Cx 4 Libris: Oskar Doty THE CLINICAL. PATHOLOGY OF THE BLOOD OF DOMESTICATED ANIMALS THE MACMILLAN COMPANY NEW YORK «+ BOSTON + CHICAGO - DALLAS ATLANTA + SAN FRANCISCO MACMILLAN & CO., LimitepD LONDON + BOMBAY - CALCUTTA MELBOURNE THE MACMILLAN CO. OF CANADA, Lt. TORONTO @ THE CLINICAL PATHOLOGY OF THE BLOOD OF DOMESTICATED ANIMALS BY SAMUEL HOWARD BURNETT, A.-B., M.S. D. V. M. PROFESSOR IN COMPARATIVE PATHOLOGY, NEW YORK STATE VETERINARY COLLEGE, CORNELL UNIVERSITY, ITHACA, N. Y SECOND EDITION REVISED AND ENLARGED WITH FOUR COLORED PLATES AND TWENTY-THREE FIGURES q 353 ; 28. 3 23. Nem ¥York THE MACMILLAN COMPANY 1917 All rights reserved Copyriant, 1908, By SAMUEL HOWARD BURNETT. Copyrigut, 1917, By THE MACMILLAN COMPANY. Set up andelectrotyped. Published October, 1917. SE bay B81 Bene Norwood jfress: Berwick & Smith Co., Norwood, Mass., U.S.A. Cu JAMES LAW A PIONEER AND LEADER IN VETERINARY EDUCATION IN AMERICA THIS WORK IS AFFECTIONATELY DEDICATED BY ONE OF HIS PUPILS PREFACE TO THE SECOND EDITION The purpose of this book is to furnish a text-book of hematology for the use of students and practitioners of veterinary medicine. It is also intended to render easily accessible for investigators data concerning the blood of the kinds of experimental animals commonly used. Compared with the investigations made on the blood of man, but little has been done on the blood of domesticated animals. Some work, however, has been done. The results obtained are of more value than their mere amount would indicate because we have learned from the studies made that similar pathological processes produce changes in the blood similar to those produced by such processes in man. So the rich results obtained from man are made available to the veterinary practitioner. Free use has been made in preparing this book of the standard works on the blood of man. Those of Ewing, Cabot, Da Costa, v. Limbeck, Naegeli and Grawitz were used most freely. Refer- ences will be found in the text to the source from which material has been taken. A list of references is given at the end of each chapter for the use of advanced students. In the first edition what was reported by different investigators as normal was given without indicating which of these the author considered more reliable. Undergraduate students and most practitioners can hardly be expected to know what to consider as normal when they find considerable variation. In this edition what is considered to be the normal for each species is stated. Tables summarizing the results obtained by the different investi- gations are also given. praca, N.Y. Feb., 1917. S. H. BURNETT. TABLE OF CONTENTS PAGE PE SCERCHMS METS TE AUET ONS 2 cys 4 1S Sihicrs ui Yoven 2 trate a nnkas Gey take meen Leola aes Xill __ DEESIDE 1 i pe ecg olas ie ee OR a oh A XV CHAPTER I METHODS OF EXAMINATION Re SaRTTAVITRH EUS OOEL ce em in, acct ee Caetano heen). 2 eth ahs Se AOE 1 Smiiinescne Ted COrPUSELES. f=. Jackie awk ala Shece hh her dso ate waade wake ene 2 MntrprrbiH AUN GUCUEOEV LES a 65) fois dogo costo nae mw hoe ele ns cee 8 Ono ihe, DGGE PHALESi io. +e 2 lates es hae seed genesis ob cs lRatees 9 Estimation of the amount of hemoglobin. ........................4.. 11 SPER CINE: a CIOL OMIEN SCAIC 58: 02h 4p estcnAaty ohana Ss ooh Ce 11 ower s hemoplobinometer: 2°48 Sachs oe lowe tee cet oe 12 Danes shemoglobimometer.c Ao c..6 sen oo ee eee all) Mleisehl-Miescher hemometer .:2). s.4...¢ 043.0) oe he nee os ns ere ee 15 Peo icminex Anni AIO 10 bats = Cae cy baa LS oc MR late 16 Reis ite Freak COMEMMOMS f-0s Soa sto kd ae ike Wert kesh eke oa ee 16 in-itxed!and(stamed preparations: =. 20.000 oe. ne Se. oe cen soe 17 Ditterentialcountime of leucocytes! mw... vhs ss oe ee eee Zyl sles tHlonsel Gores. may scam yes) hey her, Wi eed emer ctl 22 Test for fat. SR eR lon PE aA RIE peter Oe EE ALAS RA ere 23 The total volume a blood aad Its OXV ECM CAPACliy> 7i. Mares soe: 23 The relative volume of corpuscles and of plasma..................... 24 SUIS ICGLNE PCAVIN eee se 27s, 5. he, ots ae pe es one SG 4 ace ae ie 25 ESE DRETTE AGLERE TG C7 FAG SR ks nce de a eT Ee 25 hestiton specitmmagelutinins or precipitins=., 5. -.... a8 o40.225.5508 26 nacteniolopical- examination of blood: 2.2. <5...) ....06= asemren ot os Soe 26 RELCRCNGCR era eee Pr ie a heen ee MY crue Ey UIA cy. oi ole Dil Partial list of text-books on hematology...) ) 2... 20. 50 ess oe elon 27 CHAPTER II MORPHOLOGY OF THE FORMED ELEMENTS REM EP EOI SG LES cn tir arent NO am eg ee hn aha Lees so Sone « Ge SRI 29 TGC CO CNL CS tale te We ee te aN, EER RI A: 3 Tans DSN cae as 8 Aur, ee Meet ah es 30 Degenerative epee INBIGUCOCYLES a. ewes ae a eae Ree ee ee 35 LD TIS UOA ae Mes gk gee 2 en oe eee ge 36 TUDTOMDOCY LEB ew ae te. ect Ne AMS ea aa aso Stan tah reccbots lsemare es 36 “BLOGS GMS ci Ree Re a rite ae, gn re ea 37 aE RE CMs eye WIR 2 sd Gea ol hs apes hemes 2 Ree oomins AY OMe 38 x TABLE OF CONTENTS CHAPTER III NORMAL BLOOD OF THE SEVERAL DOMESTICATED ANIMALS PAGE TLOVSPS woo. ecihis 0 ok cia Baye, bce iat ioe Ana Een eit eb eee ere 40 Go) ee eee i ey MP Paired Gel etl haat ee tities 0 42 POPE ID seagate 622g oi, Peer Atroe 105 (GHITT GIRS. 6 thos UR Ay RE tee oS ae TL gee re es pene ee eer a 108 SRE CNIS EV! As Gh ots oh ge Nee cco cpa oe cacti eis ale kh deme alee ene 111 TRECISTORTDS eyes c allbcckt eee nC en” oc Sah ant i Regie ROE ee rene Rene NE re 2 112 PNG HINMOLTY COSISMEE SAP aA oy Sista, oye ceees ohm ico buctagtlests 6 ois SEA EE See eae i 113 ESO GTA OILY GOSS ces eacens aie, a9 ccuks says siekotay ucla estan oenaln srneiabe lsat seeteermeeye Nee ee, wale VERDES MUOHSUITATIS ae eters) ote fa aes oe Sia oe theyre arorele aioe Meteo isc me a alee 114 [NXE ESREVINOSEIA. 5 G3 Cg RG Cho CICERO a CIS ACRE oo, OR ae ote acee aeRO peacteens rat & 114 CHAPTER IX INFECTIOUS DISEASES DUE TO PROTOZOA SS DNL SITS Oe pales lle 2 aN es Nr ar ea Pear Shy ey ae o 2 116 pet COME Ea RM UE SG to tM ied 0. 3 ete wuts ecco ea as AMES arte 117 “TPES EISS TNH SS OI en ORS eG EOC Ne REED ORT Ue es een ae 118 RIF EGP ADTOGISIS: 35): su2 2s 2 26 tC ieralthe «we Rae DA HeS weceancin Geae wisn aa aes 123 Re RRR SURES PASHILOBIS Sp: 0 55) = (2°. s Wi SSR ne Bolt ee hacer aes thane ns tlre 129 EULESS UTAG (a Sa SS a Oe CPE ee eR 131 1Bir] OGTR Aaa NIE a Saeco aver hese ROM Oo pee ae OOO Sees lencene citaa ai aetna 131 xii TABLE OF CONTENTS PAGE Hast coast f6ver. dic ©. nicih ay on wo ~ 160 |G 000 (=>. de ee ne eine E ener t Me Ae hee Sa Se oa a neo 161 LIST OF ILLUSTRATIONS PAGE meawn rs. Normal blood of animals. 2.2 .....5. «ied see. cde ede oom oi: 29 eenepul sem eOLpHSGles., 54 bof 2s wee oa seed Lie ig Sain ha ee 30 Piers Til. Blood of the domestie fowls: onc... nce cece cae oe ede 32 Puate IV. Lymphatic and mixed celled leukemia.................... 88 FIGURES IN THE TEXT hic ienuhewkthomayhematocytometer)... 425 0... 206s acne eee 4 Biass2—55 ulmesiot countins chambers: «1.2. ..2..ss.e8ss-e eee ook 4 Bie G er balla wist aehemoplobin Seale, es.) 24's. ac\e occa ea ooh ore mlee eae 11 Haws. Gowers hemorlobinometer. 3) 25 .c6c 605 sees eeee ceca. ck ee 12 easton Hare's hemoglobinometers.... 24 60:6. oles coe os oho 14 Fic. 9. Fleischl-Miescher hemometer.....................-.+-2-8- 15 reel spreading film on glass SUG... 2. ne eile ce ce ba cts eee 17 RirGasiieelood-ofcaseof anthrax, COW 2... acta oss aes ao oes nee 106 Ree EES ANES A IPCI! 2 52 ok, on ee Som Astatte ok Saltese eee 119 Petes caret Eset CR Ioe CANIS "se 42 02 na, S Meals vce RRA ate wt By ores Bae TE 124 Peer l rren ES APICAL EHUEEANIS He e.5--,)-. « o.0!-sic's) co ptitan hn Satie o tee wet ene 132 RiGee mmeh eilertanparvarsd serv Aeore k.. tee so AIO one ie eee 133 Hee GO eelry PASGSOMaGVANSIS. ore che c.chti etree a aoe le mach eee ee 136 Geel peelryp an Osoniss DEUCE! cer gaes fl: 5 aeons oan oc eck crs tetera 137 re) Ubry panosoma Cquiperdunm: 2252). Ger snc Sef ltwrnies os dee o nae f 139 re. 19. Trypanosoma equiperdum, mice: .5....:.....00-5+600ee res 140 Bec 2 OR iry paAnOsOma CQUINUMA ts qee - hohe ina ae ee ee aioe ee 141 Dre 22 ny peOSONIA, GHMOT POM: <.). {p62 ci, sisal ogc oss «a eR 143 areca Wey AMOR Oin GHETELE ie 6a, vals alec tc oa aan eee eee 143 MiG. 2s. sBilsriy mutts in blood; dog: .)\.. eaves 2s yds se esse seer 158 1 ha, — = | —— A an Cee eae INTRODUCTION The blood may be considered as a tissue composed of a cellular part and an intercellular substance. The cellular part comprises the colored and white corpuscles and the plates while the inter- cellular substance consists of the fluid portion. The blood is a peculiar tissue in that it is in intimate relation with practically all the other tissues of the body, bringing substances to them and carrying away other substances. Thus the blood is affected by all the tissues. One might expect that the blood would give a good deal of information as to the state of the activities of the tissues; but at the present time comparatively little use is made of the chemical substances contained in the plasma. In the first place they are present in very small quantities, as an excess is prevented by prompt elimination; and in the second place many of these substances have very complicated structures. In fact the chem- ical make-up of the blood is but little known. Ina clinical examina- tion main reliance is placed on physical and histological rather than chemical methods. The value of an examination of the blood varies in different conditions. In general it may afford sufficient information to make a diagnosis in a few diseases, such as leukemia, Texas fever, surra, anthrax and filariasis. It gives more or less valuable as- sistance in a very large number of conditions, as secondary anemia, sepsis, suppurative processes, intestinal helminthiasis and hemorr- hagic diseases. Finding that the blood is normal is often a great help, as it enables one to differentiate from the diseases in which the blood is not normal. Hodgkin’s disease is diagnosed by a blood examination, yet the blood is normal in the early stages, this serving to distingiush it from leukemia, which produces marked changes in the blood but has otherwise similar symptoms. Besides its value in diagnosis, the blood frequently gives most im- portant indications as to prognosis and treatment and is of value in examinations for soundness. For example, in pneumonia there is ordinarily an increase in the number of leucocytes. If instead XV xvi INTRODUCTION of there being an increase the number is below the normal it is a bad sign. During the course of this disease the reappearance of the eosinophiles is a favorable sign, indicating that the crisis is passed. In an anemia in which the hemoglobin is much lessened while the number of red corpuscles remains nearly normal, the indications for a prompt improvement under the administration of iron are good; while but little improvement is to be expected when the amount of hemoglobin in each corpuscle is normal and practically no improvement when the hemoglobin index is above normal, iron being practically contraindicated in the more severe cases when the blood shows very large red corpuscles each having an increased amount of hemoglobin. In examinations for sound- ness the blood offers valuable aid, as has been pointed out by Moore. When an examination reveals that the blood is not nor- mal a close search for the cause is indicated. The presence of disease, unsuspected it may be by the ordinary means of examina- tion, may be shown by the blood. Though the fact that an animal’s blood is normal is not indicative that the animal is sound, it is an added safeguard, and on the other hand an animal having an abnormal condition of the blood cannot be certified as being sound. In interpreting the results of an examination of the blood it must be kept in mind that this is but one symptom. A diagnosis should be made after duly considering all of the symptoms avail- able. The blood is not supposed to supplant other means of ex- amination, but is to be used with them. In fact it will have a strong tendency to sharpen one’s powers of observation for other symptoms. THE CLINICAL PATHOLOGY OF THE BLOOD OF DOMESTICATED ANIMALS THE CLINICAL PATHOLOGY OF THE BLOOD OF DOMESTICATED ANIMALS CHAPTER I METHODS OF EXAMINATION Ordinarily a clinical examination of the blood consists of count- ing the red and white corpuscles, obtaining the amount of hemo- globin, and making a histological examination of stained speci- mens. Occasionally other information is desired, as the total volume of blood and its oxygen capacity, the relative volume of corpuscles and of plasma, the number of blood plates, the specific gravity, the time of coagulation, the presence of specific agglutinat- ing or precipitating substances, and the presence of parasites. Procuring the blood. Blood may be procured from any part of the body where the circulation is normal. Inflamed or edema- tous areas are especially to be avoided. A cold or bloodless part is unsuitable. If a cold part is warmed, sufficient time should be given to allow the circulation to become normal before procuring blood for examination. In the horse and cow the under side of the tail where free from hair, slightly to one side of the median line to avoid the middle coccygeal artery, is a convenient site, or if preferred, the rump or the side of the neck may be chosen; in smaller animals the lobe of the ear will be found suitable; in the domestic fowl the comb and in the pigeon the under surface of the wing are easily accessible and convenient. Preparation of the site. The part from which blood is to be obtained should be washed with water, in the larger animals where the incision is to be made with a fleam, disinfected, and then dried with alcohol. Vigorous rubbing of the part should be avoided as it produces a transient local change in the blood. In case the neck, or other part in the horse or cow having long hair, is selected the hair may be parted and the skin, exposed in the 1 2 METHODS OF EXAMINATION parting, washed as indicated. It is not ordinarily necessary to shave the site. Puncture. In the larger animals it is advisable to use a fleam or a scalpel for making the puncture as a blood lancet does not yield sufficient blood. In the smaller animals a blood lancet or straight surgeon’s needle is preferable. Whatever instrument is used should be sharp. A dull needle or fleam causes more pain and affords less blood than one that is sharp. In the domestic fowl blood may be readily and easily secured by snipping off the tip of one of the points of the comb. Securing the blood. Before making the puncture whatever apparatus or material is to be used should be ready and within reach so that there may be no unnecessary delay in securing the blood for examination. Slides for making smear preparations, the hemoglobinometer and the pipette for making the dilution should be clean and within reach. The bottle of diluting fluid should be unstoppered. A towel or clean cloth should be at hand. After the puncture is made the first three or four drops of blood should be wiped off. Then freshly exuded blood should be secured for examination, for counting the red corpuscles and leucocytes, for obtaining the hemoglobin value, for making smears for his- tological examination or for other kinds of examination. The more rapidly blood is secured for these several processes the better. Blood quickly changes when exposed to the air. Clotted blood is wholly unsuitable and even before it clots blood undergoes certain changes, some of which simulate changes found in patho- logical conditions. Blood should be obtained as nearly as possible in the condition in which it is in the blood vessels. After sufficient blood has been secured the edges of the wound in the larger ani- mals should be held together for a few seconds until they adhere. On the following day it will require careful search to find the wound. COUNTING THE RED CORPUSCLES The red corpuscles are so numerous in the blood that it is neces- sary to dilute it considerably to be able to count them. The diluting fluid must be of such a nature that it will prevent coagula- tion, will not change the corpuscles and should be of such a specific gravity that the corpuscles will settle, not too rapidly or it will be COUNTING THE RED CORPUSCLES 3 difficult to get an even distribution of them in the counting cham- ber, nor too slowly or one will have to wait too long for them to settle. Toisson’s fluid combines these qualities perhaps the best of the diluting fluids in use. It has an added advantage in that it stains the leucocytes, rendering it possible to count them in the same preparation with the red corpuscles. Toisson’s fluid should be filtered before using as fungus will grow in it producing spores which may be confusing in making a leucocyte count. Hayem’s fluid is also a suitable diluting fluid. Under some conditions Toisson’s fluid is not the best that may be used. Jones has called attention to the counts being lower with Toisson’s fluid than with Hayem’s in certain cases of severe anemia. He found that Toisson’s fluid gave lower counts in cases of pernicious anemia in man and in experimental cases of anemia in rabbits produced by bleeding or by phenylhydrazin. It would be better to use Hayem’s fluid in cases of severe anemia. It is not certain that the formula for Toisson’s fluid is the best for the several species of animals. Formule: Torsson’s FLuip SATE UMPC 8.08 2s heh S-okh eiciaes Soe ISG A oe Sets 8 grams psesemoumrenchilnnlelese = -7.>-\. Se 2 csetus des Ps Fo. So re ee hoe 1 gm G lyon Me minal sates see sate in A Seas Bene ahh pas 30 ce. DISEMIECIWALEKC ee ae Re einai at De ee. ie LOO ce. Niethwertolet-s lr. nr meme sts eel Nei) vena oe .025 gm. Hayem’s Fiuip Wercurierchlorid Gm aps se eerie eke ee eee hee oe eas Oe 0.5 gm. SENT 1 J TAPE Bn oops nn Ae Pee ha ea ae Re ong Tomei 5 gm. SodiurmmchlOnid cece ge ore oe ka. se a ear pene ero» Lona: Dishilledswatermme cee es Aes ee ie sisi Gees ve eae a 200 ce. While several methods of counting the corpuscles have been used, Thoma’s hematocytometer, which is a combination of the older instruments of Malassez, Hayem and Gower, has come to be used as the most reliable. The instrument consists of a counting chamber with an ac- curately polished plano-parallel cover glass and a diluting and mixing pipette. The counting chamber is a thick glass slide on which is cemented a square plate having a circular piece cut out of the center. In this circular space is fastened a small circular t METHODS OF EXAMINATION —— a7 . - pS aFiara) Rih e ei sh Baadlsand edd ST) eet mg oo re eco ta INA B Fig. 1. The Thoma hematocytometer. dise which is thinner than the surrounding plate. The distance from the top of the disc to the under surface of a cover glass placed on the plate is exactly 0.1 millimeter. The upper surface of the disc is ruled as shown in Fig. 2. The central square is ruled with 1 lines ;45 mm. apart giving 400 squares each containing zy5 Sq. mm. HH Fie. 5. Turck. Rulings of Counting Chambers. COUNTING THE RED CORPUSCLES 5 The Zappert-Ewing ruling, the one to use preferably, has besides this central 400 squares additional lines as shown in Fig. 4 giving a ruled space of nine sq. mm. With each instrument are thick and thin cover glasses with plane ground surfaces. The thick covers give more accurate results. The mixing pipette is a cap- illary tube divided into ten equal divisions and terminated by a bulb of 100 times the capacity of the tube. Thus when blood is drawn to the mark 1 on the tube and diluting fluid is drawn to the mark 101 a dilution of 1:100 is obtained. The bulb contains a glass bead to facilitate mixing the blood and diluting fluid. Attached to the pipette is a flexible rubber tube having a mouth- piece. The pipettes are made in a long and a short form. The longer form is preferable as the shorter form is constricted at the point rendering it liable to become clogged and making it very difficult to clean should it become clogged. Method of using. The bottle of diluting fluid should be within reach and should be unstoppered. In using, the point of the pipette is immersed into a freshly exuded drop of blood. With gentle suction blood is drawn up to one of the divisions on the tube, the pipette is withdrawn and the blood on the outside of the tube quickly wiped off. As quickly as possible it is placed in the dilut- ing fluid with the point well below the surface and the diluting fluid drawn in filling the pipette exactly to the mark 101. The blood and diluting fluid are then thoroughly mixed by shaking and rotating the pipette: Care must be taken to draw in fresh blood, the slightest clotting will necessitate cleaning the pipette and repeating the operation. Care must also be taken not to draw in air with the blood or with the diluting fluid. If any air should be drawn in it is well to at once draw in diluting fluid to prevent the blood clotting in the pipette as it is difficult to remove clotted blood. With the blood thoroughly mixed with the diluting fluid it is not necessary to make the count at once. If the pipette is kept in a horizontal position it may be carried some distance. It is better to make the count as soon as possible, though the blood will keep for several hours unchanged in the diluting fluid. Filling the counting chamber. When ready to fill the count- ing chamber, the pipette should be shaken and rotated to get an even distribution of the corpuscles. One minute will usually be found sufficient for this. The diluting fluid in the long arm of the 6 METHODS OF EXAMINATION pipette is then expelled by