TEXT BOOK OF MILK HYGIENE BY DR. WILLIAM ERNST Official Veterinarian and Director of the Royal Milk Control Station at Munich AUTHORIZED TRANSLATION WITH ANNOTATIONS AND REVISIONS BY DR. JOHN R. MOHLER, A. M., V. M. D. Chief of Pathological Division, United States Bureau of Animal Industry AND DR. ADOLPH EICHHORN, D. V. S. Senior Bacteriologist, Pathological Division, United States Bureau of Animal Industry With 29 Illustrations and 5 Colored Plates CHICAGO, U. S. A. ALEXANDER EGER PUBLISHER 1914 COPYRIGHTED AT WASHINGTON, D. C. BY ALEXANDER EGER 1914 ;^ COMPOSITION. ELECTROTYPINO. PRINTING AND BINDING BY THE W. B. CONKEY COMPANY HAMMOND. INDIANA Table of Contents PAGE Translator 's Preface vii Author 's Preface ix CHAPTER I. Anatomy. Pathology and Histology of the Mammary Gland 1 CHAPTER II. Physiology of Lactation and Characteristics of Milk in General 16 CHAPTER III. Microscopy of Milk in General 24 CHAPTER IV. Composition of Milk and Its Biological, Chemical and Physical Characteristics 32 CHAPTER V. Procurement of Cow 's Milk 58 CHAPTER VI. Internal Influences on the Character of Milk 62 CHAPTER VII. External Influences which Act Upon Milk 132 CHAPTER VIII. Bacteria in Market Milk ; Their Origin and Action 152 CHAPTER IX. Milk Control 203 CHAPTER X. Milk Inspection 211 CHAPTER XI. Fundamental Principles of Legislative Milk Control. ....*... 233 Translator's Preface THE importance of food hygiene in the protection and preser- vation of public health is now generally recognized. Milk constitutes one of the most important foods for the human race, and since its composition and wholesomeness are entirely dependent upon its proper handling, the necessity for a strict supervision and control is obvious. The problem of milk hygiene is very complex and must embody all phases of milk control from the time the milk is produced until it reaches the consumer. In all stages it may be subjected to wilful adulteration and to contamination with injurious and ob- noxious substances. Furthermore, the danger threatens this most valuable food not only from outside sources but also from internal influences, as the milk may leave the animal in the condition of a dangerous product, a carrier of pathogenic microbes. Various kinds of infection of the udder are frequently important factors in the contamination of milk, which would render it dangerous to the consumers. Thus in recent years numerous outbreaks of in- fectious sore throats have been caused by such conditions. It is therefore apparent that in the proper control of the milk supply it is necessary to be familiar with all conditions which may be re- sponsible for an injurious or unwholesome product. The subject is one in which every sanitarian should be thoroughly qualified. Although there are numerous splendid publications available on this subject, they are either too voluminous to be used as text- books or they fail to contain the more recent very important devel- opments made in this branch of public hygiene. The excellent German publication of Dr. Ernst entitled "Milk Hygiene" meets the requirement of a concise, up-to-date work on that subject, and it is with pleasure that in response to requests from various sources we have accepted the preparation of an English edition of this publication. We did not lose sight of the fact that it should meet with the conditions prevailing in this country and accordingly we Translator's Preface, have included much valuable information from the reports of the various Milk Commissions, and other sources. For this reason Chapter X dealing with German laws and regulations has been replaced by Chapter XI which deals solely with the conditions and standards existing in this country. We cannot refrain from expressing our sincere thanks to Dr. H. J. Washburn for his most valued suggestions and assistance in proofreading the manuscript ; also to the publisher, Mr. Alexander Eger, for his interest and courtesy during the preparation of this volume. JOHN E. MOHLER, ADOLPH EICHHOBN. Washington, D. C., July 1, 1914. Author's Preface THE increased importance of milk as human food demands more and more the application of modern accomplishments and experiences achieved by science and practice, in order to elevate the milk industry to the desired high standard. The principal stress must be laid upon production, which con- stitutes a special field of the milk industry, and which is most generally in need of elevation and improvement. The product will be without reproach only when the conditions of production correspond to the value of this food. In the field of production, veterinarians are the proper ex- perts who must stand by the side of the producers and give them the necessary advice and instruction. Only by the active and ex- pert aid of veterinarians can it be hoped to improve the good-will of the producers ; provided, at the same time, other points of milk hygiene which possess bad features — in spite of the active progress of milk control and sanitary methods which have been noted for many decades — also receive proper attention. In order to be able to offer expert advice a thorough knowl- edge of milk, its formation, procurement and characteristics, is necessary; likewise, a knowledge of conditions which have an in- fluence upon milk while still in the animal body, and the factors which change this food after its procurement. These points have received the principal consideration in the following chapters. In the plan which I have followed, those questions which treat of the judgment of milk as human food in relation to its chemical contents, were given less prominence. Certain points of this sub- ject have been mentioned only to an extent that was considered advisable for the general comprehension of the subject. More specific questions, as for instance, the preparation of certain milk mixtures for the feeding of infants, the advantages and disadvan- tages of feeding cows' milk to infants, the action of a milk diet in the treatment of adults, etc., are subjects for the physician. A Author's Preface. special chapter on the preparation of infants' milk, or certified milk, has been omitted, since the sanitarian can not make any distinction in his judgment of milk as food, but must remember that milk which is consumed by children of the masses should also come up to the requirements established for any food product from a hygienic standpoint. The chemical and physical properties of milk are only dis- cussed to an extent deemed necessary to instruct the veterinary experts in court cases in judging physiological, pathological and external influences. Since the chemical examination of milk should be placed in the hands of the food chemist, I have eliminated the analytical examination of milk and the examination for preserva- tives. For this information I would recommend the numerous publications which have appeared during recent times, as for in- stance, the works of Grimmer and Sommerfeld, Teichert, Utz and Barthel. Only those methods have been described which may be undertaken by the veterinarian and which are sufficient for a thorough preliminary test of milk for adulterations. The illustrations are taken partly from the known works of my previous teacher, Professor Dr. med. Th. Kitt (Pathological Anatomy) and from Friedberger and Frohner's Methods of Clini- cal Examination ; some were drawn by myself. The illustrations of apparatuses have been avoided, as they appear in all commercial catalogues. In dividing the subject into individual chapters repetitions, of course, could not be avoided. With the preparation of this small work I desire to show to my colleagues the road which they must follow in order to cooper- ate from a milk inspection standpoint in accordance with the call made upon their profession. A difficult point of milk hygiene lies in the changing conditions of production and not in the control of milk consumption or in the supervision of milk transportation. W. ERNST. Munich, January, 1913. CHAPTER I. ANATOMY, PATHOLOGY AND HISTOLOGY OF THE MAMMARY GLAND. Development and Gross Anatomical Structure. In the lowest form of mammalian life a group of glandular ducts becomes differentiated from the glands of the skin in the median abdominal region. These ducts exude their lacteal secre- tions upon tufts of hair of the mammary region, from which it is either licked or sucked by the young (duckbill, Ornithorhynchus paradoxus). One of the land duckbills, the spiny anteater (Echidna hys- trix), has lacteal ducts opening within an abdominal pouch formed by a fold of skin of the mammary region in the shape of a pocket, in which the young are protected and nourished during their development. This abdominal pouch is not identical with the tegumentary wall from which is developed the teats of higher mammals, but it may be taken as the point of origin of the different forms of teats. In higher marsupial animals the glandular ducts are united into a complex gland with teats which constitute the orifices of the confluent lacteal ducts. In other still higher species the most varied kinds of gland structures are observed with various forms of teat development. Among the higher mammalian forms the evolution of these anatomical structures may be followed during embryonic life. On both sides of the body, between the anterior limb-bud and the inguinal fold, the milk-ridge develops from a linear thickening of the ectoblast in the form of a ledge- like elevation of the epidermis. Along this milk-ridge a series of at first spindle-shaped, then round enlargements appear, which are separated by absorption of the intervening portions of the ridge. These enlargements consist of masses of epithelial cells, which correspond to the anlage, primordium or point of origin of the true mammary gland of the lowest mammalia. This anlage sinks into the underlying mesoblastic tissue and becomes surrounded by a proliferating integument, which forms an investment for the growing epithelial mass. From this mammary envelope which becomes more or less flat- tended the fibrous and muscular tissue of the areola and teat are derived. At its base, solid epithelial sprouts grow out from the sides of the conical epidermal plug, later be- coming the lactiferous ducts, while the club-shaped thickened extremities in the further course of their development, form the milk sinus. Subsequently, the central part of the ectoblastic ingrowth undergoes degeneration and what at first was an elevation, now be- 1 Pathology, and Histology of the Mammary Gland. comes a depression. From the middle of this depressed area there appears an elevation that later becomes the teat. In cattle a single excretory canal enters from the bottom of the mammary envelope (point of the teat), into the tissue (the milk duct), the end of which, the milk cistern, breaks up into the secondary lactiferous ducts. The lower opening of the teat con- tains unstriped muscle fibres which act as a sphincter to prevent the escape of milk. (Meckel, Kolliker, Langer, Bonnet, Profe, Schwalbe, Huss, Gegenbauer, Klaatsch.) According to the number of the glandular organs there are distinguished the oligomasts and the polymasts. Cows are nor- mally tetramasts, and usually possess four distinctly separated glandular masses, commonly termed the quarters, from each of which protrudes a long teat. The four quarters are united to- gether in pairs and are arranged symmetrically. Between their bases and the yellow abdominal fascia they have a rich layer of fat. The udder is attached along the linea alba to the yellow abdominal fascia, and to the tendons of the abdominal muscles, by two layers of elastic tissue, the suspensory liga- ment (ligamentum suspense rium mammarum) which penetrates the udder between the two halves. Although the quarters situated on one side show no visible anatomical separation, injection tests with colored gelatin, and ob- servations in cases of inflammation of the udder in natural and artificial infections have proven that the secretory canal systems of the anterior and posterior quarters are separated in the same way as those of the opposite quarters. These canal systems collect into excretory ducts and terminal tubules and finally empty into the milk cistern, which in its upper part is greatly dilated and in its lower part is more constricted. Each quarter possesses a teat (6 to 10 cm. in length) from the milk sinus of which, the duct of the teat (ductus lactifera) of about 8 mm. in length, passes to the outside. The entire udder is covered by fine, slightly hairy skin, which extends posteriorly and supe- riorly into the escutcheon or so-called milk mirror. The size of the udder varies in the different breeds and indi- viduals. In the sheep and the goat there are two milk glands, each possessing a teat which stands out in a divergent direction from the one opposite. Each teat has one excretory duct. While the teats of the sheep are finely haired, those of the goat are hairless. The blood vessels of the udder are derived from the branches of the external pudic artery and anastomose with the various venous branches, through which the blood flows posteriorly through the perineal vein into the internal pudic vein and finally into the obturator vein. The greatest part of the venous blood flows laterally into the external pudic vein and anteriorly into the subcutaneous abdominal vein, which forms the immediate continu- ation of the external pudic vein and which is known as the milk Pathological Anatomy of the Udder. vein. It runs bilaterally of the median line, penetrates posteriorly and laterally to the xiphoid cartilage of the breast bone into the deeper parts and then empties into the internal thoracic vein. The lymph vessels which are very numerous enter two large lymph glands which lie bilaterally in a depression at the posterio- superior portion of the udder and are known as the supramam- mary lymph glands. The lymph passes thence to the lumbar glands and into the thoracic duct. The nerves originate from the lumbar plexus. The udder is supplied by the internal branch of the ilio-hypogastric nerve, the external branch of the lumbo-inguinal nerve, and the external spermatic nerve. In the goat the external spermatic nerve divides in the abdominal cavity into three branches, of which the median and the caudal branches pass through the inguinal ring to the udder. The cephalic branch passes to the abdominal muscles. The caudal branch (inferior) is purely a vascular branch. The median branch passes to the udder, and ramifies to the milk ducts and the teats. Pathological Anatomy of the Udder. Of the pathological processes which are of importance from a practical standpoint, the inflammations and changes which have more or less influence on the quality of the milk are of special in- terest. The other anomalies will be mentioned only briefly. Not infrequently there may be present in cows supernumerary teats, or supernumerary milk glands, which may be considered as a reversion to early stages in the evolution of cattle. Usually two rudimentary formations occur which are generally situated behind the posterior normal glands and normal teats. These may at times yield milk (Burkart, Dauberton). These accessory glands may also occur between the normal teats. In several instances as many as four supernumerary teats were observed. If the udder is abnormally small in its development or is en- tirely absent, it constitutes hypoplasia or aplasia of the udder. According to Bosetti the absence of the mammary gland was ob- served in a cow two and a half years old. Although there were four small teats on the skin, no milk was secreted even after the birth of a healthy calf. The milk veins were well developed on both sides. The opposite condition, hypertrophy of the udder, with or without secretion, is most conspicuous in male animals. Pusch ob- served a buck which produced 70 gm. of colostrum-like milk daily, and which possessed nipples 7 to 9 cm. in length. Grurlt has re- ported that the udder of a steer was as strongly developed as in a cow, and produced daily iy2 liters of secretion. It is known that newly born kids and suckling colts occasion- ally secrete milk for several days (Gurlt, Martin, Hess, Ibel). Schmidt, of Dresden, reported a giant udder with an entirely Anatomy, Pathology and Histology of the Mammary Gland. normal milk secretion, (16 liters). A functional hyperemia in the beginning of the lactation period increased the four quarters uniformly to such an extent that a day after parturition the udder touched the .ground with its central surfaces. Before and after parturition an abnormal amount of hy- peremia occurs physiologically in the udder (hyperemia conges- tiva). In inflammations the same condition may be present, the capillaries are abnormally dilated, and greatly distended with blood. This condition may result in the exuding of fluid and the solid constituents of blood. These are known as capillary hem- orrhages. In larger extensions of these hemorrhages they are spoken of as suggilations, and when the blood is contained in a sac-like cavity or swelling it is known as hematoma uberis. If in the congestive hyperemia the fluid constituents of the blood pass into the tissue of the udder, it results in edema of the udder. The same condition may develop as a result of hydremia, as for instance after changing from dry to sloppy foods (Bang), or as a result of multiple emboli of the blood vessels, or from a varicose condition of the veins of the udder. Edema of the udder manifests itself as a soft or tense swelling of the tissue, which retains the impression of the finger. While the teats usually remain normal on account of their slight but dense connective tissue, quantities of fluid collect in the front of the udder and between its glandular substance and the skin. The edema frequently extends posteriorly to the udder and up to the vulva. In- fections of wounds with the bacillus of malignant edema may result in edema of the udder. To those engaged in milk hygiene the most important of all pathological conditions of the udder are the inflammations which result from a reaction of the glandular tissue to any inflam- matory irritant. In most instances the inflammations of the udder are produced by microbian infections of various kinds, particularly by poly-bacterial infections. The bacteria penetrate the udder either by way of the blood circulation or from the outside through the orifices of the milk ducts. In such cases it is spoken of as a hematogenic or galactogenic mode of infection. If the infection results from a mixture of bacteria, and is not caused by one kind alone, the affection is a mixed infection. The infection may result from traumatic conditions when injuries extending into the paren- chyma of the glands make the infection possible, or from galactif- erous-traumatic causes when the infectious material enters the milk cisterns upon milking tubes or straws. The infection may take place also through simple contact of the orifice of the teat with the infectious material. Thus the different forms of mastitis, the peracute, acute or chronic inflammations of the udder may arise, depending upon the character of the infectious material and upon special accessory conditions. The possibility of galactiferous infection was first experimentally proven by Frank. The character and the varieties of inflammations of the udder were further established .by the work of Kitt, Nocard and Mollereau, Lucet, Bang, Hess and Borgeaud, Guillebeau. Zschokke, Sven Wall, and others. Pathological Anatomy of the Udder. The principal producers of mastitis are the colon-paratyphoid group, staphylococci, streptococci, Bacillus pyogenes bovis, Bacillus tuberculosis, and the actinomyces. Colon infection and severe mixed infections usually result from galactiferous contact, or after the introduction of milking tubes, straws, quills, cat-guts, and hairpins. Highly acute, inflam- matory conditions develop in the affected quarters, whether af- fected throughout or only partially with parenchymatous mas- titis. Hot, painful swellings of the quarters, with collateral edema Fig. 1. Acute inflammation of the right forequarter with collateral edema. (After Kitt.) of the surrounding tissues, are the associating symptoms of this form of inflammation, which either results in recovery with atrophy of the affected parts of the udder, or with regeneration of the epithelia destroyed by the disease or on the other hand the di- sease becomes chronic and may even terminate with complete gan- grenous and ichorous destruction of the affected part of the udder. In the infectious forms of mastitis the supramammary lymph glands may swell to fist-sized nodes. If the process becomes chronic a suppurative softening of the affected parts of the tissue, or a suppurative demarcation of ne- crotic parts of the tissue results. These conditions are designated as suppurative and purulent mastitis respectively. 6 Anatomy, Pathology and Histology of the Mammary Gland. The acute forms of mastitis interest those engaged in milk hygiene but little, since noticeable changes in the milk quickly fol- low the commencement of the inflammation, and the animals soon stop their secretion. On the other hand the hidden forms of inflam- mation are of the greatest importance because the milk is fre- quently almost unchanged, and does not always indicate its ined- ible condition. Such conditions of the udder may vary from a simple catarrh to a purulent inflammation. The manifestations of these forms of inflammation vary to a great extent, and the symptoms may be only slightly pronounced, so that a single clin- Fig. 2. Fibrinous form of parenchymatous mastitis; separation of quarters plainly visible. (After Kitt.) ical examination may cause a suspicion, but a positive diagnosis cannot always be established. Literature shows that slightly marked swelling of the affected quarters, increased local temperature, nodular formation of the parenchyma, and induration of the glandular tissue, may appear in the most varied forms, sometimes with and sometimes without general symptoms. At the beginning it may be localized around the base of the teats, but the hardening of the glands then pro- gresses forward, upward and backward (Sven Wall). The examination of the milk ducts should not be neglected. The mucous membrane of the cistern may have become inflamed, Pathological Anatomy of the Udder. resulting in ulcerations, scar formations or polypoid prolifera- tions, which are difficult to recognize. Sometimes such changes of the teats are characterized by cicatricial contractions (strictures). The udder, which usually becomes affected in the individual quar- ters, may remain either normally soft, or may become somewhat harder in consistence. The yellowish-red, normal color of the cross-section disappears, and changes into a grayish-orange or brownish-gray tinge. The parts which are of a harder and tougher consistence show an increase of connective tissue; the interstitial connective tissue changes into a bluish-white thickened network. Fig. 3. Purulent mastitis showing necrotic foci. (After Kitt.) The edema of the skin which develops at the beginning of the inflammation results sometimes in extensive sclerosis, even the parenchyma of the glands being sometimes dislodged by the pro- liferating connective tissue causing the quarter to atrophy and harden. Tuberculosis although almost invariably resulting from a hematogenous infection, appears either in the form of a single focus (tuberculosis uberis circumscripta), or it may be dissemi- nated over the entire parenchyma (tuberculosis embolica dissem- inata), or the tissue may be diffusely affected, becoming infiltrated throughout almost its entire extent (tuberculosis diffusa). These 3 Anatomy, Pathology and Histology of the Mammary Gland. forms of the disease may be present in combination in the same udder. During the tuberculous invasion nodular indurations of the tissue develop, which hypertrophy and become tense, hard and knotty. The lymph glands usually manifest hard, painless, thick- ening, and nodular formations. Caverns may also develop in tu- berculosis of the udder. Actinomycosis which commonly develops from the penetration of actinomycotic barley beards, or particles of straw into the tissue, or more rarely by embolic infection, may also be produced experimentally by the injection of solutions containing actin- omyces through the milk ducts. Actinomycosis of the udder has been observed in cows by Peterson, Rasmussen, Bang, Harms, and Jensen. Nodular formations, connective tissue proliferations and softening of the tissues, localized or in larger areas, are also ob- served in this disease. Botryomycosis and glanders enter into consideration only so far as the udders of mares are concerned. For completeness, various growths may also be mentioned as anomalies of the udder, such as fibroma, adenoma, adenofibroma, adenocareinoma, chondrofibroma, chon- droma, lipoma, sarcoma, angioma, etc., which are dependent on the tissue elements and the character of the tissues of which they are composed. Cystic formations have also been observed. Not infrequently the connective tissue and the subcutis of the udder of cows may show bone formations in the form of bony hooks and plates, (ossificatio plana or racemosa). Parasites have also been found in the udders of cows, namely echinococci (Behmert and Steuding). For further information see Kitt, Pathol. Anatomy, 1910, Vol. 1, page 280. The author once concluded that a goat affected with adenoma papilliferum uberis was troubled with mastitis, basing this decision upon an examination of the milk, although the secretion contained no specific inflammatory agents. The continually increasing quantity of milk was remarkable. Postmortem and histological examination finally revealed the adenoma in the udder. Structure of the Tissue The external skin of the teats possesses neither hair nor sebaceous or sweat glands, and continues as cutaneous mucous membrane into the milk ducts, which it lines up to the cistern. The mucous membrane has no glands, possesses fine folds running lengthwise, and is covered by pavement epithelium which is supported upon a well developed papillary base, and is firm and horny next to the lumen. The papillae are extraordinarily long: they apparently branch near the base, and slant towards the orifice of the teats. This cutaneous mucous membrane of the milk ducts continues without demarcation, with the mucous mem- brane of the milk cistern, which is covered by several layers of cylindrical epithelium, and possesses accessory glands which are lodged in the connective tissue. The wall of the teats contains bundles of involuntary muscles running lengthwise and crosswise, forming a strong and elastic encasement around the canal of the teats. The supporting structure is penetrated by numerous blood Structure of the Tissue. 9 vessels and lymph vessels. Numerous and strong elastic fibres strengthen the dense fibrillar connective tissue of the teats. In order to describe the finer structure of the parenchyma of the udder it is necessary at first to touch on the further develop- ment of the organ from birth until the moment of the appearance of the secretion. (a) Normal Appearance. The milk gland is an organ which performs increased func- tions only at certain times. It does not secrete during the entire Fig. 4. Vertical section through the lower end of the teat canal which is closed by a horny plug (a). life but only when the newly born offspring is to be nourished by the milk. The udder of a virgin animal does not correspond even in its finer structure, with the appearance of a fully secreting udder, and this again varies in its finer structure from a gland which is at the beginning or at the end of the lactation period ; even this is not all, since the microscopical appearance changes in ac- cordance with the condition of activity, where a lobule or only a part of the lobule may be found on examination, depending whether the cell-complex is just forming the secretion or has al- ready discharged its secreted product. The gland of a newly born calf shows but relatively few cell tubes and cell buds, imbedded in connective tissue rich in fat and branching in all directions. These prac- tically form the basis of the glandular ducts and are without alveoli. The end of the tubes is frequently somewhat dilated, or thickened in the form of a club. 10 Anatomy, Pathology and Histology of the Mammary Gland. With puberty the alveoli appear in the cow surrounded by strong connective tissue. In older virgin individuals they sometimes show a slight amount of secretion. A considerable increase of the glandular tubes appears only after the first conception. The tubes become more dilated and branch more and more, forming alveoli, from which other ducts bud out. Although indications of secretions in the cells may not yet be visible, the cavities contain a homogenous or fine granular mass of cells or cell fragments. The gland prepares for the secretion, growing at the expense of the atrophying or expanding connective tissue, until ready to commence its secretion. Fig. 5 Superior portion of the teat canal (a) with a reflection of the cutaneous mucous membrane of the cistern (b). The cell lining of the larger glandular ducts is of double layers, as in the cistern, while that of the smaller ducts and al- veoli is composed of only a single layer. The epithelium of the latter appears cubical or flat, while the upper layer of the former is cylindrical; in the deep layer the cells are more cubical and rounded, partly wedged in between the bases of the superficial cylindrical cells. The borders of the cells are sharp and the proto- plasma is clear. The nuclei of the epithelia frequently show mi- tosis, that is, division and multiplying forms. The cells rest on the so-called basket cells and the membrana propria. The basket cells Structure of the Tissue. 11 should be considered, according to the investigations of Benda and Bertkau, as involuntary muscle cells because of their appearance and their staining qualities. They probably play a part in the emptying of the glandular ducts and the milk secretion. Blood capillaries, lymph vessels and nerves run in the inter-and intra-lobular con- nective tissue, which is strengthened by elastic fibres, and contains involuntary muscle cells. Therefore, the same tissue elements are represented as in the teats, with the ex- ception of the many-layered pavement epithelium. Fig. 6. Structure of the mammary gland in secretion, Hematoxylin. 1 X 800. (a) Secreting glandular alveoli, (b) Alveoli with dormant cells. At the end of pregnancy the picture again changes consid- erably. The protoplasm of the previously clear epithelial cells of the secretory system becomes cloudy, the nuclei larger, their chro- matin collects in flakes on the periphery of the nuclei, the borders of the cell become indistinct, the cells become swollen, the nucleus lies in the center, and the indications of the division by indirect fission of the nucleus appear relatively in groups. Some epithe- lial cells show two nuclei at this stage; towards the alveoli fat globules appear. Leucocytes with which a few eosinophiles are mixed, collect beneath the epithelial cells and penetrating the 12 Anatomy, Pathology and Histology of the Mammary Gland. epithelial layer, separate themselves from the epithelial cells and enter the alveoli, which at this stage contain fatty secretions, leucocytes and epithelial cells in all stages of degeneration. With these manifestations the gland cell commences its function. The desquamation of epithelial cells and the cell de- generation disappear; the cellular infiltration of the connective tissue recedes until it is very slight between the now greatly di- Fig. 7. 2. Chronic mastitis of cow. 1 X 800. (a) Thickened interstitial tissue, (b) Alveoli, (c) Blood vessels. (1) Epithelial desquamation. (2) Colostra! bodies. (3) Cellular infiltration. (4) Fatty degeneration and necrosis. (5) Milk concrement. lated and distended glandular ducts. The cells are finely granular on the basilar border, and at times show striation, that is, fine streaks running in parallel directions (bioplasts according to Altmann). The nucleus is large and vesicular in shape ; the upper part of the cell is granulated and shows large and small fat globules. This granulation and streaking may be seen, according to Steinhaus and Duklert, at each act of secretion. The fine fat globules collect Structure of the Tissue. 13 into larger ones, which are only separated from the lumen by fine protoplasm, or having been expelled have already entered the al- veoli. With the collection of the secretion these dilate, the cell becomes flattened during the expulsion of its products, and the part lying towards the lumen appears indistinctly bordered as if shredded after the expulsion of the fat. They soon become smooth again, and by the pressure of the alveolar contents and the dilation a Chronic mastitis of cow. 1 X 90. (a) Healthy portion, (b) Glandular portion with chronic mastitis. of the alveoli, the cells sink and become so flat that the nuclei not infrequently appear bulged out towards the lumen. After the expulsion of the secretion the formation of additional secretion again commences in the cell, the protoplasm again becomes cloudy and granular, and so on, a continuous change of the form of the cell taking place. During the entire lactation period, but more so in the later stages, manifestations of atrophy of the gland appear, at first 14 Anatomy, Pathology and Histology of the Mammary Gland. commencing at the base of the gland, and finally during the end of lactation in the entire udder. Epithelial cells are thrown off, the alveoli become fewer, smaller, and irregularly distended, the con- nective tissue increases, and cellular infiltration starts under and between the epithelial layers. The epithelium contains no fat globules, it is sharply bordered towards the alveoli and the pro- toplasm becomes pale. Finally the last remains of the secretions Fig. 9. b. a. b. a. Acute streptococcic mastitis of sheep. 1 X 1000. (a-1) Blood capillaries. 