thumb and fingers that were ulcerated." 

Mr. Tanner, a veterinary surgeon, was the first to succeed in 
experimentally transmitting horse-pox to the teats of a cow by 
inoculating some of the liquid matter from the heel of a horse. From 
handhng the cow's teats he became infected himself, and had two 
pustules on his hand, which brought on inflammation, and made 
him unwell for several days. The matter from the cow and from his 
own hand proved efiicacious in infecting both human subjects and 
cattle. 

In 1801 Dr. Loy published his experiments. A butcher had 
painful sores from dressing a horse suffering from ' grease,' and Dr. 
Loy succeeded in transmitting the disease to the udder of a cow. 
Matter was taken from the cow and inserted into the arm of a child. 
Dr. Loy also inoculated a child direct from a horse suffering from 
' grease,' and subsequently five other children from this child. 

From his experiments and observations Dr. Loy was led to 
differentiate constitutional grease from the merely local affection 
commonly known as the grease, and thus he explained the failure 
on the part of many experimenters to transmit this disease to 
the cow. 

" This fact induces me " — he says — " to suspect that two kinds 
of grease exist, differing from each other in the power of giving 
disease to the human or brute animal ; and there is another circum- 
stance which renders this supposition probable. The horses that 
communicated the infection to their dressers were affected with a 
general as well as a topical disease. The animals at the commence- 
ment of their disease were evidently in a feverish state, from which 
they were relieved as soon as the complaint appeared at their heels,. 

20 



306 INFECTIVE DISEASES. 

and an eruption upon the skin. The horse, too, from which the 
infectious matter was procured for inoculation, had a considerable 
indisposition, previous to the disease at his heels, which was attended, 
as in the others, with an eruption over the greatest part of his 
body ; but those that did not communicate the disease at all, had 
a local affection only. From this perhaps may be explained the 
want of success attending the experiments of the gentlemen I have 
mentioned." 

Experiments with horse-pox were also made about this time on 
the Continent. Sacco made some observations upon this disease at 
Milan. Several horses were suffering from what was called giardoni, 
and Sacco's servant was attacked on both arms, from dressing one 
of his horses troubled with this disease. Several children and cows 
were inoculated from the horses, but without success. In another 
instance, a coachman went to the hospital with the eruption on his 
hands, and the disease was successfully communicated to three out 
of nine children. 

In 1803 Dr. Marcet described some experiments which had been 
made at Salonica by M. La Font. The disease was known to the 
farriers in Macedonia as java/rt. In one case, a horse was attacked 
with feverish symptoms that ceased as soon as the eruption appeared. 
The fore legs were much swelled and several ulcers formed. M. La 
Font took some of the discharge from an ulcer and inoculated a cow. 
and three children, and succeeded in transmitting the disease to two 
of the latter. 

Vaccinogemc grease was observed in Paris in 1812, and Baron 
cites the case of a coachman who, after dressing a horse with the 
" grease," had a crop of pustules on his hands, from which the disease 
was experimentally transmitted by inoculation to two children. A 
series of inoculations was started from an infant who was infected 
from one of the scabs taken from the pustules on the hand of the 
coachman. 

In 1813 Mr. Melon, a surgeon at Lichfield, met with vaccinogenic 
grease in the horse, and some of the virus was sent to Jenner, who 
carried on a series of arm to arm equinations for some months. And 
again in 1817, vaccinogenic grease broke out in a farm at Wansell. 
The farm-servants and the cows were infected, and Jenner employed 
this equine matter for a series of inoculations for eight months. 

In 1817 Baron described a case of a young man who had not less 
than fifty pustules on his hands and wrists from dressing a horse with 
this disease, and in the following year Baron obtained some fresh 
equine virus from the hands of a boy who had been infected directly 



CONSTITUTIONAI/ GREASE OR HORSE-POX. 307 

from a horse. The disease assumed a pustular form, and extended 
over both arms. 

In 1818 Kahlert met with this equine disease in Bohemia, and 
confirmed the experiments made by Loy and Saccd. Kahlert noticed 
that the joint of the foot was swollen, and moisture exuded from it, 
and that the posterior part of the pastern was slightly red, swollen 
and hotter than the neighbouring parts, and a clear yellowish fluid 
with a peculiar odour escaped. At the slightest touch the animal 
showed signs of pain ; the hair was stuck together. The disease was 
successfully transmitted to cows and from cows to children. 

In 1860 the horses at Rieumes, near Toulouse, were attacked by 
an epizootic malady ; in less than three weeks there were more than 
one hundred cases. According to the veterinary surgeon, M. Sarrans, 
the animals suffered from slight fever, rapidly followed by local 
symptoms, the most marked of which were swelling of the hocks, and 
an eruption of small pustules on the surface of the swollen parts, 
which were, at the same time, hot and painful. After three to five 
days there was a discharge from the pastern which continued for eight 
to ten days, during which the inflammation gradually diminished. 
The pustules dried up, and in about a fortnight the crusts with patches 
of hair fell off, leaving more or less marked scars. The eruption 
appeared at the same time on different parts of the body, especially 
on the nostrils, h'ps, buttocks, and vulva. Sarrans beheved that the 
mares taken to the breeding establishment at Rieumes had been 
infected from the ropes which had been used in tying up other aflPected 
animals, and had become thereby infected with the virus of this 
disease. One of the mares was taken by the owner, M. Oorail, to 
the veterinary school to be examined by M. Lafosse. About eight 
days after this visit significant symptoms appeared : loss of appetite, 
lameness, stiffness of both pastern joints, and a hot, painful swelling 
of the left pastern joint. The hair was staring, and there were 
vesicles on the skin, from which a liquid exuded having an ammoniacal 
odour but less foetid than the secretion in eaux aux jarnbes. 

M. Lafosse successfully transmitted the disease to cows, and from 
cows to children and to a horse. 

In 1863 the subject of vaocinogenic grease or horse-pox again 
received great attention in France. A student named Amyot was 
engaged in dressing a horse on which an operation had been per- 
formed. The leg which had been operated on became the seat of a 
very confluent eruption of horse-pox, which was followed by such 
an abundant flow of serosity that at first the nature of the affection 
was mistaken, and it was thought to be a complication of eaux aux 



308 INFECTIVE DISEASES. 

jambes. Amyot had a wound on the dorsal aspect of the first inter- 
phalangeal joint of the Uttle finger of his right hand ; in spite of 
this he continued to dress, the horse entrusted to his care. The 
wound on his finger became accidentally inoculated with the virus, 
which flowed in great abundance from the horse's leg. 

The wound was made on August 3rd, and the next day it was 
swollen, and rather painful. On the 5th, Amyot suffered from malaise 
and great weakness; on the 6th, 7th, and 8th, vesicles appeared 
successively on the fingers of his left hand, and on his forehead 
between the two eyebrows. On the 9th, these vesicles were fully 
developed; those of the fingers consisted of very large epidermic 
bullse on a bluish-red base. On opening them, a perfectly limpid 
fluid escaped in such abundance that small test-tubes might have 
been filled with it. The vesicle on the forehead was surrounded by 
a bluish-red areola, within which, the epidermis, of a leaden-grey 
hue, was raised, and had a slight central depression. The hquid 
which flowed from it when it was opened, and which continued to 
ooze, was also very abundant and of a deep citrine colour. 

The vesicles which had developed on the dorsal side of Amyot's 
fingers were extremely painful. The incessant shooting pains, of 
which they were the seat, prevented him from getting any rest for 
three days. On the 10th, inflammation of the lymphatics followed ; 
both arms were swollen and very painful, with red lines indicating 
the course of the lymphatic vessels. The glands of the axillae were 
also enlarged. 

The lymphatic glands behind the jaws were also swollen and pain- 
ful. Amyot's chief sufferings were occasioned by the intense local 
pain caused by the vesicles on the fingers, and by the infiammation 
of the lymphatic vessels and glands, which continued in this state up 
to the 18th of August. It was only at the end of the month that the 
vesicles were completely cicatrised. 

Bouley felt very great anxiety in the presence of the grave 
symptoms which accompanied the eruption. The eruption on the 
forehead was especially a cause of great uneasiness, because glanders 
manifests itself in a similar way. 

With virus from Amyot's vesicles the disease was transmitted to 
cows and to children. 

FiTrther, this outbreak enabled exhaustive experiments to be 
made, by which it was definitely established that horse-pox is never 
infectious, but, like cow-pox, is transmitted solely by contact. 

In 1880, M. Baillet, Director of the National Veterinary School 
of Toulouse, was informed that a contagious malady had developed 



CONSTITUTIONAL GREASE OR HORSE-POX. 309 

in the mares, which had been served by the stallions at the breeding " 
establishment at Eieumes, belonging to M. Mazferes. M. Peuch 
was delegated to investigate this outbreak, and he visited for that 
purpose Berat, Rieumes, and Labastide-Olermont. 

At Berat three mares were examined. In one, there were scars 
and crusts, the remains of an eruption on the lips and in the vicinity 
of the vulva ; in another, there were several reddish circular ulcers 
in the same region ; and in a third, there were dried pustules with 
blackish adherent crusts at the circumference of the vulva and 
extending over the perinseum. On the lower part of the left flank 
a vesicle was discovered surmounted by a crust, and when the latter 
was detached a sero-sanguinolent liqiiid oozed from the exposed 
surface. M. Peuch recognised the true nature of this disease, 
having several times previously had the opportunity of examining 
mares with a vesicular eruption round the vulva after coition, 
which eruption he had studied from its first appearance to complete 
cicatrisation, and had ascertained to be horse-pox. 

On proceeding to Eieumes, M. Peuch inspected eleven stallions, 
six horses, and five asses. In one ass there were several vesicles 
on the right side of the penis scattered about from the base to the 
glans. In another ass there was a trace of a vesicle on the penis 
and a characteristic vesicle on the left nostril. 

In an old bay mare there were the remains of an eruption 
on the circumference of the vulva, and in an old white mare there 
were not only vesicles on the vulva, but in addition vesicles on the 
inner side of the lower lip. M. Peuch drew special attention to 
these cases as likely to be confounded with aphthous stomatitis, but 
the existence of the same eruption on other parts of the body is an 
important aid in making a diagnosis of horse-pox. 

At Labastide-Olermont one mare was particularly noticed. 
This mare had been served on the 19th and 21st of April, and on the 
occasion of the inspection. May 11th, there were the remains of an 
eruption around the vulva, and lymphangitis existed in the right 
posterior limb, which was engorged, hot, and painful in its whole 
extent, so that the animal walked with difiiculty. The proprietor 
had contracted the disease in attending to his mare, and exhibited 
a vesicle on the thumb of the right hand, excoriated and blackened, 
but still recognisable. 

Some of the crusts collected from the cases at Berat were used 
for inoculating a cow. The result was successful, and the disease 
was transmitted by inoculation to a heifer and several students and 
children. 



310 INFECTIVE DISEASES. 

M. Peuch ascertained that horse-pox caused considerable alarm 
from the fact that the breeders regard this eruptive affection as 
syphilitic, and this alarm consequently brings discredit upon the 
breeding establishment whence the iUness has spread. He was also 
led to appreciate the great necessity for further study of this disease 
in relation to dourine or maladie du coit. 

In 1882 M. Peuch had the opportunity of investigating a case 
of horse-pox in Algeria. The disease occurred in a thoroughbred 
Arab. There was an eruption of vesicles, and there was also an 
ulcer the size of a five-franc piece in the nosti-U. In the mouth and 
on the lips there were a number of small vesicles about the size of 
a pea. The sublingual glands were engorged, hot, and painful on 
pressure. The coat, in patches, on the lateral aspect of the neck, 
on the shoulders, the flanks, and in the hollow of the heel, was 
staring, giving the appearance of small paint-brushes. On passing 
the hand over these, vesicles could be detected partly dry and partly 
secreting. 

The disease was transmitted to cows, and from cows to about 
one thousand five hundred persons. 

Cases similar to the one just described, in which there is more or 
less marked ulceration of the nostril or nasal septum, must be care- 
fully distinguished from glanders. And again, when the sublingual 
glands are affected the disease may be mistaken for strangles. 

Nature and Affinities. 

Horse-pox and human small-pox are quite distinct diseases, 
and the theory that horse-pox is derived from grooms or other 
attendants suffering from small- pox may be disinisseji without 
further comment. 

Horte-pox is never infectious, but is communicated solely by 
contact — either by grooms inoculating the virus with their hands, 
sponges, or brushes, or by horses coming into contact with each other, 
and in breeding establishments by coition. Auzias Turenne, who 
wrote exhaustively on this subject, maintained that horse-pox came 
into the same category of diseases as syphilis in man. 

"A un point de vue, le grease pustuleux inocule offre la plus 
parfaite ressemblance avec la verole inoculee, par le produit des 
accidents secondaires. Des deux c6tes nous voyons, absence de 
contagion par la voie de I'atmosphfere, travaU local, retentissement 
lymphatique et ganglionaire, fermentation universelle de I'organisme, 
eruption g^nerale et immunite acquise contre de nouvelles atteintes. 

"A un auti-e point de vue, la ressemblance avec la variole est 



NATURE AND AFFINITIES. 311 

frappante. Mais il s'en distingue ^noriuement par I'absence de la 
contagiosite atmosph^rique.'' 

Human small-pox belongs to a different group of diseases, and 
has affinities rather with small-pox of sheep and cattle plague, 
diseases which are not only inoculable, but highly infectious. Human 
small-pox is an infectious disease characterised by sudden and severe 
fever, followed after forty-eight hours by a generalised eruption ; horse- 
pox commences as a local affection, and constitutional symptoms 
follow. Auzias Turenne, guided by analogy, described the general- 
ised eruptions following "grease" or horse-pox as greasides {'^comme 
on dit syphilides"). 

Horse-pox and human syphilis are absolutely distinct diseases ; 
and there is no more ground for believing that horse-pox originates 
in humaa syphilis than there is for accepting the theory that it arises 
from grooms suffering from small-pox. Syphilis artificially inocu- 
lated on the human subject only resembles the casual or intentional 
inoculation of virulent horse-pox. The stages of papulation, vesicu- 
lation, ulceration, scabbing, and the formation of a permanent scar, 
occur in inoculated syphihs, and if we examine Ricord's illustrations 
and study the experiments of Auzias Turenne, we cannot fail to 
be struck with the remarkable similarity to the results obtained and 
depicted by Jenner. 

But in order to follow the argument of Auzias Turenne we 
must study the natural and casual horse-pox. And if we are not 
familiar with what has been written on this subject, and if we 
restrict our knowledge to the artificially cultivated horse-pox, we 
shall fail to recognise the disease when we meet with it, and we shall 
be liable to attribute the results of the full effect of the virus to 
' accidental contamination. 

Another question of very great interest is the relation of horse- 
pox to cow-pox. Jenner first of all propounded the theory that all 
cow-pox arose from horse-pox, or as he termed it " the grease," and 
thus cow-pox and horse-pox were manifestations of the same disease. 
But it was established that cow-pox also arose quite independently 
of horse-pox, and Jenner was led to distinguish between cow-pox, 
a disease peculiar to the cow, and the eruptive affection transmitted 
to the cow from the horse, which farmei-s and others, by a strange 
perversion of terms, called the cow-pox. Whether the eruption of 
cow-pox can be distinguished from the eruption of horse-pox com- 
municated to the cow, and whether cow-pox and horse-pox are 
identical, or only analogous, are questions which call for further 
investigation. 



312 INFECTIVE DISEASES. 

Bacteria in Horse-pox. — Outbreaks of horse-pox have not 
been investigated from the bacteriological point of view, and the 
nature of the contagium is unknown. 

Cow-pox. 

Cow-pox is a vesicular disease of the teats of cows. It is never 
infectious, and only attacks cows in milk, the virus being transferred 
from cow to cow by the hand of the milker. The disease is com- 
municable to milkers, and the virus artificially inoculated produces 
what is commonly known as vaccinia. In its chnical history and 
epidemiology cow-pox is totally distinct from human small-pox, and 
the hypothetical and entirely erroneous suggestion that the disease 
arises from milkers suffering from human small-pox is responsible 
for the belief which prevailed until recently, that cow-pox was an 
extinct disease in this country ; but, by the author's researches, this 
has been shown to be a mistake. 

Cow-pox is not a rare disease, and it has never been found to 
arise from a milker suffering from small-pox. As this is a matter 
of great importance in discussing the etiology of the disease, the 
history of outbreaks and the clinical characters of cow-pox will be 
given in considerable detail. 

According to Jenner, cow-pox had been known among farmers 
from time immemorial. He refers to cases occurring in 1770, 1780, 
1782, 1791, 1794, 1796, and 1798. In 1799 cow-pox was raging in 
the dairies in London, and outbreaks were investigated by Woodville, 
Pearson, and Bradley. In the same year cow-pox broke out at 
Norton Nibley, in Gloucestershire. Pearson and Aikin referred to 
the prevalence of cow-pox in Wilts, Somerset, Devon, Bucks, Dorset, 
Norfolk, Suffolk, Leicestershire, and Staffordshire ; and Barry men- 
tioned its prevalence in Ireland. 

From this time onwards, for a long period, natural cow-pox 
received little or no attention -in this country. Fresh stocks of lymph 
were raised for the purposes of vaccination, but no further attention 
was given to studying the disease in the cow. In 1836 Leese 
described an outbreak of cow-pox, and m 1838 Estlin discovered an 
outbreak in Gloucestershire. In 1838-39 cow-pox was met with 
by Mr. Fox, of Cerne Abbas, and again in 1839, in Dorsetshire, by 
Mr. Sweeting. Ceely frequently met with cow-pox in the Vale of 
Aylesbury, and particularly refers to outbreaks in 1838, 1840, 1841, 
and 1845. But after this, outbreaks of this disease in the cow were 
not recorded, though several medical practitioners met with the 
disease and raised fresh stocks of vaccine lymph. Thus, when 



COW-POX. 313 

inquiries were made in 1857, it was found that Mr. Donald 
Dalrymple, of Norwich (on two occasions), Mr. Beresford, of Nar- 
borough, in Leicestershire, Mr. Gorham, of Aldeburgh, Mr. Alison, 
of Great Retford, Mr. Coles, of Leckhampton, Mr. Rudge, of 
Leominster, and one or two others, had met with outbreaks of 
cow-pox. 

In 1885 cow-pox was discovered by the author in Wiltshire. The 
publication of the fact led to the recognition of the disease in the 
same year in many parts of England, and cases were met with in 
man in 1888 by Mr. Forty in Gloucestershire, and by Mr. Bucknill 
near London in 1894. 

In Italy, cow-pox was found by Sacco in the plains of Lombardy 
in 1800, and by other practitioners in 1808-9. In 1812 it was 
observed at Naples by Miglietta ; in 1830 in Piedmont ; and in 1832 
and 1843 at Rome, by Dr. Maceroni. More recently, several out- 
breaks of cow-pox have been met with in this country, and the stocks 
of vaccine lymph renewed. 

In France, in 1810, cow-pox was found in the department of 
La Meurthe, and in 1822 at Clairvaux ; at Passy, Amiens, and 
Rambouillet in 1836 ; at Rouen in 1839 ; at St. Illide, at St. Seine, 
and at Perylhac, in 1841 ; in 1842 at Pagnac ; in 1843 at Deux 
Jumeaux, where, during the previous thirty years, several fresh 
stocks of lymph had been raised and circulated. The disease occurred 
in a cow belonging to M. Majendie in 1844, and it was fovind at 
Wasseloune, in the department of Bas Rhin, in 1845 ; it occurred in 
three other departments in 1846 ; at Rheims, and in the department 
of Eure et Loire, in 1852 ; in the arrondissement of Sancerre, and at 
Beziers in 1854; and at Guyonville in 1863. It broke out on farms 
in three villages near Nogent in 1864 (the disease was introduced by 
newly purchased cows ; milkers were infected, and from one of these 
milkers a lymph stock was established) ; it also occurred in 1864, at 
Petit Quevilly, near Rouen; and in April 1866 at Beaugency; in 
1881 at Eysines, near Bordeaux, and again at the same place in 
1883 ; and in 1844 at Cerons. 

In Germany, as soon as attention had been drawn to the disease, 
cow-pox was frequently discovered. There were as many as thirty- 
eight outbreaks reported in one year in Wurtemberg. 

It is hardly necessary, after reciting these instances, to insist 
that cow-pox is far from being a rare disease, as many have sup- 
posed who are unacquainted with the literature of the subject and 
unfamiliar with the appearances of the natural disease in the 
cow. 



314 INFECTIVE DISEASES. 



Natural and Casual Cow-pox. 



To appreciate the characters of the natural disease in the cow, 
we must dismiss from our minds the artificial disease vaccinia, for 
the ordinary results of vaccination stand in much the same relation 
to the natural disease cow-pox as the benign vesicle of variolation to 
natural small-pox. 

The description of cow-pox given by Jen'ner, in 1798, was the 
first pubhshed account. The disease in the cow was described as 
consisting of irregular pustules on the teats, of a palish blue colour, 
surrounded by an erysipelatous inflammation, and characterised by 
a tendenoj' to degenerate into phagedaenic ulcers. The animals were 
indisposed and the secretion of milk lessened. 

In referring to an outbreak which occurred epizobtically in 
London in February 1799, Dr. Bradley gave a coloured plate of the 
disease on the arm and fingers of a milker. The cow-pox, he said, 
in this instance, " appears to have been very mild, for no loss was 
experienced by the fai-mers from the deficiency of mUk, as usually 
happens.'' 

These early descriptions were supplemented by an account of 
cow-pox by Mr. Lawrence, author of A Philosophical and Practical 
Treatise on Horses, and on the Moral Duties of Man towa/rd the Brute 
Creation. Lawrence's article on cow-pox not only affords evidence 
that this disease was known to those who had the care of cattle 
before Jenner's paper was published, but it shows that it had also 
been made the subject of practical observation and study by veteri- 
narians. Lawrence concluded by saying : " "Whatever may be the 
fate of cow-pox inoculation, it has and will give further occasion to 
a pretty large and open discussion, which is always beneficial as 
having a tendency to produce discovery and promote improvement ; 
and when the public ardour for the present topic shall have become 
a little cool and satisfied, I hope it will be turned by enlightened 
men towards, another, perhaps of nearly as great consequence — 
namely, the prevention of the original malady in the animals them- 
selves. Those who have witnessed it and only reflected upon the 
excessive filth and nastiness which must unavoidably mix with the 
milk in an infected dairy of cows, and the corrupt and unsalubrious 
state of their produce in consequence, will surely join me in that 
sentiment.'' 

Lawrence was almost a century before his time. Cow-pox was not 
again brought for-Js^ard in this light until 1887-88, when the author 
reported the contamination of the milk at the Wiltshire farms and 



NATURAL AND CASUAL COW-POX. 315 

advocated the advisability of placing this disease under the Con- 
tagious Diseases (Animals) Act. 

The numerous pathological details wanting in the early accounts 
of cow-pox were supplied by the painstaking and laborious re- 
searches of Robert Oeely. From his classical papers in the Trans- 
actions of the Px'ovincial Medical Association, we can obtain a 
complete picture of the natural disease in the cow. 

In Ceely's experience in the Vale of Aylesbury, outbreaks 
occurred at irregular intervals, most commonly appearing about 
the beginning or end of the spring, rarely during the height of 
summer. There were outbreaks at all periods from August to May 
and the beginning of June, cases being met with in autumn and 
the middle of winter, after a dry summer. The disease was occa- 
sionally epizootic, or occurring at times at several farms at no great 
distance from each other, but was more commonly sporadic or 
nearly solitary. It was to be seen sometimes at several contiguous 
farms ; at other times at one or two farms. Many years might 
elapse before it recurred at a given farm, although all the animals 
might have been changed in the meantime. Cow-pox had broken 
out twice in five years in a particular vicinity at two contiguous 
farms, while at an adjoining dairy, in all respects similar in local 
and other circumstances, it had not been known to exist for forty 
years. It was sometimes introduced into a dairy by recently 
purchased cows. Twice it had been known to be so introduced by 
milch heifers. It was considered that the disease was pecuhar to 
the milch cow ; it came primarily while the animal was in milk, 
and it was casually propagated to others by the hands of the milkers. 
Sturks, dry heifers, dry cows, and milch cows milked by other hands, 
grazing in the same pastures, feeding in the same sheds, and at 
contiguous stalls, remained exempt from the disease. 

For many years, the " spontaneous " origin of cow-pox had 
not been doubted in the Vale of Aylesbury. In all the cases that 
Ceely had noticed he could never discover the probability of any 
other origin. 

Condition of Animal primm-ily affected. — There was much diffi- 
culty in determining at all times, with precision, whether this 
disease arose primarily in one or more individuals in the same dairy. 
Most commonly, however, it appeared to be solitary. The milkers 
believed they were able to point out the infecting individual. In. 
two instances, there could be very httle doubt on this point. In 
August 1838, three cows were affected with the disease. The first 
was attacked two months after calving and seven weeks after 



316 INFECTIVE DISEASES. 

weaning. This animal was considered in good health, but it looked 
out of condition. Heat and tenderness of the teats and udder were 
the first noticed signs. The other two were affected in about ten 
days. In December 1838, in a large dairy, a milch cow sHpped her 
calf, had heat and induration of the udder and teats, with cow-pox 
eruption, and subsequently leucorrhoea and greatly impaired health ; 
the whole dairy, consisting of forty cows, became subsequently 
affected, and also some of the milkers. In another dairy, at the 
.same time, it first appeared in a heifer soon after weaning, and in 
about ten or twelve days extended to five other heifers and one cow, 
milked in the same shed, affecting the milkers. And in another 
dairy thirty cows were severely affected, and also one of the milkers. 
It appeared to originate in a cow two months after calving. The 
only symptoms noticed were that the udder and teats were tumid, 
tender, and hot just before the disease appeared. 

Condition of Animals casually affected. — In some animals, it was 
less severe than in others, depending on the state and condition of 
the skin of the parts affected, and the constitution and habit of the 
animal. It was sometimes observed to diminish the secretion of 
milk, and in most cases it commonly did actually affect the amount 
artificially obtained ; with this exception, and the temporary trouble, 
and accidents to the milk and the milkers, little else was observed ; 
the animal continued to feed and graze apparently as well as before. 
The topical effects varied ver}' much in different individuals ; the 
mildness or severity being greatly influenced by temperament and 
condition of the animal, and especially by the state of the teats and 
udder, and the texture and vasculaiity of the skin of the parts 
affected. Where the udder was short, compact, and hairy, and the 
skin of the teats thick, smooth, tense, and entire, or scarcely at all 
chapped, cracked, or fissui'ed, the animal often escaped with a mild 
affection, sometimes with only a single vesicle. But where the 
udder was voluminous, flabby, pendulous, and naked, the teats 
long and loose, and the skin corrugated, thin, fissured, rough, and 
unequal, then the animal scarcely ever escaped a copious eruption. 
Hence, in general, heifers suffered least, and cows most, from the 
milkers' manipulations. 

Progress of the Disease. — Cow-pox once arising or introduced, and 
the necessary precavitions not being adopted in time, appeared in ten 
or twelve days on many more cows in succession, so that among 
twenty-five cows perhaps by the third week nearly all would be 
affected ; but five or six weeks or more were required before the 
teats were perfectly free from the disease. 



NATURAL AND CASUAL COW-POX. 317 

Propagation hy the Hand of the Milker. — Ceely was able to confirm 
the way in which the disease was said to spread. In December 
1838, on a large dairy farm, where there were three milking-sheds, 
cow-pox broke out in the home or lower shed. The cows in this 
shed being troublesome, the milker from the upper shed, after 
milking his own cows, came to assist in this for several days, morning 
and evening, when in about a week some of his own cows began to 
exhibit the disease. It appears that, having chapped hands, he 
neglected washing them for three or four days at a time, and thus 
conveyed the disease from one shed to another. During the progress 
of the disease through this shed, one of the affected cows, which had 
been attacked by the others, was removed to the middle shed, where 
all the animals were perfectly well. This cow, being in an advanced 
stage of the disease, and of course difficult to milk and dangerous 
to the milk-pail, was milked first in order by the juvenile milker for 
three or four days only, when, becoming unmanageable by him, its 
former milker was called in to attend exclusively to it. In less than 
a week, all the animals of this shed showed symptoms of the disease, 
though in a much milder degree than it had appeared in the other 
sheds, fewer manipulations having been performed by an infected 
hand. 

Topical Sym2}toms of the Natural Disease. — For these, Ceely was 
almost always, in the eax-ly stage, compelled to depend on the obser- 
vations and statements of the mUkers. They stated that for three 
or four days, without any apparent indisposition, they noticed heat 
and tenderness of the teats and udder, followed by irregularity and 
pimply hardness of these parts, especially about the bases of the 
teats and adjoining the vicinity of the udder ; these pimples on skins 
not very dark are of a red colour, and generally as large as a vetch 
or a pea, and quite hard, though in three or four days many of 
these increase to the size of a horse-bean. Milking is generally 
very painful to ' the animal ; the tumours rapidly increase in size, 
vesicate, and are soon broken by the hands of the milker. Milking 
now becomes a ti-oublesome and occasionally a dangerous process. 
Ceely adds : " It is very seldom that any person competent to 
judge of the nature of the ailment has access to the animal before 
the appearance of the disease on others of the herd, when the cow 
first affected presents on the teats acuminated, ovoid, or globular 
vesications, some entire, others broken, not infrequently two or three 
interfluent ; those broken have evidently a central depression with 
marginal induration ; those entire, being punctured, diffuse a more 
or less viscid amlior-coloured fluid, collapse, and at once indic3,te the 



318 INFECTIVE DISEASES. 

same kind of central and marginal character. They appear of 
various sizes, from that of a pin's head, evidently of a later date, 
either acuminated or depressed, to that of an almond or a filbert, or 
ever larger. Dark brown, or black, sohd, uniform crusts, especially 
on the udder near the base of the teats, are visible; at the same time, 
some much larger are observed on the teats; these, however, are 
less regular in form and less perfect. Some are nearly detached, 
others quite removed, exhibiting a raw surface with a slight central 
slough. On the teats, the crusts are circular, oval, oblong, or 
irregular; some flat, others elevated, some thin and more trans- 
lucent, being obviously secondary. The appearance of the disease 
in different stages, or at least the formation of a few vesicles at 
different periods, seems very evident. The swollen, raw, and en- 
crusted teats seem to produce uneasiness to the animal only while 
subjected to the tractions of the milkers, which it would appear are 
often nearly as effectual as usual." Referring again to the character 
of the vesicle, Ceely says, that " those fortunate enough to have an 
opportunity of watching the disease in its progress may observe 
that, when closely examined, they present the following characters : 
In animals of dark skin, at this period, the finger detects the 
intumescent indurations often better than the eye, but when closely 
examined the tumours present at their margins and towards their 
centres a glistening metallic lustre or leaden hue ; but this is not 
always the case, for occasionally they exhibit a yellowish or yellowish- 
white appearance." 

In describing the crusts in detail, Oeely says that " large black 
solid crusts, often more than an inch or two in length, are to be 
seen in different parts of these organs, some firmly adherent to a 
raw elevated base, others partially detached from a raw, red, and 
bleeding surface; many denuded, florid, red, ulcerated surfaces, with 
small central sloughs secreting pus and exuding blood, the teats 
exceedingly tender, hot, and swollen. ... In some animals, under 
some circumstances, this state continues Kttle altered till the third 
or fourth week, rendering the process of milking painful to the 
animal, and difficult and dangerous to the milker." 

" In many, however, Kttle uneasiness seems to exist. The parts 
gradually heal ; the crusts, although often partially or entirely 
renewed, ultimately separate, leaving apparently but few deep 
irregular cicatrices, some communicating with the tubuli lactifei'i, 
the greater part being regular, smoothly depressed, circular, or oval." 

Ceely illustrated his classical memoir with a series of valuable 
coloured drawings. One plate is a faithful picture of the disease on 



NATURAL AND CASUAL COW-POX. 319 

the teats as it is ordinarily met with ; the other is a composite picture, 
consisting of the disease as ordinarily observed in the cow, to which 
is superadded a number of depressed vesicles as they occur in inocu- 
lated cow-pox. It is, however, an improvement on a plate published 
by Sacco. The latter is an elaborate drawing, representing the udder 
and teats of a cow, with an eruption purpoi'ting to be the natural 
cow-pox. Jenner had described a bluish tint in the vesicles in 
natural cow-pox, and Sacco deliberately represents the natural disease 
by a highly coloured diagrammatic illustration in which he depicts 
clusters of vesicles of inoculated cow-pox, coloured blue, and with 
a silvery lustre. 

Hering has given a coloured plate of the natural cow-pox. On 
the teats are a number of oval and circvilar bullous vesicles and 
crusts. More recently, Layet has pointed out the same characters 
in the cow-pox discovered near Bordeaux in 1883 and 1884. The 
classical characters of the inoculated disease were wanting, particu- 
larly the central depression. In Wiltshire, the author could only 
distinguish, on the cow's teats, globular and broken vesicles and 
thick prominent crusts and ulcers, appearances which had very 
little in common with the ordinary results of vaccination. 

The early accounts of the severe character of the disease will 
appear by no means exaggerated to those who have had an oppor- 
tunity of studying the effects on the hands of the milkers, or indeed 
to those who have made themselves familiar with the descriptions 
given by Jenner, in some of his cases : — 

" Joseph Merret had several sores on his hands, swelling and stiffness 
in each axilla, and much indisposition for several days. 

" Mrs. H. had sores upon her hands which were communicated to her 
nose, which became inflamed and very much swollen. 