compressing the rubber tube and twist- ing it. One should never blow in the tube as that would be apt to cause saliva to mix with the diluted blood. After a few drops have been expelled a medium sized drop, obtained by simply compressing the rubber tube, is placed on the center of the counting chamber and the cover glass adjusted. In placing the cover glass care must be taken to avoid including air bubbles. If one side of the cover is placed in position and the other gradually lowered there will be no danger of including air bubbles. The cover glass should be placed as quickly as possible. A delay will result in an uneven distribution of the red corpuscles, the surface of the drop before being covered being rounded the corpuscles fall on the ruled dise from unequal thicknesses of fluid. The drop should be large enough to fill or very nearly fill the central dise without running into the trench or between the cover glass and the plate. The proper size of the drop must be learned by experience. With a little practice one can get the proper sized drop at the first trial. If the cover glass fits closely a play of colors (Newton’s rings) will be seen between the cover glass and the plate. If they are not seen it is because the cover glass is not in place. The layer of blood is consequently more than 0.1 mm. in thickness. Gentle pressure on the corners of the cover glass may facilitate the ap- pearance of the colors, but they must remain after the pressure is removed. The counting chamber and cover glass should be cleaned and another drop adjusted if the fluid runs into the trench or be- tween the cover glass and the plate, if the disc is not well filled with fluid, if air bubbles are included, if the interference colors (New- ton’s rings) are not seen, or if the blood is found to be unevenly distributed. Time is saved by brushing off dust or lint from both counting chamber and cover glass with a small camel’s hair brush before filling the counting chamber. Counting. The counting chamber should be kept level after the drop of blood is placed in it. The count is made on a micro- scope with a level stage. It is convenient to use a x 4 (2 inch) ocular and a five mm. (4 in.) objective or a special four mm. (% in.) objective with a long working distance, N. A. 66. After the counting chamber is placed on the stage one must wait until the corpuscles have settled on the lines. Count the corpuscles in one COUNTING THE RED CORPUSCLES t hundred squares. It is well to count blocks of 25 squares in each corner of the ruled space. Clean the counting chamber and cover glass, place another drop and count the corpuscles in another 100 squares. The pipette should be shaken for a minute and a few drops expelled before placing the second drop in the counting chamber. If the two counts do not agree closely count another 100 squares in another drop. Computation. Divide the number of corpuscles counted by the number of squares counted, divide this by the dilution and multiply this quotient by 4000 as each square represents z555 of a cubic millimeter of blood. The result will be the number of corpuscles per cubic millimeter. N O. Corpuscles counted x dilution x 4000 _ No. of corpuscles per y No. of squares ith For example suppose 1321 corpuscles were counted in 200 squares with blood. diluted .5:100. The computation would be: 1321 x 1000 x 4000 200 x 5 = 5284000 The blood examined has 5,284,000 red corpuscles per cmm. Limit of error. Using such a high dilution and such a small quantity of blood the error is at best rather high. Thoma and Lyon obtained an error of five per cent. in counting 200 corpuscles, two per cent. with 1250 corpuscles and one-half per cent. with 20,000 corpuscles. A variation of 100,000 corpuscles is not or- dinarily important. This is close enough for clinical purposes. Cleaning the apparatus. It is important to clean the ap- paratus as soon as the counts are made. If blood dries in the pipette it may take several hours to remove, while if cleaned promptly, it requires only two or three minutes. The counting chamber is to be cleaned with pure water only. As the plate and disc are cemented to the slide by Canada balsam, alcohol or any- thing that will act on balsam must not be used. The cover glass may be cleaned with water, then with alcohol. For drying the counting chamber and cover glass about the best thing is a linen handkerchief that is practically worn out. A new one is too harsh. Japanese lens paper may be used but it is not so convenient as soft linen. Anything more harsh than lens paper or old soft linen should not be used. The pipette should be cleaned, after expelling cal 8 METHODS OF EXAMINATION the remaining diluted blood, with pure water, drawing in water and expelling it several times, then with alcohol, then with ether, finally forcing air through until it is thoroughly dry. When the pipette is dry the glass bead will roll about freely in the bulb with- out adhering to any part. An aspirating bulb will be found time saving in cleaning the pipette. To draw fluid into the pipette the rubber tube may be attached or the aspirating bulb may be com- pressed and the thumb placed over the valve. Care must be taken not to allow the aspirator to become moistened during cleaning as the last step is to force dry air through the pipette. Even when care is taken to clean the pipette well it will be found necessary at more or less long intervals to fill with strong nitric acid and let it act for several minutes. All trace of the acid must be removed by drawing water through the pipette several times, following the water with alcohol, ether and dry air as ordinarily. COUNTING THE LEUCOCTYES If the counting chamber with the Zappert-Ewing ruling is used the leucocytes may be counted in the same preparation as the red corpuscles. The leucocytes in the entire ruled space, nine sq. mm., are counted. The leucocytes are readily distinguished as they have a bluish tinge in contrast to the straw color of the — red corpuscles. A dilution of 1:100 is more accurate when the leucocytes are to be counted. Repeat the count with a second drop as with the red corpuscles. : Computation. As with the red corpuscles divide the number of leucocytes counted by the volume of blood counted (.9 emm.), and this by the dilution. The count should be repeated with a second drop. The formula for leucocytes counted with a dilution of 1:100 is: No. of leucocytes counted x 10 x 100 ae eas, ——_—— = No. of leucocytes per'emm. When the counting chamber having only the central square millimeter ruled is used, a pipette giving a dilution of 1:10 should be used. As it costs more to get the two pipettes and is less con- venient to use it is recommended to get the counting chamber with the Zappert-Ewing ruling. With the special pipette for leucocytes it is necessary to have COUNTING THE BLOOD PLATES 9 a larger drop of blood than for the red corpuscles. As the fluid will run out of this larger pipette it is necessary to keep it as nearly horizontal as possible. Acetic acid one per cent. (glacial one-third per cent.) is preferable as the diluting fluid. With this fluid the red corpuscles are made transparent, the nuclei of the leucocytes standing out in bold relief. With a dilution of one part of blood to ten of the acetic acid it is necessary to mix the blood and dilut- ing fluid quickly; otherwise the leucocytes will be found in clumps, making an accurate count impossible. The method of adjusting the drop in the counting chamber is the same as for counting red corpuscles. The leucocytes in the entire ruled area of the Thoma’s counting chamber, 400 squares, should be counted. In computing the results multiply the number of leucocytes counted by 10, since- the 400 squares represents .1 emm. of diluted blood, and multiply this product by 10, if the dilution was 1:10, to give the number in undiluted blood. COUNTING THE BLOOD PLATES The blood plates show a marked tendency to undergo dissolu- tion soon after the blood is taken from the blood vessels and to adhere to each other and to foreign substances. Special precau- tions must be taken in enumerating them. Several methods have been used, three of which are given in detail. Pratt’s method. Pratt used the following diluting fluid, which keeps indefinitely unless moulds or bacteria develop: Sodium metaphosphate (Merck)..................00-- 2 grams “S PEICTST OE DUT Ts a a ee a A a Pee SP 0.9 gm. stile d wy aber s esas ca stat of cks sala en a ayer oo 100 ce. The number of erythrocytes is determined in the usual manner with the Thoma hematocytometer. A few cubic centimeters of the diluting fluid are placed in a watch glass. All the glassware used must be perfectly clean. Blood is obtained from a puncture free enough to allow the blood to flow freely. A sterilized platinum loop, as used in bacteriological work, with a diameter of about three mm. is filled with diluting fluid and the center of the loop brought in contact with a fresh drop of blood. There should be three or more parts of fluid to one of blood. A portion of the mixture is at once placed on a slide and covered with a cover glass. 10 METHODS OF EXAMINATION The diluted blood should spread so that the erythrocytes are well separated. It is not necessary to mix the blood and diluting fluid by long stirring. Two preparations should always be made. If the count varies much in the two, other preparations should be made. Examination is made with an oil immersion objective. A square diaphragm in the ocular, easily made of stiff paper, facilitates counting. Both the blood plates and erythrocytes are counted in fields taken at random in different parts of the specimen until 250 to 500 erythrocytes have been seen. This will give the ratio of plates to erythrocytes. The number per emm. is obtained by multiplying the number of erythrocytes per emm. by this ratio. Kemp and Calhoun’s method.— Kemp and Calhoun used the following diluting and fixing fluid: Pormialin: (AOS eee esto oehe ORs Corer tel eater an ee Tee OE 10 ce. Sodium, chioride (1%, sq: soln.) s 2257 osc ee ae ee ee 150 ce. (Color with methyl green or methyl violet if desired). In this method the blood comes in contact with the fixing fluid before touching anything else. The site of puncture is carefully cleaned and dried. Puncture is made, the first drop wiped off and diluting fluid placed on the site of puncture so that the next drop as it emerges flows into the diluting fluid. Mix thoroughly for a few seconds with a clean glass rod then transfer a large drop with the glass rod to the Thoma counting chamber and cover with a thin cover glass. If the corpuscles are fairly evenly distributed, let the chamber rest quietly for about five minutes. Count the red corpuscles and blood plates in about six frames of 16 squares each. This usually gives about 100 blood plates. With a small number of blood plates or with not so even a distribution more than six frames should be counted. The number per cmm. is obtained by multiplying the number of erythrocytes, obtained in the usual manner, by the ratio of blood plates to erythrocytes. Wright and Kinnicutt’s Method.— The diluting fluid is com- posed of two parts of an aqueous solution of “‘brilliant cresyl blue ” (1:300) and three parts of an aqueous solution of potassium eyanid (1:1400). These two solutions must be kept in separate bottles and mixed and filtered immediately before using. The blood is diluted 1:100. The process should be done as rapidly as possible. Count in the ordinary counting chamber, using a high power dry objective. The authors used the thin ESTIMATION OF HEMOGLOBIN 11 cover glass of Zeiss with the central excavation. After the count- ing chamber is filled, it is let stand 10-15 minutes to let the plate- lets settle. The red corpuscles are decolorized and appear as “‘shadows.”’ The nuclei of leucocytes are stained dark blue. The platelets appear as sharply outlined round or oval or elongated lilac colored bodies. After the blood and diluting fluid are mixed precipitate does not form. The count may be made after some hours if necessary. The cresyl blue solution is stable but should be kept on ice to prevent the growth of mould. The cyanid solution should be made up at least every ten days. ESTIMATION OF HEMOGLOBIN There are several instruments for obtaining the amount of hemoglobin, the more important of which are Tallqvist’s, Gower’s, Dare’s and Fleischl-Miescher’s. Tallqvist’s hemoglobin scale. This apparatus consists of fifty leaflets of absorbent paper bound in a booklet with a scale of ten standard tints correspond- ing to the color of blood stains having a hemoglobin value of 10, 20, 30, 40 to 100. In making the test a small piece of the absorb- ent paper is touched to a drop of blood which is allowed to soak in gradually. As soon as the blood has lost its humid gloss and be- fore drying has taken place the stain is placed against a white background and compared with the tints of the standard scale. The tints should be compared by daylight. This is one of the most convenient of the hemoglobinom- eters. The.booklet is of a size convenient to be carried in the pocket, requires only a few seconds to make a test and has no pipettes or other parts to be cleaned. Fia. 6. Tallqvist’s hemoglobin scale. 12 METHODS OF EXAMINATION The error in use does not amount to more than ten per cent. This instrument is sufficiently accurate for ordinary clinical work. Gower’s hemoglobinometer. This instrument consists of two sealed glass tubes containing glycerin tinted with picrocarmine to represent the tint of a one per cent. solution of normal blood. One of these tubes, marked with a white dot, is for use by daylight and the other, marked by a black dot, for use by candle light. Besides these tubes there is a tube of similar size graduated into 140 parts, each of which contains 20 cmm., a capillary pipette marked at 20 cmm. and a block for holding the tubes when making the comparison. Method of using. The blood is drawn into the pipette to the mark 20 cmm., the outside wiped off and the contents expelled into the diluting tube, which should contain a lit- tle distilled water. The inside of the pipette is rinsed out by alternately drawing in and expelling water from the tube. While expelling blood or water from the pipette the point of the pipette should be raised slightly above the surface of the water in the tube to avoid blowing bub- bles. Rinsing the pipette also serves to mix the blood with the water. After rinsing the pipette, water is added gradually to the diluted blood in the tube until it is of the same tint as the standard tube. The blood and water should be mixed by closing the tube with the thumb and inverting it several times. Do not shake the tube and produce bubbles in it as bubbles will render it difficult to read the amount. Comparison is made by placing both the standard tube and the diluted blood in the block and viewing them by reflected light. It is better to hold a paper or other white background behind the tubes and let the light fall Fic. 7. Gower’s hemoglobinometer. ESTIMATION OF HEMOGLOBIN 13 over the shoulder while making the comparison. When they are of the same tint the hemoglobin value is read from the dilut- ing tube, reading at the middle of the meniscus. As the readings on the lower part of the scale are not accurate, it is advisable to use two pipettes of blood for low percentages, dividing the result obtained by two. Gower’s hemoglobinometer is used extensively, though not so extensively as a few years ago. A decided disadvantage is that the tint of the standard solution is not permanent, becoming darker after a time. It does not give the hemoglobin value closer than about five per cent. The instrument is not enough more accurate than Tallqvist’s to make it advisable to use. There are modifications of Gower’s hemoglobinometer in which the standard solution of picrocarmine is replaced by modified hemoglobin, permanent solutions it is claimed. This is an ad- vantage in that the same things are compared since the blood is modified in the same manner as the standard solution. As the same substances are compared any kind of light may be used. In Haldane’s modification the standard tube contains CO hemoglobin. The sample of blood is treated, before making the comparison, with illuminating gas. Some of the blood may easily be lost, bubbles form and illuminating gas is not always available clinically. In Sahli’s modification the standard solution is hemoglobin in glycerin. The blood to be tested is treated with a one-tenth normal HC! solution. Sahli’s standard solution is claimed to be perma- nent; but on standing for some time the hemoglobin sometimes settles and cannot be made a uniform solution again. Dare’s hemoglobinometer.—The instrument consists essen- tially of a capillary pipette and a standard color scale representing the hemoglobin values from 10-120. The pipette is composed of two glass plates, one transparent and the other white which has a depression of measured depth ground in one end so that when the plates are clamped in the holder the depression forms a capillary chamber which fills automatically when either of the three sides is touched to a drop of blood. The standard color scale consists of two prismatic glass plates tinted to give the color of undiluted blood by candle light and arranged in the form of a semicircle, the outer part of which increases in depth of color. The percent- 14 METHODS OF EXAMINATION age scale is etched into the edge of a corresponding semicircle of glass and is placed directly opposite the standard scale. These semicircular plates are fastened to a white disc and are protected by a circular rubber case. The standard is rotated by a milled head placed on the upper part of the case. The speci- men of blood is compared with the standard scale through two small holes horizontally placed and Zz viewed through a camera tube, the eyes of the oper- ator being protected from the light by a shield. Light is afforded by a candle at- tached to the instrument. Method of using. The in- strument is prepared for use by screwing the camera tube into place and rotating the shield so that the two holes through which the blood and standard scale are viewed are uncovered. The candle in its holder is then placed in position. The candle wick should be straight or the candle rotated so that it will illuminate both apertures equally. The pipette is removed from the instrument. It should be scrupulously clean and dry. The pipette is filled by touching it, plain glass uppermost, to a drop of blood. As soon as it is filled it is placed in position. It is not necessary to wipe blood from the edges of the pipette, none should however be on the surface of either of the plates. The instrument is held horizontally and should be pointed toward a dark surface. Comparison is made by rotating the milled head, making rather quick turns. When the tint of the standard scale is made to exactly match that of the blood the hemoglobin value is read from the edge of the instrument, the reading being the one indicated by the beveled edge of the opening. Dare’s instrument gives the amount of hemoglobin with greater accuracy than Tallqvist’s or Gower’s. It has given in my ex- Fig. 8. Dare’s hemoglobinometer. ° ESTIMATION OF HEMOGLOBIN 15 uum a + £2 i Hi eed |) | | | “al i = Tn f] ee Se CC Fic. 9. Fleischl-Miescher hemometer. perience practically the same readings as the more expensive instru- ment of Fleischl-Miescher and is much easier to manipulate and to clean. One hundred per cent. in Dare’s hemoglobinometer represents a mixture of 13.77 grams of hemoglobin diluted with 100 ec. of serum. Fleischl-Miescher hemometer.