2. Thrombosis by disseminated streptococci, (b) Glandular alveoli, with clumps of streptococci. (c) Migration of leucocytes into the infected alveoli. disappear, the plasma cells and leucocytes taking care of the resorption. At the end of this process the gland is at rest, and the cow is dry. Of course these processes are not always so schematically uni- form as they have been described. During the entire lactation period, colostrum-forming, and retrogressing lobules may be ob- Structure of the Tissue. served; likewise certain parts of the udder may remain in secre- tion during retrogression until storing of the secretion, leucocytic resorption activity and connective tissue proliferation cause them to cease their activity. (b) Pathological Appearance. Any kind of irritation of the gland, such as stasis of the milk, especially in chronic catarrhs and inflammations, may result in the most varied kind of pathological conditions, either in mixed form or individually. The manifestations vary, depending upon whether degeneration and destruction of the tissue, or reparation and recovery gain the predominance. Sometimes desquamation of epithelium, with or without fatty degeneration, occurs together with cellular infiltration of the in- terstitial connective tissue and capillary engorgement as the only indications of inflammation ; or, on the other hand, the changes in the interstitial parts may be very pronounced, while the changes of the parenchyma may be less prominent. The inter- and intra- lobular connective tissue extends forming thick indurations, from which the separated epithelium is compressed to small necrotic nests. In other stages of inflammation the cellular infiltration of the tissue predominates. The alveoli and the milk ducts are plugged up thickly with leucocytes, and dilated with the pus. In highly acute inflammations the rapid breaking down of cells, de- struction of epithelium, serous and cellular infiltration of the tissues even to their dissolution, are the principal manifestations. The ducts and the alveoli are inundated with serous, bloody co- agulated masses. In stasis of the milk, and in all inflammatory manifestations, especially of the acute form, the alveoli contain hyalin and con- crement arranged in layers, in addition to inflammatory cells and broken down cellular products. CHAPTEB II. PHYSIOLOGY OF LACTATION AND CHARACTERISTICS OF MILK IN GENERAL. As already mentioned the udder secretes only in certain lactation periods between births. The lactation lasts under nat- ural conditions in healthy animals as long as the young needs the glandular secretion for its nourishment, and stimulates the lactation by the irritation of the intermittent suckling. Shortly before parturition, or at the time of parturition, the glandular tissue terminates its increase in development, and the milk secre- tion starts and becomes actively established. The causes of the increased cell production during pregnancy, and for the secretion after this time, are variously explained. Nervous irritation from the genitals to the milk glands may by means of reflex action stimulate the glands into activity. That such reflexes on the genitals may originate from the milk gland is proven (Pfaundler). Reflex actions in the opposite way, however, have not been proved (Halbau). It has been impossible either experimentally (extirpation of the lumbar cord) or by accident (fracture of the spine), to pro- duce a complete " nervous isolation," since as emphasized by Pfaundler, there are still remaining the nervous connections through the vasomotors. However, the re-section of nerves, oper- ations on the spinal cord, transplantation experiments, etc., by Eckhardt, Rohrig, Sinety, Busch, Mirnow, Piaster, Ribbert, Golts and Ewald would suggest that besides the nervous influences, which undoubtedly exist, there must be some other agent which stimulates continuous growth during pregnancy, terminates the same with the end of parturition, and inaugurates the secretion. Hematogenic influences may be readily accepted, as they may be led to exert their action either by the quantity or by the quality of the blood. After parturition the body and the milk gland have at their command great quantities of blood which was previously utilized by the gravid uterus. The plethora which appears at this time may be held responsible for the inauguration of the secretion, after the udder has been rendered ready for action by the increase 16 Milk Secretion. of its growth through nervous influences. On the other hand it has been observed that in other conditions, in which there exist also a diversion of great quantities of blood from the genital parts for the supply of other organs, as for instance after operation on very large tumors in the region of the genital organs, no secre- tion appears even when the udder is prepared for the secretion. As a matter of fact the secretion may commence before birth, and even in early abortions, or if the fetus dies. At times when the uterus is only so slightly distended that the quantity of blood set free after abortion is hardly sufficient for an effective hypere- mia of the milk gland, the secretion of milk may result (Sinety, Kreidl, Mandl). Therefore the explanation that the quantitative influences of the blood may give rise to a stimulation of the milk secretion (Freund), can scarcely be accepted. Consequently the qualitative changes of the blood must be considered as more prob- able factors. Authors have diversified opinions upon this question. While some accept the view that substances are eliminated from the impregnated organs, or by the fetus itself into the blood of the mother by internal secretions, and that these act as stimu- lants on the milk glands, others believe that the factors causing lactation lie in the assimilation of certain nutritive substances. The supporters of the theories of "stimulation substances" (Sinety, Halban, Starling) take the stand that stimulating sub- stances which cannot be utilized for the cellular growth and cellu- lar activity, contrary to the nutritive substances, cause the devel- opment of the gland during pregnancy, and at the same time pre- vent it from secreting (stimulines, hormones [I stimulate], sub- stances of pregnancy). Development of the gland and prevention of secretion may, of course, be the action of one and the same sub- stance (Hildebrand, Starling), or its development, as long as the growth continues, may retard secretion. With birth the stimula- tion of growth and development ceases, and secretion commences. Contrary to this, the theories of nutritive substances empha- size the fact that the glands at times may start the specific activity without the presence of certain stimulines, probably through nutri- tive substances which are present in the blood at various times. Eauber attempts to explain the activity of the gland after birth by declaring that after the expulsion of the fetus a nutritive material becomes available, which has served prior to birth for the preparation of nutriment for the offspring. While the ex- planation of the author that the lymph cells play the most impor- tant part in this can no longer be considered, still it furnishes the basis for all new theories relating to the action of nutritive substances. These views were strengthened in 1908 by Schein by the state- ment that during pregnancy the mother animal, in order to meet the requirements of the fetus and of the impregnated organs, en- 18 Physiology of Lactation and Characteristics of Milk in General. riches her blood with the so-called "milk producing substances." Pfaundler recommends the designation "offspring nutritive pro- ducing substances." Since during pregnancy the continuously developing placenta utilizes and consumes these substances for use in the nourishment of the young, there remain for the milk gland only slight remnants, just sufficient to result in the necessary stimu- lation for the cellular increase in the gland. After parturition when the activity of the placenta is completed, the milk gland takes up the released nutritive substances for its own use (specific af- finity of the substances to the cells of the milk gland), and is stimulated to secretion by the quantity of the disposable material. Schein's milk producing substances in the blood constitute the initial material for the formation of specific components of the milk, milk sugar, casein and milk fat. The material acquired by the mother, through placental con- tact with the fetus, while aiding in the development of the latter is also of benefit to the activity of the milk gland, whose product adapts itself exactly to the requirements of the young, as far as it concerns the material which the young uses for the growth of its body. If conception again takes place the developing placenta of the new fetus enters into competition with the lactating gland, and draws from it milk producing substances for its own use, whereby the secretion of the milk gland becomes reduced or ended. Influences exerted on the milk gland by oestrum or puberty, and also the impulse of pregnancy, have not yet been sufficiently explained through this theory. Pfaundler enlarges upon and ex- plains these phenomena by stating that the withdrawal of certain nutritive substances, through the germinal gland, embryo and ovum, and not the appearance of milk producing substances alone, periodically disturb the equilibrum of physiologically acting sub- stances in the blood, and thereby the antagonizers of those sub- stances (the stimolines, harmones of other authors), are enabled to find specific receptors (affinities) in other organs of the genital apparatus. After birth, continuing intermittent stimulation may retain or increase the lactation of the milk glands for a longer or shorter time. Stasis of the milk diminishes and retards the secretion. Rievel opposes Schein's view, since in his opinion it does not explain how udders of animals in which neither pregnancy nor birth has preceded, could start secretion (lactation of milk glands of the newly born or virgins, occasionally even of male animals). According to the author's view these facts would not oppose the theory of nutritive substances. Schein, himself, aims to bring these observations into harmony with his views, and asserts that the newly born may give a secretion from their milk glands, when towards the end of pregnancy the activity of the placenta is dis- turbed, and as a result small quantities of the "milk producing Milk Secretion. substances" enter without changing directly into the blood of the fetus, and thence into its milk gland. Sufficient stimulation for the secretion and formation of the so-called "witches milk" re- sults. Schein explains the formation of milk in virgin mammae, or in milk glands of individuals which have passed their climacteric, by the fact that through the stimulation produced by sucking, the secretory cells are awakened from their dormant state and then utilize the milk producing substances in the blood for the perform- ance of their functions. Finally (1910) he concludes that the oc- currence of milk secretions in nullipera and in wromen who have passed the climacteric, which differs from the gradually inaugu- rated normal lactation as a result of pregnancy, and also the ob- served secretion by the breasts of newly born and of male indi- viduals, represents a continuous secretion analogous to the normal secreting process in other glands, in which the product is as a rule, however, re-absorbed by the glandular elements. In pregnancy and at birth the secretion is increased to the greatest extent, but other stimulants may under certain conditions stimulate the activity of the gland. Duval's more recent observations contain data relative to the occurrence of milk secretion by women outside of their normal lactation periods. It is not uncommon to observe secretions in virgin animals especially when young animals which are present stimulate the udder intensively by sucking. It should be emphasized however that the udder secretion of virgin animals distinguishes itself in its appearance from the milk of mature milking animals ; it repre- sents a secretion which does not even deserve the name of milk. The experiments which were conducted by various authors in support of their lactation theories appear of interest. The experiments of Starling aim to show the presence of bodies in the blood during pregnancy which prevent secretion, in which claim is made that an interruption of pregnancy in rabbits at a time in which alveoli capable of secretion were not yet present, led to a retrogression of the milk gland, while in the later periods of pregnancy secretion was induced. According to Pfaundler's view the harmone theory could be effectively supported by the fact that an existing secretion may be successfully interrupted or prevented by the introduction of serum of pregnant animals of similar species. The author does not believe that this proof is satisfactory and mentions observations made in a case in which the secretion ap- peared at birth of twins which were born at long intervals, thaHs, the pregnancy continued after the first birth, yet the milk secretion continued unchecked. Wucherer observed a case in which a sow gave birth to nine, and seventeen days later to six other pigs. At the birth of the second lot the first born pigs were taken from the sow. These continued to thrive, but of the second lot only three remained alive. He emphasizes the opinion that a transitory 20 Physiology of Lactation and Characteristics of Milk in General. action of blood serum, as used in Pfaundler's experiment, which corresponds only slightly in its composition with the normal blood serum, can never be favorably compared with natural influences in the body. This exception must hold also for the indecisive ex- periments of Starling, who by injections of juices from rabbit em- bryos, but not with injections of preparations from rabbit ovaries, placentas and mucous membrane of the uterus, produced a devel- opment of the glands, and at times a degree of milk secretion. He believes that the true cause of the secretion may be found in the chemical changes which are produced by the growing embryo and are brought to the glands through the placental circulation. Ac- cording to Basch, secretion may be established in the mammary glands of virgin rabbits by injecting them with placental extract (serum of unlike origin, from man), which was so powerful that it also brought on a secretion of milk in mother animals without the intervention of pregnancy. The placental extract could induce the secretion only when the teats of these animals were stimulated to hyperplasia by the implantation of ovaries from pregnant animals. According to the author's observations these questions can only be determined through experimentation, when by uniting two female individuals of like species a basic condition is established, by which the activity of the glands of one of the impregnated indi- viduals as a consequence of its pregnancy may be observed upon the other, and the result of the impregnation of the latter on the lactation of the first mother may also be determined, Such experi- ments have already been made by Cristea of Vienna, by coliotomy of a virgin and a pregnant animal, and uniting both by suturing of the peritoneum, the musculature and the skin, the author establish- ing a double individual, united by a broad peritoneal communica- tion. Of eighteen such pairs (rats and rabbits) six remained alive. In the experiments after parturition of the gravid animals the milk secretion also appeared in the virgin animals to which they were united. Cristea therefore believes in a slow transition of a secretion from the gravid animal into the non-impregnated animal, namely by the way of the lymphatics, since there existed no blood vessel union between the individual animals. With this result the hypoth- esis that the changed distribution of the blood after birth pro- duces the milk secretion collapses, since on account of the lack of communication of the blood vessels it is not possible that an in- creased blood supply of the mammae of the virgin animal would result from parturition of the attached animal. It can make no difference whether milk producing substances or substances which are not assimilable and are not consumers of energy (stimulating and inhibiting substances), stimulate the glands to activity. Recently Basch observed an abnormal birth to one of a pair of twins (the Blazek sisters showing a condition of pygopagus, union of the pelvis and sacrum with a common introitus vagina?, and a common rectum), in which after the birth of a child to one, lacta- Milk Secretion. 21 tion commenced also in the virgin sister. In this instance nervous connections may exist in the genitals of both individuals. Accord- ing to the author's view this case is not an absolute proof of the stimulation of the gland by hematogenic means. The lactation theories may be laid aside, and consideration only be given to the fact that at birth, puberty, pregnancy, at the conclusion of parturition and also in the disturbances of the gen- itals influences are exerted on the milk gland the character and action of which are still uncertain, although the results manifested by the production of milk may readily be observed. Especially typical and striking are the phenomena seen at puberty and during and at the end of pregnancy. Exceptionally a condition may ap- pear outside of these normal periods of the organs in females, and in single cases even in male individuals, which permits the conclu- sion that the glands react to special stimulation. Abnormalities may occur in the anatomical structure of the gland, pathological manifestations in the sense of inflammatory reactions, etc., may also be observed, and exceptionally the usual functions may be present or may develop, without their being accompanied by gross anatomical changes of the gland ; these however are usually pres- ent at the same time. These influences on the gland originate partially in the gravid genital organs and the fetus; in other instances the germinal glands and the disturbances of their functions are the cause of these influences. Such influences may be classed according to the impulses which lead to glandular activity, as follows (Halban) : 1. Embryonic impulse — action very transitory — mastitis neonatorum — witches milk. 2. Puberty — lasting effect — development of the gland. 3. Oestrum — action rapidly transitory — hyperemia, inter- stitial hemorrhages, disturbances to physiological lactation, secre- tion. 4. Impulse of pregnancy — lasting between parturitions. Lactation may be sustained for a long period of time by the regular drawing of the milk, and ceases in healthy udders only when after frequent and absolute stasis of the milk (after about eight days), the tissue becomes affected by inflammatory irrita- tions (absorbtion and change of the condition of the epithelium), or when the animals are soon to give birth to young. If no re-im- pregnation takes place the lactation period may last longer, even from one to two years although not to an unlimited extent. The activity of the gland may be retained for a long time through the sucking of the young, stimulation by milking, or artifical with- drawal of milk. Frequent periodical and complete emptying of the milk cis- terns acts favorably on the amount produced. In the cow two to three milkings per day are sufficient to retain the udder in secretion. 22 Physiology of Lactation and Characteristics of Milk in General. The milk formation occurs between the milking periods and during the milkiiigs ; therefore of these two periods, the first lasts for many hours, the second with more intensive production is com- pleted in a few minutes. The first phase is the work of continued activity of the gland, the second is brought on under the stimula- tion of the sucking, or milking, on the secretory nerves, and as a result of the increased blood supply (stimulation of the vasodila- tors). The capacity of all the milk ducts of the udder represents less than half of the quantity of milk obtained in one milking. According to Fleischmann the volume of the entire udder of a cow with the teats is 6700 c. c. Of this 3000 c. c. is represented by the cavities ; the secretion obtained in one milking may never- theless amount to 7000 c. c. Niiesch substantiates Fleischmann 's statements by an experi- ment; a cow gave daily before slaughter 10 liters of milk of which 5 . 5 liters was the amount of the morning milking. After slaughter before milking in the morning 2.7 liters of milk could be proven in the udder (catheterization and calculation of the amount remaining in the udder), which proves secretion during the process of milking. The two phases may be considered as though the glandular cells which tire after the milking gradually recover (increased blood supply) and recommence their secretions. The collecting secretion will increase until a certain relative pressure between the collective quantity of secretion and the tissue with the blood vessels is established, when the secretion is retarded or ceases until renewed stimulation of the glands by milking, emptying, massage, (electric irritation), or stimulation of the central nervous system from milk accumulations causes the milk to fill the cavities of the udder again. If the usual milking time is omitted a flow of milk may result, that is the pressure under which the secretion is held finally over- powers the resistance of the sphincter muscles at the opening of the teats (directly or by reflex), whereupon formation of milk again takes place. Nervous influences on the secretion are exerted by the sper- maticus externus and by the sympathicus. Experiments which were conducted for the study of the ener- vating influences on the secretion produced contradictory results. Rohrig severed the ramus inferior of the nervns spermaticus externus (vessel branch), and observed an acceleration of the secretion, while the severing of the glandular branch (part of the median branch) resulted in inhibition. Eckhard failed to observe any influence on the quantity of milk after the severing of the ner- vus spermaticus extermis. Heidenheim and Partsch demonstrated an increase of the quantity of milk from the cutting of the nervus spermaticus externus, but only when strychnine or curare had been administered at the same time (test by Sinety on guinea pigs). Milk Secretion. 23 Although Basch could not establish a quantitative increase by sev- ering the nervus spermaticus oxternus, he found qualitative changes (formation of colostrum). Pfaundler concludes from these and other experiments that an action of the peripheral nerves on the development of the gland and its functions, especially from a qualitative point of view, must figure in the consideration, but that these influences have only slight importance. Insignificant as well in their results on the secretion were the severing and re-sectioning of the spinal cord, or interference with the sympathetic system. Basch again observed the formation of colostral milk after re-sectioning of the coeliac ganglion. From this he concludes that the regulating influence of the nervous system exists through reflex action, especially from the sympa- thetic, but that at the same time the gland is also capable of independent secretion. As a matter of fact far reaching influences of a nervous character are observed. 1. Psychic influences. 2. Eeflexes, which are caused by local stimulations (sucking — milking — electrical stimulations, etc.). 3. Reflexes from the genital region. These points are only briefly mentioned here, since the various conditions will be discussed in subsequent chapters, when consider- ation will be given to the quantitative and qualitative changes which appear under varying influences. An active part in the emptying of the milk from the cisterns, and in the passage from the upper part of the duct and alveolar sys- tems, is played by the sucking and pressure exerted during the. milking (pressing outwards, sucking from the gland), massage of the udder (pressing out into the cistern), the contractility of the tissue (elastic fibres, involuntary musculature, filling of the blood vessels), and the vis a tergo of the newly formed secretion. CHAPTER III. MICROSCOPY OF MILK IN GENERAL. If milk is examined through a microscope one chiefly sees numerous small fat cells floating in the fluid or milk plasma. These will be considered later, but at first the cells and cell fragments originating from healthy and affected udders will be discussed. Between the milk globules, by which term the small fat droplets are designated, bodies may be seen which are hard to define unless stained. After special treatment, however, they may be readily recognized as cells or their fragments, or as a precipitation of soluble or suspended substances. Since the external skin of the udder, and the lining of the milk passages and milk secretory ducts in the udder are of similar for- mation, we naturally are only concerned with the upper layers of pavement epithelium, cylindrical epithelium, and the deeper cubical epithelium of the terminal ducts and alveoli, and only in severe tissue changes would cells of other parenchymatous parts appear in the milk. Naturally in such an actively working organ, even in a physiological normal condition, leucocytes of the most varied kind, and even red blood corpuscles may be found. In cases of special stimulation from physiological or pathological causes, the resulting cell mixture may be of a most varied character depending upon the location of the stimulation, and its quality and duration; hence at times certain leucocytes, and again red blood cells or epithelia, may predominate in the mixture. 1. Cells from compound pavement epithelium. Following the intensive manipulation and stimulation of the teats by milking, the appearance of cells from the upper layers of the pavement epithelium of the outer skin, and the milk ducts is natural. As a matter of fact in the fresh milking periods during which irrita- tion from the extraction of the milk is especially evident, the milk always contains fine folded platelets of round, oval, or irreg- ularly distended and curved borders, which frequently when folded in several layers, appear as small clasped cysts without special structure. These bodies have been described by Winkler, and were con- sidered by him as indications of pathological changes. The author 24 Cellular Content of Milk. 25 took a stand against this view of Winkler, as he had observed them in the milk of entirely healthy animals, but not until the present time has he been able to offer an explanation of the nature of these bodies, designated as "skinlets" or " shell." They represent desquamated cells of the stratum mortificatum of the pavement epithelial layers singly or in groups. Although usually no parti- cular structure is manifested yet in single instances typical flat, round nuclei can be seen. If the teats of a slaughtered cow are taken and the cistern and milk duct are carefully cut open, and from the surface of the milk duct a small quantity of the cellular layer is scraped Fig. 10. off, an examination by the usual method discloses the typical "shells." 2. If cells from the cistern are prepared and examined, elongated or oval, or quadrangular cells with oval nuclei, frequent- ly elongated at the base, will be found, singly or in groups. Single fat drop- 1 e t s may frequently be seen in the plasma sur- rounding these cells. Sim- ilar cells may also be found in normal milk. They are usually single, although sometimes united in groups arranged like flow- ers. In stimulation, which brings on a desquamation from the mucous mem- brane of the cisterns, or from the parenchyma in catarrhal conditions of the milk passages, they of course appear in masses. Such reactions occur in the cistern for instance as a result of the so-called kneading. 3. Cells from the secreting milk ducts and the alveoli, ap- pearing large or small according to the quantity of fat globules collected in them, often become tremendously distended and bloated (foam cells). Their structure is mostly honeycombed or mulber- ry-shaped when they contain fat ; without fat the cell is surrounded with only a narrow^border of protoplasm. The nucleus is usually in good condition. Film of sediment from milk of a fresh milking cow. Cells from the stratified layer of pavement epithelia of the teat canal. Thionin. 1 X 1000. 26 Microscopy of Milk in General. These cells are the large colostral bodies. They are in their entire structure and in their staining characteristic epithelial cells and not leucocytes ; the amoeboid movements observed in them, if these observations were beyond questioning, do not prove that all colostral bodies represent leucocytes. This point will be again taken up during the discussion of colos- trum. While such cells only appear occasionally in ripe milk they are extremely numerous at the beginning and termination of secre- tion, and in pathological processes, in the latter especially in sub- acute and chronic forms, but not in peracute and acute inflammatory conditions of the paren- chyma. Such cells may occasionally be noted collected in groups. The author believes that their appearance in masses in the milk, that is, the condition increas- ing the expulsion of these epithelia, results from the fact that each cell, which in its singu- lar activity precedes or follows the other cells of the union, becomes desquamated. It does not correspond func- tionally, with the other cells, and is therefore removed from the rows of cells which are de- veloping for a definite purpose or are working for that purpose. Only when uniform work is performed by all of the cells working in unison, .„ and bringing about a uniform condition, will the organ cease to free itself of incapable elements. In inflammation the inflammatory irritation and its consequences soon drive the cells to overproduction. At other times it paralyses or destroys them, even before the formation m?' 5epending on the d.uration of the inflammation. Ihe form of the epithelium varies in accordance with the con- tent ot fat The collection of fat is not the result of fatty degenera- tion, but is produced when the cell is thrown off before its time for secretion, or while still capable of taking up material and produc- ing tat but without strength for the separation of fat. Therefore Cells from the lining membrane of the wall of the cistern Sediment in catarrh of the cistern. Thionin. 1 X 1000. ' Cellular Content of Milk. 27 such cells may be found even in the epithelial groups, which is an additional proof that they with certainty represent epithelial cells. The cells are from 5 to 25, even to 47 p> in size (Schulz). Not infrequently 2 to 3 nuclei of oval or roundish shape are present. The author has never observed more than one nucleus, and be- lieves, with Popper and Schulz, that the appearance of more than one nucleus results from two cells lying on each other, in which case the cell thus formed may appear to possess two or more nuclei. Migrated macrophages may also simulate a polynuclear appearance. Notinfrequentlyepi- Fig 12< thelial cells are thrown ' off, with a single large fat globule in the body of the cell, known as " seal-ring cells." In such cases the fat glob- ules have a "moon" or "cap" appearance. 4. Leucocytes of all forms are frequently met with in milk such as mononuclear basophiles, eosinophiles, polynuclear basophiles, acidophiles, or cells with neutrophilic and eosinophilic granules in the protoplasm. If the polynuclear cells show no nuclear bridges, they may be found with % or more spherical shaped nuclear granules (spherical gran- ule polynuclear leuco- cytes, Babs.) The nu- cleus is usually in the shape of a ribbon, or clover leaf, or heartshaped. The protoplasm usually contains fat globules, which in stained preparations appear as fine vacuoles. The lymphocytes are small cells with round nuclei and a very small border of protoplasm. According to Schulz they never con- tain fat. Large mononuclear leucocytes are also supposed to be present in the milk. If they gorge themselves with fat they are filled to their fullest extent, and can no longer be distinguished from fat-containing epithelial cells. 5. The red blood cells may be seen as small, round or thorn- Sediment in milk of a cow after milk stasis. Numer- ous desquamated epithelia, among these an "albu- minophore," and polynuclear leucocytes. 28 Microscopy of Milk in General. apple shaped bodies, with metachromatic staining substances. They may be readily recognized as erythrocytes. 6. Degeneration of these various kinds of cells may result in the finding of the most peculiar formations. The protoplasm of the epithelial cells becomes shredded ; the nucleus splits up and eliminates its chromatin into the plasma in the form of dust or flakes. It diffusely passes into the cell pro- toplasm, which appears darkly stained, and in the place of the Fig. 13. The formation of large colostral spheres and desquamation of "seal-ring cells." 1 X 800. nucleus a pale vacuole appears. If the breaking down continues there may appear a disintegration of the cell and of its nucleus into small droplets and fragments of roundish appearance, either with or without a lightly stained border around a small darkly stained center of chromatin (Heidenheim, Cohn, Popper, Schulz). These chromatin flakes are probably identical with the so-called free nuclei (Michaelis), which were also observed by Lenfers. The flakes which result from chromatolysis have been des- Cellular Content of Milk. 29 ignated up to the present as "Nissen's Globules." According to Ottolenghi they are derived either from leucocytes or from epithelia. If fat-containing cells break down in this manner, fat globules in the shape of grape-like bunches, and single fat globules result, which are united by a mesh of fine protoplasm, or they are sur- rounded in the form of a moon by a narrow border of protoplasm, which crowded to one side rests like a cap on the fat globules. Such moon and cap formations may also result in another way. The leucocytes (mac- rocytes), crowd on to the dead or dying cells, eat their way into the cell bodies and establish in the more and more distending cell actual Fig. 14. * * *n * *• * >'; •** * V . C.' lacunae, in which the de- vouring leucocytes lie. The remains of the pro- toplasm and of the cell and nuclear membranes float in the shape of caps and moons in the milk protoplasm until the swelling or further breaking down converts them into spheres or globules. At the same time of course the mac- rocytes may them- selves degenerate in the cell, and no longer pre- sent a recognizable nu- cleus. In such cases its respective lacuna con- tains homogeneous, sharply circumscribed proteid globules. The author considers these epithelial cells which have been destroyed by macro- phages, as identical with the albuminophores of Bab and Schulz which they described as large lymphocytes, (15 to 20/u.), containing fat and 1 to 4 or more proteid globules. Besides these regularly formed constituents of the milk, its sediment contains flaky constituents, small irregular shaped coag- ula, which readily tinge with basic anilin dyes, or with nuclear staining substances. Frequently they are without any structure. At times they appear in individual milkings, almost completely dominating the microscopical field. They are the early stages of the corpora amylacea, soon to be described, which appear either * '«• Budding globules, free nuclei, Nissen's globules, that is cell fragments, in the sediment of cow's milk. 1 X 1000. 30 Microscopy of Milk in General. round, oval, bean-shaped, or nodular, ranging from very small (1 to 2 /A), to an enormous size (5 to 200 ^ according to Zimmer- mann). These bodies show no concentric formation, or radial stripes. They usually appear during abnormal activity of the gland, and are found in colostrum, in stasis of the milk, in mastitis, in the inactive glands of older animals, etc. Their varied thickness makes active turning of the micrometer screw necessary. These corpora amylacea (according to Siegert, corp. flava in contra-distinction from corp. versicolorata, are the same as amylacea) were seen by Herz, Ottolenghi, Iwanoff, and later described by Martin, Lenfers, Winkler and Zimmermann. Wederhake Epithelia in different stages of destruction by macrocytes, that is so-called albuminophores. 1 X 1000. confirmed their occurrence in the colostrum of women, and compared them with the corp. amylacea of the prostate gland. A section offers the best opportunity for the microscopical study of the nature of these bodies. In preparations of acute mastitis, their development is especially clear. Around small flakes of proteids, possibly precipitated nuclear or cell fragments, layer after layer will be formed until a concrement results, which may even fill the entire alveolus. Lime and salts of magnesium are later absorbed by this basic structure of concentric layers, and fine Milk Concrements. 