" Sarah Wynne had cow-pox in such a violent degree that she was 
confined to her bed, and unable to do any work for ten days. 

" William Rodway was so affected by the severity of the disease that 
he was confined to his bed. 

" William Smith had several ulcerated sores on his hands, and the 
usual constitutional symptoms, and was affected equally severely a second 
and a third time. 

" William Stinchcomb had his hand very severely affected with several 
corroding ulcers, and a considerable tumour in the axilla. 

" Sarah Nelmes had a large pustulous sore on the hand, and the usual 
symptoms. 

"A girl had an ulceration on the lip- from frequently holding her 
finger to her mouth to cool the raging of a cow-pox sore by blowing 
upon it. 



320 INFECTIVE DISEASES. 

" A young woman had cow-pox to a great extent, several sores which 
maturated having appeared on the hands and wrists. 

" A young woman had several large suppurations from cow-pox on the 
hands." 

Pearson in his investigations encountered, and was informed of, 
similar experiences. 

" Thomas Edinburgh was so lame from the eruption of cow-pox on 
the palm of the hand as to necessitate his being for some time in hospital. 
For three days he had suffered from pain in the armpits, which were 
swollen and sore to the touch. He described the disease as uncommonly 
painful, and of long continuance. 

" A servant at a farm informed Pearson that in Wiltshire and 
Gloucestershire the milkers were sometimes so ill as to lie in bed for 
several days. 

" Mr. Francis said that cow-pox was very apt to produce painful sores 
on the hands of milkers. 

" A servant of Mr. Francis said that cow-pox affected the hands and 
arms of the milkers with painful sores as large as a sixpence. 

" Mr. Dolling describes the disease as ' a swelling under the arm, chilly 
fits, etc., not different from the breeding of the small-pox. After the 
usual time of sickening, namely, two or three days, there is a large ulcer, 
not unlike a carbuncle, which discharges matter.' 

"Dr. Pulteney described the disease as causing 'a soreness and swell- 
ing of the axillary glands, as under inoculation for the small-pox, then 
chilliness and rigors and fevers, as in the small-pox. Two or three days 
afterwards abscesses, not unlike carbuncles, appear generally on the hands 
and arms, which ulcerate and discharge much matter.' 

" Mr. Bird wrote a short account : ' It appears with red spots on the 
hands, which enlarge, become roundish, and suppurate, tumours take 
place in the armpit, the pulse grows quick, the head aches, pains are felt 
in the back and limbs, with sometimes vomiting and delirium.' 

" Annie Francis had pustules on her hands from milking cows. These 
pustules soon became scabs, which, falling off, discovered ulcerating and 
very painful sores, which were long in healing. Some milk from one of 
the diseased cows, having spurted on the cheek of her sister and on the 
breast of her mistress, produced on these parts of both persons pustules 
and sores similar to her own on her hands." 

In more recent times these descriptions have been corufirmed. 

In 1836 cow-pox was discovered at Passy, near Paris. A black 
cow, in very poor condition, had cow-pox six weeks after calving. 
Bousquet had no opportunity of seeing the eruption in the early 
stage, but on examination he found reddish-brown crusts on the 
teats, which later gave place to puckered scars. The milk-woman, 
Fleury, who had had small-pox, nevertheless contracted the disease 
from the cow. She had several vesico-pustules on the right hand 



NATURAL AND CASUAL COW-POX. 321 

and on her lips. A vesico-pustule, when opened with a lancet, 
discharged like an abscess. 

In a letter to Mr. Badcock, dated April 3rd, 1845, Ceely referred 
to another new stock of lymph raised from a milker's hand. He 
added : — 

" In the enclosed lymph I see nothing unusually severe, except 
on very thin skins ; although the milker's hand exhibits now rough 
ulcers, one on the hand deep enough to encase a bean." 

Recent discoveries of cow-pox in England. — After Ceely's cases in 
1840-41, no cases of casual cow-pox on the hands of milkers were 
recognised as such and recorded in this country for nearly fifty years. 
In the outbreak of cow-pox discovered by the author in December 
1887, in Wiltshire, the disease was communicated to nearly all the 
milkers. The reader is referred to the account of this outbreak, 
which has already been given in the chapter on scarlet fever (p. 274). 

The author's researches were confirmed by Mr. Forty in 1888, 
and Mr. Bucknill in 1895. 

In June 1888 Mr. Forty, in practice at Wotton-under-Edge, 
Gloucestershire, reported to the Local Government Board, that at 
a farm at Alderley, an eruptive disease on the udder and teats was 
occurring amongst cows, and that the farmer's son, and other persons 
engaged as milkers, had contracted an eruption like that of the cows. 
The farmer's son had been under Mr. Forty's care suffering from an 
eruption, and circum-anal piles. Mr. Forty had watched the course of 
the eruption from papules to vesicles and scabbing, and concluded 
that the eruption could not be distinguished from vaccinia. Klein 
visited the farm, and found a number of cows with sores on the teats 
and udders. The sores were of various sizes and outline, mostly 
irregular, and covered with brown or black scabs. Those on the 
teats were larger and more irregular than those on the udder. 
Klein was shown several milkers who had had sores on one or more 
fingers ; one had had a bad arm with swollen axillary glands. 
The farmer had also contracted the eruption ; but in these persons 
only scabs were visible as the remnants of their sores. 

A girl of about twenty had taken the place of an incapacitated 
milker, and noticed a red pimple form on the dorsal surface of her 
right thumb. Eight days afterwards there was a slightly raised 
circular vesicle, with dark centre and pale periphery ; the centre of the 
vesicle was slightly depressed. It was just under half an inch in 
diameter ; there was peripheral redness, but no marked areola. The 
girl had three good vaccination marks. 

Klein experimented on calves with lymph from the vesicle and 

21 



322 INFECTIVE DISEASES. 

crusts from the cow's teats, with the result that from both sources 
an eruption was produced, which in appearance and course was like 
vaccinia. With lymph from one of the calves, a public vaccinator 
inoculated a number of infants, and fine vesicles developed, indis- 
tinguishable from vaccinia. 

In 1894 Mr. BuckniU met with a case in a milkman. He had 
been milking a cow affected with cow-pox, and on the ninth day after 
exposure to infection, and the seventh day after the eruption of the 
first papule, there were three pocks on the fore-arm. The pocks 
were elevated, circular, and umbilicated, with a dull, creamy-white 
ring at the circumference, and there was well-marked induration 
and extensive areola. There were four excellent marks of primary 
vaccination. The vesicles contained clear lymph, and re-inoculation 
of the arm failed to take. An attempt to re-vaccinate the man 
with current calf lymph produced only topical irritation. 

Inoculated Cow-pox. 

Natural or Virulent Lymph. — Severe symptoms are not limited to 
milkers casually infected from the cow. Under certain conditions, 
artificial inoculation of fresh virus from the cow reproduces the 
disease without any mitigation. Thus, in Jenner's cases : — 

"James Phipps. The incisions assumed at their edges rather a 
darker hue than in variolous inoculation, and the efflorescence around 
them took on more of an erysipelatous look. They terminated in 
scabs and subsequent eschars. 

" Susan Phipps was inoculated from the cow by inserting matter 
into a superficial scratch on December 2nd. The child's arm now 
showed a disposition to scab, and remained nearly stationary for 
two or three days, when it began to run into an ulcerous state, 
and then commenced a febrile indisposition, accompanied with an 
increase of axillary tumour. The ulcer continued spreading near 
a week, during which the child continued ill, when it increased to 
a size nearly as large as a shilling. It began now to discharge pus ; 
granulations sprung up, and it healed." 

Jenner's lymph was employed by Mr. Chne with similar results. 

" The child sickened on the seventh day, and the fever, which 
was moderate, subsided on the eleventh. . . . The ulcer was not 
large enough to contain a pea." 

Precisely similar experiences have since been encountered, in the 
early removes of fresh stocks of virulent lymph. Bousquet in France, 
in his first trials with a new lymph, in 1836, made three punctures, 
but he had soon to abandon this practice, because the intensitv 



INOCULATED COW-POX. 323 

of the inflammation was sometimes so great that it spread over the 
entire arm as far as the glands of the axilla. In one case, the 
vesicles were enormous, and the inflammation so violent, that baths, 
poultices, fomentations, and antiphlogistic diet scarcely sufficed 
to reduce it. The crusts when they fell off left ulcerations which 
were very slow to undergo cicatrisation. In some cases, the vesicles 
which resulted hollowed out the skin so' deeply that they left regular 
holes. 

In the following year Esthn, in England, started a stock of fresh 
vaccifle virus from the cow, and found on inoculating childi'en that 
the new lymph was extremely active. 

In 52 the disease was regular, 

1 severe erysipelas, 
4 erythematous eruptions of a violent character, 

2 highly inflamed ulcerated arms, 
1 no effect after twice vaccinating, 
8 result unknovm ; supposed to have been favourable. 

68 

Cultivated or Attenuated Lymph. — When cow-pox lymph has 
been mitigated by successive transmission through the human subject, 
or by cultivation on the belly of the calf, with careful selection 
of vesicles, it will produce effects which are as follows : About 
the end of the second day after insertion, or early on the third day, 
a slight papular elevation is noticeable. By the fifth or sixth day, 
it has become a distinct vesicle, of a bluish-white colour, with raised 
margin and central cup- like depression. By the eighth day, the 
vesicle is perfect. It is circular, pearl-coloured, distended with clear 
lymph, and the central depression is well marked. On the same day, 
or a httle earlier, the areola begins to appear, and gradually extends 
to a diameter of from one to three inches, accompanied with 
induration and tumefaction of the subjacent connective tissue. After 
the tenth day, the areola begins to fade, and the vesicle at the same 
time begins to dry in the centre ; the lymph becomes opaque and 
gradually conci'etes, and by the fourteenth or fifteenth day, a hard 
mahogany-coloured scab is formed which contracts, dries, blackens, and 
falls off between the twentieth and twenty-fifth days. A circular, 
depressed, foveated, and sometimes radiated scar remains behind. 
By selecting characteristic vesicles on the calf or on the human 
subject, and by collecting the lymph at an early stage on the fifth, 
sixth, or seventh day, this artificial disease, commonly known as 



324 INFECTIVE DISEASES. 

vaccinia, can be kept up in this comparatively mild form. But 
under certain conditions, such as a peculiarity in the subject inocu- 
lated, or if lymph be taken too late, there will be, just as in variolation, 
tendency to revert to the full intensity of the natural virus. 

Bacteria in Vaccine Lymph — Oohn, Sanderson, and Godlee 
described micrococci in vaccinal vesicles. Quist and Ferre in 1883 
investigated the same subject. Voigt in 1885 distinguished three 
species of micrococcus— a diplococcus, a large coccus, and a third form. 
Bauer in the same year described the presence of bacilli and sphsero- 
cocci. Marottain 1886 regarded a tetracoccus as the specific micro- 
organism, and Tenhot in 1887 distinguished a dozen micrococci, two 
bacilli, and two yeasts. In the same year Garre isolated a micrococcus 
which appeared to him to be the contagium, but inoculated on a child 
it neither produced local vesicles nor immunity ; while Guttmann 
pointed out three micro-organisms which appeared to be rather more 
constantly present than others. Pf eiffer much more fully investigated 
the bacteriology of vaccine lymph, and found Saccharomyces vaccinae, 
which was seldom present in human lymph but constantly found in 
calf lymph ; sarcin^, both in human and calf lymph, including Sarcina 
lutea, Sarcina tetragonus, Sarcina aurantiaca, Sarcina muscopus ; 
bacteria and bacilh were found only exceptionally in hviman lymph, 
but frequently in calf lymph. These included a bacterium corre- 
sponding with Proteus vulgaris. 

Three mice were inoculated subcutaneously with a drop of the , 
liquefied gelatine, but the result was negative. The injection of a 
considerable quantity proved fatal to guinea-pigs and rabbits, a 
result which was probably diTO to ptomaine poisoning. 

There were also several bacilli which did not liquefy gelatine ; 
these were not investigated. 

Staphylococcus cereus albus was found very frequently, and 
Staphylococcus pyogenes aureus occasionally. Pure-cultivations of 
these micrococci inoculated on the skin of calves produced a rapid 
local irritation, followed by vesiculation, but without the classical 
characters of the vaccine vesicle. The inoculated part was com- 
pletely healed in three to five days. According to Pfeiffer they 
explain the so-called false vaccine. 

Micrococcus pyogenes albus was almost constantly present. 
Numerous other micrococci were found, but not constantly present ; 
vaccine lymph being a splendid medium for the growth of micrococci. 
Pfeiffer pointed out that the effects of Staphylococcus pyogenes aureus, 
albus, and citrous, and of Streptococcus pyogenes on rabbits had an 
important bearing upon the practice of vaccination, and he recom- 



INOCULATED COW-POX. 325 

mended that calf lymph should be tested before use vipon children 
by inoculation of the ear of a rabbit. If after two days no erysipelas 
occurs in the inoculated rabbit, the absence of streptococci may be 
considered as almost proved. Two or three rabbits should be inocu- 
lated at the same time. 

The author's researches into the bacteriology of vaccine lymph 
extended over some years. They independently confirmed and 
extended the results obtained by Pfeifler. Having on several 
occasions examined vaccine lymph and vaccine pns, and failed to find 
a specific bacterium, the author proceeded to make a more systematic 
examination of the different species of bacteria in samples of current 
vaccine lymph. Pure-cultivations were obtained by plate-cultivation, 
and inoculation of the surface of nutrient agar, obliquely solidified 
in test tubes. Various current stocks of lymph were used in the 
investigation. Among the specimens of calf lymph. No. 1 yielded a 
torula. Bacillus pyocyaneus and Bacillus subtilis; No. 2, a bacterium, a 
variety of proteus, Staphylococcus pyogenes aureus, and yellow bacte- 
rium ; No. 3, a bacterium, micrococcus, yellow bacterium, and torula ; 
No. 4, yellow micrococcus, white micrococcus, white torula, yellow 
sarcina, white diplococcus. Staphylococcus cereus albus, and a mould 
fungus; No. 5, yellow sarcina. Staphylococcus pyogenes aureus, yellow 
micrococcus, white bacillus. Staphylococcus pyogenes albus, large 
white micrococcus, yellow bacterium, and a white micrococcus. Among 
the specimens of human vaccine lymph. No. 1 contained a white 
micrococcus, proteus, and Staphylococcus pyogenes avu'eus ; No. 2, a 
micrococcus, a tetracoccus, a white liquefying mici'ococcus, and a 
yellow bacterium ; No. 3, white micrococcus, yellow micrococcus. 
Staphylococcus aureus and flavus, a bacterium, a white micrococcus, 
a bacillus resembling Bacillus subtilis. Staphylococcus pyogenes cereus 
and a brown tetracoccus. The au.thor is familiar with these different 
species of bacteria, and not one of them is peculiar to vaccine lymph ; 
there was no bacterium constantly present in human and calf vaccine, 
and there was not one which could be regarded as the contagium. 
To sum up, most of them are well known saprophytic bacteria, and 
some were identical with bacteria commonly found in suppuration. 
Yaccine lymph is a most suitable cultivating medium for micro- 
organisms, and bacteria invariably got access to the contents of the 
vaccine vesicle. There is no evidence to be obtained by the present 
methods of research as to the bacterial nature of the contagium of 
vaccine lymph. Copeman obtained similar results, and thus con- 
firmed the author's conclusions. 

Klein and Copeman have also observed minute bacilli in calf- 



326 INFECTIVE DISEASES. 

lymph and in variolous lymph. Numerous attempts to cultivate them 
in nutrient media and in the living animal failed entirely, and the 
identity of the bacilli could not be determined. Pfeiffer, Guarnieri, 
Monti, Euffer, and Plimmer have drawn attention to structures in 
lymph, which they believe to be of the nature of parasitic protozoa. 
These bodies have been studied, more especially in the tissues. They 
are four times the size of ordinary micrococci, and are found in the 
clear vacuole in the protoplasm of epithelial cells. Whether they 
are really parasites or altered anatomical elements has not been 
determined. No other conclusion can be drawn from all these 
observations, except that the nature of the contagium of cow-pox 
is unknown. 

Origin of Cow-pox. 

Jenner's original theory was that cow-pox was derived from 
"grease," but subsequently he distinguished between cow-pox, a 
disease peculiar to the cow, and "grease," a disease transmitted to 
the cow from the horse, and the mistake of confounding these two 
diseases was attributed to farmers and farriers. Thus he wrote : — 

" Prom the similarity of symptoms, both constitutional and local, 
between the cow-pox and the disease received from morbid matter 
generated by a horse, the common people in this neighbourhood 
when infected with this disease, through a strange perversion of 
terms, frequently called it the cow-pox." 

Jenner's theory of the origin of cow-pox has been discouraged ; so 
also has the view of its being a " spontaneous " disease in the cow, 
though Oeely, after many years of research in the "Vale of Aylesbury, 
could never discover the probability of any other origin. Both 
opinions have given way to the theory that cow-pox is small-pox 
transmitted to the cow — an opinion advocated by Baron, and 
supported by an erroneous interpretation of Oeely's and Badcock's 
variolation experiments. Thus the cow-pox and grease of farmers 
and fairriers no longer attracted attention in this country, and as 
natural cow-small-pox has never been discovered, cow-pox has been 
credited with being extinct. 

For a full discussion of this subject the reader is referred to the 
work by the author on the History and Pathology of Vaccination, 
but the variolation experiments alluded to will be briefly mentioned. 

In 1801 Gassner inoculated eleven cows with small-pox lymph, 
and succeeded in one in producing phenomena indistinguishable from 
the results of ordinary vaccination with cow-pox, and children were 
inoculated from the cow. 



ORIGIN OF COW-POX. 327 

In 1828 Dr. McMichael reported that several physicians in 
Egypt had obtained similar resvilts, and children were successfully 
" vaccinated." 

In 1836 Dr. Martin, in America, inoculated the cow's udder with 
variolous lymph, and by inoculating children produced an epidemic 
of small-pox with fatal cases. In 1839 Reiter of Mu'nich, after fifty 
unsuccessful attempts, succeeded in producing a vesicle, and a chUd 
inoculated from the vesicle contracted small-pox. 

In 1839 Dr. Thiele, after a number of unsuccessful attempts to 
inoculate cows vsdth variolous virus, succeeded in producing a vesicle 
with the physical characters of the vaccine vesicle, and from it a 
stock of lymph was raised from which over three thousand persons 
were inoculated. Thiele's method was to inoculate the udder with 
lymph, and to select for the purpose young cows which had recently 
calved and had delicate skins. In England Ceely succeeded by 
inoculating the vulva of a heifer. One of the punctures developed 
into an enormous vesicle, which was undoubtedly variolous. His 
assistant punctured his hand with the lancet which had been used 
to open the vesicle, and febrile symptoms appeared, followed by an 
eruption on the face, neck, trunk, and limbs, at first papular, then 
vesicular, and finally pustular. The lymph was used in children, 
and " vaccine " vesicles were produced. One child suffered from 
vomiting dehrium, and extensive roseola, but there was no eruption 
in any other case. 

In 1840 Badcock of Brighton inocvalated a cow successfully, and 
later succeeded in variolating thirty-seven out of two hundred cows 
upon which he experimented. 

In 1847 variolation of the cow was successfully performed at 
Berlin, but the vii-us produced variola, and one of the children 
inoculated died of confluent small-pox. 

In 1864 Ohauveau inoculated seventeen animals with virulent 
small-pox lymph. Very small papules resulted, and the virus from 
the papules produced variola in a child, which was infectious to others. 
Kiein in this country until recently was uniformly unsuccessful. 
Voigt, Fischer, King, Eternod, Haccius, Hime and Simpson, have 
all succeeded in inoculating cows and producing variola-vaccine. 

The results of these experiments have been very generally misin- 
terpreted, and claimed by some as conclusive evidence of the identity 
of cow-pox and small-pox. Instead of the vesicle being regarded as 
the most attenuated form of variola, the experimenters are said to 
have succeeded in producing cow-pox. 

It is quite true that they produced phenomena indistinguishable 



328 INFECTIVE DISEASES. 

from the phenomena of an ordinary vaccination, but that does not 
mean that they produced the disease cow-pox. The vesicle which 
followed the inoculation, whether papular or vesicular, was srriall-pox. 
Oeely, Badcook, Yoigt, and others, succeeded in ingrafting the cow 
with small-pox, and when suitable lymph and suitable subjects were 
employed, the virus was so attenuated that a benign vesicle resulted. 
Similar results were obtained by Sutton and Dimsdale, and identical 
results by Adams, Guillou, and Thiele, by inoculating the human 
subject with variolous lymph without first ingrafting the disease on 
the cow. 

Vaccination with variola-vaccine is simply a modification of the 
Suttonian system of small-pox inoculation, only in the first remove 
the cow is substituted for the human subject. All those who were 
inoculated with Ceely's, Badcock's, or Simpson's variola- vaccine, 
were not in the usual meaning of the word vaccinated ; they were 
not inoculated with cow-pox but they were variolated, and in such 
an extremely attenuated form that the persons so variolated do 
not convey the infection. By judicious selection it is thus possible 
to obtain a strain of lymph from variola which, by direct inoculation 
of the hviman subject or by first inoculating a cow, is deprived of 
infectious properties, and produces on the arm the physical characters 
of an ordinary vaccine vesicle. This has been regarded as a proof 
of the identity of small-pox and cow-pox, but it is not so. Variola 
and cow-pox are not the only diseases caused by a virus which can 
be attenuated until only a vesicle is produced with the characters of 
an ordinary vaccine vesicle. The results which have been obtained 
with the virus of cattle plague and of sheep-pox and horse-pox have 
been given in previous chapters ; and no one would urge on this 
account that human small-pox, cattle plague, cow-pox, sheep-pox, 
and horse-pox are all manifestations of the same disease. Cow-pox 
has never been converted into human small-pox, and, in their clinical 
history and epidemiology, natural cow-pox and human small-pox 
are so different, that the comparative pathologist is no more pre- 
pared to admit their identity than he is prepared to admit the 
identity of cow-pox and sheep-pox, or small-pox and cattle plague. 

Protective Inoculation.— Whether vaccination of all heifers 
on a farm would protect them from cow-pox when they came into 
milk is not known, the duration of the immunity in calves afforded 
by vaccination having not been determined. Calves undoubtedly 
have an immunity after vaccination, lasting for some weeks. 

In 1896 Becl^re, Chambon, and Menard experimented upon 
the immunising power of the serum of vaccinated calves. They 



COW-POX AND SMALL-POX. 329 

concluded from experiments on animals and children that the serum 
of a vaccinated calf from ten to fifty days after vaccination will give 
immunity against inoculated cow-pox. ' They further stated that, 
whereas the immunity given by vaccination in the ordinary way 
is not complete until the eighth day, the immunity obtained b}' 
injection of the immunising serum is immediate. The serum has 
also been credited with therapeutic properties and has, it is said, 
proved efficacious in cases of small- pox. 

Jenner believed that cow-pox did not protect against itself but 
protected against small-pox, and for a century this has been a subject 
of much controversy. The reader is referred to the Reports and 
conclusions of the Koyal Vaccination Commission. 

Stamping-out System. — It would undoubtedly be an advan- 
tage if cow-pox were scheduled under the Contagious Diseases Animals 
Act. Cow-keepers and dairy-men, being anxious that their trade 
should not be interfered with, very commonly conceal the existence 
of the disease, and perhaps nothing is known about it, unless a milker 
infected from the cows seeks for medical advice. The contamination 
of the milk with lymph, pus, crusts, and sometimes blood, renders 
it unwholesome, and therefore precautions ought to be taken to 
prevent its occurrence. If the infected cows in a herd are the last 
to be milked, and the milker washes his hands after the milking, the 
disease will not spread. 



CHAPTEK XXIII. 

DIPHTHERIA. 

Diphtheria is a specific infectious disease, especially of children, 
characterised most commonly by inflammation, and infiltration with 
lymph cells and fibrine, of the mucous membrane of the fauces, 
pharynx, larynx and trachea, followed by necrosis of the mucous 
membrane and the formation of a greyish-white false membrane, 
the diphtheritic membrane. In some cases a diphtheritic membrane 
forms in the stomach, intestine, the urinary organs and in wounds. 
After the separation of the membrane an ulcer remains, which may 
gradually heal. In the superficial part of the diphtheritic membrane 
there are masses of bacteria including cocci, streptococci, and baciUi. 
The diphtheria bacilli are not found in the blood or in the 
internal organs. There is no doubt of the fact that diphtheria 
is a disease which can be communicated from one person to 
another ; but the question of its origin is still a vexed one. There 
is a close association with insanitary conditions and decaying 
animal and vegetable refuse, and dampness. Localities with damp 
houses, defective drainage, and a cold exposure, are favourable 
to the development of diphtheria; but that does not necessarily 
indicate that these conditions can originate it. On the other 
hand, assuming the disease to be due to a living contagium, 
these insanitary conditions would afford a suitable environment 
predisposing to the development, and facilitating the spread, of the 
disease. Scarlet fever and measles predispose to diphtheria ; and 
defective sanitary conditions, causing sore throat, may indirectly 
act as a predisposing cause. A great many cases have been quoted 
to illustrate the possibility of the conveyance of diphtheria by milk, 
and the theory which best harmonises with all these observations is 
the existence of a specific bacillus, which may be readily transferred 
from the throat of the diseased to the healthy ; which finds also in 
milk a suitable soil for its growth, and by its agency may be trans- 
mitted to the consumer. Such a bacillus was discovered by Lbfiler, 

330 



DIPHTHERIA. 



331 



and may be easily 
obtained from tbe 
throat of dipbtlieritic 
patients in the fol- 
lowing manner : — 

Culture Outfit. — 
Steel rods like or- 
dinary knitting 
needles, about six 
inches in length, are 
beaten out or rough- 
ened at one end, and 
a pledget of wool is 
twisted round so as 
to form a swab. 
These swabs are 
placed in clean test- 
tubes, which are then 
plugged with cotton- 
wool. The test-tubes 
and swabs are steri- 
lised by heating in 
the hot air steriliser 
for an hour at 150° C. 
The so-called culture 
u t fi t consists of 
a small box con- 
taining a test-tube of 
blood serum and a 
swab. The}^ can be 
always kept ready for 
use, and after use 
should be conveyed 
by hand for further 
examination. The 
danger of trans- 
mitting virulent 
diphtheritic material 
by post is obvious. 
When the examina- 
tion of the tube has 
been completed, the 



^ 



>* 























Fig. 126.— Free Subface of Diphtheritic Larynx 
X 350 (Hamilton).— X, Deposit of diphtheria bacillus 
on surface of false membrane ; B, false membrane ; 
C, mucosa ; I, lymph-cells and false membrane 
surroimded by meshes of fibrine ; c, surface of mucosa 
deprived of its epithelium ; l,v, lymph-cells containing 
shed epithelium. 



oo2 INFECTIVE DISEASES. 

culture outfit and its contents should be disinfected or destroyed. 
To inoculate the tubes the patient, if it is possible, should be turned 
to the light, the mouth well opened, the tongue depressed, and the 
swab, without touching the teeth or the tongue, should be pa.ssed 
straight to the tonsils or pharynx, and especially to the membranous 
exudate. The swab is carefully and quickly withdrawn, and at 
once very gently rubbed over the surface of the blood serum. The 
culture outfit is then sent to the laboratory with full particulars, 
and the tubes are placed in the incubator at .37° C, and can be 
examined after twelve hours. If the throat has been disinfected 






« 






m^ -■ -^ 




^ 



^Uk- 






'f5^ 

^"* *"' "». 

Fig. 127.— B.\oiLLrs of Diphtheria ; fkom a Cultivation on Blood 
Sebum, x 1000 (Fbaxkel and Pfeifpek). 

before examination, this must be taken into account, as the failm-e 
to find bacilli would not then necessarily indicate a wrong diagnosis. 
In all undoubted cases of diphtheria, growths will be obtained either 
in the form of a pure-culture of the bacillus, or far more commonly 
there will alsoibe colonies of various bacteria, especially Streptococcus 
pyogenes. 

Bacillus of Diphtheria — Eods, straight or slightly curved, 
•3 to -8 /t in breadth, and I'S to 6-5 /a in length. They occur 
'singlyi in pairs, sometimes in chains, and sometimes as short 
leptothrix forms. In some cultures very irregular forms are 
observed, the bacilli being swollen at one or both ends or thicker 
in the middle portion, or the bacillus may contain oval or spherical 



DESCEIPTION OF PLATE VIII. 
BacillTis diphtheriaB and Bacillus typhosus. 

Fig. 1. — Co%er-glass preparation from a pure-cultivation of Bacillus diph- 

therise on blood serum ; obtained from the throat in a typical case of 

diphtheria. Stained with gentian- violet, x 1200. 
Fig. 2. — Cover-glass preparation from a pure-cultivation of Bacillus typhosus 

on nutrient-agar ; from the spleen in a, case of typhoid fever Stained 

with gentian-violet, x 1200. 



Plate VIII. 






•J... 



I--/ 



•'..:v:; i^::. ..-.v. ; V * -{^M 



/ •. 






-.. \ 




■ -^ • •■?' 






;-' r.: 



Fiol. BACILLUS DIPHTHERIyE 

CD 




•7\ 



Fiji 2. BAG ILL us TYPHOSUS 



^■M.Cropfofi^rJf, 



■■.f&cil. 



fi.fi.:€nt B'-aok:.%nay ^Sr^n.lith. 



DIPHTHERIA. 



333 



fJM 



elements. The}' differ greatly in size and shape, often in the same 
cultures, and still more in cultures obtained from different sources. 
Spore formation is unknown. In unstained preparations thei'e are 
highly refractive elements which correspond with the deeply stained 
parts of the bacillus. They stain readily with the ordinary aniline 
dyes. At certain stages of their growth they stain irregularly, the 
protoplasm of the rod being broken up into irregular segments. 
The bacillus is non-motile, and does not liquefy gelatine ; it grows 
at 20° C, but much more readily at higher temperatures. Colonies 
in gelatine plate-cultivations are 
yellowish-brown, and opaque, 
granular, and circulai-, but with 
more or less irregular margin. 

In jDlate-cultivations on agar 
and on glycei-ine agar the same 
description applies. 

On the surface of gelatine 
the appearances depend greatly 
on the method of' inoculation. 
The growth may occur in the 
form of a whitish film, but if a 
sub-culture has been prepared 
from broth the growth is often 
composed of a number of iso- 
lated white colonies (Fig. 128, a). 

On blood serum, after twelve 
hours the colonies appear in the 
form of little elevated greyish- 
white or pearl-grey dots, which 
coalesce, forming a film if the 
serum is moist. On the surface 
of 1 per cent, alkaline glycerine 
agar, the appearances are found 
to vary, and this medium is not so suitable for the cultivation of the 
bacillus. In slightly alkaline broth, with or without the addition of 
1 per cent, grape-sugar, the culture is cloudy, or a fine granular 
deposit occurs along the sides and bottom of the tube, while the 
broth remains clear. 

On potato the growth is almost invisible, in the form of a dry, 
thin glaze. Irregular forms are very numerous on microscopical 
examination, whilst the rods are thicker than usual (Welch and 
Abbott). In milk the organisms grow readily. 



Fig. 128. — PunK-crLTUKES of Bacillus 
DiPHTHEtii/E ON Gelatine: a, isolated 
colonies ; h, iilmy growth. 



334 INFECTIVE DISEASES. 

Dried diphtheritic membrane and cultures dried on silk threads 
retain their vitality for several months. 

A brotli-culture in forty-eight hours may be used for inoculating 
guinea-pigs. A few drops will cause death in from three to five 
days ; there is hypersemia and cedema at the seat of inoculation, the 
lymphatic glands are enlarged, there is fluid in the peritoneal, 
pleural and pericardial cavities, and the lungs are congested. The 
bacillus is found at the seat of inoculation, but not, as a rule, in the 
blood or internal organs. Inoculation of rabbits produces extensive 
local oedema, enlargement of the lymphatic glands, and death in 
from four days to three weeks. Roux and Yersin pointed out that in 
less acute cases there was paralysis of the hind limbs. Mice and 
rats have an immunity. Cultures lose their virulence with age, but 
the filtrate from old cultures contains more toxic substance than 
that from fresh cultures. The toxin has been described in a 
previous chapter (p. 46). 

Old cultures sterilised by heating for an hour to 60° 0. or 
70° C. will render guinea-pigs immune in two weeks. The toxic 
substance is believed to be destroyed by this process, while according 
to Frankel the immunity-giving substance which is also present in 
the culture is not affected. 

According to Behring's researches, the blood of immune animals 
contains diphtheria antitoxin, consequently the blood of an im- 
mune animal is caj)able of neutralising the toxic properties in a 
filtered culture, not only in the living animal but when added to the 
culture in a test-tube. These researches led to the employment of 
the serum of an immune animal as a therapeutic agent in the treat- 
ment of diphtheria in man (p. 58). 

Bacteriological Diagnosis. — The diphtheritic bacilli are not only 
found in the throat while the lesions exist, but they are found after 
all sign of the disease has disappeared. In some cases they 
persist for a few days, in others for three or four weeks, and 
in rarer cases they have been found several months afterwards. 
They have also been found in the throats of persons in health, 
especially of those who have been in contact with cases of diph- 
theria, such as healthy children in' infected families and healthy 
nurses. 

The bacilli which persist in the throat after recovery may be 
virulent up to the time of their disappearance, or they may gradu- 
ally become attenuated, and entirely lose their pathogenic properties. 
The value of a microscopical examination as an aid in the diagnosis 
of diphtheria has been considerably exaggerated, and unless the 



» DIPHTHERIA. 335 

bacillus when isolated is tested by inoculation the test may prove 
to be entirely fallacious. 