—This instrument consists of a stand having a stage with a circular opening in which a metallic mixing cell fits. Beneath the stage a standard colored glass wedge with a graduated scale works by means of a rack and pinion. A graduated pipette marked +, ; and 5 serves to dilute the blood, 1:200, 1:300 or 1:400. The mixing cell has a glass bottom. A metal partition which projects slightly above the surface divides the cell into two vertical halves, one for diluted blood, the other for water. A grooved glass disc covers the mixing cell and this is in turn covered by a metal cap having a rectangular opening which serves to cut off from view all but a narrow strip of the colored wedge. Method of using. The blood is procured with the usual pre- 16 METHODS OF EXAMINATION cautions and is drawn into the pipette to the mark +, 4 or 4. Diluting fluid, a filtered one per cent. solution of sodium ear- bonate, is drawn up to the mark as in the hematocytometer. It is well shaken, and the diluting fluid expelled from the long arm of the pipette. Then one compartment of the mixing cell is filled with the diluted blood, the other compartment filled with water, the glass dise slid into place and the metal cap adjusted. The cell is placed in position on the stage so that the part containing water is over the colored wedge. Comparison should be made by the light of a candle or petroleum lamp placed at some distance from the hemometer. A better result will be obtained in match- ing the blood and scale by moving the scale by short quick move- ments. A table accompanies each instrument in which is given the number of grams of hemoglobin corresponding to the reading obtained with that instrument. The Fleischl-Miescher hemom- eter is one of the most accurate of those used for obtaining the amount of hemoglobin. The feature of being able to get the number of grams of hemoglobin is a very desirable one and should be given in all hemoglobinometers. It is more reasonable to learn the amount of Hb, given in grams per 100 ee. of blood, than the percentage of a supposed normal, which is fixed at different values in different makes of hemoglobinometers. HISTOLOGICAL EXAMINATION For histological examination blood is examined fresh and in fixed and stained preparations. Fresh preparations are made by touching the center of a thoroughly clean cover glass to the top of a freshly exuded drop of blood then dropping the cover gently on a clean slide. If a proper sized drop is obtained the blood will spread between the cover and slide in a thin layer. The cover glass should not be pressed down as pressure may rupture or distort the red corpuscles. If it is desired to keep the specimen longer than a few minutes the edges of the cover glass should he sealed with liquid paraffin or castor oil. For fixed and stained preparations the blood should be spread in a thin smear. The method of making smears on glass slides will be found a simple and convenient one. The slides should be HISTOLOGICAL EXAMINATION 17 thoroughly clean. Unused slides may be cleaned in strong soap or “gold dust” solution, well rinsed in water, then placed in al- cohol from which they are wiped and polished. Slides with ground edges are preferred. The edge of a slide is touched to the top of a fresh drop of blood then applied to another slide at an angle of about forty degrees (Fig. 10). As soon as the blood has spread along the line of contact of the two slides, the smearer is drawn Fig. 10. Spreading film on glass slide. along with very gentle pressure slowly leaving a thin smear of blocd on the other slide. The smear should cover one-half or two-thirds of the slide. : per PLATE I NORMAL BLOOD OF ANIMALS CHAPTER i MORPHOLOGY OF THE BED CORPUS Gao eters ete, : : is In the circulating bicwd th ‘man, horse, cow, shew, », clteg. APAIT 2 Lewis) and presunatiy im ether & : e. “_ - eamels. DDE toner isl eroaog ,bookd hua na , | ‘ a * Poisson’ 9 fuig is ota aot herd ‘aaitag booid fuse saloarigtos hast q ; - ~ physiological oa Yeo (staly bodld baa sefsecpon bat + ie ; solution of soda Inca pie ‘iete ody VT es pohtiq book gar *hen canneine nm tha front “artiste atin 0 yoetol Jodgondqrayd rh bon: = ie Ted eo i, pitiate 8 stair A, woo ehwedqaryh : . Be ask pose x ait ia soning euloalova gorwords ae bende aah} oe ote ities aonb. serord Agslounonont axe Te Bie he COPA ies Fost ods ‘esloutloltoat bgutd Varese ln differen! conc lata creates peas syred™ tively large. The mistee'seanebsziquwscing wolsumonos syd OL © oe ia abfegbe pinecone Resiemiaeni ereaeRe ab, naiwods rgelnynocum,syiial IL ~33 “Sapall het aite vic i nue es: aidghn getodl_givclspandgaaryloT ot 2 ’ 5 4h a erent HOS 1b abpunianigronny lod ef a enlating blood im the yo ‘it a'adyin to teslopaontionrrlod +1 a > puscles are elliptical auclenterlanges'sennst wos" slisdgontioot 21 wy i puscles varies vath the dither waheninalaaaal aton jslidqenieedl (Of 3 is for each a ill Le Loupe a -Ciptag Sdn an slulgonieott «if pter TH fu li x aint eo ,bomsigim olidqenwed .21 “ “fide easel, orton disa Jag. Li me! © stain, the chferenne between. tf Cais ran | wo bs se ‘be PP) colored periphers! pari beiteg cess se pe | ae eonditions marked changes in the sizé deive and staring proper ad rer : die Rare + ‘ al ties are BOmetimes shown, Tnstend of being about the e Bixe, a varia ion. TRAY per rin ade di aumiia. Itiste he noted ie os that pie] the SCANS ther. a t pins: ; Heaieramte carnation aS me e Pe ae be ' ry ‘> ‘ . sie, Weuy pervall: sorpransdee aba coped! the otros « called sasepneytin, poslinnitne poe hail ees ; 4 hae. ae _* meat of Bo a of the average ape cain! Se Sas Kemaics od eae “various shapes tray fe, fas, ees» Satie sake Yo +H a te Thee are’ called pedhlories aot the chtu@tiins & Bummer sa pve ay Wes ODNAA Pwd = . Red corpuscles and blood plates, horse, Jenner’s stain. . Red corpuscles and blood plate, cat, Wright’s stain. . Blood plates, cat, Wright’s stain. . Lymphocyte, horse, Wright’s stain. . Lymphocyte, cow, Wright’s stain. . Lymphocyte showing nucleolus, guinea pig, Wright’s stain. . Large mononuclear, horse, Jenner’s stain. . Large mononuclear, cow, Jenner’s stain. . Large mononuclear, guinea pig, Jenner’s stain. . Large mononuclear showing degeneration, guinea pig, Jenner’s stain. . Polymorphonuclears, horse, Wright’s stain. . Polymorphonuclear, cow, Jenner’s stain. . Polymorphonuclear, cat, Wright’s stain. . Eosinophile, cow, Jenner’s stain. . Eosinophile, horse, Jenner’s stain. . Eosinophile, cat, Wright’s stain. . Eosinophile ruptured, cat, Wright’s stain. . Mast cell, horse, Jenner’s stain. . Mast cell, cow, Jenner’s stain. PLATE I Normal blood, camera lucida drawings. x 1200 Large mononuclear, cow, Jenner’s stain. CHAPTER II MORPHOLOGY OF THE FORMED ELEMENTS RED CORPUSCLES (ERYTHROCYTES) In the circulating blood these elements have a cup shape in man, horse, cow, sheep, dog, cat, rabbit, guinea pig (Weidenreich, Lewis) and presumably in other domesticated mammals excepting camels. This shape may be seen in the counting chamber when Toisson’s fluid is used as a diluent, or when blood is examined in physiological salt solution; but better when examined in a 2.5 to 5% solution of sodium metaphosphate freshly prepared. Ordinarily when examined in the fresh condition or in dried preparations the red corpuscles appear as bi-concave discs of a pale straw color, the color being deeper in the peripheral part of the disc and nearly absent in the central part. The size of this central clear area varies in different conditions. In cases of anemia it may be rela- tively large. The red corpuscles in mammals are not nucleated as a rule in the circulation except during the intra-uterine period. Small numbers of nucleated red cells may be found in the cir- culating blood in the young. In the domestic fowl the red cor- puscles are elliptical nucleated cells. The size of the red cor- puscles varies with the different species of animal. The averages for each animal will be found in the tabulated summaries in Chap- ter III. In stained preparations the red corpuscles take the acid stain, the difference between the central clear area and the deeper colored peripheral part being clearly shown. In _ pathological conditions marked changes in the size, shape and staining proper- ties are sometimes shown. Instead of being about the same size, marked variation may appear in cases of anemia. It is to be noted that in the young there is normally a considerable variation in size. Very small corpuscles, about one-half the average size are called microcytes. Corpuscles one-half larger to twice the diameter of the average are called megalocytes. In anemia corpuscles of various shapes may be found, the most usual being pear shaped. These are called poikilocytes and the condition is known as poi- 29 30 MORPHOLOGY OF THE FORMED ELEMENTS kilocytosis. Nucleated red cells (erythroblasts) occur in certain abnormal conditions, as in severe cases of anemia and after con- siderable hemorrhage. In the very young nucleated red cells may be normally found, they may be numerous in the dog and cat. A few nucleated red cells have been found by Sherrington and by Traum in adult dogs and by Sherrington in adult cats. Nucleated red cells of the average size are called normoblasts, those considerably smaller than the average are microblasts, those considerably larger than the average are megaloblasts. Under certain conditions one meets with red corpuscles that take the stain irregularly. They may take the basophile stain in small puncte, the corpuscle presenting a coarsely stippled appearance, which is called punctate basophilia. Sometimes corpuscles appear darker, taking some of the basophile stain diffusely. This condi- tion is known as polychromasia or polychromatophilia. Punctate basophilia and polychromasia have been observed in the circulat- ing blood of young animals that were apparently in perfect health. In adults it is not usual to find these changes present except in cases where there is a rapid formation of red corpuscles, as after severe hemorrhage. Walker regards the red corpuscles that take a basophile tint as being younger forms than those having a greater amount of hemoglobin. LEUCOCYTES In fresh blood the leucocytes are distinguished by being color- less, somewhat refractive bodies of a spherical or irregular form. Some have a rounded nucleus and hyaline or refractive cell bodies. These ordinarily show but little if any ameboid movement. Others have irregularly shaped nuclei and granular cell bodies. The granules in some are minute, showing as dark points; in others the granules are larger, appearing as refractive bodies having a greenish tint. The cells with granules possess active ameboid movement. The several varieties are best distinguished in stained preparations. NORMAL VARIETIES OF LEUCOCYTES Five normal varieties are found in the circulating blood of the domesticated animals and are differentiated in stained prepara- tions by the following characters. PLATE II Red corpuscles, camera lucida drawings. «x 1200 . Polychromasia, guinea pig. e . Punctate basophilia, guinea pig. . Erythroblast, puppy. . Erythroblast, puppy. . Erythroblast, dog. . Erythroblast, dog. . Megaloblasts, one showing punctate basophilia, homo. . Poikilocytosis and polychromasia, homo. . Pernicious anemia, homo, showing megalocytes, microcytes, poikilocytes, 77 scuedablsati and punctata basophilia, Jenner’s stain. CONOR WNH ge a) | — . ‘© } ) Oe ‘vs ’ ae » . IL STAI . sai Por J ri tal —_— Le ’ ; t : y " " ’ ; “ ot estivlere * ua - - = “ & a” a *) > a5 rei > . z : ¢ oe ae : | 7 . “y = iva y i - i NORMAL VARIETIES OF LEUCOCYTES 31 Variety I. Lymphocytes. This variety includes cells usually about the size or smaller than the average red corpuscle. Each has a relatively large nucleus that occupies nearly all of the cell. The nucleus is usually round but may be incurved or show a deep notch or sinus at one side. The cell body usually shows as a nar- row rim about the nucleus. Both nucleus and cell body are coarsely reticular. With careful staining a nucleolus may be seen. The cell body has a strong affinity for basic stains, often staining a deeper blue with Jenner’s stain than the nucleus. With Wright’s stain the cell body has a greenish blue while the nucleus has a dark violet tint. Cells falling in this group have practically the same appearance in the several domesticated animals. Variety II. Large mononuclears (hyaline cells, Kanthack and Hardy; large monocytes, Pappenheim). Cells belonging to this variety are larger than those of Var. I, usually about twice the diameter of the average red corpuscle. The nucleus usually oc- cupies only about one-half of the cell and is situated at one side of the center. Its shape is oval or curved (kidney or horse-shoe shaped). Both nucleus and cell body are finely reticular and stain less deeply than do those of lymphocytes. The cell body is faintly basophile. These cells have much the same appearance in the several species of the domesticated animals. In the guinea pig, however, a large number of cells of this variety contain within the cell body one or more rounded bodies that are in some cases ap- parently clear vacuoles and in other cases appear as homogenous or occasionally reticular bodies (see Plate I, Fig. 11) of a purplish tint with Wright’s or Jenner’s stains. They vary from about one micron in diameter to a body occupying nearly one-half the cell. The true nature of these bodies is not certainly known. Kurloff, who first described them, regarded them as vacuoles containing a secretory product. That they are products of secretion or de- generation has been followed, at least passively, by the majority of those who have written on the subject (Burnett, ’04, Staubli, ’06, Howard, ’07, and Canavan, 712). By some, however (Ledingham, ’06), the bodies are regarded as possibly organisms, parasitic in nature. In the majority of instances there is no difficulty in distinguish- ing the cells belonging to Var. I and Var. II, but a certain number of intermediate forms do occur. In fact one can find all stages 32 MORPHOLOGY OF THE FORMED ELEMENTS between typical lymphocytes with a small amount of strongly basophile, coarsely reticular cytoplasm and typical large mono- nuclears with a much larger amount of faintly basophile, finely reticular cytoplasm. Variety III. Polymorphonuclears (polynuclears, polynuclear neutrophiles, polymorphonuclear neutrophiles, finely granular oxyphile cells). The nucleus in this variety is several lobed, the different lobes being connected by slender portions. Rarely the nucleus consists of several separate parts. In shape the nucleus is polymorphous; it may be twisted, spirally coiled, S-shaped, U-shaped, Z-shaped or elongated. It is usually well stained and is coarsely reticular. The cell body contains many fine granules, so small that they appear as mere points. These granules show a rather weak affinity for acid stains, showing as reddish points with Jenner’s or Wright’s stains. The cell body is usually un- stained. In size these cells vary from the size to about twice the size of red corpuscles. In the domestic fowl the cells belonging to this variety differ strikingly as to the granules from polymorphonuclears in mam- mals. The nucleus varies in shape as in mammals and stains similarly. The cell body contains many large granules, spindle shaped with tapering ends, rod shaped with rounded ends, club shaped or oval, that stain a reddish color with Jenner’s stain or eosin and methylene blue and a dark reddish with Wright’s stain. The tint with Wright’s stain is darker with a little violet than the clear reddish of the eosinophile granules of fowl’s blood. With Ehrlich’s triacid stain the polymorphonuclear granules take a deep reddish purple. The granules vary in size from one to three micra in length by about one micron or less in width. This cell has generally been classed as an eosinophile * but it *It has had different names, more or less descriptive as,— crystalloid eosinophile,”’ “polynuclear leucocyte with eosinophilic rods.” The last name is inexcusable. Polymorphonuclear is used as a descriptive term. The word polymorphonuclear was coined, I believe, as the specific name of a certain kind of leucocyte. At any rate it is now used as a specific term. Polymor- phonuclears are not merely leucocytes with polymorphous nuclei. There would be little difficulty if teachers would have beginning students use Roman numerals or something as little descriptive to designate the several varieties of leucocytes until their distinguishing characters are learned. The names then would be recognized not as descriptive but as specific terms. That this PLATE III Domestic fowl, normal blood, Jenner’s stain, from photographs . Red cells, young cell in center, x 780. 2. Red cells and thrombocytes, x 780. . Lymphocyte, thrombocyte and red cells, x 640. Large mononuclear, x 800. Lymphocyte and polymorphonuclear (ruptured), x 640. . Polymorphonuclear and red cells, x 640. . Eosinophile and red cells, x 640. . Mast cell and red cell, x 640. . Thrombocyte and red cells, x 1280. . Lymphocyte and red cell, x 1280. . Large mononuclear and red cell, x 1280. . Lymphocyte and polymorphonuclear, x 1280. . Polymorphonuclear, x 1560. . Eosinophile, x 1560. . Mast cell, x 1280. . Mast cell (ruptured), x 780. ri , ine bh purple. The x soils vary in size from one to three Af n leneth by about one mucror r teas in width. , eff neraily bean. « Sian sa as an eosinoph ile * bat it © Mierent nae es, more or fess ce meripyt ive m4," Meryntallond ©) le? Jenuclear leucocyte with commophare rode” the lost nme Polvmerpheonricar is ary ‘sh downey re term. The word 3 vue wae coped, ¢ believe, as the apatite came of « certain” A any rate it is mow vew! axa specific erm. Polymor ne jot merey leucoeytes With polymearpbous nuclei, “There x Wieulty if teachers would hate heagnning stadents use Rorean onsmbvine at ite descriptive to designate the several “varieties y «o)- theit distinguishing chameters are learned. The nama ™ eview) wot as ddgeoriptive bat gp byes tarms. That thie ue ae r howls sisson ato an ‘amano, boold Snsryos ap : ce pa sr \peyOLOGY OF THE FORMED RUEMENTE == tee = a ital Inmphocy tes with & stiall kanes of pe, weely reticula eytoplasta and typicsl large rion) pine) jasger amount of faintly basophile, finely” : iy neagyi yar orphonuclears {potynuclears, polynicleat s wrorph mucear new rophates, finel iy grontilar” < “he noche in this variety ois eeverel lobed, the sing connected by aleniet portwana. Rarely the. ceveral separate parts, In shape the. nucteus | ue: if may: be twisted, spiraiy: opiled, Sahaped, wad ie eh nated, TREE AdAally: well : stained and teine many fine granules, wee OS eahitas aifion : yarro for G0) LOAWME Bor £ wih allen’ sed brik latyociehbeivdl! yet You -) 008.«: rrelonndvont onal che Oto, ms a Te! I Bites Baca has otysodqery.I .3 | 7 OJ %, elles ber bas 128 iqucostqaostys q 2 PSIG OWE t Sees » GE" Sia bert bas afi ona iter to the granules: trom PP UBS BSE Ora ss eas nucleds vaties in 028i ,ailso' bor has BPA gins N hody contel@Siallee terhaasnpaiqag ote © ae ga eon f stage mie mst Sib bag ix 1e0loyn deromrglog. pee 77 * wh atl ag : "OBE f5 bas p itor ' | if " : air : hi ne Hive ; a 41, ame it righ ; Ann deren oa (2 “eka eae en 3 if the posinophi enh | = bet sorphoauctear granules take # PATHOLOGICAL VARIETIES OF LEUCOCYTES 33 evidently does not belong to that variety. The domestic fowl has an eosinophile leucocyte with round granules, similar to the eosinophile found in mammalian blood. As stated above the nucleus of the avian polymorph varies much exhibiting the same shapes asin mammals. In affinity for stains the granules resemble the polymorphs rather than the eosinophiles. The cells have ac- tive ameboid movement. In its physiologic properties, this cell resembles the polymorphonuclears. This is the cell found in abundance in purulent exudates and is the one that reacts in acute inflammatory conditions as does the mammalian polymorphonu- clear. Variety IV. Eosinophiles (coarsely granular oxyphile cells). The nucleus is polymorphous, ordinarily being bi- or tri-lobed. The lobes are coarsely reticular and usually stain well. The cell body contains many coarse, strongly acidophile granules which are commonly round in outline though oval, ovate or oblong ones are found. In the cat these granules are ordinarily short rod- shaped with rounded ends. The granules vary in size in the dif- ferent species. In the horse they are very large, generally one to one and one-half micra in diameter. These cells are of about the size of the polymorphonuclears. Variety V. Mast cells (coarsely granular basophile cells, Kanthack and Hardy). In this variety the nucleus usually takes the stain so faintly that it is difficult to make out its shape. It varies in shape from rounded or curved to bi-, tri- or many lobed. The cell body contains many strongly basophile rounded or oval granules that take a deep violet tint with Jenner’s and a royal purple with Wright’s stains. Mast cells are as a rule slightly larger than eosinophiles. PATHOLOGICAL VARIETIES OF LEUCOCYTES Besides the varieties just described which are found in normal circulating blood there are other kinds that are sometimes found in pathological conditions. In some cases of disease severe enough leucocyte is homologous with the mammalian polymorphonuclear was first stated in a paper read before the American Association of Pathologists and Bacteriologists on May 7th, 1907. It was first published in the first edition of this book, March, 1908. 