31 radiated stripes appear upon its surface in consequence (Fig. 13, Fig. 16 and Table L). While the alveolar epithelium succumbs to the pressure of the growing concrement, and may be absorbed for some time, the concrement resists the influences of the organs, and finally is surrounded by connective tissue. Zimmermann states that the bodies may be either in the alveolus or on or under the epithelial layer, and even free in the con- nective tissue. These observations have been confirmed by the author. They stain with methylene blue, iodine green, and gentian violet, similar to other amyloid substances, but do not give the starch reaction with iodine solution and sul- Fig. 16. phuric acid (Zimmer- mann, and author's ob- servations). Wederhake and Winkler claim to have obtained a bluish violet coloration with iodine. The corpora amy- lacea of the mammary glands resist few acids (sulphuric acid, hydro- chloric acid). Otto- lenghi and Zimmermann obtained a solution with pure sulphuric acid. They are therefore pure concrements of se- cretion which form un- der peculiar conditions. Their quality varies, depending on the char- acter of the precipita- tions, which combine to lOrm them. Lime concrement in the milk sediment of a cow. 1 X 1000. What remarkable significance may be attached to such conditions may be indicated by the views of Herz, who considers them as the initial formation of casein, and those by Winkler, who believes that they change into fat or that they are degenerated epithelium. Leucocytes also crowd upon these bodies, and attempt to dis- solve them just as osteoclasts attack bones. Under their influence, combined with that of the body juices, a destruction, solution and absorption of the concrements may take place, or on the other hand new layers of thickened secretion may form around the old debris, and a new concrement develops. This describes, with the exception of the fat globules, the cell elements which may be demonstrated under the microscope, as far as they originate from the udder of the cow. The fat globules will be discussed under the heading of milk fat. CHAPTEE IV. COMPOSITION OF MILK AND ITS BIOLOGICAL, CHEMICAL AND PHYSICAL CHARACTERISTICS. There is very little known with, absolute certainty relative to the development of the individual constituents of milk. The theories in this regard are almost entirely hypothetical. It is certain that milk constitutes the specific product of cell activity of the glandular parenchyma, and does not represent a simple trans- udation of the constituents of blood, with a mixture of broken down products of cells (nuclear masses of leucocytes and epithelia, and fatty detritus), nor the fatty breaking down of the epithelium (Reinhardt, Virchow, Skanzoni, Koelliker), nor partial epithelial degeneration of the parts lying adjacent to the lurnen (Heiden- hain), nor transformation of leucocytes and lymph cells (Rauber). None of these is the basic pheonomenon in the formation of milk, but it is due instead to the assimilating activity of the cells, which send their secretion into the lumen of the cell tube (Ottolenghi). A breaking down of cells of course occurs to a greater or lesser extent, in accordance with their increased activity, and therefore the milk contains cells and cell fragments in varied quantities, without this throwing off of cells or breaking down of cells having anything to do directly with the secretion proper. The throwing off of useless material, and its natural replacement by functionat- ing elements are only signs that the organ desires to maintain itself in a condition capable of continued secretion. Our attention has previously been directed principally to the functions and activities of the milk gland from a physiological point of view; the morphological condition of the udder and some con- stituents of the secretion have also been noted. In this chapter the chemical qualities of the milk will be considered, as far as this is necessary for the most ordinary conception of these properties. The quality of the milk — in the broadest sense — adjusts itself to the requirements of the young. The milk gland offers it nutri- tive and protective material in a form which most favorably meets the requirements of the off-spring. In order to give only a few examples attention should be directed to the estab- lished facts, which show that there exist absolute relations between the time required 32 Composition of Milk. 33 for the doubling of the weight of the young and the percentage of proteids in the milk; between the proportion of certain salts and the ash constituent, and the rapid growth of the young ; between the growth of the brain and the supply of proteids and lecithin. Milk consists of dissolved constituents, and this solution con- tains substances in suspension ; in the entire mixture there are also undissolved substances in emulsion. The dissolved and suspended substances are designated as milk plasma, which after coagulation separates in milk serum and coagulum. The fat is present in an emulsion; there are in addi- tion to this several salts, coagulums, cells, etc., undissolved or in a precipitated condition. In coagulation the casein which at first is in suspension, thickens, and carries down the undissolved sub- stances, separating more or less from the milk serum in which the soluble salts, milk sugar, certain proteids, ferments, coloring mat- ter, etc., remain. The principal constituents of the milk, which constitute as well the principal properties of the glandular secretions, are the parts which have received the most thorough study. The proteids. Casein, milk albumen, and milk globulin (traces of lactomucins, and possibly traces of other proteid substances, which remain after acid precipitation and boiling, being known collectively as lactoproteins) are the protein constituents of milk. The fat; the milk sugar. The milk further contains lecithin, sarcin, kreatinin, nuclein, urea and sulphocyanic acid. Nothing is known at the present time of some of these constit- uents, whether they occur originally in the milk, or whether they are only split products, which result during the final production of the various principal constituents, or through bacterial action in the milk ; of such substances may be mentioned peptone, ammonia, leucin, etc. Of non-nitrogenous substances milk also contains citric acid, cholesterin and under certain conditions free lactic acid, alcohol and acetic acid. Gases which occur free in milk are oxygen, nitrogen, and oc- casionally carbonic acid ; the salts are combinations of the bases of sodium, potassium, magnesium, calcium, and iron, with hydro- chloric acid, sulphuric acid, phosphoric acid, carbonic acid, and citric acid. Principal Constituents. Casein is a proteid especially characteristic of milk, occur- ring almost exclusively in the milk gland secretion of mammalia, in quantities of from 2 to 4 per cent. (It is supposed to occur also in the secretion of the sebaceous glands of mammalia and in the coccygeal gland of birds.) The origin of casein is unknown. It was formerly supposed that it originated from an enzymic change of serum albumen produced by the action of enzyme-like bodies upon the albumen. However, since it has been found that the assertion of Kemmerich, relative to the increase of the casein at the expense of the lactalbumen, after the di- 3 34 Biological, Chemical and Physical Characteristics of Milk. gestion of milk at blood temperature for several hours was incorrect (Schmidt and Tier- felder, likewise that casein is not produced by mixing blood serum and macerated milk gland structure, or milk gland juice and ovalbumin, and especially since it is known that casein represents a nuclear albumin containing phosphorus, the enzymic origin of the casein in the above sense is denied. For a time Basch 's hypothesis relative to the origin of the casein was accepted, namely that the nucleic acid which is set free in the alveoli by the activity of the gland, combines with the transuded blood serum, forming the nucleo-albumin, the "casein." Investigation of the experiments of Basch by Odenius, Mendel, Levene and Lobisch proved however that Basch 's hypothesis cannot stand. At the present time it must be admitted that the cells of the milk gland break up the proteids into more simple bodies, and then build up the casein from these products. The casein is distinguished from other proteids containing phosphorus, as for in- stance from the nucleo-proteids, by the absence of the xanthin group, the pyrimid.ins, and the pentose group. The consistence of casein from various species of animals varies chemically to a considerable extent. By special reactions with casein anti-serum (precipitation, complement fixation), the caseins from different species of animals may be differentiated one from the other. In the splitting up of casein into its various con- stituents, quantitative differences in these split products are found which indicate the differences in the individual caseins. The cow casein contains according to C. H. S. P. N. 0. Tangl 52.99 6.81 0.832 0.877 15.65 23.141% Ellenberger .... 53.07 7.13 0.76 0.80 15.64 22.60 % Burow 52.825 7.095 0.725 0.808 15.64 22.906f0 Hammarsten ... 52.96 7.05 ^ 0.758 0.847 15.65 It is insoluble in water and in alcohol, but with bases forms solutions, the so-called casemates. Alkali-caseinates form opales- cent solutions, while solutions from casemates of earthy alkalies represent cloudy, milky fluids. Casein is slightly acid, the solution of which with the bases is accompanied by the formation of salt-like compounds. The characteristics of casein are of especial interest, as they give to the milk its well known properties of rennet-coagulation, and easy acid coagulation, etc. Casein is present in the milk as caseinate of lime, in suspended condition as dicalcium-caseinate, which gives an acid reaction to phenolphthalein, and a neutral reaction to litmus. Acid abstracts calcium from the caseinate, the casein being precipitated (that is casein from the milk of cows and other rum- inants) as coarse, flaky material, while the casein from the milk of solipeds and women is precipitated as a fine, flaky substance. This difference in its properties is traceable to the physical condition which is mani- fested by the casein molecule of the various kinds of milk (Fuld and Wohlgemut) ; but it 7iiay also be the result of a variation in the quantity of salt and proteid present in the milk. In the presence of di- and tri-phosphates the casein dissolves by combining with a part of the bases, so that the neutral and alka- line phosphates change into monophosphates (Hammarsten, Arthus). Casein. 35 Casein is also soluble in other salts, but not, or only to a very slight extent in NaCl, Na2 S04, NaN03, KC1 and others. In the presence of an excess of acid the casein which is first precipitated is again dissolved into a syrup-like mass, but may be again recovered as casein after neutralization. Neutral calcium casein suspensions do not coagulate in boiling, but they form a pellicle on the surface. (The nature of this manifestation is not entirely clear, but depends probably on the drying and transforma- tion of the casein into a more solid form.) Casein is precipitated even in the presence of relatively small quantities of acid while boiling under this condition changes it slowly into a body not susceptible to the action of rennet. In over- heating and likewise in boiling and over-heating with small excesses of alkali, casein is split up through hydrolysis. Even under the action of water, casein is split up into a pro- teid body which is coagulated by heat, passes through a filter and is probably identical with whey casein. The latter substance is formed after the precipitation of the cheesy substance, through the action of rennet, and is a mixture of reduction products of the casein originating through the action of the rennet (Raudnitz). One characteristic property of casein is its precipitation by rennet in the presence of earthy alkali salts. The precipitation of casein has no connection with the action of the rennet as such. This may occur even without having precipitation as a result. If for instance a casein solution is mixed with active rennet, and another solution mixed with inactive boiled rennet, then in the mixture con- taining active rennet, para-casein is formed without any action being noticeable. Only after the addition of soluble calcium salts will precipitation of the para-casein calcium result in the glass which contains the active rennet, but not in the glass containing rennet which has been inactivated by heating. In the change of casein by the rennet ferment, there results in addition to the substance designated as para-casein, another proteid body free of phosphorus, with the properties of albumose, the whey -proteid (Hammarsten). The change of the casein to para-casein, and whey proteid may be a splitting up of the casein, or it may depend on a change in the grouping of the molecules, or it may correspond to a change in its physical condition. The action of rennet in the curdling of milk is practically the same as in casein solutions ; however it is influenced by the other (dissolved) substances, by the other proteids and salts, and pos- sibly also by the physical condition of the fatty emulsion. Curdling with calf rennet develops in accordance with definite laws. In milk that has been brought to low temperatures (refrigera- tor) the action of the rennet may be established by subsequent heat- ing; the precipitation, however, will not take place until the mix- ture is heated to 37 deg. C. (Morgenroth.) 36 Biological, Chemical and Physical Characteristics of Milk. Coagulation may not always appear if the milk is immediately heated to 37 deg., which would indicate that some of the rennet is destroyed at 37 deg. If the same milk is utilized under the same experimental con- ditions, it can be seen that the amount of rennet necessary for the coagulation of the milk is nearly proportionately opposite to the length of time necessary for the coagulation to be completed ; this fact is expressed by Storch and Segelke as follows : "The product from the quantity of ferment and time of coagulation is constant." Each kind of rennet has a certain strength which of course is changeable, and rela- tive for each sample of milk. In strong dilutions of the rennet the action does not corre- spond with the time rule, the time of coagulation becoming continually longer unto infinity; that is, coagulation finally no longer takes place. The action of rennet depends on the most varied factors, which may either hasten or retard its action and influence the precipitation. Acids for instance strengthen the rennet action, likewise earthy alkali salts, while alkalies, albumoses, neutral salts of high- er concentrations, heating of the milk, talcum, caolin, and muci- laginous substances retard the rennet action. Shaking reduces the strength of the rennet if it is in solution. The following data are taken from a work of Smeliansky in order to show the in- fluence of various additions on the rennet coagulation of cow 's milk. It appears that: 1. Heating the milk results in retarding the action. The longer the heating lasts the softer and smaller are the flakes. 2. Addition of water likewise retards the action. 3. Mucilaginous substances retard the rennet action from taking place, and the flakes formed are soft and loose. Barley water especially influences its consistence while corn water principally alters the time of coagulation. If boiled milk is diluted with equal parts of a mucilaginous infusion and water, the mucilaginous portions coagulate more quickly than the watery parts. 4. The addition of soda solution renders the flakes soft, and retards coagulation. Milk containing 0.5% of soda is entirely prevented from coagulating even after standing for 24 hours. Four per cent of table salt renders the flakes softer. Potassium carbonate acts the same as soda while the other salts respond according to their alkalinity. 5. Milk of lime retards the action ; chlorate of lime accelerates it. If boiled milk for instance coagulates after 6% hours, the time required for coagulation after the addi- tion of Ca C12 is only 8 to 15 minutes. It causes the flakes of raw milk to become loose and soft. According to Smeliansky, the reaction indicates the character of the coagulation, and the time required for it. Sugars exert no influence. On the other hand Reichel-Spiro have determined a slight retarding of coagulation in the presence of a high content of cane sugar. Cooking the milk retards the process (lowering the acidity as a result of the loss of CO2 and precipitation of lime salts, Eaud- nitz). In overheated milk no coagulation or only poor coagulation takes^place. The addition of water retards coagulation (Weitzel), likewise physiological salt solution or whey which is free of ren- net (Reichel-Spiro). Hammarsten, Lorcher, Peters, Weitzel, Gerber and Eaudnitz conducted experiments relative to the action of salts on coagulation, the results of which according to Eaudnitz may be interpreted as follows : Rennet. 37 1. The chemical reaction of rennet is hastened by the distri- bution of the rennet and its quantitative relation to the casein, pos- sibly also by elevated temperatures up to an unknown limit. Alka- line earths and acids probably act in a similar manner by activat- ing the rennet. 2. The chemical reaction is retarded : (a) By the destruction of the rennet: temperatures over 41° C., free hydroxylions ; (b) by inactivation of the same: anti-rennet; (c) by changes of tho casein : temperatures over 80 deg. ; formalin. 3. The physical reaction is hastened by higher temperatures, free hydrogenions, and the neutral salts up to a certain concentra- tion, especially the salts of alkaline earths. 4. The physical reaction is retarded by reducing the concen- tration of the mentioned salts below a certain point, especially of the alkaline earths ; therefore heating the milk and the salts which precipitate lime, and calciumions will produce this result. Higher concentrations of neutral salts have the same effect. It may also be possible that some of the alkaline action should be considered here. It is known that by the injection of rennet into an animal an anti-rennet may be produced. The rennet acting as antigen induces in the body of the rabbit the formation of a specifically acting anti-body, which works against the action of the antigen in the re-agent glass, very likely through fixation. Normal serum also contains rennet-inhibiting substances. The action of the rennet may be inhibited or entirely prevented by the addition of horse blood as has been proved by Hammarsten, and later by Roden. The same inhibi- tion is exerted on the action of trypsin and pepsin and is referred to as an anti-ferment action of the blood serum. Blood of cattle added to cow's milk also shows this char- acteristic (Schern). Inhibition action is traced back to the anti-ferment substances of a specific nature contained in the blood, and the presence of an anti-rennet is considered probable. It should however be noted that Eaudnitz and Jakoby prevented inhibition by neutralizing the serum with acid. The strength of the rennet may be tested in various ways. That quantity of milk is measured which is coagulated by one part of rennet in 40 minutes at 35 deg. Market rennet has a strength of 1:10,000 to 1 :100,000 (fluid rennet and solid rennet). Meunier ascertains the quantity of milk which is coagulated by one c. c. of undi- luted gastric juice in ten minutes. Schern employs solutions of rennet (standard rennet prepared according to Morgenroth) of varying density (1:100:200:300, etc). One part of these rennet dilutions is mixed with nine parts of milk, so that milk-rennet dilutions of 1:1000:2000:3000, etc. are obtained. After an action of two hours the samples are placed in the incubator. The dilutions in which coagulation may now be demonstrated give the relative value of the rennet for the respective milk, and if a mixed milk of healthy animals had been used it establishes the " rennet-titer. " It is to be regretted that the standard rennet solutions are not constant, and that they weaken by storing, etc. For this reason it is necessary to establish the rennet-titer before each test on the milk of healthy animals, or on casein solutions. In addition to the rennet of calves, extracts and ferments from other organs of these animals act on milk in a similar manner, such as extracts of spleen, kidney, liver, lung, thymus, intestine, ovaries, testicles and muscles. Rennet from the stomach of a calf is known as chymosin ; rennet from the stomach of a hog, and from the gastric juice of man as parachymosin (Bang). Rennet enzymes may also be demonstrated in the bodies of other animals, fish, birds and snails. gg Biological, Chemical and Physical Characteristics of Milk. Enzymes with the action of rennet have been found in various plants and parts of plants, such as the artichoke, branches of fig trees, candytuft (Iberis pinnata), yellow mustard (Isatis tinctoria), etc., also in bacteria (proteolytic) and in yeast. The individual kinds of rennets vary considerably in their sensitiveness to various influences. Whereas the rennet of calves is very susceptible to heat, and exerts its action readily in alkaline solutions, the parachymosin is less influenced by the harmful action of heat, but is greatly affected in its action by the presence of alkalies. The rennet enzymes obtained from plants act in an optimal way at high tem- peratures (sykochymas at 65-70 deg. C. for raw, at 85 deg. C. for sterilized milk). Aside from casein, milk contains proteids which are coagulable by heat. (1) Lactalbumin which is related to the serum albumin but is not identical with it (it has a slight optical polarization: — 36.4 to —38 against— 60.1 to —62.6, Sebelien). (2) Lacto-globulin may be precipitated with the aid of mag- nesium sulphate. It is contained in milk in quantities of about 0 . 1 per cent, of the total proteids. The lacto-albumin is obtained from the residual solution after saturation with magnesium sulphate and acidifying it, or by almost complete saturation with ammonium sul- phate. 3. Lacto-mucin has been also demonstrated in milk by Storch, Siegfeld, Voltz and Eosengren, whereas other proteid substances such as albumose, peptone, albuminose, lacto-protein, gelatin, galactozymase and opalisin, are considered more recently as products of the preparation of other proteid bodies, at least so far as their appearance in ripe milk is concerned. The proteids which remain in the fluid after precipitation with acid and boiling are collected under the term "lacto-protein." The milk fat consists of a mixture of triglycerides, choles- terin, lecithin, and a coloring substance, and distinguishes itself considerably from the fat of the body and from the nutritive fat by its chemical and physical characteristics. Although the milk fats manifest considerable dependence upon the nutritive fat, as will be seen from the later chapters, nevertheless a transition of the nutritive fat into milk fat cannot be asserted. The same state- ment would also apply to the transition of body fat, although in this instance a closer relationship between the substances must be admitted. It is possible that transitory relations exist, by means of which split up body fat may be converted in the milk gland into milk fat, and thus the nutritive fat takes part indirectly in the formation of milk fat after first having been deposited as body fat. It should be considered however, that the specific activity of the cell builds up the fat from the constituents at hand, and utilizes whatever material is placed at its disposition, such as nutritive fat, when such is present, or body fat in emergencies. The product will approach in its properties the material which has been utilized, but will always remain peculiar to the species of animal producing it. A formation of fat from proteid is possible, as may be seen when cows are fed with substances free of fat, and after the body fat deposits have been used up. It is probable that the carbohy- drates of the food here take part in the formation of fat. Fat Content. 39 The fat which is contained in milk in the form of very fine globules, causes in part the white color of the milk through the reflection of light. The size of the fat globules varies in the milk of the same cow and depends upon the individual, length of the period of lactation, the race, feeding, and upon whether the first, middle or the last part of the milking is examined. According to Woll, D'Hunt, Schellenberger and Gutzeit the diameter varies between 0.8 and 22 n with an average of 2.2:2.5:2.9:3.6 /*. Variations in the percentage of fat are caused by change of food, etc. These changes also have an influence on the size of the fat globules, and according to Woll the fat globules become larger with dry feeding, a statement which could not however be confirmed by Schellenberger and Pankowsky. According to the investigations of these authors the feeding of green forage, especially clover, produces large-sized fat globules. The length of the period of lactation should be considered since the variations of size at the beginning of lactation are more con- siderable than in ripe milk, in which the milk globules appear more uniform and mostly of medium size. In colostrum they vary from the sizes of dust to 20/j. and over. Donne and Schulz found that colostrum contains large, broad oil drops in addition to the small and minute fat globules, which show a less uniform appearance and contour, when compared with the usually spherical fat globules of ripe milk. In interrupted milking the size of the milk globules bears a certain relation to the fat content. With the increased quantity of fat which obtain in the milk toward the end of a single milking, the size of the fat globules also become larger (Schellenberger, Woll). With the extension of the lactation period the size of the fat globules decreases, but their number increases. According to Gutzeit and Schellenberger the following values were obtained in milk from different breeds: Si?e in 1/1000 mm. No. per cc. in millions. Gutzeit : Schellenberger : Voigtlander 2.73 1944 to 4476.9 Jersey 3.5 2.95 2064.1 to 4643.3 East Friesian 2.30 2521.0 to 5911.0 Angus 2.95 2.20 2886.0 to 6200.0 Simmenthal 2.56 2995.0 to 5210.3 Dessau 3070.0 to 6308.6 Swiss 2.33 4008.0 to 5326.7 Shorthorn 2.76 Montavoner 2.62 Holstein 2.58 Breitenburger 2.46 According to Grimmer the number of milk globules fluctuated in 21 tests on three herds of blackish-brown lowland cattle in Pomerania, from 1,330,000 to 3,073,000 per cubic millimeter, having an average diameter of 2.6-3.7/u. The milk globules retain their form through their surface tension and are not surrounded by special capsules which could be considered as membranes, as has been thought by former authors. 40 Biological, Chemical and Physical Characteristics of Milk. Although the milk globules cannot be entirely freed from proteids by washing (covering the milk with water and allowing the separation of fat), the demonstration of the remains of proteids cannot be considered as proof of an actual "haptogen mem- brane" which must be broken down during the butter-making process, in order to make possible the flowing together of the milk fat, but it does constitute a proof that rem- nants of proteids, even after the most careful washing of the cream, remain around the fat globules. At least it has never been possible to demonstrate membranes of the fat globules, neither in boiled milk, in which during continuous heating larger fat clumps develop, nor in fat extractions (Soxhlet, Quincke, Morres). Milk sugar is also a specific substance of milk. It is formed in the gland and is found only in its secretion. If sucking is in- terrupted, it may be present in the urine, from which it immedi- ately disappears upon amputation of the lactating gland, or it may not appear at all when the gland is amputated before the appear- ance of lactation (Sinet, Magnus-Levy, Zuntz). After the com- plete removal of the gland in goats and cows, however, a temporary hyperglycosemia and glycosuria appear. If parts of the gland remain, lactosuria results. After the injection of glucose, lactose appears in the urine (Porcher), likewise after the ingestion of large quantities of dex- trose. Since the blood in the mammary vein before parturition and during lactation contains considerably less glucose than the blood of the jugular vein (Kaufman and Lagne), it may be accepted that glucose has been utilized in the gland, and further that glucose is the material from the constituents of which the lac- tose is formed in the gland. Of the various salts milk contains compounds of potassium, calcium, magnesium, iron, traces of manganese, aluminum, phos- phoric acid, hydrochloric acid, carbonic acid, sulphuric acid, citric acid, fluorine and iodine. Carbonic acid, oxygen and nitrogen have been demonstrated as gases in the milk. Besides these substances, lecithin, cholesterine and coloring matter are present in the milk, besides ferments and substances which are collected as residual substances ; these have been pre- viously mentioned. Baudnitz and Grimmer have recently published compiled arti- cles relative to the individual constituents and chemical properties of milk which contain the collected material of many experimental results, and at the same time show how much is still unsettled in regard to the composition of milk and the characteristics of the substances which it contains. Certain physical characteristics of milk correspond to its chemical condition. These adjust themselves according to the pro- portion of the various constituents, and to the conditions attend- ing the mixing of the different component parts. The appearance of the milk is influenced by the suspended casein and the proportion of fat. Skimmed milk, which is almost free from fat constitutes a non-transparent, somewhat bluish fluid, as compared with the whitish yellow color of whole milk. The ad- Specific Gravity of Milk. dition of alkalies to milk free of fat renders it transparent. Ham- marsten furnished the proof that a calcium caseinate solution which corresponds to the composition of milk is almost as non-trans- parent as milk. The milk becomes less transparent the smaller the fat globules are. This is most strikingly apparent when the fat globules are broken up to dust-sized bodies (for instance through homogenization). The appearance of fresh milk is also influenced by the coloring matter present in the milk plasma and in the fat. It is known that the skimmed milk of certain cows varies considera- bly in color ; at times it is bluish white, sometimes more yellowish green, again transparent, other times of a non-transparent whitish color, and also the fat has a more yellow color during the pasturing of the animals than at the time of stable feeding. The non-transparency as mentioned above is no proof of the presence of fat in the milk; therefore all methods which are destined to establish the quantity of fat or addition of water by the establishment of the whiteness, are of no use, as for instance, Heeren's pioscope, Feser's lactoscope, etc. If milk is allowed to stand for a time, cream forms on the surface ; the fat globules rise and collect usually as a distinct layer of cream above the milk. The rapidity of the separation depends on the temperature, the size of the fat globules, and the density of the milk plasma. The quantity of the cream is not in parallel rela- tion to the quantity of fat ; it depends on the size of the fat globules. The separation of cream may be hastened and increased by centrifugalization. During separation while allowing to stand, about 85% of the fat rises to the surface, while by a perfectly operating centrifuge the separation of cream may be accomplished up to 0. 01% of its fat. The specific gravity of the milk depends on the solid sub- stances, the relation of the mixture and the condition of the sus- pended, dissolved, and emulsified constituents of the solid sub- stances. Corresponding to the variable composition of cow's milk it is natural that the specific gravity of the milk should vary. It fluctuates considerably, varying from 1.027 to 1.034 at a temperature of 15 deg. Similar to the impossibility of speaking of milk of normal composition, one cannot speak of milk of normal specific gravity, and even to give average figures would be of very problematical value; but to take such average figures or even smallest values as a basis for the calculation of falsification would be a gross error. Milk from many cows would under ordinary con- ditions have a specific gravity of 1,029 to 1,033. The specific gravity is measured, or is calculated from the values of fat contents and solids, according to formulas, which, depending on the milk from certain breeds, or certain localities, show slight variations. This formula made on the basis of the value of the specific gravity of the milk fat (about 0.93), and the solids or dry substances (1.6001), which is quite constant, is according to Fleischmann : 42 Biological, Chemical and Physical Characteristics of Milk. 1000 B== • 1000—3.75 (d— 1.2 f) In these equations s stands for specific gravity, d for dry substances or solids, and f for fat. The following values may also be calculated from the fat con- tents of the milk and its specific gravity. 1. Total solids: d=1.2f+2.665X100s-10° s 2. The fat-free solids are shown by deducing the percentage of fat from the percentage of the total solids. 3. The specific gravity of the solids sXd _ " sXd— (100 s— 100) 4. Finally the fat contents when the solids and specific grav- ity are known: The values obtained from formulas are of course not abso- lutely correct, but represent the results only approximately with the analytical methods of weights, the fat-free dry substance of the milk is not of absolute constant composition, but varies, so that its specific gravity which is based upon the sugars, proteids and salts, varies more or less from the number which has been accepted by Fleischmann as the average value (1.60). The equations hold only for cow's milk. If milk is freshly drawn, and immediately tested it shows a considerably lower specific gravity (0.0008-0.0015), than after cooling. The milk "contracts" and becomes constant in its specific gravity only after standing for several hours. The cause of this manifestation is not yet entirely clear. Toyonaga aims to explain it by the fixing of previously uncooled and fluid fat globules, which is the most plausible explanation ; other authors believe that the contraction is the result of a cessation of the expansion of the casein. The density of the milk varies in accordance with the tempera- ture. The maximum (for water at 4 deg.) lies almost near its freezing point, namely at 0 . 3 deg. C. The freezing point of milk is somewhat lower, namely — 0 . 54 to — 0 . 57 deg. This is especially influenced by the presence of salt, less by the sugar contents of the individual samples of milk, and it is induced by the relatively constant amount of soluble salts in the salt contents, which is subject to only slight fluctuations in the milk of healthy animals. For the sake of completeness the electrical conductibiiity of the milk should also be mentioned. This varies according to the re- sistance which is offered by the fluid to the current. It fluctuates Polarization of Milk. 43 within wider borders than the freezing point, and is influenced ac- cording to Zanger by general diseases, through local affections of the udder, by estrum, pregnancy, feeding, etc. The conductivity is diminished by the fat globules ; therefore skim milk conducts better than whole milk or cream. The conductivity of the different quar- ters is inversely proportional to the quantity of milk, in milk from different quarters of one cow (Schnorf). The viscosity of milk is a factor which principally depends on the condition and on the quantity of the casein and the fat. Higher temperatures reduce the viscosity, likewise shaking; quiet standing increases it. The surface tension of milk is lower than that of water (0.053 against 0.075). Of the physical properties the specific gravity of milk and its serum, and the polarization of milk serum, are of special impor- tance for the practical testing of milk (see technique). For practi- cal results, however, the determination of the fat contents is also necessary. As it has been shown the total solids may be determined by the aid of the fat contents and the specific gravity and the fat-free solids may be established by deducting the percentage of fat, these four factors are generally sufficient for the preliminary tests. For more accurate study these preliminary tests are com- pleted by the establishment of the specific gravity of the milk serum, or still better by the ref ractoscope to determine the chlorids of calcium serum, which renders more rapid work possible. This is a method whose satisfactory use in practice has been proven by the numerous works of Mai and Eothenfusser. Publications relative to the polarization of milk were issued by Valentin in 1879, and later continued by Villiers and Bertault, Braun, Utz, Lam, Radulesku, Eipper, Schnorf and others, on ren- net serum, acetic acid serum and milk serum, which had been pre- pared by voluntary coagulation. The given values of the authors varied in accordance with the method of preparation of the serum ; nevertheless it could be estab- lished that comparatively uniform figures were obtained whenever the work was carried out under similar experimental conditions. In 1908 Cornalba showed that contrary to the variance in the amount of colloidal substances dissolved or suspended in milk, the sum of the dissolved constituents of milk is very constant. Whereas in samples of mixed milk the sum of the first substances varied between 5 and 8.585 per cent, the differences for the total dissolved substances were only 6.05 to 6.25 per cent. Milk serum which contains the dissolved substances, offers therefore constant results in the examinations, the same as the examinations which lead to the establishment of the fat-free solids, which still include the casein. Examinations of serum are therefore of the highest practical value for the demonstration 44 Biological, Chemical and Physical Characteristics of Milk. or establishment of the addition of water, provided that the serum is always prepared in the same way. Ackermann, Mai and Roth- enfusser have in their fundamental works, determined the practi- cal importance of the polarization of the proteid-free serum, and have proved that with the polarization of the chloride of calcium serum we possess means which are better adapted than any other to the detection of the adulteration of milk by water. Refrac- tion is the most valuable accessory to the various methods of tests of recent times. Ackermann found in 2,800 samples of normal milk, variations in the scale division of Zeiss 's immersion ref ractometer, from 38 . 5 to 40.5. Even slight additions of water reduce the refraction con- siderably; the addition of 5% of water results in a 1.3 lowering of the scale division, while 10% lowers it 2.3. According to Mai and Rothenfusser the original refraction of 39 scale divisions is lowered to a refraction of : 37.9 with about 4% addition of water 37.7 " " 5% " < 37.5 37.3 37.1 36.9 36.7 36.5 36.3 36.1 35.9 35.7 35.5 35.3 35.1 35.0 34.8 34.0 33.3 32.6 32. 30.9 070 7% 8% 10% 11% 12% 13% 14% 16% 17% 18% 19% 20% 25% 30% 35% 40% 50% It (t 1 1 14 1 1 (t 1 1 1 1 1 1 « i I 1 1 t < (4 it it it 1 1 < t 1 1 ( l In the establishment of the refraction index of the chloride of calcium serum it was also discovered that it is impossible to estab- lish normal values for the chloride of calcium serum, as well as for other constituents of milk. Mai and Rothenfusser also estab- lished the general rule for milk, that only in the presence of rigor- ous controls of the same origin can the addition of water be satisfactorily determined, and the extent of the adulteration established. Ferments in Milk. 45 The experiments of Weigner and Yakuwa are of interest since they demonstrate that the refraction and specific gravity of the chloride of calcium serum are theoretically of equal value. Mai and Rothenfusser, on the other hand, emphasize the fact that of two theoretical methods of equal value the man in practice has to prefer the method which offers, with the same certainty of the results, greater advantages in regard to rapidity, convenience, and saving of material, advantages which the method of refrac- tion possesses. The investigations of Mai and Rothenfusser prove that the variations in the results of continued tests, from day to day may reach in mixed milk of one stable 0.1 to 0.55, and in longer periods ( 22 days ) , up to 1 . 