Loffler and Von Hoffman both found bacilli in healthy throats, 
and thus created doubt as to the importance of the Loffler bacillus. 
Hoffman found this bacillus in the throats of twenty-six out of 
forty-five individuals, some of them suffering from scarlet fever, 
measles or some other non-diphtheritic affections, while the rest were 
healthy. The bacilli from these sources showed slight differences 
in morphological and cultural characteristics, and Hoffman was 
unable to decide whether these bacilli were diphtheria bacilli, which 
had become harmless, or whether they were accidental epiphytes, 
belonging to a closely allied but different species. 

Roux and Yersin confirmed these observations. In a hospital 
for ehildi-en in Paris without any question of the existence of 
diphtheria they found the so-called pseudo-diphtheria bacilli in 
fifteen cases out of forty-five. In a school, in a seaside place 
entirely free from diphtheria, the same bacilli were found in 
twenty-six out of fifty-nine children. They were also found in 
children with simple sore throats, and in five out of seven cases 
in measles. Roux and Yersin concluded that these bacilU were 
not distinct from the Loffler bacillus. There were slight variations, 
but there was no constant difference except in their pathogenic 
properties. The appearance of the colonies, the growth in broth, 
and the peculiar morphological elements showed chal'acters common 
to both, and there was, in fact, less difference than there is 
between attenuated anthrax and virulent anthrax in forn^ and in 
cultures; but inoculations of the bacillus did not cauSe death, 
though in some cases in guinea-pigs there was marked oedema at 
the seat of inoculation. On the other hand, Lbffler's bacilli 
possess different degrees of virulence, some cultures producing only 
temporary oedema, while others cause death in twenty-four hours. 

Virulent diphtheria bacilli subjected to a current of air can 
in two weeks be deprived of their virulence partially, and in four 
weeks entirely. Weakened bacilli can be raised in virulence by the 
simultaneous injection of the streptococcus of erysipelas, but bacilli 
deprived of their virulence and bacilli originally non-virulent cannot 
be made to assume virulent properties. Escherich maintained 
that they could be distinguished by comparative cultures ; that the 
pseudo-diphtheria bacilli made broth alkaline, so that in forty-eight 
hours litmus was turned red by Lbffler's bacilli and blue by the 
false bacilli. The bacilli themselves, according to Hoffman, are, as 
a rule, shorter, wider and more uniform in size. 



336 INFECTIVE DISEASES. 

Parke and Beebe, with a view to clearing up this question, made 
cultures from three hundred and thirty healthy throats. They 
found bacilli of three varieties : bacilli characteristic in growth 
producing acid reaction in broth, but having no virulence; bacilh 
not characteristic in growth producing an alkaline reaction in broth, 
not virulent ; and bacilli producing acid reaction in broth and 
virulent. The virulent characteristic diphtheria bacilli were found 
in eight cases, non- virulent diphtheria bacilli in twenty-four, and 
non-virulent false diphtheria bacUli in twenty-seven. They con- 
cluded that the eight cases must have been in contact with diphtheria, 
although the throats were healthy. With regard to the bacillus 
in the twenty-four cases they regarded it as the true diphtheria 
bacillus which had lost its virulence, and the bacillus found in the 
twenty-seven cases showing differences in size and manner of staining 
and the reaction produced in broth was properly designated pseudo- 
diphtheria bacillus. 

Diphtheritic Diseases in Animals. 

There are diptheritic diseases of the lower animals which are in 
some respects similar to, and, some observers maintain, identical 
with, human diphtheria. 

In pigeons there is a disease accompanied with the formation of 
false membranes associated with a bacillus described by Lbffler. 

Bacterium of Diphtheria of Pigeons (Bacillus colwnibarum, 
Lbffler). — Short rods with rounded ends, mostly in irregular masses. 
In plate-cultivations on nutrient gelatine they formed whitish patches 
on the surface, and compact, ball-like masses when embedded in the 
gelatine. They were also cultivated on blood serum and potatoes. 
Subcutaneous inoculation of a pure- cultivation produced in pigeons 
local inflammation and necrosis ; inoculation in the mucous mem- 
brane of the mouth gave the appearances of the original disease. 
Other animals were only locally affected, except mice, in which 
characteristic symptoms and death resulted. They were isolated 
from the diphtheritic exudations in pigeons, and in sections were 
found in the vessels of the lungs and liver. 

A similar disease is known to attack fowls, and there are also 
diseases with development of false membranes of the respiratory 
passages in horses, cats and swine. Outbreaks of these diseases 
have been said to occur in times of prevalence of diphtheria in 
man, and their intercommunicability has been suggested. 

Dr. Turner supposes that diphtheria in man originates in diseases 



DIPHTHERITIC DISEASES IN ANIMALS. SST 

simulating diphtheria in cats, pigs, and horses ; and Klein, wha 
accepts this theory, maintains that cats suffer from genuine 
diphtheria, and that after death the lungs are found full of grey, 
consolidated lobular patches, and the kidneys are ehlarged and 
white. 

Human diphtheritic membrane inoculated subcutaneously in cats 
produces a painful swelling in the groin, and fever, and a fatal 
termination in a week. The subcutaneous and muscular tissues at 
the seat of inoculation are hsemorrhagic and oedematous. The 
internal organs are congested, and in the kidneys the medulla is 
congested, while the cortex is fatty. 

A recent culture produces illness in twenty-four hours, a painful 
tumour forms at the seat of inoculation, and death ensues in from 
two days to a week. Pneumonia and fatty white kidney are found 
after death, and the tissues at the seat of inoculation are ha3mor- 
rhagic, and in parts almost gangrenous. 

Klein found that diphtheritic membrane or a pure-culture in- 
oculated into the cornea after removal of the superficial epithelium 
produced ulceration, and in two cases perforation of the cornea, 
and purulent panophthalmitis. Bacilli were again recovered from 
the ulcer similar in cultural characters, but conspicuously shorter 
and thinner. 

An epidemic occurred amongst cats at the Brown Institution. 
Five out of fourteen died. The symptoms were, running from the 
eyes, sometimes a muco-purulent discharge, sneezing, coughing, and 
pulmonary trouble, resulting in emaciation and death in from one 
to three weeks. After death lobular pneumonia and large white 
kidney were found ; and in one case a diphtheritic condition of the 
trachea, preparations of which showed diphtheria bacUU in crowds 
under the microscope. 

Klein regarded this disease as an epidemic 6i cat-diphtheria, 
and believed that the disease was possibly induced accidentally by 
the cats drinking milk, which was infected in the course of some 
other experiments with diphtheria. He states that on account 
of the very definite results obtained by inoculating diphtheritic 
membrane and cultures of the bacillus, subcutaneously and on 
the cornea, and of the condition of the lung and kidney in cats 
naturally or experimentally infected, the disease must be con- 
sidered as equivalent to human diphtheria, and the cat capable 
of communicating the disease to other cats, and also to human 
beings. These conclusions have not yet met with the acceptance cf 
veterinary aiithorities. The results of the experimental inoculations 

22 



338 INFECTIVE DISEASES. 

are certainly by no means conclusive. It does not follow from these 
experiments that the disease diphtheria naturally occurs in the 
oat or that under ordinary circumstances cats may contract the 
disease from the human subject ; but the experiments show that, 
like' guinea-pigs and rabbits, cats are susceptible to the toxic effects 
of the extremely poisonous principles developed during the growth 
■of LbfHer's bacillus. 

Milk Diphtheria. 

It has been shown that milk infected with diphtheria has been 
"the cause of epidemics among the consumers ; there have also been 
epidemics apparently associated with the milk supply, in which it 
Jias not been possible to trace the source from which the milk was 
infected. A difficulty in tracing the origin in no way excludes the 
possibility of contamination from a human source. In the light of 
recent researches we should expect that it would be easy to overlook 
the source of the virus, if it be true that diphtheria may exist without 
^ny symptoms indicating its presence, and be unrecognised until the 
throat has been examined for diphtheria bacilli. As this fact was 
unknown until quite recently, the absence of an acknowledged 
■case of diphtheria was taken as evidence that no diphtheria existed, 
and consequently that the milk must have been infected by a 
■diseased condition of the cow. Mr. Power, whose views upon milk 
scarlatina have already been referred to, endeavoured to trace the 
■origin of a milk epidemic to the very common disease of " garget," 
or mammary abscess. This idea may be dismissed without further 
■consideration ; but the theory of some disease existing in the cow 
capable of producing diphtheria in man was resumed by Dr. 
Cameron, who suggested that there might be an eruptive disease of 
the teats producing diphtheria, and by Mr. Power, who supported the 
theory in an investigation of a milk-diphtheria outbreak in 1886 
.at Oamberley. Diphtheria in this case existed in the neighbourhood, 
but as the source of human infection could not be traced, attention 
was drawn to two cows in the herd which had recently calved, and 
•especially to one with chapped teats. Following this line of inquiry, 
Klein investigated the behaviour of milch cows to the diphtheria 
bacillus. Two cows were injected subcutaneously under the skin of 
the shoulder with a Pravaz' syringe filled with a sub-culture in broth. 
There was a rise of temperature, and on the third day a painful 
tumour, which enlarged to the size of a child's head. In about a 
fortnight the tumour began to decrease, and ultimately one cow 



MILK DIPHTHERIA. 339 

died and the other was killed. Such results might have been 
anticipated as the result of injecting a large quantity of the toxic 
products of the bacillus, but certain other phenomena were observed 
to which importance was attached. On the fourth day, on one of 
the cows an eruption on the teat was first noticed, consisting of 
small vesicles passing into pustules and crusted ulcers. Examina- 
tion of the contents of the vesicle revealed the bacillus. With 
matter from the vesicles and pustules two calves were inoculated, 
and a similar vesiculation produced at the seat of inoculation. 
The milk of the cows was inoculated on nutrient gelatine, and 
produced a culture of Bacillus diphtherise. The question naturally 
arose whether this eruption had any connection with the original 
experimental inoculation. No other cows in the locality from 
which these cows were obtained had a similar eruption, and it was 
taken for granted that it was the result of the experimental inocu- 
lation. By accepting the possibility of this eruption being identical 
with the chaps on the teats of the Camberley cows, the theory was 
gradually built up that cows suffer from diphtheria, which manifests 
itself in the form of an eruptive -disease of the teats, and that the 
disease is conveyed in the milk to the consumers. ^ 

In the original experiment the bacilli were found to have 
multiphed abundantly in the tumour at the seat of inoculation. The 
eruption might have been, as admitted by Klein, a symptom of the 
work of the chemical poison, and the elimination of the bacilli by the 
milk is also possible ; but that there is in cows a vesicular disease of 
the teats which is the origin of human diphtheria is not accepted by 
veterinarians, and there is not sufficient evidence to justify the con- 
clusion that the infectivity of the milk in epidemics of milk diphtheria, 
has been proved to be due to a morbid condition of the cow. 



CHAPTER XXIV. 

TYPHOID FEVER. 

Typhoid Feveh, is a specific febrile disease peculiar to man, with 
characteristic pathological lesions in the intestine, mesenteric 
glands, and spleen. The Peyer's glands pass through three stages. 
They become swollen from infiltration of round cells in lymph 
follicles, due, it is supposed, to the presence of the typhoid fever 
bacillus. The enlargement of the lymph follicles is followed by 
coagulation necrosis until the entire patch becomes necrosed, and 
sloughs away, leaving an ulcer. The disintegration of the patch 
may extend in depth, and result in perforation and peritonitis, or 
the ulcer may heal, and a pigmented scar take the place of the 
Peyer's patch. The lymphatic glands are found more or less 
enlarged, and may be easily felt in the groin, axilla, and neck. In 
some cases there is a tendency, to haemorrhage, followed by infarctions 
in the spleen and lungs, which may develop into pysemic abscesses 
In the mesenteric glands similar changes take place, but without 
ulceration. Pneumonia may occur as a pulmonary complication. 
The bacteria of pneumonia and Streptococcus pyogenes may be 
found in association with the bacillus of typhoid fever. It is 
now generally accepted that the disease is conveyed by water and 
food which have become contaminated with the virus contained 
in typhoid evacuations. This has been practically proved by the 
number of cases which have been shown to have been intimately 
connected with contamination of drinking water from wells and 
other sources, by sewers, cesspools and faulty drains, the sewage 
presumably having been infected with typhoid excreta ; but whether 
sewage independently of typhoid contamination can originate 
typhoid is still an open qiiestion. Accepting the former theory 
as a working hypothesis, we must assume that a typhoid fever 
bacillus exists in the intestinal evacuations, and that it must be able 
to retain its vitality under very varying conditions until it gains 

310 



TYPHOID FEVER. 



341 



access by the mouth to a fresh host, and by its development in the 
intestine, and by the absorption of its toxic products, produces the 
phenomena which we recognise as typhoid fever. 




Fia. 129.— Typhoid Fever. Ileum of Adult, showing Sloughy and 
Infiltrated Patches (Hamilton). 

Typhoid fever is also disseminated by milk ; sewage- contaminated 
water having been added to the milk, or used for washing the milk 
cans and other vessels. 



342 



INFECTIVE DISEASES. 



Typhoid fever cannot be communicated to the lower animals. 
iSTiimerous experiments have been made bv feeding and by injecting 
typhoid stools, but with absolutely negative results. Murchison 
gave typhoid fever discharges to pigs, Klein experimented with 
rabbits, monkeys, and other animals. Motschutkowsky injected the 
blood from cases of typhoid into monkeys, rabbits, and other animals, 
but with negative results. 




Tig. 130.— Typhoid Bacilli from a Colony on Nutrient Gelatine, x 1000 
(Fkankel and Pfeiffek). 



hem 



Various micro-organisms have been described in typhoid, but the 
one to which most importance is attached is a bacillus which was 
first discovered by Eberth, but cultivated and fully described by 
Gaffky. Gaffky cultivated it from typhoid 
evacuations, from typhoid ulcers, from 
the mesenteric glands, and from the 
spleen. It is found in scattered colonies 
in the spleen, and is rarely if ever present 
in the blood. 

Bacillus of Typhoid Fever — Kods 
1 to 3 /i in length, and -5 to '8 /x in breadth, and threads (Plate VIII., 
Fig. 2). Spore-formation has not been observed, but the protoplasm 
may be broken vip, producing appearances which may be mistaken 
for spores. They are actively motile, and provided some with a single 
and others with very numerous flagella, which are from three to five 
times as long as the bacilli. They stain well with aqueous solutions 



Fig. 131. — Typhoid Bacilli, 
X 950 (Baumgaeten). 



TYPHOID FEVER. 



343 



of aniline dyes, and grow well at the temperature of the room. In 
plate-cultivations minute colonies are visible in thirty-six to forty- 
eight hours ; they are circular or oval, with an irregular margin ; they 
appear granular by transmitted light, and are yellowish-brown in 
colour. Cultivated in the depth of gelatine a well-defined shiny film 
forms at the point of puncture, and a greyish-white filament, com- 
posed of closely packed colonies, develops in the track of the needle 
(Fig. 1.34). On the surface of gelatine a greyish-white translucent 
film forms, with sharply defined margin (Plate II., Fig. 2). On agar 
there is a whitish transparent layer. They flourish in milk. On 
potato at the temperature of the blood there is no culture visible, but 




Fig. 132. — Flagella of Typhoid Bacilli, x 1000, stained by Lofflek's 
Method (Feankel and Pfeiffek). 



the inoculated area appears moist and shining, and cover-glass pre- 
parations made from the potato will demonstrate that there is really 
a copious growth of the bacillus. This almost invisible growth is not 
peculiar to this micro-organism. 

Whether this bacillus is really peculiar to typhoid is much dis- 
puted. Bacilli very closely resembling it, if not actually identical, 
have been found under other conditions. These pseudo-typhoid bacilli 
are regarded by some bacteriologists as varieties resulting from the 
different environment afforded by a saprophytic existence. Numer- 
ous experiments have been made on animals with p)ure-cultures of 
the bacillus, but in the production of typhoid fever they have 
been no more successful than the experiments with typhoid stools. 



344 



INFECTIVE DISEASES. 



Frankel and Simmonds inoculated a number of rabbits in the vein of 
the ear, producing death, in some eases in forty-eight hours. Seitz 
administered broth-cultures by Koch's method of introducing them 

into the stomach after 
the administration of 
opium in guinea-pigs, 
and death resulted in 
several instances. 
But in all these cases 
the results depended 
upon the jjoisonous 
products found in the 




Fig. 133. — Colonies or Typhoid Bacillus. 
Three days old. x 100 (Fbankel and Pfeiffek). 



■cultivations, a similar result following the 
injection of sterilised cultures. An account of 
the products has already been given (p. 41). 

Cassedebat isolated three species of bacilli 
from water, which could be distinguished 
with great diiBculty, and only after the most 
■careful comparison. The bacillus which most 
■closely resembles it is the Bacillus coli com- 
munis ; in fact, Ptoux regards it as a non- 
pathogenic variety of the typhoid bacillus. 
Others claim to be able to distinguish it by 
careful comparison and the application of 
tests. Special importance is attached to 
potato cultures, the typhoid bacillus forming 
an invisible film, and Bacillus coli communis 
a well-marked yellowish growth. Terni 
pointed out that Bacillus typhosus retains its 
motility in media containing hydrochloric 
acid, while Bacillus coli communis and other 







Fig. 134. — Purb-Cultuke 
OF Typhoid Bacilli 
inoculated in the 
Depth of Nutrient 
Gelatine (Baumgab- 
ten). 

bacilli resembling those 



of typhoid lost their motility. Schild maintained that Bacillus typhosus 



TYPHOID FEVER. 345 

was destroyed by exposure to the vapour of formalin, while Bacillus 
coli communis and similar bacilli isolated from water gave subcul- 
tures after exposure for two hours. Typhoid bacilli do not give the 
reaction for indol, and there is no development of gas in cultures in 
the depth of nutrient agar containing 2 per cent, of grape-sugar. 
According to Miiller, sterilised milk is coagulated in twenty-four 
hours, at 37° C, by Bacillus coli communis, but not by the Bacillus 
typhosus until several weeks have elapsed ; and, further, cultures on 
acid potato give different results. The typhoid bacillus on micro- 
scopical examination shows marked polar staining, but Bacillus 
coli communis only shows a shght tendency of the protoplasm to 
break up. 



I-. 



fm^ 






^©S^ 







Via. 135. — Typhoid Bacilli in a Section of Spleen, x 800 (Flugge). 

Kitasato suggested the negative indol test, and recommended 
Salkowski's method. Broth-cultivations are treated with a solution of 
sodium or potassium nitrite : 1 cc. of the nitrite solution ('02 gr. to 
100 cc. of water) is added to 10 cc. of a broth-culture after twenty- 
four hours in the incubator, and on adding a few drops of strong 
sulphuric acid, the typhoid cultures i-emain colourless, but cultures 
of bacilli apparently identical give the red colour. On the other 
hand, Losener maintains that he has cultivated from earth, water 
and healthy human evacuations bacilli which could not be distin- 
guished from typhoid bacilli by any of these tests. 

The detection of the typhoid bacillus in water has been 



346 



INFECTIVE DISEASES. 



described in another chapter (p. 147) ; but sufficient has been said to 
show that bacteriological reports in which it is stated that the typhoid 
fever bacillus has been found in water causing typhoid epidemics 
must be accepted with great reserve ; and further, no one is justified 
in stating that the typhoid fever bacillus is undoubtedly the cause 
of typhoid fever. It is not found in every case of typhoid, it is 
not found in the blood, but it is found in those tissues which are 






'■z%*'^^y--^^S^^B^MM 




Tig. 136. — Typhoid Bacilli in a Section oi' Intestine, invading theSobmucous 
(T.J.) AND MusouLAK Layers (M.), x 950 (B.aumgaeten). 



commonly the seat of secondary invasion of epiphytic bacteria, 
whose normal habitat is the intestinal canal. 

Lastly, as the disease does not exist in the lower animals, the 
crucial test cannot be applied. The etiology of typhoid fever is 
still enveloped in doubt, and the nature of the contagiam has not 
yet been detei'mined. 



CHAPTER XXV. 

SWINE FEVER. 

Pig Typhoid, or swine fever, is a highly contagious disease peculiar 
to swine, causing death in from ten to thirty days, associated with 
a fibrinous pneumonia, enlargement of, and haemorrhage into, the 
lymphatic glands, and characteristic ulcers of the mucous membrane 
of the stomach and intestines. The lesions may assume the form of 
extensive croupous or diphtheritic deposit, which may fill the intes- 
tinal tube. But the most characteristic appearance results when 
the lower part of the ileum and commencement of the colon is 
dotted all over with elevations of the mucous membrane, resembling 
leather buttons or nux vomica seeds, and sometimes with concentric 
rings, so that they have been compared to sKces of calumba root. 

Swine fever is difficult to detect in the early stage, and sometimes 
symptoms are absent altogether in animals suffering from the 
disease and quite capable of transmitting it ; or nothing may be 
noted except cough, and possibly enlargement of the inguinal glands. 
In typical cases the animals are noticed not to feed, to exhibit 
dulness, and to have occasional rigors. Partial paralysis may 
follow, producing unsteady gait or loss of power over the hind legs. 
Diarrhoea sets in, and the evacuations become blood stained. There 
is occasionally a diffused or patchy reddish or purplish rash on 
the skin. After death the appearances most commonly found are 
inflammation of the peritoneum, and redness and enlargement of the 
mesenteric glands and the lymphatic glands in the lungs. There 
is generally lalceration, especially of the colon and ileo-csecal valve, 
or a diphtheritic exudation, sometimes pale yellow, more commonly 
greyish or black, similar to the centres of necrosis within the ulcers. 
The spleen is enlarged and liver congested, and there are haemorrhages 
in the kidneys. As the lungs are so commonly affected, Klein 
proposed the name pneumo-enteritis ; but the pulmonary lesions 
are not constant. Indeed, the cases in which the intestines and 

•347 



348 I?JFECT1VE DISEASES. 

lungs are simultaneously affected are not numerous, and sometimes 
the lungs may be foimd to be perfectly healthy in cases with 




Fig. 137. — Ulceeation of the Intestine in a Typical Case op Swine-Fevee. 

long-standing lesion of the intestine. Old pigs may linger on for 
weeks, and ultimately recover, and in the meantime act as centres 
for the dissemination of the disease. 



DESCRIPTION OF PLATES IX. AND X. 
Swine Fever. 

Plate IX. — Part of intestine from a typical case of swine fever, showing 

scattered ulcers and ulceration of the ileo-osoal valve. 
Plate X. — From the same case of swine fever. The lungs were extensively 

inflamed and partly consolidated, and the lymphatic glands were enlarged 

and of a deep red or reddish-purple colour 



Plate^ DC. 




y < 



■asf" 




■^' X J 



SWINE FEVER. 



KmcM £rooi,,i)a, 4ion,J.H. 



PlateX, 




SWINE FEVER. 



ti. Orodkthi.nic.fiid't. 



Knc^nt Brrjoks.Day 4Jo)i.,.LitJL, 



SWINE FEVER. 



349 



/ 



<iP 




Fig. 1.3S.- 



-B.iciLLUs 01'' Swine-Fever 
No. 1. (Klein.) 



Budd first pointed out that this disease might be compared 
to human typhoid, both diseases being attended by a peculiar 
ulceration of the intestinal folhcles ; but the diseases are not to 
be considered in any sense identical or interchangeable. 

Bacteria in Swine Fever. 
— In 1877 Klein published a 
research in a Report to the Local 
Government Board, in which 
he claimed to have discovered 
bacilli characteristic of the 
disease. They were described as 
similar to Bacillus subtilis, or 
Bacillus anthracis, but smaller 
in size. These bacilli developed 
into long leptothrix filaments, and formed spores. It was further 
asserted that on inoculation, cultures produced lesions indicative of 
swine fever ; the bacilli were also pathogenic in mice and rabbits. 

Later this bacillus was re- 
nounced in favour of another. 
In the following year Det- 
mers described a bacillus, but 
subsequently renounced it in 
favour of a micrococcus. 

In 1882 Pasteur maintained 
that the virus of swine fever in 
France (rouget) was a dumb-bell 
micrococcus, which produced the 
same effect in pigeons as the 
microbe of fowl-cholera. Though 
rouget or swine measles is probably a different disease, the occurrence 
of this micro-organism is of interest in this connection. 

In 188.3 Klein again investigated swine fever, and discovered 
Bacillus No. 2, and maintained 

that these bacilli were found in ■^'■^"'^ 

the blood, in the pieritoneal and ^ ^ ^ .="',,, ^ 

bronchial exudations ; and in the '* v.^ ==--.■. ^^ 

air vesicles of the lungs, in the '~^*' i== 

form of leptothrix filaments ten 
or twenty times the length of 
single rods. Cultivations were 
made on solid media. The organisms in these cultures were minute 
rods actively motile, occurring singly or forming chains, two or three 




Fig. 139. — Bacillus No. 2. Fitosi a 
Pkepakatiok or Bronchial Muons 
OF A Pig. (Klein.) 



Fig. 140.— Bacillus No. 2. From an 
Artificial Culture. (Klein. ) 



350 



INFECTIVE DISEASES. 






times as long as Bacterium termo; and in preparations made from 
diseased organs they were found to possess a very narrow trans- 
parent halo, a sort of hyaline gelatinous capsule, Inoculation of 
cultures failed to produce the lesions found in animals naturally 
infected. Two pigs were inoculated, one with a sub-culture from the 
swollen bronchial gland of a pig that had died of pig-typhoid, and 
a second with a culture obtained from the spleen of a mouse that 
had been inoculated from another case of swine fever. After two 
days the inguinal glands near the seat of inoculation became swollen, 
and the temperature rose slightly. After three or four weeks the 
aiuimals recovered. 

' Mice on the fifth or sixth day after inoculation showed symptoms 

of illness, then respiration became 
superficial and slow. Death 
occurred on the sixth or seventh 
day. 

Rabbits showed a rise of 
temperature, and death followed 
between the fifth and eighth days, 
the temperature falling before 
death. At the post-mortem ex- 
amination there was usually 
peiitonitis, with copious exuda- 
tion. The kidney, spleen, and 
liver were enlarged and dark in 
many cases, there was red hepa- 
tisation of the lobes of the lungs, 
and generally pericarditis and hfemorrhage under the pericardium. 

In 1885 Salmon, in the annual report of the United States 
Bureau of Animal Industry, published the result of his investigations 
into American hog cholera, which is identical with English pig typhoid. 
A motile figure-of-eight bacterium was isolated, each part being 
about twice as long as broad. The bacterium grew on nutrient 
gelatine without liquefying it, and on potato produced a brownish 
growth ; broth tubes became turbid on the following day. Colonies 
in plate- cultivations were oval or circular, and brownish in colour. 
Six pigs inoculated subcutaneously were all said to have died of hog 
cholet-a, and the bacterium was again obtained from the blood of 
the heart and spleen. The bacteria proved fatal to mice, rabbits, 
guinea-pigs, and pigeons. 

In 1893 Welch and Clement described the hog-cholera bacillus 
as variable in form, and they further stated that a culture obtained 




Fig. 141.— Blood oi? Feesh Spleen of 
A Mouse, aftee Inoculation with 
Bacillus No. 2. (Klein.) 



SWINE FEVER. 351 

from Klein, while not possessing the characters originally ascribed 
to it, could not in its form, biological characters, or pathogenic pro- 
perties, be distinguished from the American hog-cholera bacillus. 

In 1887 pig typhoid was investigated at Marseilles by Rietsch, 
Jobert, and Martinaud, and a bacillus found. This grew rapidly 
on all the nutrient media. In gelatine a growth was obtained in 
twenty-four hours at 18° 0. ; on blood serum and agar an opaque 
growth developed ; and on potato the growth was yellowish. It 
was asserted that 'a young pig was killed by a culture in twenty- 
two days, and that the characteristic ulcerations were observed in 
the intestines. 

In 1887 pig typhoid was prevalent in Sweden, and Bang and 
Selander experimented with cultui'es from a rabbit that died after 
inoculation with a fragment of spleen from a diseased pig. The 
bacilli were motile, varying from rods to cocci, without spore-formation 
and pathogenic in mice, guinea-pigs, and rabbits, but not in pigeons. 
Pigs fed on broth-cultures were said to have succumbed to genuine 
pig typhoid. In the blood they were generally found in the form 
of short oval bacteria, but in the blood of the heart longer rods 
were sometimes found. MetchnikofI described a bacillus isolated by 
Chantemesse from an outbreak in France, as highly polymorphic. 

Smith identified the hog-cholera bacillus with the bacillus found 
by Schiitz, and this in turn has been identified with the bacillus 
of hsemorrhagic septicaemia. 

From these researches it would appear to be probable that one of 
the bacteria isolated by Klein, and those found by Salmon, Smith, 
Bang, Welch and Schiitz are identical ; and further, that they have 
been identified with the bacillus of hsemorrhagic septicaemia. We 
may sum up the characters thus : — 

Bacillus of Klein, Salmon, Smith and Schiitz Very small 

rods, actively motile ; spore-formation not observed ; colonies circular 
and brown by transmitted light. Inoculated in the depth of gelatine 
faintly yellowish-white colonies develop along the track of the needle ; 
on the surface an opalescent film ; on potato they produce a straw- 
coloured layer, changing to brown. There is absence of indol in 
cultures containing peptone; the bacilli are fatal to mice, guinea- 
pigs, rabbits, and pigeons. Swine die after intravenous injection, 
l)ut not, as a rule, after subcutaneous injection. •• 

According to Caneva, the bacillus obtained from the Marseilles 
epidemic would appear to be closely allied, if not identical, with 
the bacillus of ferret disease (Eberth and Schimmelbusch), and the 
bacillus of Texas fever (Billings). 



352 INFECTIVE DISEASES. 

Billings appears to have isolated two bacilli, one identical with 
the Marseilles bacillus, and the other with the hog-cholera bacillus. 

Bacillus of Rietsch and Jobert. — Rods about twice as 
long as broad, rather shorter than the bacillus of typhoid fever, 
longer and thicker than the American, bacillus. They exhibit 
end-staining. They possess flagella, and are actively motile. 

They grow rapidly in nutrient media. They are only feebly 
pathogenic. They are also said to be distinguished from the bacilli 
of hog-cholera by producing indol in solutions containing peptone, 
and by causing an acid reaction in milk. 

Bacillus of M'Fadyean. — ^M'Fadyean investigated swine fever 
in 1895, and found bacilli which he differentiated from hog-cholera 
bacUli. The method employed was to inoculate the surface of 
nutrient agar and of potato, with fragments torn out of the centre 
of a lymphatic gland, with specially constructed forceps. Inocula- 
tions were also made from the spleen pulp, and blqod, in the usual 
way. They are from 1 to 2 //. in length, and •& fj, in breadth. 
They never grow into filaments, they do not form spores, and 
they are actively motile. They are readily stained by the watery 
solutions of the aniline dyes, and are decolorised by Gram's 
method ; with methylene-blue they show end-staining. The bacilli 
grow on gelatine without liquefaction, forming a thin white line 
along the needle track. On agar a thin, transparent pelUcle forms, 
which is not easily visible at first, but gradually acquires a faint 
greyish tint. More characteristic appearances result in plate- 
cultivations of gelatine-agar, at 37° C. The colonies are distinctly 
visible in eighteen hours, appearing when viewed by transmitted 
light as bluish-wliite, circular specks ; each colony has a dark centre 
and a granular margin. In broth the bacilli produce turbidity after 
twenty-four hours. On potato there is no visible growth, even when 
the surface is inoculated with an abundance of material. On sohd 
blood serum the growth is scanty. They grow in milk without 
producing coagulation. They are harmless to guinea-pigs and feebly 
pathogenic to rabbits. 

Several experiments were carried out upon swine. In the first, 
series the most rigid precautions were taken to prevent accidental 
infection with swine fever. Four young pigs were inoculated, upon 
a farm where there was no previous history of the disease. These 
pigs were killed, and the post-mortem examinations were said to 
show indications of swine fever, principally patches of diphtheritic, 
material in the colon, and healing ulcers. The next series of pigs 
was inoculated at the Royal Veterinary College. Cultures were 



SWINE FEVEK. 353 

administered with milk to two pigs. Five days after wai-ds one pig 
died ; and the mesenteric glands were congested, and the mucous 
membrane showed spots of necrosis. The other pig was killed, and 
there were ulcers in the colon. In a third experiment, eight pigs 
were fed with milk and broth cultures. These pigs were all killed 
at different dates, and most of them had ulceration of the colon ; 
in control experiments the intestine was normal. 

MTadyean compared his bacillus with a culture of the hog- 
cholera bacillus, and found that the American organism grows at a 
lower temperature in gelatine, and colonies appear in plates much 
earlier. They produce a less transparent and thicker growth, and 
much greater turbidity in broth and a more abundant sediment. 
On potato they form an abundant growth at 37° C, at first 
yellow, later brown, with considerable resem.blance to a glanders 
culture. 

Colonies xipon gelatine-agar are distinguished by their opacity 
and sharp outKne. Agar, potato, and broth cultures of the American 
organism consist of short ovoid forms like the bacilli of fowl cholera, 
while the bacillus isolated by M'Fadyean has a closer resemblance 
to the bacillus of glanders. M'Fadyean asserts that the American 
organism is not pathogenic to the pig. Pigs after feeding on broth 
cultures remained healthy, and showed no trace of swine fever when 
killed from one to three weeks afterwards. On the other hand, 
broth cultures of his bacillus produced the characteristic ulceration 
of the bowel. M'Fadyean claims, therefore, to have discovered the 
true pathogenic organism of swine fever. He does not appear to 
have compared this bacillus with that obtained from the epidemic 
of swine fever at Marseilles. From ttie description of the mor- 
phological and other details there seems to be a close resemblance 
between the two. 