34 MORPHOLOGY OF THE FORMED ELEMENTS to affect the blood forming organs immature forms of leucocytes may pass into the blood stream. In certain cases of leukemia not only immature but even parent forms of leucocytes may be found in the circulating blood. The immature leucocytes occur in the circulating blood only in small numbers except in cases of leukemia. lt is important to recognize them when they do occur as their presence indicates a serious condition, involvement of the blood forming organs. It is sometimes difficult to distinguish immature cells belonging to the myelocytic from those of the lymphocytic series. The granules of the myelocytes usually do not stain well with the ordinary blood stains. Graham recommends the use of his alphanaphthol-pyronin method, by which myelocytic cells are made easier to recognize as the granules are made prominent. Myelocytes are cells whose cell bodies contain fine or coarse granules like those of polymorphonuclears or eosinophiles. The nucleus is rounded or may be indented. According as the granules are fine or coarse the cells are known as finely granular myelocytes (usually simply myelocytes) or eosinophilic myelocytes. The eo- sinophilic myelocyte is found in the circulating blood practically only in cases of mixed celled leukemia. Metamyelocytes or transitional leucocytes are intermediate between myelocytes and polymorphonuclears. The nucleus in- stead of being round or oval as in a myelocyte is bent or twisted. It differs from a polymorphonuclear in that the nucleus does not consist of lobes connected by slender portions or of separate divi- sions. Metamyelocytes and myelocytes are sometimes found in small numbers in the circulating blood in severe cases of infection or intoxication. Large numbers of myelocytes and metamyelocytes are found in the circulating blood in cases of mixed celled leukemia. The leucoblast is a still more primitive type of cell. Its neuclus is like that of a myelocyte, but the cell body is feebly basophile and is without the granules that distinguish the myelocytes, metamyelocytes and polymorphonuclears. Leucoblasts closely resemble large lymphocytes. They are found in cases of acute mixed celled leukemia. So far as I know leucoblasts have not been recognized in the circulating blood of the domesticated animals. Plasma cells (reizungsformen, Tiirk) are non-granular, mono- DEGENERATIVE CHANGES IN LEUCOCYTES 35 nuclear cells usually somewhat smaller than large mononuclears, but are sometimes very large. The round or oval nucleus is rela- tively small and is generally excentric. The cell body is strongly basophile and frequently contains minute vacuoles, giving the cell a honey-comb structure. These cells are apparently de- generative forms of mononuclears and are found in the blood in various inflammatory conditions. DEGENERATIVE CHANGES IN LEUCOCYTES It is important in making a histological examination of blood to note the degenerative changes in the leucocytes as well as in the red corpuscles. Among the more common changes observed are those in the nuclei. Sometimes they appear swollen and stain less deeply; again the lobes of the nuclei are shrunken and have an irregular contour. The nucleus may consist of several separate divisions, each of which stains deeply. The nucleus may show hydropic degeneration. In severe leucocytosis one often finds leucocytes with the cell bodies ruptured, the granules scattered and the nucleus pale. Vacuoles may be found in the cell bodies. Small or larger, rounded, purplish granules with Wright’s or Jenner’s stain may be observed in lymphocytes and in polymorphs as well as in large mononuclears, as was mentioned in describing the latter variety. In acute leucocytosis a diminished number of the granules in polymorphs is often seen. Glycogenic degeneration is observed in certain conditions. It consists in the presence of glycogen granules in the leucocytes, especially the polymorphs. Locke and Cabot have applied the term iodophilia to this condition. A certain amount of extra- cellular glycogen may be found in normal blood. The reaction consists in finding glycogen in leucocytes and in an increased amount as extracellular masses. A positive reaction according to Locke and Cabot signifies a general toxemia such as might be produced by abscess, gangrene, uremia or malaria and has been observed in local and general infection with pyogenic organisms, in toxemia of bacterial origin, in non-bacterial toxemia, e. g., uremia, in disturbances of respiration and in grave anemia both primary and secondary. The reaction (Barnicot) does not run parallel to leucocytosis other than having a common cause; its 36 MORPHOLOGY OF THE FORMED ELEMENTS continued presence in pneumonia after the crisis is further aid to other clinical signs that lead one to suspect delayed resolution or other complications; when accumulations of pus are suspected the absence of the reaction is of very great negative value. BLOOD PLATES The blood plates or platelets in mammals are flattened, color- less, very finely granular bodies, usually about one-third the di- ameter of red corpuscles, that show a marked tendency to collect in clumps. They are very vulnerable, changing quickly when blood is drawn. On Deetjen’s agar the blood plates show ameboid processes. According to Wright, confirmed by Bunting, Ogata and others, the blood plates are formed by detached fragments of certain giant cells, the megakaryocytes (Howell), of the bone mar- row and spleen. There has been a great deal of confusion as to the nature of the blood plates because bodies extruded from red corpuscles and somewhat similar to the appearance of the blood plates may be seen in fresh blood and in films prepared in the ordinary manner. The extruded bodies, however, differ from true blood plates, as pointed out by Sacerdotti, by being not flattened, by being homogenous and by often having a faint hemoglobin tint. The blood plates may be studied in freshly drawn blood diluted with physiological salt solution (0.9% NaCl solution) or preferably 10% sodium metaphosphate solution which prevents the clumping of these and avoids the formation of the extruded bodies from red corpuscles. In fixed and stained smears the blood plates take a bluish tint with eosin and methylene blue; with Wright’s stain the central part of the blood plate shows granules of a reddish purple tint while the outer portion appears homog- enous and has a bluish tint. Thrombocytes (spindelzellen, v. Recklinghausen), Pl. III, Figs. 2 and9, in the blood of the domestic fowl are elliptical, oblong or spindle-shaped cells with an elliptical to broadly oval nucleus. In size the cell has nearly the length and about one-half the width of the average red cell. The nucleus occupies about one-half the length and nearly the entire width of the thrombocyte and is usually situated in the central part of the cell. The cell body is pale and often contains one or more clear vacuoles and occa- BLOOD PLATES 37 sionally one or more compact, rounded, deeply staining (deep purple with Wright’s stain) bodies about the size or somewhat larger than a mast cell granule. These bodies are probably a result of degeneration. The thrombocytes show a marked tend- ency to collect in clumps. In fresh blood and in the less thinly spread parts of films, they collect in masses in which it is difficult to distinguish the outline of individual cells. This indistinctness of cell‘outline and structure shows another property of these cells, that is their vulnerability. They change quickly when taken from the blood vessels, passing through a characteristic series of changes. Both cell body and nucleus become less distinct, the cell body losing its structure first. Finally both become structure- less, appearing in stained preparations merely as a diffusely stained mass, the nucleus being distinguishable by having a slightly deeper stain. Hayem and Goodall call these cells hematoblasts. Their func- tion and properties, however, show them to be similar to the blood plates in mammals. As the thrombocytes are nucleated they cannot be homologous with blood plates. Wright considers them homologous with the megakaryocytes from which the blood plates are derived. For his opinion he gives two reasons, first that the forerunners of the megakaryocytes are circulating cells, as may be seen in embryo guinea pigs, and second that in certain Amphibia the thrombocytes regularly lose their cytoplasm by a pinching off process similar to that which takes place in the for- mation of blood plates. Blood dust (hemokonia). These are minute spheroidal or spindle-shaped bodies of one-fourth to one micron in diameter found in the blood. They were first described by Bizzozero and later by Miiller. These bodies are insoluble in acetic acid, alcohol and ether and are not blackened by osmic acid. Stokes and Wege- farth found them to vary in size in different animals according to the size of the granules of leucocytes. They regard them as extruded granules of polymorphs and eosinophiles. These bodies have not been shown to have any special clinical sig- nificance. 38 MORPHOLOGY OF THE FORMED ELEMENTS REFERENCES 1. Barnicot, J. The iodine reaction in the leucocytes. Journ. of Path. and Bacter., xix, 1906, p. 304. 2. Bizzozero, J. Ueber einen neuen Formbestandtheil des Blutes und dessen Rolle bei der Thrombose und der Blutgerinnung. Arch. f. path. Anat., xc, 1882, S. 261. 3. Buntina, C. H. Blood-platelet and megalokaryeyte reactions in the rabbit. Journ. Exp. Med., xi, 1909, 541. 4. Burnett, 8S. H. A study of the blood of normal guinea pigs. Journ. of Med. Research, xi, 1904, p. 537. 5. Canavan, M. M. The blood cell picture in horse serum anaphalaxis in the guinea pig: note on Kurloff’s inclusion bodies. Journ. Med. Res., xxvii, 1912, 189. 6. Cutten, E. K. A morphological study of the blood of certain fishes and birds, ete. Bul. Johns Hopk. Hosp., xiv, 1903, 352. 7. DeretseN. Untersuchungen iiber die Blutpliittchen. Arch. f. path. Anat., elxiv, 1901, S. 239. 8. Dexuuyzen, M. C. Ueber das blut der Amphibien, Verhandl. d. anat. Gesellsch., 6 ten Versamml., Wien, 1892, 8. 90. 9. DexHuyzen, M. C: Ueber die Thrombocyten (Blutplittchen). Anat. Anz., xix, 1901, 8. 529. 10. Goopaut, A. The numbers, proportions and characters of the red and white blood corpuscles in certain animals. Journ. of Path. and Bacter., xiv, 1910, 195. 11. Granam, G.S. The oxidizing ferment of the myelocyte series of cells and its demonstration by an alphanaphthol-pyronin method. Journ. Med. Res., xxxv, 1916, 231. 12. Grinsera, C. Beitrige zur vergleichenden Morphologie der Leuk- ocyten. Arch. f. path. Anat., elxiii, 1901, 303. 13. Howarp, C. P. The relation of the eosinophile cells of the blood, peri- toneum and tissues to various toxins. Journ. Med. Res., xvii, 1907, 237. 14. Howrtt, W. H. The life history of the formed elements of the blood especially of the red blood corpuscles. Journ. of Morphol., iv, 1890, Dol. 15. KanrHack AND Harpy. The morphology and distribution of the wandering cells of Mammalia. Journ. of Physiol., xvii, 1894, p. 81. 16. Kuruorr.. Cited by Ehrlich und Lazarus, Die Andmie, Nothnagel’s Specielle Pathologie und Therapie, viii, 1898, 8. 56. 17. Lepincuam, J.C. G. On the vacuolated mononuclear cells in the blood of the guinea pig. Lancet, i, 1906, 1675. 18. Lewis, F. T. The shape of mammalian red blood corpuscles. Journ. of Med. Research, x, 1904, p. 513. 19. Locke aNp Caxsor. Iodophilia. Journ. of Med. Research, vii, 1902, p. 25. 20. Masstow, G. Einige Bemerkungen zur Morphologie und Entwicke- lung der Blutelemente. Arch. f. mikr. Anat., li, 1897, 61. REFERENCES 39 21. Mtuter, H. F. Ueber einen bisher nicht beachteten Formbestandtheil des Blutes. Centralbl, f. allge. Path. u. path. Anat., vii, 1896, S. 529. 22. OaaTa. Untersuchungen iiber die Herkunft der Blutplittchen. Beitr. z. path. Anat., lii, 1912, 192. 23. PappENHEIM, A. Atlas der menschlichen Blutzellen. 1905-1912. Gustav Fischer, Jena. * 24. PappeNHEIM, A. Uber verschiedene Typen von Lymphozyten und Monozyten, zum Teil im scheinbar normalen Blut. Fol. Haemat., xii, 1911, 26. 25. PappeNHEIM, A. Clinical examination of the blood and its technic. Translated and adapted by R. Donaldson, 1914. Wm. Wood and Co., New York. 26. v. RECKLINGHAUSEN. Ueber die Erzeugung von rothen Blutkér- perchen. Arch. f. mikr. Anat., ii, 1866, 8. 137. 27. Sacervorri, C. Erythrocyten und Blutplittchen. Anat. Anz., xvii, 1900, S. 249. 28. Srausui, C. Klinische und experimentelle Untersuchungen iiber Trichinosis und iiber die Eosinophilie im allgemeinen. Deutsch. Arch. f. klin. Med., \xxxv, 1906, 312. 29. Strokes AND WrEGEFARTH. The presence in the blood of free granules derived from leucocytes, ete. Bull. Johns Hopk. Hosp., viii, 1897, p. 246. 30. Turx. Klinische Untersuchungen iiber das Verhalten des Blutes bei akuten Infektionskrankheiten. Wien, 1898. 31. Waker, E. L. A comparative study of the red corpuscles of Verte- brates. Journ. of Med. Research, xiii, 1904, p. 61. 32. WeipenrercH, F. Studien iiber das Blut. I Form und Bau der rothen Blutkérperchen. Arch. f. mikr. Anat., Ixi, 1902, S. 459. 33. Wricut, J. H. The origin and nature of blood plates. Bost. Med. and Surg. Journ., cliv, 1906, p. 643; also Virchow’s Arch., 186, 1906, 55. 34. Wricut, J. H. The histogenesis of the blood platelets. Journ. of Morphol., xxi, 1910, 263. CHAPTER III NORMAL BLOOD IN THE SEVERAL SPECIES OF THE DOMESTICATED ANIMALS Different investigators have reported widely varying results of the examination of the blood of normal animals. More examina- | tions have been made of laboratory animals,—dog, rabbit and guinea pig. Consequently what is the normal for these animals is better known than for other domesticated animals. More work is needed to determine the normal for the majority of the domes- ticated animals, especially for the numbers and percentages of the several varieties of leucocytes. For convenience in reference the normal condition of the blood is presented in tabular form for each species—horse, cow, sheep, goat, dog, cat, swine, rabbit, guinea pig and domestic fowl with references to the investigators who have worked with each species. HORSE There is considerable normal variation in the number of red corpuscles in the horse. The normal number for the stallion is from 7 to 10 millions per emm. with an average of 8 to 814 mil- lions; for the gelding 51% to 9 millions with an average of 714 to 8 millions; for the mare 514% to 71% millions with an average of 614 to 7 millions. Perhaps observations on a large number of normal horses will show that the smaller numbers, those below 6 millions, are not of strictly normal animals. The hemoglobin varies fully as much as the red corpuscles. Moore, Haring and Cady give the normal as 85 to 100 with an average of 94. Wetzl found the normal to be from 62 to 80. Sabrazés, Muratet and Durroux give the normal as from 68 to 84. More examinations of the hemoglobin of healthy horses are needed. Prus gives the number of blood plates as 500,000 per emm. The normal number of leucocytes seems to be from 5,000 to 10,000 with the average about 8,000. Widely varying results are reported by different investigators. Wiendieck found differences 40 HORSE 4] in the leucocytes of stallions, geldings and mares. In the stallion he found 8,400 to 11,000 with an average of 9,300 per cmm. In the gelding there were from 6,900 to 9,400 with an average of 8,200 per cmm. In the mare there were from 6,500 to 9,000 with an average of 7,900 per emm. Gasse also found differences in the leucocytes in stallions, geldings and mares. He found the leucocytes in stallions to vary from 8,500 to 9,900, with an average of 9,000 per emm., in the gelding 7,800 to 9,300 with an average of 8,500 and in the mare 5,400 to 8,300 with an average 6,900 per emm. Moore, Haring and Cady obtained much lower counts. They found from 4,400 to 6,800 leucocytes with an average of 5,600 per cmm. Taste I.—SuMMARY OF EXAMINATIONS OF THE BLoop oF NorMAL HorsES BY DIFFERENT INVESTIGATORS Red Sr ee ey ee a | ates per cmm. 11,000 - a o ae 2 Bidault. 6.5-9.2 | 7,200—-8,800 | ——— a a Franke. 9.41 9,000 —_—— -——— ee Gasse. 8.28 ” 8,500 — —— —_—— Gasse. Geo 6,900 ——. — — Gasse. en —— 5 Gulliver. TA 9,500 —_——- — 5.58 Hayem. 6.3 —— —_—— yao Malassez. 7.95 8,500 _——— — - —. Meier. 7.198 12,000 6.5 eS 5.78 Mikrukow. 7.9 5,625 94 —- Moore, Haring and Cady. —_- 7,000 es —. ——_— Nicholas et Cour- mont. Ue 15,000 — — ee Prus. 6.75-8.66| 3,720-11,500| 68-84 1,049 4.5-7.8 | Sabrazés, Mura- tet et Durroux. 7.639 4 10,460 ——. —- —- Storch. 9.395 14,034 oa — Storch. bed —- 1,060 5.8 Sussdorf. (a! —_—— oe — Trasbot. 8.45 — oe — — | Wendelstadt und Bleibtreu. 6.16-8 .69 62-80 a a Wetzl. 5.4-10.3) 6,500-11,000) —-— —— Wiendieck. 1 Stallions, 2 Geldings, * Mares, 4 Adults, * Colts. 42 NORMAL BLOOD IN DOMESTICATED ANIMALS Wide differences occur in the findings of different investigators in the numbers and percentages of the several varieties of leu- cocytes. The results obtained by different investigators are given in the following tables II and III. Tasie IIl.—NumBERS or LevucocyTES AND PERCENTAGES OF THE VARIETIES IN THE BLoop or NormMau Horses Percentage of Varieties Leucocytes per cmm. I II III | IV V Lymph». Large M.| Polymor.} Eosins Mast Author ——-— 11,090 37 57 5 —— | Bidault. —— 34-38 60-65 1-2 —— | Cozette. = 22.5 ao 67 5 —— | Fischer. 7,200-8,800 15-35 |2.5-12 | 60-70 |0.8-5. | 0-0.4 | Franke. 5,400-9,900 |23.7-41.29) 0-3 52-73 |0.5-5.8) 0-0.4 | Gasse. 8,500 30 Bp 63.5 3 —— | Meier. 5,625 30 6 59 4 1 Moore, Haring and Cady. 5,400-10,300 35-45 |1.5-3.5} 50-70 |1.5-4 |0.2-0.7| Wiendieck. TasLe II]—NUMBERS OF THE VARIETIES OF LEUCOCYTES PER CMM. IN THE Bioop or NormMat Horses I II IIL IV Vv Lympho. Large Polymorphs Eosins Mast Author Mon. Cells 1053-2431 | 137-518 | 2414-4107 | 161-364 | 6-125 | Moore, Haring and Cady. 1687 340 3316 228 51 | M. H.C. Average. 1600-4300 78-376 | 3208-7964 26-553 | 0-126 | Wiendieck. 2500-3500 | 150-300 | 4000-5000 | 200-350 | 20-60 | Wiendieck Average. COW The red corpuscles vary from about five to eight millions. The average in this country has been found to be about six millions. Wetzl found an average of seven millions (9 cattle). The hemo- globin ranges from 45 to 85 with an average of 60 to 65. Hibbard and Neal found an average of seven million red corpuscles and 97 per cent hemoglobin in five yearling heifers. COW 43 Considerable variation in the leucocytes is reported by different investigators. The normal seems to be from 5,000 to 10,000. A very great variation in the numbers of the several varieties has been found by Dimock and Thompson in apparently normal cattle. Further investigation of the blood of normal cattle is needed, especially as to the numbers of the several varieties of leucocytes. TaBLe I1V.—SumMMARY OF EXAMINATIONS OF THE BLooD OF NORMAL CATTLE BY DIFFERENT INVESTIGATORS Red See een Sree ee | autos per cmm. 6.2 = — —— ——— | 467.2¢| Bethe: Gal 5,486 59.7 ——. eo Dimock and Thompson. —. —— — 5.95 Gulliver. 7.03 4 8,950 97 —- Hibbard and Neal. 4.2 —— —- —_—-— 6 Mallassez. 6.0 9,730 —— — 5-6 Smith and Kilbourne. 5.0 — — ae + Stéltzing. Gees 7,841 — — Storch. 6.6? 9,367 —= —- ——— Storch. 5.43 8,241 = ——— —-- Storch. AVE 11,614 —— — —— Storch. 8.55 15,739 - oa a Storch. 7.0 65 —- ——- Wetzl. 1 Bulls, 2 oxen, * cows, * young cattle, ° calves. Dimock and Thompson obtained the following numbers and percentages of the several varieties of leucocytes in the blood of normal cattle: Averages Minimum Maximum Lymphocytes....... 2,992 per cmm. 54.2 % 31 % 76 Large mononuclears 86 1.4 0.2 33° Polymorphs........ 1,786 30.5 13 45.8 Hosins ey el eees eee 772 13.15 3.8 26.5 Masticells= 25522. 52 31 0.59 Ost 12 44 NORMAL BLOOD IN DOMESTICATED ANIMALS Refik-Bey gives the normal number of leucocytes for cattle as 7,000-11,000 per cmm., the number of mononuclears (includ- ing lymphocytes) as 4,500-6,500 per emm. (57-84%), the number of polynuclears as 1,500-3,500 per cmm. SHEEP The average number of red corpuscles is given by Woltmann as 8,000,000 and of leucocytes as 8,000 peremm. Wetzl found the red corpuscles in normal sheep to vary from 8,008,000 to 9,272,000 per cmm. and the amount of hemoglobin to vary from 47 to 63. Giltner found higher amounts in two normal males. The red corpuscles were 9,120,000 and 11,500,000, the hemoglobin 85 in each and the leucocytes 5,330 and 10,900 per emm. Woltmann found the following percentages of the several varieties of leu- cocytes. Giltner found widely varying percentages in two males. Each had many more lymphocytes than were found by Wolt- mann. More work is needed on the blood of normal sheep. Taste V.—SuMMARY OF EXAMINATIONS OF THE BLoop oF NORMAL SHEEP BY DIFFERENT INVESTIGATORS Red Corpuscles Leucocytes Hemoglobin| Specific | Size of Red canes per cmm., per cent. | Gravity Corpuscles ashore 9.133 4,140 —— — 3.9-5.9 Bethe. 12.090 es a —- 4.9 Cohnstein. 9.1-11.5 5,300-11,900 85 —- we Giltner. —————— | ————— a —- 4.79 Gulliver. — aa --— 1038 Miintz. 10.472 1 9 420 oe a —— Storch. 11.032 2 10,198 ee a —— | Storch. 1225 7,000 a a Warthin. ——-— —— — 5 Welcker. 8-9 .27 So 47-63 ee Wetzl. 8.0 8.000 ——- — ——— Woltmann. 1 Adults, ? lambs. The percentages of the several varieties of leucocytes of the blood of normal sheep obtained by Woltmann are as follows: DOG 45 Average gmnnOCytese 8s farce u ore pa eiele sen k S Bele a 40-60% 53% aree;mononucleats. a7... -sa 5 40 co ey ics ae ees 3-11 8 12a) haiti eric oinice atine ob odcag ace Meee nao pST 30-55 37 iBlosino piles! ree, etree eee er tats e sete 0.2-8 ver | IVE ASESCOLS mera were rar rey) Sr ae ce eater erate NA ease aba ea 0-2 0.3 GOAT Not enough examinations have been made of goats’ blood to know what the normal is. The red corpuscles are high, the aver- age apparently 9-10 millions. The hemoglobin has not been determined for normal animals. Mohler and Washburn state that they were unable to obtain correct hemoglobin readings on ac- count of the color of the plasma. We have had no difficulty, how- ever, using Dare’s hemoglobinometer. The leucocytes are about 10,000 per emm. More work needs to be done to determine them and the numbers of the several varieties. Taste VI.—SumMary OF EXAMINATIONS OF THE BLoop or NorMAL GOATS BY DIFFERENT INVESTIGATORS Red Cor- a ee | ee ee eae per cmm. ——. ———. ——— 3.94 Gulliver. 19.0 30,000 —— ——— 4.25 u Hayem. 18.0 ——. —— 3.5 Malassez. 9.9 9,200 —— — Mohler and Washburn. 14°52) 12057 ——. —— - —_— Storch. 10.12) S91,358 ——— —---- Storch. 9-10.0 —— —— 1042 4.14 Sussdorf. 16.0 8,000 === Warthin. —- ——- —_—— 4.14 Welcker. 1 Goats, ? kids. DOG The red corpuscles are normally from about 5 to 8 millions with an average of about 6 millions. The hemoglobin has been found to vary from 55 to 104. The average is about 90. Considerable variation in the normal number of leucocytes has been found by different investigators. Six thousand to 12,000 with an average of about 8,000 seems to be the normal for adults. The percentages 46 NORMAL BLOOD IN DOMESTICATED ANIMALS of the several varieties are approximately these:—lymphocytes 11-29, average 20; large mononuclears 3-10, average 6; polymorphs 60-76, average 68; eosins 1-10.5, average 6 and mast cells rare. Taste VII.—SumMmary oF EXAMINATIONS OF THE BLOOD oF NorMAL Docs BY DIFFERENT INVESTIGATORS Red Cor- purer | lager ene et ae aoe per cmm 7.0 ——_—_— Uff ——— —— | Breuer und y. Seiller. 5.9 8,221 90 ——— —— | Burnett and Traum. Gie2 9,526 ~ ——. —— | Busch and Van Bergen. 6,000-10,000 | ——— —— —— |Courmont et Lesieur. ez 19,300 87 — —— | Dawson. 