0. Changes of feeding have no marked influence. The milk of individual cows failed to show any important fluctua- tion during the time in which the tests of the entire stable were made (0.2 to 0.6). More considerable may be the fluctuation between the find- ings of normal milk and the secretion from cows with an affected udder, and the variation between the findings of milk from the same animal while healthy, and within 24 hours after the udder becomes diseased. The milk of individual animals with affected udders shows, not infrequently, values which are considerably below the values of normal milk. This has been proved by the work of Metzger, Fuchs, Jesser and Henkel, and from the experience of the official milk control station. These abnormal values, however, do not affect the worth of this method, if the results are compared through the use of satis- factory control tests, and confirmed by other methods. Ferments in Milk. Immune Bodies. Milk as Antigen. For the testing of milk special characteristics which it pos- sesses, which may be collected under the name of reaction manifes- tation of ferment action, and for which at present there is still no satisfactory explanation, are of importance. Under the term ferments (enzymes) those substances are included which hasten chemical changes with an explosion-like rapidity (Uexkuell), and without using themselves up they act in relatively minimal quantities. Their activity is inhibited by the products of the reaction. Higher degress of heat and certain toxins (ferment toxins, as for instance hydrocyanic acid) inhibit their activity, the ferments being thermolabile. The author desig- nates as ferments all of those bodies with ferment-like action, with- out consideration as to whether the nature of the ferment is known or not. A careful distinction must be made between original fermen- 46 Biological, Chemical and Physical Characteristics of Milk. tative action and ferment-like bacterial activity sometimes taking place in milk. The original "ferments," the nature of which is disputed, originate from the blood, or are formed from the cells of the blood and the parenchyma of the udder. They are either eliminated the same as products of metabolism into the surrounding parts, or they are anchored to the cell and are only set free in the breaking up of the cell (ecto- and endo-ferments). The ferments in their action are destined to certain substances to which they fit, "as the key fits the lock" (Fischer). They act either through hydrolytic splitting, through oxidation, or through reduction. Those ferments are of importance to the milk inspector, where diminished or increased presence or complete absence offers cer- tain conclusions as to various conditions in the udder or in the milk. These are the amylase (diastase), the indirect oxydase (per- oxydase), the superoxydase (catalase) and the indirect reductase ( aldehydreductase, ' l aldehydcatalase " ) . Besides those mentioned, milk also contains other bodies which are included among the ferments; for practical milk examinations, however, they have little or no bearing. Mention need only be made here of the proteolytic ferment, ' ' Galactase, ' ' found by Babcock and Eussell and bodies acting like pepsin or trypsin (Jensen, Freudenreich, Spolverini and others). These are only present in very small amounts. Kinase and fibrin ferment have also been demonstrated in milk. The proteolysis could be explained through the presence of leucocytes in the milk. Similar to the proteolytie ferments which cannot be utilized for diagnostic purposes, the lipase and the salol-pplitting salolase (the existence of which as a ferment is dis- puted by Desmouliere, Miele and Willen; the alkaline reaction of various kinds of milk is sufficient to split up the salol) can not be likewise utilized for the purpose of di- agnosis. Rullmann in 1910 proved by the examination of aseptically drawn milk, that salolase is not an original ferment; the author considers the splitting of the salol to be the result of bacterial action. Of the ferments in milk which split up the carbo-hydrates, the amylase (diastase, galactoenzyme), whose action is similar to the ptyalin of saliva splitting up the polysaccharid starch into dex- trose and maltose, is of the greatest interest (Moro). This fer- ment was first found by Bechamp in the milk of women, later by Zaitscheck, Koning, Seligmann and others in cow's milk. One hundred c. c. of mixed milk can be split up by 0.015 to 0.020 gm. of amylase. Amylase is destroyed by heating for 30 minutes at 68 deg. C. (Koning) ; the optimum of its activity lies at 45 deg. C. The substances designated as oxydase and peroxydase exert a special action. They transmit the oxidation either by "activation of the oxygen of the air," (direct oxida- tion) or by abstracting the active oxygen, for instance from peroxide of hydrogen (per- oxydase). Substances acting as reagents indicate their oxidation by the formation of coloring matter. The occurrence of direct oxydase in milk, the action of which appears even without peroxide of hydrogen, is uncertain. Bull- man has found traces of direct oxidation in milk drawn under sterile conditions ; the quantity however is almost nil for practical purposes. The indirect oxydase acts only after the addition of hydro- gen peroxide or other oxygen carriers (for instance super-borates), Catalase. 47 by abstracting active oxygen after the formula H202=H20+0 (Jensen). The active oxygen oxidizes the added "chromogenic" substances, as guaiacol, ursol, paraphenylendiamin, etc., to coloring matter. The peroxydase is injured by long heating, even at the relatively lower temperature (50-60-70 deg. C.), and is destroyed at about 75 deg., so that boiled or pasteurized milk may be dis- tinguished from raw milk by the non-appearance of the color reaction. The action of the superoxydase (Raudnitz) or catalase (Loew) develops in a different way. It splits the H202 according to the formula 2 H202=2 H20+20, which join to a molecule of 02. Other authors include the superoxydase with the oxidizing fer- ments, as the freed oxygen is utilized in the body for the oxidation (Seligmann). According to others it is included with reductase, as the action of the ferments on H202 equals a reduction of 2 H20, and molecular oxygen 02 which passes out without being utilized for oxidation, whereas the oxygen freed by peroxydase is imme- diately utilized for further oxidation changes; therefore the per- oxydase is an oxidyzing, while the catalase is a reducing ferment (Grimmer). Original catalase has been demonstrated in the milk of all animals ; it originates in the cells of the milk gland, especially from the leucocytes. It is secreted, but may be set free in the breaking down of cells or may appear bound to the cell. That catalase is derived from the cells (especially leucocytes) is not contradicted by the fact that cream is richer in catalase than skim milk since leucocytes and other cells are also included in the separation of the cream. These conditions were indicated by Friedjung, Hecht and Pallazzi, and later confirmed by Koning. This also explains the reason for the centrifuge foam, rich in leucocytes, giving such a strong reaction. Since the formed elements (cells) are precipitated with the casein, and probably a part of the free ferment is also drawn down with it, milk serum is always poorer in catalase than the original milk. Catalase passes through infusorial earth filters, but consider- able quantities are retained. Light, storage, etc., affect catalase, even if it is relatively resistant. A leucocytic extract, which was kept exposed to the light in the laboratory of the author, showed even after months, an unweakened action to H202, while hydrogen sulphide, hydrocyanic acid, potassium cyanide, mercuric cyanide, barium nitrate, hydrochloric acid, sulphuric acid, acetic acid, oxalic acid, and potassium nitrate affected its action (Faitelowitz). It appears noteworthy that H202 inhibits the ferment in its action. In the presence of excessive amounts of peroxide of hydrogen the ferment splits up less H202 than if the diluted peroxide of hydro- gen is gradually added. Heating to 62-70° C. destroys the original catalase in a short time. The optimum temperature appears to be about 37 deg. C. 48 Biological, Chemical and Physical Characteristics of Milk. An original " ferment," the nature of which is by no means definite, is Schardinger's formalin methylene blue reductase, which according to Trommsdorff, will be designated as Schardinger's ferment (synonyms are indirect reductase, aldehydcatalase, alde- hydreductase). Fresh milk in a mixture of formalin and aqueous methylene blue solution (Schardinger's re-agent), is decolorized inside of a few minutes. Smidt explains the action of Scharding- er's ferment by the fact that the formalin changes into formic acid and thereby reduces the methylene blue. The character of its action however is not yet solved. The Schardinger ferment exerts its best action, at 65 to 70 deg. C., it is destroyed above 70 deg. As has already been indicated by Smidt and confirmed by Tromms- dorff, Schardinger's ferment is very sensitive. It is injured by small excesses of formalin, and by relatively larger quantities it is destroyed. Bonier and Sames established more recently, the interesting fact that boiled milk with 0.3 e. c. of a 1% of ferrosulphate solution also gives the reaction, and this disap- pears again when the mixture is boiled for a half hour. The authors point to the care which must be taken in .'judging the so-called enzyme reaction, since it is possible, with the aid of simple chemical reagents, to produce similar effects to those obtained in the supposed enzymatic reaction. Very little is known relative to the origin of the formalin re- ductase in milk. This ferment is not in every sample of milk, being frequently absent in milk from an animal whose off-spring is still sucking, and in animals which are just fresh in milk (Schern). It is absent when the time of milking is over-extended, and in stasis of the milk (Romer and Sames), and it does not decolorize, or only incompletely so, in the first part of the milking, better in the middle of the milking, and rapidly in the last portion of the milk- ing. This also corresponds to the relative frequency of fat in milk but no one however has been able to establish a complete par- allelism. The authors conclude from this that the same conditions under which the gland excretes especially large amounts of fat, cause the quantity of Schardinger's ferment to be likewise in- creased. Milk as Antigen and Carrier of Anti-Bodies. Since the fundamental experiments of Ehrlich relative to the formation of immune substances in the animal body, we possess an explanation for manifold manifestations between the inter-action of the disease-producing agent and the animal's power of protec- tion, known as Ehrlich 's theory of immunity. The substances which are formed in the body in the com- bat against certain invaders are the anti-bodies ; the harmful sub- stances which are capable of stimulating the body to the formation of anti-bodies are the antigens. Antigens may be substances of the most varied kinds ; animal proteid, animal cells, plant cells, plant proteid, living and dead bac- teria, bacterial substances, toxins, etc. The antigens are distin- Antigen Action. 49 guished by groups, which make possible their combining with cer- tain groups of the cell substances of the body. The "haptophore" groups of antigens under certain conditions fit as a key fits the lock, into the haptophore group of the "receptors," thus making possible the binding of the antigens to the cell. These terms were applied to these bodies by Ehrlich. The simplest way of explaining the mechanism of the antigen action and the anti-body formation is by using toxin as an example. A toxin is an antigen with a haptophore binding group, and a poison-producing group, the toxophores. If the toxin enters the body of an animal it may find groups on the cell to which it fits, the so-called receptors, which bind its haptophore group. If this has been the case the toxophore group exerts its action, the effect of the toxin becomes noticeable and the animal suffers as a result of the toxin. If there are no receptors present for the specific toxin it is impossible for the toxin group to exert its action, and the animal is therefore resistant against this respective toxin. It is possible that as a result of the receptors of the cell com- bining with the toxin, the cell molecule is destroyed. But if the damage is not too serious, the protoplasm is stimulated to produce numerous receptors, — an over-production in fact. As not all of these are necessary for the performance of the cell function, the superfluous ones are rapidly thrown off into the body fluids. If such free receptors combine with the haptophore groups of the toxin, the latter is no longer able to combine with the protoplasm of the cell. These free receptors therefore protect the body against renewed action of the toxin, that is they act as antitoxins, and con- stitute the antitoxic part of the serum. Besides the antitoxins, the action of which lies principally in the neutralization of the binding group of the toxin (anti-bodies of the first order), there are still more complicated receptors, for instance those which possess an active or ferment-producing group ; they are anti-bodies of the second order. Finally there are anti-bodies of the third order, which are unable to act by them- selves, but must utilize a third body in order to exert an action on the antigen. Immune bodies of the third order become complete in their action only through the utilization of the complement. These im- mune bodies of the third order possesss therefore a binding group for anchoring the antigen, and a binding group for the complement. They are amboceptors, in contradistinction to the uniceptors of the first and second order. Some anti-bodies resist heating for a half hour at 56 deg. C. ; they are thermostabile, as for instance the antitoxins, the agglu- tinins, the amboceptors, while others, as for instance the comple- ment, are destroyed at this temperature, as they are thermo-labile. Tf, for instance, hemolytic anti-bodies are produced in a rabbit by treating the rabbit with red-blood corpuscles of another animal, then the hemolytic rabbit serum lo^es its action by heating to 56 de^. C. 50 Biological, Chemical and Physical Characteristics of Milk. The red blood corpuscles however are again dissolved when to the heated, "inacti- vated" rabbit serum, guinea-pig serum containing complement is added. Therefore, whereas neither the amboceptor in itself, nor the complement in itself can dissolve blood corpuscles, the combination of the two is capable of doing it. The action of the anti-body is specific for the substance which induced its formation, on homologous antigen. Diphtheria anti- toxin acts only on the toxins of the diphtheria bacillus, and not on the toxins of the tetanus bacillus. The specificity is a very high one, nevertheless it is only relative; that is, a similar, although somewhat weaker action is exerted on related antigen, as compared with the specific antigen. The chemical structure of the anti-bodies is unknown, and they are generally designated according to the action which the/ exert in the animal body. Anti-toxins neutralize toxins, agglutinins agglutinate (stick together) animal cells and bacteria, and drag them to the bottom, precipitins and coagulins produce precipitation in antigen solu- tions, hemolysins dissolve erythrocytes, bacteriolysins dissolve bacteria, cytolysins dissolve animal cells, etc. If anti-bodies are produced by injecting antigens into an an- imal, then the animal is actively immunized against the antigen. On the other hand, if another animal is injected with the produced anti-bodies it is given a passive immunity. The active immunity lasts for a long time, the passive immunity does not last beyond several weeks. If anti-bodies are present in the blood in certain quantities they are excreted by the milk gland, and may be demonstrated in the milk. Ehrlich succeeded in proving the passing of anti-toxins into the milk of anti-toxic immune mothers, by showing that young mice from non-immune mothers acquired a high degree of resis- tance against the toxin when they were allowed to suck actively- immunized mothers. The passing of anti-bodies into the milk even in the presence of passive immunity was proven by Ehrlich, Schmidt and Pflanz, although the passage was only slight. The action of such milk was 15 to 20 times weaker than that of the blood. Similar to the action of the anti-bodies of the first order are those of the uniceptors of the second order; for instance, bacterial agglutinins and precipitins. The passage of agglutinins from the blood into the milk has been demonstrated by Kraus, in goats which had been immunized against colon-bacilli, typhoid and cholera. The later works of Bensaude, Bertarelli, Bamberg and Briigsch, de Blasi, Courmont, Cade, Figari, Maragliano, Eodella, Staubli and others confirm the findings of Kraus. The agglutination value of milk, as compared with blood, may be lower or identical, or it may even be greater than that of the blood. As it has been found that bacterial agglutinins may pass into the milk, so it also has been proved that agglutinins against animal cells may do likewise. Complement Content of Milk. That under certain conditions amboceptors, as immune bodies of the third order, may pass into the milk, is proved by Bertarelli's experiment on a sheep treated with the red blood corpuscles of a chicken. The specific hemolytic amboceptor which resulted could be demonstrated in the milk. Therefore although amboceptors may pass into the milk and although normal milk contains non- specific hemolytic amboceptors in small amounts, nevertheless the passing of hemolysins into the milk is very uncertain. According to Kraus, Kopf and others hemolysins do not occur in milk ; like- wise bacteriolysins are absent, or their presence is very doubtful, according to the investigations of Bab. Of course one of the hemolytic factors, the amboceptors, might be present in the blood, while the complement under the special conditions present in milk, may be inactive. While Pfaundler and Moro state that hemolytic and bacterici- dal complement may be found in cows' milk, Bauer and Kopf, and Bauer and Sassenhagen, on the other hand showed that in normal, ripe milk complements are not present; that is, even by special examinations only traces could be established. On the contrary in samples of colostral milk, and milk from udders affected with mastitis, both amboceptor and complement may be demonstrated. The complement content of milk drops with the duration of time which has elapsed between parturition and the taking of the sample, until from the sixth to the twenty-seventh day after calving the amount of the complement disappears. This observation may possibly be of great practical value in ascertaining whether or not a cow is fresh in milk. Mastitis milk, which bears a close relation to colostral milk, showed a relative richness in amboceptor and in complement, thereby making it pos- sible to establish the affection of the udder by the demonstration of the complement. Of course it is not certain that the comple- ment occurs early enough to enable this method to be utilized more readily than for instance the Trommsdorff test, the catalase test, or microscopic examination of the centrifuged sediment, and others. Sassenhagen found in one case that the presence of mastitis could be determined by complement-fixation 18 days before the first clinical appearance of the disease, even when the quantity of sediment, after the Trommsdorff reaction was insufficient to afford a basis for a diagnosis of mastitis. Bauer further proved that complement inhibiting substances are present in milk ; Hausmann and Pascucci traced this inhibition of hemolysis to the presence of lecithin or cholesterin in the milk. According to Kopf the complement passes from the colostral milk into the blood of the calf ; it may be demonstrated in the serum of the calf from the third to the fourth day, before which time the blood cells of guinea pigs were not dissolved. 52 Biological, Chemical and Physical Characteristics of Milk. As proved by Kraus bacteriolytic immune bodies also pass into the milk, in artificially immunized animals, and into the body of the suckling consuming the milk, provided the mother possesses active immunity (de Blasi). Of other immune bodies which are present in the blood and have been demonstrated also in the milk of the same animal, should be mentioned the opsonins (Wright), which influence the bacterins in such a way that they may be readily assimilated by the phago- cytes (Turton and Appleton, Eisler and Sohma). Other substances which induce the so-called hypersensitive- ness (anaphylaxis), have also been demonstrated (Otto). At least it has been proven in the study of hypersensitiveness, that the off-spring of hypersensitized guinea pigs possess an increased sensitiveness for homologous antigens, and this may not only be the result of the intra-uterine transmission of the anaphylaxis from the mother to the young, but also of the transmission of the immune bodies, through the milk of the mother. To the subject of immune substances belong possibly the ob- servations made by Tage, Duhat and Dobrowits, during the treat- ment of nursing syphilitic mothers with salvarsan which shows its effect upon the untreated syphilitic children. Syphilitic chil- dren thrive splendidly after the treatment of their mother. It was impossible to demonstrate arsenic in the milk, either in or- ganic or non-organic combination. Ehrlich explains the action by the fact that a rapid breaking down of the syphilitic spiro- chaetes in the mother is produced through the action of the new syphilitic remedy, and thereby an elimination of the endo-toxins is induced. The antitoxins which develop in the mothers pass into the milk, and cause a passive immunization of the child, through the gastro-intestinal tract ; Jesionek, on the other hand claims the passage of the arsenic from the blood of the treated mother to the milk, and explains thereby the remarkable results in untreated children which are nursed by the treated mothers. Very little is known with certainty relative to the quantita- tive relation which exists between the immune bodies appearing in the circluating blood, and those in the milk. The views expressed are too widely divergent. It is known of the anti-toxins in which this relation has been mostly studied, that of 15 to 30 parts of the anti-bodies which are demonstrable in the blood a certain amount appears in the milk. These relations are still somewhat vague, since the passing of the anti-bodies which are bound to the albumins and globulins depends on the quantitative relation of these proteids in the milk, and the experimental results therefore must vary in accordance with the species of animal used, the stage of lactation of the respective individual, diseases of the udder, etc. In infections of the udder, for instance with colon bacilli, anti- Agressins. 53 bodies accumulate in the glands so that as a result the milk serum agglutinates more readily than the blood serum. Not only anti-toxins and other protective immune bodies pass into the milk, but substances also which inhibit the protective power of the body, for instance aggressins, at least so long as the body has not formed anti-aggressins. The aggressins for instance act against the dissolving of bacteria. Schenk demonstrated anti- staphylolysins and anti-vibriolysins in the milk of goats, cows and women. Otherwise the passage of toxic substances of the character of antigen, which are closely allied to proteids, could be just as plausible as the passage of the constituents of the blood which are indispensable in the composition of the milk. The passage of toxins into the milk has not yet been satisfactorily proven for all toxins. A large number of known substances from animal and plant life are known as toxins, that is, bodies which do not act like chemical poisons, but exert their toxic action only after a period of incubation, in which time fixation takes place. These toxins do not affect all animals in a similar degree, but only those which are susceptible. Certain species of animals are not susceptible to certain toxins ; they are immune. This im- munity may also be artifically established in susceptible animals. The toxin is an antigen, and under certain conditions it produces an anti-toxin contrary to the toxins which act purely chemically. Among toxins acting in this manner may be mentioned the products of metabolism of the Bacillus diphtheria, the Bacillus tetani, the bacillus of certain forms of meat poisoning — the Bacillus botulinus and the Bacillus pyocyaneus, the bacillus of blackleg, and the body substances of certain bacteria (endo-toxins). They may be of animal origin : snake toxins, spider toxins, scor- pion toxins, turtle toxins, toxin of the blood of eels, salamander toxins, wasp toxins, or of plant origin, such as the abrin, robin, krotin, ricin, etc. If it is considered that the gastro-intestinal tract of very young individuals is readily penetrable for proteids, although pro- teids of unlike origin pass with greater difficulty than those of like origin, the question as to whether the milk of the mother may con- tain toxins when toxins are circulating in her blood, assumes prac- tical importance. This becomes, however, unimportant when it is considered that even in severely affected individuals only very small quantities of toxins are circulating free in the blood. Should a part of these minute amounts be secreted in the milk, this quan- tity itself is of only little practical importance even when the great susceptibility of the intestines of the suckling is considered. It is true that Miessner succeeded in proving that mice die from tetanus when they are fed with raw milk from a cow affected with tetanus, whereas the feeding of meat has no influence on the 54 Biological, Chemical and Physical Characteristics of Milk. health of the animal; this proves the passage of the tetanus toxin into the milk. Older animals do not become affected even after the adminis- tration per os of large doses of toxins, at least not from diphtheria or tetanus toxins, and the Bacillus botulinus, the toxins of which are absorbed by the stomach but the bacillus does not thrive in the body; therefore the possibility of secreting these toxins through the milk gland is from the first of small importance, especially since in severely affected animals the secretion ceases. From a practical consideration of the question of toxin elimi- nation, the plant toxins come principally into consideration, espe- cially ricin, as food adulterations to a great extent take place with ricinus seed and its flower. An elimination of ricin with the milk, however, has not been observed up to the present time. Ehrlich was unable to observe an elimination of ricin in mice which were under the action of ricin; the offspring of these mice wrere not actively immunized against ricin but acquired only a passive im- munity of short duration. Of more importance however are the bacterial toxins, and products of decomposition acting like toxins, which subsequently develop in the milk after certain fermentation processes. The above-mentioned immune substances are probably of great importance for the nourishment of the young and the sucklings. The passage of genuine proteids in very young individuals with injured mucous membranes, is an established fact, and with the globulins anti-bodies also pass into the blood of the young, while in older individuals the relatively labile anti-bodies are changed or destroyed by the splitting up of the proteids. The absorption of anti-toxins through the intestines of the young has been proved by the classical experiments on sucklings by Ehrlich. Other works by Brieger, Ehrlich, Salge and Eomer prove that certain immune substances of milk of like origin pass through the intestines, while in feeding sera or anti-substances of like origin contained in milk of unlike origin the quantity passed was only very slight. Thus Rb'mer succeeded in demonstrating passive immunity in foals after feeding them with anti-toxin milk of like origin, but was unsuccessful after feeding anti-toxic sera of like origin. ^ In calves of course the results were positive even when the anti-toxin was mixed with the milk as a serum of unlike origin, but the quantity of immune bodies of unlike origin absorbed was smaller than that of like origin. The absorption diminishes with the increase of the age of the animal. Relative to the passage of other immune substances from the milk into the blood of the suckling, the same experiences hold as a rule as in the case of milk containing anti-toxin. Milk is not only a carrier of anti-bodies, and possibly of anti- Arule purulent fibrinous mastitis. Dilation of the blood vessels (c) ; with exudate of numerous cells into the alveoli and excretory duct. Hematoxylin — Sudan III. Concrement formation in milk stasis (a), and in mastitis (b) ; inactive portion of the gland (c). Ernst, Milk Hygiene. Precipitin. 55 gen, but as a proteid-containing material it is an antigen in itself, or rather a collection of antigens, which may again produce anti- bodies in the body of an animal. These anti-bodies against milk not only develop in artificial administration by injections, but also under certain conditions during the natural ingestion of the milk per os as a food. Although usually such anti-bodies against nutri- tive proteids of unlike origin appear only in intensive over-feeding of proteids (Ascoli, Michaelis and Oppenheimer, Uhlenhuth, and others), nevertheless in the presence of an injured intestinal mucous membrane the absorption of proteids of unlike origin may, under natural conditions, take place, and thereby induce the for- mation of anti-substances. Moro succeeded in finding cow milk precipitin in two instances, and milk proteid in one instance in an examination of 22 anemic bottle-fed children. Bauer found precipitating substances of cow's milk in the blood of an emaciated man. Kentzler, with the aid of the precipitation test, demonstrated milk proteids in the blood of six human subjects in which the gas- tric secretion was disturbed, out of 61 cases that he examined two to three hours after feeding. Although milk is absorbed through the intestines of older individuals only after the splitting up of the proteids, nevertheless in case of an injured mucous membrane, or in greatly emaciated and in very young individuals the direct absorption of unchanged proteids is possible. Ganghofner and Langer succeeded in proving this on very young rabbits, on pigs and on newly born cats, and they succeeded also in demonstrating a precipitin formation in the blood. Schkarin describes similar results after the feeding of cow's milk to young rabbits. Lactoserum. It will be advisable and appropriate to include at this place a subject which as a matter of fact belongs to the chapter dealing with the characteristics of the milk of various species of animals. Milk is an antigen and contains various antigens. After injecting the milk of species A into an individual of species B, the formation of various anti-bodies, precipitins, amboceptors, etc., may be ob- served in the blood serum of the treated individual, which gives to the blood serum the specific characteristics of lactoserum. This specific characteristic is shown by the fact that the cow lacto- serum of rabbits produces a precipitation only when cow's milk is used for the precipitation, but not with milk of women or goats. Works of Bordet, Fish, Morgenroth, Wassermann and Schiitze show the specific action of lactosera. With the aid of such sera the possibility is afforded of differentiating the milk from various species of animals. ~)Q Biological, Chemical and Physical Characteristics of Milk. This however does not end the degree of the specificity, as it is possible with the aid of the precipitation method to differentiate various kinds of proteids of one and the same milk. If the soluble proteid bodies are separated from the undissolved casein by filtration (Schlossmann), then the rabbits which are treated with soluble proteid bodies furnish sera which react only to milk albumin and globulin (Hamburger). It is of further interest that cow-casein sera gave precipitation with cattle blood (Hamburger), the same as is the case with lac- tosera (Landsteiner, Halban, Dungern, F. Meyer, L. Aschoff). Moreover lactosera immobilizes spermatozoa of bulls, and dissolves red blood corpuscles of cattle. However no reaction results from the addition of cattle blood serum containing anti- bodies to cow milk (Meyer). The anti-serum sensitized against cattle blood only gives slight precipitation when it possesses espe- cially high value (Uhlenhuth and Sclmtze). The same conditions were found in preparations of human blood, and women's milk, by Halban and Landsteiner. Uhlenhuth and Sclmtze proved that the differentiation of various kinds of milk shows that the biological method succeeds even when the milk is heated to a high temperature (114 deg. C. in an autoclave) ; if the milk antigen was heated 20 minutes at 120 deg. C., lactosera resulted, which only contained coagulins, but no hemolysins. Sion and Laptes showed that the most varied splitting and decomposing changes of cheese-making and cheese-ripening do not influence the antigen to such an extent that the kind of milk used in making the cheese could not be determined by the biological method. This specificity is of course manifested even in the use of lactosera, but is not absolute, only relative. Lactosera also gave a reaction with the milk of closely related animals, the same as has been established for blood sera, meat sera, etc. Thus for in- stance it is impossible to differentiate sheep's milk from goat's milk (Uhlenhuth, Moro, Gengou), although it is possible to draw conclusions from the comparison of the intensity of the reaction in the homologous milk. Other authors, as for instance Bauer, succeeded in demonstrat- ing by the so-called complement-fixation method, the presence of cow's milk in woman's milk, even when only 1 c. c. of the former had been added to 1000 c. c. of the latter. If specific serum which has been heated for a half hour at 57 deg. C. is mixed with milk and as much complement is added as is necessary for the dissolving of the subsequently added blood corpuscle suspension, with the aid of certain quantities of hemo- lytic amboceptors, then the amboceptors of the lactoserum bind the complement, provided they find in the milk the specific antigen (cow lacto serum-cow milk), and the subsequently added hemolytic system, free of complement, no longer finds complement, so hemo- lysis does not occur, but instead fixation of complement results. If there is no specific antigen present (if the milk to be examined con- tains no cow milk), the complement remains free to be utilized later by the hemolytic amboceptors and the blood cells, for the Anopliylaxia. 57 functionating liemolytic system, and a solution of the blood — liemo- lysis — results. The appearance of "anaphylaxia" may also be produced ex- perimentally with milk (Arthus and Besredka). It may be brought on by raw as well as by boiled milk. The phenomenon of hypersensitiveness as is known, results when a proteid of un- like origin is injected into an animal and later after a period of time the same proteid is re-injected. At the second injection (or only after later ones, depending on the ex- perimental animal and the quantity of proteid), the experimental animal reacts violently with indications of extreme sickness (Von Behring, Eichet and Arthus), which may even result in death from convulsions and pulmonary edema. Miessner succeeded in producing a hypersensitiveness against homologous kinds of milk in guinea pigs, sometimes after one in- jection, but more markedly after repeated subcutaneous injec- tions, and with the greatest certainty after intra-abdominal in- jections of small quantities of raw milk. The best reaction was obtained in the animals after three intra-abdominal injections of . 5 c. c. of milk, on three successive days. After the preparation of the animal 40 to 50 days should elapse before the test which is made by intra-cardial injections. After repeated injections it is possible to demonstrate an ana- phylaxis in most cases even with boiled milk. In order to utilize the biological test for milk differentiation several rabbits should be prepared. This is carried out by in- travenous injections of small quantities (5 or more c. c.) of milk heated for a long time to 65 deg. C. The injection is repeated 5 to 8 times at intervals of 1 to 4 days. In from 14 to 20 days after the last injection the lactoserum may be tested for its effectiveness and if found suitable, may then be drawn. For this purpose the animal is kept without food for one-half day, (in order not to obtain a cloudy serum), a venous hyperemia of the ear is produced by intensive lighting of the ear with the aid of an electric globe, or by rubbing it with xylol, and the vein is then punctured with a fine hypodermic needle. Several cubic centimeters of blood are drawn, which is allowed to coagulate; the blood clot is separated and allowed to stand for 24 hours in an ice chest. The test is made as follows: 1. Establish the dilutions of the serum which are capable of producing a visible precipitation in 3 c. c. of milk dilution, with 1 :60 physiological salt solution, or 2. Establish the dilution of milk with 1 :10 physiological salt solution, in which when mixed in the relation of 1:6, the lacto- serum still produces a precipitation. The most active lactoserum is the best adapted for use. The rabbit is bled to death (slight anesthesia-opening of the thorax-puncturing of the heart), the blood for the collection of the serum is allowed to stand, the serum is drawn off in quantities of 2 to 5 c. c. into small vials, and placed in an ice box for safekeeping. CHAPTER V. PROCUREMENT OF COW'S MILK. As has already been mentioned milk secretion may be retained for a long time by proper emptying of the gland and by the stimu- lation exerted on the gland in the process of emptying. In the presence of incomplete milking, in over-extending the time of milking, and in stasis of the milk, a condition of the gland results, which finally passes into a state of inactivity, when the stimulating condition which is exerted by the retention of the secretion on the secreting epithelia does not again appear with the act of a com- plete milking. The milk secretion therefore is largely dependent on the activity which is exerted on the gland from the outside, such as the sucking act of the calf, or artificial milking. Artificial emptying is carried out in various ways: 1. By closing the upper portion of the teat with the aid of the thumb and index finger, and pressing out the contents of the cis- tern by gradual closing of the hand to a fist in such a way that first the middle finger, then the ring finger, and finally the small finger presses the milk downward and from the opening of the teat. The open hand is passed up again, forcing the milk into the cistern from the upper part of the quarter, the thumb and index finger again squeeze the cistern at its base, and the procedure ends as before. This manipulation is known as "fisting" or full handed milking. 