Not less than three and possibly four species of bacilli have been 
cultivated from swine fever, two at different times by Klein, one 
by Reitsch Jobert and Martinaud, and one by M'Fadyean ; and 
cultures of all these bacUK have been credited with producing swine 
fever in experimental animals, and each one has been pronounced 
to be the contagium of the disease. We must conclude either that 
contaminated cultures were inoculated in some cases, or, what is 
far more probable, the swine fever which resulted in experimental 
animals was due to accidental infection; and until a bacillus has 
been cultivated from swine fever from which another investigator 
can prepare sub-cultures, and with those sub-cultures produce the 
typical ulcerations of swine fever in pigs on a farm, or on premises 

23 



354 INFECTIVE DISEASES. 

in which swine fever is unknown, we are justified in concluding that 
the contagium has not yet been discovered. 

The mistakes which are likely to occur when the same investi- 
gator isolates bacilli from cases of swine fever, and subsequently 
inoculates or feeds healthy swine, cannot be better illustrated than 
by quoting from a leaflet issued by the Board of Agriculture, 
pointing out the great precautions necessary to prevent accidental 
infection. 

"There seems reason to beUeve that the disease is not infre- 
quently introduced by means of persons who have been in contact 
with diseased animals. Pig owners, therefore, are advised to prevent 
strangers from at any time approaching their pigs, and should the 
admission to the premises of spaj'ers or castrators be necessary, 
those persons should be required, before approaching the animals, to 
thoroughly wash their hands with soap and water, and to wash and 
■disinfect their boots with a solution of carbolic acid and water, or 
some other suitable disinfectant. Such persons might also with 
advantage be required to wear, while operating, a waterproof apron, 
which should be washed and disinfected before the wearer is per- 
naitted to approach the animals to be operated on." 

Protective Inoculation. — The experiments of Salmon and of 
Schweinitz have been referred to in another chapter (pp. 41, 46). 
A method of protective inoculation was attempted in America, but 
the experiments were unsuccessful, and the plan was abandoned. 

Stamping-out System Notification is compulsory, and the 

order in force is the Swine Fever Order of 1896, but the stamping- 
out system has not been applied in a thoroughly satisfactory manner, 
and the disease is still very prevalent. 



CHAPTER XXVI. 

swine measles. distemper in docxs. — epidemic disease of 

ferrets. — epidemic disease of mice. 

Swine Measles. 

Swine Measles, or swine erysipelas, is described as an acute, in- 
fectious disease of swine which is very prevalent in France and 
Germany, biit is included in this country in the term " swine fever." 
According to some, it is a distinct disease, and distinguished from pig- 
typhoid by absence of the ulceration of the intestines which is so 
characteristic of that disease ; while, according to others, ulceration 
of the intestine and ileo-ccecal valve may be found post-mortem. The 
onset of the .symptoms, as in pig typhoid, is very rapid ; the animals 
cease to feed, and show other general signs of illness ; the voice is 
hoarse, and there is a rapid rise of temperature. On the neck, 
chest, and abdomen, red patches make their appearance, which 
extend and coalesce, and change to a dark reddish or brownish colour. 
These symptoms may be followed by convvilsions, and sometimes by 
paralysis of the hind legs ; and death occurs in from one to four 
days. It is especially a disease of young pigs, and from 50 to 60 
per cent, of infected animals die. 

On post-mortem examination there is hsemorrhage and oedema 
in the patches of the skin, the lymphatic glands are swollen and 
dark red, the peritoneum is ecchymosed, the intestinal mucous 
membrane is congested and swollen, and the solitary follicles and 
Peyer's patches are prominent, and in the neighbourhood of the ileo- 
coecal valve there are, according to Fliigge, ulcers of considerable size. 
The liver and spleen are congested and enlarged. Pasteur investi- 
gated swine measles or rouget, and described a figure-of-eight micro- 
coccus, which he believed to be the contagium of this disease. This 
organism appears to be identical with the bacterium of hsemorrhagic 
septicaemia, which is also commonly found in pig-typhoid. 

In experimenting with the virus obtained from the spleen 

365 



356 INFECTIVE DISEASES. 

Pasteur found that, by successive inoculation of rabbits, the virulence 
was exalted for rabbits, but attenuated for swine, and the virus 
which had thus been passed through the rabbit was used as a 
vaccine for swine, to protect them against virulent erysipelas. 

Pasteui' found that by passing the wus through pigeons it was 
made more virulent for swine. 

In the blood, and the juice of the internal organs, and of the 
lymph glands, Schiitz found a minute bacillus identical with the 
bacillus of mouse septictemia. 

Bacillus of Swine Erysipelas (Schiitz). — Extremely uiinute 
rods '6 to 1'8 ft in length, morphologically and in cultural charac- 
ters identical with the bacillus of mouse septicaemia. Filaments 
and involution forms. Spore-formation present. 

House mice if inoculated with a pure culture die in two to four 
days. Pigeons are also \evy susceptible. Fowls and guinea-pigs 





Fig. 142. — Bacilli of Swike Fig. 143. — Blood of Pigeon iNOcUL.iTED 
Feysipelas (Baumgaeten). with Bacilli of Swine ERrsiPELAS, x 600 

(SCHIJTZ). 

are immune. E-abbits after inoculation of the ear suffer from 
erysipelatous inflamm;ition, identical with that produced by inocu- 
lation of the bacillus of mouse septicaemia. The bacilli are also 
pathogenic in swine and sheep. 

Protective Inoculation. — With Pasteur's \accine immunity is 
said to be produced which lasts about a year. Schiitz and Schottelius 
found the minute bacilli in Pasteur's vaccine, which they had already 
found in cases of swine erysipelas in Germany. 

The results of vaccination in France are said to be very satis- 
factory, but in test experiments in Germany they were not so 
favourable. Out of 119 vaccinated swine 5 per cent, died a.s the 
i-esult of the inoculation, while the average loss in the ordinary way 
is 2 per cent. 

Metchnikoff found that the blood of immunised raljbits was 
antitoxic, and Lorenz maintains that the serum of swine which 



SWINE FEVER. 



357 



and aftei' 
294 pigs; 



have recovered from swine erysipelas is also antitoxic, and will 
produce immunity in other animals. The treatment introduced 
by Lorenz is to inject serum in the proportion of 1 co. to every 
10 kilogrammes of the weight of the animal's body. Two days 
afterwards '5 to 1 cc. of virulent culture is injected 
twelve days the dose is doubled. Lorenz inoculated 
12 were suffering from swine erysipelas, and 
of these 6 recovered and 6 died. 

In the opinion of the author this disease 
requires re-investigation, for if it be true that 
rouget or schweinrothlauf is associated with 
ulceration of the intestines, the recognition 
of it as a thsease distinct from our English 
swine fever apparently rests npon the pres- 
ence of a bacillus, which cannot be distin- 
guished from the bacillus of mouse septicEcmia. 
The question arises whether this bacillus 
is really the cause of a distinct disease, swine 
erysipelas, or, on the other hand, whether 
the bacillus is really the bacillus of mouse 
septicaemia which has been isolated from 
certain cases of swine fever. The bacillus of 
mouse septicaemia is widely distributed, aiad 
it may only lie an accidental concomitant in 
rouget or sclaveinrothlaitf. The pi'esence of 
the bacterium of hsemorrhagic septicemia in 
both rouget and pig typhoid would not prove 
identity, as this micro-organism is un- 
doubtedly only secondarj' in both diseases. 
There is great need, therefore, for further 
careful investigation. Clinical and jjatho- 
logical observations must be made in this 
country, to determiiie whether there are 
reaUy two diseases included under the name 

" swine fever." If this prove to be the case, we must ascertain the 
chnical and pathological differences between rouget and pig typhoid. 
How can rouget be distinguished from cases of swine fever in which 
there is a patchy rash, paralysis of hind legs, but no ulceration of the 
intestine ? Further, how is swine erysipelas with ulceration of the 
intestine and ileo-ccccal valve to be distinguished from an ordinary 
case of pig typhoid ? 




Fig. 14-1. — Puke-chltube 
IN Nutrient Gela- 
tine OF Bacilli 
IN Swine Erysipelas 
(Baumgakten). 



358 INFECTIVE DISEASES. 



DiSTEMPEE IN Dogs. 



Distemper is an infectious febrile disease of dogs, characterised by 
bronchial catarrh and discharge from the eyes. Bronchitis and 
pneumonia may supervene, or there may be intestinal catarrh ter- 
minating in dysenteric diarrhoea, sometimes complicated by jaundice. 
The disease may affect the nervous system, and produce convulsive 
contractions of the muscles of the nose, ears, lips, and Umbs. 
Occasionally there is an eruption, especially in animals which are 
out of condition. The virus exists in the discharge from the nostrUs 
and eyes, and is given off from the lungs and the skin. 

One attack of the disease does not confer entire immunity ; 
and some dogs are completely insusceptible. 

Bacteria in Distemper. — Millais has isolated a micro-organism 
resembling the pneumococcus of Friedlander, which he beheves to be 
the cause of the disease. The bacillus occurs with other bacteria and 
micrococci in the nasal discharge. 

Protective Inoculation Mixed cultures of these bacteria 

liquefy the gelatine, and the liquid has been used as a vaccine. 
When applied to the nose, it is said to produce a mild attack of 
distemper, which protects as much as an attack of the disease 
contracted naturallj'. These results require confirmation. 

Inoculation of the nasal discharge in healthy dogs . has been 
practised, so that they may have ^he disease under favourable con- 
ditions ; but the system should not be encouraged, as dogs need not 
necessarily contract distemper. Vaccination with cow-pox lymph 
has been advocated, but it is perfectly useless. 

Stamping-out System — Dogs suffering from distemper must be 
completely isolated. Any straw or litter which has been in contact 
with a diseased dog should be burnt. Clothing, collars, chains, and 
the kennel or premises inhabited, must be thoroughly disinfected. 
The animal after recovery should be washed with carbolic soap. 

Epidemic Disease of Ferrets. 
Eberth and Sohimmelbusch investigated an epidemic disease of 
ferrets {fretfchen-seuche), and isolated a bacillus, which in mor- 
phology and cultivation agrees very closely with the bacillus of 
hEemorrhagic septicaemia. 

Epidemic Disease of Mice. 
Lbfiler investigated an epidemic disease which occurred in mice 
kept in confinement, and isolated a bacillus resembling Bacillus 
typhosus. 



EPIDEMIC DISEASE OF MICE. 359 

Bacillus Typhi Murium. — Rods varying in length ; and fila- 
ments ; motile ; flagellated. The colonies are circular, brownish 
and granular on the surface of obliquely solidified gelatine. The 
bacteria inoculated on the surface produce a greyish-white semi- 
transparent growth, and on agar and potato the appearance of the 
growth is very similar. They can be cultivated readily in milk and 
in broth. White and field mice are killed in from one to two 
weeks, when given bread moistened with a culture. 

Lbfiler claims to have used this method with success in Thessaly, 
where there was a plague of field mice causing great losses to 
agriculturists. 



CHAPTER XXVII. 

ASIATIC CHOLERA. CHOLERA NOSTRAS. — CHOLERAIC DIARRH(EA 

FROM MEAT-POISONING. — DYSENTERY. CHOLERAIC DIARRHCEA 

IN FOWLS. 

Asiatic Cholera. 

There are several diseases in man associated with diarrhoea, which 
have certain characters in common, but are totally distinct. They 
include Asiatic cholera, cholera nostras, dysentery, and choleraic 
diarrhoea. Asiatic cholera is an endemic disease of the Delta of 
the Ganges, a locality which has become notorious as the home 
of cholera. Cholera is a filth disease ; and the accumulation of 
filth on the banks of the Ganges, with contamination of the 
water, and the climate, afford most favourable conditions for the 
development of the cholera virus. 

Four great cholera epidemics have originated in, and spread 
from, India: in 1817, in 1826, in 1846, and in 1865. Cholera 
follows the routes of pilgrims and caravans, and now, owing to 
the rapid means of communication by steamers and railways, it 
spreads to the most distant parts of the world, covering in a few 
weeks or days distances which in former times couldonly be traversed 
in several months or even years. 

In 1892 the epidemic passed from India, through Afghanistan, 
to Russia in Asia, and quickly spread westwards along the route 
of the trans-Caspian railway ; and all this occurred within the space 
of a few weeks. By Russian emigrants it was carried to Hamburg 
and Antwerp ; and the virus, finding a suitable environment in the 
former place, produced a severe epidemic there. Thus, in about three 
months, it was brought into close proximity with England. Mecca 
is one of the great infective centres of the world, for there all 
the conditions are found for the propagation of cholera, including 
filth, overcrowding, and the water of the famous Holy Well, which 
is used for ablutions and drinking purposes. The return of 

3G0 



ASIATIC CHOLERA. 361 

the pilgrims to Egypt, and the proximity of England to Egypt, 
necessitate the greatest possible precautions to prevent the intro- 
duction of the disease into this country. 

In 1884 a German Commission was sent out to India, and Koch 
discovered a micro-organism which he described as a curved or 
comma-shaped bacillus, and pronounced to be the contagium of 
this disease. 







Fig. 145. — Covee-glass Pkeparation op a Drop of Meat Infusion, containing 
a pure-cultivation of comma-baciUi, with {a) spiriUiform threads, x 600. (Koch.) 

Spirillum cholerse Asiaticse {Comma-hacillus, Koch). — Curved 
rods, spirUla, and threads. The curved rods or commas are about 
half the length of a tubercle-bacillus. They occur isolated, or 
attached to each other forming S "Shaped organisms or longer 
screw-forms, the latter resembling the spirilla of relapsing fever. 




>~:^^^ 



Pig. 146. — Aethkospores ; (a) Cotoma-bacillu.5 breaking up into spheres; (6, c), 
formation of spheres in spiral forms ; (d, e), groups of spheres ; (/) spirilla 
with spheres from an old cultivation; (g) germination of the spheres. 

(HUEPPE.) 

Fiaally they may develop into spirilhform threads. In old cultiva- 
tions threads are found with swellings or irregularities (Eig. 148). 
The commas are actively motile, and possess flagella (Fig. 147). 
Their movements] and development into spirilla may be studied in 
drop-cultivations. Arthrospore formation has been described by 
Hueppe (Pig. 146). In plate-cultivations, at a temperature of from 



362 



INFECTIVE DISEASES. 



16° to 20° C, the colonies develop as little specks, which begin to 
be visible aftei- about twenty-four hours. Examined with a low 
power, and a small diaphragm, these colonies have the following 
characteristics. They appear as little masses, granular, and 



r^f 






<f>*. 






» <A 



ir7 




Fig. 147. — Flagella of Comma-bacilli ; stained by Lofflek's Method 
(Fkankel and Pfeiffee). 

yellowish-white in colour, and sometimes very faintly tinged with 
red, which have hquefied the gelatine, and sunk down to the bottom 
of the resulting excavations. 

In test-tubes of slightly alkaline nutrient gelatine (10 per cent.). 









/'■■ 








Fig. 148.— Involution Forms, x 700 
(Van Ermengem). 



Fig. 149.— Colonies of Comma-bacilli 
ON Nutrient Gelatine, Natural 
Size (Koch). 



the appearance of the growth is very striking. In typical cultures 
it begins to be visible in about twenty-four hours. Liquefaction 
sets in very slowly, commencing at the top of the needle track 



ASIATIC CHOLERA. 



363 





around an enclosed bubble of air, and forming a funnel continuous 
with the lower part of the growth ; the latter preserves for several 
days its resemblance to a white 
thread (Plate II., Fig. 1). In 
about eight days, however, lique- 
faction takes place along the 
whole of the needle track. 

On the surface of agar- agar 
the cviltivation develops as a 
white, semi-transparent layer, 
with well-defined margin. The 

appearance on blood serum is very similar ; liquefaction very slowljr 
takes place. In broth they form a wrinkled film on the siirface, 
there is a rapid and abundant growth at the temperature of the 



Fig. 150.— Colonies or Koch's Comma- 
bacilli, X 6U. 



O— 




J/. .,>'' .-' -\ •«'. . ^' 




Fig. 1.51.— Covee-glass Pkepaeation 
FKOM THE Contents of a Cholera 
Intestine, x 600. (a) Remain.sof the 
epithelial cells ; (h) Comma-bacillas ; 
(c) Group of comma-baciUi (Koch). 



Fig. 152.— Covee-glass Peepakatiok 
OP Choleea Dejecta on Damp Linen 
(two days old), x 600. Great prolife- 
ration of the bacilli with spirilla (a) 
(Koch). 



blood, and the same applies to sterihsed milk ; and they will even 
multiply in sterilised water. In potato-cultivations the microbe 
will only grow at the temperature of the blood (37° 0.), forming a 
slightly brown, transparent layer. Inoculation of a cultivation of 
the bacillus in the duodenum of guinea-pigs, with and without 



364 



INFECTIVE DISEASES. 



ligation of the bile-duct, has given positive results. More recently 
these results have been confirmed by the following method : Five 
cc. of a 5 per cent, solution of potash were injected into the 
stomach of a guinea-pig, and twenty minutes after, 10 cc. of a 
cultivation of comma -bacilli, diffused in broth, were similarly intro- 
duced. Simultaneously with the latter, an injection of tincture of 
opium was made into the abdominal cavity, in the proportion of 
1 cc. for every 200 grammes weight of the animal. Those who 
have had success with inoculation experiments maintain that choleraic 
symptoms were produced without any trace of peritonitis or putrid 
infection, and that the comma-bacilli of Koch were again found 
in the intestinal contents, and fresh cultivations established. 




Fig. 153.— Section of the Mucous Membeane of a Choleka Intestine, x 600. 
A tubular gland (a) is divided transversely ; in its interior (6) and between 
the epithelium and the basement membrane (o) are numerous comma-baciUi 
(Koch). 

On the other hand, these results have been disputed, the fatal 
effects of the inoculation attributed to septicaemic poisoning, and 
the proUferation of the bacilh considered to be dependent upon an 
abnormal condition of the intestines, induced by the injection of 
tincture of opium. It has, however, been shown that these organisms, 
like several others which have been isolated from intestinal dis- 
charges, produce definite poisonous substances. The comma-bacUli 
were found in the superficial necrosed layer of the intestine, in 
the mucous flakes and liquid contents of the intestinal canal of 
cases of Asiatic cholera. It is stated that they were also detected 



ASIATIC CHOLERA. 



365 



in a tank which contained the water supply of a neighbourhood 
where cliolera cases occurred ; but comma-shaped organisms are 
frequently present in sewage-contaminated water. Koch's comma- 
baoilh are aerobic, and their development is arrested by deprivation 
of oxygen. They are destroyed by drying on a cover glass, but 
retain their vitality longer when dried on silk threads. Cultures 
are sterilised by exposure for fifteen minutes to 5.5° C, and by 
various antiseptic substances. 



Ill 

V.l 



I I 



u 



', I 



w 



ll I 






Fig. 154. — Pube-cultivatioks in Nutkient Gelatine. «, Koch'.s Choleka 
Bacillus, twenty-tour hours old. b, Finklee'h Bacill-us, twenty-tour 
hours old. 

Methods ui' Staining the Comma-bacilli of Koch. 



In cover-glass preparations they may be well stained in the ordinary 
way, with an aqueous solution of methyl-violet or fuclisiue, or by the 
rapid method, without passing through the flame (p. 85, Babes' method). 

Nicati and EeUxch's Method. 

A small quantity of the stools, or of the scraping of the intestinal 
mucous membrane, is spread out on a glass slide and dried, theu steeped 
during some seconds in sublimate solution, or in osmic acid (1 to 100). 
It is then stained by immersion in fuchsine-aniline solution (1 or 2 
grammes of Bale f uchsine dissolved in a saturated aqueous solution of 
aniline), washed, dried, and mounted in Canada balsam. 



366 INFECTIVE DISEASES. 

In sections of the intestine their presence may be demonstrated by : — 

(a) Koch's method. 

Sections of the intestine, which must be well hardened in absolute 
alcohol, are left for twenty-four hours in a strong, watery solution of 
methylene-blue, or for a shorter time if the solution is warmed ; then 
treated in the usual way. 

(6) Babks' method. 

Sections, preferably from a recent case of cholera, and made as soon as 
possible after death, are left for twenty-four hours in an aqueous solution 
of f uchsiue, then washed in distilled water, faintly acidulated with acetic 
acid, or in sublimate solution 1 in 1000, passed rapidly through alcohol, 
and finally treated in the usual way. 



Klein investigated cholera in India, and does not accept Koch's 
conclusions. With regard to the inoculation experiments, Klein 
believes that the living choleraic comma -bacilli, even if introduced 
in large numbers into the small intestine, are quite innocuous, 
but capable of great multipliqation if the intestine is previously, 
from some cause or another, diseased ; the chemical products of the 
comma-bacilH then act as poisons analogous to the ptomaines 
obtained from other putrefactive bacteria. The observations made 
by Roy, Brown, and Sherrington, in Spain, tended to confirm 
Koch's views. Comma-bacilh were found to be present, in some 
cases, in enormous numbers, and the frequency of their occurrence 
led these observers to believe that they must bear some relation to 

the disease. At the same time, 
as they failed to find them in all 
cases, they regarded the existence 
of a causal relation as not proven. 
They failed to find the Naples 
bacterium, or a small, straight 
bacillus noted by Klein ; and 
they drew attention to certain 
Tig. 155. -Comma-shaped Organisms peculiar mycehum-hke threads 

WITH OTHKK BaCTERIA IN SeWAGE- . ,, , „ ,, 

CONTAMINATED Watee, X 1200. ^^ ^'^^ mucous membrane of the 

intestines ; but these cannot be 
considered to have any significance. Methylene-blue has been 
employed by Koch and others, including the author, for staining 
sections of the intestine from cholera cases, and had they been 
constantly present, it is hardly possible that such striking objects 
could have been overlooked. Again, we must bear in mind that 
hypho-mycetous fungi occasionally have been found to occur sapro- 
phytically in the intestinal canal, as well as in the lungs, external 
auditory meatus, and elsewhere. Cunningham, of Calcutta, maintains 




ASIATIC CHOLERA. 367 

that Koch's comma-bacilli are not constantly found ; and that 

the comma-bacilli obtained from typical cholera cases show a 

great variation in cultivation, and cannot be distinguished from 

comma-bacilli from other sources. 

Ounninsrham asserts that ,,, ., 

comma-bacilli resembling Koch's ifr^^T ''2>.'" 

are found in the intestine in '^C^'^^S^C^V'..'!!, T*-. 

health. Sternberg, on the other 'i.y if V-t -'S V* 

hand, made a number of examina- '^'C-' 

tions of the evacuations of yellow 

, . Fig. 156. —Comma-bacilli OF THE MOUTH, 

fever patients and healthy mdi- ^ ^qq (Van Ekmengem). 

viduals, and failed to find any 

micro-organism resembling the cholera spirillum. 

Various comma-bacilli have been isolated from different sources 
and compared with Koch's comma-bacillus. Comma-bacilli have been 

found in the mouth by Lewis ; 
'X in cholera nostras by Finkler and 

^~Ji^!^i Prior ; in cheese by Deneke ; 

',i.i|iiS;^ V. in hay infusion and sewage by 

^ V.) '■^ ^-^/M^N-^W Weibel ; in the intestines of fowls 

'('' ' f^^%^iK;?^?J''' by Gamaleia, and in water by 
^■'yffs- <'■' Sanarelli. 

3*''*' Whether the comma-bacillus 

Fig. 157.-Finkleb's Comma-bacilli ; is the cause of cholera or 
fbom Cholera nosteas, x 700 not, its detection is an aid in 
(Flugge). diagnosis. If we are dealing 

with a case alleged to be one of Asiatic cholera, and a micro- 
organism is found in the intestinal evacuations, which can be 
differentiated from the comma-bacillus described by Finkler in 
cholera nostras, and identified 

with the comma-bacillus de- y - ^ . 

scribed by Koch, we are justified "" , " ^ ^-- ^,* 

in regarding the case as one of X ^ , <//^o , ~ 

Asiatic cholera. But we cannot ^-~ ' K ^ ^ ^~' ^>'' ^ 

diagnose Koch's comma-bacillus, ' '^^-iT -._ r -^^ 

with certainty, unless we know ^~ 

the source of the culture. The ^^^ 158.— Dbneke's Comma-bacilli, 
clinical symptoms of cholera in j-eom Cheese, x 700 (Flugge). 

man, and especially the presence 

of rice-water stools, must be taken into account, together with 
the biological, morphological, and chemical characteristics of 
the baciUi which are found to be present. There are several 



368 INFECTIVE DISEASES. 

chemical tests which can be applied to cultures. According to 
Frankel, the Bujwid-Dunham test can be relied upon to distin- 
guish Koch's comma-bacillus from the comma-bacillus of Einkler- 
Prior (cholera nostras), and from those found by Gamaleia. The 
comma-bacilK are inoculated in broth containing peptone, and, 
after twelve hours in the incubator, a drop of strong sulphuric 
acid added to the culture will produce a red colour, owing to the 
presence of indol. A test which distinguishes Koch's comma- 
baciUus from Finkler- Prior's and Deneke's was introduced by Cahen. 
A solution of litmus is added to the broth, and the culture placed 
in the incubator, until the following day ; in the case of Koch's 
commas, the colour will have disappeared. 

Koch points out that in the bacteriological diagnosis of cholera 
the first step is to examine the mucus in the evacuations, or in the 
intestine if the examination is made after death. Cover-glass 
preparations should be stained with dilute Ziehl-Neelsen solution. 
Cultures are next made in peptone, and in eight hours will give the 
indol reaction. In twenty-four houi-s the colonies may be examined 
on plate-cultivations. The peptone cultures are prepared by adding 
a trace of the choleraic evacuations, or of mucus containing the 
bacilli, to a sterilised 1 per cent, solution of peptone, with '5 to 1 
per cent, of common salt. The solution must be alkahne, and the 
culture is placed in the incubator at 37° C. The pathogenic effects 
can be ascertained by diffusing the bacilli from an agar-culture in 
broth, and injecting it into the peritoneal cavity. 

Toxic Products. — Brieger isolated several toxic products which he 
had found in association with putrefaction, such as cadaverin and 
putrescin ; but there were also present two new toxic substances, one 
producing cramps and muscular tremors in inoculated animals, and 
the other lowering the temperature and depressing the action of 
the heart. Later, Brieger in conjunction with Frankel, succeeded 
in isolating a tox-albumin from pure cultures. Guinea-pigs were 
killed in two or three days, but rabbits had an immunity. Pfeiffer 
found that cultures contained a poisonous principle which proved 
fatal to guinea-pigs in extremely minute doses. It is broken up 
by alcohol and by boiling, and secondary products formed, of very 
much mitigated virulence. Similar toxic products were obtained 
from cultures of both Finkler-Prior's and Metchnikoff's commas. 

Protective Inoculation — Haffkine has introduced a system of 
protective inoculation, which is founded on the principle of inducing 
the formation of antitoxins, or defensive proteids. Comma-bacilli 
when first cultivated from a cholera patient are not sufficiently 



ASIATIC CHOLERA. 369 

virulent, and the virulence is increased by cultivation in the 
peritoneal cavities of a succession of guinea-pigs. This successive 
cultivation is carried on until a virus is obtained which proves fatal 
in a few hours when inoculated into the peritoneum. A culture 
from the peritoneum is obtained on an agar plate-cultivation, and 
a pure sub-culture on agar is thoroughly shaken up with broth. 
This constitutes the vaccinating fluid. It may be used as a living 
vaccine, or the comma-bacilli killed by the addition of carbolic 
acid. 

Haflfkine, having studied the pathological and physiological effects 
on some sixty persons, mostly scientists interested in the subject, and 
finding the treatment to be harmless, transferred his operations to 
localities in India affected by cholera. The inhabitants of the 
northern part of India were the first to come forward and submit 
themselves to the inoculation. In the course of the first year 
22,703 were inoculated in the North-West Provinces and Oudh, 
and in the Punjab. All classes of] the population were included. 
In the second year operations were carried out in those parts of 
the country where cholera always prevails, and where, therefore, the 
method could be more satisfactorily tested. 

From March 1894, to July 1895, 19,473 individuals were 
inoculated in some of the most affected localities. 

Prom observations made at Calcutta by Dr. Simpson, from March 
1894 to August 1895, cholera occurred in 36 houses containing 
inoculated people. There were 521 inhabitants in the infected 
houses, of whom 181 were inoculated from 1 to 459 days before the 
occurrence, while 340 remained uninoculated. The uninoculated had 
45 cases with 39 deaths from cholera ; the inoculated had 4 deaths, 
1 occurring 451 days after the first inoculation, and 3 others from 
1 to 4 days after the first inoculation. These four cases had not 
been re-inoculated. If the occurrences in inoculated and non-inocu- 
lated during the first 10 days were set aside, and those considered 
that occurred after the 10 days expired, then, according to Dr. 
Simpson, the proportion of cases was 19-27 and that of deaths 17'24 
times smaller in the inoculated then in the uninoculated. 

Cholera broke out in the Gya gaol, and inoculations were made 
after 6 cases, with 5 deaths, had occurred. During the stay of the 
prisoners in the gaol, there were 209 uninoculated, with 7 cases and 
5 deaths, and 211 inoculated, with 5 cases and 4 deaths. 

In July and August in the same year cholera attacked the East 
Lancashire Regiment. Out of 773 men there were 133 inoculated 
and 640 uninoculated, 

24 



370 



INFECTIVE DISEASES. 



The occurrences of cases and deaths were : — 

In 640 umnocuLited 120 cases (18-75 %), 79 deaths (12-34 %). 

In 133 inoculated 18 cases (13-53 %), 13 deaths (9-77 %). 

These results were, it is said, due to the weakness of the vaccines 
procurable at that period of the work, and to the small doses used. 

There wei-e a great many records kept of the results of inocula- 
tion of coolies on tea estates in different localities. After a summai-y 
of the results Haffkine concludes, in his Report to the Government 
of India, that, in his opinion, the experimental stage was not yet 
in so advanced a condition as to be comjjletely closed ; but that 
the observations made and records collected justified steps being 
taken to give the inoculations a more 
extended trial. 

Cholera Nostras. 

Cholera nostras, English cholera, or 
English dysentery, produces an inflamma- 
tion of the mucous membrane of the 
bowels with croupous exudation. The 
large intestine is commonly affected, and 
the mucous membrane may be covered 
with small superficial ulcers. The disease 
is associated with severe diarrhcea. 

Finkler and Prior obtained a comma- 
bacillus from the evacuations, which they 
believed to be identical with the comma- 
bacillus found by Koch in Asiatic cholera. 
Koch pointed out that there were marked 
differences in the biological character of 
the two micro-organisms. 

Spirillum Finkler-Prior {Comma- 
bacilhisin Cholera nostras). — Curved rods, 
thicker than the comma-bacillus of Koch, 
and spirilla. The colonies on plate- 
cultivations are very much larger than 
those of the comma-bacillus of Koch of the same age. They have a 
very faint yellowish-brovm tinge, a well-defined border, and a distinctly 
granular appearance. They liquefy nutrient gelatine very rapidly, 
so that the first plate of a series is, as a rule, completely liquefied on 
the day following inoculation, and the second plate in two or thi-ee 
days more. In a test-tube cultivation in nutrient gelatine the 
appearances are especially characteristic : the gelatine is very rapidly 



1 
1 



II iii||i 



Fig. 159.— Pure-cultivation 
OF THE Spirillum Finklek- 
Pkiok, in Nutkie.xt Gela- 
tine. In thirty-six hours. 



I 



CHOLEEAIC DIABRHOflA FROM MEAT POISONING. 371 

liquefied along the whole track of the needle, so that the cultivation 
resembles a conical sack, or the finger of a glove turned inside out. 
On a sloping surface of nutrient agar-agar a white moist layer forms 
very quickly. On potato they grow at the ordinary temperature 
of the air, producing a bi-ownish layer and corrosion of the surface 
of the potato. They have been shown to be pathogenic. 

Oholeraio Diarrbcea from Meat Poisoning. 

There are two varieties of choleraic diarrhoea from meat poisoning, 
and both are associated with vomiting, diarrhoea, pain in the abdo- 
men, in severe cases followed by suppression of urine, collapse, and 
death. These conditions are brought about by poisonous foods, and 
include those cases of poisoning by tinned meats, pork pies, hams, 
cheese, sardines, and other articles of food improperly prepared. In 
most cases putrefaction has taken place, owing to the action of 
various bacteria. Associated with their growth we find highly 
poisonous substances, but no bacteria are found in the body in these 
cases. They are all due to chemical poisoning ; but Klein has also 
described cases of poisoning due to the growth of bacteria without 
the presence of putrefaction. The latter were of the nature of an 
infectious disease. In the Welbeck . poisoning cases, described by 
Ballard, the poisonous hams contained a short bacillus, which was 
also found in the kidney and spleen in the fatal cases in man. In 
the Carlisle epidemic, which was due to poisonous pork pies, the pork 
and gravy stock proved fatal to mice, and from the infected mice 
a bacillus was cultivated, which, administered to mice by feeding or 
subcutaneous inoculation, produced enteritis, diarrhosa, and congestion 
of the lungs. 