4.75-5.5 | 11,200-31,200 | 75-110 | ——— —— | Goodall. 15,800 —. —— |Goodall, Gul- land and Pa- ton. se -- 7.17 | Gulliver. 6.6 10,000 ae —— 7.2” | Hayem. (hs 21,058 Ss — —— | Hinerfauth. 7.4 11,757 —— —_— ——— | Lyon: ——— —— 6.95 «| Manassein. 7,762 —— —— —— | Nicholas et Cot. 6:11 14.08 — — | Otto. 5.71 a 13272 —— —— .||\Otto: 14,182 —— —— | Pohl. 7:3 8,686 117 1063 —— | Rieder. 6.2 7,440 93 —— | Sabrazés et Muratet. 5.5 9.438 58 1058.8 —— |Sherrington. 4.0-5.6 —— |Stéltzing. —— — 1060 7.3 | Sussdorf. —— | 8,100-15,800 | —— —— | Tallqvist und v. Willebrand. 4.4 as we a 7.3 | Vierordt. = ~ ——— 7.3 | Welcker. 4.8-8 ——— 55-104 —_— —— | Wetzl. 6.4 — - os —— | Worm-Miiller. 6.1 7,000 97 —— | Zenoni. 1 Male, 2 female. CAT 47 Taste VIII—Nuvumsers or LevucocyTES AND PERCENTAGES OF THE VARIETIES IN THE BLoop oF NorMAL Docs Percentage of Varieties Leucoeytes | AAA — ee per cmm. |, i |. ar IV. v. Lympho | Large M | Polymor| Eosins Mast 8,221 19.4 6.3 | 68 6.1 rare | Burnett and Traum. 9,526 | 21 G28 Oded 5.3 rare |Buschand Van Bergen. 6-10,000 ——— | 69 ae Courmont et Lesieur. 19,300 | 22.17 | 4.42 | 64.56 | 8.55 | ——— | Dawson. 19,500 30 63 iG ——— | Goodall. 15,800 | 18.5 Geom OOnS 14.5 | ——— | Goodall, Gulland and Paton. 25-36 75-80 ——— | Kriiger. 7,762 | 26.6 4.2 | 69 0.2 | —— | Nicholas et Cot. 9,699 10 Wists || 7A 3.3 | —— | Nicholas et Dumoulin. 7,440 | 33.12 1.32 | 51.07 | 13.97 | ——— |Sabrazés et Muratet. 9,438 17 75 7.8 Sherrington. 12,409 5-10 | 10-15 | 70-80 | 4-8 -0.5 | Tallqvist und v. Wil- lebrand. 7,000 28 62 10 ——— | Zenoni. CAT There are but few examinations of the blood of normal cats re- ported. The normal number of red corpuscles seems to be from 4.8 to 7.6 millions per emm. with an average of about 6.5 millions. Goodall found the normal to vary from 7.28 to 8.6 millions with an average of 8 millions. The leucocytes range from 7,000 to 19,000 with an average of 13,300. Goodall found higher numbers of leucocytes. He reports the normal as from 15,800 to 38,800 with the average as 18,000. He reports the following percent- ages :—mononuclears 37, polymorphonuclears 54, and eosinophiles 9. 48 NORMAL BLOOD IN DOMESTICATED ANIMALS TaBLe 1X.—SuUMMARY OF EXAMINATIONS OF THE BLoop or NorMAL Cats BY DIFFERENT INVESTIGATORS Red Corpuscles | Leucocytes | Hemoglobin Specific Size of Red ‘AGih Millions per cmm. per cent. Gravity Corpuscles oe per cmm., 6.6 13,331 ——. — |Busch and Van Ber- gen. 7.28-8.6 18,000 65-80 ——_——— —— |Goodall. 13,500 aaa — Goodall, Gulland and Paton. —- wes 5.76 |Gulliver. 9.9 7,200 oe ——-- 6.2 » |Hayem. ae —-—— 5.77 |Manassein. 7.9 14,000 6 5.49 |Mikrukow. 6.8 14,017 ABS 1052.6 Sherrington. 1054 6 Sussdorf. OM 6.5 |Welcker. The percentages of the varieties of leucocytes in the blood of normal cats obtained by Busch and Van Bergen are as follows: Tasmallamononuclears aes eerie 34.38% 4 . 3 /O Dy barce;mononuclearcmress ain eerie reer 4.89 IIIa. Polymorphonuclear without granules......... 54.15 ) 55.5 Illb. Polymorphonuclear with fine granules........ 1.36 i IVa. Polymorphonuclear with large coarse rod- shaped oxyphile granules............... 0.9 IVb. Polymorphonuclear with large medium round t 5.2 oxyphulejpranules rap ani eee 4.35 Vis WMlasticells® Sister, ate earns tee crass tate ee roe 0.035 In making two varieties each of polymorphs and eosins, Busch and Van Bergen have followed Hirschfeld. There does not seem to be sufficient reason for giving the eat more varieties of leucocytes than the other mammals. In other animals than cats one can find polymorphs in which the granules are indistinguishable or barely visible. Very slight differences in the technic of staining have been observed to produce similar differences in the staining of these granules. The great majority of eosins contain rod-shaped granules, though round and oval forms may be found in the cells containing mostly rod-shaped granules. Variations in the shape of the granules are seen in other animals than cats. SWINE 49 SWINE A great deal of variation is reported by different investigators in the blood of normal swine. Some of the variation is probably due to the different age and sex of the animals examined. The normal count of red corpuscles seems to be about 8 millions. It is less for young pigs. The average hemoglobin value is about 85. The leucocytes seem to average about 19,000 with more than half of them lymphocytes. Gititig found normoblasts numerous in some of the four weeks old pigs examined by him. The following tables, X and XI, give the results of examinations by different investigators: TABLE X.—SUMMARY OF EXAMINATIONS OF THE BLOOD oF NORMAL SWINE BY DIFFERENT INVESTIGATORS Hed rae Hemoglo- P 3 eee | ee ce ER te audios per cmm. 4 6.9 7,840 — a 5.28-7.9 «| Bethe. 7.9 19,000 88 —— Giltner. a — as 6K Gulliver. 29,000 2 =, a 6.2 Giitig. 3.28-9.6 19,900 83 1059 — | King and Wilson. 5.4 == = Stéltzing. 8.0 = —— ——— Storch. 4.9} 11,518 = Storch. —- 1050 6 bw Sussdorf. 8.6 —— = Wendelstadt und Bleibtreu. 7.0 os 60 a 4.7 8,000 | Nasal discharge, sears. 14 seth Le? 7.42 12,000 | No clinical symptoms. 15" | Golds) 103; 6.5 10,000 | Lymphatic swellings. 16 | °G. 8 | norm. 6.97 8,444 | Nasal discharge, indur. lymph gland. TUBERCULOSIS 111 TaBLE XX XII.—TueE LevcocyTes IN CASES OF GLANDERS IN Horses (BurR- NETT AND PEARCE). Varieties of Leucocytes Case} Leuco- No. | cytes per Lymphocytes|Large Mon.} Polymorph. | Eosins_ |Mast Cells cmm. % No. eNO: Cr aeNO: % No.|% No. 1 6,640 | 30 1,992} 5.8 385|/61.5 4,084/2. 133] -4 27 2 9,555 |18 WPA)! Be Sn yey. M7 Ah) Bae || ail 10 3 | 20;750 |15. 3,118) 4.5 934) 80. 16,600 ae 4 6,000 | 30 1,800 | 11. 660) 58. 5 35480) 29 54) -3 18 5 9,422 |22. 2,074) 10. 9AD6D. G6 124 12 94) .1 9 6 C00 | 2625" 976 a8. 5938) 55. 4.180) 92 61455 38 7} 12,000 | 14. - 1,680] 4.7 564/79. 9,480)1.3 156] .3 36 eel OOO pi 2 ease Ie SOOl is 3: 450/84. 12,600} .09 14] .16 24 Sie GO OOOhs Miles 1760) 5: SOONS2) 135120) (45) 72)|25 80 9 6,500 |} 24. 1,560] 6.9 449/61. 3,965|6.3 410) .59 38 HOD) 14.000) | 19 = 2'660)) (3.7 S18) 745 103560) 27 98-7 98 vil 8,666 | 23. OOS | ORON eAWG | OSeam a 894)| le 2a OAs 7 61 eo OOF Ore 22250) ort te S50r | Soa2 216250 13 8,000 | 23. 1,840} 5.7 456/68. 5,440)2.2 176) .5 40 14 | 12,000 |} 13. ILo0)|| 4b GSP. CHOON sey (at) 8? 36 15 | 10,000 | 12. 1200} set 340) Sle 200) eon OOl el Onl 16 8,444 123.8 2010] 5.2 439)]67.6 5,708|3.2 270) .2 17 TUBERCULOSIS Much fewer cases of blood examination have been reported for tuberculosis in animals than for glanders. An anemia usually moderate but occasionally very severe is ordinarily found in un- complicated eases of the chronic disease. The Hb suffers relatively greater diminution than the red corpuscles. In man the majority of the cases of chronic phthisis show but little change in the number of red corpuscles and amount of Hb, though the patient may be pale and emaciated. In such cases the normal counts are explained by there being a loss of fluid from the blood which masks the loss of red corpuscles and Hb. This oligemia is due (Ewing) ‘‘to the specific lymphogogic action of the toxins of the tubercle bacillus, by which there is established a continuous excess in the balance of fluids which leave the tissues through the lymphatics.” ‘In the majority of cases of well-advanced phthisis (in man) therefore, approximately normal blood indicates con- 112 SPECIFIC INFECTIOUS DISEASES siderable absorption of the toxins of the tubercle bacillus” (Ewing). In cases of tuberculosis with secondary infection the blood may exhibit the changes found in infection with pyogenic or- ganisms. These, however, are not so common in the domesticated animals as in man. Ordinarily only slight changes in the size and form of the red corpuscles are seen. Poikilocytosis and marked changes in the size of the red corpuscles may be found in cases of severe anemia. Nucleated red cells are as a rule not found. The fibrin is not increased except in cases of secondary infection. The blood plates were found by Webb, Gilbert and Havens to be increased in guinea pigs to 468,000-776,000 with an average of 622,000 per cmm. in eleven cases. Ordinarily the number of leucocytes is diminished with a relative lymphocytosis. In a proportion of generalized cases the leucocytes may be increased, though as a rule there is leucopenia. The proportion of polymorphs is sometimes above the normal. In a dog with generalized disease I found 3,050,000 red corpuscles and 4,100 leucocytes. A differential count of the leucocytes gave 15.6% lymphocytes, seven per cent. large mononuclears, 74.4% polymorphs, 2.8% eosins, and 0.2% mast cells. Courmont and Lesieur found 88% polynuclears in a tuberculous dog. Webb, Gilbert and Havens found the leucocytes increased in tuberculous guinea pigs to 10,000-15,000 with an average of 13,200 per emm. in eleven cases. Moore and Ward examined the blood of tuber- culous fowls and found from 1,010,000—2,600,000 red cells and from 35-70% hemoglobin. The leucocytes appeared to be slightly increased. Ellermann and Bang state that there is ordinarily an increase in the number of leucocytes in tuberculous fowls. The increase is in the polymorphonuclears. Arloing and Courmont found that the serum from tuberculous subjects would agglutinate cultures of tubercle bacilli. Similar results have been obtained by other investigators; but agglutina- tion could not be obtained by several others. At present the agglutination test has not been sufficiently elaborated to be of clinical value. TETANUS In two cases in the horse (Meier) and in three in man (Cabot) a leucocytosis was present. In the horse the red corpuscles and ACTINOMYCOSIS 113 hemoglobin were normal. In the human cases the hemoglobin was moderately reduced. The following table shows the result of the examinations in Meier’s cases: TaBLE XXXIII.—TerTaNus 1n Horses (Meter) @ Red | re | nee Varieties of Leucocytes ss eatahe Pall al Area | RS eh ebb Te how Mare No. 51} 25-X 2138.5} 7.0 90; 12,400 dischargedas : cured 3-XI. 1-XI 8.65 |115) 12,240)25.2] 4.4/68.8] 1.2 Mare No. 52| 3-XII |18 9.47 |110| 24,698] 4-1] 0.9/4.8] | —_ died 14-XIL. Three fatal cases in man treated with antitoxin showed the following (Cabot) : Pee Pe as Pea go kG d Seo 6, 21-VI 11,100 70 23-VI 11,900 0 ai cea a 19,600 ja) [LT Weg anos Ae Baier 18,200 80 ACTINOMYCOSIS In a case of actinomycosis of the jaw in a cow, Dimock and Thompson found 5,443,000 red corpuscles, 58% Hb. and 7,222 leucocytes. Of the leucocytes there were 43.6% lymphocytes, 0.36% large mononuclears, 53.1% polymorphs, 2.27% eosins, and 0.54% mast cells. Ewing found 21,500 leucocytes in a case of pulmonary actinomycosis in man. Bierfreund found marked anemia of chlorotic type with 30-50% hemoglobin in man. Cabot examined the blood of four cases in man and obtained the following counts of leucocytes: IMGseICHeMAIC seit e mee, oe base: 31,700 18-VI 28,400 19-VI 28,200 25-VI autopsy. Nos 2 spulmonainy. meen 4) aie 20,800 Apr. 23,000 Aug. autopsy. IPT ONS fe) 072 Oe 12,500 INGs: 4; pUlMOnaye cise oucvels. cece 12,200 11-VIII 21,000 15-VIII 26,000 17-VIII 114 SPECIFIC INFECTIOUS DISEASES BOTRYOMYCOSIS In two cases of extensive botryomycosis of the skin in horses, reported by Mielke, the red corpuscles were 8 and 8.8 millions and the leucocytes 24,000 and 23,200 per cmm. Mielke states that the red corpuscles were normal. In one case the varieties of leu- cocytes per cmm. were, lymphocytes 5,208, large mononuclears none, polymorphs 18,408 and eosins 384; while in the other case there were, lymphocytes 3,016, large mononuclears 464, poly- morphs 19,720 and no eosins. There were no mast cells in either case. These two cases showed a polynuclear leucocytosis. HERPES TONSURANS Meier reports a case in a gelding five years old, with 8,120,000 red corpuscles and 8,500 leucocytes. Of the leucocytes there were 16.1% lymphocytes, 3.5% large mononuclears, 75% polymorphs, and 5.4% eosins. REFERENCES 1. ARLOING ET CourmontT. Etude sur la recherche et la valeur clinique de l’agglutination du bacille de Koch par le serum sanguin de homme. Con- gres pour l'étude de la tuberculose, 1898, p. 586. 2. Bipauut, C. Recherches sur les leucocytes du sang du cheval et sur certains leucocytoses experimentales. Arch. d. med. exper., xvi, 1904, p. 354. 3. Brerrreunp. Arch. f. Chirurg., xli, cited by Ewing. 4. BurNeTT AND Pearce. A clinical examination of the blood of glandered horses. Proceed. Am. Vet. Med. Assn., 1908, 270. 5. Curisror rT Kifiner. De la présence des bactéries et de la leucocytose concomitante dans les affections farcino-morveuses. Compt. rend. Acad. d. Sciences, Paris, xvii, 1868, p. 1054. 6. CoLEMAN AND Ewrna. A ease of septicemic glanders in the human subject. Journ. of Med. Research, ix, 1903, p. 223. 7. CourmMonT ET LesteuR. La polynucléose de la rage. Journ. d. Physiol. et d. Path. gen., iii, 1901, p. 599. 8. Dimock AND THompson. Clinical examination of the blood of normal cattle. Am. Vet. Rev., xxx, 1906, p. 553. 9. Duvau, GASNE ET GuILLemMor. Observations de morve aigue humaine. Arch. d. med. exper., viii, 1896, p. 361. 10. Kiern. Cited by Ewing. 11. V. Limpeck. Klinisches und Experimentelles tiber die entzundliche Leucocytose. Zeitschr. f. Heilk., x, 1889, S. 392. REFERENCES 115 12. Maccnta. Untersuchungen uber Leucocytose. Il nuovo Ercolani, 1903, p. 468. Abstr. in Jahresbr. Vet. Med., xxiii, 1903, S. 38. 13. Merer, P. Beitrage zur vergleichenden Blutpathologie. Zeit. f. Tiermed., x, 1906, 8S. 1. 14. Mre.xr, Geore. Blutkérperchenzalungen bei Rotz und differential- diagnostisch in Betracht kommenden Krankheiten des Pferdes. Monats. f. prakt. Tierheilk, xxiv, 1913, 1. 15. M’Fapyran, J. A peculiar staining reaction of the blood of animals dead of anthrax. Journ. of Compar. Path. and Therap., xvi, 1903, p. 35. 16. M’Faypean, J. A further note with regard to the staining reaction of anthrax blood with methylene blue. Journ. of Compar. Path. and Therap., xvi, 1903, p. 360. 17. M’Fapyran, J. Preliminary note on the sero-diagnosis of glanders. Journ. of Compar. Path. and Therap., ix, 1896, p. 322. 18. Mixruxow. Ueber die Veranderungen der Zahl, Form und Wider- standsfahigkeit der rothen Blutkérperchen unter dem Einfluss des Rotz- contagiums. Compt. rend. Chark. Vet. Inst., 1891 (abstr. in Jahresbr. d. Vet. Med., xi, 1891, S. 33.) 19. Monier aND WasHpurN. ‘Takosis, a contagious disease of goats. Bull. 45, Bur. Anim. Ind. U.S. Dept. Agr., 1903. 20. Moors, V. A. Infectious leukemia in fowls. 12th and 13th Ann. Rep. Bur. Anim. Ind., U. S. Dept. Agr., 1895-96, p. 185, also Path. of Infect. Dis- eases, 3d edn. 1908, p. 114. 21. Moore, TayLor AND GitTNER. The agglutination method for the diagnosis of glanders. Am. Vet. Rev., xxx, 1906, p. 803. 22. MoorE AND WarD. Avian tuberculosis. Proceed. Am. Vet. Med. Assn., 1903, p. 169. 23. Nonrewicz. Bakteriologische Untersuchungen des Blutes beim Rotz. Arch. f. Vet. Med., 1891 (abstr. in Jahresbr, Vet. Med., xi, 1891, S. 34.). 24. Prus, J. Uber die Wirkung des Malleins auf das Blut und iiber seinen di- agnostischen Werth. Oesterreich. Zeitschr. f. wissen. Tierheilk., vi, 1894, 8. 106. 25. ScHinpELKA, H. Hamometrische Untersuchungen an gesunden und an kranken Pferden. Oesterreich, Zeitschr. f. wissen. Tierheilk., ii, 1888, 8. 119. 26. Scuntrer. Zur diagnostischen Verwerthung der Rotzagglutination. Centralbl. f. Bakter., xxxix, 1905, S. 180. 27. Scuttz unp Miessner. Zur Serodiagnose der Rotzkrankheit. Arch. f. wissen. Tierheilk., xxxi, 1905, 8. 353. 28. Srorcu, A. Untersuchungen tiber den Blutkérperchengehalt des Blutes der landwirtschaften Haussiugetiere. Inaug. Diss., Bern, 1901. 29. Taytor, W. J. A report upon an outbreak of Fowl Typhoid. Journ. Am. Vet. Med. Assn., xlix, 1916, 35. 30. Warp, A. R. Fowl cholera. Bull. 156, Agr. Exp. Sta., Coll. of Agr. Uni. of Cal., 1904. 31. Wess, GitBeERT AND Havens. Blood platelets and tuberculosis. Trans. Natl. Assn. for Study and Prevent. of Tuberc., 10, 1914, 180. 32. Wuerry, W. B. Glanders: its diagnosis and prevention. Bull. 24, Bur. Govt. Lab. Dept. of Interior, Phil. Isl., 1904. CHAPTER IX INFECTIOUS DISEASES DUE TO PROTOZOA SPIROCHETOSIS Launoy and Levy-Bruhl infected fowls intramuscularly with cultures of Spirochaeta gallinarum. Spirochetes appeared in the peripheral blood in 48 hours. ‘The septicemia lasted three days when virulent cultures were used and five days with attenuated cultures. There was an early very intense anemia followed by regeneration of the red cells in the cases that recovered. The greatest anemia they found to be usually at about the fifth day. The following are examples of their experimental cases, showing the changes in the number of red cells, given in millions per emm. The first one (R‘) was infected with strong virus, the second (Z) with weaker. After Infection Before Infection | 2 days | 3 days | 4 days | 5 days | 6 days | 7 days | 9 days Re Sy 2.27 1.96 — — 2.76 2.85 Z 2.89 | 2.54 2.28 == 1.47 1.26 == — The leucocytes were increased, from 35,000 to 40,000, excep- tionally to 50,000. In some fatal cases the leucocytes reached 80,000. The leucocytic formula is reported to vary but slightly in different fowls. There seems to have been an increase in the polymorphs during the period of septicemia followed by an in- crease of lymphocytes. A few myelocytes were observed during the height of the septicemia. The eosinophiles decreased during the septicemia. The following table shows the changes in the leucocytes in the case reported by Launoy and Levy-Bruhl. What they call ‘“polynucléaires pseudo-éosinophiles” and ‘“neo- polynucléaires”’ are evidently polymorphonuclears. 116 RABIES 117 TaBLE XXXIV.—Fowt INrecteD Marca 6 wits 0.2 cc. or A CULTURE OF SPIROCHAETA GALLINARUM INTRAMUSCULARLY (LAUNOY AND LEvy-BRUHL) Condition of infection 0 + +H | tit 0 ee ES --——- — | — — —- Variety of leucocytes ise if 8 9 | 10 11 12 14 17 19 Lymphocyte......... 49 29 22 16 14 34 56.5] 63 59 45 Medium mononuc.....} 14 8.5 il 24 25 35 31 15 18 11 Large mononuc....... ite 1 2 1 5 & 0.5 0 0 0 Polynucl. pseudo eosino| 24 56 24 10 6 2 0 0 15 35 Polynucel. eosino...... 7 S250 ae5 0 1 ff) ft) 1 8 8 Neopolynucliaires..... | 0 0 39 47 48 23 11 20 9 0 Basophiles) 25252: =<: 5 Z 0.5 2 0 1 1 1 1 1 Myelocytes.......... 1 Leucocytes, total no. . .|20,500}21,000 24,000)21,000 24,000/ 21,000) 27,000) 36,000) 32,000) 25,600 RABIES Courmont and Lesieur made a study of rabies in three cases in man and five cases in the dog of natural infection and also experimental cases in dogs, guinea pigs and rabbits. They found the percentage of polymorphonuclears high in all cases, beginning in experimental cases with the appearance of symptoms. The total number of leucocytes was increased in nearly all cases. In a woman 29 years old, 83 days after having been bitten and one hour before death, there were 3,300,000 red corpuscles and 24,800 leucocytes of which six per cent. were lymphocytes, three per cent. large mononuclears, three per cent. intermediate forms, and 88% polymorphs. In a child ‘six years old, not given Pasteur treatment, the percentages of leucocytes were, lymphocytes 13.5, large mononuclears 0.5, intermediate forms two, and polymorphs 84. In a man 49 years old the leucocytes were: 29 hours before death 5,000 with polymorphs 84%. 2D ce “ce ‘e 7,000 ce ins 83%. 8 “ce ce “c 12,000 “c cc 5%. 1 “ce “ “c 21,000 “ “ 85%. In five cases of natural infection in dogs they found the following: Dog No.1 2 hours before death 98% polymorphs. 2 2 oe “ ce 88% ce 3. al 13 iT: bc 93% 6c 4 1 “cc “cc “cc 96% “ee 5 1 “ce “c “c 90% “c 118 INFECTIOUS DISEASES DUE TO PROTOZOA In experimental cases in dogs the total number of leucocytes increased suddenly on the appearance of symptoms, reaching 15,000-19,000, then falling somewhat on the day of death. The percentage of polymorphs exceeded 90% from the time of the appearance of symptoms to death. In experimental cases in rabbits and guinea pigs there was a polynuclear leucocytosis from the time symptoms appeared and an increase in the number of leucocytes in some of the cases. In some there was no increase in the number of leucocytes, in a few there was a leucopenia. TEXAS FEVER The protozoon causing this disease was first observed by Babes (1888) in Roumania in the blood of cattle affected with hemo- globinuria. In 1889, Smith found the organism in the blood of cattle suffering from Texas fever and named it Pyrosoma bige- minum. The generic name has been changed to Babesia. In the acute form of the disease the parasites may be found in the circulating blood during the febrile period. When the fever subsides and the number of red corpuscles has been found greatly reduced, the parasites disappear rapidly from the peripheral blood, the reduction of temperature usually coinciding with the more or less rapid disappearance of the parasites. An occasional parasite may be found for some days or even a week after recovery has set in (Smith and Kilbourne). Kossel and Weber report that the parasites are abundant during the time hemoglobinuria is present. When hemoglobinuria disappears the parasites disappear or are very scarce in the circulating blood. In one case Kossel and Weber found individual large typical pyriform parasites in the blood seven days after the disappearance of hemoglobinuria. In acute cases the parasites appear in fresh blood as two pale, pear-shaped bodies situated within red corpuscles. The broader end of the parasite is rounded, the other end long and tapering. The tapering ends are directed toward each other and are ordinarily close together. Sometimes a thin film of protoplasm may be made out connecting the two tapering ends. The broad ends may occupy various positions as regards each other; sometimes they are rather close together so the pyriform bodies lie nearly parallel, then they may point away from each other, the pyriform bodies TEXAS FEVER 119 forming a straight line. The smaller forms appear as a rule homogenous, the large forms contain a minute spherical body, not over 0.1-0.2 in diameter, which appears darker than the re- mainder of the body of the parasite. In the largest pyriform bodies, a larger rounded or oval body 0.5-1.0u in diameter may be ob- 10 Fic. 12. Babesia bigemina. 