2. By stroking with the closed thumb and index finger from the base of the teat to its point the milk may also be pressed out ("stripping" or "tipping"). This method of milking requires much less strength than the full handed milking, but causes a lengthening of the teats, and is a painful operation for the animal, as it is frequently accompanied by injuries to the tissue, and tear- ing of the mucous membrane. The full handed milking may be carried out by dry milking, while "stripping" succeeds only when the teat and hand are moistened (moist milking), since the necessary smoothness and slipperiness of the skin result only from moistening. If the teat is not pressed with the extended thumb (a brace for the index finger), but the thumb is crooked and the teat is pressed and stringed with the bent index finger against the nail surface of the thumb and the knuckle of the joint, this is spoken of 58 Methods of Milking. 59 as " streak milking" or stripping with bent thumb. The "streak milking" may be completed by stripping or by full handed milking. Full handed milking and this method combined with ' ' streak milk- ing" are according to Henkel permissible; the other kind of milking should be prohibited since the teats are too much extended. The udder should be milked by the dry method since this method of milking is more cleanly than moist milking, in which the fingers become moistened by the milk, and although they slip easily, at the same time they wash off the dirt from the entire teat. It is to be regretted that moist milking and stripping because of their labor saving advantages, are preferred by many milkers on account of their convenience, and even if they are urged to carry out the ordinary dry milking, as soon as they are left with- out supervision they will at once fall into the same fault. The order of milking the various teats differs. Milking from the same side is supposed to induce the development of the side first milked, since the half of the udder first milked is worked with fresh strength while the subsequently milked quarters are not emptied as well on account of the beginning weariness of the milker and therefore they develop less perfectly. The hind quar- ters are either not emptied entirely when the milkers have com- pleted the milking of the fore quarters, or else one hand of the milker rests while he finishes milking the hind quarter with the other. The same applies in milking the teats crosswise, when the hind quarter of one side of the udder is milked at the same time as the fore quarter of the other side. Therefore it is advisable to milk the fore quarters together and the hind quarters together, and the milking should be undertaken first on those quarters which appear to be most distended. With the drawing of the milk from the udder through milking the teats, the complete act of milking is not concluded, as the udder has not yet been sufficiently exhausted in its production. The cause of this may lie in the fact that the milk cannot be emptied by the simple sucking action from the smallest milk ducts and alveoli, or that after the apparent entire emptying, the milk pro- duction still goes on if the gland cells are properly stimulated in their functions. As the flowing in of milk may be accomplished through the so-called "preparation," that is stroking or massage of the bases of the teats and quarters, the same result is possible through the so-called "clean milking," or "after milking," to obtain an addi- tional quantity, which is especially rich in fat. These methods vary and are practiced in different ways in dif- ferent localities. The best known method of "clean milking," and one which has been mostly studied, is that practiced by Hegelund, a Danish veterinarian. This method is divided into the following phases : Procurement of Cow 's Milk. 1. Milking through simultaneous full handed milking, first of the fore and then of the hind teats, until the milk flows no longer. 2. This milking is followed by the " clean milking," which consists in massaging the udder, beginning at the teat up to the base of the teat, and as high as possible extending on to the paren- chyma. While the first act corresponds with the usual full handed milking, the second act massages with a milking motion, the base of the cistern, and the third is carried out by surrounding between the thumb and the hand, and stroking down the lower part of the quarter, that is, through simultaneous pressing against each other of both quarters of opposing sides. 3. The first manipulation of the after milking is carried out by pressing the right quarters of the udder against each other, the left hand being placed on the hind quarter and the right hand on the fore quarter. In case of a large udder, only one quarter is grasped at one time. The hands are then pressed upwards with a rubbing motion on the gland which exerts a massage on the par- enchyma of the udder, this being repeated three times, followed by milking out the cistern. This manipulation is repeated until no more milk is obtained, when the left quarters are treated in a similar manner. In the second manipulation the fore quarters are milked by placing one hand on the outside of the quarter and the other in the division between the two fore quarters. The hands are pressed against each other followed by milking of the teats. Then the hind quarters are milked by placing a hand on the outside of each quarter in such a way that the fingers are turned upwards and the thumb placed in front of the hind quarter. The hands grasp the quarter and are pressed upward; then they are lowered and the milking follows. This is also repeated until no more milk is obtained. In the third manipulation the milker imitates the butting mo- tions of a calf during sucking. The hands loosely surround the teats and the quarters are lifted and pushed against the abdominal wall so that the gland tissue is shaken. This lifting and pushing motion is repeated three times and the teats are then milked out. Following this procedure on the fore quarters the hind quarters are treated in a like manner, until no more milk is ob- tained. The works of Aashamar, Alfonsus, Woll, van der Zande and Henkel, and Wenk, speak of the excellence of the method of Hegelund. According to Henkel the increase of the milk yield in 37 Simmenthal cows was 217.4 gm. (3.4%) per milking on an aver- age. Wenk succeeded in obtaining from 24 cows 4.5 kg. of milk per day more than with the ordinary method of milking. Of course against the increase of yield must be placed the Methods of Milking. 61 additional work and time, which is an additional expense and considerably diminishes the profit derived from the increased yield of milk, and may even nullify it, since it involves the employment of additional help. The principal advantages of Hegelund's meth- od lie in the fact that the milkers are held down to thorough work, and the milk glands are subjected to more correct and appropriate handling. A modified form of Hegelund's method is the so-called "New Algauer milking method, ' ' which combines the acts of the Algauer method with those of Hegelund. The massage of each quarter is carried out with both hands. The method of Sondergaard aims to simplify the time-consuming work of "clean milking" inasmuch as the residual milk is obtained by a wide extensive hold of the halves of the udder at their bases, and pressing at the same time and stroking downwards. The principal factor in each method of milking is that the udder should be thoroughly emptied, and this can only take place when each part of the milk gland is stimu- lated by massage to the limit of its production. CHAPTER VI. INTERNAL INFLUENCES ON THE CHARACTER OF MILK. The influences which must be considered in the formation of milk may be separated into internal influences which lie in the individual characteristics and in the immediate condition of health of the animal, and in outside influences, such as stabling, feeding, *"etc., which again act only in that they influence the internal condition. As internal influences may be considered the characteristics of the breeds, strains, family, individual, age, influences of the lactation period, pregnancy, and the general and local conditions of health. The outside influences may be considered under care and attendance, feed, medical treatment, climatic influences, meth- ods of milking, etc. Following this outline the internal influences on the formation of milk will be considered first. Breed, Family, Heredity, Individual Characteristics. Age, Lactation and Other Special Conditions of the Individuals. The influence of the breed on milk formation is generally known. There are beef breeds which fatten especially well, milk breeds in which milk production is especially prominent, and breeds which possess the ability to produce both milk and meat. Breeds of low lands and their crosses produce more milk with lower percentage of fat than breeds from the highlands. Ac- cording to the quantity of milk produced, the breeds are headed by the Holsteins, Angler, Oldenburger, East Friesian, Breiten- burger, Wilstermarscher, Dithmarscher with 20 to 25 liters of milk per day per animal at the height of production, with percentages of fat from 2.5 to 3 to 3.4. Smaller quantities of milk are given by the grayish-brown mountain cattle, the Swiss and Algauer, with 3.6 to 3.7% of fat, and the spotted mountain cattle, for instance Simmenthal, Misbacher and Pinzgauer, with 3 . 5 to 4% of fat. The fat content of the Westerwalder and of the Schlesian red cattle 62 Influence of the Breed. 53 varies between 3.5 to 3.6%, while the English Shorthorns and Ayrshires give 3.7 and 3.8% ; the richest in fat is the milk of the Vogelsberger and Harz cattle (3.9 to 4.2%) and that of the Jer- sey with 5 to 5.4% (Ramm). Reference should be made here to the following results of tests of production made with Simmenthal, Vogelsberger, Westerwaldern and Lahn breeds of cows. Simmenthal Lahn Vogelsberg Westerwald Average weight of the tested animals 578* 502 427 395 kg Average production 2495 2650 1919 1878 Fat contents 4.001% 4.001% 3.74% 4.1% Individual maximum production .... 4562 3955 3800 3234 kg For 100 kg. body weight 431.1 528 450 477.4 " *The cow with maximal production stood last year in ninth place. 17 cows pro- duced over 3000 kg. each. The fluctuation is not in fat content alone, but also in the other solid substances. Milk which is rich in fat as a rule contains more of the other solid constituents as well. The proportion of individual factors of the dry substances is variable ; in cattle from the highlands for instance, the casein was 76.24% of the dry sub- stance, in cattle of the lowlands it averaged 73.78% (Fischer). Fat from the milk of the mountain breeds is generally under otherwise similar con- ditions, the richest in fatty acid of molecular weight; the fat globules of the breeds of the lowlands are smaller than the fat globules of the cattle from the highlands. Babcock mentions that Jersey and Guernsey cows produce larger and more uniform fat globules than the Holsteins, while Ayrshires have small, irregular fat globules in their milk. Milk with large fat globules is preferable for butter making, since these produce butter of good consistence and good taste with a low melting point. The ash content of milk from highly bred animals is some- times somewhat lower in CaO and P205 than that of the common breeds, but the fluctuation is such that definite deductions cannot be established. In highly improved breeds Pages found: CaO in 0.143 to 0.227% and P265 in 0.18 to 0.273%; in common breeds the same elements amount to 0.15 to 0.204, and 0.153 to 0.296, respectively. These characteristics of breeds are general, but they are not so constant that individual strains, individual families, and espe- cially particular animals may not present exceptions. This fact forms the basis of breeding for increased milk production within individual breeds. Good milk cows should be bred to bulls, whose mothers and sisters were or are known to be good milkers, and then it may be expected that the good characteristics of the family will be inherited. In experiments extending over eight years Hogstrom tried to discover whether the characteristics of producing milk of certain fat contents could be transmitted by bulls to future generations. The large majority of female progeny produced a higher fat content than the milk of their mothers, which points to a positive influence from the male animal. In cases in which the fat content of the mother was 3.08 to 3.77% the fat content of the milk of the daughter increased mate- rially;- but as soon as the fat reached or exceeded 3.77%, the influence of the bull was no longer sufficient to further increase the percentage of fat, and the percentage re- mained lower than in the mother. (The experiences of Hogstrom 's were confirmed by the rule laid down by Galton.) 54 Internal Influences on the Character of Milk. The great variation which occurs in the milk production of individuals of the same breeds depends on hereditary qualities. According to the statistics of the dairy control station at Algau, animals of the Algauer breed produce : Quantity of Fat Percentage Quantity of Milk Fat Maximal production 5201 kg. 4.603 181.93 kg. Minimal production 1255 kg. 2 . 493 45 . 31 kg. Difference 3946 kg. 2.11 136.62 kg. Among 50 of the Jeverland breed Maximal production 8699 kg. 3.713 286.76 kg. Minimal production 2449 kg. 2.482 75.21 kg. Difference 6250 kg. 1.231 211.55 kg. The production of single individuals during the lactation period is sometimes remarkably large. Some of the following data relative to production is taken from Kirchner's handbook. Percentage Quantity Author Cow Quantity of Milk of Fat of Fat Kirsten Wesermarsch .. 11291 kg. 2.78 324 kg. 5th calving Kirsten East Friesian . . 9047.75kg. 3.07 277.77kg. 6th calving Woll Guernsey 6768 kg. 5.745 388.8 kg. "Yeksa Sunbeam" Kirchner also quoted a case in which a farmer observed that a seven-year old cow in the second month after calving still pro- duced 50 liters of milk per day. Just as the quantity of milk and percentage of fat may vary, so also the fat-free solid substances may vary in the individuals, although only within narrow limits. The rule also holds good here that an individual with milk rich in fat will at the same time pro- duce more fat-free solids. Normally fed animals which are not individually large pro- ducers cannot be brought up to a remarkable increase of produc- tion through any agency. The elimination of the poor assimilators of food in favor of good producers, which is a matter of economic necessity, should be based upon the capacity of the individual cow to properly utilize her food. Only through a systematic test of milkings and production records can the profits of the dairy be increased. In judging individuals as milk producers by their external conformation, the following rules of the German Society for Breeds and Breeding may serve as a basis : Influence of Age. 65 (a) Heavy milk production is usually associated : 1. With low body weight, 2. With low measurement at the shoulder, 3. With a straight back, although slight deviations should not be considered as signs of small milk productiveness. 4. With more or less prominent hips and rump according to the characteristics of the breed. 5. With the more pronounced depth of thorax ; heavy milkers are often narrow and flat chested ; 6. With long shoulders, 7. With long rumps, 8. With long, narrow head, 9. Generally with fineness of horn, 10. With fine bony structure ; 11. The most important is the udder. The best cows have large udders of spongy-granular consistence, with large tortuous mammary veins, large milk wells, and easily movable skin. The skin should lay together over the perineal surface of the udder in 4 to 6 or more large, well developed folds. The udder should collapse thoroughly after milking, and the animals should be easy milkers. Relatively early calving seems to have a good in- fluence. The possibility of estimating the qualitative production of milk from external conformations is only very slight. As a rule, as shown by investigations, the smaller and shorter animals with fine long bones produce milk of higher quantity, and above all milk with a large yield of fat. The productiveness of one and the same individual varies, especially with age and the lactation period. Cows with the first calf, provided normal conditions prevail, do not produce as much milk as after subsequent calvings ; as a rule when cows reach the age of 7 to 9 years with the fifth and sixth calf, the maximum pro- duction is obtained. With the advance of age the production again gradually recedes. The proportion of solids is higher in cows with the first calf than in those which have calved several times; the quantity of fat on the other hand, as compared with that of older cows, is smaller (Teichert, Hittcher, Hogstrom, Vieth and others). The variations which are manifested in the production of milk during single lactation periods are considerable, and depend en- tirely upon the individual, as does the length of the milking period. For a few days after parturition a product is secreted which has very little in common with milk, and which may be considered as a product of glandular inflammation as a result of physiological irritation. It corresponds strikingly in its appearance and com- position, as well as in the microscopical appearance of its cream and sediment with the inflammatory product of the milk gland. This product called colostrum is a yellowish or even yellowish-red, slimy fluid, with an acid reaction. Corresponding to the increased content of albumen, globulin and 55 Internal Influences on the Character of Milk. fat in colostrum, as compared with ripe milk, the amount of dry substances in colos- tral milk is very high, and its specific gravity is increased. The amount of urea, cre- atinin, cholesterin, and lecithin in colostrum is increased. The milk at this stage is rich in fat-containing glandular epithelium in the form of foam cells, and seal-ring- shaped cells with so-called caps and moons, and in albuminophores. Numerous leuco- cytes are to be found, and during the first days red blood corpuscles are also present in great numbers. According to Emmerling, cow colostrum, on the rnorning after the birth of the calf, consists of 76.14% of water, and 23.86% of dry substance, of which 4.705% is casein, 0.58 albumen, 8.320 globulin. Compared with normal milk, the fat content is increased or diminished, the milk sugar diminished, and the ash contents increased. Engling found the following values for colostrum: Immediately after After 10 After 24 After 48 After 72 calving hours hours hours hours Specific gravity 1.068 1.046 1.043 1.042 1.035 Solids 26.83 21.23 19.37 14.19 13.36 Casein 2.65 4.28 4.5 3.25 3.33 Albumin and globulin 16.56 9.32 6.25 2.31 1.03 Fat 3.53 4.66 4.75 4.21 4.8 Milk sugar 3.0 1.42 2.85 3.46 4.1 Ash 1.18 1.55 1.02 0.96 0.82 The composition of the ash differs from that of ripe milk, as may be observed from the findings of Schrodt and Hansen: Ripe Milk (10 days Colostrum after calving) 17.4 24.12 10.10 8.72 22.99 22.69 6.88 2.92 0.42 Traces 2.82 4.10 34.30 30.73 6.85 8.30 The ferments in colostrum also deserve special consideration. The amylase content is considerably increased, also the amount of catalase. During the colostral period the milk further con- tains hemolytic amboceptors and increased complement. Formalin methylene blue is not decolorized. The reaction of the colostrum is acid. The colostral period lasts from 3 to 5 days after calving. In heifers the transition period results more slowly than in old cows (Deisman, Hittcher). Up to the end of this period there is a constantly increasing approach to the properties of ripe milkr together with an increase in the yield, which continues to increase until the first or second month, and then gradually recedes and finally rapidly diminishes towards the end of the lactation period. The reduction of the milk yield corresponds with an increase in the percentage of fat. The fat globules become smaller and more numerous. At the end of the lactation period the milk again assumes the character of colostrum, becoming especially rich in chlorine, and sodium oxide, while the phosphoric acid and the potassium contents appear diminished. The milk becomes salty, bitter, and its reaction alkaline. The entire time of lactation or one lactation period, usually lies between the birth of two calves, Influence of Disease. 67 and is divided into the lactation period and the dry period. Good milk cows give milk on an average for 300 days. Cows which are not bred again, or which cannot be impregnated, may have a con- siderably longer lactation period. During estrum a considerable diminution of milk in quantity and quality may be observed in cows. Sucking calves may at this time become affected with digestive disturbances. Hittcher and Neumann state that the quantity and the proportion of fat dimin- ish, while the casein contents and the specific gravity of the milk are increased. There are however no set influences in one and the same animal, and still less so in different individuals. Sometimes the quantity of milk even increases, and not infrequently the milk becomes abnormally rich in fat (Martiny). Fascetti and Bertozzi found diminished quantity, increased specific gravity, and in- creased dry substance, which they supposed resulted from the in- creased proteid contents, especially from the increase of fat. The volatile fatty acids in the fat according to Nilsen are diminished, and the degree of acidity of the milk is frequently increased (Mez- ger). As a whole, however, the milk is not materially changed (Weber). No observations have been made on the influence of coition and the beginning of another pregnancy. However the milk of cows far advanced in pregnancy frequently has a lower value. It coagulates sometimes as early as in the sixth, seventh, or eighth month of the gestation period. Finally it becomes slimy, yellow, and shortly before the cow goes dry it shows a similarity to colos- trum. The amount of phosphoric acid and lime, contrary to that in colostrum immediately after calving, is diminished, and the taste is bitter and rancid (Backhaus). If cows are spayed 5 to 6 weeks after calving the milk is supposed to be richer in fat, casein, and ash. The lactation period of such cows is considerably lengthened, according to Gouin ex- tending to 6 years. Lajoux on the other hand states that in healthy animals the quality of the milk remains the same, but during the course of lactation the cows do not dry off so rapidly and the yield is therefore greater. Milk which is produced after abortion is supposed to be sim- ilar to that of ripe milk (Schaffer and Hess). The lactation period however is short, and the milk yield small. If the cow remains farrow for a long time a greater yield is obtained. This however is only slight, and does not compensate for the shrinkage during the latter part of the milking period. Influence of Diseases. Relatively little is known of the chemical changes which milk undergoes from the influence of general affections of ani- mals. We are in possession of better information relative to the gg Internal Influences on the Character of Milk. occurrence of specific disease agents in milk, and it is known that these pass into the milk either directly from the blood being then eliminated with the milk, or else they reach the milk through sub- sequent contamination of the milk with excretions. A rapid diminution of the milk yield is characteristic in all acute diseases associated with great pain and fever, and in some cases a sudden cessation of the secretion may be observed. Whenever the yield of milk of a cow suddenly shows a con- siderable diminution, all of her milk should be excluded from mar- ket, even though the animal shows no visible affection and before the disease can be recognized as a general or specific affection. In the sense of the pure food law the milk of every severely affected cow should be considered unfit for food without any further con- sideration. Sometimes this unfitness of the milk is manifested by strong objective perceptible changes, as compared with the secretion of healthy animals. The milk may become bitter, salty, have an increase of ash and albumin, and coagulate more rapidly than healthy milk (Jensen). The fat content of the milk is at the same time diminished or in- creased, while the sugar and ash contents may show fluctuation. The amount of catalase present, according to Spindler, may in- crease considerably, especially in cases of peritonitis and tubercu- losis. The reaction of the milk remains acid or becomes slightly alkaline. According to Schnorf, most of the internal affections, even when the udder is not involved, produce a diminution of sugar and proteid contents as a result of increased metabolism. The electrical conductivity of the milk of animals with general affections is subject to great fluctuations. After tuberculin injec- tions with subsequent fever, the milk shows a slight increase in its electrical conductivity. The index of refraction in pathological milk is normal, and not diminished; the temperature at which freezing occurs is not infrequently higher. During the course of individual diseases the following should be considered: An elimination of toxins and toxic products of metabolism with the milk is to be feared in all septic and pyemic diseases. If with this there is a possibility of contamination with pathological excretions, as for instance in septic metritis, hemorrhagic or ichorous enteritis, or in the retention of putrid afterbirth, the milk should be considered harmful. In septic metritis the infective agents pass from the uterus into the meat and into the udder, from which they may be eliminated. Basenau demonstrated the Bacillus morbificans bovis, a meat poisoning organism of the colon typhoid group, in the meat during the existence of septic metritis. The stapylococci and streptococci which are frequent participants in mixed infections of the uterus, are also eliminated with the milk, provided the udder has not already ceased its secretion. Influence of Disease. Milk from cows affected with acute and sub-acute intestinal inflammations should be judged in the same way as milk from animals affected with septic metritis. The ingestion of milk from cows affected with bloody or fetid diarrheas should be especially guarded against. The appearance of sickness in man after the ingestion of such milk has been satis- factorily proved by Gaffky and Follenius. Two assistants and a helper of the Hygienic Institute of Giessen drank milk of this character and became sick with dullness, headaches and chills. After two days diarrhea, vomiting and high fever appeared. The clinical manifestations in the two assistants simulated those of typhoid fever, while in the helper they were similar to those of Asiatic cholera. The milk originated from a cow affected with hemorrhagic enteritis. Gaffky demonstrated rapidly growing and strongly virulent colon bacilli both in the bloody excrements of the cow and in the stools of the affected patients. In the presence of infectious diseases the milk of the en- tire stable should be withdrawn from use, or should be rendered safe by suitable treatment, as for instance by pasteurization. Such milk should never be sold as certified or infants' milk. Jensen ex- tends this prohibition even to milk from stables in which white scour of calves, and other calf affections of an infectious nature have occurred. Dangerous properties of the milk should also be considered in the appearance of other diseases, as for instance malignant catarrh- al fever, purulent broncho-pneumonia, traumatic pericarditis, rin- derpest, etc. (Bongert). In all cases of hemorrhagic, purulent, acute or chronic inflammations of the kidneys the milk should be judged similarly to milk from animals with intestinal inflamma- tions. In such affections the freezing point of the milk approaches zero, and the refraction index is lower. At the same time these values in animals affected with inflammations of the kidneys vary extensively. Special Infectious Diseases. Tuberculosis of animals, especially its hygienic importance, is considered here in connection with tuberculosis of the udder. It should be mentioned at this point that some investigators be- lieve that the toxins of the tubercle bacillus pass into the milk. A change of the quality of the milk will occur only in cases in which the advanced chronic affection of the animal results in lasting emaciation, or when an acute attack of the disease, associated with fever, appears during the chronic course of the disease. In tuber- culosis the milk may become bluish, and poor in fat, the sugar and proteid substances may be diminished, or the latter may be even in- creased (Storch). Several tables, which indicate the experimental results of Monvoisin, are taken from Grimmer 's "Chemistry and Physiology of Milk." Internal Influences on the Character of Milk. 1000 gm. of milk contained Tuberculous cows without Healthy cows tuberculosis of the udder. Acidity as lactic acid 1 . 543 0 . 664 1 . 292 Total nitrogen 5.87 8.67 4.21 Fat 46.5 29.6 59.7 Sugar 43.5 29.8 43.9 Solids 142.3 126.05 147.5 Ash 7.3 8.2 6.7 Chlorin (sodium chloride) ... 1.4 4.13 1.05 Freezing Point — 0.55 Eefraction at 15 deg 1 . 3434 1 . 3416 1 . 3442 In rinderpest, according to Busson, the amount of fat and sugar diminished rapidly, whereas the casein, albumin and salt increased. The passage of the contagion of rinderpest into the milk in a direct way from the blood is probable; the milk, how- ever, can be contaminated with certainty through infectious secre- tions and excretions. Einderpest is of no practical importance from the standpoint of milk hygiene, to most of the European countries (with the exception of Turkey), since it has been eradi- cated with the aid of veterinary police measures and even in the event of any possible introduction, it will be immediately sup- pressed. Milk from cows affected with contagious pleuro-pneumonia is supposed to have caused the death of children (Randou, Lecujer and Wiedemann). Secretion of milk is immediately reduced at the onset of this disease, it becomes poor in fat and sugar, richer in albumin and ash, its appearance resembles that of colostrum, and its taste is peculiar. The contagion of pleuro-pneumonia appears to pass into the blood but rarely, and therefore its elimination in the milk can occur only exceptionally, if at all. Contagious pleuro- pneumonia is also subject to the most stringent veterinary police measures, and therefore has but little practical importance for milk hygiene. Similar conditions prevail with pox of cattle. This disease however demands our interest for the reason that the infectious agent of cow pox must be considered as a mild form of smallpox of man. Cattle usually become affected through transmission of the disease from naturally infected men, or from those vaccinated with cow pox. The infection occurs if during milking the contagion of pox is rubbed into visible or invisible wounds of the skin of the udder. The infected teats manifest roundish or oval, hard papules of the size of a pea, which after 1 to 2 days change into yellowish- white vesicles of a mother-of-pearl luster. After ripening into pus- tules which requires from 8 to 10 days, the lesions show a charac- teristic depression in their center, the so-called navel of the pox. They either rupture and suppurate, or dry and heal, leaving a superficial scar. The udder becomes sensitive to pain, the milk is thinner, and Influence of Disease. of lower specific gravity, but richer in albumin (Jensen). The injection of the contagion of pox into the ducts of the udder results in the development of pox vesicles on the walls of the milk ducts. After 2 to 3 days a swelling of the udder, with increased sensitive- ness, develops and the secretion is changed. It becomes purulent and bloody on the eighth to the tenth day (Lienaux and Hebrant). Transmission from animal to animal may be brought about by milking, and the entire herd in a stable may rapidly become af- fected. The course of cow pox is usually benign. According to Herz the milk becomes rich in cells, contains colostral bodies, and it has an unpleasant taste. Careful examination showed the fol- lowing results : Beginning of After 13 After 40 observations days days Specific gravity of the milk 1 . 0265 1 . 0270 1 . 0215 Specific gravity of the whey 1 . 0245 1 . 0235 1 . 0209 Acidity according to Soxhlet-Henkel . 5.3 6.6 4.1 Fat 5.36% 4.02% 5.54% Solids 13.31 11.82 12.25 Fat-free solids 7.95 8.81 6.72 Ash 0.72 0.72 0.8 Transmission of pox from cattle to man is of course very readily possible, and is not at all uncommon as a result of milking affected animals. After the ingestion of infected raw milk the pox exanthema may develop on the face (Jensen). The so-called false or gangrenous variola which may be fre- quently observed on the teats of fine-skinned, fresh milking animals should not be mistaken for true pox. These eruptions are pro- duced by the ordinary pus-producing organisms, which have been rubbed into the skin during milking or have penetrated the skin by means of various injuries. Small furuncles and skin abscesses result, which heal without influencing the formation or secretion of the milk. Healing is of course retarded through the act of milk- ing, and during the presence of the pus cells, blood and pyogenic organisms may pass into the milk in small quantities. These false pox lesions are not very important. Of much greater importance than cow pox is foot-and-mouth disease which sometimes appears extensively. This is a highly acute febrile disease which is transmitted to cloven-footed animals with remarkable ease. The most striking symptom which occurs in association with the disease, the vesicular eruptions, may also affect the udder, and especially the teats. The udder swells, becomes painful, and red-bordered vesicles develop in sizes up to that of a walnut, which burst during milking or spontaneously, leaving painful ulcers. During the beginning of foot-and-mouth disease the yield of milk is considerably dimin- ished, sometimes one-quarter less than the usual yield, as a result 72 Internal Influences on the Character of Milk. of the febrile affection and on account of the inappetence due to the pain caused by the vesicles in the mouth and on the feet. The effect of the disease upon milk secretion varies according to the individuals, the age and the lactation. Siedamgrotzky, Weber and Born have published the effects of the disease on milk secre- tion during outbreaks in certain herds. In 43 cows the quantity of milk at the height of the disease dropped from 745 to 364 liters, and again rose after the eradication of the outbreak to 522 liters. Thirty cows of another herd gave only 30 liters instead of 300 liters of milk during a period of eight days. In a third herd the quantity of milk dropped from 510 to 260 liters, later rising to only 350 liters. Other figures showed a decrease from 750 to 280 liters, with a subsequent rise to 400. The diminished yield per cow per day was from 5 to 6 liters and even more. In cows that have been milking for a long time the loss in milk reaches as high as 75 %, in animals in the middle of the lactation period up to 43%, while in fresh milkers it may amount to 55% (Hutyra and Marek). Sugar and fat contents diminish, but at times the amount of fat may become considerably higher. The volatile fatty acids are diminished, but the milk contains more albumin and salts, an in- creased amount of thrown-off epithelium, colostral cells, pus cells, and also red blood corpuscles (Lavena, Kalantar, Herberger, Kreis, Vogler and others). The catalase content is increased even if the udder manifests no changes (Bertin-Sans and Gaujoux). Honigmund examined five cows affected with foot-and-mouth disease, one of which was not visibly affected on the day of the examination although already infected. The individual data in- side of nine davs were as follows : Quantity Tempera- Specific Fat Con- Nitrogen- of Milk ture Gravity tents ous Subs. Sugar Solids Ash 15 L. 38.7 1 .032 3.05 2 .99 4.24 11 .62 0.74 6 — 7 " 39.6 1 .031 5.4 2 .97 3.63 13 .00 0.63 6 — 7 " 38.9 1 .030 4.3 2 .99 3.80 12 .81 0.89 about 8 " 39.0 1 .030 3.43 3 .04 3.91 11 .33 0.65 8 — 10 " 38.6 1 .031 3.06 3 .04 4.15 11 .01 0.70 8 — 10 " 38.4 1. ,029 2.9 3 .1 4.49 12, .03 0.69 about 10 " 38.6 1 .030 2.84 3 .19 4.57 11 .21 0.71 10 - 11 " 38.5 1 .032 3.45 3 .24 4.41 11 .77 0.67 about 12 " 38.4 1 .031 3.25 3 .33 4.38 12 .30 0.70 It appears also from the other investigations of Honigmund, in which the animals showed symptoms of the disease as early as on the first day of the examination, that the fat and ash content is greater in the first day than in normal conditions. The total solids and also the fat-free solids fluctuate considerably. When catarrh of the milk ducts becomes associated with foot- and-mouth disease, the milk becomes yellowish, of a rancid, bitter taste, colostrum-like, and similar to the secretion during other in- Foot-and-Mouth Disease. 