Gartner cultivated Bacillus enteritidis from the spleen in a 
fatal case of meat poisoning. Gaffky obtained a similar bacillus in 
cases of gastro-enteritis, following the consumption of meat and 
sausages, which had been made of horseflesh. 

Bacillus of Choleraic Diarrhoea from Meat-poisoning 
(Hein). — Bods from 3 to 9 /x, in longth, 1'3 //, wide, rounded at their 
extremities, singly or in chains of two. Spore-formation occurs, 
the spores being 1 in, thick, oval, and situated in the centre or at 
the end of the rod. 

Feeding mice with the bacilli and inoculation produced positive 
results. At the autopsy, pneumonia, peritonitis, pleuritis, enlargement 
of the Uver and spleen, and hcemorrhages were observed, and baciUi 
were present in the blood and exudations of these animals. They 



372 



INFECTIVE DISEASES. 



occurred in the blood and juices, and especially in the glomer'uli of the 
kidneys, of several fatal cases of choleraic diarrhoea. 

Bacillus enteritidis (Gartner). — Short rods in pairs, and short 
chains. They are motile ; spore-formation not observed. Colonies 
are granular, and old colonies at the margin have an appearance of 




Fig. 160. — Teopical Dysentery. Mucous membrane of large intestine some 
months after an acute attack : a,a, representing remains of mucosa ; b,h, inter- 
vening parts corresponding to the muscularis (Hamilton). 



powdered glass. On the surface of gelatine a thick greyish-white 
film develops, which in time becomes wrinkled. In the depth of 
gelatine a white filament forms. The gelatine is not liquefied. On 
agar the film is slightly yellowish. On potato it is similar in colour, 
moist and shining. On blood serum it is very similar, Mice fed 



CHOLERAIC DIARKHCEA IN FOWLS. 373 

with the bacilli die in one or two days. Subcutaneous injection is 
fatal in guinea-pigs and rabbits in from two to five days. Dogs, 
cats, and fowls are immune. 

The bacilli were obtained from a cow suffering from a disease 
associated with diarrhoea, and from the spleen of a man who died 
twelve hours after partaking of the flesh of this animal. 

Dysentery. 

Dysentery is a disease of tropical climates associated with in- 
flammation and ulceration of the large intestine (Fig. 160). At first 
the discharge from the bowel is a whitish or brownish mucus, 
which soon becomes blood-stained; later the evacuations become 
thin and watery, with altered blood clots, fragments of mucous 
membrane, and pieces of false membrane ; and in some cases they 
become purulent. The virus is believed to be in the intestinal 
discharges, which by contaminating water or soil may give rise to 
other cases. 

Micrococci have been found in dysentery, but the micro-organism 
which has received most attention is a protozoon, the Arnxba coli, 
which will be described in another chapter. 

Choleraic Diarrhosa in Fowls. 

Choleraic diarrhoea in fowls, or gastro-enteritis cholerica, is an 
infectious disease of fowls, occurring in Russia during the summer. 
The disease is very like fowl-cholera. The birds are sleepy, and 
suffer from diarrhoea, but the temperature is not raised, as in 
fowl-cholera. After death there is usually an abundance of grepsh 
liquid in the small intestine, which is stained with blood. It was 
investigated by Gamaleia, who found a comma-bacillus, to which he 
gives the name Vibrio Metchnikovi. 

Spirillum of Towl-enteritis (Vibrio Metchnikovi). — Curved 
rods and spirilla ; thicker, shorter, and more curved than Koch's 
commas. They are motile, and possess a single flagellum at one end. 
They stain with the usual dyes. Spore-formation doubtful. In plate- 
cultivations minute white colonies appear in from twelve to sixteen 
hours, and the gelatine is liquefied. The colonies in about three 
days resemble those of both Finkler-Prior's and Koch's comma-bacilli, 
some colonies being more like the one kind, and some like the other. 
In the depth of gelatine the growth is very much like that of Koch's 
comma-bacillus, possessing the characteristic air-bubble appearance. 



374 INFECTIVE DISEAvSES. 

On agar a slightlj- yellowish growth is obtained, resembling that 
of Koch's commas ; on potato a yellowish-brown or chocolate layer 
develops after incubation at the temperature of the blood, very 
similar to cultures from Asiatic cholera. Broth becomes turbid, and 
a wrinkled film forms on the surface ; the addition of sulphuric acid 
gives the indol test. The spirilla grow in milk, and coagulate it ; 
the milk becoming strongly acid, and the casein being precipitated. 
They are pathogenic in chickens, pigeons, and guinea-pigs. Pigeons 
die in about twelve hours after a subcutaneous injection ; and the 
spirilla are found abundantly in blood from the heart. Guinea-pigs 
die from acute septicaemia in about twenty-four hours. The spirilla 
are found in the blood and internal organs. Inoculation of pigeons 
and guinea-pigs with sterilised cultures will produce immunity. 



CHAPTEE XXVIII. 

TUBERCULOSIS. 

Tuberculosis is a communicable disease of man and animals, charac- 
terised by the formation of new growths associated with the presence 
of the tubercle bacillus. Ton Bayle, in 1810, was the first to describe 
little growths like millet seeds, which were considered to be character- 
istic of consumption or phthisis. Laennec, in 1834, attached much 
more importance to the existence of caseous matter and classified 
miliary tubercle, crude tubercle, granular tubercle, and encysted 
tubercle, as varieties of tuberculosis. Virchow would not accept 
all these varieties as tubercular, and only regarded those conditions 
associated with the presence of miliary tubercles as genuinely 
tubercular. Laennec's so-called crude tubercle, for example, was 
simply due to pneumonic caseation. Villemin threw entirely fresh 
light upon this controversy by proving that tuberculosis was a 
communicable disease. Rabbits and guinea-pigs, inoculated with 
tubercular sputum or caseous tubercle, developed miliary tubercle in 
a few weeks. Sanderson confirmed these experiments, and pointed 
out that foreign bodies would produce experimental tuberculosis 
in rabbits. Oohnheim also confirmed the experiments of Villemin, 
and maintained that tuberculosis was a specific inoculable disease, 
and, therefore, everything was tubercular which, on inoculation, 
produced tuberculosis. Koch, in 1882, announced the discovery of 
the tubercle bacillus, and expressed the opinion that without the 
tubercle bacillus there could be no tuberculosis. Tubercle was 
defined as tissue containing the tubercle bacillus, whatever might 
be the clinical manifestations of the case, or the microscopical and 
naked- eye appearances of the diseased parts. 

A tubercle is a small growth about the size of a millet seed. In 
the early stage it is circular, hard, grey in colour, and lustrous ; but 
when it undergoes necrosis and caseation it becomes soft and yellowish. 
In the very early stage it consists of a little collection of round 

375 



376 



INFECTIVE DISEASES. 



cells, in -which it is possible, though often with extreme difficulty, 
to demonstrate the tubercle bacillus. The cells originate in the 
proliferation of endothelial connective tissue and white blood cells. 
Later on, large oval or circular multi- nucleated cells, or giant cells, 
make their appearance. The tubercle bacilli are only occasionally 
found in the interior of human giant cells, whereas in the lower 
animals, in equine and bovine tuberculosis more especially, the bacilli 
are often present in great numbers, and very commonly in the form 
of conspicuous rings, visible under a low power of the microscope. 



1 



mil 11 

^/i-@o>' ■■■■ 




9 h9/ 






i'lu. 101,— TUBKECLE Of THE LuNU IN A VEUY EAHI.Y STAGE, X 4(J0 : a. An alveolar 
wall; 6, blood-corpuscles in capillaries of the same; c, Ijlood-corpuscles 
extravasated into the alveolar cavities ; d, alveolar capillaries filled with 
blood-corpuscles carried forward by the tubercle which is growing into the 
alveolar cavity ; e, large endothelium-like cells, of which the tubercle in this 
stage IS mainly composed ; /, portion of a branch of the pulmonary artery 
injected (Hamilton). 

Whether the absence of blood-vessels or the action of the bacillus is 
the main factor in producing caseation, is an open question. When 
suppuration follows caseation, as commonly happens in tuberculosis 
of the lungs in man, and in experimental tuberculosis in animals, an 
abscess forms. In cattle there is a remarkable tendency to the 
formation of calcareous deposit in the caseous masses. 

The tubercle may not degenerate and die, but live and develop. 



TUBERCULOSIS. 



377 



The giant cells, which are more or less central, have been described 
as sending off processes, which, by dividing and subdividing, and 







Fici. 162. — Peimary Tubercle of Lung two to theee weeks old, x 50 : 
«, Portion of wall of a branch of the pulmonary artery ; ^,Z/, giant cella with 
concentric arrangement of fibrous tissue ; c, centre of tubercle beginning to 
caseate ; ri, small branch of pulmonary artery seen on transverse section ; 
^■, injected capillaries of the alveolar walls (Hamilton'). 



interlacing, form a reticulum, or 
meshwork. Towards the peripliery 
of the tubercle the reticulum may 
become arranged in the form of a 
capsule as the age of the tubercle 
advances, and the reticular giant 
cell becomes eventually converted 
into fibrous tissue. The bacillus has 
disappeared, and the tubercle has 
healed. 

Giant cells cannot be relied upon 
to indicate tuberculosis. They are 
not always present in tiibercu- 
losis, and they are not peculiar to 
tubercle, being found, for example, 
in actinomycosis. The only certain 
indication of tuberculosis is the pre- 
sence of the tubercle bacillus, which 




Fig. 163.— Laege Oval Giant Cell 
fkom tubekcle of lung x 300 '■ 
a, Granular centre; 6, nucleated 
periphery forming a mantle-like 
sheath ; c, processes from the 
same. 



378 INFECTIVE DISEASES. 

can be revealed either by microscopical examination of the suspected 
tissue, or after inoculation in guinea-pigs. 

Bacillus Tuberculosis (Koch). — Eods, 2 to 4 //, and occa- 
sionally 8 fj. long, very thin, and rounded at the ends. They are 
straight or curved, and frequently beaded, and occur singly, in pairs, 
or in bundles ; there are also involution forms and short branched 
threads. Spore-formation is observed in old cultures. They are 
non-motile. Tn the interior of giant cells they are often accompanied 
by grains which exhibit the same colour reaction. 

The bacilli in tissue sections of bovine tuberculosis are shorter 
and less granular than those in human tubercular sputum, but in 
milk they are quite as long, and even longer, and very distinctly 
granular or beaded, and are thus brought much closer, morpho- 
logically, to the bacilli in human sputum. Speaking generally, 
however, the average length of the human bacilli is greater than 
the average length of the bacilli in cow's milk, but the longest of 
the bovine bacilli cannot be distinguished in length from the longest 
human bacilli. There are, however, exceptional cases, for in some 
preparations of pus from human lungs the bacilli are remarkable, 
not only for their thinness, and their uniformly beaded character, 
but more particularly for their extraordinary length. They should 
be compared with other preparations, in which the bacilh, though in 
human sputum, are sometimes much more distinctly rod-shaped, 
much shorter and thicker, with complete absence of any beaded 
appearance. Neither length nor granularity is a characteristic 
sufficient to denote any specific difference between human and bovine 
bacilli. The author has examined minutely the bacUli in tuberculosis 
of other animals, such as the horse, pig, and cat ; and of birds — the 
fowl, guinea-fowl, pheasant, and ostrich. Here, again, minute 
morphological differences can be observed. For example, in many 
cases in fowls the bacilli are conspicuously short and straight. In 
the liver and lungs of an ostrich, packets of short rod-forms are 
found, while in other parts of the same sections the baciUi attain 
a very great length. Many of the long, sinuous forms exhibit a 
peculiar terminal enlargement. There are also short rods with a 
similar appeai'ance, and free ovoid bodies, singly and in groups, which, 
from their connection with the bacilli, and their sharply defined 
outline in the free state, are similar to spores in old cultures. 

Thus, morphological differences are found under different circum- 
stances, and within limits the morphology of the tubercle bacillus 
varies with its environment. 

Koch first succeeded in cultivating the bacillus by employing 



DESCEIPTION OP PLATE XI. 

Bacillus tuberculosis. 

The figures in this plate represent- the bacilli of tuberculosis in 
different animals, examined under the same conditions of amplifica- 
tion and illumination, x 1200. Lamp-light illumination. 

Fig. 1. — Bacilli in pus from the wall of a human tubercular cavity. In 
this specimen the bacilli are shorter than those in tubercular sputum, 
and are very markedly beaded. 

Fig. 2. — Bacilli in pus from a tubercular cavity from another case in man. 
They are present in the preparation in enormous numbers, The proto- 
plasm occupies almost the vfhole of the sheath, and the bacilli are 
strikingly thin and long. 

Fig. 3. — Bacilli in sputum from an advanced case of phthisis, showing 
the ordinary appearance of bacilli in sputum ; some beaded, others 
stained in their entirety ; occurring both singly and in pairs, and 
in groups resembling Chinese letters. 

Fig. 4. — Bacilli in a section from the lung in a case of tuberculosis in man. 
The bacilli in human tuberculosis are found in, and between, the tissue 
cells ; and sometimes, as in equine and bovine tuberculosis, in the 
interior of giant cells, but not so commonly. 
I Fig. 5. — From a cover-glass preparation of the deposit in a sample of milk 
from a tubercular cow. The bacilli were longer than the average 
length of baoiUi in bovine tissue sections, and many were markedly 
beaded. 

Fig. 6. — From a section of the brain in a case of tubercular meningitis in a 
calfi showing a giant cell containing bacilli with the characters usually 
found in sections of bovine tuberculosis. 

Fig. 7. — From a section of the liver of a pig with tubercle bacilli at the 
margin of a caseous nodule. 

Fig. 8. — From a cover-glass preparation of a crushed caseous mesenteric 
gland from a rabbit infected by ingestion of milk from a cow with 
tuberculosis of the udder. 

Fig. 9. — B'rom a section of lung in a case of equine tuberculosis, showing a 
giant cell crowded with tubercle bacilli. 

Fig. 10. — From a section of lung from a case of tuberculosis in the cat, with 
very numerous tubercle bacilli. 

Fig. 11. — From a cover-glass preparation of a crushed caseous nodule from 
the liver of a fowl, with masses of bacilli. These are for the most part 
short, straight rods ; but other forms, varying from long rods to mere 
granules, are also found. 

Fig. 12. — From sections of the liver and of the lung in a case of tubercu- 
losis of a Bhea. Isolated bacilli are found, as well as bacilli packed in 
large cells, colonies of sinuous bacilli, and very long forms v, ith terminal 
spore-like bodies and free oval grains. 

The preparations from which these figures were drawn were all 
stained by the Ziehl-Neelsen method, vdth the exception of the 
first, which was stained by Ehrlich's method. 



^x 



Plate XL. 



_i#^ 



' V 



/ \ 



%i^>'' / 



v_, 



Figl. 



/ 



Pig 5^ 



^ 



Fig 2 






Fig 6 







Pigs 



y 



Pig 7 



^ 



.=^ife_ 






Pi^4 



-i> 



:u/ 



^ 



i^^^'^ 






P:g9. 









^^^ 



Fig 3D. 



,/rv 






/ 




Pi^ll Pig 12 



BACILLUS TUBERCULOSIS 



E, M. Croaks hs-nkftcLt . 



Viru-ent Brooka.Day &Son.,UtK. 



TUBERCULOSIS. 379 

blood serum. Solid blood serum, with or without the addition of 
gelatine, was employed, and the cultures incubated at 37° C. The 
growth takes place very slowly, and only between the temperatures 
of 30° 0. and 41° C. In about eight or ten days the growth appears 
as httle whitish or yellowish scales and grains. 

The bacillus can also be cultivated in a glass capsixle, on blood 
serum, and the appearances of the growth studied under the 
microscope. The scales or pellicles were described by Koch as made 
up of colonies of a perfectly characteristic appearance, which may 
be still further studied by making a cover-glass impression. They 
are then seen to be composed of bacilli, arranged more or less 
with their long axis corresponding with that of the colony itself, 
and with an appreciable interval between the individual bacilli. 
The colonies themselves appear as fine curved lines, the smallest 
being mostly S -shaped. Longer colonies have serpentine twistings 
and bendings, which often 
recall the curves of fancy 
lettering. The ends of the 
lines run to sharp points, 
but the middle of the 
growth is spindle-formed. 
The youngest colonies are 
extremely delicate and 
narrow, but the older 
colonies increase in size, 

are thicker across, and, l^i^- 164. - Bacillus tcbmculosis feom 

TUBEKOULAB SPUTDM, X 2500. From Photo- 
blending with each othei', graphs. 

gradually obliterate the 

characteristic appearances; a lamellated growth results, which 
increases, and gives the appearance to the naked eye of the scale 
or pellicle already described. The blood serum is not liquefied 
unless putrefactive bacteria contaminate the culture. A fresh tube 
can be inoculated with one of the little scales, and a new generation 
started. The scales gradually increase in size, and consist entirely 
of bacilli. In about three to four weeks the cultivation ceases to 
increase, and it is then necessary to inoculate a fresh tube. 

In liquid blood serum a film forms on the surface of the liquid, 
and is easily broken by agitation. In neutralised broth there is 
very little indication of success. When a triturated culture is added 
to the broth, a granular, sandy, whitish deposit collects at the 
bottom of the vessel, with indications of an increase in amount. 
Koch also tried nutrient agar-agar, which did not prove to be at 





380 



INFECTIVE DISEASES. 



BACILLUS TUBERCULOSIS. 




Fig. 1C5. — Puhe-cultivations on Glycerine-agab from Human Tubercular 
Sputum, a, After six months' growth. (Fifth sub-culture.) b and c, After 
ten months' growth. (Fourth sub-cultures. ) 



TUBERCULOSIS. 



381 




all a favourable medium. Some increase took place, but there 

was no continuous growth over the inoculated area. 

Glyceritie Agar-agar. — Nooard and Koux were among those who 

worked at the subject and confirmed Koch's 

observations. Nooard attempted to get 

cultures of avian tuberculosis on blood serum 

to which peptone, salt, and cane sugar had 

been added. The results were more success- 
ful than with ordinary serum. But he 

encountered a difficulty in the rapid drying 

of the surface of the medium, which rendered 

the tubes unfit for use. It occurred to 

Nocard and Roux to obviate this by the 

addition of a hygroscopic agent, and accord- 
ingly they added sterilised glycerine. The 

result, which far exceeded their expectation, 

evidently was not solely attributable to the 

prevention of desiccation. Following up 
their discovery, and 
being anxious to find 
a medium moi-e easily 
prepared than blood 
serum, they added 

6 to 8 per cent, of glycerine to ordinary 
nutrient agar-agar. The bacillus grew so 
abundantly in this mixture that a culture 
in fifteen days equalled in extent a culture 
on blood serum which was several weeks old. 
The bacillus was found to grow abundantly 
in veal broth, to which glycerine had been 
added in the proportion of 5 per cent., the 
bottom of the flask being covered in about 
three weeks with a flooculent deposit, having 
some resemblance to anthrax cultivations 
in liquid media. In beef broth, chicken 
broth, and in Cohn's liquid, cultures were 
obtained after the addition of glycerine. 

Description of Cultivations on Glycerine 
Agar-agar. — The cultivations on the .sloping 
surface of obliquely solidified glycerine agar- 

agai' begin to appear in from four to six days as very minute white 

colonies. These steadily increase in .size, and either look moist and 



Fig. 166. — Puke-culti- 
vation IN Glycerine 
Agae-ah.vb, after ten 
months' growth. 



Fig. 167. —Pure-culti- 
vation IN Glycerine 
Agae-agar, — A Sub- 
culture FROM A PURE- 
cultube in Glycerine- 
milk. In two months. 



382 INFECTIVE DISEASES. 

smooth, or, even at this early stage, appear dry and crinkled. 
According to the number of bacilli inoculated, the colonies will 
either remain isolated or coalesce and form a more or less continuous 
film. If the nutrient agar-agar has only recently been prepared, 
there is usually a quantity of liquid present, and the bacillus forms 
a white coating over the inoculated area and beyond it. The 
appearances are much more characteristic when this medium is, 
comparatively speaking, dry. A semi-transparent membranous 
growth develops, thickens, and assumes a characteristic lichenous 
appearance. Such a culture, examined with a pocket lens, resembles 
a model in wax in miniature of the folds of the gastric mucous 
membrane. In about six weeks to two months the culture has fully 
developed. In old cultures, especially when the individual colonies 
remain isolated, the appearance is very characteristic. Some cul- 
tures in appearance closely resemble cultivations on blood serum. 
The consistency of the growth depends upon the character of the 
soil and the age of the culture. When the medium is moist the 
growth is moist and viscous, but more often it is distinctly tallowy, 
and in old and dry cultures scaly and friable. 

Cultivations in Glycerine Broth. — In a few days minute flakes are 
visible, which steadily increase in size, and subside to the bottom of 
the flask, forming in time a very copious deposit. On shaking the 
flask, this deposit, which is extremely tenacious, rises in stringy 
masses, and gives an appearance which is more or less character- 
istic. If the flask is left undisturbed, a delicate veil-like film forms 
over the surface, which can be readily broken up by gentle agitation, 
forming flakes which gradually sink in the liquid. If undisturbed 
for several weeks this film increases in thickness, is irregularly 
fissured, and has more the appearance of masses of tallow floating 
on the surface.. The growth also may be seen to extend up the side 
of the flask above the liquid. Pasteur or Erlenmeyer flasks can be 
employed for these cultures. Solidified egg-albumin added to the 
glycerine broth seems to increase the amount of growth, which 
clings to the albumin, and waves to and fro in the liquid when the 
flask is gently shaken. The author has confirmed the observation 
of Nocard and Roux, that sub-cultures from glycerine agar-agar, or 
from glycerine broth, will give cultures in ordinary broth without 
glycerine. Ordinary broth with egg-albumin, and without glycerine, 
will also give a good growth when inoculated from previous sub- 
cultures, although the attempt to produce primary cultures in these 
media has hitherto failed. 

Cultivations in Glycerine- Milk, and other Media. — In milk the 



TUBERCULOSIS. 383 

author found there was only a slight increase in the number of bacilli 
inoculated, but milk with glycerine in the proportion of 5 per cent, 
forms a more favourable medium. The author has also cultivated the 
bacillus on sterilised urine and glycerine, and ordinary nutrient 
gelatine with 5 per cent, of glycerine. On potato the growth of 
the bacillus is extremely slow. Beevor succeeded in obtaining 
cultures at the ordinary temperature of the room. 

Examination of Cultivations. — To examine the bacilli in these 
various preparations the author prefers to use Neelsen's method, 
floating the cover-glasses for from five to ten minutes on warm 
carbolised fuchsine, and passing them through dilute sulphuric acid. 
In some cultures the bacilli are shorter and thicker than is commonly 
observed in human sputum, and they are for the most part without 
the beaded appearance. In old cultures on glycerine agar-agar the 
number of granular or beaded bacilli increases, and there are also 
numerous peculiar forms. There are bacUli, sometimes two or 
three times the length of an ordinary bacillus, provided with a 
club-shaped enlargement at one or both extremities, and in rare 
cases with lateral branches. They are no doubt identical with the 
bacilh with swollen extremities and the branched forms observed 
by Nocard and Roux. 

In milk the appearance is very striking, many bacilli attaining 
in old cultures a great length, and all are more uniformly beaded 
than in any other cultivations. Staining preparations by the method 
of Gram in all cases exaggerate this appearance. 

The important part played by the environment is shown by the 
morphological differences observed in artificial cultivation under 
varying conditions, and by the fact that by successive cultivation 
the bacillus can be educated to grow upon a medium which is un- 
suitable for obtaining primary cultures. 

Impression preparations of the growth of the bacillus on the 
surface of glycerine agar-agar in capsules show a tendency to the 
formation of serpentine colonies, composed of bundles of more or 
less parallel bacilli. 

Spore-formation. — In old cultivations true spore-formation can 
readily be observed, both in stained and unstained preparations. In 
the latter case they are recognised in the form of one or two highly 
refractive bodies in individual bacilli. 

Toxic Products of Cultures. — The poisonous substances found in 
cultures, and the composition and use of tuberculin, have already 
been described (p. 43). 

Inoculation Experiments.— A relatively small portion of a culti- 



384 INFECTIVE DISEASES. 

vation inoculated into the subcutaneous tissvie, into the peritoneal 
or pleural cavities, into the anterior chamber of the eye, or directly 
into the blood stream, produces after three or more weeks artificial 
tuberculosis in guinea-pigs and rabbits. Dogs and cats can also be 
infected by experimental inoculation. 

When a trace of tubercular virus is inserted subcutaneously in 
the thigh of a guinea-pig, in about a week or ten days a chain of 
enlarged glands will be easily felt in the vicinity of the seat of 
inoculation. This affords an unfailing test, which can be applied 
when there is difficulty in ascertaining by the microscope the presence 
of the bacilli in the material under examination. It also affords a 
valuable method for testing the effects of antiseptics on tubercular 
virus. The appearances observed at the autopsy are swollen 
lymphatic glands, in the neighbourhood of the inoculation, followed 
by softening and abscess ; enlargement of the spleen and liver, with 
formation of caseous tubercles ; and tubercular deposits in the lungs, 
bronchial glands, and peritoneum. 

After inoculation of the eye, grey tubercles appear on the iris, 
and undergo enlargement and caseation, followed by tuberculosis 
of the eyeball and organs generally. 

The bacilli appear to be the direct cause of tuberculosis, and 
the presence of the bacillus in the sputum of patients is a distinctive 
sign of the existence of this disease. The detection of the bacillus 
has, consequently, become a test which is constantly applied. 

The bacilli are found in all tubercular growths of man, monkeys, 
cattle (jPerlsucht), birds, and many other animals, and in cases of 
artificial tuberculosis, in rabbits, guinea-pigs, cats, etc. In man the 
bacillus can be detected in the tissues, in the sputum, in the blood, 
and in the urine. 

Tuberculosis may also be produced by inhalation and feeding 
experiments. The channels of infection in man are also most 
probably the pulmonary and intestinal mucous membranes. The 
possibility of inoculation of skin wounds is open to doubt. The 
bacilli or their spores are inhaled from the air, or taken in with 
food. Morphologically identical bacilli have also been observed, but 
very sparsely, in sections of lupus. 

Methods of Examining the Tubercle Bacillus. 

Numerous methods have been recommended for examining the 
tubercle bacillus. A few of these will be described, as many are 
only of historical interest, 



TUBERCULOSIS. 385 

The Ziehl-Neelsen method is preferred by the author both for 
sections and cover-glass preparations. 

Koch's original method. — Cover-glaas preparations or sections are laid 
in Koch's solution (No. 23, c) for twenty-four hours, or for one hour if 
the solution is warmed to 40° 0. Rinse in water ; immerse in a watery 
solution of vesuvin for two minutes ; rinse again in water, and examine ; 
or, after rinsing in water, treat with alcohol, clove-oil, and Canada 
balsam. 

Ehrlich's method. — Cover-glass preparations are allowed to float in a 
watch-glass, containing a solution of gentian- violet or fuohsine, added to 
aniline water. A saturated alcoholic solution of the dye is added till 
precipitation commences (10 cc. anihne water, and 10 to 20 drops of the 
colour solution). The cover-glasses are left in the solution for about 
half an hour ; then washed for a few seconds in strong nitric acid (one 
part commercial nitric acid to two of distilled water), and rinsed in 
distilled water. After-stain with vesuvin or methylene-blue, rinse in 
water, dry and preserve in Canada balsam. 

Ehrlich-Koch method. b 

Saturated alcoholic solution of methyl- violet or fuchsine 11 
Aniline water . . . 100 

Absolute alcohol . . .10 

Preparations are left for twelve hours in this solution (colouring of 
the cover-glass preparations can be expedited by warming the solution). 

Treat the preparations with (1 to 3) solution of nitric acid a few 
seconds. 

Wash in alcohol (60 per cent.) for a few minutes (cover-glass prepara- 
tions need only be rinsed a few times). After-stain with diluted solution 
of vesuvin or methylene-blue for a few minutes. 

Wash again in 60 per cent, alcohol, dehydrate in absolute alcohol. 
Clear with cedar-oil, mount in Canada balsam. 

Rindfleisch' s method. — Prepare a solution composed of 

Saturated alcoholic solution of fuchsine . 10 drops 

Aniline water . . . .2 drams. 

Pour it into a watch-glass, and float the cover-glass ; warm the watch- 
glass over a spirit-lamp until steam rises. Remove it from the flame, 
and set it aside for five minutes. Take out the cover-glass, and transfer 
it for a few seconds to acidulated alcohol (two drops of nitric acid in a 
watch-glass full of alcohol). Wash in distilled water, dry, and preserve 
in balsam. After-stain, if necessary, with Bismarck-brown, or methylene- 
blue. 

Oibbcs' mertod— Cover-glass preparations are placed in Gibbes' double- 
staining solution which has been warmed in a test-tube, and, as soon as 
steam rises, poured into a watch-glass. They are allowed to remain for 
five minutes, and then are washed in methylated spirit till no more colour 
comes away, dried in the air or over a spirit-lamp, and mounted i 
Canada balsam, If the solution is used without warmiug, the cover-glassei 

2§ 



386 INFECTIVE DISEASES. 

must be left in it for an hour. Sections are treated on the same 
principles, but must be left in the solution for several hours. The 
crumpling of the sections by the action of nitric acid is avoided. 

Baumgarten' s method. — Cover-glass preparations of sputum are made 
as already described, and immersed in a very dilute solution of potash 
(1 to 2 drops of a 33 per cent, solution of potash in a watch-glass of dis- 
tilled water). The cover-glass is pressed down on a slide, and examined 
with a high power. The bacilli can be thus examined in the unstained 
condition, and to avoid any mistake from confusion with other species, 
the cover-glass can be removed, dried, passed through the flame, and 
stained with a drop of an aqueous solution of fuchsine, or gentian- violet. 
The putrefactive bacteria are stained, but the tubercle bacilli remain 
absolutely colourless. 

Baumgarten's new method. — A solution is prepared as follows : Drop 
4 to 5 drops of concentrated alcoholic methyl-violet solution into a small 
watch-glass full of water, (a) Stain the sections in this solution, wash 
them in water, and decolorise in absolute alcohol (five to ten minutes) ; 
or, before treating with alcohol, immerse the sections for five minutes in 
a half-saturated solution of carbonate of potash. Pass through clove-oil, 
and mount in a mixture of Canada balsam, free from chloroform, and 
clove-oil (equal parts). The object of this process is to differentiate the 
tubercle bacilli from chance bacteria, inasmuch as the tubercle bacilli 
are gradually decolorised by the clove-oil. (&) Sections stained in the 
above solution are placed for five minutes in alcohol, and then in a 
concentrated solution of Bismarck-brown in 1 per cent, solution of acetic 
acid. The after-treatment may be conducted as already described. 

Ziehl-Neelsen method. — Cover-glass preparations may be quickly stained 
in Neelsen's solution warmed in a watch-glass till steam rises. Sections 
are left for from five to ten minutes in the solution, and then washed in a 
watery solution of sulphuric acid (25 per cent.); rinsed in distilled water, 
and immersed in methylene-blue solution. After two or three minutes 
they are passed through alcohol and oil of cloves, and mounted in Canada 
balsam. 

FrdnkeVs method. — Sputum preparations are rapidly double-stained 
by the following method : Prepare a solution by adding concentrated 
alcoholic methyl-violet or fuchsine solution, drop by drop, till opalescence 
arises, to 5 ccm. of aniline-water heated to 100° C. Float the prepared 
cover-glasses two minutes in the warmed solution. The process of after- 
staining and decolorisation is effected by placing the preparation for one 
to two minutes in one of the following solutions : for fuohsine-stained 
preparations, a saturated solution of methylene-blue in a mixture of 
Alcohol . . 60 

Distilled water . . 30 

Mtric acid . . . 20 

which is filtered before use ; for preparations stained in methyl-violet, a 
saturated solution of vesuvin may be used in 

Alcohol . . 70 

Nitric acid . ... . . 30 



TUBERCULOSIS. 387 

Ehrlicli's Method and Eosin. — The author has found that after sections 
have been stained with methyl-violet and Bismarck-brown by Ehrlich's 
method, as described by Koch, they may with advantage be immersed in 
a weak alcoholic solution of eosin, then rinsed in clean absolute alcohol, 
clarified with clove-oil, and mounted in Canada balsam. The giant cells 
are then stained pink, while their nuclei are brown, and the bacilli blue. 

Tuberculosis' in Man. 

The disease manifests itself in various forms in man, and most 
frequently in the lungs, producing phthisis or consumption. The 
sputum contains the bacilli in large numbers, and is extremely 
virulent. Scrofula and lupus are forms of tuberculosis ; they are 



Fig. 168. — Section thkough a Lupds nodule oi' the Nose. 

probably produced by an attenuated variety of the tubercle bacillus. 
Lupus can be distinguished from tuberculosis of the skin ; and 
scrofulous lymphatic glands are distinguished from tubercular glands 
by the tendency of the latter to produce generalised tuberculosis. 
This difference in the intensity of the virus in the two cases, 
Lingard illustrated by the effect upon inoculated guinea-pigs. 

Cavities in the lungs are often thickly lined with bacilli. They 
are present in great numbers in the caseous matter, though in 
equine and bovine tuberculosis this is not the case. 

Whether the disease in man is contagious is an open question, 
though numerous cases of supposed communicatioa between husband 
and wife, brothers and sisters, have been reported, and Rausome 



388 



INFECTn'E DISEASES. 



showed that tubercle bacilli were present in the breath in phthisis. 
On the other hand, the experience in consumption hospitals does not 




Fig. ICH.— T-uberculae Uloeeatios iof Muoosa of Human Ileum. 
Between the ulcers there are tubercular lymph-follicles (Hamilton). 

support this view, there being no evidence of the communication of 
the disease to nurses and hospital attendants, 



TDBERCDLOSIS. 