1, blood obtained postmortem; 2, before death (Smith and Kilbourne); 3-10, different forms of the parasite (Kossel). served situated in the center of each broad rounded end. The infected red corpuscles show crenated margins and may be darker than the uninfected corpuscles; they have lost their flexibility, retaining the disc form even after uninfected corpuscles have become shrivelled and folded in preparations that have been made 120 INFECTIOUS DISEASES DUE TO PROTOZOA for some time. Besides the pyriform parasites a considerable number of single forms somewhat irregular in outline may be found. These forms undergo ameboid movement. Alkaline methylene blue stains the parasites a blue color leaving the red corpuscles unstained. Methyl violet gives much the same result. The parasites are stained by carbol fuchsin; but this stain also stains the corpuscles deeply. Much better results are obtained by using the Romanowsky method of staining or one of its modi- fications. Chromatin bodies within the parasites are brought out by the use of one of these stains. With the Romanowsky method of staining the smallest forms appear as very small rings about one-eighth the diameter of a corpuscle. The periphery of the ring takes a red color in a greater or less extent, while the re- mainder appears blue. Other very small parasites have a very irregular outline and contain two, sometimes four, chromatin bodies of a red color. In the large double pyriform parasites a red chromatin body is found usually at the broader pole, some- times in the middle (Kossel). Ziemann observed forms with the chromatin bodies at the pointed end. The remainder of the body of the parasite stains blue. The chromatin body is ordinarily rounded. The number of infected corpuscles is usually about one-half to one per cent. of the entire number of red corpuscles. Some- times a long search is necessary to find a parasite. When the number becomes larger, death may be expected within 24 hours. Toward the fatal termination five to 10% infected corpuscles may be found. Very rarely large numbers of parasites may be present and yet the animal recover. Smith and Kilbourne ob- served one such case (No. 49), in which hemoglobinuria was present. Parasites are present in the internal organs in much larger numbers than in the peripheral circulation. The distribu- tion of the parasites is well shown in one of Smith and Kilbourne’s cases, No. 163. This cow was killed when the temperature was 107° F., red corpuscles 2,645,000 per ecmm. Four days before the temperature was normal, the red corpuscles numbered 5,000,000 per cmm. Before she was killed there were two to three per cent. infected corpuscles in the peripheral blood. Examination of the internal organs showed 50% in heart muscle and hyperemic fringes of the omentum, 10-20% in liver and in kidney, five per cent. in id BY fi PS) 4 # te | ee TEXAS FEVER 121 spleen, two to three per cent. in blood of left heart and lung, very few in marrow of sixth rib and in skeletal muscles. At post- mortem examination the corpuscles have a rounded form. Free forms of the parasite were not observed by Smith and Kilbourne in the peripheral blood; but were found in the blood from the heart. In the mild type of the disease the parasites are as a rule in- visible in fresh blood, rarely one may be observed on the edge of a corpuscle as a pale spot about 0.5 in diameter. In prepara- tions stained by alkaline methylene blue, they appear as round coccus like bodies 0.2—0.5u in diameter, situated within red cor- puscles. Ordinarily but one is found in a corpuscle. In many cases division of the parasite was observed into two (Smith and Kil- bourne). This form of the parasite is characteristic of the mild autumnal form of the disease. Smith and Kilbourne found that three groups of animals have this type of the disease, (1) those exposed late in the season (October and November), (2) those that have passed through an acute attack earlier in the season; the second attack or relapse in October or November, (3) those that contract the mild form during or previous to the season of the acute form of the disease. In groups 1 and 2 the disease is mild and may pass unnoticed. Infected corpuscles appear in the blood as the number of red corpuscles begin to diminish and dis- appear as the number of red corpuscles begin to increase. Rarely a corpuscle containing a large pyriform parasite is found. A few animals (group 3) showed infected corpuscles several weeks before fever appeared, the disease changing into the acute type with pyriform parasites instead of those of the coccus form. In the mild type of the disease there may be five to 50% of the red cor- puscles in the circulatory blood infected for a period of one to five weeks. Changes in the blood.—There is a progressive loss of red cor- puscles until an extreme oligocythemia is reached. In some cases as high as 1,000,000 corpuscles have been lost per day. In many cases the count of red corpuscles decreases nearly to 1,000,000 to 2,000,000 per emm. Counts, taken from Smith and Kilbourne, are given of two cases,—one fatal, the other recovering. With the reduction in the number of corpuscles marked changes are found in their size and staining. When the number of corpuscles 122 INFECTIOUS DISEASES DUE TO PROTOZOA TaBLeE XXXV.—Texas Fever (SMITH AND KILBOURNE) Date Steer No. 56 20-IX 22-IX 29-IX 9-X 22-X 25-X 30-X 6-XI 8-XI 13-XI 15-XI 17-XI 21-XI 26-XI 2-XII 11-XII 7-VII 13-VIII 14-VIII 15-VIII 16-VIII 18-VIII 19-VIII Cow No. 95 20-VIII 22-VIII 25-VIII 31% 104. 104. 100. 104. “J Red C : Mill PRO Ne own wo ek, WO Ff 36 .09 or. Parasites negative negative negative negative 10-20% peri- pheral cocci. 10-20% peri- pheral cocci. 10-20% peri- pheral cocci. 20-30% peri- pheral cocci. 30% peripheral cocci. 10% peripheral cocci. 10% peripheral cocci. 1-2% periph- eral cocci. a few periph- eral cocci. negative negative 1% peripheral cocci. 5% peripheral cocci. 20% peripheral cocci. 20% peripheral cocci. 15% peripheral coccl. 10% peripheral cocci. 10% peripheral cocci. 10% peripheral cocci. 144-1% large parasites. 10% peripheral cocci, 44-1% large parasites. 10% peripheral coccl. 144-1% large parasites. 5% large para- sites; 5% peripheral cocci. exposed 8-IX. many macrocytes; some punctated red corpuscles 30% macrocytes; 10-20 % tinted corpuscles. 30 macrocytes; 10-20 % tinted corpuscles. some erythroblasts. 20% macrocytes; 10% punctated; 5% tinted corpuscles. 5% tinted and punctated corpuscles. some macrocytes. some macrocytes; recovery complete so far as ap- pearances go. exposed 4-VII. a few punctated corpuscles. 1% macrocytes; some punctated corpuscles. some punctated corpuscles. many macrocytes; some punctated corpuscles. many macrocytes; some punctated corpuscles. evidently dying; some macrocytes; both tinted and punctated corpus- cles. has fallen to about 3,000,000, enlarged corpuscles (six to eight in diameter), showing punctate basophilia and polychromasia, are found. When the number falls to 2,000,000 erythroblasts appear. The amount of hemoglobin seems not to have been determined. While the rapid destruction of red corpuscles is going on hemo- Hemoglobinuria, however, is globin may appear in the urine. not present in all cases. CANINE PIROPLASMOSIS 123 It has been found by Hibbard and Neal that there are changes present in the blood in mild chronic cases of Texas fever, as in cattle raised within tick infested regions or that have been im- munized by having mild attacks of the disease. The red cor- puscles and hemoglobin are diminished by mild attacks, but increase to normal or nearly so after a time. The leucocytes are increased and remain higher than normal. The increase is mostly in the lymphocytes. Examinations of 41 cattle in the general herd of the Mississippi Agricultural College showed the red cor- puscles 4.5-7.8 millions, average 6.05; the hemoglobin 50-98, average 83; and the leucocytes 4880—26,220, average 12,360. The varieties of leucocytes were: lymphocytes 3,218—24,622, average 9,578; the large mononuclears 0-851, average 155; the poly- morphonuclears 440-5,052, average 1,456; the eosinophiles 60- 3,320, average 947 and the mast cells 0-249, average 59 per cmm. The following table shows the results of examinations of five yearling heifers before treatment and after inoculation with Texas fever organisms. The animals had recovered from three attacks and been passed on as in good condition a month before the second examination (Nov. 18th) was made. TABLE XX X VI.—EXAMINATIONS OF THE BLoop oF FIVE YEARLING HEIFERS - IMMUNIZED TO TEXAS FEVER (HIBBARD AND NEAL). After Treatment Betore. er eS See Treatment | Nov. 18 Dec. 1 Jan. 31 Apr. 14 Red corp. millions 7.3 ee 9.3 72 6.64 Hemoglobin 97 90 86 865.5 88.5 Leucocytes 8,950 24,444 | 22.804 | 23005 | 21,721 Lymphocytes 7,163 20,280 19,003 18,243 15,994 Large mononuclears 171 383 700 858 519 Polymorphs 1,443 2,384 2,310 3,444 _ 2,930 Eosins 253 586 756 1,296 2,215 Mast cells 37 52 —— 87 68 CANINE PIROPLASMOSIS (MALIGNANT JAUNDICE IN THE DOG) The specific cause of this disease, Babesia canis, first described and figured in 1895, by Piana and Galli-Valerio in Italy, is a hematozoan found in the blood of diseased dogs. It occurs mostly 124 INFECTIOUS DISEASES DUE TO PROTOZOA within red corpuscles though parasites are found free in the blood. Parasites were found by Nocard and Motas within 36 hours after inoculation in the most rapid case, usually, however, it was two days before they were found even after intravenous inoculation; after subcutaneous or intermuscular inoculation it was usually Fiaq. 13. Babesia canis (Nuttall and Graham-Smith). five or six days before the parasites were found. Robertson did not find them before the fourth day after inoculation though the blood from a dog three days after inoculation proved to be virulent. Nuttall and Graham-Smith found that the parasites made their appearance in appreciable numbers in the peripheral blood im- mediately before the onset of fever. In one case parasites were first found in films on the sixth day of the disease; in most of their other cases parasites appeared between the eighth and twelfth days after infection. Babesia canis has a striking resemblance to the parasite of Texas fever. Piana and Galli-Valerio from its morphology named it Pyrosoma bigeminum var. canis. Nocard and Motas state that morphologically the hematozoan is identical with that of cattle. bat ct: iil CANINE PIROPLASMOSIS 125 The changes which the parasite undergoes in ameboid movement are best seen during the febrile period. The most varied ameboid forms are found toward the end of the febrile period (Nocard and Motas). The infected corpuscles are larger and paler than other corpuscles. The parasites appear as small rounded bodies having a dark contour and refractive central part. Very soon after the febrile period the parasites lose their ameboid properties, take on a rounded form and remain immobile in the center of the affected corpuscles. At the beginning of the disease only a single parasite is found in each infected corpuscle; later infected corpuscles often contain several parasites, when pyriform parasites may be found, though they are rare (Nocard and Motas). Though the parasites may be found in fresh blood when they are numerous, they are much more readily found in stained preparations. Some one of the modifications of the Romanowsky method is_ preferable. Nuttall and Graham-Smith obtained excellent results using Leish- man’s modification. With this stain the parasite appears as a blue body containing usually a single homogenous chromatin body (karyosome) which stains a bright red. In larger forms the cell - body has frequently a vacuolated or trabecular structure and appears condensed at the periphery. Small spherical forms often appear as rings, resembling young malarial parasites in man. In dividing forms a delicate protoplasmic thread may for some time join the daughter cells. With pyriform parasites the connecting thread is usually at the pointed ends, with other forms the con- necting threads are somewhat irregularly situated. The chro- matin body is usually spherical and is usually centrally situated, though it is sometimes eccentric or peripheral. In dividing forms the chromatin body becomes elongated and separates into two portions, immediately followed by division of the cell body. Both spherical and pyriform parasites undergo division (direct division). In the acute form of the disease several parasites, two to 16, may be found in a corpuscle; in the chronic form it is rare to find several parasites in the same corpuscle. Corpuscles may be found containing one to 16 or even more parasites. In the very beginning of the disease only a single parasite is found in a corpuscle, later the infected corpuscles often contain several. Those containing several parasites, eight to 16 or more, are found more often in blood from the internal organs, brain, lymphatic glands, bone 126 INFECTIOUS DISEASES DUE TO PROTOZOA marrow, etc. The parasites are smaller in the corpuscles contain- ing many. Free parasites are also found, especially in advanced stages of the disease (Robertson). Graham-Smith reports that free forms are seldom seen in the earlier stages of the disease but later become more numerous. On the day of death he found one free parasite to 18 infected corpuscles. The number of infected corpuscles varies considerably. They are more abundant in the acute form, in which they are in con- siderable numbers during and immediately after the febrile period. The following table shows the percentage of infected corpuscles and the duration of the disease in 11 cases of the acute and sub- acute forms of the disease (from Graham-Smith). TaBLE XXXVII.—CaNINE PiropLasmosis (GRAHAM-SMITH). Percentage of Infected Corpuscles Duration of Disease Day Before Death Autopsy Dog VI 6% 4% 13 days VII 4 5 32 days. I 1.4 1 25 days. x 1.4 B 23 days. II 8 2.8 23 days. XI 5 12 17 days. Iil 4 8 22 days. Ix 3 at 24 days. VIII 2 4 47 days. IV a 4 23 days. V t 05 13 days. In dog I, the proportion of infected to non-infected corpuscles varied from .3-.6% from the first appearance until the day before death. In dog II, only .05% were infected five days before death, two days later .8% were infected. In the chronic form the parasites are often scarce. Usually examinations made on successive days will reveal their presence. In some cases it is necessary to inoculate several c.c. of blood from the suspected case into a young dog which will often develop an acute case in which the parasites may easily be found. The parasites are more numerous at autopsy in the capillaries of the internal organs. In the majority of cases Graham-Smith found - CANINE PIROPLASMOSIS 127 parasites in great numbers in the small capillaries of the internal organs, in kidneys 95% of the corpuscles were infected, while in the larger vessels there was a small proportion of infected cor- puscles. Changes in the blood.—A severe, progressive anemia is the chief feature of the disease. From the time of the appearance of the first symptoms the number of red corpuscles decreases slowly and regularly, then at the time of the hemoglobinuric crisis falls suddenly to two millions and below (Nocard). The Hb decreases parallel to the red corpuscles from 12-13%, the normal, to six to four and three and one-half per cent. The greatest reduction found by Wright was from 110-18%; the smallest amount. of Hb 17%. Nocard and Motas give the following as a typical acute case: Day Red Corpuscles 5,240,000 intravenous inoculation 2 c.c. virulent blood. 5,560,000 well. 5,960,000 temperature 40° C, few parasites. 5,240,000 many motile parasites. 2,600,000 35.6° C, hind limbs paralyzed. 2,200,000 33.5°C, worse. found dead. aon fr Wh = Db ee The following are two cases reported by Wright: TasBLeE XXXVIII—Two Casrs or CANINE PIROPLASMOSIS (WRIGHT) Dog III Bed Leu- | Varieties of Leucocytes Day| Temp : SSS SSS SS Corpuscles cocytes| 7; | 17 |tIr/1v | Vv 15 | 104.2 | 5,050,000 20,000 | 18 | 10 | 70 | 2 16 | 104.3 18,570 17 | 104.2 | 4,800,000 12,500 18 | 103.3 | 3,800,000 13,000 19 4,800,000 10,000 20 | 105.7 | 4,400,000 11,000 parasites found, Ze ed OD: 1,300,000 9,900 22) 104.3 | 1,850,000 17 10,000 | 15 | 11 | 74 128 INFECTIOUS DISEASES DUE TO PROTOZOA Dog XI 6} 103. 4,900,000 | 100 28 70 | 2 7 | 106.5 10 | 103. 4,000,000 | 70 | 4,500 14 |} 101.8 | 1,800,000 | 32 | 50,000 parasites found. 16 | 101.4 | 1,400,000 26 | 59,000 17 | 100.8 950,000 18 | 49,000 23 76 1 The blood is pale as if diluted with water. Coagulation is slower than in normal blood. The serum is tinted with Hb. The red corpuscles present considerable changes, some are larger (from one-third to two-thirds) and paler than normal; some are smaller than normal. At the time parasites appeared erythro- blasts were also found present (Wright). Erythroblasts in some cases are present in large numbers. The leucocytes are usually much increased. Nocard found as high as 40,000. Wright found nearly 52,000 in one and 60,000 in a second case. In one dog Wright found that the leucocytes dropped gradually from 20,000 on the 15th day to 10,000 on the 22d day. The increase, according to Nocard and Motas, is nearly always in the polymorphonuclears. In Wright’s case both poly- morphonuclears and mononuclears were increased. In the chronic form of the disease there is marked anemia, the red corpuscles decreasing to about two millions. In one of Nocard and Motas’ cases they fell to 1,200,000. The return to normal is very slow. They are scarcely returned to normal before two or three months (Nocard and Motas). As recovery progresses the number of red corpuscles increases; erythroblasts become rare. The Hb suffers much less than the red corpuscles. In one of Nocard and Motas’ cases with only 2,760,000 red corpuscles there was 914% Hb. Thechanges in the size and staining of red corpuscles are more marked than in the acute form. Some corpuscles are two or three times the normal diameter and stain less deeply. Many erythroblasts are present at the beginning of the decrease in red corpuscles. The number of leucocytes is increased, ordinarily from 15-30,000. Nocard and Motas record one case where they reached 54,000. The leucocytosis involves polynuclears and mononuclears equally. EQUINE PIROPLASMOSIS 129 Phagocytosis of infected red corpuscles is frequently observed in the days following the febrile period. The phagocytes are exclu- sively mononuclears. _ The following is a case, dog No. 61, given by Nocard and Motas as typical of the chronic form of the disease: TABLE XX XIX.—CuHrRoniIc CANINE Prropuasmosis (NocarD AND Moras). Red Cor- | Day ices Leucocytes 1 5,840,000 temperature 38.7° C. , few parasites found. 33 more parasites. 4 numerous motile parasites. 6 4,040,000 few parasites; mucosz pale. 8 | 2,820,000 very few parasites; profound anemia. 10 1,520,000 54,000 | erythroblasts numerous. 15 1,200,000 10,000 ‘| no parasites found. 18 | 2,120,000 slight improvement; no parasites. 20 =| 2,480,000 few erythroblasts; distinct improvement; no parasites found. 25 | 4,380,000 improving rapidly; mucose pink; no para- sites found. ae 5,100,000 one parasite found. dog considered cured. EQUINE PIROPLASMOSIS; EQUINE MALARIA; SOUTH AFRICAN HORSE SICKNESS The specifie cause, Babesia equi, discovered by Guglielmi, closely resembles Babesia bigemina. The parasites are found without difficulty in the peripheral blood during the febrile period but disappear later. They are transmitted as shown by Theiler by Rhipicephalus evertsi. The parasites appear as small spherical or elongated, oval or rarely pyriform bodies and are nearly always within red corpuscles. It is not common to find them free in the plasma. The parasites are from .56-2)4mu in diameter, the most common being 1-1)p. Stained by Romanowsky’s or Laveran’s method the cell body takes a bluish tint, the karysome a red-violet color. The following forms of the parasite are described by Bowhill: (1) large and small 130 INFECTIOUS DISEASES DUE TO PROTOZOA spherical forms, karysome situated at edge of parasite; (2) large and small pyriform parasites single and in pairs; (3) large and small rod-like bodies, some of them extending across the entire diameter of the corpuscle, sometimes in pairs; (4) rosette form, consisting of four bodies connected in center by fine threads, each body usually containing a karysome at its extremity; (5) flagellate forms.