73 flammatory conditions of the udder, that is, slimy, watery, and in- termixed with coagulum. It is an important fact that milk from animals which are af- fected with foot-and-mouth disease will contain the virus of foot- and-mouth disease, if it has been contaminated by the vesicular contents. Nocard succeeded in proving, however, by careful ster- ile drawing of the milk from cows affected with foot-and-mouth disease, that the milk does not contain the virus of foot-and-mouth disease as it leaves the udder. Nevertheless it is not satisfactorily proved that a direct elimination of the virus may not take place at the beginning of the febrile state, as at this time the virus is present in the blood. If the udder itself is affected by the eruptions of foot-and-mouth disease it is hardly possible to avoid contamination of the milk with the vesicular contents. Considering the ease with which the virus of the disease is spread, it may be assumed that the entire milk of a herd affected by the disease, under ordinary conditions of milk production, contains the contagion of foot-and-mouth disease. Strict veterinary police measures must be inaugurated to prevent the spread of the disease. Sale of the milk should be permitted only after sufficient heating. The maintenance of a temperature of 70 deg. C. for one-half hour will make the milk perfectly safe. Milk containing the living virus of foot-and-mouth disease must be considered deleterious to human health, since it has been established by experiments and observations that the disease is transmissible to human beings. Vesicular and ulcerated inflam- matory changes of the buccal mucous membrane with fever and general symptoms develop with possible vesicles and ulcers on the hands, arms, breast, lips, ears, and in the throat. Vomiting and diarrhea may be associated with symptoms of a gastro-intestinal inflammation, and the affection may even terminate in death. (Bus- senius and Siegel, Jensen, annual reports of the Imperial Board of Health, Hertwig, Stickler, Schreyer, Krajewski, Walkowski, and others). Bongert suggests the separation in dairy stables of the non-affected, slightly and severely affected animals into isolated groups, and in order to reduce the economic losses as low as pos- sible, the milk of these groups should be treated in different ways. Heated milk from the non-affected animals for instance, could be utilized as infant's milk. The milk from slightly affected animals could be marketed as ordinary milk [after pasteurization], while the milk from the severely affected cows or milk changed in its consistence, should be excluded from consumption even in a heated condition. Even with this separation the losses will necessarily be high as a result of the enforcement of stringent sanitary regu- lations. According to Ebert sour milk 3 to 4 days old is no longer capable of transmitting the infection. The transmission is possi- 74 Effect of Internal Influences. ble through cheese and butter (Frohner, Ebstein, Thiele, Schnei- der, Frick, Frohlich). The general rules which have been indicated above obtain also in changes of the milk in malignant oedema, blackleg, or parturient blackleg of cattle. Transmission of these diseases through the consumption of milk from affected cattle, or through the diseased products of contaminated milk, is not to be feared; besides milk production ceases very rapidly in the affected animals. The same rules should apply in judging milk from animals affected with hemorrhagic septicemia, a disease which is pro- duced by a bi-polar bacterium. This disease is transmissible to calves, through sucking or feeding milk from affected animals. Anthrax of cattle should also be mentioned. This runs in an acute or sub-acute form, and as a rule is associated with a sudden cessation of the milk secretion, which occurs even as early as at the beginning of the fever. The anthrax bacilli only multiply towards the end of the disease sufficiently to cause a direct passage from the blood into the milk. If the secretion has continued to some extent this direct passage is possible even if no hemorrhages, such as are typical during the course of anthrax, have developed in the parenchyma of the udder. The demonstration of anthrax bacilli in milk has been accomplished micro scopically, and by inoculation and cultural experiments, but not in all the cases which have been examined (Bollinger, Chambrellent and Mous- sou, Feser, Monatzkow). In severe cases the milk becomes yellowish, bloody and slimy. At the appearance of the fever the fat and sugar contents are in- creased, while the proteid contents are diminished. The danger of infection through the ingestion of raw milk con- taining bacilli is slight, since the anthrax bacilli are digested by the gastric juice. More dangerous than the bacilli which may pass into the milk from the blood are the anthrax spores which may reach the milk through contamination with manure of affected animals, or through straw and stable dust, since the resistant spores are not destroyed by the gastric digestion. The virus may also be present at times in normally healthy animals after they ingest food containing anthrax spores.. The milk may become infective through contamination with feces from such bacilli car- riers. In spite of the fact that there are remarkably frequent opportunities to obtain milk with bacilli and spores from localities in which anthrax persists epizootically as a disease of the soil, yet only one anthrax infection of man is known to have occurred through the ingestion of milk. This resulted in a patient with typhoid fever, who after drinking iy2 liters of milk became affected with intestinal anthrax. The milk was derived from a cow with a malignant pustule on the udder, which had died in the meantime from anthrax. Rabies. 75 Lehnert states that the calf of a cow affected with anthrax re- mains "well, although it may suck the mother through the entire course of the disease. Even though milk offers a splendid nutritive medium for the anthrax bacillus, an increase of bacilli only occurs during the first three hours. Keeping the milk at room temperature for 18 to 24 hours, is followed by the death of the bacilli (Caro). At the beginning of souring the vegetative forms of the virus are quickly destroyed; the spores however remain active (Inghilleri). If anthrax bacilli are cultivated in milk, coagulation occurs under the rennet action of the peptonizing bacterial ferments. The coagulum again slowly dissolves, and the milk separates into fat and whey. Less important than anthrax is rabies, as this disease occurs much more rarely in cows. According to Nocard and Bardach the milk of animals affected with rabies contains the virus. Never- theless the danger to man from the ingestion of such milk is hardly probable, since it is impossible to affect experiment animals by feeding fresh milk (exceptions are rats and mice). A nursing infant of a woman affected with rabies remained well, although it was fed with the milk of the patient until one day before her death (Bardach). The uninjured mucous membrane of the mouth, pharynx, and the intestinal tract does not offer opportunity for infection. This opportunity is afforded only when destruction of tissue and small wounds permit the entrance of the contagion. Thus for instance Galtier succeeded in producing rabies through rubbing brain material of rabid animals into the mucous mem- brane of rabbits. According to the observations of Virschikowsky the rabid virus is destroyed by the gastric juice. Very little, or nothing at all is known relative to the special relationship of other infectious diseases to milk, as for instance malignant catarrhal fever, croup of cattle, the blood diseases of cattle caused by spirochaetes, trypanosomes and piroplasma, or in- fectious vaginal catarrh and infectious abortion. In the presence of infectious vaginal catarrh and contagious abortion the milk secretion is supposed to be diminished. It should be remembered that in such affections the passing of the disease agents from the blood into the milk is possible. [That the bacillus of infectious abortion is eliminated by the milk has been definitely established. See Bureau of Animal Industry Circular No. 216]'. In a case of icterus in a woman Mayer observed the passage of bile acids, especially taurocholic acid into her milk. Finally two other diseases should be mentioned which may be transmitted from animal to man: 1. Milk Sickness. A rather peculiar disease, called "milk sickness," is found in the central part of the United States, where it at times occurs as an epidemic among cattle and people. In cattle, the first indication of disease is dullness, followed by violent Effect of Internal Influences. trembling and great weakness, which increases during the suc- ceeding day until the animal becomes paralyzed and dies. Through the ingestion of flesh, milk, or dairy products of an affected animal the disease is transmitted to man or to another animal, and at- tacks produced in this way most frequently prove fatal. In man the disease develops with marked weariness, vomiting, retching, and insatiable thirst. Respirations become labored, peristalsis ceases, the temperature is subnormal, and the patient becomes apathetic. Paralysis gradually follows and death takes place quietly without rigor mortis. Many efforts have been made to elucidate the question re- garding the nature and cause of this disease, but although many theories have been discussed none of them has so far been general- ly accepted. Some investigators hold that the disease is of micro- organismal origin, some that it is due to auto-intoxication, while others think it is caused by vegetable or mineral poisons All seem to agree, however, that the disease is limited to low, swampy, uncultivated land, and that the area of the places where it occurs is often restricted to one or a few acres. Furthermore, when such land or pastures have been cultivated and drained the disease dis- appears completely. The discovery of a new focus of this disease in the Pecos Val- ley of New Mexico in November, 1907, gave Jordan and Harris the opportunity of studying this peculiar affection by modern bacter- iological methods. As a result they have succeeded in isolating in pure cultures from the blood and organs of animals dead of this disease a spore-forming bacillus which they name "Bacillus lactimorbi." With this bacillus they have reproduced in experi- ment animals the symptoms and lesions peculiar to milk sickness or trembles, and from these animals the same organism has been recovered in purity. It therefore appears to have been demon- strated that the bacillus in question is the probable cause of the disease. As Jordan and Harris have already indicated, more com- prehensive studies, based on a larger supply of material, are desirable in order that the many obscure and mystifying features connected with the etiology of this rapidly disappearing disease may be elucidated. From the above facts it seems evident that milk sickness is an infectious disease communicable to man, and the cattle owners should therefore not be permitted to make use of the meat or milk of affected animals for human consumption. Trans.] 2. Malta Fever. On the coast of the Mediterranean, in South Africa, India, China, Philippines, America, and especially on the Island of Malta, there occurs in goats a disease which exists in the animals without producing any or at most only very slight symptoms. Cows may also possibly be affected. 'The infected animals eliminate for months, frequently at intermittent periods, the virus of the disease (Mlcrococcus melitensis, Bruce). Follow- Mastitis. 77 ing the ingestion of such milk " Malta Fever" develops in man. It lias a protracted course with recurrences, and is accompanied by anemia, headaches, rheumatic pains, constipation and swelling of the joints. Malta fever terminates fatally in about 3% of the cases. The goats show on postmortem, swelling of the spleen and lymph glands, frequently also inflammations of the kidneys and lobular pneumonia. The virus is relatively resistant against souring of the milk, but at 70 deg. C. it dies in 10 minutes. According to Zammit about 10% of all the goats on the Island of Malta eliminate the virus, while 50% of the animals show by the agglutination test that they are or have been under the influ- ence of the Micrococcus melitensis. We are. in possession of better information concerning the changes which milk undergoes in inflammations of the udder than we have regarding the effect on the milk secretion as a result of general diseases, or regarding the importance of milk from affected animals from a hygienic standpoint. Changes in Appearance, Consistence, Contents, Etc., During an Attack of Mastitis. Relatively very little is known as to the influence of the dis- eases of the udder on the chemical and physical character of the milk, although it is well known that with the changes in function and condition of the organ the product is also changed, as com- pared with the product of the normal gland. Even in the same disease the product varies in accordance with the intensity, dura- tion and the extension of the disease, the same as it naturally varies in accordance with the nature of the injury to which the parenchyma is subjected. As a result of these conditions the re- sults of the data of different authors vary considerably. It may be said in general that in affections of the udder the proportion of the proteids, sugar, salt, fat, and enzymes in the milk becomes altered and that the relation of the individual pro- teids, the salts and the enzymes, also undergoes fluctuations. In acute and greatly extended chronic inflammations, both fluid and cellular constituents of the blood may pass into the milk, cells of the parenchyma are thrown off, coagulation sets in, and briefly, the milk changes more or less rapidly in appearance, taste and contents, so that it deviates considerably from the milk of healthy cows. At times none of these characteristics appears, especially in the early stages of chronic inflammations of the udder, or after the subsidence of the acute symptoms, and it is then only possible with the aid of certain methods of examination to differentiate such affected milk from normal. Therefore of special importance to milk hygiene are the chronic inflammations, and inflammatory stages in which the 78 Effect of Internal Influences. changes of the secretion appear slowly, and relatively late, while inflammations of an acute character very quickly produce a tre- mendous change in the secretion, the mixing of which with market milk would be the grossest negligence. It is to be regretted that such cases occur. Appearance of affected milk : In forms of inflammation which are associated with rapid development, painful swelling and in- creased temperature of the udder, the milk usually has a bloody discoloration, later becoming yellow (colostrum-like), and finally changes into a custard, or honey-like secretion, in which thick, yellow and yellowish-brown flakes are suspended in a more or less clear serum or plasma. Such changes are observed in samples of milk in acute forms of mastitis, through infection of bacteria of the colon group, in mixed infections, in acute attacks or in great extension of strep- tococcic mastitis, and in infections with the Bacillus pyogenes, etc. In chronic affections the milk changes only slightly or not at all during the beginning of the disease, or it may appear normal long before the disease as such is considered cured. If such nor- mal appearing milk from affected quarters is allowed to stand for several hours a white, yellowish-white or yellowish sediment settles to the bottom. At the same time the quantity of cream is increased and changed, appearing yellowish, tenacious, and when shaken it assumes a cloudy or wavy appearance. If the migration of the pus corpuscles from the blood vessels becomes more inten- sive the milk appears thick, yellowish, cream-like, and after stand- ing separates into a yellowish-white to ocher colored sediment, which may amount to two-thirds or more of the entire mass, and into a dark, transparent, yellowish-grey to greenish-yellow skim milk. The sediment layer is at times increased, at other times decreased. The cream becomes granular, shredded, and tenacious. If red blood corpuscles are eliminated in great numbers they col- lect in the form of a red disc on the yellow to yellowish-brown base, which is composed of leucocytes and coagulation masses. In hemorrhagic stages of the inflammation the milk is pinkish or brownish-red; by sedimentation it separates into a Bordeaux-red or rust-colored precipitate, and a pinkish-red layer of cream over the reddish-gray skim milk. In other cases the milk becomes grayish and watery, and only a few thin conglomerates and fat globules indicate the layer of cream. Cream and sediment are especially rich in cells in all forms of inflammation. Epithelial cells are desquamated into such milk in the form of colostral cells, or entire epithelial bands, and numer- ous polynuclear leucocytes, besides single epithelial cells, into which macrocytes penetrate (albuminophores), erythrocytes, cell debris, fragments of nuclei, as well as Nissen's globules are found. Besides concrements of the most varied quality, casein and Mastitis. 79 fibrinous flakes appear (Zschokke, de Bruin, Kitt, Sven Wall, Doane, Eussell and Hoffman, Ruhm, Ernst, Bahr and others). The taste of milk from affected quarters of the udder is also affected markedly, the milk becoming salty, bitter, and pungent. According to Craandijk in 67% of cases the taste of the milk changes in streptococcie mastitis. From the appearance which the affected quarter manifests, as compared with healthy quarters, from the change in the behav- iour of the animal, from the varying quantity of the secretion against the quantity from healthy quarters or the previous yield of the same quarter, the milker becomes suspicious of the existence of an abnormal condition in the suspected quarter, and the tasting test reveals a salty, bitter taste which assures him of the appear- ance of a change in the activity of the gland. If the udder secre- tion could be examined on the hollow of the hand before being milked into the pail, in order to determine the possible presence of flakes, etc., as should be the duty of the milkers, then the mix- ing of such milk from affected quarters would not occur to the extent that it does at present, as has been proved on numerous occasions. A great deal would be gained if the milk from those quarters which produce a milk so changed that its abnormalities can be recognized by its appearance or taste could be totally de- stroyed. As a matter of fact milkers can much more readily rec- ognize developing inflammations of the udder (as for instance streptococcie mastitis) from the varying conditions of the udder, or quarter, the quantity of milk, and the behavior of the animal, than the veterinarian can by a single clinical examination. Therefore the method applied in practice consisting of a single clinical examina- tion of the cows producing infant milk at the time of purchase, or every 3 to 4 weeks is not sufficient to determine the presence of ud- der affections. Periodical examinations of all cows producing certi- fied milk, supplemented by tests of the milk obtained at the time of the examination, are necessary when the inspection is to serve its purpose. At the examination in the stable a comprehensive history should be taken from the milkers relative to the general condition of the cows, their action during milking, the condition of the teats and the gland tissue, the inflammatory changes noted, in fact all points which may offer valuable supplements to physical examina- tion. One may learn from questioning that the cow milks very hard from one quarter, that she sometimes refuses to "give down" her milk, or that she "draws up" the milk or that recently the cow has shown a tendency to kick during milking. At other times one may hear that the parenchyma contains knobs or lumps or that the teats contain beads or warts, or are "fleshy," the quantity of milk is diminished, the milk is sometimes hot, "heated," or that the cow has the "cold garget" without any inflammatory indications of streptococcie infections. The milk is ropy, the 80 Effect of Internal Influences. quarter is " blind," the milk contains stringy clots and other things. The keeping of milk records and the taking of milk samples at least every four weeks, should be required of all owners of animals which produce milk for city consumption and those fur- nishing it to wholesale milk dealers. Together with the visible changes in the milk, changes of the value of the chemical and physical properties occur which have been especially studied by Guillebeau and Hess, Schaffer and Bondzynski, E. Seel, Mezger, Fuchs and Jesser and Mai and Rothenfusser. These changes in the contents and properties are therefore especially important since frequently values are obtained which suggest adulteration with water. Irreproachable comparative tests of milk obtained directly from the stable may indicate however that in the specific cases the investigations were being made with abnormal milk. .According to Schaffer and Bondzynski 's examinations the milk from cows affected with mastitis showed the following values: Water Solids. Fat In non-infectious garget 92.83 7.17 0.82 In yellow gait 89.34 10.66 1.99 In parenchymatous mastitis 90.26 9.74 2.16 In comparison with healthy milk ..87.75 12.25 3.4 Pro- teid 4.01 6.00 4.21 3.5 Milk Sugar 0.53 1.84 1.01 4.6 Total % Ash P2O4 0.79 0.83 0.97 0.75 7.35 Cl. 35.76 19.21 20.0 27.79 14-.0 The milk sugar content was also considerably diminished ; the amount of mineral substances on the other hand was increased. Guillebeau and Hess give the following values in milk from cows with affected udders: Duratio 1% d£ 2 5 7 2 2 1% ll/2 1 8 2 21 n of the Disease and Origin of the Milk lys . . . Specific Solids. Gravity 7.45 5.15 udder recovering 9.80 1.0314 11.28 7 69 23.58 from from two two affected affected quarters quarters 15.88 9.66 from from two two affected affected quarters quarters 15.88 1.0430 20.94 1.0379 18.18 Fat 0.52 0.22 1.95 2.72 1.09 9.30 4.50 2.09 0.53 4.50 0.97 2.80 Nitrog- enous subst. 6.17 4.26 2.98 3.50 5.74 8.53 5.37 6.74 5.13 5.37 16.65 7.93 Milk Sugar 0. 4.06 4.35 0. 4.68 5.14 2.09 5.14 2.61 2.04 Ash 0.85 0.82 0.81 0.70 0.87 1.07 0.87 0.85 0.87 0.71 0.91 In most cases the specific gravity is lower (Seel, Mezger, Fuchs and Jesser) and approaches the normal only towards the end of the disease. In mixed milk from all four quarters the specific gravity is less influenced. The quantity of fat, according to Seel, and in some cases of Mezger, Fuchs and Jesser, is very much reduced. The latter authors emphasize the fluctuation of the fat in sudden jumps. The experience of the official milk control station in Munich also gives similar results, at the beginning of the affection frequently finding abnormally high or again abnormally low fat contents of the milk. Mastitis. g]^ The amount of milk sugar as a rule is reduced, and rises only with the appearance of recovery. The solids are likewise usually diminished ; the fat-free substance is also and only becomes increased after signs of recovery have been noticed. The amount of proteid coagulable by heat frequently increases enormously, as compared with the contents of casein which diminishes. The ash content on the average is increased. Eelative to the composition of the ash the data appear to te contradictory. Although Seel found in 15 cases out of 18 a diminution of chlorides against an increased quantity of P2O5 Mezger, Fuchs and Jesser observed an increase of the chlorine content and a diminution of the phosphoric acid, while Steinegger and Allemann found that the amount of P2O5, CaO, KoO and MgO diminishes, in general, while the quantity of CL, No2O and SO3 increases. According to Hashimoto the ash of abnormal milk closely approaches the ash of blood serum (0.78%) consisting of 8.9863 K2O; 36.544 Na2O; 7.44 CaO: 1.738 MgO: 17.380 P2Or, and 33.627% CL The reaction of affected milk is mostly alkaline (Seel, Merger, Fuchs and Jesser, Hoyberg, Auzinger, Ernst), or more rarely acid (Zschokke, Henkel, Wyssmann and Peter, Ernst). The determination of acidity is recommended by Plant as a means for the diagnosis of udder affections. Independently from the degree of acidity, the coagulability on the addition of alcohol is frequently considerably increased but not always, and in some cases not con- stantly. (Henkel, Rullmann and Trommsdorff, Auzinger). The refractability of the calcium chloride serum not infrequently suffers con- siderable changes upward and downward. Eipper, Ertel, Mairhofer, Schnorf, Mai and Rothenfusster, Henkel, Mezger, Fuchs and Jesser found considerable changes in this respect and proved that the daily variations in the refraction may be very great in milk of individual quarters, and even in the full milk of an animal. Frequently however the refraction of the calcium chloride serum shows no change when compared with the milk of healthy animals. The same variation obtains in the lowering of the freezing point of milk from affected quarters; the values may be considerably higher than that of healthy milk, or on the other hand they may be lower. More frequently a high value is observed (Schorf, Quirand and Laserre, Crispo, Bertozzi, Pins). According to Schnorf the elec- trical conductivity is always increased, never normal or lower. • According to Bonnema the increase of chlorides results in an increase of the electrical conductivity. A change in the contents of original ferments appears very early during the affection, together with an increase of cellular elements, especially leucocytes (Zschokke, Bergey, Trommsdorff and Rullmann), and fibrinous flakes (Doane, Eussell and Hoffmann). According to Konirig the increase of the catalase content in freshly obtained milk is an indication of the affection of the ud- der, provided the colostral period has passed. The publications of Spindlers and Rullmann (who were enabled to obtain asepti- cally milked samples with which to work) and the author's obser- vations confirm Koning's findings. The author observed that in slight, local affections of a chronic nature, without febrile mani- festations, the content of catalase usually runs parallel with the cell content, and it rises when there is an especially marked throwing off of epithelia (presence of typical colostral cells). The faculty of splitting up added starch solution likewise in- creases in milk from affected udders as compared with that from healthy udders. There are no observations relative to the quanti- tative effects of peroxydase. According to Weichel the peroxy- dase content of affected milk from an artifically affected goat dis- appeared, whereas the healthy milk gave the guaiac reaction. 6 Effect of Internal Influences. The reaction again appeared when the secretion became of a milk- like consistency. Affected milk behaves in various ways on the application of f ormalin-methylene blue solution ; frequently a very rapid decol- oration of Schardinger's reagent may be observed (Rullmann, Sas- senhagen, Rievel). Sometimes in typically changed samples the reduction does not take place (author's observation). As the above-mentioned enzymes (not amylase), at least in part may appear to be brought on by bacterial action, their abnor- mal presence in milk has a diagnostic importance only in freshly milked samples. The case is different with the complement con- tent. As indicated in the chapter on antigens, blood constituents pass directly into the milk during periods of physiological and pathological irritations. Therefore in mastitis, as proved by Bauer and Sassenhagen, complements are demonstrable in the milk. This, according to Sassenhagen, is possible even in affec- tions in which the Trommsdorff value of the centrifugalized cells per 1000 parts of milk is still remarkably slight. The alkaline reaction of affected milk, the altered proportions of mineral salts, at times the passing into the milk of bloody parti- cles, and the diminution of casein, reduce the coagulability of the milk towards added rennet. Affected milk therefore generally utilizes a considerably larger quantity of rennet than normal milk before it becomes coagulated (Schern). The spontaneous coagulation of affected milk also appears to be considerably delayed. Infectious Agents of Mastitis. Nocard and Mollereau, Kitt, Lucet, Bang, Guillebeau and Hess, Zschokke, Jensen, Streit, Glage and Sven Wall have offered sufficient information regarding the infectious agents of the dif- ferent forms of mastitis. Most cases of mastitis are produced by streptococci; they consist of chronic inflammations of one or more quarters of the ud- der. The disease is of relatively small influence on the general condition of the animal. Bacilli of the coli-aerogenes-paratyphus-paracolon groups pro- duce highly acute, parenchymatous lesions. The general condition is severely influenced through infections by bacteria of the paratyphus-B group. Locally a gangrenous, septic mastitis de- velops with this infection, and the milk is markedly ichorous, while in colon infections the secretion is of a serum-like character (Weichel). A third form of inflammation of the udder, also of a chronic nature, is produced by a representative of the group of the Bacillus pyogenes, Sven Wall's pyobacillosis of the udder. The Bacillus pyogenes colonizes with a special predilection in the pres- Streptococcic Mastitis. 33 ence of streptococci or staphylococci, and in these mixed infections causes severe necrotic inflammations of the udder, and may con- tinue to produce chronic mastitis in the affected udder tissue after the disappearance of the other bacteria. Other forms of mastitis are produced by tuberculosis and actinomycosis, and they usually result through emboli of the in- fective agents. They may be of a traumatic origin (actinomycosis) induced by irritation with particles of straw, or barley beards. Furthermore all possible infective agents, as for instance the Bacillus necrophorus, may be found in inflammations of the udder, either independently or as mixed infections. Only the more important infections of the parenchyma will be described here. Streptococcic Mastitis. By far the most widely spread type is the Streptococcic mas- tisis, described by Sven Wall as streptomycosis of the udder. The works of Bergey, Craandijk, Trommsdorff and Rullmann, Kunze, Eussell and Hoffmann, Savage, Riihm, and Ernst give gen- eral information on this condition. The disease is either sporadic or epizootic among the animals of a stable according to the stable conditions. The disease may attain an especially wide distribu- tion when the secretion of the affected quarter is milked upon the floor or into the bedding, and the milkers fail to wash their hands, both bad practices which, it is to be regretted, are quite common. Zschokke, Jensen, Bang, and Sven Wall proved experi- mentally that bacteria injected into the cistern penetrate even into the farthest alveoli in from 2 to 24 hours. By inoculating with strains of streptococci of different ori- gin varying reactions may be produced in the udder (Bang: Strep- tococci equi and Streptococci agalactice; Gminder : streptococci of the stable air and of infectious vaginal catarrh). The manifesta- tions also vary after the injection of individual strains into the same animals, and from injections of the same strain into various animals. In other words the course of the disease varies in ac- cordance with the virulence of the organism, the resistance of the body, and the extent of the local invasion which again is influenced by the lactation period. According to de Bruin fresh milking animals more frequently become affected with the acute form having inflammatory manifestations, while in old milking animals the disease confines itself mostly to the altered appearance ot the secretion. The result of the disease is that sooner or later the affected part of the gland becomes destroyed. Sometimes the streptococci remain for weeks in the folds of the mucous membrane of the cistern without infecting the par- enchyma ; in other cases again the entire quarter quickly becomes affected. Unfortunately the disease does not often subside even g4 Effect of Internal Influences. through the physiologically dry period, and the affection re-ap- pears immediately after parturition. The destruction of all streptococci involves a difficult task for the entire body. The dissolution of the streptococci progresses only very slowly even in actively or passively immunized animals. Living streptococci may be demonstrated in the abdominal cavity of test animals, many hours after an intra-peritoneal injection. Not infrequently a delayed death appears in apparently recovered animals (v. Lingelsheim). The long streptococci appear to rep- resent specially adapted forms which have great tenacity. Never- theless at times recovery takes place. According to Zschokke the relation between recovered and unrecovered cases is as 7 :5. According to the experience of the author in practice, infectious mastitis is not curable, or only with the greatest difficulty, and if so, always with a loss of productiveness, which even remains after the physiologically dry period. The chronic irritation causes a change in the connective tissue structure of the parenchyma of the udder so that the usual development of the gland during pregnancy cannot take place. The principal aim in treatment therefore should be prompt drying of the suspected udder, in order to make possible the most rapid and most complete recovery, which, ac- cording to Zschokke may be expected only when the quarter has been allowed to remain dry for a long time. This is also necessary in order to prevent a spread of the disease, which is to be feared since the hands of the milkers and milking upon the straw may transmit the infective agent to other quarters. Care should be taken therefore to keep the milk from the healthy quarters of the udder separate from the secretion of the affected quarter. As long as the most primitive requirements of clean milk production on the part of the milkers are so carelessly neglected, which unfortunately has been the case up to the present, the im- mediate drying of the affected quarter offers the only means of preventing the further spread of the disease. If, however, there is assurance that the affected animal or the affected quarter, respectively, is individually milked, and the milkers follow instructions, an attempt may be made by special frequent milking (into a jar) to produce a hyperemia of the udder. With this method success can only be expected in the early stages. The extent of the spread of the disease may become obvious by the findings after examination of individual herds. In such cases it is necessary to milk each cow, or still better each quarter, separately. The results vary, depending on the technique of the examination. The lowest number of affections is obtained when only a clinical examination is made. This therefore does not suffice in order to eradicate the disease effectively, or to single out the affected animals. The data of the different authors vary relative to its occurrence. The following figures are given which were obtained by systematic examinations of entire herds. Streptocoecic Mastitis. §5 Out of 260 animals Trommsdorff found 15.6% affected, Eiilim 31.25% out of 16 animals, Eussell and Hoffmann found in 188 sam- ples 50% with " streptococci." Savage found similar values (55%). The author examined from April 1, 1907, to November, 1908, 1697 samples of milk from individual cows, and found in 348 sam- ples the typical signs of streptococcic mastitis. In 1908 and in the following years he has demonstrated : 1908. No. of animals, 1695. Streptococcic cases 353 1909. No. of animals, 738. Streptococcic cases 301 1910. No. of animals, 597. Streptococcic cases 203 1911. No. of animals, 876. Streptococcic cases 279 Therefore 20.6; 20.9; 40.6; 34 and 31.8%, respectively, of the animals were found to be affected with streptococcic infections of the udder. If the milk of the individual quarters of the affected udder is examined various stages of the affection in the different parts of the udder may frequently be found. Out of 528 quarters of animals with affected udders 276, or 52.2%, showed lesions in individual quarters. 39.2% of the cows had the disease in one quar- ter, 25.9% showed it in two quarters, 18.5% in three quarters, and 16.2% in all four quarters. According to Zschokke out of 662 affected quarters 193 occurred in one quarter of the animal, 211 in two, 109 in three, and 149 in all four quarters. The contamination of market milk with the secretions from animals with udder affections is relatively high. In spite of the fact that proof of the mixing of milk from affected udders with market milk is possible only in very pro- nounced cases (typical streptococcic chains with characteristics of animal origin), nevertheless the following results, obtained in examinations, demonstrated conclusively that the secretion from quarters affected with streptococcic mastitis had been added to the whole milk: 1908, in 352 out of 1578 samples=22% 1909, in 501 out of 1629 samples=40.5% 1910, in 243 out of 1211 samples=20% 1911, in 432 out of 1273 samples=33.9% The hygienic importance of the affection to the consumers of milk may be illustrated from the following data. I. Hoist, in 1894, had the opportunity of examining in Chris- tiania four series of affections of acute gastro-intestinal catarrh. I. Four grown persons and four children out of three families in the same street became affected four hours after the drinking of milk which originated upon one farm. Those persons who drank no milk or only that which had been boiled were spared with the exception of a child who became affected, although only slightly, after drinking boiled milk. gg Effect of Internal Influences. The appearance of the milk showed nothing abnormal, but it coagulated on boiling and showed a tremendous number of bacteria, especially streptococci, which could not be distinguished from the Streptococcus pyogenes. The veterinary examination confirmed the suspicion that a pus-containing secre- tion was being yielded by one cow. The milk from the cow with mastitis on the day in question was added to the whole milk through the neglect of a newly hired attendant. 