389 



Tuberculosis of Cattle. 

In cattle the disease may occur as the result of inhaling bacilli, 
or of ingestion with food. It is very frequently found in the lungs ; 
and calves may be infected by milk from cows with tubercular 
udders. Calves may also suffer from congenitiil tuberculosis, the 
bacilU having been transmitted from the mother during gestation. 

Breeding in-and-in, over-production of milk, and confinement with 
insanitary surroundings, predispose to tuberculosis. The disease is 
known in Germany as " Perlsucht " ; and in this country the lesions 
on the pleura are known as 
" grapes," and the animals 
themselves are commonly 
called " wasters." 

The disease may also exist 
in the lungs or in other 
organs, in a limited form, 
without any indication of 
ill health. In such oases 
the disease can be detected 
by injection of tuberculin, a 
marked rise of temperature 
occurring in tubercular 
animals. 

In advanced cases, the 
symptoms commonly observed 
•ire cough, difficulty in breath- 
ing, staring coat, wasting, and 
diarrhoea ; and if the udder is infected, nodules in the gland, and thin 
bluish milk. In the lungs, after slaughter, a few small cheesy tubercles 
may be found in animals apparently in perfect health and in prime 
condition for the market. In advanced cases, the lungs on section 
show large yellow masses, containing calcified matter, and the 
bronchi may be full of yellowish pasty contents. The disease will be 
found to involve the bronchial glands. The serous membrane may 
be covered with little warts or grape-like masses. The lymphatic 
glands may be enlarged to an enormous size. Tubercular ulceration 
of the intestine is sometimes found, but not commonly. In tubercular 
disease in the udder, a painless swelling is found which may affect 
one or more quai'ters of the gland. 

Transmission of Tuberculosis from Man to Cattle.— It is 




Fig. 170. —Section of Lupus of the Skin, 
X 700. Giant cell containing a tubercle 
bacillus (Flugge). 



590 



INFECTIVE DISEASES. 



for obvious reasons impossible to ascertain by experiment whether 
tuberculosis can be transmitted from cows to man by milk or other- 
wise- but some light may be thrown upon this important, question 
by ascertaining the result of inoculating bovines with human tuber- 
culosis If calves can be infected with tuberculosis from a human 
source by inoculation or ingestion experiments, and especially if 
the effect of administering human and bovine tubercle to calves, 
by these means is found to be the same, such experiments will 
not only serve to dispel any doubt there may be as to the identity 
of the two affections, but they will strengthen the hands of those 




Fig. 171.— Tuberculosis of Pleuea ; ".Gkape-disease.' 



who insist upon the necessity of more thorough inspection of dairy 
cows, and of power to deal with tubercular animals. 

Inoculation of a Calf tvith Human Tuhercular Sputum. — The 
author obtained sputum containing numerous bacilli from an 
advanced case of phthisis. The sputum was shaken up with sterihsed 
salt solution and injected into the peritoneal cavity. A few weeks 
afterwards the calf showed signs of illness. The animal looked dull, 
did not feed well, had a slight cough, and showed less inclination to 
move about than usual. These symptoms gradually increased, and 
death occurred forty-two days after inoculation. Extensive lesions 



TUBERCULOSIS. 391 

were discovered at the post-mortem examination. The mesentery 
was adherent to the abdominal wall, at the seat of the inoculation, 
and to the rumen ; the liver was adherent to the diaphragm. There 
was extensive tubercular deposit at the seat of inoculation, and 
an abscess the size of a walnut. Extending over the mesentery 
from this point there were hundreds of wartlike, fleshy, new growths, 
some quite irregular in form, others spherical or button-shaped. 
There were similar deposits on the under surface of the liver, on the 
spleen, in the gastro-splenic omentum, and on the peritoneal surface 
of the diaphragm. The spleen was adherent to the rumen, and 
on dissecting away the adhesions another abscess was opened. The 
lungs were congested and the pleurse thickened. On microscopical 
examination of sections extremely minute tubercles were found to 
be disseminated throughout the whole of the substance of the lungs 
. and liver, and tubercle bacilli were found in these and in the 
peritoneal deposits. The abscesses contained Streptococcus pyogenes. 
The calf died of pysemia, but sufficient time had elapsed for marked 
local infection leading to generalised miliary tuberculosis. 

Tuberculosis in Relation to the Public Milk Supply. 

There is not the slightest doubt that when the udder is involved 
the mUk is highly virulent to the lower animals, and presumably 
is, therefore, dangerous to man. The virulence of the milk was 
first insisted upon by Klencke in 1846, and confirmed by Gerlach in 
1869, and later, by others. 

This subject was again brought forward with the discovery of 
the tubercle bacillus, and the demonstration of its existence in the 
milk in certain cases of bovine tuberculosis. Koch pointed out that 
the milk only contained bacilU, and was only infective, when the 
udder itself was tubercular. By this he explained the contradictory 
results obt.ained by various experimenters with milk from cows un- 
doubtedly suffering from " Perlsucht." Koch considered that positive 
effects were obtained with milk when it happened to contain tubercle 
bacilli, and negative with milk from which they were absent. Bang 
in a number of cases verified the presence of tubercle bacilh in milk, 
and, owing to the contradictory results of previous investigations, 
repeated the ingestion experiments. The milk was found to be 
virulent both to pigs and rabbits. 

In this country Woodhead and M'Fadyean tested milk for 
tubercle bacilli. They examined six hundred cows in the Edinburgh 
dairies, and found thirty-seven suffering from mammitis, but in only 



392 INFECTIVE DISEASES. 

six were they able to demonstrate the presence of tubercle bacilli in 
the milk, and then only in small numbers. 

Hirschberger found in twenty cases of tuberculosis in cattle 
that the milk of eleven was virulent to guinea-pigs. Three cows out 
of nine in which the disease was restricted to the lungs gave infected 
milk. On the other hand, Nocard inoculated milk from eleven 
tuberculous cows, of which only one had diseased udder, and only 
this one gave infective milk. Bang injected rabbits with milk from 
twenty-one cases of tuberculosis, with the udders apparently normal, 
and the milk was virulent in two. 

The author had two cases of udder tuberculosis under observation, 
and as no experiments had at the time been made in this country 
with milk known to contain tubercle bacilli, it was decided to study 
the effect on rabbits, and test the results obtained by Bang. These 
cases were both interesting and instructive, and may be referred to 
in detail. 

One was a case of advanced general tubercnlosis. There was extreme 
emaciation, general apathy, and a peculiar dull expression of countenance. 
The skin was dry and harsh, the coat staring, and there was loss of hair 
in patches about the face and neck. There was dulness on percussion 
over a large area of the thorax, and the respirations were increased in 
rapidity. There was also occasional cough and some diarrhoea. But the 
most interesting condition was observed on examination of the udder. 
The gland was swollen, especially posteriorly, and distinct induration 
could be felt on examination. The deposit appeared to be more or less 
limited to the posterior quarters. The cow evinced no pain during the 
examination of the udder, not even on the application of firm pressure. 

The author took samples in test-tubes of the milk from all four teats ; 
when freshly drawn, it differed noticeably from the normal secretion. It 
was a thin, watery, turbid fluid with whitish flakes in suspension, but it 
was not gelatinous or muco-purulent in character, and was free from any 
markedly yellow colour. After being set aside in the laboratory for 
some hours it separated into a layer of cream and a turbid liquid of a 
yellowish tint, while at the bottom of the test-tube there was a whitish 
flocculent deposit, especially in the samples from the posterior quarters. 

There were tubercle bacilli both in the cream and in the deposit. In 
the cream they were only present in small numbers, and were detected, 
therefore, only after careful search. But in the deposit they were readily 
found, as in a cover-glass preparation there were sometimes four or five 
in the field of the microscope. 

The method adopted for the examination of this deposit was as 
follows : The whole of the liquid in the test-tube was carefully poured 
off, and a trace of the sediment spread out on a cover-glass. This was 
allowed to dry, and passed through the flame, and stained in hot Ziehl- 
Neelsen solution in the usual manner. 



TDBERCULOSIS. 



393 



The other cow was also a case of general tuherculosis, and presented 
somewhat similar lesions of the udder. The induration of the gland was 
readily detected, and examination of the milk showed, as in the previous 
case, the presence of tubercle bacilli. 

It will be observed that in neither of these cases was the disease 
limited to the udder ; in both the implication of the gland was part of 
general tuberculosis. 




Fig. 172.— Tubercul.\k Ulcebation of the Intestine of a Cow. 

The first cow was killed, and the following lesions were found at the 
post-mortem examination. 

Thorax.—Ihe lungs and bronchial glands were extensively invaded 
with tubercular deposit. The glands were greatly enlarged and densely 
fibrous, in many cases with central, stone-like masses, grating on section 
against the edge of the knife. In the lung there was every stage, from 
the early deposit to purulent cavities, cheesy masses, and calcified cUhris. 

Abdomen.— There were a few caseous nodules in the liver, but none 
in the spleen. The mesenteric glands formed an almost continuous chain 



394 INFECTIVE DISEASES. 

of large tumours, mostly with central cretification. Tubercular deposit 
in the intestines could be recognised from the outside, and on laying 
them open the mucous membrane was found to be studded with tuber- 
cular ulcers. These ulcers were most numerous in the large intestine, 
and varied in size from a sixpence to a florin. Some were circular, others 
slightly irregular in form, and others again distinctly oval. In the latter 
case they were generally situated with their long diameter transversely. 
The base of the ulcer involved the muscular coat, and was irregularly 
radiated. The margin was broad, and elevated above the general surface, 
producing a ring-like appearance. 

Mammary Gland.— The udder was infiltrated throughout with tuber- 
cular new growth, but the invasion was most marked in the posterior 
quarters. There was apparently very little tendency to caseation. 

Microscopical Examination of the Udder.— la order to study the histo- 
logical characters of the gland, and the distribution of the bacilli, sections 
were stained with logwood and rubin, and others again with fuchsine 
and methylene- blue. The tubercular new growth consisted of the usual 
histological elements, round cells, epithelioid cells, and giant cells. 
Healthy lobules here and there were sharply marked ofE from those in 
which the growth was compressing and obliterating the alveoli in its 
progress. Bacilli were present in the giant cells, and also distributed in 
vast numbers throughout the tubercular tissue generally. Bacilli were 
found in epithelioid cells close to the alveolus, and also between the cells 
lining the alveoli. In parts also the new growth had involved the milk 
ducts, and therefore it was easy to account for the presence of the bacilli 
in the milk. 

The bacilli were found in considerable numbers also in sections of the 
intestinal ulcers. 



Experimental Infection of Rabbits. 

Ingestion. — A rabbit received the contents of a test-tube which 
had been filled with milk from one of the posterior teats, mixed with 
a small quantity of bran. In four weeks there was commencing 
emaciation ; later, diarrhoea set in, and death occurred exactly fifty- 
eight days after administration of the milk. At the post-mortem 
examination the mesenteric glands were found to be much enlarged 
and caseous. A cover-glass preparation from a crushed gland 
revealed numerous tubercle bacilli. On opening the intestines there 
was a patch of ulceration, showing the point of access of the bacilli. 
The intestinal ulceration was a reproduction, to a certain extent, of 
the condition in tlie cow which had been the source of the virus. 

Subcutaneous Injection. — A second rabbit was injected under the 
skin of the back by means of a capillary pipette with about ten 
drops of milk, including some of the deposit from the bottom of the 
test-tube. The sample of milk had in this case also been taken from 



DESCRIPTION OP PLATE XII. 
Tuberctilar Mammitis. 

Fig. 1. — From a section of the udder of a milch cow. The tubercular deposit 
is seen to invade the lobules of the gland. Lobules comparatively healthy 
are marked off, more or less sharply, from the diseased ones in which the 
new growth in its progress compresses and obliterates the alveoli. Stained 
by the Ziehl-Neelsen method and with metbylene-blue. x 50. 

Fig. 2. — Part of the same preparation. On the right of the section part of a 
healthy lobule is seen. On the left a lobule is invaded by tubercular new 
growth composed of round cells, epithelioid cells and typical giant cells. 
Tubercle bacilli can be seen both singly and collected in groups. They 
are found in and between the cells, and in the interior of giant cells. 
Bacilli may be seeii between the cells Uning an alveolus and projecting 
into its lumen, x 800. 






:>--l-, 



f ■■•■■41' |*^S^^^^viS- 

vP-#fS'^'-y' :-■-■;-: 

;i ... fij:^-^-: ■';■•:. -';^^>v ' 



J '-:;^ 



^'3( VI. Ji n 





«iS> 


■<V"Vf .;-,;■ 


'■^\---(.-. 










i? \ 


,i.i^- 


!;j^;^^' 






'■■■' 


S-*-?^ . 








•■--'■'•C 


|«- 


^-\:'-\ 


*."'-. i"";". 



v: ■(.. 







■■.■"■*-7".: . ,■.' 






i^^-^ 




'"■'"" ''/'■' 




^ii^;^«*ifrS4fe 


^^■;£>.^;- 


iaC J 




■■-^■■s^ 






>"'■>'■■'" '-■■.'^^1^':'- 


,,i4::>/"^i; 














It-' ~^^S^ 



©imrv^o^'^f' ^ 






^S^l^t?^ r^^'-aa 



a 



I 









O 






Q.lo, 




nTKKRCULAU 



#■ ^ 

Fife 2. 



TUBERCULOSIS. 



395 



one of the posterior teats. The rabbit wa.s placed in a separate 
hutch, and death from general tuberculosis occurred ninety- two days 
after inoculation. 

The diaphragm and mesentery were studded with tubercles the 
size of a pin's head. The kidneys superficially showed whitish 
rounded nodules projecting above the surface. These were found 
on section to be continuous, with wedge-shaped deposits in the sub- 
stance of the kidney. The lungs presented a very striking appear- 
ance, being, in short, a mass of tubercular deposit; and the bronchial 
and tracheal glands were similarly affected. In sections of the 
kidney and lung the bacilli were present, but they were distributed 
irregularly ; in one part of a section it was difficult to detect a 
single bacillus, in other parts they were present in large numbers. 

The milk from the two cows, 
previously to their coming under 
observation, had been mixed with 
the general supply of a dairy. There 
is indeed ample evidence that, both 
in this and in other countries, the 
milk of tuberculous animals finds its 
way into the market. The question 
which naturally arises is the possi- 
bility of any manifestation of tuber- 
culosis in man, arising from the 
consumption of unboiled milk con- 
taining tubercle bacilli. We must 
admit that there is no direct 
evidence of the transmission of 
tuberculosis by milk from cow to 
man ; but this may arise from the difficulty in tracing such a 
source of infection, owing to the long time which elapses before 
symptoms manifest themselves in man. Yet, if milk be a source of 
infection, we should naturally expect that primary tuberculosis of the 
intestine would be by no means an uncommon manifestation of the 
disease; and this in the adult is not in accordance with clinical 
experience. Such an argument would tend to contra -indicate 
danger to adults ; but, on the other hand, the possible danger to 
children has been rightly insisted upon by the earliest writers on 
this subject. Woodhead has recently stated that, from his experi- 
ence in two large hospitals, he has been much struck by the fact 
that, in children who had died from other diseases during the course 
of tubercular disease of the abdominal gland.-i, there was frequently 




Fig. 173.— Tubekculab XJlceka- 
tion of the intestine of a 
Rabbit. 



396 



INFECTIVE DISEASES. 



not any trace of tubercular disease in other parts ; thus pointing to 
the intestine as the channel by which the bacillus made its way into 
the body. Woodhead also remarks that in a large number of cases 




Pig.' 174. — Tubeeoulosis oi' the Lungs. 

From a photograph of the lungB of a rabbit which had been injected snb- 
cutaneously with about ten drops of milk, including in suspension a small 
quantity of the deposit at the bottom of a sample of milk from a cow with 
tuberculosis of the udder. Death occurred from general tuberculosis ninety-two 
days afterwards. The appearance of the lungs was very striking. They were 
almost completely composed of tubercular deposit. The bronchial glands, 
as well as the tracheal, of which one is seen in the photograph, were also 
enlarged and caseous. There were tubercular deposits in the kidneys and 
other organs, and also at the seat of inoculation. 



of general tuberculosis, where the possibility of infection by the 
pulmonary passages was evidently excluded, the tubercular process 



TUBERCULOSIS. 397 

appeared to have invaded the body by the intestinal canal. These 
facts, taken in connection with the occasional existence of tubercle 
bacilli in milk, went far to prove, in his opinion, that milk was a 
source of tubercular infection, especially to young children. 

From his own experiments and observations the author has 
drawn the following conclusions : — 

1. Cows with tuberculosis of the udder are to be found in dairies 

in this country. 

2. The milk of thftse cows is, as a rule, mixed with the general 

supply. 

3. The milk in cases of udder tuberculosis contains tubercle 

bacilli. 

4. Rabbits inoculated with, or fed upon, milk containing tubercle 

bacilli contract tuberculosis. 

5. Direct evidence of transmission of tubercidosis by mUk to man is 

wanting, but from the effect of such mUk on the lower animals 
it is reasonable to conclude, in the present state of our know- 
ledge, that there may be danger in using the milk of cows 
with tubercular udders, and therefore strict inspection of 
dairies should be enforced ; and boiling of milk before 
use will, as a rule, be a wise, if not absolutely a necessary 
precaution. 

Bollinger has shown that the virulence of cow's milk is reduced 
by dilution with water in the proportion of 1 in 40 and even of 
1 in 100, and that therefore there would be much less danger 
in consuming tubercular milk which had been mixed with the 
milk of healthy cows, than there would be in taking it direct 
from the infected cow. This is a matter of scientific interest ; but 
it would be no justification for a dairyman to mix the milk of a 
tubercidar cow with milk of cows known to be healthy. The milk 
of cows suffering from tuberculosis should undoubtedly be rejected. 



Tuberculosis and the Public Meat Supply. 

The question of the advisability of allowing the flesh of tuber- 
cular animals to be sold for food, especially when the disease exists 
in a very small degree, is a vexed one. Numerous experiments 
have been made upon the infectivity of the flesh of tubercular 
animals. Kastner inoculated the juice expressed from the flesh of 
ti;it§rcular cows. Sixteen guinea-pigs were unaffected after injection 



398 INFECTIVE DISEASES. 

of 1 to 2 cc. into th.e peritoneal cavity. Nocard injected ten to 
twenty drops of the muscle juice of the hearts of tubercular cattle, in 
which the disease was well marked, and none of the guinea-pigs were 
infected. With juice of the muscles of the thigh derived from ten 
tubercular cows Nocard inoculated forty guinea-pigs, and one only 
showed signs of tubercle. Nocard concluded that if there was any 
danger in the flesh of tuberculous animals, it was the exception and 
not the rule. On the other hand, Ohauveau and Arloing produced 
tuberculosis in two guinea-pigs out of ten inoculated with muscle 
juice from a tubercular steer. 

In 1890 a Royal Commission was appointed to investigate this 
subject, and the report was issued in 1895. Martin, on behalf of 
the Commission, tested the flesh of twenty-one tubercular cows. In 
two cases only was evidence obtained of the presence of the bacillus 
by inoculation of guinea-pigs. The flesh of eight cows affected with 
mild tuberculosis produced tubercle in one instance by inoculation, 
but the ingestion experiments were negative. The flesh of five cows 
severely affected with tubercle gave the disease in four cases, either 
by feeding or inoculation, but only one gave the disease both ways. 
Martin thought that some of the results were due to the butcher 
infecting the meat in the process of dressing the carcase, either by 
his hands or knives. Woodhead made a series of experiments to test 
the effects of roasting and boiling on the tubercular virus in meat. 
It was found that in boiling and roasting experiments, as ordinarily 
carried out in the kitchen, the temperature, however high it may be 
on the surface, seldom reaches 60° C. in the centre, except in the 
case of joints less than about six pounds in weight. Boiling and 
roasting were found insufiicient to destroy tubercular virus enveloped 
in rolls of meat. 

The following were among the conclusions of the Commissioners : 

We have obtained ample evidence that food derived from tuberculous 
animals can produce tuberculosis in healthy animals. The proportion of 
animals contracting tuberculosis after experimental use of such food is 
difEerent in one and another class of animals ; both carnivora and 
herbivora are susceptible, and the proportion is high in pigs. In the 
absence of direct experiments on human subjects, we infer that man also 
can acquire tuberculosis, by feeding upon materials derived from tuber- 
culous food-animals. 

The actual amount of tuberculous disease among certain classes of 
food-animals is so large as to aflrord to man frequent occasions for 
contracting tuberculous disease through his food. As to the proportion 
of tuberculosis acquired by man, through his food or through other means, 
we can form no definite opinion, but we think it probable that an 



TUBERCULOSIS. 399 

appreciable part of the tuberculosis that affects man is obtained through 
his food. 

The circumstances and conditions with regard to the tuberculosis in 
the food-animal which lead to the production of tuberculosis in man are, 
ultimately, the presence of active tuberculous matter in the food taken 
from the animal, and consumed by the man in a raw or insufficiently 
cooked state. 

Tuberculous disease is observed most frequently in cattle and in 
swine. It is found far more frequently in cattle (full grown) than in 
calves ; and with much greater frequency in cows kept in town cow- 
houses than in cattle bred for the express purpose of slaughter. Tuber- 
culous matter is but seldom found in the meat substance of the carcase ; it 
is principally found in the organs, membranes, and glands. There is 
reason to believe that tuberculous matter, when present in meat sold to 
the public, is more commonly due to the contamination of the surface 
of the meat with material derived from other diseased parts, than to 
disease of the meat itself. The same matter is found in the milk of cows 
when the udder has become invaded by tuberculous disease, and seldom 
or never when the udder is not diseased. Tuberculous matter in milk is 
exceptionally active in its operation upon animals fed either with the 
milk or with the dairy produce derived from it. No doubt the largest 
part of the tuberculosis which man obtains through his food is by means 
of milk containing tuberculous matter. 

Provided every part that is the seat of tuberculous matter can be 
avoided and destroyed, and provided care be taken to save from contami- 
nation by such matter the actual meat substance of a tuberculous 
animal, a great deal of meat from animals affected by tuberculosis may 
be eaten without risk to the consumer. 

Ordinary processes of cooking applied to meat which has got con- 
taminated on its surface are probably sufficient to destroy the harmful 
quaUty. They would not avail to render wholesome any piece of meat 
that contained tuberculous matter in its deeper, parts. In regard to milk 
we are aware of the preference by English people for drinking cow's milk 
raw — a practice attended by danger, on account of possible contamination 
by pathogenic organisms. The boiling of milk, even for a moment, would 
probably be sufficient to remove the very dangerous quality of tuber- 
culous milk. 

Tuberculosis in Equines. 

Tuberculosis is not very common in the horse, but when it does 
occur, it is frequently mistaken for glanders. There may be miliary 
tuberculosis in the lungs, or nodules disseminated throughout the 
lungs, liver, spleen, and bones. In a number of cases investigated 
by Nocard, the disease commenced in the abdominal organs, and 
the affection of the lungs appeared to be secondary. The author 
has examined several cases of equine tuberculosis. In some cases 



400 INFECTIVE DISEASES. 

the lungs were affected with the disease in a miliary form. The 
bacilU could not be distinguished from bacilli in sections of the bovine 
disease. Giant cells were extraordinarily numerous, and in many 
cases were densely packed with bacilli, so that they could be recog- 
nised en masse under a low power. The bacilli were also distributed 
in the tissue generally, but were much more numerous in the giant 
cells. 

Tuberculosis in Doas. 

Peters described a case of tuberculosis in a pet dog, from eating 
spiitum from a tubercidar patient. This is said to be a not 
uncommon cause of canine tuberculosis. 

Tubebculosis in Oats. 

Nocard reported a case of tuberculosis in a cat from eating tuber- 
cular sputum. The abdominal organs were diseased. Bollinger 
has described two cases of miliary tuberculosis. MTadyean also has 
described a case. The bacilU are very plentiful in the lung. A 
minute examination of the individual micro-organisms by the author 
did not reveal any distinctive character. 

TUBEECULOSIS IN SwiNB. 

The author examined the tubercular liver of a pig. The pig 
was about six months old, and after suffering from cough and 
emaciation, died. 

The liver had caseous nodules scattered throughout its substance, 
some the size of a pea, and others larger. Tubercle bacilli without 
distinctive characters were found on examination of sections ; but 
it was in some parts of a preparation difficult to detect any bacilli, 
and in other parts there were not more than five or six in the 
field of the microscope. Tuberculosis in swine is said to be very 
rare in America. 

Tuberculosis in Birds. 

Hens, guinea-fowls, turkeys, pheasants, and partridges, are sub- 
ject to tuberculosis, and ostriches and other birds kept in confinement 
may contract the disease. 

Tuberculosis in fowls appears to be introduced principally with 
the food, the disease occurring commonly in the intestines and 
liver. 



DESCKIPTION OF PLATE XIH. 
Tuberculosis in Swine. 

Section of liver of a pig with scattered tubercular nodules. Miorosoopioa 
sections of the liver showed tubercle bacilli in very small numbers. 






O 

w 

> 



TUBERCULOSIS. 401 

The author has examined several cases of so-called spontaneous 
tuberculosis in fowls. Sections of the liver were in one case remark- 
able on account of the extraordinary invasion of the caseous deposits 
with bacilli. Cover-glass preparations had been made from the 
liver in the following way for diagnostic purposes : A tubercle was 
readily picked out on the point of a scalpel and crushed between two 
slides, and the cover-glass preparations stained with the Ziehl- 
Neelsen solution. The bacilli are for the most part very small. A 
few attain a considerable length, but the majority are in the form 
of small, straight rods, with many sizes intervening between, these 
rods and isolated granules. 

In July 1888 the author received from Mr. Bland Sutton the 
liver and lungs of a Rhea, which had died in the Zoological Gardens. 
The lung was infiltrated with caseous deposits, and there were 
scattered caseous nodules in the liver varying in size from a pea 
to a marble. The naked-eye appearance of a section of the liver 
through these nodules, at once recalled to mind the naked-eye 
appearance of the deposits in the pig's liver already described. But 
whereas in microscopical preparations of the pig's liver, bacUli were 
very scantily present, the sections of the lung and liver of the Rhea 
•contained bacilK in such extraordinary numbers that, under a power 
of fifty diameters, the collections of bacilli could be recognised as red 
granular masses. These red masses under a high power were re- 
solved into dense colonies of bacilli. In their number and their 
distribution in the tissues, in their varying size, and in the extra- 
ordinary length of the longest forms, they presented very interesting 
points for observation. From the naked- eye appearance of the 
disease and the general microscopical characters, as well as the 
presence of bacilli agreeing in their staining reactions with the 
classical tubercle bacilK, the author had no hesitation in pronouncing 
the disease to be avian tuberculosis. 

Klein, who had examined a similar case, alluded to it in a 
description of leprosy; but this disease is unknown in the lower 
animals, and all attempts to infect them from man have been 
almost, if not entirely, negative. 

The bacUli in the Rhea are principally collected in the caseous 
parts, but they are also found in the tissue generally, and often 
collected in large cells. In size they vary to, a marked extent. In 
the cells they often form compact masses of short bacilli, but in 
other parts, both in collections and singly, they attain a greater 
length than is observed in any other form of tuberculosis. Some of 
the bacilli present a vsry interesting appearance. They are provided 

26 



402 INFECTIVE DISEASES. 

terminally with a sharply defined ovoid body. There are also 
collections of short bacilli, many with these spore-like appearances. 
The author has also seen free ovoid forms, sometimes singly, some- 
times in groups. From their connection with the bacilU and their 
sharply defined outline they are very suggestive of spores. 

Johne examined the livers of a number of fowls accidentally 
infected by phthisical sputum. Nocard reported an outbreak in 
a poultiy-yard where the man in charge had consumption. He 
also found the disease amongst fowls fed with the infected organs 
of tubercular cattle. Subcutaneous inoculation, and feeding of fowls 
with sputum or bovine virus, will produce the disease. 

Experimental inoculation of tubercular virus from different 
sources affords an illustration of the different pathogenic effects 
obtained by varieties of the same species of bacillus. The bacillus 
of fowl-tuberculosis is a distinct variety. A very small proportion 
of guinea-pigs, inoculated in the peritoneal cavity with fowl-tubercle, 
succumb to the disease, though so susceptible to the effects of human 
or bovine virus. Maffucci maintains that guinea-pigs have an 
immiinity, and that rabbits rarely develop a generalised tuberculosis. 
Cultures are not identical in appearance with those obtained from 
man, and on microscopical examination show many long, thick, and 
branched forms, which are only rarely found in cultures from a 
human source. 

Stamping-out System. — In 1888 a Departmental Committee 
was appointed to inquire into pleuro-pneumonia and tubercitlosis, 
and they considered that legislation ought to be directed not only 
to the protection of cattle from tuberculosis, but also to prevent 
the possibility of the disease being communicated to man. 

The following extracts are from the recommendations of the 
Committee, which were made on the lines of : — 

A. Peevention. 

B. Extirpation. 

A. — Preventive Measures. 

These should include provision for : — 

Improved hygiene of cattle sheds, etc. (especially in the direction of 
providing proper ventilation, pure water supply, and adequate disinfection 
of stalls, etc., wherein tubercular animals have been kept). This has 
been partly met in the Dairy and Milk Shops Order, but its administra- 
tion by the local health authorities is at present imperfect ; and we would 
suggest that it should be much more stringently enforced, and that 
veterinary inspectors should be given more extended powers of entry into 
all places where animals are kept. 



TUBERCULOSIS. 40S 

Improvement in the hygienic surroundings of animals should include 
isolation of all suspected cases, precautions against the flesh or milk of 
diseased animals being given as food to others— e.(/., to pigs, fowls, etc.— 
and care that fodder, litter, and water should not be taken from one 
animal or stall and given to another. 

Our attention has been drawn to the frequency with which animals,, 
obviously diseased, sometimes even in the last stage of the malady, are 
sold in open market. 

Although in England and Ireland, under the provisions of the 
Nuisances Removal Act aa embodied in the Public Health Act, 1886,. 
the medical officer of health or inspector of nuisances may seize such, 
animals, yet such seizure is rarely performed. 

We find the veterinary inspector has no power to prevent such sales, 
or to seize the beasts for slaughter, since tuberculosis is not included in 
the Contagious Diseases (Animals) Act of 1878. 

We further find that there is actually a regular trade in such stock 
infected with tuberculosis, and that they go by the name of " wasters " 
and " mincers," being frequently slaughtered in the neighbourhood of the 
larger towns, to which such portions of the meat as are likely to escape 
the observation of the inspector of nuisances are sent, for the purposes 
of sale among the poorer inhabitants, and especially for the making of 
sausages. 

We are, therefore, very strongly of opinion that power should be given 
to the veterinary inspector to seize all such animals in fairs, markets, or 
in transit. 

Notwithstanding the uniform prevalence of the disease in Europe and 
elsewhere, there seems to be no reason to apprehend that, with our 
present regulations for the slaughter of animals at the port of debarka- 
tion, and for quarantine of those imported for breeding, there is any 
special danger of increasing the infection in England by introduction 
from abroad. The danger, however, exists in regard to the stock brought 
from countries, which are exempt from slaughter on landing, and sub- 
jected to the ordinary veterinary inspection during the present period of 
detention of twelve hours. 

It is, therefore, evident that the present rules for the prevention of 
the introduction of disease into the United Kingdom from abroad, are 
incomplete. 

Since all authorities are agreed that the disease is very marked by 
heredity, we think it highly desirable that breeders should in their own, 
as well as in the public interest, discontinue breeding from tuberculous 
stock. 

B. — Extirpation. 

In order to insure the gradual extirpation of tuberculosis, we are of 
opinion that it should be included in the Contagious Diseases (Animals) 
Acts for the purposes of certain sections of those Acts, so as to provide : — 

(a) For the slaughter of diseased animals, when found diseased oa 
the owner's premises. 



404 INFECTIVE DISEASES. 

(6) For the payment of compensation for the slaughter of such 
animals. 

(c) For the seizure and slaughter of diseased animals exposed in fairs, 

markets, etc., and during transit. 

(d) For the seizure and slaughter of diseased foreign animals at the 

place of landing in this country. 

Notification of this disease should not be compulsory, because it may 
exist without developing any suflfioient outward evidence to enable the 
owner to detect it, and its gi-owth is so slow, that non-notification of its 
existence, even in a large number of cases, would do little to nullify the 
stamping-out effect of the Act of 1878. 

The powers and responsibilities of inspectors in ordering the slaughter 
of diseased animals should be the same for tuberculosis as for pleuro- 
pneumonia, according to section ,51 (5) of the Act of 1878. 

Further, tubercle, though hereditary, is nevertheless much less conta- 
gious than the other diseases included under the Act of 1878, and it is 
clear, therefore, that the immediate slaughter of diseased animals would 
go far to stamp it out, though, doubtless owing to heredity, this stamping- 
out process would be gradual in its effect. 