* Laveran found multiplying forms more common in the spleen than in peripheral blood. Multiplication is by direct division, usually into two, sometimes into four bodies. The karysome elongates, then divides into two parts; these separate, followed by division of the cell body. The two parasites often divide giving four, within the red corpuscle. Sometimes the karysome divides into four parts before the protoplasm divides. The disposition in fours is one of the most striking morphological characters of Babesia equi (Laveran). The number of parasites in the peripheral blood varies a great deal. They are present during febrile stages but disappear after the fall of temperature. During the stage of high fever they are numerous. Theiler has found them in unstained blood as abundant as a parasite to five to 40 red corpuscles. Williams states that the number of parasites in the peripheral blood is proportional to the severity of the disease and that from one to 10% or up to 30% of the corpuscles may be infected. Baruchello and Mori found 50-60% of the red corpuscles infected in some cases; they state that the parasites are found most easily during the early stages of the disease. The “rosette” form is present in varying numbers. Sometimes the parasites are so scarce as to be found only after a long search. After the administration of quinine the rosettes are very scarce (Theiler). With the fall of temperature the number of parasites decreases from day to day; by the time the temperature has fallen they can be found only with difficulty. Changes in the blood.—The disease is characterized by a progressive anemia. In the beginning of the disease the infected red corpuscles ordinarily show no microscopical change; they are * Nuttall and Graham-Smith observed dumb-bell forms and flagella like processes while studying canine piroplasmosis. They found that these forms and processes were due to overheating the red corpuscles and were able to produce similar forms by overheating films of normal blood. BABESIA MUTANS, THEILER 131 infrequently larger than the non-infected corpuscles. Later when the anemia is marked very many relatively large, pale corpuscles are found (Theiler). Williams found variation in the size and shape of red corpuscles, (megalocytes, microcytes, poikilocytes) and polychromatophilia and an increase of mononuclear and polymorphonuclear leucocytes; in some smears also a marked increase in the eosinophiles. Counts were not given. OVINE PIROPLASMOSIS (CARCEAG) Babesia ovis, Starcovici, discovered by Babes in 1892, is very similar to Babesia bigemina. The form of the parasite is round or pyriform. It is not rare to find two to four in the same red corpuscle. The liver seems to be the principal depot of the para- sites, which occur within red corpuscles and free (Motas). Divid- ing forms are frequent in the capillaries of the liver. The injection of a large quantity of virulent blood into cattle, goats, dogs, cats and rabbits was without effect (Motas). Complete examinations are not recorded. The number of red corpuscles falls from eight or nine millions to four millions or less. There is a marked diminution in the amount of hemoglobin, from 13 and 14 grams per 100 cc. to seven or eight grams per cubic centimeter and a lessened coagulability (Motas). The red cor- puscles vary greatly in size (Bonome). BABESIA MUTANS, THEILER Theiler has recently described another species of Babesia in the blood of cattle in South Africa. He named the parasite causing it Piroplasma mutans. The organism is found, though never present in large numbers, in the peripheral blood of the affected animals. In the cases observed by Theiler, B. bigemina has also been present. The disease is moculable, thus differing from East coast fever. Theiler inoculated 15 calves with the blood of cattle immune to red water and obtained reactions with small organisms in the blood from the 25th to the 41st day after inoculation, be- tween the secondary and tertiary reactions due to B. bigemina. The parasites were present in the blood in small numbers. The importance of this disease is that it is liable to be mistaken for 132. INFECTIOUS DISEASES DUE TO PROTOZOA Hast coast fever, a much more serious disease, from the similarity of the parasites. Theiler states that B. mutans may easily and has constantly been mistaken for Th. parva. Tor diagnostic purposes where small numbers of piroplasmata are found in the Fic. 14. Babesia mutans (Theiler). blood, the examination must be repeated. In East coast fever the organisms will usually increase rapidly in numbers, whereas B. mutans increases slowly and is never present in large numbers. Examinations of the blood, except for parasites, were not given. EAST COAST FEVER (RHODESIAN FEVER) East coast or Rhodesian fever is a destructive disease of cattle in parts of South Africa caused by Theileria parva, Theiler. A disease very similar, if not identical, has been reported in Tunis (Ducloux) and in Egypt (Bitter). The parasites during the first stages of the disease appear as very small rod-shaped or ring forms. Exceptionally, larger and pyriform parasites are found (Schilling). The parasites are found usually in large numbers in the circulating blood. The disease is not inoculable by blood containing the parasites, but is transmitted by ticks. TRYPANOSOMATOSIS 133 ‘ Notwithstanding the large number of parasites in the blood, anemia is not very marked. Often the red corpuscles remain at the normal number, in some cases fall to not less than 4,500,000, in only one case fell to 2,380,000 (Koch). Dschunkowsky and Luhs have described a disease of cattle in Fig. 15. Theileria parva (Theiler). Transcaucasia, due to an organism very similar to Th. parva, which Dschunkowsky named Piroplasma annulatum. The para- sites have been found in large numbers in the blood in the acute form of the disease, up to 95% of the red corpuscles containing from one to eight parasites. The disease may be transmitted by the inoculation of blood containing the parasites. In the chronic form from 10-40% of the red corpuscles contain parasites. Anemia is marked in chronic cases, as low as 800,000 red corpuscles having been found. TRYPANOSOMATOSIS The organisms causing the several varieties of trypanosomatosis are so similar in morphology that a description of one with slight modifications will serve for the others. The trypanosomes are 134 INFECTIOUS DISEASES DUE TO PROTOZOA unicellular organisms found in the blood of the animals infected by them. The body of a trypanosome is fusiform, provided with a lateral undulating membrane, the thickened free border of which terminates in the posterior part of the body in a centrosome or blepharoplast and is prolonged anteriorly as a free flagellum. The nucleus is generally in the anterior part of the body. Multiplica- tion is by longitudinal division and by segmentation, which differs only in that the cell body divides much more slowly than the centrosome (blepharoplast) and nucleus, thus four, eight or 16 small trypanosomes are formed attached by their anterior ends. Conjugation is at present undetermined. ‘Trypanosoma lewisi was cultivated by MacNeal and Novy in 1903. Since then several other species have been cultivated by them and others. A positive diagnosis is made by finding the trypanosomes. In the majority of cases this may be done without difficulty by making a microscopical examination of the blood of the affected animal. The parasites are most numerous in the blood during the febrile periods. During the intermissions the blood is virulent though the parasites may be so few as to escape even a careful microscopical examination. Several preparations should be made and carefully searched. The blood may be examined in the fresh condition for living parasites, or smears may be made and stained, preferably by one of the modifications of the Romanowsky method. When the parasites are scarce it is of advantage to centrifuge the blood or fluid to be examined. If blood, the red corpuscles may be rendered invisible by the addition of dilute acetic acid (one- third per cent. glacial). If the organisms are not found the exam- ination should be repeated each day for several days. Consecu- tive examinations each day for six or seven days will usually reveal the parasites; but in some cases their presence is more easily determined by inoculating a susceptible animal with some of the blood (from a few drops to one cc.) of the suspected case. Mice, rats or dogs are the most susceptible to nearly all of the trypanosomes (not for Tr. theileri), the disease running an acute course in them. The parasites will be found in large numbers in the blood of these animals inoculated with blood or infective material containing pathogenic trypanosomes. At the present time opinions differ as to whether the disease occurring in different countries is caused by different species of SURRA 135 trypanosomes. A discussion of the relationship of the several trypanosomes is outside the scope of this work. The following diseases caused by trypanosomes affect the domesticated animals: TABLE XL.—TRYPANOSOMATOSES : : . Geographical Animals D db BaSeae Specific Cause |iscovered by Distribution Affected Tr. evansi Evans 1880 | India, Indo- Horse, ass, Surra Steel (1885) China, East mule, Indies, Persia, camel, Philippines, elephant, Mauretius, dog. North Africa. Nagana Tr. brucei Bruce 1894 Africa Horses and (Plimmer most and Brad- mammals ford 1899) Dourine Tr. equiper- Rouget 1894 | N. Africa, Per- Horse, ass dum (Dof- sia, Turkey, lein 1901) South France, North Spain. Mal de Tr. equinum | Elmassian S. America Horses, dogs Caderas (Voges 1901)| 1901 Gambian Tr. dimorphon| Dutton and | Senegambia Horses horse (Dutton and| Todd 1904 sickness Todd 1904) In all of these, except dourine, transmission is supposed to be by means of biting insects; in dourine the disease is transmitted by copulation. SURRA The greatest losses from surra are among horses, mules and asses. It is also naturally acquired by the camel, elephant, dog and cat; cattle, buffaloes and caraboa are less susceptible. The 136 INFECTIOUS DISEASES DUE TO PROTOZOA disease is transmissible by inoculation to nearly all mammals, rats and mice being the most susceptible. Tr. evansi is a motile trypanosome 20-30 p in length, including the flagellum, by one to two p in breadth, somewhat blunt at the posterior end and tapering gradually at the anterior end. The undulating membrane is well defined, beginning at or near the centrosome (blepharoplast) in the posterior portion of the parasite and ending anteriorly in a long, free flagellum. The parasites are transmitted by flies, Tabanus tropicus (Rogers), Stomxys calcit- rans, (Curry), possibly other flies and by fleas (Musgrave and Clegg). In the majority of cases the parasites are easily found in the peripheral blood. They are usually seen during the febrile periods; but decrease in numbers during the afebrile periods so that a prolonged search may be required or the microscopical examination may even fail to reveal the trypanosomes. The periods during which the parasites are scarce or are not found on microscopical examination vary from one to six days (Lingard); but the blood is virulent as shown by inoculation into susceptible ani- mals. Mice, rats and dogs are the most suitable for inoculation. In mice and rats the parasites are present in the peripheral blood, two to three days after subcutaneous inoculation, 24 hours after intraperitoneal inoculation and increase in numbers till death, which occurs three and one-half to five days after subcutaneous and two and one-half to three days after intraperitoneal inocu- lation. Changes in the blood.—There is a progressive diminution in the number of red corpuscles and in the amount of hemoglobin. Smith and Kinyoun state that in a horse sick seven days there were 3,500,000 red corpuscles and 14,500 leucocytes, and in an- other, sick six weeks, 3,200,000 red corpuscles and 13,900 leu- cocytes per emm. The hemoglobin was slightly diminished, Fia. 16: Trypanosoma evansi, rat’s blood (Nocht and Mayer). NAGANA 137 about 85%. Hemoglobinuria has been observed in some cases. Musgrave and Clegg state that it may occur temporarily in the first stage of the disease. Leucocy- tosis is given as being constantly present. A detailed study of the changes in the blood has apparently not been made. NAGANA Trypanosoma brucei is very similar to Tr. evansi. Laveran and Mesnil consider that there are certain differ- ences,—that Tr. evansi is more slender, has a longer flagellum and is more ac- tively motile in hanging drop and that animals immunized to one are not to the other species. The parasites are present in the blood in sufficient numbers to be found on microscopical examination during the febrile periods; but during the intermissions may be very scarce. Inoculation of susceptible animals however, shows them to be present even when the microscope fails to re- veal their presence. Toward the end of the disease they are often present in both febrile and afebrile periods in considerable numbers. There may be 50,000-70,000 per cmm. on the day of death (Theiler). Kanthack, Durham and Blandford found the trypanosomes present in Fic. 17. Trypanosoma brucei, rat’s blood (Nocht and Mayer). the right inguinal lymph glands in animals inoculated subcutane- ously in the right side one to three days before they were dis- coverable in the blood. 138 INFECTIOUS DISEASES DUE TO PROTOZOA TasLe XLI.—NuMBER OF TRYPANOSOMES IN THE BLoop (AFTER SAUERBECK) ee Kanthack, Durham and Laveran and Mesnil, Proportion a Blandford of Parasites to Red Corpuscles Horse scarce Dog 300,000 1:50-1:100 Mouse, | 2-3,000,000 tel Rat } Guinea pig 500,000 Rabbit 60,000 1:50 Changes in the blood.—The disease is a progressive anemia. Bruce reported a diminution of red corpuscles in one horse from 5,000,000 to 3,800,000 and in another case from 5,500,000 to 2,500,000. The red corpuscles decrease while the parasites be- come more numerous. As low as 848,000 red corpuscles are reported in the dog on the day of death. Erythroblasts are often present. Schilling found a reduction of red corpuscles to 2,270,000 and of hemoglobin to 25%. The red corpuscles lose the power of forming rouleaux and form clumps; the serum of such blood mixed with normal blood of the same species of animal caused the red corpuscles to clump (Kanthack, Durham and Blandford). According to Kanthack, Durham and Blandford leucocytosis is not constant. The highest number they found was 15,000-34,000. Schilling found a slight increase, 11,000 per emm. DOURINE This disease affects equines, mainly horses. Asses are less sus- ceptible. Infection occurs naturally by coition. Dogs, rabbits, rats and mice are easily infected by inoculation. Trypanosoma equiperdum closely resembles Tr. evansi and Tr. brucei. Laveran and Mesnil state that the most important dif- ference is in the absence of the protoplasmic granules such as are present in Tr. brucei. Baldrey states that Tr. equiperdum is smaller than Tr. evansi, the posterior extremity is not so long and is less sharp, and the centrosome and nucleus are more elongated. The parasites are found most easily in blood taken from the MAL DE CADERAS 139 center of newly formed plaques. If the plaques are old (24 hrs.) there may be no parasites visible; the further removed from the center the puncture is made the fewer are the parasites likely to be (Baldrey). Before the forma- tion of plaques diagnosis is diffi- cult. The parasite may often be found by making a microscopical examination of scrapings of the urethra in males or the vulva in females (Baldrey). The parasites are present in the blood in small numbers. Ordinarily they are found only by inoculating a sus- ceptible animal. They are pres- ent in the exudate of the male urethra and of the vagina, in the fyg. 18. Trypanosoma equiperdum, seminal fluid and in the fluid of blood, horse (Rouget). edematous swellings. Changes in the blood.— Detailed examinations of the blood have apparently not been made. From the time of the appearance of the plaques there is a progressive anemia which seems to be more rapid from the time of the appearance of nervous symptoms. There is an increase in the number of leucocytes. Pease states that there is a large increase in the number of eosinophiles. MAL DE CADERAS Trypanosoma equinum is similar in form and size to the other pathogenic trypanosomes, but is readily distinguished morpho- logically by the apparent absence of a centrosome, which is so inconspicuous that its existence has been denied by some. In Tr. equinum the centrosome measures about one-fourth to one- third p. The number of trypanosomes in the peripheral blood varies a good deal in cases of natural infection. Sivori and Lecler state that in cases of progressive anemia they are nearly always found and are numerous, while in cases of parexia they are found from time to time in small numbers, and in cases of paraplegia they are 140 INFECTIOUS DISEASES DUE TO PROTOZOA not generally found on microscopical examination. When not found on microscopical examination the injection of blood into susceptible animals (horses, dogs, mice, rats) proves their ex- istence. Fia. 19. Trypanosoma equiperdum, blood, mice; a, four days after inoculation, b, eight days after inoculation (Rouget). Changes in the blood.—There is a progressive diminution in the number of red corpuscles and in the amount of hemoglobin. The red corpuscles may fall to 3,000,000 or less some days before death. Voges found them reduced to 800,000 in one ease. The hemoglobin may be reduced to five or six per cent. (grams per 100 ce.). The leucocytes appear to be increased. Sivori and Lecler found 10,000 on the fourth day after inoculation. Of these MAL DE CADERAS 141 there were 70% polymorphs, 20% large and medium mononu- clears, five per cent. lymphocytes, four per cent. eosinophiles and one per cent. transitionals. Before inoculation the same horse had an average of 54.5% polymorphs, 31.25% large and medium & = Fic. 20. Trypanosoma equinum, horse (Sivori and Lecler). mononuclears, nine per cent. lymphocytes, 3.25% eosins and two per cent. transitionals. The leucocytes vary corresponding to the appearance and increase of the trypanosomes in the blood. At the end of the disease they found 10% myelocytes. The following case illustrates the progressive diminution of the red corpuscles and of the hemoglobin. 142 INFECTIOUS DISEASES DUE TO PROTOZOA TasBLe XLII.—Mau pre Capreras (EXPERIMENTAL Horse No. 3 Stvorit AND _———?}———<——— | — ————————— |__| —_—________ Date Temp. | Temp A. M. P.M 16-VIII 37. By eel 17-VIII 38. 38. 18-VIII 38. 39.6 19-VIII 39. 39.3 — 20-VIII 38.8 39. 22-VIII 37.1 37. 24-VIII 36.5 39. 1-IX 38. 37.6 6-IX 39.4 40.6 8-IX 37.2 38.5 15-IX 37.5 37.9 10-X 37.5 o7.7 14-X 38.6 39.3 19—X 37.8 38.4 25-X 38. 38.5 4-XI 38. 38.1 17-XI 39.4 38.3 22-XI LECLER) Red Cor- Hb. % Trypanosomes puscles 6,000,000 | 14. 5,900,000 | 14. 5,800,000 | 14. 2-3 per field, first seen. 5,500,000 15.5 | 2-3 per field. 6,100,000 14. 2-3 per field. 5,500,000 10.5 | none observed. 5,400,000 | 10.2 | none observed. 4,900,000 9.8 | none observed. 5,000,000. 9.8 | 1 per field. 4,500,000 8.8 | none observed. 3,400,000 7.9 |1 to 2-3 fields. 3,000,000 6.7 | 5-10 per field. 2,800,000 6. 10-20 per field. 2,500,000 6. 1-2 per field. 2,300,000 5.7 | none observed. 2,000,000 4.5 | none observed. 1,800,000 3.5 | 5-10 per field. 5-10 per field, died. GAMBIAN HORSE SICKNESS Horses are affected naturally; horses, rats, mice, guinea pigs, Fig. 21. Trypanosoma dimor- phon (Laveran and Mesnil). rabbits, dogs, cattle and goats may be given the disease by inoculation. Trypanosoma dimorphon was found by Dutton and Todd in three forms: (1) in the beginning of the disease small forms 11-13 by 0.8-1 p, with very small undulating mem- brane, short flagellum and blunt pos- terior end; (2) long form 26-30 by 1.6-2 p, with very long flagellum; (3) “stumpy form” 16-19 by 3.4- 3.5 @, with short thick body and very long flagellum. Transitional forms between these occur. The long form (2) is very numerous during the last stages of the dis- ease. Laveran and Mesnil did not find the ‘‘stumpy form” TRYPANOSOMA THEILERI 143 and found that the long form had no free flagellum, but that in the long and short forms the protoplasm extended to the end of the flagellum. The parasites are found in small numbers (to 10 in a field of the microscope) in the peripheral blood, or are not found for several days at a time. TRYPANOSOMA THEILERI The parasite, named Trypanosoma theileri, was found by Theiler in 1902 in the blood of cattle. It is the largest of the pathogenic trypanosomes, measuring 20-70 by 2-6 p, and is di- morphic. In the ordinary form (Theiler) the centrosome is situ- ated at the posterior end of the body at some distance from the nucleus; morphologically it resembles Tr. brucei except that it is much larger. In the rarer form the centrosome is near or even attached to the nucleus. This form is broader and shows ab- normal shapes,—round, oval, lacerated, etc., while the nucleus is larger and generally less compact. The parasites are most quickly found in living blood. They are present in the blood a varying length of time. The longest period observed by Theiler was 13 days, the shortest one day, the average nine days. The number of parasites in the peripheral blood varies a good deal. In one case Theiler found 30 per field (objective No. 6 Zeiss); in other cases none could be found on microscopical examination. Theiler considers about five per field as a fair average. Frequently the parasites have been transmitted in the process of immunizing cattle against rinderpest, the blood injected containing the trypanosomes. Theiler showed experimentally that flies, Hippobosca rufipes and H. maculata, F'6- 22. Try- may transmit the disease. Tr. theileri is at pres- ?@”080"@ Mee : Z . (Theiler). ent regarded to have very little if any pathogenic properties. A similar trypanosome, perhaps the same species, has been found in the blood of apparently healthy cattle in America. Changes in the blood.