2. Several hours after the drinking of raw milk five persons, and as found later other cases also became affected with acute gastro-intestinal catarrh. In this case a milk dealer was implicated, and it was found on inquiry that the milk con- tained secretion from a cow affected with streptococcic mastitis. 3. According to the observations of Johannesen two persons (mother and child) became sick after the drinking of milk. The milk was thin, flaky, and contained pus-like lumps. In the herd from which this milk originated two cows were found to be affected with streptococeic mastitis. 4. After the drinking of freshly drawn raw milk four children of the same family became affected with acute gastro-intestinal catarrh. The milk appeared ap- parently normal, but contained large quantities of streptococci. It originated in a stable from which on the day in question a cow was sold on account of mastitis. The milk from this cow appeared to have been mixed with the whole milk due to the neglect of a new milker (the regular one being sick). The affections which occurred in Stockholm with symptoms of fever, dullness, attacks of fainting, nausea, vomiting, diarrhea and cramps in the calf of the leg (nine families being involved), cannot according to the obtainable reports, be de- clared to be streptococcic infections. The milk, through the drinking of which the cases could be traced, originated from a dairy of 14 cows, among which one cow had mas- titis. It is possible that in this case an infection with bacteria of the paratyphus group, which plays an important part in the development of acute mastitis, was con- cerned. Further contributions to the casuistics of "milk- poisoning" were published by Jakobsen and Weigmann and Gruber. II. In 1905, Jakobsen observed symptoms in several persons which he traced to the drinking of milk from one stable. The symptoms were diarrhea, vomiting and fever. Out of 17 persons, 10 who drank the milk became affected, while 7 who did not take any remained well. On May 30, 1905, other persons became affected. The examination of the 32 cows of the dairy showed a streptococcic mastitis in one animal. The cow was slaughtered and no further cases were reported. III. Edwards and Severn described an epidemic of follicular tonsilitis which de- veloped from the drinking of milk. They found in the exudate of the throat, and in the milk, in addition to other bacteria pyogenic streptococci which as shown by the in- vestigation were also contained in the secretion of a cow affected with mastitis. [In various cities of the United States epidemics of sore throat with swelling of the cervical lymph' glands, colic, diarrhea and fever lasting several days have occurred which were traced to the use of milk from cows affected with streptococcic mastitis. Such milk when examined was found to contain pus and streptococci in great abundance. — Trans.] IV. Lameris and Harreveld observed an outbreak of diarrhea among the inmates of a hospital after the drinking of boiled milk, which in part was obtained from cows affected from streptococcic mastitis. Whether the authors of the last cases are correct in their view that very likely a heat-resisting toxin brought on the disease, or whether the streptococci might remain alive in the milk foam or in the formed membrane, etc., as suspected by Jensen, it becomes evident that boiling does not carry with it an assurance that the danger from streptococcic milk is eliminated. If Jensen's sus- picion is correct a proof would be offered that even the smallest Table II. Ernst, Milk Hygiene. Streptococcic Mastitis. 87 quantities of mastitis streptococci are sufficient for the production of severe intestinal affections. V. On December 17, 1907, a sample of boiled milk was brought to the official milk control station of Munich; about a half hour after the drinking of this milk the man who delivered the milk, and his family, as well as a neighboring family using milk of the same origin, became sick. The milk contained 1.5:1000 streptococcic pus. It orig- inated from a large dairy. Three producers and one distributor were suspected. In tracing down the cause of the trouble two producers were found whose herds contained six animals with affected udders, their milk being mixed with the whole milk. The affection was marked by chills, diarrhea, headaches, and lasted not quite an entire day. The milk constituted the only common food partaken by all, and there- fore could be considered, although not with absolute certainty, as the probable cause. From the examples cited it may be seen that the drinking of milk which contains the secretion of streptococcic infected udders is capable, under certain conditions, of producing injurious effects upon the health of human beings. Considering the fre- quency of the disease, and the numerous cases where the prohi- bition of milking affected udders into the whole milk is disre- garded, it is to be wondered at that affections which could be traced to the drinking of such milk are not observed with greater frequency. This may be due to the fact that the secretion of affected 'quarters is usually very greatly diluted with the milk of healthy quarters, showing that the harmful actions are not necessarily induced by the predomination of the injurious material, and fur- ther it may also be due to the fact that the milk is mostly used after being boiled (Trommsdorff, Jensen). That the boiling of the milk is not always sufficient to destroy the injurious properties may be seen from the cases of Hoist and Lameris and van Har- reveld; the milk of course is marketed in a raw state, and must therefore be judged in the condition in which it is sold. The factors which induce the harmfulness of the milk from streptococcic animals are not known. Whether the injurious factors are due to the toxin produced by the streptococci of mastitis, or to the products of the disease, as for instance pus (Jensen), or to streptococci which are pathogenic to man as such, cannot at the present time be definitely determined. This, how- ever, is of little importance from a practical standpoint. Some authors, such as Petruschky and Kriebel, consider that infected cows are the sources of milk streptococci, and that these are the principal cause of the summer mortality of children. Seiffert considers the streptococci originating from affected udders as more dangerous than the saprophytic streptococci which contam- inate the milk as a result of unclean milking. This view was also expressed by the author in May, 1908, and was confirmed by Trommsdorff. Neither studies nor animal experiments have succeeded up to the present time in proving the harmfulness of the streptococci by themselves, or the relationship of the mastitis streptococci to human pathogenic strains of streptococci, the animal experiments gg Effect of Internal Influences. offering only a relative conclusion on the susceptibilityof the respective species of animals. The differentiation of the mastitis streptococci fron other milk streptococci is however, absolutely necessary for milk3ontrol since only in the presence of typical mastitis streptococci -an the milk dealers be held responsible, and be obliged to prevnt the contamination. Escherich and Hoist found streptococci in almost every sample of tilk, and Hellens repeatedly isolated them from milk. In 1840 in samples of market tilk from Munich and vicinity the author succeeded in isolating streptococci either hy culti- vation or recognizing them by bacteriological examination in 100% of the ca». Other investigators confirmed a positive finding in a strikingly high percentage. Beck— Market milk of Berlin 62 % Savage — 17 samples of market milk 100% 10 samples of market milk 100 % Kaiser — Market milk of Grar 76.% Briining — 28 samples of Leipsic market milk 93% Easten — 186 samples 57 Eastles — 185 samples from all parts of England 75 % The simplest proof of the constant occurrence of stre]tococci in market milk is the usual acid fermentation of cow milk dduced by streptococci. A method for distinguishing these frequently observer strep- tococci from mastitis streptococci has not yet been disoveredr either through the fermentation of various kinds of sugarsby the streptococci, or through the investigations of creatinin forma- tions, hemolytic action, acid formation or their actions in th pres- ence of various temperatures of cultivation. It should bi borne in mind that the behavior of the various strains of mastiti strep- tococci has been described in such a variety of ways, that eiter the presence of remarkably numerous strains or a strong installity of characteristics, or confusion with saprophytic forms, rast be accepted. The formation of acid by the streptococci is sometiies de- scribed as strong (Zschokke, Nenski, Groning, Kaiser, Heinmann, Miiller, Koning), at other times it appears insignificant (Sven Wall,Kullmaim). Lohnic classes the streptococci of mastitis with the gamp of lactic acid streptococci, especially with the group of Strefococci gimtheri, with close relationship to the group of Strefococci rosenbach, having the following characteristics : "Form of the cells variable; capsule formation is frequent and appears o be as- sociated in certain forms with the presence of sugar in the nutritive media, bores are not formed; the bacteria are Gram-positive; the intensity of the growth hi; no sig- nificance. Coagulation of the milk results, in these varieties, either thrcgh acid formation or through a rennet-like ferment; gas formation is rare; the patigenicity varies remarkably." Miiller in his work on comparative examinations of lactic acid bacteria fyp. giin- theri, etc.) presents the following: 1. "The strains studied manifest marked differences either in their cbtural or morphological characteristics with the exception of the strain causing "SOT ; brood'1 among honey bees. ' ' 2. ' ' The action on carbohydrates is practically uniform. ' ' Streptococcic Mastitis. 89 Fig. 17. 3. ' Influencing individual strains relative to their acid formation in the sense of increasing decreasing it, is possible. The characteristics which the freshly isolated strains jxsess are more or less permanent." 4. • There exists a relation between the group of Streptococcus giintheri and the Streptocuws agalactiae since their capability of forming acid is about the same. ' ' 5. ' The oft recurring confusion of the two may be explained by certain similar forms o%rowth which both possess. ' ' 6. ^The supposition that the pathogenic streptococci represents lactic acid bac- teria of to Typ. giintheri which have adapted themselves to parasitic conditions, is sub- stantiatecby the findings, since it was possible in the various strains of streptococci to produce tinsition forms, which correspond to the Typ. giintheri." Threfore from these few examples it may be seen that it is impos^le to separate the streptococcus of infectious mastitis from the grap of the lactic acid streptococci. Nevertheless it would be a greaterror to identify the orihary lactic acid streptcocci with patho- genic siteptococci of man and aniials. If he fact is taken into consideration that some steptococci, as for instancTi that the milk contains the secretion of an animal af- C1> W11 fected with streptococcic mastitis in spite of the oc- currence of other forms of streptococci (d and e). is sometimes more, at other times less pro- nounced. This sometimes swells to a broad mucin capsule (com- pare Lingelsheim on streptococci, Wassermann-Kolle's Hand- book of pathogenic micro-organisms III, pp. 309 and 310, and Sven Wall, p. 29). The endococcus, especially in short chains is spherical or swollen to a club shape. With slight practice one almost invariably succeeds in dis- tinguishing, by one or the other given characteristics, the "animal" mastitis streptococci from streptococci which have gained access to the milk accidentally (even though they may also possibly be de- scended from ' ' animal ' ' mastitis cocci) . In this way the author succeeded, from April, 1907, to Novem- ber, 1908, in demonstrating by the aid of smears that secretions from cows with streptococcic mastitis wrere mixed with market Streptococcic Mastitis. 91 milk. Out of 1840 microscopically examined samples 336, or 18. 26% showed the presence of such an infection. In 91 cases, or 4.945% the changes were not very pronounced; later control however proved that milk from cows affected with yellow garget had been mixed with these shipments. 18 . 26 % + 4 . 945 % = 23 . 205 % , proved contaminated with streptococcic pus out of 1840 milk samples. Miiller intended in his work to distinguish milk streptococci, especially the streptococci of mastitis, from strains of streptococci pathogenic to man. In confirmation of the work of Nieber, Fischer and Berger, Miiller came to the conclusion that the Fis- 19- recognition of milk strep- tococci pathogenic to man is impossible. Although milk streptococci as a rule coagulate milk some- what more quickly, there are also strains which coagulate milk somewhat more slowly, and strains which dissolve the blood cells in Schottmiiller's blood agar, and these in. their agglutination value stand very close to the pathogenic streptococci of man, that is, they ag- glutinate even in dilu- tions of the serum of 1:400—800. At the same time several of the abso- lutely pathogenic strains fail to give any agglu- tination, and other ap- parently saprophytic va- rieties give a higher ag- glutination value. Bau- mann proved that there is no uniform agglutination value of the individual kinds of streptococci, and that spontaneous agglutina- tion frequently appears in tests of their cultures. Together with Horauf, the author found that mastitis strains show similar characteristics on Schottmiiller's blood agar to the less pathogenic strains of man, a fact which has recently been con- firmed by Grminder. Lingelsheim makes the statement that strepto- cocci producing toxins are always obtained from subacute and chronic processes. Acid formation and milk coagulation are common to the entire SS& *<* Sediment of red milk. Many red blood corpuscles, several polynuclear leucocytes and colostral cells. Streptococcus brevis with capsules. 1 X 1000. 92 Effect of Internal Influences. group of pathogenic streptococci. Of the pyogenic strains of man, according to Andrewes, the Streptococcus pyo genes and the Strep- tococcus mitis produce acidity without coagulation. Sven Wall proved these characteristics from the mastitis strains isolated by him. According to Adametz, the mastitis cocci sometimes coagulate very intensely, the same as is the case with the streptococci of enteritis of sucklings (Petruschky). The fermentation in various sugars using Gordon's bouillon mixtures, varies greatly with the different pathogenic streptococci and milk streptococci, so that the possibility of differentiation by this means is quite impossible, which is likewise the case by testing their virulence on small test animals. Pathogenic strains may at times show great variations of viru- lence, while according to Heinemann strains of the Streptococcus lacticus may become virulent by passage through rabbits, until they will produce changes in rabbits which correspond in their ap- pearance, extension and character, to those caused by pyogenic strains re- covered from man. Through their action on animal bile, or on sodium taurocholate, Mandel- baum differentiates the Streptococcus mucosus and Pneumococcus from Streptococcus pyogenes and other streptococci (Neufeld, v. Levy). The author used the mastitis strains he had on hand on cattle, hog, horse and chicken bile, but failed to observe either a clearing of the culture media or an influence upon the form and appearance as shown by the microscope. The establishment of the "virulence number" through phag- ocytic experiments also fails to yield the desired result. In short up to the present time the absolute separation of culture strains of varied origin, the differentiation of saprophytic streptoccoci from mastitis streptococci, and these from pathogenic streptococci of man has not been successfully accomplished. We have, however, in certain morphologic indications, for instance Sediment of milk from an udder with acute inflammation. Short forms of streptococci. 1 X 1000. Streptococeic Mastitis. the cross-position of the segments, the capsule-like covering ancf other characters, a way of distinguishing streptococci originating in the udder of an animal from such as have subsequently gotten into the milk. If these distinguishing signs are present then smears from the sediment of market milk permit the deduction that secre- tion from an affected udder has been included in the milk. If, on the other hand, these signs are not present in the streptococci of the milk, it cannot be asserted that the milk is not contaminated with the secretion of affected udders. Although it is not known what factors Fis- 21. (streptococci, toxins, in- flammatory products) convey the unwholesome- ness to the milk, and al- though in spite of the fre- quent occurrence of mas- titis injurious effects re- sult with relative rarity, nevertheless the secretion from udders affected with streptococcic mastitis, and mixed milk which is contaminated with such secretion should be con- sidered capable of im- pairing the human health, since 1. There are known cases in which severe dis- turbances of health re- sulted from the ingestion of such milk. 2. The investiga- tions of streptococci scientifically justify the suspicion of harm arising from their ingestion. What are the conditions of the mastitis streptococci among themselves? Formerly a Streptococcus brevis and a Streptococcus longus were distinguished, but it was shown that these distinguish- ing features were not absolute (Staeheli). The differences of the in- dividual strains and the forms of their growth in culture are as in- constant as their pathogenicity, acid formation, and other biological characteristics, so that, as expressed by Kitt, it would be necessary to distinguish as many varieties as there are mastitis cases if it was desired to accept the differences of the individual mastitis strains as indicative of different varieties. All the smaller and greater differences should be considered as indications of adapta- Streptococcic pus from milk of a cow with streptococcic mastitis. Streptococcus longus. 1 X 1000. Effect of Internal Influences. tion to the various energies of reaction of the various animals and organs, and as the investigations of the author showed, to energy reaction of the same milk gland at different times. For instance it appears that certain changes in form bear a definite relation to the number of leucocytes in the milk. Thus the author obtained the following results in the same quarter of a cow examined at dif- ferent times : L. — Leucocytes. Str. — Streptococci. Dec. 17 , 1908. Jan. 4, 1909. Cow No. 29 L. Str. L. Str. 1 right fore 0.3 0 .1 0 2 left fore 0.2 0 0.9 Dipl. 3 right hind 500.0 longus 2.0 brevis 4 left hind 500.0 longus 20.0 longus Cow No. 34 Dec. : L7, 1908. Jan. 5, 1900. 1 right fore 0.1 0 0.2 0 2 left fore 0.2 Dipl. Drops 3 right hind 0.4 Dipl. 0.3 Dipl. 4 left hind 0.3 Dipl. 0.5 brevis Dec. 18, 1908. Cow No. 31 L. Str. 1 right fore 0.5 Dipl. 2 left fore 1.5 0 3 right hind 1.2 Dipl. 4 left hind 1.3 brevis Cow No. 33 1 right" fore 2 0 brevis 2 left fore 0.5 0 3 right hind 1.3 brevis 4 left hind 0.9 brevis Cow No. 58 Dec. : 19, 1908. 1 right fore 0.1 0 2 left fore 0.4 brevis 3 right hind 0.3 0 4 left hind 0.1 0 Jan. 20, 1909. L. Str. 0.5 0 0.4 brevis 1.9 brevis Jan. 25, 1909. L. Str 0.2 0 2.0 brevis 3.0 brevis 0.5 brevis 10.0 longus Jan 0.1 0.3 3.0 4.0 . 16, 1909 Jan. 29. 1909. 0 0.2 0 0 0.5 0 longus 50.9 brevis longus 60.0 brevis Jan. 4, 1909. L. Str. 0.3 brevis 200.0 medium sized 1.8 brevis 0.9 Dipl. 0.3 Dipl. 0.2 0 20.0 longus 10.0 longus Jan. 5, 1909. 0.5 Dipl. 0.3 Dipl. 0.1 0 0.1 0 These differences in the forms of streptococci may be seen dur- ing one milking on the same animal, if they are compared at the beginning, the middle and at the conclusion of the milking. These differences are only slight, so that no definite conclusions should be drawn from them. If however the results from various animals are compared it may be seen, as already indicated above, that certain relations exist between the number of leucocytes and the forms of the streptococci since the streptococci become longer as the number of leucocytes increases. The experiments extended from December 11, 1908, to February 8, 1909, and include three stables with a total of 149 cows. Of these 149 cows 59, or 39.6% were more or less affected. In most animals (140), all quarters were separately examined and Streptococcie Mastitis. 95 showed that out of 560 quarters 112, or 20% were affected. Forty- two of these gave at times a greater, at other times a smaller secre- tion with a distinctly changed consistency. The other 70 manifested the infection only after sedimentation, or only through microscopi- cal examination. Some of these 12 quarters were successively (see above) examined, so that the material used for smears from af- fected quarters, and which had been microscopically examined, amounted during the period mentioned, to 134. Fifty-five affected quarters showed the Streptococcus brevis, 32 the Streptococcus longus; in 47 the infective agent was recog- nized in the form of a diplococcus. The 47 quarters with diplococci had as a rule a very small leucocytic number. In values of over 2.0, longer coccus-chains were always observed. 32 quarters out of the 47 had 0 . 5 :1000 leucocytes 12 quarters out of the 47 had 1.0:1000 leucocytes 3 quarters out of the 47 had 2 . 0 :1000 leucocytes In the 55 cases with Streptococcus brevis the change in the leucocytic number varied to a greater extent. In 18 it represented 0.5:1000 or 32.73%. In 14 it represented 1.0:1000 or 25.47%. In 8 it represented 2.0:1000 or 14.55%. In 3 it represented over 2.0:1000 or 5.45%. In 12 it represented 5-20 and more :1000 or 21.82%. The 32 longus-cases were divided as follows : Leucocytic Quantity. Number of Cases. Percentage. Under 0.5:1000 1 3.125 up to 1.0:1000 1 3.125 up to 2.0:1000 3 9.375 up to 5.0:1000 3 9.375 up to 20.0:1000 3 9.375 up to 100 and more:1000 21 65.625 In other words : In leucocytic quantities Up to 0.5, 63.00% showed 35.0% brevis, 2.00% Dipl. longus Up to 1.0, 44.40% showed 51.9% brevis, 3.70% Dipl. longus Up to 2.0, 17.65% showed 64.7% brevis, 17.64% Dipl. longug Up to 5.0, 0.00% showed 50.0% brevis, 50.00% Dipl. longus Up to 20.0, and more 0.00% showed 33.0% brevis, 66.60% Dipl. longus The leucocytic values will be taken up again later in the dis- cussion of the "Trommsdorff" test. From this tabulation it may be seen that the length of the chains actually grows with the increase of leucocytes, or with the amount of sediment. In high leucocytic values and short forms of the infective agents, the latter are frequently present in exceed- ingly large qantities. *The opinion that the form of the streptococcus represents an adaptation to the energy reaction of the respective animal and or- gan is thereby substantiated especially when the streptococci are 96 Effect of Internal Influences. found in the secretion of one and the same part of the udder of a cow at different times. In the same way it is impossible to establish rules for definite differentiation of the streptococci of the yellow garget among them- selves through the study of their morphological relations, by com- parison of their biochemic characteristics or the pathogenic viru- lence of individual strains, since the acid production and acid susceptibility which are present in mastitis streptococci at first may be easily changed by artificial means, and individual strains have proven the possession of stronger, others a weaker pathogenic action for test animals (Groning, Sven Wall). The author does not desire by any means to establish a theory of unity for mastitis streptococci. To be sure there are marked differences in the various strains, especially in regard to the pro- duction of clinical symptoms, which cannot be attributed alone to the variation of virulence, and to unequal resisting powers. It is possible that with the aid of newer methods of differ- entiation (blood media, etc.) it will be possible to establish a fun- damental type of mastitis streptococci in strains freshly cultivated from animals. Even if with the continuance of cultivation new characteristics, as for instance hemolytic properties, may be acquired by the cultures, and the earlier characteristics become lost, the characteristics acquired by the respective strains of streptococci in their former growth may remain constant for a sufficient length of time to permit the establishment of the type of varieties, as has already been the case with the streptococci of man (Petruschky, Schottmiiller, Baumann, Schulze and others). Enrst, Gminder and others have demonstrated that the mastitis streptococci mostly correspond to the mitior seu viridans or mucosus horn, group, respectively. Based on the grounds previously described, milk hygienists, bacteriologists, children specialists and veterinarians sometimes more and at other times less imperatively have demanded the ex- clusion of cows with streptococcic mastitis from the production of milk (Jensen, Weigmann, Blevel, Sven Wall, Ruhm, Trommsdorff, Seiffert, Ernst and others). This requirement is natural from the hygienic standpoint, but its practical execution is rendered very difficult by the remarkable prevalence of the disease, and as a matter of fact as long as the general control of production and the examination of milk of indivi- dual cows are not required a thorough enforcement cannot be hoped for. For the present the exclusion from the market of all milk which shows changes in a recognizable way, as for instance through a collection of yellow sediment, should be considered satisfactory. At the same time from an economic standpoint only milk from affected quarters should be excluded, while the sale of milk from healthy quarters should be allowed. Udder Tuberculosis. 97 TUBERCULOSIS. Occurrence of Tuberculosis in Cattle. Tuberculosis of the udder in cows appears with relative fre- quency, corresponding to the frequent occurrence of tuberculosis in cattle. Tuberculosis in the udder is manifested in different forms, the circumscribed, lobular, focal tuberculosis and the diffuse tuber- Fit?. 22. Tuberculosis of the udder. Progressive miliary form. culosis extending over and infiltrating the entire udder. Two of the different forms may be present at the same time in the organ, and from one form through accentuation of the infection the other forms may result. Clinically tuberculosis is manifested by nodular swelling or hard enlargement of the affected quarter, and with enlargement and painless nodular swelling of the supramammary lymph glands. These changes, however develop very slowly and the milk from such infected quarters for weeks and months may contain millions of 98 Tuberculosis. tubercle bacilli without the udder indicating any special lesions, and without the milk showing any noteworthy changes. The quantity of the secretion from a tuberculous quarter is at first uninfluenced, or only slightly so ; later it is considerably dimin- ished. For a long time* it is of a normal appearance ; later it generally becomes thick, transparent, watery, intermixed with small flakes, or again it may become thick, yellow, pus-like, depending on the intensity and extension of the lesions in the udder and the in- fluence of the general health of the cow. Tuberculosis of the udder as a rule is of an embolic character, and rarely represents the result of a galactiferous infection. No- card, Meyer, Calmette and Guerin, and Zwick succeeded in pro- ducing tuberculosis of the udder of various characters, by injections of bovine and human tubercle bacilli into the milk duct of the teats. The pathologico-anatomical appearance in these instances was the same. The extent of the spread of tuberculosis in cattle in general is best indicated by the statistics of abattoirs in the different states. In Germany in 1904, 17.89%, in 1905, 19.16%, in 1906, 20.66%, and in 1907, 21.21% of cattle were retained on account of tuber- culosis, and 0.26%, 0.30%, and 0.35%, respectively, on account of tuberculosis of the udder. In Bavaria in 1898, 5.7% ; 1900, 6.0% ; 1902, 6.8%; 1904, 9.2%; 1906, 10.31%, were tuberculous. The spread of tuberculosis is especially assisted by exposure in stabling (in 1907, 7.28% of young stock, 18.54% bulls, 22.55% steers, and 29.62% of cows were found to be tuberculous on slaughter, against 5.3%, 13.9%, 18.3% and 25.3%, respectively, in 1904), and forced feeding while the percentage in pasture ani- mals diminishes considerably, and in range cattle tuberculosis is practically unknown. In Prussia the total infection from 1898 to 1906 is estimated at 16.09 to 23.4%, in Saxony from 30.46 to 37.58%, from 1898 to 1908. In France in certain localities the infection ex- ists in 30 to 40% of the stock, an average of 10%. In other countries similar conditions obtain. If the abattoir findings are not considered as indicative of real conditions, and delicate biological methods are employed which prove that an animal is infected with the tubercle bacillus (with- out however manifesting anatomically demonstrable changes) the increase of the numbers is considerable. In Saxony from 1891 to 1897, in round numbers two-thirds of the cattle were found to be infected through the aid of tuberculin. Ostertag accepts 25% of the cattle as infected in northern Germany and around Stuttgart, basing his estimate on sample testing in the various localities. In France from 50-80% of the animals reacted, in Great Britain 27%, in Austria 14-60%, in Hungary up to 35 . 18%, Belguim 48.8%, Norway 22.8%, Sweden about 31%, Finland 13.7%. In Denmark during the first years of the tuberculosis eradication 38.5 to 40% of the animals gave tuberculin reactions, while later only 8.5% reacted. Frequency of Tuberculosis. 99 [The extent of tuberculous infection among cattle in the United States is indicated by the statistical reports of the Federal Bureau of Animal Industry. According to these figures out of 400,000 cattle tested, there were 37,000 reactors or 9.25%. The majority of these animals were dairy cattle, from which fact the conclusion has been drawn that approximately 10% of the dairy cattle in this country are affected with tuberculosis. On the other hand the meat inspection statistics show that 6,978,293 cattle were slaughtered in official establishments during 1913, of which 75,870 were found tuberculous, suggesting that probably 1% of beef cattle are affected with tuberculosis to some degree. — Trans.] The relative frequency of tuberculosis of the udder corres- ponds to the numerous occurrences of bovine tuberculosis. According to Bergmann 3.5% of the tuberculous cows slaugh- tered at Malmo were also affected with tuberculosis of the udder. Ostertag estimates the appearance of tuberculosis of the udder in 0.1 to 0.3% for Germany. These figures of course increase in lo- calities in which the other forms of tuberculosis occur with greater frequency. Thus Meyer-Stendal report that out of 818 cows 4% showed udder tuberculosis. Meissner found in 1910 from all cows examined in Posen 0.32% affected with udder tuberculosis. Bugge mentions out of 16,050 cows included in the eradication work at Schleswig-Holstein in 1906, the presence of tubercle bacilli in 30 out of 318 examinations of mixed milk, and in 27 out of 562 individual milk tests of suspected cows. In 1907 the tests for tuberculosis of 258 samples of mixed milk, revealed tubercle bacilli in 35 while out of 597 individual milk samples, tubercle bacilli were found in 32, which corresponds to 0.14% of open udder tuberculosis in the 23,278 examined animals (pulmonary tuberculosis 5.1%). In 1908 these numbers amounted to 0.124 to 2.644% respec- tively out of 38,454 animals. In the presence of such an extension it should be not at all surprising that market milk contains tubercle bacilli with extraor- dinary frequency. Examinations for this purpose were under- taken as early as 1893 by Montefusko in Naples, in 1894 by F'riis in Copenhagen, in 1895 by Obermuller and Fiorentini, in 1898 by Petri, 1900 by Beck in Berlin, in 1895 by Zacharbeko in Petersburg, 1897 by Massone in Genoa, Buege in Halle, Boyce and Delepine in Liverpool, Klein in London, Nonewitsch in Wilna, Stepanow in Kasan, Bujwid in Krakau, and in 1905 to 1906 by Eber in Leipzig, with varying results. The numbers of tubercular infection of market milk obtained (other investigators for instance Brusaferro, Roth, Schuchardt, Groning, Petri, Eabinowitsch, Hermann and Morgenroth, Ascher, Coggi, Bonhoff, Herbert, Markl, Herr and Beninde, Eber and others experimented with butter and cheese) fluctuated between 0 and 100%. 100 Tuberculosis. It is to be expected that all market milk, no matter of what origin, may occasionally be infected with tubercle bacilli ; all milk in the production of which no special care is taken in the selection of the milking animals and no clinical examination or tuberculinization of the animals has been undertaken, should be suspected of contain- ing tubercle bacilli, and the larger the herd which produces the milk, the greater the danger. German investigators established the following figures for the presence of tubercle bacilli in market milk : Author. Samples. Place. Tubercular %. Obermuller 13 Berlin 61 Buege 6 Halle 33 . 3 Petri 64 Berlin 14 Beck 56 Berlin 30.3 Proskauer 9 Market milk Berlin 55.5 Seeligmann 5 Dairies under veterinary control . . 0. Croner 13 Danish milk .' 38.5 Eber 210 Leipzig 10.5 [The percentage of tubercle bacilli found in the milk supply of large cities in this country has been accurately determined in only a few instances. In 1907 Anderson proved that in Washing- ton, D. C., 10.7% of the dairies supplied milk containing virulent tubercle bacilli, Schroeder found 7.7% of the 26 dairies examined were distributing infected milk to Washington, D. C., while still later Mohler showed that about 3% of the 73 samples of milk ex- amined contained tubercle bacilli. The apparent discrepancy in these results may be readily explained by the fact that during the last 6 years strenuous efforts have been carried on by the Bureau of Animal Industry to eradicate tuberculosis from among these herds, with the result that in the District of Columbia the number of tuberculous animals has been reduced from 18.8% to 1.2% in 1914. Hess has found that 17, or 16%, of 107 samples of milk dis- tributed in New York City contained virulent tubercle bacilli, while Campbell made extensive investigations of the occurrence of tubercle bacilli in the market milk of Philadelphia, and found 18 or 13 . 