A supplementary report was made by Professor Horsley, in 
which he expressed the opinion that there ought to be legislation 
to prevent breeding from diseased animals, and compulsory notifi- 
cation : — 

1. Breeding. 

Tuberculosis is notorious, even among the laity, as a disease which is 
transmitted from parent to offspring. This is a fact with which cattle 
breeders are specially familiar, and which finds strong expression in the 
evidence attached to this report. Further, this generally received truth 
has been completely confirmed by the results of scientific investigation, 
as is also duly set forth in the report. Considering, therefore, the 
extreme importance of this point, I think that the act of wittingly breed- 
ing from animals so affected should be made an indictable offence. The 
only objection that can be raised to such legislation, which if effected 
would prevent the dissemination of the disease among cattle in this 
country, is that, owing to the present state of want of knowledge among 
cattle owners, and even veterinary surgeons, of the early symptoms, and 
physical signs on examination, of this disease, prosecutions would occa- 
sionally occur in cases in which no fault could properly be attributed to 
the owner, and that, therefore, such prosecutions would be needlessly 
vexatious. 

Considering, however, the extreme rarity with which such cases would 
occur, and that, as in the matter of non-notification, each case would be 
tried before district magistrates on its own merits, this objection is 
deprived of the force it might have possessed. 



TUBERCULOSIS. 405 

2. Notification of the Existence of the Disease. 

This point requires no explanation, since it is clear that, unless the 
veterinary inspectors or authorities receive information of occurrence of 
diseases, it is impossible to ensure the thorough carrying out of the 
provisions of the Contagious Diseases (Animals) Act. 

That deliberate nou-notification should be punished cannot be doubted 
by any one. Objection, however, to legislation in this direction has been 
put forward, on the same grounds as those upon which the prevention of 
breeding from diseased animals was contested. As, however, I consider 
that these objections have been already shown to have no weight, I 
recommend that both J;he forbiddance of breeding from diseased animals, 
and the notification of the disease, should be included in any legislation 
for tuberculosis. 

The difficulty referred to by the Committee, is presented by 
cases of the disease ■which cannot be detected by the ordinary 
methods of examination, and might possibly be overcome by the 
use of tuberculin as a diagnostic agent. 



CHAPTEE XXIX. 

LEPROSY. — SYPHILIS. — EHINOSCLEROMA. — TRACHOMA. 

Leprosy. 

Leprosy occurs in three forms — tubercular, anaesthetic, and mixed 
tubercular. It may be classed with the granulomata, as the most 
common form of the disease is characterised by deposits in the skin, 
mucous membrane, and internal organs. These deposits are composed 
of small cells, and large cells resembling giant cells. The cells 
become deposited in the surrounding tissues, and so the tubercle 
enlarges, involving the epidermis and developing into an ulcerating 
sore ; or, after a certain stage of development, beginning to decline, 
and finally leaving a puffy discoloration. In the anaesthetic form 
the cells invade the connective tissue of nerves. In the mixed 
form the varieties occur together, but the tubercular character 
predominates. 

Tuberpular leprosy commences with the development of an 
erythematous patch, which becomes infiltrated, and finally tubercu- 
lated, the tubercles varying in size from a millet seed to a marble, 
or even larger. The eruption on the head and face produces a 
characteristic leonine expression. The progress of the disease is very 
slow. After death the following changes may be found in the 
internal organs : Ciri'hosis of the liver and spleen, enlargement of 
the lymphatic glands, and a condition of the lungs corresponding 
to cheesy bronchial pneumonia. 

In the anaesthetic form patches develop on the skin, which 
become anaesthetic ; ulceration follows, and the fingers and to§s, or 
the entire hand and foot, may slough off. 

The disease is undoubtedly communicable, but the infectivity is 
of a very low type. 

The infectiousness is illustrated by the well-known case of Father 
Damien. Arning inoculated a man named Keanu, a condemned 
criminal,, and leprosy developed three years afterwards, but this case 

406 



LEPROSY. 407 

is not regarded as conckisive, as the man had a family history of 
leprosy. The disease has never been known to spread from patients 
in this country, who have contracted the disease abroad. 

The bacilK of leprosy were first observed by Hansen in 1874, and 
siTbsequently fully described by him, and his observations confirmed 
by Neisser, in 1879. 

Bacillus Leprae— Eods 5 to 6 ^u. in length and 1 /x, in breadth. 
The bacilli are straight or curved, resembhng very closely the 
tubercle bacilli. They are present in the leprous tubercles of the 
skin and mucous membrane, in the lymphatic glands, and in the 
liver, testicles, and kidney ; and in the nerves in the anaesthetic 
variety. They are found between the cells, and in colonies in the 
cells. They stain readily with the aniline dyes, especially by the 
Ziehl-Neelsen and Gram's methods. The bacilH are found in extra- 
ordinary numbers in the skin, and they are rather straighter than 
tubercle bacilli, and stain more readily. 

Numerous unsuccessful attempts to cultivate the bacillus have 
been made by many bacteriologists. The author has made repeated 
inoculations upon glycerine-agar, upon which the tubercle bacillus 
grew abundantly, but always with disappointing results. On the 
other hand, Bordoni-UfEreduzzi showed the author a cultivation 
which he had obtained from the bone marrow of a leper. The 
cultivation was made on blood serum and glycerine, and cover- 
glass preparations resisted decolorisation with acid. There were 
sKght morphological differences when compared with the appear- 
ance of bacillus leprae in the tissues, and the results were hardly 
conclusive. 

The English Leprosy Commission also reported successful cultiva- 
tion of the leprosy bacUlus. The author had the opportunity of 
examining one of the first cultures received in this country, and 
found that the bacilli stained deeply in ordinary cover-glass pre- 
parations, they did not resist decolorisation by the Ziehl-Neelsen 
method, and they corresponded in culture with one of the varieties 
of Bacillus subtiKs, commonly found on the skin. 

Inoculation of animals has given equally unsatisfactory results. 
Numerous experiments have been made by Beaven Rake on small 
animals and birds, with invariably negative results. The blood of 
leprous patients, tubercles from the living subject, fragments of the 
skin and of the internal organs after death, have been inoculated by 
different observers without result. Melcher and Ortmann alone claim 
to have produced really definite resvilts. These observers excised 
leprous tubercles from the living subject, and inoculated fragments 



408 INFECTIVE DISEASES. 

in the anterior chamber of the eye of rabbits. The animals died 
after some months with extensive deposits in the coecum, lymphatic 
glands, spleen, and lungs. 

These tubercles varied in size from a pin's head to a millet seed, 
and contained bacilli, resembling leprosy bacilli in their staining 
reactions. The question naturally arises whether the lesions were 
really indicative of leprosy or tuberculosis. Until the experi- 
ments are independently confirmed, and the result of inoculation 
differentiated from tuberculosis, it would be rash to accept these 
experiments as conckisive. 

It has been suggested that tuberculosis and leprosy are identical. 
There is a similarity in the bacilli and in the lesions of leprosy and 
tuberculosis, the injection of tuberculin produces a reaction in leprosy 
nodules, and many lepers die from tubercular disease of the lung. 
But while tuberculosis is very readily transmitted to guinea-pigs 
and rabbits by inoculation of fragments of tubercular tissue, leprosy 
is inoculable, if at all, in most exceptional instances. The bacilli 
of tubercle are cultivated with the greatest facility, the bacilli of 
leprosy, if at all, only with exceptional difficulty; tubercle bacilli 
are found in giant cells, leprosy bacilli in the so-called leprosy 
cells. Leprosy bacilli are straighter than human tubercle bacilli,^ 
and differ slightly in their behaviour to staining reagents. On 
the other hand, the morphological differences are not greater than 
those existing between different forms of tubercle bacilli obtained 
from tuberculosis in animals and birds. It would be premature 
to regard leprosy as a variety of tubercle until cultivations of 
the bacillus have been obtained, and carefully compared with 
those of the tubercle bacillus. Differences in morphological details 
and results of inoculation would then carry less weight as a means 
of differentiation. 

The tubercular pneumonia of lepers would be regarded, if the 
bacilli are identical, as a development of leprosy in the lungs, and 
not, as at present, a result of double infection with tuberculosis. 

Methods of Examining the Bacillus of Leprosy. 

Cover-glass preparations may be made in the ordinary way, or by a 
special method, which consists in clamping a nodule with a pile clamp 
untU a state of anaemia of the tissue is produced. On pricking with a 
needle or sharp knife a drop of clear liquid exudes, from which cover- 
glass preparations may he made, and stained by Neelsen's method. 

For sections the author prefers Neelsen's method and methylene-blue. 
They can also be stained by Gram's method, which, as a rule, brings out 
very clearly the beaded appearance of the bacilli. 



DESCRIPTION OF PLATE XIV. 
Bacillus LeprsB. 

Fig. 1. — From a section of the skin of a leper. The section is, almost in 
its entirety, stained red, and, with moderate amplification, has a finely 
granular appearance. Stained by the Ziehl-Neelsen method (carbolised 
fuchsine and methylene-blue). x 200. 

Fig. 2. — Part of the same preparation with high amplification, showing that 
the appearances described above are due entirely to an invasion of the 
tissue by the bacilli of leprosy, x 1500. 




i'lgl 




ri£? 2 



BACILLUS LEPR^ 



LEPROSY. 4()9- 

Method of Bahh. — Preparations are stained in rosaniline hydroohlorate- 
in aniline water, decolorised in 33 per cent, hydrocliloric acid, and after- 
stained with methylene-blue. 

Stamping-out System The history of leprosy in the British 

Islands during the Middle Ages, and the conditions under which it 
both increased and declined, have been discussed by several writers. 
A large number of institutions of a charitable and ecclesiastical 
character were established in endemic areas and were occupied by 
the lepers either voluntarily, or compulsorily by means of the Act 
De le]}roso amovendo. These institutions were to a very small extent 
a means of segregation. According' to Dr. Newman the disease, 
which had reached its zenith about the twelfth or thirteenth century, 
began to decline from that time owing to '' a general and extensive 
social improvement in the hfe of the people, to a complete change 
in the poor and insufficient diet (which it is evident consisted far 
too largely of bad meat, salt, putrid and dried fish, and an almost 
entire lack of vegetables) and to agricultural advancement, improved 
sanitation and land drainage." Of all the unfavourable conditions 
it woiild appear that food in some way was especially associated with 
the cause of the disease, either by introducing the bacillus or by 
rendering the tissues a suitable soil for its reception and development.. 

In other countries segregation has been attempted voluntarily 
or compulsorily, but it has never been completely carried out. There 
can be very little doubt that the presence of a leper in a healthy 
community is no greater source of danger than the presence of 
an individual suffering from tuberculosis, but, for other reasons,, 
voluntary isolation should be carried out as completely as local 
circumstances will permit. 

The Leprosy Commission in India recommended — ■ 

(a) That the sale of_ articles of food and drink by lepers should 

be prohibited, and that lepers should be prevented from 

following certain specified occupations. 
(6) That the concentration of lepers in towns should be discouraged, 
(c) That Leper Asylums should be established in which lepers 

might live voluntarily. 
{d) That Leper Farms scattered over the country should be 

encouraged, 
(e) That the few children who are born of lepers should be 

removed to Orphanages. 

They concluded that by means of improved sanitation and good 
dietetic conditions a diminution of leprosy will result. 



410 INFECTIVE DISEASES. 

Syphilis. 

Syphilis is a disease peculiar to man, and communicable oidy by 
inoculation. The local infection is followed by a period of latency, 
and by a period during which generalised eruptions appear. One 
attack confers immunity from future attacks'. The virus in its 
most virulent form is found in the primary seat of inoculation, and 

in the indurated glands which 
follow. It is also supposed to be 
present in the blood and secretions. 
Lustgarten, Eve and Lingard have 
found bacteria which they beUeved 
to be specific. 



'^'iWfih^M'. Bacillus in Syphilis (Lustgar- 

(^(g,^^^^ ten). — Rods resembling the bacilli 




N^?* 




of leprosy and tuberculosis, 3 to 4 yu, 
Fig. 175. — Covee-glass peepaea- , „ , , . i m 

TIONOpPuSFEOftACHANCEE, X ^°^S' '^ /^ ^^^<'^- ^WO Or mOre 

1050 (Lustgabten). colourless, ovoid points in the course 

of the rod are visible with a high 
power ; it is thought that possibly they are spores. The bacilli 
are always found in the interior of nucleated cells, which are 
more than double the size of 
leucocytes. They have been ob- 
served in the discharge of the ?■' §^ '^ii^^-- / 
prima]*y lesion, and in tertiary '" -' •-"^ ■ 
eummata. 

Alvarez and Tavel state that ;*:: *^';,.^4-.';'^Sr^:/'?f ;, '.■'■.•.V',';; 

an identical bacillus is found in '^i-'* •■: .^ ■'■'-' •"■,K^^^rT J: ''X 

•: -'I' '- i/ ■'"'■■.': ;■>.''■)' '-iV"'"*-*.' 

normal secretions (smegma). Eve ■:;^i^'■C<*•"v■i^J: „,'.:■; '.,.,;;• 

and Lingard have described a bacil- -^f-;.- ;:'!■, ^1' ■,■"•, ---'■ "^:'- 

lus associated with specific lesions, =•.%-• ■^„,.'?"-'-^"y.t' -■•.:!.>■•. ii-',?' 
which differs from the above in its 

morphology and behaviour towards 3^ig. 176.— Wandeeing Ceil con- 

staining reagents. ^^'^^-^ 2"^'^'''" (Lustgarten). 

Methods of Staining the Bacillus of Syphilis. 

21ethod of Lustgarten : — 

Sections are placed for twelve to twenty-four hours in the following 
solution, at the ordinary temperature of the room, and finally the solution 
is warmed for two hours at 60° C. : — 

Concentrated alcoholic solution of gentian-violet . 11 

Aniline water . . .... 100 



SYPHILIS. 41 1 

The sections are then placed for a few minutes in absolute alcohol, 
and from this transferred to a 1'5 per cent, solution of permanganate of 
potash. After ten minutes they are immersed for a moment in a pure 
concentrated solution of sulphurous acid. If the section is not completely 
decolorised, immersion in the alcohol and in the acid bath must be 
repeated three or four times. The sections are finally dehydrated with 
absolute alcohol, cleared with clove-oil, and mounted in Canada balsam. 

By this method the bacillus is distinguished from many bacteria, but 
not from the bacilli of tubercle and leprosy which are stained by this 
process. 

Method of De Giacomi : — 

Cover-glass preparations are stained with hot solution of fuchsine 
containing a few drops of perchloride of iron. They are then decolorised 
in strong perchloride of iron, and after-stained with vesuvin or Bismarck- 
brown. 

Method of Doutrelepont and Schutz : — 

Sections are stained in a weak aqueous solution of gentian-violet and 
after-stained with safranin. 

The nature of the contagium in syphilis is unknown. 

Protective Inoculation. — Inoculation of the virus, or sypMlisa- 
tion, as a protective measure, was at one time practised and strongly 
advocated ; but it is rightly regarded in this country as dangerous 
and unjustifiable. From the experiments of Ricord it would appear 
that the local results in the vesicular stage resenable the results of 
the inoculation of virulent vaccinogenic grease or horse-pox. The 
inoculation goes through the stages of papule, vesicle, ulcer, scab, 
and scar. The accidental inoculation which occurs in cases of 
vaocino-syphihs may so closely resemble the results of inoculation 
with very virulent cow-pox, that it is sometimes difficult to decide 
as to the exact nature of these cases. 

Ehinoscleboma. 

E/hinoscleroma is a rare disease, resembling lupus, and pro- 
ducing in the nostrils and neighbouring parts nodular swellings, 
composed of granulation-tissue. The disease is met with in 
America, Egypt, Austria, and Italy. There are no giant cells, but 
peculiar large cells, which were first described by Mikulicz. Frisch 
discovered bacteria in sections, and Cornil and Alvarez pointed out 
the existence of a capsule. In morphology and cultivation they 
resemble, according to Dittrich, Friedlander's pneumococcus. They 
are probably identical with this micro-organism, and Paltauf and 
Eiselsberg, and others, found that they produced septicaemia in rabbits 
and guinea-pigs. 

Bacterium of Rhinoscleroma {Bacillus of RUnosolero-ma, 



412 INFECTIVE DISEASES. 

Cornil and Alvarez). — Cocci and short rods, 1 '5 to 3 /j, in length,. 
•5 to "8 fi thick. Deeply coloured points or granules may occur 
in the course of the rods when stained, but it is very doubtful 
whether these can be considered as spores. The bacteria are en- 
capsuled, the capsule being round when enclosing a coccus, and 
ovoid when enclosing a rod. The capsule is composed of a tough 
resisting substance; two or more capsules may unite by fusion, 
enclosing two, three, or a greater number of rods. The bacilli were 
observed in sections of the tumours, which developed on the lips 
and in the nasal and pharyngo-laryngeal regions. 

Method op Staining the Bacillus of Rhino- Scleroma. 

Method of Cornil and Alvarez : — 

Sections are immersed in a solution of methyl-violet (B) for twenty- 
four to forty-eight hours, with or without the addition of aniline-water ; 
and are then decolorised after treatment with the solution of iodine in 
iodide of potassium. If the sections are left to decolorise in alcohol for 
forty-eight hours, the capsule is rendered visible. 

Trachoma. 

Trachoma is a disease of the conjunctiva, common in Egypt. 
The new growth is composed of round cells, and may be regarded, 
according to Kartulis, as the chronic stage of either gonorrhoeal or 
Egyptian ophthalmia. Koch failed to find any micro-organisms 
in the swollen lymph follicles. Sattler asserted that he had culti- 
vated a micrococcus which produced the disease when inoculated 
on the conjunctiva. Other observers have found the common 
pyogenic micrococci in the secretions, especially Staphylococcus 
pyogenes aureus and albus. 



CHAPTEE XXX. 

actinomycosis. — madura disease. 

Actinomycosis. 

Actinomycosis belongs to the class of infective granulomata. It is 
a chronic inflammatory affection characterised by the presence of a 
special microphyte, which by irritation produces a neoplasm, composed 
of roiind cells, epithelioid cells, giant cells, and fibrous tissue. These 
neoplasms form nodular tumours of various sizes. In some cases 
there is a tendency to develop very large tumours, and in others to 
break down early and suppurate. In cattle, cretification takes 
place in the fungus tufts. Actinomycosis closely resembles tuber- 
culosis in its histological characters. The disease attacks man, 
horses, cattle, and pigs. 

Many interesting observations have been made upon the origin 
■of this disease in man. Two cases have been recorded in support of 
the theory of direct infection from the cow. Stelzner described a 
case of actinomycosis in a man who had had the care of animals, 
some of which had suppurating glands. Hacker had a case of 
actinomycosis of the tongue in a man who had charge of cows, one 
of which had a tumour of the jaw which he had opened. On the 
other hand, Moosbrugger found that out of 75 cases, 54 were in men, 
and 21 in women, including 2 children. In 11 of these men the 
occupation was not stated. In 33 their occupation did not bring 
them into contact with diseased animals ; they were, for example, 
miUers, glaziers, tailors, shop people, and students. Only 10 cases 
occurred among farmers, peasants, and farm-labourers, and in only 
one case out of the 10, had the patient been brought into contact 
with diseased animals. 

Out of the 21 women, there were only 4 peasants, and none 
of them had been associated with diseased cattle. 

Infection by the flesh of diseased animals has also been dis- 
■cussed. But there is no evidence of prevalence of the disease 

413 



414 INFECTIVE DISEASES. 

among slaughterers and butchers, who would be particularly liable 
to it if flesh were a source of infection. The chances of infection 
by ingestion are minimised by the flesh being almost always cooked. 
Actinomycosis occurs also in pigs, and pork is very often eaten in an 
uncooked state ; but Israel has pointed out that this may probably 
be excluded, as many of the cases occurred among strict Jews. 

The evidence points to the disease originating in man and lower 
animals from the same source, and there is a very strong suspicion 
attached to cereals. This view is supported by important obser- 
vations, with reference to the part played by cereals in inducing 
the disease in cattle, and it gains additional support from a case 
described by Soltmann, where the disease resulted from an awn of 
wall barley. A boy, aged eleven, accidentally swallowed an awn 
of Hordeum murinum. He became very ill, and suffered great pain 
behind the sternum, extending to the back. An abscess formed,, 
covering an area extending over six intercostal spaces, and when 
opened, the awn of this grass was found in the evacuated pus. 
The pain, however, continued, and fresh deposits occurred, and when 
the boy was taken to the hospital, the ray-fungus was detected. 
Possibly the spores of the fungus can be conveyed both by air and 
water. 

This disease in cattle has long been known in this country, but 
its various manifestations were either mistaken for other diseases, 
or simply received popular names. Indeed, the various forms are 
still familiar to many as wens, clyers or crewels, scrofulous, tuber- 
cular or strumous abscesses, polypus, lymphoma, cancer of the 
tongue, scirrhous tongue, indurated tongue, ulcerated tongue, cancer 
of bone, bone tubercle, osteo-sarcoma, fibroplastic degeneration of 
bone, spina ventosa, and carcinoma. 

Bovine antinomycosis is especially prevalent in river valleys, 
marshes, and on land reclaimed from the sea. The disease occurs 
at all times of the year, but general experience leads to the belief 
that it occurs more commonly in the winter. 

It is more frequently met with in young animals, and usually 
occurs between one and three years, but it may be found at almost 
any age, and probably affects equally both sexes. 

There is little if any evidence to show that the disease is heredi- 
tary. In numerous cases, the family history has been most carefully 
inquired into by the author ; and in the case of some imported 
pedigree animals, the disease was quite unknown on the farm wher& 
they had been bred. 

The tongue is so commonly the seat of the disease, that suspicion 



ACTINOMYCOSIS. 415 

at once falls on food as the means by which the parasite is conveyed. 
Skin wounds produced by rubbing against the mangers, posts, or 
wire fencing, may also become infected. 

The evidence is very strong in favour of believing that the micro- 
organism gains access to the system through wounds or lacerations 
of the mucous membrane and skin, or through carious teeth. It 
has been pointed out that the common occurrence of the disease 
at the time of the second dentition may be owing to the wounds 
produced in the alveolar mucous membrane by the shedding of 
the teeth. Experience also points to straw being sometimes a 
factor in the production of the disease, and it is possible that thistles 
and frozen roots also, by wounding the mucous membrane, may 
afford a way for the entrance of the micro-organism. The disease 
in the jaws, both in man and in cattle, is very commonly associated 
with carious teeth. 

The cowsheds, pastures, and drinking tanks may become infected 
with the discharges from diseased animals. The discharge con- 
taminates the fodder in the sheds, and falls on thistles and siKceous 
grasses in the pasture, which may first wound, and then introduce 
the micro-organism. The discharge is also coughed out of the 
mouth, and expelled from the nose, in cases in which a tumour in 
the pharynx, or the nasal chambers, has undergone suppuration. 

Jensen believed that the disease was produced by different kinds 
of grain, especially when cultivated on ground reclaimed from the 
sea. He mentions an instance of a farm, where nearly the whole 
of the young stock, about thirty in number, had actinomycosis after 
feeding on mixed forage, grown on a certain field. Two years after- 
wards the same disease occurred in the same stalls in four animals, 
after being fed on barley-straw from the same field. According to 
Jensen, the fungus grows on grain, husks, and straw of different 
cereals, but most abundantly on barley, which is also the most 
likely to wound the mucous membrane. Johne's observations tend 
to corroborate this view, for in twenty-two out of twenty-four cases 
in which he found barley sticking in the tonsils of pigs, he found 
the beard thickly beset with a fungus very similar to, if not identical 
with, the ray-fungus; These observations are of great interest in 
connection with Soltmann's case. 

Experience points to the belief that the disease is not readUy 
communicable from animal to animal, and it is possible that when 
it affects a large number of cattle in a herd, the same causes have 
been acting to produce the disease in a number, which in another 
instance may only produce it in one. At the same time, isolated 



416 INFECTIVE DISEASES. 

cases are possibly not quite so common as they are reported to be. 
It is well known that, as a rule, the services of a veterinary surgeon 
are not called for except in hopeless or very severe cases. The 
cowmen themselves, in many districts, treat the cows successfully, 
and then send them into the market, and thus the existence of 
previous cases may not have come to the knowledge of the veterinary 
surgeon. 

Historical. — In 1845 Professor von Langenbeck, of Kiel, made 
notes of a ease of vertebral caries in a man, and prepared drawings 
of peculiar bodies in the pus from an abscess. The drawings were 
published together with a reference to the case by Israel in 1878. 
There can be little doiibt that these structures were the fungi of 
actinomycosis. But the first to publish observations was Lebert 
in 1848. 

Lebert received from M. Louis some pus, of a thick, almost 
gelatinous consistency, which had been obtained from an abscess of 
the thoracic wall in a man aged fifty. The patient had been attacked 
four months previously by a pulmonary affection, which was 
suspected by M. Louis to be cancerous in nature. The pus contained 
a very considerable number of little spherical bodies of a slightly 
greenish-yellow colour, about the size of a pin's head. They 
could be readily crushed between two strips of glass, and on 
examination with a power of fifty diameters two elements could be 
distinguished : a soft connective substance, and many hard, narrow, 
wedge-shaped corpuscles, arranged in a radiating manner. Under 
a high power these bodies were observed to be -Jg- to j\^ of an inch 
in length, -j^^ in width at the base, and ^-J^ in width at the 
apex. Some of these corpuscles were regular, while others showed 
one or two constrictions, with intermediate flask-shaped swellings. 
Lebert tested these structures with reagents, with the following 
results. The bodies were found to remain unaltered by concentrated 
mineral acids. Acetic acid freed them . from foreign elements 
adhering to their surface. Solution of caustic potash did not affect 
them if used cold, but a boiling solution reduced the cuneiform 
structures to a fine greyish powder without dissolving them. Ether, 
alcohol, and chloroform had no effect upon them when used either 
hot or cold. Solution of potash, in which these bodies had been 
heated, mixed with a solution of sulphate of copper and brought to 
boiling point, did not offer any uniform red colour, which would 
have been the case if they had contained albumin. Thus, the chief 
chemical characters of albuminous and fatty substances were 
wanting, and they resembled chitine in their behaviour to reagents. 



ACTINOMYCOSIS. 



417 



Lebert bore in mind the possible existence of some helminthic debris, 
of which these bodies might be hooklets, but he sought in vain for 
echinococci and cysticerci. 

Actinomycotic pus was later described and figured by Robin. 
In the illustration accompanying the description, the fungi are most 




Fig. 177.— Section of Livek fkom a Case oi' Actinomycosis in Man. 

accurately depicted. Eobin states that he had found, in two or 
three cases in the pus of deep-seated chronic abscesses, yellowish grains 
attaining a diameter of one-tenth of a mm., surrounded by a sort of 
halo or thin, viscous, finely granular stratum, containing leucocytes. 
These grains were composed of elements 2 to 6 mm. in length, swollen 

27 



418 INFECTIVE DISEASES. 

at one end and tapering off at the other, arranged in a regular series, 
radiating from a common centre which consisted of granular matter. 
They were highly refractive, possessed a brilliant centre and sharply 
■defined outline ; they were dissolved, or at least rendered indistinct, 
by acetic acid, and proved insoluble in ammonia and ether. 

The disease in man was next described by Israel in the paper 
mentioned above. Ponfick was the first to clearly recognise the 
identity of the disease in man with the disease in cattle, and he 
described a number of cases in man. Israel subsequently published 
a work on the subject. The various cases which had been observed 
up to that date were described, and the disease classified according 
to the seat of invasion. 

From this time onwards numbers of cases in man have teen 
described, and various important researches published, of which 
those of Bostrom and Moosbrugger may be especially mentioned. 

In England, Acland recognised a case on examining the liver 
after death (Fig. 177). H. Taylor was the first,, in this country, 
to detect the fungus during the life of a patient. Shattock found 
specimens of the disease in museums. Skerrit, Powell and Godlee, 
Eve, Delepine, Eansome, Poore, Malcolm Morris and others have 
published cases. 

In Italy, Perroncito studied the sarcomata of cattle, and claims 
to have first observed the micro-organism in 1863. In 1876 he 
described it in the Encyclopaedia Agraria, and, from the negative 
results obtained by inoculation experiments, was led to regard it, 
not as the cause, but as a result of the disease. 

Rivolta of Turin also claims to have been the first to have 
discovered the fungus in actinomycosis bovis. As early as 1868 
he published a paper on a sarcomatous tumour of the jaw of 
an ox. 

Hahn of Munich, in 1870, undoubtedly met with the fungus, for 
he states that in a case of "wooden-tongue " he found characteristic 
organised structures, which he provisionally described as a species of 
mould fungus. 

Bollinger was the first to recognise the nature of this disease in 
cattle. In 1876 he pointed out that new growths occasionally 
occurred on the upper and lower jaws of cattle, which either started 
from the alveoli of the back teeth, or from the spongy tissue of the 
bone, and by increasing in size loosened the teeth. In their progress 
they destroyed bone, muscles, mucous membrane, and skin. After 
-some time they frequently broke down, forming ulcers, abscesses, 
-and fistula ; but in some cases tumours were formed, which attained 



ACTINOMYCOSIS. 419 

the size of a child's head. BoUinger stated that this disease had 
been known by various na,m.es,—Osteosarkome, Winddm-n {Spina 
venipsa), Knochenkrehs, Knochenwurm ; in other instances it had been 
regarded as bone tuberculosis, or mistaken for a simple chronic 
glossitis. Among breeders of cattle and owners of stock in Germany 
it had been known under the following names : Ladend/ruck, Laden- 
geschwulst, dicker Backen, Boickel, Einnheule, Kiefergeachwulst, etc. 

Bollinger pointed out that these swellings 'consisted of several 
centres of growth, bound together by connective tissue. They were 
often as large as a walnut or a hen's egg, and of a pale yellow 
colour and moist appearance. The cut surface presented yellowish- 
white, suppurative foci, while in other cases the growths had a 
spongy texture, owing to the formation of lacuna or hollow spaces 
in a fibrous stroma, which contained a turbid, thick, yellow, caseous 
pulp. 

Microscopical examination of the tumour showed that it had a 
structure like a sarcoma, while the squeezed-out pulp consisted 
principally of pus cells, granulation cells, fat granules, and granular 
detritus. In addition, there were numerous opaque, pale-yellow, and 
coarsely granular bodies of different sizes, which had a mulberry- 
like appearance, and were sometimes encrusted with chalk. After 
careful examination Bollinger found that these bodies were true 
fungi, and he further maintained, from the constancy of their 
appearance in all parts of the sarcomatous growth, that they were 
not accidental, but of pathogenic significance. This was found to 
be the case, not only in fresh preparations, but in old specimens 
preserved in the museum. This remarkable form of mycosis was 
observed by Bollinger, not only in the upper and lower jaws, but 
also in the tongue. It had long been observed that the tongue was 
sometimes covered with more or less tubercular growths, scattered 
abundantly over the surface of the mucous membrane, mostly the size 
of a millet seed or hemp seed, but often reaching the size of a cherry 
or walnut, or even larger. In the fresh state these nodules were 
greyish-white, and semi-transparent, but they soon became cloudy 
or distinctly purif orm in the centre ; they were surrounded externally 
with a connective tissue capsule. If the nodules were situated on 
the surface of the tongue, destruction of the mucous membrane very 
readily followed, leading to the formation of ulcers. The tongue 
also might become affected with an interstitial glossitis, which often, 
in spite of the partial atrophy of the muscular fibres, led to a great 
enlargement and wood-like hardness of the tongue. On account 
of this peculiar character, such a tongue was long known in South 



420 INFECTIVE DISEASES. 

Germany as Eolzzunge. In other cases the condition was regarded 
as " tubercle of the tongue," " chronic sarcoma," " chronic interstitial 
glossitis,'' or simply " degeneration of the tongue.'' 

Bollinger described this disease as occurring in cattle of aU ages, 
developing itself gradually, and being always incurable. As a rule, 
the animals were slaughtered, because the diminished mobility and 
enlargement of the tongue interfered with feeding. He also pointed 
out that this disease of the tongue was by no means rare, as he had 
had no less than six such tongues from diflferent parts of Bavaria in 
the space of a year, and he also had been able to prove the existence 
of the disease in museum specimens. 

On further continuing his researches, Bollinger found the same 
fungus in tumours which occurred in the pharynx, larynx, - and 
the mucous membrane of the stomach. These tumours were very 
common in the throat in some parts of North Germany, where as- 
many as 5 per cent, of the animals had been known to be affected. 
The disease frequently occurred in the form of subcutaneous 
neoplasms, called Lymphome, Hohzgeschwiilste, Fibrome, Tuberkd, 
Tuherhel-scroplieln. 

This disease also appeared in the form of abscesses, which were 
called, in many districts, Schlundheulen. These growths were found 
in the neighbotirhood of the parotid gland, the larynx, and pharynx, 
and were similar in every respect to the affection of the jaw. They 
were described as starting apparently from lymphatic vessels in 
these parts. Bollinger discovered the fungus in a case of so-called 
fibroid of the second stomach of a cow, a spongy growth nearly the 
size of the fist ; and he believed that in another case the disease 
manifested itself in the form of tubercular ulceration of the 
intestines. 

BolHnger submitted the fungus to Dr. Harz, a botanist, who 
described the fungi as mulberry-like masses from '5 to 1 mm. in 
diameter. They appeared to the naked-eye as opaque, white grains, 
and when calcified were difficult to recognise. On slight pressure 
the tufts of the fungi fell apart into segments of unequal size, each 
of which appeared to correspond to an individual fungus. The 
latter was described as beginning at the pointed end of the wedge, 
with a somewhat cone-shaped basal cell, which, in the absence of a 
mycelium, perhaps took its place, and bore a great number of short 
linked hyphse. At the ends of the hyphse there were oval, globular, 
or elongated club-shaped bodies, the reproductive cells or gonidia. 