—The effect on the animal varies greatly. In some cases no symptoms are produced; in others there is a marked reduction in the number of red corpuscles. In one of 144 INFECTIOUS DISEASES DUE TO PROTOZOA Theiler’s experimental cases the red corpuscles fell from 6,780,000 to 3,000,000 per emm. Poikilocytosis was observed. In more severe cases the number may be reduced still lower, with marked pathological changes in the red corpuscles, the presence of numer- ous corpuscles showing punctate basophilia, many erythroblasts (normoblasts and megaloblasts), microcytes and megalocytes. In cases of light infection the number of leucocytes is scarcely increased; but when the trypanosomes are frequent, the leucocytes seem to increase in direct proportion. The eosinophiles are usually increased in number and are sometimes very numerous (Theiler). REFERENCES 1. Bases, V. Etiologie de l’hémoglobinurie bactérienne du boeuf. Ann. d UInst. d. path. et. d. bact. d. Bukarest, i, 1888-89. 2. Bases, V. Die Aetiologie der seuchenhaften Hiimoglobinurie des Rindes. Arch. f. path. Anat., exv, 1889, 8. 81. 3. Bases, V. L’étiologie d’une enzootie des moutons denommée Carceag en Roumanie. Compt. rend. Acad. d. Sciences, exv, 1892, p. 359. 4. Bauprey, Ef. 8. W. Dourine. Journ. of Compar. Path. and Therap., xviii, 1905, p. 1. 5. BARUCHELLO UND Morr. Untersuchungen iiber die in Italien vorkom- mende Piroplasmose des Pferdes. Centralbl. f. Bakt., I Abt. Orig., xliii, 1907, S. 593. 6. Brrrer. Cited by Theiler. 7. Bonomr, A. Ueber parasitiire Ietero-Hematurie der Schafe. Beitrag zum Studien der Amoebo-Sporidien. Arch. f. path. Anat. exxxix, 1895, 5S. 1. 8. Bownttu, T. Equine piroplasmosis. Journ. of Hyg., v, 1905, p. 7. 9. Bruce, Davin. Preliminary report on the tsetse fly disease or nagana in Zululand. Durban, 1895. 10. Brucr, Davin. Further report on the tsetse fly disease or nagana in Zululand. Ubombo, Zululand, 1896. 11. Bruce, Davip. Note on the discovery of a new Trypanosoma. Pro- ceed. Roy. Soc. London, |xix, 1902, p. 496. 12. Courmont ET Lesteur. La polynucléose de la rage. Journ. d. Physiol. et d. Path. gen., iii, 1901, p. 599. 13. Crawtey, H. A Trypanosome which appears in cultures made from the blood of American cattle. U.S. Dept. of Agr., Bur. Anim. Ind., Bul. 119, Oct. 1909. 14. Curry, J. J. Report on a parasitic disease of horses, mules and caraboa in the Philippine Islands. Am. Med., iii, 1902, p. 512, also Vet. Journ., liv, p. 292. 15. Dortern, F. Die Protozoen als Parasiten und Krankheitserreger, Jena, 1901. REFERENCES 145 16. Dortern, F. Lehrbuch der Protozoenkunde. 3te Auf. 1911. Gustav Fischer, Jena. , 17. DscHUNKOWSKY UND Luus. Die Piroplasmosen des Rindes. Centralbl. f. Bakt., I Abt. Orig., xxxv, 1904, S. 486. 18. Ductoux, E. Sur une piroplasmose bacilliforme de bceuf en Tunisie. Compt. Rend. Soc. d. Biol., lvii, 1905, p. 461. 19. Durron anD Topp. Researches on trypanosomiases in West Africa. Brit. Med. Journ., 1903, II, p. 650. 20. Exmasstan, M. MaldeCaderas. Anal. d.l. Univ. Nacional, Asuncion, i, 1901. 21. Eumasstan, M. Mal de Caderas. Berlin. tierdirtz. Woch., 1901, S. 604. 22. Evans, GrirritaH. Report on “surra” disease in the Dera Ismail Khan district. Military Dept., Nov. 13, 1880. 23. Evans, Grirrtid. On a horse disease in India known as ‘‘surra,”’ probably due to a Hematozoan. Vet. Journ., xiii, 1881, pp. 1, 83, 180, 326 and xiv, pp. 97, 181. 24. GrawaM-Smitu, G. S. Canine piroplasmosis III, morbid anatomy. Journ. of Hyg., v, 1905, p. 250. 25. GueuieLmr. Un caso di malaria nel cavallo. La Clinica veterinaria, 1899, p. 220. 26. Hipparp anp Neau. Some observations on the blood of dairy cows in tick-infested regions. Journ. Inf. Dis., ix, 1911, 324. 27. KanTHACK, DURHAM AND BLANDForRD. On nagana or tsetse fly disease. Proceed. Roy. Soc. London, |xiv, 1898, p. 100. 28. Kocu, Ropertr. Rhodesian investigations. Agr. Journ. Cape of Good Hope, xxiv, 1904. 29. Kocu, Rosperr. Vorliufige Mitteilungen iiber die Ergebnisse einer Forchungreise nach Ostafrica. Deutsch, med. Woch., xxxi, 1905, S. 1865. 30. Kosset, H. Die Haimoglobinurie der Rinder. Handb. d. path. Mikroog. von Kolle u Wassermann, Bd. I, 1903, S. 841. 31. KossEL unp WesBer. Ueber die Himoglobinurie der Rinder in Fin- land. Arbeit. Kais. Ges-Amt., xvii, 1900. 32. Launoy eT Livy-Bruuxu. Les variations numériques et morphologiques des globules blanes chez les poules infectées de Spirochaeta gallinarum. Compt. rend. Soc. d. Biol., xxiv, 1913, 754. 33. Launoy, L. et Lfivy-Bruat, M. Sur L’anémie observée chez la poule au cours de l’infection par la Spirochaeta gallinarum. Compt. rend. Soc. d. Biol., lxxv, 1913, 250. 34. LaveraNn, A. Contribution a l’étude de Piroplasma equi. Compt. rend. Soc. d. Biol., liii, 1901, p. 385. 35. LAVERAN, A. Sur un nouveau trypanosoma des bovidés. Compt. rend. Acad. d. Sciences., exxxiv, 1902, p. 512. 36. LAVERAN ET Mesniu. Recherches morphologiques et expérimentales sur le trypanosome du nagana ou maladie de la mouche tsetse. Ann. d. UInst. Past., xvi, 1902, p. 1. 37. LAVERAN ET Mesniu. ‘Sur un trypanosome d’Afrique pathogéne pour 146 INFECTIOUS DISEASES DUE TO PROTOZOA les équidés. Tr. dimorphon Dutton et Todd. Compt. rendes Acad. d. Sciences, exxxvill, 1904, p. 732. 38. LaverRAN AND MeEsniu. Trypanosomes and trypanosomiases, transl. by D. Nabarro. London, 1907. 39. Lincarp, A. Annual report of Imperial Bacteriologist, 1900-1901, Calcutta, p. 3. 40. MacNrnau anp Novy. On the cultivation of Trypanosoma lewisi. Contributions to medical research, dedicated to Vaughn, 1903, p. 549. 41. Moras, C. 8. Contribution a |’étude de la piroplasmose ovine (Car- coag). Arhiva Veterinara Bucarest, 1904. Abstr. in Fol. Ham. i, 1904, p. 607. 42. MusGraAvE AND Cieca. Report on trypanosoma and trypanosomiasis, with special reference to surra, in the Philippine Islands. No. 5, Bur. of Govt. Laboratories, Manila, 1903. 43. Nocarp, E. Sur la fréquence en France et sur le diagnostic de la piro- plasmose canine. Bull. d. l. Soc. centr., 1xi, 1902, p. 716. 44. Nocarp ret Atmy. Une observation de piroplasmose canine. Bull. d. l. Soc. centr., Ix, 1901, p. 192. 45. Nocarp ET Moras. Contribution a l'étude de la piroplasmose canine. Ann. d. UV Inst. Past., xvi, 1902, p. 257. 46. Nocut unp Mayrr. Trypanosomen als Krankheitserreger. Handb. d. path. Mikroorganismen, Kolle und Wassermann, Erst. Ergdnzsungsbd., 1907, S. 1. 47. Nurrauy, G. H. F. Canine piroplasmosis I. Journ. of Hyg., iv, 1904, p. 219. 48. NurraLL AND GRAHAM-SmiTH. Canine piroplasmosis. Journ. of Hyg., v, 1905, p. 237; vi, 1906, p. 586 and vii, 1907, p. 232. 49. Pease, H. T. Surra and dourine. Vet. Journ., N.S., ix, 1904, p. 187. 50. Prana BT GALLI-VALERIO. Su di un’ infezione del cane con parassiti endoglobulari nel sangue. Jl moderno Zooiatro, 1895, p. 163. 51. PLimmer AND Braprorp. A preliminary note on the morphology and distribution of the organism found in the tsetse fly disease. Proceed. Roy. Soc. London, |xv, 1899, p. 274. 52. Roperrson, Wm. Malignant jaundice in the dog. Journ. of Compar. Path. and Therap., xiv, 1901, p. 327. 53. Roacrrs, L. The transmission of the Trypanosoma evansi by horse flies, ete. Proceed. Roy. Soc. London, \xviii, 1901, p. 163. 54. Roucrr, J. Contribution a l'étude du trypanosome des mamiféres. Ann. d. UInst. Past., x, 1896, p. 716. 55. SauperBEcK, E. Die Trypanosomiasis vom Standpunkt der allgemeinen Pathologie. Lubarsch und Ostertag-Ergebnisse der allgem. Path., 10te Jahrg., 1904-05, S. 305. 56. Scuriytinc, C. Piroplasmosen. Kolle und Wasserman. Handb. d. path. Mikroorganismen, Erst. Ergdnzungsbd., 1907, 8. 76. 57. ScuriuiNG, C. Bericht iiber die Surra-Krankheit der Pferde. Centralbl. f. Bakt., xxx, 1st Abt. Orig., 1901, S. 545. 58. Stvort prt Lecter. La surra américain ou mal de Caderas. Anal. d. Minist. d. Agr. Argentina, i, No. 1, 1902. REFERENCES 147 59. SmitH, THEOBALD. Preliminary observations on the micro-organism of Texas fever. Med. News, Dec. 21, 1889. 60. SmitH AND KILBOURNE. Investigations into the nature, causation and prevention of Texas or Southern cattle fever. Bull. No. 1, Bur. An. Ind., U.S. Dept. of Agr., 1893. 61. SmirH AND Kinyoun. A preliminary note on a parasitic disease of horses. Army path. laboratory, Manila, P. I., 1901. 62. Starcovict, C. Bemerkungen iiber den durch Babes entdeckten Blut- parasiten und die durch denselben hervorgebrachten Krankheiten, etc. Cen- tralbl. f. Bakt., xiv, 1893, S. 1. 63. STEEL, J. H. An investigation into an obscure and fatal disease among transport mules in British Burma. 1885. 64. THemerR, A. Die Pferde-Malaria. Schweizer-Arch. f. Tierheilk., xliii, 1901, S. 253. 65. THEILeER, A. Die Tsetse-Krankheit. Schweizer-Arch. f. Tierheilk., xliii, 1901, S. 97, 153. 66. THEmLeR, A. A new trypanosome and the disease caused by it. Journ. of Compar. Path. and Therap., xvi., 1903, p. 193. 67. THeILER, A. Piroplasma mutans noy. sp. A new species of piroplasma and the disease caused by it. Report of Govt. Vet. Bacteriologist for 1905-06, Dept. of Agr. Transvaal, 1907, p. 33. Also Journ. of Compar. Path. and Therap., xix, 1906, p. 292. 68. Vocres,O. Das Mal de Caderas. Zeitschr. f. Hyg., xxxix, 1902, 8. 323. 69. Wituiams, A. J. Indian equine piroplasmosis. Journ. of Compar. Path. and Therap., xx, 1907, p. 23. 70. Wricut, J. A. Canine piroplasmosis IV. On certain changes in the blood. Journ. of Hyg., v, 1905, p. 268. 71. ZreMANN, H. Ueber Malaria und andere Blutparasiten u. s. w. Jena, 1898. 72. ZieMANN, H. Neue Untersuchungen iiber Malaria und den Malariaer- regern nahestehenden Blutparasiten. Deutsch. med. Woch., xxiv, 1898, 8. 125. CHAPTER X DISEASES WHOSE CAUSE HAS NOT BEEN DEFINITELY DETERMINED DOG DISTEMPER Sabrazes and Muratet examined the blood of four cases of dog distemper at Bordeaux where the nervous form of the disease is frequently observed and is very severe, the mortality exceeding 60%. The counts of red and white corpuscles and the amount of hemoglobin were not reported. In one case there were signs of anemia with normoblastic reaction. In another case there were slight anisocytosis and polychromatophilia. The changes in the leucocytes were more pronounced. Polynucleosis with lessened number of eosins was the rule. The absolute number of leucocytes was increased in three cases. The iodine reaction was present each time it was made (three cases). The following table gives the differential counts in the four cases examined. Taste XLIII.—Doc Distemper (SABRAZES AND MURATET) . 2 . | Age Large ain T ar , = = No. |Sex Mo. Lymph. Maes Polym. | Eos. 38 I m| 10 5.% |11.% | 83.5% |0.54% symptoms for 24 hrs. Te et 6.) 27AL |S OA ee 0.85 |0.41 |sick 6 hrs. temp. 39.2° C. IV |m]} 3 6. St 91. sick 5 days; temp. 37.6°. V f | 14 } 138.84 85. 0.84 |sick 114 months. RINDERPEST Though the specific cause of rinderpest has not been determined, the blood of animals sick of the disease is virulent. Jobling states that it has been proven experimentally that 0.1 ce. of blood from a sick animal injected subcutaneously will produce the disease. 148 RINDERPEST 149 Nicholle and Adil-Bey have shown that the virus is able to pass through a Berkefeld filter. The changes in the leucocytes in the course of the disease have been studied by Réfik-Bey. He found that in cases of fatal infection there is an initial increase in the number of leucocytes followed by a decrease, constantly present; then there is a secondary increase, constantly present. The initial increase takes place the second or third day after inoculation; the number may reach 18,300 per emm. The mononuclears, including the lymphocytes, increase in about half the cases, maximum ob- served 12,300 percmm. Réfik-Bey states that the normal number of mononuclears in cattle is from 4,500 to 6,500 per emm., 57-84%; the normal number of polynuclears 1,500 to 3,500 per emm. Gen- erally the polynuclears take part in the initial increase, maximum observed 8,000. Sometimes the eosinophiles are increased, maxi- mum observed 3,500. The number of leucocytes commences to decrease the fourth day (sometimes the third), the minimum being ordinarily reached the fifth day (sometimes the fourth, exceptionally the sixth or seventh). The lowest count obtained was 2,000. The minimum is generally observed the day of the rise of temperature, rarely the day before and sometimes the second or third day of fever. In this stage all the varieties are diminished. The minimum number of mononuclears observed was 1,000 per emm., of polymorphs 200 per cmm., of eosinophiles 200 per emm. The eosinophiles then disappear and do not reappear. The secondary increase begins generally the eighth day, some- times the seventh and exceptionally the ninth. Usually the tem- perature falls at this time; rarely it has fallen the day before or falls the day following the increase. The leucocytes in the cases observed have increased, exceeding the normal; in one case there were 45,000 per cmm. Toward the time of death the leucocytes commence to decrease. The mononuclears increase, in half the cases observed not reaching the normal; in the other half reaching or exceeding the normal, maximum 27,000 per cmm. The poly- morphs increase, sometimes simply returning to the normal number and sometimes exceeding it. The polynucleosis is more marked when life is prolonged, maximum observed 18,000 per cmm. 150 DISEASES WHOSE CAUSE IS UNKNOWN HOG CHOLERA The cause of hog cholera has been determined to be a virus that will pass through a bacteria proof filter. King, Baeslack and Hoff- mann state that they have constantly found by dark field examina- tion a spirochete in the blood of hogs (40 cases) affected with hog cholera but did not find it in the blood of hogs (48 cases) not affected with hog cholera. The spirochetes were not found in large numbers, but were found with less difficulty in the acute form of the disease and when it was at its height. The spirochete, which they named Spirochaeta su7s, later Sp. hyos, is relatively large, averaging from 5-7 micra in length by one micron in width. The blood was drawn aseptically from the caudal artery into sterile sodium citrate solution and kept in the incubator until the examination was completed. Dilution was in the proportion of about one part of blood to ten or fifteen of citrate solution. The changes occurring in the blood in hog cholera were studied by King and Wilson. They found the red corpuscles and hemo- globin decreased. The red corpuscles varied from 2.8 to 9 millions with an average of 5.84 millions. “The hemoglobin varied from 50 to 82, average 72. There was an increase of ore minute in the average time of coagulation. The leucocytes were decreased. The number varied from 7,200 to 23,600 with an average of 15,515. The decrease was in the lymphocytes, polymorphonuclears and eosinophiles. The large mononuclears and mast cells were increased. The lymphocytes varied from 16.5 to 74.2 per cent., average 50.3; the large mononu- clears 2 to 42 per cent., average 11.3; polymorphonuclears 10 to 70 per cent., average 31.8; eosinophiles 0 to 16 per cent., average 3.5 and mast cells 0 to 18.1, average 3.5. EQUINE INFECTIOUS ANEMIA The cause of equine infectious anemia has not been discovered. It is so minute it will pass through a bacteria proof filter (Carrée and Vallé). The disease may be transmitted by the inoculation of blood or urine (Carée and Vallé, Francis and Marstellar and others) of a diseased animal. It is probable that the disease in America known as swamp fever is the same. EQUINE INFECTIOUS ANEMIA 151 Infectious anemia was at one time called ‘pernicious anemia” because of the profound anemia produced by it. The red corpuscles and hemoglobin are much reduced. As low as one million red corpuscles and 25 per cent. hemoglobin are reported (Meier). Marked changes in the size, shape and staining of the red cor- puscles are found in the more marked cases of the disease. Many poikilocytes, microcytes and megalocytes may be found. Nu- cleated red cells sometimes occur, normoblasts and microblasts. Megaloblasts have not been observed. The leucocytes are decreased. Sometimes they are increased, but possibly the leucocytosis is due to some other condition. The polymorphonuclears do not seem reduced so much or so constantly as the lymphocytes. Further investigation is needed to determine the changes in the leucocytes. The following tables are of cases in Switzerland reported by Meier. TaBLeE XLIV.—Cases or Equine Inrectious ANEMIA (MEIER) ae Date = Sex ape gh Hb. Leucocytes 13 13 g 1.452 25 5,000 14 10-X-04 5% m 1.8 25-30 5,880 11-X 1.9 25-30 6,200 17-X 1.74 25-30 5,500 15 13-I-05 11 m 4.0 60 8,000 17-1 3.22 45-50 20,600 16 7-IlI a m 1.76 30 4,500 17 3-XI hs m 2.376 30-35 21,870 26-XI 2.4 30-35 6,600 8-XIT 1.3-1.9 30 4,026 10-XII 2.4 30 50-7,812 11-XII 3.3 62-5,312 12-XII 10,920 18 9-VI 9 m 3.4 45 4,200 19 10-VI 6 g 1.0 25 10,800 152 DISEASES WHOSE CAUSE IS UNKNOWN TaBLE XLV.—Tue LevucocytTes IN Cases oF Equine INFEcTIOUS ANEMIA (MErER) L VARIETIES OF LEUcocyTES, NUMBER AND PERCENT euco- No. | Date ee cytes |T,ympho.|Large Mon.|Polymorphs| Eosins |Mast Cells 13 5,000 | 1,640 587 2,675 42.8 1157 33.5 “14 | 11-X| 6,200] 389 375 5,439 39. 6.2 5.4 87.7 «|) 1086 | 17-X |} 5,500) 389 223 4,865 10. 7.0 4.0 88.4 | 0.17 “15 | 13-1] 8,000 | 2,249 1,435 4,160 21. 28.0 17.9 52. 0.2 ~~ | 17-1 | 20,600 | 3,333 1,517 15,610 16 7.2 tise lee 16 | 7-IIl| 4,500] 932 151 3,406 7 20.7 Pa 75.60 | 0.1 “17 | 26-X1] 6,600 | 1,485 704 4,400 11 22.5 10.6 66:6. |. ===") ene | 12-X1T | 10,920 | 3,594 1,168 6,136 32.91 10.7 ae a es “1g | 9-VI| 4,200] 946 295 2,956 22.5 6.8 70S | eee “19 | 10-VI | 10,800 | 5,810 614 4,150 ive 53.8 5.6 58.4. | 0001 RHEUMATISM There are but few cases reported in animals in which examina- tions of the blood have been made. In man there is an oligo- cythemia in some cases. Cabot in 163 cases had 13 with less than 4,000,000 red corpuscles. The lowest count was 2,528,000 with 45% hemoglobin. The average number for the entire series was 4,300,000. The hemoglobin suffers more than the red corpuscles. In Cabot’s cases the average was 63% with a color index of .73. The fibrin is much increased. There is usually a moderate leu- cocytosis but in mild cases without exudation there is usually no inerease in the leucocytes. The average number of leucocytes in 243 cases reported by Cabot was 13,800. Turk insists that when REFERENCES 153 the leucocytes reach or exceed 20,000 it is nearly always due to complications (pleuritis, peritonitis, pneumonia). Ewing supports Turk’s statement. When the leucocytes do not exceed the normal limit, there is little change in the proportion of the varieties; but with distinct leucocytosis, there is an absolute increase in the polymorphs. The eosinophiles are scanty or may be absent during the very early stages, but later are present in moderate numbers. After defervescence the eosins are usually increased. Meier reports a case of muscular rheumatism in a mare, five and one-half years old, with 8,420,000 red corpuscles, 90% hemo- globin, and 20,600 leucocytes, of which there were 8.1% lym- phocytes, 2.5% large mononuclears, and 89.3% polymorphonu- clears. Dr. Ward Giltner, in a personal communication which he has kindly permitted me to use, gives the counts in a gelding about 15 years old, taken with acute inflammatory rheumatism. On Nov. 9th, with a temperature of 105° F., there were 5,250,000 red cor- puscles and 20,000 leucocytes, of which 2.6% were lymphocytes, three per cent. large mononuclears and 94.4% polymorphs. No eosins were found. Two days later the animal was much improved; eosins were present in normal numbers. On Dec. 19th, the disease had changed into the chronic form; the horse was down, unable to rise. There were 6,200,000 red corpuscles, 11,000 leucocytes and 70% hemoglobin. Of the leucocytes 13.6% were lymphocytes, one per cent. large mononuclears, 84.8% polymorphs, 0.4% eosins and 0.2% mast cells. The horse died the following day, the post- mortem showing lesions of articular rheumatism, ulcers on the articular cartilages and fibrosis of left biceps. REFERENCES 1. Francis AND MarsTeELuarR. Infectious anemia of the horse. Tezras Agr. Exp. Sta., Bul. 119. 2. Josyine, J. W. A preliminary report on rinderpest of cattle and carabaos in the Philippine Islands. Bull. No. 4, Bur. of Govt. Lab., Manila, 1903. 3. Kina, Barstack AND HorrMaNN. Studies on the virus of hog cholera. Journ. Inf. Dis., xii, 1913, 206. 4. Kine, W. E. anp Witson, R. H. Studies on hog cholera and preventive treatment. Kans. State Agr. Col. Exp. Sta., Bul. 171, 1910. 5. Kina anp Witson. Spirocheta suis, its significance as a pathogenic organism. Journ. Inf. Dis., xii, 1913, 463. 154 DISEASES WHOSE CAUSE IS UNKNOWN 6. Mack, W. B. Equine anemia. Univ. of Nev., Agr. Exp. Sta., Bul. 68, Mar. 1909. 7. Metigr, P. Beitriige zur vergleichenden Blutpathologie. Zeitschr, f. Tiermed, x, 1906, S. 1. 8. NicHOLLE ET ApiL-Bry. Etudes sur la peste bovine. Ann. d. l’Inst. Past., xvi, 1902, p. 56. 9. Rérik-Bry. Modifications leucocytaires dans la peste bovine. Ann. d. lV Inst. Past., xvi, 1902, p. 163. 10. SaBprazis ET Muratet. Contribution a l’étude du sang et du liquide cephalo-rachidien dans la ‘‘ Maladie” des chiens. Rev. gen. d. Med. Vet., viii, 1906, p. 663. 11. TurK. Klinisch. Untersuchungen. Wien, 1897. 12. VauLtfe eT Carr. Sur la nature infectieuse de l’anémie du cheval. Compt. rend. Acad. d. Sc., exxxix, 1904, 331 and 1239. 13. VaLLie UND Carr. Die infektidse Natur der Animie der Pferde. Berl. tierartz. Woch., Aug. 18, 1904. 14. VaLLfie eT Carrté. Recherches expérimentales sur |’anémie pernicieuse du cheval (Typho-anémie infectieuse). Rev. gen. med. vet., viii, 1906, 593. 5 4 4 : et j Pa Pa eee CHAPTER XI DISEASES DUE TO ANIMAL PARASITES Acariasis.—