Cultivation experiments, and inoculation of the tongue of a calf 
with liquid containing the micro-organism, failed. Harz proposed 



ACTINOMYCOSIS. 421 

to call the fungus, from its ray -like appearance, actinomyces ; but 
what the position of the fungus in nature might be, was difficult to 
determine. It did not, he believed, belong to the yeast fungi, but to 
the mould fungi, and might be compared to Botrytis, Monosporium, 
and Polyactis. 

Bollinger concluded that there could be no doubt that actino- 
mycosis occupied an important position in the pathology of cattle 
diseases. As further evidence of the prevalence of the affection, he 
remarked that Zippehus of Obernburg had observed in the course of 
about ten years' practice not less than 254 cases of lymphoma, in 
the neighbourhood of the larynx and pharynx, besides 157 cases 
of disease of the jaw ; and Bollinger says that he had very little 
doubt that the greater part of the former, and very likely all the 
cases in the jaw, were due to the fungus which he had discovered. 
In certain parts of Franconia, according to a communication received 
from Professor Frank, these tumours of the throat were extremely 
abundant in cattle. 

Bollinger's researches were followed by those of Siedamgrotzky, 
and later by a communication from Johne. Johne described the 
various forms of the disease which had up to that date been 
recognised, including a description of actinomycosis of the ' bones 
of the jaws, of the fauces, of the larynx, of the oesophagus, of the 
stomach and intestinal canal, and of the udder. He carried out a 
series of experiments, by which ib was clearly estabhshed that the 
disease could be communicated from cattle to cattle. Previously 
Bolhnger, Harz, Perroncito, Ponfick, Siedamgrotzky, and Johne had 
failed, but subsequently by employing fresh material from the 
living animal, both Johne and Ponfick succeeded. 

Siedamgrotzky not only confirmed BolKnger's researches, but he 
described the presence of the fungus in so-called " multiple sarcomas " 
of the mucous membrane of the oesophagus. Babe described the 
presence of the fungus in tumours known as Winddorn, and pointed 
out that, in at least one case, he considered that the disease had 
been carried by the lymphatics. There were eleven subcutaneous 
tumours in a row on the face, which were connected by swollen, 
rope-hke, lymphatic vessels. They appeared to be secondary to a 
growth on the nostril, the size of a hen's egg. 

Perroncito described a case of " sarcoma " of the intestines and 
stomach, which proved to be actinomycosis. 

Many additional communications were made on the subject of 
this disease. Ponfick produced it in the lungs by intravenous 
injection, and subsequently three cases occurring naturally in the 



422 INFECTIVE DISEASES. 

practice of veterinary surgeons were published. They not only 
deserve especial mention, but as this form of the disease appears to 
be so seldom recognised, they will be given in detail. 

Plug described a case in the lungs. The cow had been out of 
health for four weeks, did not eat, and had a cough, and two days 
previous to the visit had become rapidly worse. Schmidt found 
dyspnoea with abdominal respiration ; the nostrils were dilated, the 
head protruded, and the mouth kept open. There was dulness on 
percussion, and crepitation. The animal was killed, and the lungs, 
which alone were diseased, were sent to Plug. The pleura on exami- 
nation was normal, but beneath it were numbers of miliary 
tubercles, many equal in size to a pin's head. On section the lung 
had a granular appearance from the presence of countless numbers 
of minute deposits, which all had the appearance of grey tubercles ; 
in none was there any central softening. They were present in 
enormous numbers around the bronchi, and in the vessels of the 
interlobular tissue. Microscopical examination showed, in the middle 
of most of these nodules, the presence of greenish-yellow, radiating 
bodies, which under a high power appeared to be undoubtedly 
actinomycotic granules. In many there were only rudimentary fungi 
consisting of four or five clubs ; there was only one rosette in each 
tubercle. The fungus was ' surrovmded by round cells and fibrous 
tissue. Larger nodules resulted from the agglomeration of several 
tubercles, or from diffuse infiltration of round cells in the neighbour- 
hood of a tubercle. 

Hink met with a somewhat similar case. A ten-year-old cow 
was slaughtered, and in the middle lobe of the right lung there were 
yellowish nodules about the size of a pea, scattered over an area 
the size of the palm of the hand. These nodules were not at 
first sight distinguishable from ordinary tubercles, but on closer 
inspection they appeared to be somewhat different, and could be 
easily shelled out from the thickened lung tissue. On making a 
section, pus welled up at several points, and contained yellowish, 
calcareous particles. These particles, on microscopical examination, 
were found to be strongly calcified tufts of the actinomyces embedded 
in granulation cells. Addition of hydrochloric acid dissolved the 
calcareous matter, but had no action on the fungus. 

Pusch described a third case. The lungs of a cow, which had 
been killed on suspicion of having pleuro-pneumonia, were sent for 
examination. The front lobe of the left lung was collapsed and 
firm, the pleura was thickened and opaque ; the larger bronchi were 
enlarged, filled with pus, and their walls thickened. In the posterior 



ACTINOMYCOSIS. 423 

lobe of the left lung there was a cavity the size of the fist, which 
had been opened, and the contents had, for the most part, escaped ; 
what remained was a greyish, purulent liquid, full of yellowish 
bodies. By the side of this cavity there was another collection of 
pus, the size of a walnvit. In the lower part of the second lobe of 
the right lung there was a firm, grey tumour, the size of a hen's 
egg, over which the pleura was much thickened. On section this 
was cavernous, with similar purulent contents, and yellow grains. 
These grains under the microscope proved to be ray-fungi. The wall 
of the cavity consisted of dense connective tissue lined with a soft 
granulation tissue, bathed in pus. There was no disease of any 
other parts in this case, so that it corresponded in this respect with 
the two previous ones. Pusch adds that it was difficult to determine 
whether the organism had gained access to the lungs by the blood- 
vessels, or by the inspired air. In his case he inchned to the latter 
view, and concludes by saying that the organism is probably very 
common and attached to the most varied objects, from which it 
may be conveyed by the air. 

Pusch refers in the same paper to an interesting case which 
occurred in the practice of Eggeling. The latter had under his care 
a cow with extensive paralysis. The spinal cord was compressed 
by a compact swelling in the neck, consisting of the nodules of 
actinomycosis. There were no manifestations of disease in any other 
part of the body. 

Prevalence of the Disease. — The author found that the disease 
was not generally recognised as a common affection of cattle 
in this country, in spite of the interest excited by the work of 
Fleming, to whom is due the credit of first recognising a case in 
England. In 1887 there was a disease prevaihng in Norfolk, and 
in the following year outbreaks were investigated by the author 
in Essex, Hertfordshire, Cambridgeshire, and Middlesex. In the 
Norfolk outbreak the author found on one farm 8 per cent, of the 
beasts affected with the so-called " wens " or " sitfasts," which 
proved on microscopical examination to be oases of actinomycosis. 
These growths had previously been described in veterinary text-books 
as the result of strumous or scrofulous inflammation ; but in all 
the specimens of wens received from this country and the colonies, 
the author has been able to demonstrate the presence of the ray- 
fungus. 

A case of pulmonary actinomycosis, with grape-Hke growths on 
the pleura, indicated that wens were not the only manifestation of 
this disease, which had been lost sight of under the designation of 



424 



INFECTIVE DISEASES. 



tuberculosis. Many other cases were examined, and the disease 
was shown to be prevalent in this country. 




Fig. 178. — From a photograph of a Norfolk steer. There is a growth about the 
size of an orange in front of the throat, an example of a so-called " .scrofulous " 
or "strumous" tumour. This growth was associated with a large polypoid 
growth in the pharynx which, by interference with deglutition, produced 
emaciation (Fig. 180). 

In Australia actinomyco.sis commonly occurs in the form of 
tumours of the upper and lower jaw, which were attributed to 

" cancer " or to " scrofulous 
inflammation." The diseace 
is still commonly known in 
Australia as " cancer " and 
" lumpy jaw." 

Reports of the prevalence 
of actinomycosis in the United 
States have been published 
by the Board of Live Stock 
Commissioners for the State 
of Illinois. In their Report 
for 1890 several intere.sting 
communications were pub- 
lished. Mr. Casewell, State 
Veterinarian, investigated an 
outbrejik of this disease, known also in America as " lumpy jaw,' 





Fig. 179.— a Noki'OLk Heifer with \ 
L.iKGE ' Wen " is the P.vhotiu Region. 



ACTINOMYCOSIS. 



425 



on a farm in Yates City, where there were 80 head of cattle, and 
16 were found to be suffering from actinomycosis. Mr. Casewell 




-n^j*--. -^^ 



Tig. 180. — Photograph of a steer nearly three years old, but about the size of a 
yearling. The emaciation and deplorable aspect recall the api^earance of 
" a piner " or " waster " (tuberculosis). 

reported that the disease was prevalent in nearly every county in 
that State, and that in his opinion it was spreachng. In one 
instance 109 cases were slaughtered. 

Actinoimjcosis in Relation to Tuberculosis. — When we considei' 
the very high percentage of cases 
of tuberculosis which has been 
reported in some localities, the im- 
portance of differentiating actino- 
mycosis from tuberculosis cannot 
be over-estimated. The very 
great contrast in the appearance 
of the micro-organisms in the 
two cases renders this a very 
«asy matter for the pathologist. 
But practical veterinarians and 
breeders of cattle are liable to 
mistake some manifestations of 
actinomycosis for tuberculosis. 

It is of the greatest im- 
portance to bear in mind that 
wens or clyers are really not tubercular, but actinomycotic ; and 



m 


i 


^^ 


k 


■^^9 


wl 


^-^ . 


' ■-«f^. 


|: m-^S. '■■-'' 



Fig. 



181.— Actinomycotic Nouules 

FKOM THE PlEUKA. 



426 INFECTIVE DISEASES. 

that a condition of tlie lungs, may occur as the result of actino- 
mycosis, which from the naked-eye appearances may be mistaken for 
"grapes" or " angleberries." It will be well also to remember in 
connection with the above remarks, that extreme emaciation may 
result in actinomycosis, producing a condition which, without a 
post-mortem examination, would probably be attributed to tuber- 
culosis, the animal being regarded as a " piner " or " waster." 
If these possible fallacies are taken into account, the excessive per- 
centage of tubercular cases so commonly reported will be very 
considerably reduced. 

There is no evidence to show that the flesh of animals suffering 
from actinomycotic tumours is unfit for human consumption. In 
very severe cases it is iinwholesome, but there is no evidence that 
it can produce actinomycosis in man. 

Manifestations of Actinomycosis in Man. 

(I.) Invasion hy the Mouth and Pha/rynx. — The fungus may gain 
access through carious teeth, or wounds or fistulse of the jaw, and 
very possibly by inflammatory processes in the pharynx and tonsils. 

The disease attacks the 'lower jaw most frequently. The tumour 
is found in close connection with the bone, or in the sub-maxillary 
or sub-mental regions, and also in the prse-tracheal region. It occurs, 
though rarely, in the interior of the bone. 

In a case described by Israel, which occurred in a woman aged 
forty-six, there was a small tumour about the size of a cherry 
attached to the external surface of the lower jaw, with an opening 
through which a probe could be passed into the bone. The tumour 
was incised and scraped away, and a cavity discovered in the bone, 
admitting a small sharp-spoon. Later, a further operation was 
performed : the periosteum was detached, the cavity of the bone 
enlarged, and the contents scraped out, consisting of granulation 
tissue, fragments of bone, and the yellowish fungi. At the bottom 
of the cavity the fang of the canine tooth was found. No return of 
the growth occurred. 

The first cases of actinomycosis which were observed in America 
were connected with the jaw. In 1884 Dr. Murphy described two 
cases at Chicago. The first was that of a woman aged twenty-eight. 
Two weeks previously she had sxiffered from severe toothache, with 
swelHng in the throat and great pain in swallowing. It disappeared 
after poulticing, but she was again attacked with toothache, and a 
swelling appeared on the angle of the jaw on the left side. The 



ACTINOMYCOSIS. 427 

mouth could not be opened without difficulty ; the tonsil was much 
enlarged, and pus was set 'free on incision. She still suffered with 
toothache, and a small swelling now occurred on the left side of the 
neck below the jaw. She had several carious teeth. The swelling, 
which was about the size of a walnut, was punctured, and a drainage 
tube inserted ; a creamy-looking discharge containing yellow granules 
continued to escape, but the swelhng and induration increased. A 
further operation was decided upon. The carious tooth was removed, 
and a probe passed into the alveolus showed a communication with 
the external wound ; the angle of the jaw was chiselled away, and 
the alveolus scraped out. lodoformed gauze was applied, and the 
case recovered. 

The second case was a man aged eighteen, who had also suffered 
with severe toothache and swelling at the angle of the jaw. On 
examination a carious tooth was noticed. The swelling was well 
marked, and there was fluctuation ; it was as large as a pigeon's 
egg, and situated below the jaw. When punctured, thick creamy 
pus escaped containing the fungi ; the sinus was scraped out, and in 
ten days the wound was healed. Another swelling appeared, and 
this was treated as before, and the case recovered. 

The peculiar feature of these growths is their apparent migra- 
tion. Israel states that in one case a tumour occurred on the alveolar 
process, close to carious teeth, and later was close to the edge of 
the jaw in the sub-maxillary region. From thence it disappeared, 
and a large swelling formed below the hyoid bone, and after this had 
been incised and had healed, an abscess formed above the clavicle. 

Actinomycotic tumours in this region would sometimes appear to 
correspond very closely with wens or clyers in cattle : they may dis- 
charge through the skin, and the opening close, or a fistula result ; 
but they differ, from their tendency to form burrowing abscesses 
instead of recognisable tumours. In this respect they recall chronic 
inflammation rather than the sarcoma-like growths in cattle. 

Cases in which the upper jaw is attacked are not so frequent as 
those in the lower jaw. The progress is usually described as slow, 
and there is a tendency for the deep-seated soft parts to be involved, 
while in the lower jaw there is a tendency for the tumour to come 
to the surface. There may be burrowing suppuration, or small 
tumours, which, after a time, fluctuate and form distinct abscesses. 
These may involve the skin, discharge their contents, and leave 
fistulous openings. 

In other cases the disease has been described as extending from 
the alveolar process to the temporal bone, or the base of the skull. 



428 INFECTIVE DISEASES. 

destroying bones and even reaching the brain ; or the growth may 
descend by the spinal column, implicating the vertebrae, and travel- 
ling and pointing in various directions. 

(II.) Invasion by the Respiratory Tract. — In one recorded case 
the disease existed for seven years, vras localised to the bronchi 
{Bronchitis actinomycotica), and did not extend into the lungs. 
The sputum was examined, and contained the characteristic fungus. 

If the micro-organisms are inhaled they pass into the bronchioles 
and alveoli, and produce proliferation of round cells, which undergo 
fatty degeneration. The resulting patches of peri-bronchitis or 
pneumonia become yellowish - wh ite ; suppuration and hsemorrhage 
from the capillaries follow, and small cavities result, containing pus 
■cells, fat granules, blood, and the fungi. In the neighbourhood of 
the new growth there is compression of the alveoli, and ultimately the 
formation of a dense stratum of connective tissue, separated from the 
cavities by a lining of granulation tissue containing the character- 
istic fungus. The symptoms are usually obscure ; but the sputum 
may contain the fungi, which are often visible to the naked eye. 
The apices of the lungs are not, as a rule, affected. There is con- 
siderable clinical resemblance to chronic phthisis : cough, night- 
sweats, pallor, shortness of breath, and hsemoptysis are symptoms 
common to both. Light may be thrown upon the case by the examina- 
tion of the sputum. The presence of the actinomyces will be positive 
■evidence as to the nature of the disease. The existence of these 
symptoms, with absence of tubercle bacilli, would lead to the 
suspicion of actinomycosis, even failing the discovery of the fungus 
in the sputum. 

In the second stage the symptoms are more characteristic. The 
disease spreads to neighbouring parts, and pleurisy commonly super- 
venes. This extension may involve the peri-pleural tissues. Thus 
the disease may follow the prse-vertebral tissues, descend behind the 
insertion of the diaphragm, and point as an ordinary psoas or 
lumbar abscess ; it may perforate the diaphragm and reach the 
abdominal cavity. Peritonitis or sub-phrenitic abscess may then 
result. In some cases adhesions have formed, and the disease has 
extended to the liver or spleen, or other abdominal organs. The 
disease may also extend forwards in the direction of the anterior 
mediastinum and the pericardium. 

The primary affection of the lung becomes of secondary import- 
ance. Grave symptoms occur, hectic fever, night-sweats, rigors, 
and marked pallor. In the third stage, the disease comes to the 
.surface, either over the chest, or in the neighbourhood of the dorsal 



ACTINOMYCOSIS. 42^ 

or lumbar vertebrae ; a swelling appears of a livid colour, and if 
punctured no fluid escapes, but if allowed to make its own way to the 
surface, the skin gives way, a muco-purulent discharge mixed with 
pieces of the growth escapes, and the fungi can readily be recognised. 

(III.) Invasion of the Digestive Tract. — In a case under Chiari, 
death, with general marasmus, took place at the age of thirty-four, 
after two years' illness. The mucous membrane of the intestines was 
almost completely covered with whitish patches, raised in the centre, 
and covered with yellow and brown granules closely adherent to 
the adjacent tissues. The teeth were carious. 

Small nodules about the ^ze of a pea may be found in the sub- 
mucous tissue, and in the mucous membrane itself. They soften 
and form ulcers with undermined edges, the base reaching the 
muscular layer. They may undergo cicatrisation, but generally 
the disease extends through the peritoneum to the abdominal cavity,- 
and perforates the bladder or the intestines, or makes its way through 
the abdominal wall. Symptoms are either a,bsent or not character- 
istic. The fungus may sometimes be found in the evacuations, or 
by exploratory puncture. 

(IV.) Undetermined. — In, addition there are a number of recorded 
cases presenting very varied symptoms and anatomical relations, in 
which it has not been possible to satisfactorily determine the path 
of infection. Delepine has described a most interesting case of an 
actinomycotic tumour of the brain. 

Manifestations of Actinomycosis in Cattle. 

(I.) In the Digestive system we find the disease attacking : — 

(a) The lips, gums, buccal mucous membrane and palate, and 
appearing as nodules, wart-like growths, or ulcers. The nodules 
and ulceration of the palate were well shown in a specimen sent to- 
the author for examination, under suspicion of being the result of 
severe foot and mouth disease. 

{b) The upper and loioer jaw, where it probably originates in 
carious teeth, and extending and invading the neighbouring cavities 
and sinuses destroys the tissues with which it comes in contact,, 
expanding the bones into thin plates or reducing them to the 
appearance of pumice-stone. 

(c) The tongue, where we see it most commonly in the form of 
nodules or wart-like patches under the mucous membrane, with a 
special tendency to ulcerate, through the irritation of the teeth. These 
nodules may extend into the deep muscles, and often collect in row& 



430 



INFECTIVE DISEASES. 



more or less parallel to the superficial muscular fibres. Complete 
tranverse sections of the tongue, double-stained, readily sho^\' this 
arrangement, even to the naked eye. Induration of the tongue 
results from secondary interstitial glossitis. The author has seen, 
in one case only, a tumour embedded in the substance of the tongue 
about the size of a small Tangierine orange, and more or less isolated 
from any surrounding growth. 

{d) The pharynx, where the disease may occur in the form of 
polypoid growths producing asphyxia. 




Tig. 182. — A Nohfolk Steeh with extensive Actinomycotic Ulceeation of 
THE Skin op the Flank. 



(II.) In the Respiratory system we may meet with the disease in : — 

(«) The nasal cavities, originating primarily there or resulting 
from extension of a growth from the lips, or the pharynx. 

(6) The larynx and trachea, generally in the form of polypoid 
growths, sessile or pedunculated, which arise primarily or occur 
secondarily, by extension from the tissues in the neighbourhood. . 

(c) Tlie lungs, where the cliflFerentiation of the disease is most 
important, as neoplasms in the lungs, especially in the early 
stages, and nodular gi'owths on the pleura, may be mistaken for 
tuberculosis. 

The disease is very rarely found in connection with the 
Xervons system (HI-); ^^ut probably does not so rarely attack the 
Reproductive system [YV.). 



ACTINOMYCOSIS. 431 

(V.) The skin and subcuta/neous tissues are a favourite seat of this 
disease, producing the so-called wens or dyers so commonly seen in the 
fen country. A wen is first recognised as a small tumour, the size 
of a marble or walnut, which increases in size sometimes with great 
rapidity, and breaks down and discharges its muco-purulent contents 
through the inflamed and ulcerated skin ; or it may go on increasing, 
and form a large compact growth, the size of a child's head. These 
growths when excised, hardened, and cut, have a characteristic 
honeycombed appearance, produced by the interlacing bands of 
fibrous tissue, which form a spongy structure, from the interstices 
of which the fungus tufts and thick yellowish pus have for the 
most part dropped out. 

Actinomyces Hominis. — Careful examination of pus from 
a case of actinomycosis in man will reveal to the naked eye little 
yellowish-white or yellow bodies, which a casual observer might 
mistake for grains of iodoform. On collecting some of the discharge 
in a test-tube, and holding it between the light and the eye, the tufts 
of fungi appeared as brownish or greenish-brown grains, embedded 
in a muco-purulent matrix. 

On spreading some of the discharge on a glass slip, the largest 
tufts of the fungus are found to be about the size of a pin's head. 
They have a distinctly sulphur- yellow colour by reflected light, but 
appear of a yellowish or greenish- brown tint by transmitted light. 
With a sewing needle, or a platinum wire flattened at the end into 
a miniature spatula, the grains can be readily picked out of the 
discharge, or taken off the dressing, transferred to a clean slide, and 
gently covered with a cover-glass. Examined with an inch objective, 
they have the appearance of more or less spheroidal masses of a 
pale greenish-yellow colour. On removing the preparation from the 
microscope, and gently pressing down the cover-glass with the finger, 
the grains flatten out like specks of tallow ; and on again examining 
with the same power they are found to have fallen apart into a 
number of irregular and sometimes wedge-shaped fragments of a 
faintly bi-own colour, affording a characteristic appearance. By 
preparing another specimen, and covering it with a cover-glass 
without completely flattening out the grains, the spherical, oblong 
and reniform masses of which the tufts are composed can be 
recognised with a -j-in. objective as rosettes of clubs. By examining 
the peripheral part of a rosette with a, Y^-in.., and especially after, 
pressing the grains into a thin layer, with or without the addition 
of a drop of glycerine, the characteristic clubs are most readily 
demonstrated, and the most varied shapes observed by carefully 



432 INFECTIVE DISEASES. 

examining the form of tlie individual elements. As in tlie bovine 
fungus, every variation in form is found, from single clubs to clubs 
vsrith lateral offshoots, clubs bifid at the extremity, palmate or 
fan-shaped groups, and banana-like bunches. In many cases the 
clubs are divided by transverse fission into two, three, or more 
segments. As a rule, the clubs are irregular in shape, and of about 
equal size, while a few are conspicuous by their length. In other 
parts of the preparation the clubs are replaced by long slender 
forms, which are sometimes transversely divided into a number of 
short links. With suitable illumination many clubs are seen to 
taper off into slender filaments. In addition there are free filaments, 
which are twisted, branched, and sometimes distinctly spirillLform. 
Many of the clubs arei composed of layers differing in their refractive 
power, and many have the appearance of a central channel. There 
are also in the preparation small, highly refractive bodies, fat 
granules, granular detritus, round cells, pus cells, and sometimes 
blood corpuscles. 

The grains differ, as a rule, from those from a bovine source, in 
the absence of that sensation of grittiness so often transmitted to the 
finger when pressing the cover-glass upon them, and in the slightly 
greater tendency of the tufts to retain their compact form. By 
teasing the grains in a drop of water on a slide, . and examining 
the preparation with a ^ or a Jj- objective, the explanation 
of the latter is forthcoming ; for by this process the clubs are 
gradually washed away, and a central core remains, which is com- 
posed entirely of a dense network of filaments. This can readily be 
observed by using a small diaphragm, and it will be found that 
the rosettes of clubs are now replaced by tangled masses, having some 
resemblance to miniature tufts of cotton-wool. These filaments 
constitute the delicate network which is seen in sections stained 
by the method of Gram. This can be readily verified by making 
a cover- glass preparation of the grains, and staining by that method. 
The characters of the fungus can readily be studied by proper 
illumination, without staining. The clubs have a faintly greenish 
tint, and in form and arrangement are quite characteristic and easily 
recognisable. Permanent preparations may be made by mounting 
the fungus in glycerine. 

Description of Stained Specimens. 

The fungus may be stained in alcohoUc solution of eosin in the 
manner to be described for the bovine organism, or in orange-rubin, 



DESCRIPTION OF PLATES XV. AND XVI. 
Actinomyces. 

PLATE XV. 

Fig. 1. — From a preparation of the grains from an actinomycotic abscess in 
a boy ; examined in glycerine. The drawing has been made of a com- 
plete rosette examined by focussing successively the central and peripheral 
portions. Towards the centre the extremities of the clubs are alone 
visible ; they vary in size, and if pressed upon by the cover-glass give the 
appearance of an irregular mosaic. Towards the periphery the clubs are 
seen in profile, and their characteristic form recognised. At one part 
there are several elongated elements, composed of separate links, x 1200. 
Fig. 2. — Different forms of clubs from preparations in which the rosettes have 
been flattened out by gentle pressure on the cover-glass, x 2500. 

(a) Single club, (i) Bifid club, (c) Club giving rise to four 
secondary clubs, (d) Four clubs connected together, recalling 
the form of a bunch of bananas, (e) Mature club with a lateral 
bud. (/) Apparently a further development of the condition 
represented at (e). (^) Club with a lateral bud and transverse 
segmentation. (A) Single club with double tranverse segmenta- 
tion, (i) Club with oblique segmentation, (j) Collection of 
four clubs, one with lateral gemmation, another with oblique 
segmentation. (,A) Clnb with lateral buds on both sides, and 
cut off square at the extremity, (l) Club with a daughter club 
which bears at its extremity two still smaller clubs, (m) Club 
divided by transverse segmentation into four distinct elements. 
(») Elongated club composed of several distinct elements, (o) and 
(p) Clubs with terminal gemmation, (j) Palmate group of clubs, 
(r) Trilobed club, (s) Club with apparently a central channel. 
(t) Filament bearing terminally a highly refractiveoval body. 

PLATE XVL 

Fig. 1. — From a section of a portion of the growth removed from a boy 
during life. The tissue was hardened in alcohol, and cut in celloidin. 
The section was stained by Gram's method and with orange-rubin. x 50. 

Fig. 2. — From the same section. A mass of extremely fine filaments occupies 
the central part of the rosette. Many of the filaments have a, terminal 
enlargement. The marginal part shows a palisade of clubs stained by the 
orange-rubin. x 500. 

Figs. 3 and 4.— From cover-glass preparations of the fungus teased out of the 
new growths produced by inoculation of a calf with pus from a boy 
suffering from pulmonary actinomycosis. Stained by Gram's method and 
orange-rubin. The threads are stained blue and the clubs crimson (a) 
In the younger clubs the thread can be traced into the interior of the 
club (ft). In some of the older clubs the central portion takes a yellowish 
stain, and in others the protoplasm is not continued as a thread, but is 
collected into a spherical or ovoid or pear-shaped mass. In others, again 
irregular grains stained blue are scattered throughout the central portion 
(Fig. 4). X 1200. 

Fig. 5. From a pure-culture on glycerine-agar. (a) branching filaments, (i) a 
mass of entangled filaments. Gram's method, x 1200. 

Fig. 6. — From a similar but older cultivation, (a) a filament with spores 
(b) chains of spores simulating streptococci. Gram's method, x 1200. 



J 



v 




Jt\ 



z 



ii/: 2. 



/\C'l'ISO\l\r,ES 





Pig 5 



ACTINOMYCOSIS HOMINIS 



Fm..vif Bn.'oks.D-r/ &Son..l'-th. 



. ACTINOMYCOSIS. 433- 

and in either case mounted in glycerine. But althougli the fungus- 
can be detected without any staining process, there may sometimes 
be doubtful appearances, and then cover-glass prepai'ations should be- 
made and stained by the method of Gram with eosin. The filaments 
can be readily recognised, and this is of great value, as it forms 
an additional means for the diagnosis of the disease. In combination 
with orange-rubin we have a test that is as characteristic and useful 
as staining for tubercle bacilli. The discharge, scraping from a 
growth, sputum, or the isolated fungus is squeezed between two- 
cover-glasses, which are then slid apart ; they are allowed to dry, 
passed through the flame in the ordinary manner, and then stained 
The cover-glasses can be cleared in clove-oil, the excess of clove-oi] 
being removed by gentle pressure between pieces of blotting-paper, 
and then the preparation can be mounted in balsam and rendered per- 
manent. On examination of these specimens the masses of filaments- 
will be found to be stained blue, and the tissue elements pink. These 
filaments vary very much in extent and character in different pre- 
parations. In some cases there are masses of short threads, which 
are either straight, sinuous, or twisted, and branched. In other- 
parts the field is occupied by very short, straight, or curved and 
sometimes spiral fragments ; in others, again, there are comparatively 
long strands. On examination with a high power, and with careful 
illumination, some filaments will be observed to be momliform,. 
while others are provided with a terminal oval body. There are 
■also free spherical, and oval, bodies stained blue, which represent the- 
spores of the organism. When orange-rubin is used instead of 
eosin, the clubs will be stained and easily recognised. This method 
enables one to determine the exact relation of the threads to the- 
club-shaped bodies ; and this is an interesting point, as it has been 
suggested that the threads are not connected with the clubs, but are 
merely an adventitious micro-organism growing in the track of the 
ray-fungus. The threads are stained blue and the clubs crimson. 
In the younger clubs the protoplasm of the thread can be traced into 
the interior of the club. In some of the older clubs the central 
portion takes a yellowish stain, and in others the protoplasm is not 
continued as a thread, but is collected into a spherical, ovoid, or 
pear-shaped mass. In others again, irregular grains, stained blue, are 
scattered throughout the central portion. The sheath of the thread 
is stained pink ; and the protoplasm, stained blue, fills the sheath, or- 
consists of small spherical or irregular grains, giving a distinctly 
headed appearance. 

The effect of various reagents should be tried upon the isolated 

28 



434 INFECTIVE DISEASES. 

grains. The grains are picked out of the pus and transferred to 
watch-glasses containing strong potash, xylol, and benzol. If 
returned to a slide and covered with a cover-glass, the clubs are 
found unaltered. Water or weak potash washes away the clubs, 
and the filaments become easily distinguished ; ether and strong 
acids have no effect upon them. Oorallin soda, Hanstein's violet, 
and iodine zinc-chloride fail to give any particular reaction. 
Hoffman's blue stains the clubs, but without bringing out any 
structural details which could not be observed in the unstained 
specimens. 

Actinomyces bovis. — The fungus in cattle may in the same 
way be detected with the naked eye in the muco-purulent discharge, 
■or in a scraping from the cut surface of a growth. The tufts of the 
fungus vai'y in size under different circumstances, from that of a grain 
of fine sand to that of a pin's head. If the pus or scraping be spread 
out on a slide and examined against a dark background, the grains 
appear to be white or yellowish-white in colour ; but if examined by 
transmitted light, they appear distinctly brownish. On pressing the 
■cover-glass on the slide the grains readil}^ flatten out, being of a 
soft, tallowy consistency ; or in the process of gently pressing the 
cover-glass on the slide with slight lateral movement, a distinct 
gritty sensation is transmitted to the finger, owing to the presence 
■of calcareous matter. On examination with a low power the 
fungus will be recognised in the form of irregular patches scattered 
over the field, which might readily be regarded as collections of- 
granular debris of a brownish or yellowish-brown colour, but on 
careful examination they are observed to have a more or less 
characteristic appearance. On examining with a higher power, 
spherical, ovoid, or reniform bodies are to be seen, which are 
■either typical rosettes of clubs or granular masses, with here 
and there a club-shaped body at the periphery. Pus cells, round 
cells, fat granules, and minute spherical bodies may also be 
■distinguished. If the grains consist of typical rosettes, and be 
merely covered with the cover-glass, and examined without being 
flattened out between the cover-glass and the slide, they will recall 
to mind, on focussing alternately the centre and the periphery, 
the appearance of the capitulum of a composite flower. The central 
portion appears to consist of spherical forms ; these are the ex- 
tremities of the component elements, and as we focus the edge of 
the rosette these elements are seen laterally, and their characteristic 
club-form is readily distinguished. The central portion may be 
flattened against the cover-glass, and as the individual clubs vary 



DESCRIPTION OF PLATES XVII. AND XVIII. 

Actinomycosis Bovis. 

PLATE XVII. 

Section of an actinomycotic tongue stained by tlie method of 
Gram and witli eosin. 

Fig. 1. — This illustrates the appearance which is usually seen under a low 
power, when a section is stained by Gram's method and with eosin. The 
central portion of a mass of the fungus is either unstained or tinged with 
eosin, while the marginal portion is stained blue. The reverse is seen, as a 
rule, in sections from man ; although under a low power the general appear- 
ance of sections from these two sources is somewhat similar, x 50. 

Fig. 2. — «, &, 0, A, represent the earliest reoognisabla forms of the ray fungus 
in the interior of leucocytes. In e the club-forms can be recognised. In 
f and g there are small stellate groups of clubs, x 500. 

Fig. 3. — ^A part of the section represented in Fig. 1, under a high power. The 
marginal line of blue observed under a low power is now recognised as the 
result of the stain being limited to the peripherally arranged clubs. At 
(a) part of a rosette has undergone calcification ; the clubs are granular, 
and have not retained the stain. At (J) and close to it there are the 
remains of rosettes in which the process of calcification