8% of the 130 samples examined to contain living bacilli of tuberculosis. — Trans.] Under What Conditions Do Tubercle Bacilli Enter the Milk? The infection with tubercle bacilli is natural when the animal is affected with tuberculosis of the udder, or may occur indirectly when through contamination of the udder with feces in pulmonary or intestinal tuberculosis, urine or vaginal secretion in kidney or uterine tuberculosis, or with infected straw, tubercle bacilli are brushed off from the soiled udder into the drawn milk, or when in Infectiousness of ?l\tilkV-» ' i *' \, >''/> I '••] \ / 101 open tuberculosis of the lungs the bacteria get into the milk through the air or straw. As early as 1869, prior to the discovery of the tubercle bacil- lus by Koch in 1882, Gerlach proved the infectiousness of milk from highly tuberculous animals through feeding and inoculation experiments. This was also emphasized by Ziirn, Klebs, Sommer, and in 1880 by Bellinger, who first pointed to the fact that the milk of a tuberculous cow in which the udder is not noticeably affected may contain tubercle bacilli. The same results are shown by the works of Stein, Bang, Hirschberger, Ernst, Schroeder and Fiorenti, who succeeded in producing tuberculosis in test animals with milk of tuberculous cattle, even though there was no udder tuberculosis present. Milk from tuberculous udders always has been proved to be especially dangerous (May, Bang). If special care were exercised in milking, it not infrequently happens even in extensive, generalized tuberculosis that the in- oculated animals remain well ; thus Nocard from injecting milk of 54 cows affected with generalized tuberculosis succeeded only in 3 cases in producing inoculation tuberculosis. Therefore it ap- pears that in spite of a generalized tuberculosis when udder tuber- culosis is not present, tubercle bacilli are not always excreted with the milk ; the possibility of elimination however, that is, that the milk of such a tuberculous cow may contain tubercle bacilli, should at all times be given consideration. Is milk liable to be affected when tuberculosis cannot be clin- ically demonstrated in suspected cows, or wrhen they appear healthy and yet react to tuberculin? Are tubercle bacilli eliminated only with the milk from animals affected with tuberculosis of the udder, or also in cases where the udder is not affected by tuberculosis? These questions may be answered at the present time with great certainty, namely, that tubercle bacilli of cattle are eliminated with the milk as a rule only in animals which are affected with tubercular mastitis. This question is of especial interest in the eradication of tuberculosis in the dairy herd, since it is well known that calves and hogs are highly susceptible to the tubercle bacillus of cattle. It will be advisable therefore to refer to the more important publica- tions on the elimination of tubercle bacilli with the milk, before entering into the question of the harmfulness of such for man. Delepine, Ravenel, Rabinowitsch and Kempner, Gehrmann, Gehrmann and Evans, Moussu and Mohler obtained positive results from milk of animals not clini- cally affected, but reacting to the tuberculin test, through inoculating or feeding of test animals. Other investigators, as Martel and Guerin, also Hirschberger, aimed to solve the question by inoculating the milk of slaughtered animals or glandular substance from udders of reacting animals. Their work also frequently gave positive results. All of these authors therefore, conclude with great certainty that tubercle "bacilli may "be eliminated with the milk even from animals which are not clinically affected with tu- berculosis. Other views are supported by Archer, Muller, Ostertag, Stenstrom, McWeeney, Pusch and Hessler as the result of their negative findings, namely, if the experiments were conducted under the most painstaking requirements and all contaminations through 102 ' ' ' tuberculosis. infected straw, etc., were avoided as much as possible, they failed in spite of numerous experiments in producing tuberculosis with milk from a tuberculin reacting animal, and frequently not even with the milk from an animal clinically affected, but free from tuberculosis of the udder. From the standpoint of milk hygiene the fact is important that in the work of tuberculosis eradication by the agricultural societies the examinations for tubercle bacilli in the mixed milk of individ- ual herds were mostly negative after the clinically affected tuber- cular animals had been eliminated. According to the works of Muller and Hessler until July 1907, 2,949 samples of mixed milk of individual herds were examined; all of these herds were sub- jected to the Siedamgrotzky-Ostertag method of eradication. From 30 to 200 cows participated in each test and 156 herds gave milk free of tubercle bacilli. As eliminators of tubercle bacilli were found : Two cows with udder tuberculosis, 8 times each. One or more COWTS with tuberculosis of the uterus, 16 times. One or more cows with tuberculosis of the uterus, 6 times. Once kidney and uterine tuberculosis. Once pulmonary and intestinal tuberculosis. Once a cow whose saliva contained tubercle bacilli and 19 times open pulmonary tuberculosis. In five positive tests there was no clinically demonstrable form of tuberculosis, and the subsequent tests of immediately drawn con- trol samples remained negative. These five cases wrere observed in the beginning of the eradication work. The remaining 2,793 milk samples were free of tuberculosis in spite of the fact that among the animals of these herds there were surely a great number which would have positively reacted to tuberculin tests. The five cafes observed at the beginning of the eradication work, in which the milk contained tubercle bacilli, although clinically open cases of tuberculosis could not be found on stable examination, are explained by Hessler in that the milk became contami- nated with particles of the feces from pulmonary cases of tuberculosis, which had not yet developed clinically. This is also suggested by the smaller number of bacilli found in the milk. Tubercle bacilli therefore occur in the milk in great numbers when animals with open tuberculosis, and tuberculosis of the udder stand in the stable. Milk from animals which manifest their tuber- culosis by a positive tuberculin reaction, will usually be free from tubercle bacilli. Such cows belong to the least dangerous class. Nevertheless the investigations of Rabinowitsch, Kempner, Rav- enel and others, who obtained positive inoculation results with milk from reacting animals, prove that such milk may at times contain tubercle bacilli in small numbers. Ordinarily however this will not be the case, and the milk of such animals may, as a whole, be considered free from tubercle bacilli. Therefore it appears evident that under present conditions of milk production the ingestion of tubercle bacilli with milk is possi- ble at almost all times. Tubercle Bacilli Types. 103 What Danger Threatens Man Through Ingestion of Milk Which Contains Bovine Tubercle Bacilli? In order to answer this question it is necessary to refer to the development of tuberculosis in man, and to consider the fac- tors which are necessary for an infection of his body. These factors in addition to the toxicity of the infective agent, and the quantity in which it has the opportunity to enter the body, depend upon the avenues of infection which it takes and the local and general resistance of the attacked individuals towards the spe- cific infective agent. There is perfect agreement relative to the virulence of the bovine tubercle bacillus for man. The bovine tubercle bacillus is a strain of the tubercle bacillus with such pregnant characteristics that it is almost invariably possible to classify it separately from other strains when obtained in culture, that is, to distinguish the bovine tubercle bacillus from the bacillus of the human type. These bacilli are distinguished as the typus bovinus and typus humanus (only these two types need to be considered from the standpoint of milk hygiene) which are characterized by the fol- lowing peculiarities : Typ. Bovinus. The growth is delicate and in the thin film small wart-like colonies develop; on bouillon a mesh-like fine membrane with wart-like prominences develops proliferating downward, or a membrane of tissue-paper thick- ness results; the bouillon is probably neutralized and finally becomes alkaline. The bovine tubercle bacil- lus is as a rule of greater toxicity for smaller mammalia. Rabbits develop through intra- venous injections of the bacillus (0.001 gm.), a generalized tu- berculosis from which the ani- mals succumb inside of three weeks. When 0.01 gm. is in- jected under the abdominal skin, it produces in a short time, gen- eralized tuberculosis. Cattle succumb readily to Typ. Humanus. The growth is a luxuriant, uniformly thick and wrinkled membrane, which proliferates on the wall of the tube. The growth is the same in bouillon; the degree of acidity of the bouillon is usually at first diminished, later increased. 0.001 gm. of bacilli of the human type intravenously in- jected produces after months only a chronic form of tuberculo- sis (joints, kidneys, lungs, testicles). The injection under the ab- dominal wall produces only local lesions. Inoculated into cattle the bacilli of the human type (0.05 gm. subcutaneously), produce only slight or no pathogenic ac- tion. The process remains local, and extends only to the 104 Tuberculosis, infection with the typus bovin- us, from an extensive form of tuberculosis. Guinea pigs die more quickly from an inocula- tion with bovine tubercle bacilli than from an inoculation with the bacillus of human type. neighboring glands; a great healing tendency prevails. Intravenous injections of 1 mg. of the typus humanus into the vein of a mouse will show it to possess a greater resistance than when inoculated with the typus bovinus. In the last 10 years about 2000 strains of tubercle bacilli from man and cattle have been cultivated and studied. In these studies even further differences were found which, however, are not as constant as those given above; for instance, the bacillus of the human type in glycerin bouillon cultures is delicate, slender, slightly curved, and of beaded staining qualities, whereas the bovine strain is regular, plump, thick without granular differentiation in staining, and frequently with swollen ends. The pigment formation on glycerin potato is more typical of the human type than the bovine type. The former when placed on serum in hermetically sealed glass tubes remains viable for twelve months, the latter for over a year. Contrary to the views of many investigators of tuberculosis, Nocard, Hueppe, Von Behring, Komer, de Jong and others support the theory that the tubercle bacillus adapts itself to the infected animal and becomes transformed as a result of its environment. Rabinowitsch, Dammann and Eber have also supported this trans- formation theory. The latter especially attempted to prove by ex- tensive experiments and investigations that Bacillus humanus, by passage through cattle, changes into the bovine type. This question however appears at the present, to be decided in favor of the stability of the bacillus. It has at least been shown with six various strains of the human type, that in passage experiments through 2 to 7 cattle, in from 247 to 512 days, the character of the bacillus was not changed (English Commission and Weber), and that bacilli of the human type by eight subsequent passages through goats, in 516 days, and by four passages through cattle in 685 days, were not influenced in their typical characteristics. The immunization experiments which were undertaken with the bacillus of the human type on cattle showed no changes whatsoever in the human type after the pres- ence of the bacteria in cattle for a year and seven months (Baldwin) in spite of their propagation in the udder of the cow. The same results were obtained in three experi- ments by Weber, Titze and Joern, who allowed the bacillus of the human type to exist in the body of cattle for two years and one month and for two years and six months. Eber's experiments found no confirmation in the Imperial Board of Health; the experiments however are being continued in strict co-operation with Eber. On the contrary it is shown that the bovine bacillus constantly retains its characteristics within the human body. From a boy who was affected since his second year with tuberculosis of the fourth digital bone of his hand, it was possible during surgical interference to obtain material from the same place at five different times, during his age from 8 to 13 years. The in- fection was caused by the bovine bacillus. In spite of their existence for ten and a half years in the human body these bovine bacilli had retained their characteristics. A marked influence in their virulence was manifested, however, since the bacilli, from the second operation, in quantities of 2 mg. could no longer kill rabbits even after intravenous inoculations. From the sub- sequent operations it was found that the virulence was again increased. Griffith obtained the same results with bovine cultures from lupus from which the bacilli were isolated six months, two and three and a half years, respectively, after the first examination. It is true that the virulence was several times lower than is or- dinarily the case with the bovine type; the other characteristics of the strain however Transmission of Bovine Tuberculosis. 105 were tenaciously retained. In one of the cases the bacillus persisted in the human body for 18 */2 years. Passage through rabbits and cattle again increased its virulence. Al- though through animal passage a change of the virulence is possible, nevertheless this change results only inside of the borders of the type and in the direction of the type. The bovine bacillus therefore does not pass into the human type, nor the latter bacillus into the bovine type. Only a brief statement will be made relative to the so-called atypical strains. It has been demonstrated that there are cultural strains which cannot be classified as belonging either to one or the other type (Kossel, Weber and Heuss, Lydia Rabinowitsch, de Jong and others). These strains proved to be mixed cultures of both types. In the same person not only mixed infections of both types may exist in the affected organs, but also a double infection may occur in such a way that in one organ the Typus humanus, and in the other organ the Typus bovinus, may be found in pure culture (Weber, Weber and Taute, Griffith, Park and Krumwiede, Steffenhagen). In 1901 Koch explained at the International Tuberculosis Congress at London, that tuberculosis of man is produced by a tubercle bacillus which differs from the bovine tubercle bacillus, and expressed himself as opposed to the general prevailing opin- ion of that time, regarding the great danger of the cattle tubercle bacillus for man, and as believing that the transmissibility of bovine tuberculosis to man was so slight compared with the dan- ger which threatens man from tuberculous human beings, that its practical importance was negligible. Although Koch's statement cannot stand in the directness of his declaration, nevertheless at the present time it is generally accepted from the above mentioned differential characters, that marked differences exist between the bacillus of bovine tubercu- losis and that of man, and it is a fruitless work to dispute whether they are differences of varieties or peculiarities of the different strains, which lead to the variations, if we accept the fact that the differences of the strains are obstinately retained. The results are of especial value in differentiating the two types of tubercle bacilli. In association with Shiitz, Koch under- took some experiments to establish points of differentiation. Nineteen calves which were infected intravenously, subcu- taneously, intraperitoneally, by inhalation or feeding experiments with the Bacillus humanus, showed no manifestations of disease, in- creased in weight, and on autopsy conducted several months after infection, showed only caseous purulent changes at the point of inoculation. On the other hand, after the inoculation of bovine tubercle bacilli, severe febrile symptoms and extensive tubercu- losis, especially of the lungs, liver and spleen resulted. The same results were obtained from the experiments of Kossel, Weber, Heuss. Bacilli of the human type were retained in the regional lymph glands ; the changes induced by them gradually retrogressed, whereas infection with the bovine type of the bacillus led to a pro- 106 Tuberculosis. gressive tuberculosis. Inhalation and feeding experiments showed the slight virulence of the human tubercle bacillus for cattle. In the experiments of Nocard, Meyer, Calmette and Guerin and Zwick, the inoculation of tubercle bacilli of bovine origin into the milk ducts resulted in a tuberculosis of the udder with rapid emaciation of the animal, terminating in death ; whereas the bacilli of human origin produced only a passing inflammatory irritation, and an interstitial atrophy of the udder. Calves which nursed on these latter infected udders remained healthy (Zwick and Maier), or on the other hand (in one case of Zwick) intestinal tuberculosis, with tuberculosis of the mesenteric lymph glands, developed. At autopsy undertaken 20 weeks after the infection, the udder of the cow showed atrophy with miliary tuberculosis, without however typical tuberculous changes in the regional lymph glands. From these results the conclusion may be drawn that tubercle bacilli of human origin are only very slightly dangerous for cat- tle. It should be considered however that occasionally after artifi- cial infections the bacilli may persist in the infected region, with or without marked local or at times even generalized changes. Calves may develop intestinal tuberculosis or tuberculosis of the mesenteric lymph glands as a result of ingesting a large amount of tubercle bacilli of the human type. Almost the same relation exists in man towards the bacillus of bovine tuberculosis. The principal dangers threatening man are through the possibility of infection from affected human beings, and less so to the possibility of infection with diseased products of animal origin, as for instance milk. The possibility of tuberculosis infection through animal products is presented with remarkable frequency, as may be seen from the above statements; still the rarity of infection with the bovine type is quite striking. Hogs which become readily infected with the bovine type are very frequently af- fected by the ingestion of skimmed milk containing tubercle bacilli. In northern Germany some of the herds show an infection of 50-60, occasionally even up to 90%. The experience at the tuberculosis eradication stations indicated that by the elimination of cattle affected with open tuberculosis a marked reduction was obtained in tuberculosis of hogs, and that this measure in association with pasteuriza- tion of the skimmed milk, offers a certain remedy against the spread of tuberculosis of hogs. The same opportunity which is afforded hogs to contract- tubercle bacilli from the feeding of skimmed milk, would apply to man. The relative infrequency of the infection of man with the bovine type of tubercle bacillus is not the result of a milder virulence of the bacilli but is due to the previous boiling of the milk. Convincing observations have also been made on this point. However before entering into a discussion of these, it will be advisable to illustrate further the possibility of infection for man from the standpoint of the port of entry, and also show the relative condition existing between the necessary infective quantity of bacteria and the establishment of the disease. Ingestion Tuberculosis. The development of the affection depends on the most varied conditions, on the quantity of the introduced virus, condition of the port of entry, general resistance, etc. Frequency of Tuberculous Infection Through the Alimentary Tract. If the lesser virulence of the bovine type for man, as compared to the human type, is left out of consideration, which fact is con- sidered satisfactorily proven, the experiments of Ostermann, Schroeder and Cotton show what great quantities of infectious ma- terial are necessary in order to produce tuberculosis by ingestion. Schroeder and Cotton fed milk artificially infected with tuber- cle bacilli, and proved that infected milk which invariably produced tuberculosis when inoculated intraabdominally in 5 c. c. doses, could be fed for 30 days without causing the disease in the ex- perimental animals. The dilutions were prepared (1) by adding one platinum loopful of a cloudy sus- pension of tubercle bacilli to 10 c. c. of milk. (2) by adding one loopful of the orig- inal suspension to 10 c. c. of sterile water, and of this dilution one loopful was placed into 10 c. c. of milk, (3) by adding one loopful of the original suspension to 100 c. c. of sterile water and of this dilution one loopful was placed into 10 c. e. of milk. It was not possible to produce ingestion tuberculosis with either the second or third dilution during the period of the experiment, although the dilutions were not as high as they occur in the milk of tuberculous animals (with the exception of tuberculosis of the udder). Ostermann by comparing the average number of tubercle bacilli in cow's milk with the minimal dose necessary for producing ingestion tuberculosis in guinea pigs, rabbits and goats, came to the conclusion that an alimentary infection is exceedingly rare. Nevertheless the danger of an alimentary infection with bovine tubercle bacilli, even in high dilutions of the tuberculous material in market milk, cannot be disregarded. The danger of infection to which small children are exposed from the ingestion of food (without attempting to distinguish "bo- vine tuberculosis" from "human tuberculosis") is best illustrated by the clinical cases and also those cases of intestinal and mesen- teric tuberculosis which are found on autopsy. Edens, from October 1, 1904, to September 30, 1905, found 12%, and from this time until September 30, 1906, 13.6% of the bodies of children which he autopsied at the ages of 1 to 15 years, affected with primary intestinal tuberculosis or tuberculosis of the mesenteric lymph glands, whereas in man from 15 to 19 years of age only 3.8% and 2.6%, respectively (all autopsies), showed the disease. The intestinal tract of children appears therefore to be a prominent port of entry for the tubercle bacillus, which is also proven by the works of Orth, Henke, Chiechanowski, Hamburger, Nebelthau, Lubarsch, Bruning, Fibiger and Jensen, Symes and Fischer, Price and Jones, Kingsford, Harbitz, Ogyia, Edens, Wagener and Heller, who demonstrated primary intestinal tuber- JOS Tuberculosis. culosis in varying proportions, up to 47.6% of the tuberculous children. The frequency, however, with which tubercle bacilli actually pass through the intestinal wall without producing demon- strable changes in the intestines and mesenteric lymph glands cannot be stated. The intestinal tract may be the avenue of infection without itself or its regional lymph glands becoming in- fected. The percentage given above should therefore be higher. The works of McFadyean, MacConkey, Harbitz, Weichselbaum, Bartel, Rosenberger, Rabinowitsch, Ipsen and others offer proof for this contention, as they mention cases in which apparently healthy mesenteric lymph glands contained tubercle bacilli which although appearing to be in a latent form at the time of finding, produced tuberculosis when inoculated into experimental animals. It appears also to be proven experimentally that through the feeding of tuberculous material tuberculosis of the lungs may develop without the presence of intestinal tuberculosis or tuber- culosis of the mesenteric glands (Bartel, Bongert, Kovacs and others). In this regard the question may be raised as to whether there is any possibility of the tubercle bacilli working up from the intestines into the esophagus, and into the buccal cavity, from which inhalation tuberculosis could result (Uffenheimer, Dieterlen). This objection would not enter into consideration for the pur- pose of milk control, since it is immaterial for the hygienist work- ing along practical lines, whether the infectious agent causes dis- ease in the body by way of the circulation or through inhalation. In this instance it is only necessary to keep in view preventive measures, which should completely prevent the body from com- ing in contact with producers of the infection. Alimentary Infection of Man With Bovine Tuberculosis. After the supposed cases of transmission of the bovine tuber- cle bacillus cited in former years failed to withstand critical observations, Koch at the International Tuberculosis Conference, held in Berlin, in 1902, urged the following up of all cases of established tuberculosis of the udder, the determination of how long the disease persisted, who consumed the milk and milk pro- ducts from these cases, whether the milk had been boiled and whether the respective persons became affected with tuberculosis. This request was fruitful of results, and in 1910 Weber pub- lished the results of his compilation investigations, which were carried out by the aid of official statistics from Prussia, Bavaria, Saxony, Wurttemberg, Baden and Hessen. The investigations extended over the time between the be- ginning of 1905 to April, 1909; the investigation of some of the individual cases however is still being continued, since in the chronic courses of tuberculosis it must be considered that the Bovine Tuberculosis in Man. 1Q9 results of infection with bovine tubercle bacilli may under certain conditions only manifest themselves after years have elapsed. In the given period 113 cases were reported, of which 68 were from Prussia, 14 from Bavaria, 6 from Saxony, 6 from Wurttem- berg, 10 from Baden, and 9 from Hessen. At least 628 persons came under consideration in the inges- tion of such milk, possibly even more, since at times only the term 11 family" is designated, and the milk was not infrequently deliv- ered to dairies with a large patronage. These cases were not in- cluded, although every person is exposed to an infection who par- takes of such milk and dairy products. In the case of 9 persons no age is given; 284 were children, and 385 adults. The value of the individual cases must of course be judged in different ways. In 44 cases it is stated that the milk had been consumed only as an addition to coffee, or mixed with milk of healthy animals, or the data were otherwise not accurate. Of especially great interest are those cases in which it was emphasized that the milk was consumed in a raw state, unmixed, mixed with milk of only a few cows, or in which such milk was used in the preparation of butter, buttermilk, sour milk, or had been consumed for a long period. In such cases tremendous num- bers of tubercle bacilli must have been taken into the digestive tract. According to Bang and Wall the milk from tuberculous udders may retain a normal appearance for months, being used as food without any objection, and yet such milk contains millions of tubercle bacilli. Bang found in smear preparations of such milk, in a single field as many as 200 bacilli. In all 69 cases were reported, in which it was stated with cer- tainty that raw milk of animals with udder tuberculosis, or pro- ducts prepared from such milk, were consumed. The milk was taken for a longer or shorter time, in large quantities, by 151 children, 200 adults, and 9 persons whose age was not mentioned. These persons are divided by Weber according to the results of the investigations, into four groups, namely: 1. Cases, in which an infection occurred of a bovine type. 2. Those in which a suspicion of an infection exists, but on account of insufficient bacteriological examinations has not yet been determined. 3. In affections in which the bacteriological examination relative to the suspicion of tuberculosis was negative, or in which the human type was found exclusively, and 4. Cases in which no affections whatsoever have been dem- onstrated up to the present time. In Group 1 an infection with the bovine type was demonstrated in two families, affecting one child in each. 110 Tuberculosis. In both cases it was possible to trace the consumption of mill containing the bacilli up to the nursing age. In one case it laste( for one and a half years, in the second case one year, and in th< latter case the udder affection had been recognized for thre< months during the period that the child had been using the milk In both cases the respective cow was affected with a sever* tuberculosis of the udder in all four quarters ; the milk had beei consumed at all times mixed with the milk of a second cow, in th< first case boiled or raw but in the second case only raw. The other members of the family remained well in spite o: the consumption of this milk ; in both instances only the younges child became affected with tuberculosis of the cervical glands. In the first case another child of four, and one of five years was included in the family ; in the second case children of the ag< of 3, 4, 7, 8, 9 and 12 participated in the consumption of the milk all remaining normal. The tuberculosis of the cervical glands healed in the tw( youngest after abscess formations, leaving several slightly en larged small glands in the surrounding parts. One of the boys appears to be in the best of health, the other is somewhat behind ii his development (at the age of 2 % years he weighs 25fbs.) ; in tin last six weeks however his weight increased slightly more thai 1 lb. In the cases of the second group there exists suspicion of { bovine type of infection. In six children and one adult there are swellings of the cervi cal lymph glands and in four children and one adult a suspicioi of abdominal tuberculosis is given. One child is affected witl scrofula. In the four children the manifestations of disease re trogressed, while in the adult it appears doubtful, according t( Weber, whether the affection is of a tuberculous nature. Forty-one persons are included in Group 3, who consumec milk in a raw state from cows affected with tuberculosis of th< udder. This was at times mixed with milk of other cows. A girl o: 16 years of age and a boy four years old, who died of tuberculosis were included in this group. The producers of their infections were bacilli of the human type. A man and a woman who wer( affected with pulmonary tuberculosis (human type), a boy witl suppuration of the middle ear and cervical lymph glands (no' tubercular) ; an 18-year old boy with rheumatism of the joints and valvular heart trouble, chronic diarrhea and pulmonary symp toms (not tubercular) ; a woman with catarrh of the apex of th( lungs (not tubercular) ; a woman with swelling of the glands diarrhea, cough, night sweats and emaciation (inoculation o1 sputum without results), and a woman and a man with pulmonary symptoms (not tubercular)* were also in the total of forty-one. The cases in which a boy and a girl died from tuberculosis are of especial importance. In spite of the prolonged consump Table III. Milk from a tuberculous udder. 1 X 1200. Ernst, Milk Hygiene. Danger from Bovine Tuberculosis. tion of raw milk from a tuberculous udder by the children who were already infected with the human type of the disease, it was impossible to isolate from the tuberculous glands of the neck and mesentery any bacilli of the bovine type. Weber concludes from this that a body already infected with the human type of the dis- ease is resistant rather than susceptible towards an infection with the bovine type. The fourth group contains by far the greatest number of cases in which children and adults consumed raw milk from cows affected with tuberculosis of the udder, or milk products prepared from the same, and includes those cases in which no disturbances of the health resulted from such consumption. It was especially stated relative to the children that they all appeared thriving and healthy. Among these persons are included those who for a long period ingested especially great quantities of bovine tubercle ba- cilli ; thus a 13-month old child has been brought up exclusively on raw and boiled milk from a cow affected with udder tuberculosis, and up to the present remains healthy. Other cases may be considered as presenting complete ex- periments with the necessary controls, since the persons who drank the milk remained healthy, whereas calves and hogs fed with the same milk developed severe ingestion tuberculosis. The ob- servations of such cases may be traced back incompletely for 3 or 4 years. A 25-year old waitress, and a 28-year old dairy hand drank mugfuls of freshly drawn tuberculous milk, frequently without any other milk being added without be- coming affected; the calf of the cow which produced the milk had to be slaughtered after four weeks, and showed tuberculosis of the mesenteric lymph glands, liver, lungs and kidneys, a severe ingestion tuberculosis. The milk of another highly affected tuberculous cow also suffering from udder tuberculosis, was mixed with the milk of two other cows, and was consumed frequently in a raw condition, by two adults and a 13-year child; a child 1% years of age was given the milk only in a boiled condition. The adults remained healthy; the calf from this cow had to be slaughtered after five weeks, and showed generalized tuberculosis. From the stable of a herdsman the milk of a cow affected with udder tuberculosis was mixed with the milk of three other cows, and the cream and butter prepared from this was consumed by four persons, aged 31 to 59 years, without producing any ill effects. The five hogs of the herdsman were found on postmortem to be tuberculous. Similar results were reported from the ingestion of milk from a tuberculous udder of a goat, which was consumed as raw and boiled milk by three adults and four chil- dren of ages from 5 to 16 years. The persons remained well while a hog became af- fected with ingestion tuberculosis. In two other cases the milk was consumed in a raw or unmixed state, as milk, buttermilk and butter. It was consumed by seven adults, in one case for a period of four months, in another case even longer. In spite of the fact that the family has been kept under observation for four years no disturbance in health can be detected. It is proven by the collected material of Weber that even though tremendous quantities of tuberculous material are con- sumed, still more favoring accessory conditions are necessary in order to produce an infection with the bovine type of tubercle bacilli. Of course it is not yet known, as indicated by Weber, how many of the children which show swelling of the cervical lymph glands and symptoms of suspected abdominal tuberculosis, are 112 Tuberculosis. affected with the bovine type of tuberculosis, or how many of the persons who fail to show any disturbance of health may harbor one or more infected mesenteric glands ; likewise it is not known how many children with a latent form of the disease may through a special weakening, or under the influence of other infections, break down later with tuberculosis, possibly even with a fatal termination. Through the compilation investigations we know only of the time (which extends over a period of 1-3-4 years in the individual cases), the opportunity and the immediate results of the infection, and not the further development of the same, but we do know that in two children a true bovine type of tuberculosis existed. Therefore, although a possibility of infection was present in a great number of persons, the infection has positively occurred up to the present only in two children in infancy. This constitutes proof that "the danger which threatens man from the consumption of milk and milk products from cows affected with udder tubercu- losis is very slight when compared with the danger of man affected with open pulmonary tuberculosis to his fellow men." This conclusion of Weber may be supported without further consideration. Nevertheless the danger still prevails, and al- though it is slight in comparison with the danger through infec- tion with the human type, it should be by no means under-esti- mated; it should be considered that the danger of infection with human tuberculosis is amazingly great, and the opportunity of ingesting the bovine type of tubercle bacillus with milk is similarly great. Bovine Tuberculosis in Man in General. Although the attention of pathologists of all countries has been directed for the last ten years to infections of man with bovine tuberculosis, up to the present time there are collected only 117 cer- tain cases of bovine tuberculosis in children and 21 cases in adults (over 16 years of age). Of the 117 cases in grown children 105 are accurately de- scribed, and invojve the following organs : 60 cases of abdominal tuberculosis 25 cases of tuberculosis of the cervical glands 4 cases of tuberculosis of the tonsils 7 cases were generalized 3 cases were localized in the bones and joints 6 cases represented lupus Two cases should also be included in which bovine bacilli were found in unchanged lymph glands. The 60 cases of abdominal tuberculosis are again divided into 34 severe cases in which the mesenteric lymph glands, the intes- tines and the peritoneum showed changes. Thirty of these after generalization of the affection, terminated in death. Bovine Tuberculosis in Man. Twelve of the 60 patients had tuberculosis of the mesenteric glands, slight intestinal tuberculosis, and tuberculous meningitis. The 12 cases were severe fatal affections. In 14 cases the autopsy revealed tuberculosis of the mesen- teric lymph glands with the bovine type of bacilli, but this was found accidentally following other causes of death, as diphtheria, scarlet fever, measles, and pneumonia. In the 21 adults the bovine type of the disease was established three times in pulmonary tuberculosis with expectorations, once in a primary abdominal tuberculosis and pulmonary tuberculosis, once in an infection of the buccal mucous membrane and cervical lymph glands, once each in tuberculosis of the knee joints, the kid- neys and the peritoneum, and finally the bacillus of bovine type was isolated from three cases of lupus, two cases of skin tuber- culosis, and five cases of tuberculosis verrucosa cutis in butchers. Besides these instances the bovine type of tubercle bacillus was isolated three times from the mesenteric glands of adults. In two cases of phthisis the bovine tubercle bacillus was found in association with the human type. Of the total of 138 cases, 56 were fatal, and 89 could be ex- plained with certainty or with the greatest probability as inges- tion tuberculosis. The other forms of tuberculosis, with the ex- ception of the skin tuberculosis of the butchers and of one milker, may also probably be traced to the same mode of infection. Weber deduces from his findings that the danger of becoming infected with tubercle bacilli of cattle is great for the individual, but is only slight for the human race as a whole. Kossel reports in the German Medical Weekly relative to the number of cases of animal tuberculosis in man as compared with the human type of tuberculosis, and observed that in 1602 cases of human tuberculosis the bovine type appeared as the infective agent in 126 cases, the human type alone in 1464 cases, the human and bovine type in association nine times, and the avian type of tubercle bacillus three times. Therefore in about 8.6% of human tuberculosis, bacilli of animal origin were found, and in about 8% of these they were of the bovine character. If however the most frequent form of tuberculosis of man is considered, namely pul- monary tuberculosis, then the bovine type can be demonstrated only in about .6% of the cases, whereas in the other forms of tuberculosis it may be found in 16% of the cases. Tuberculosis of bovine origin occurs most frequently in chil- dren in which tuberculosis of the cervical glands is caused in about 40% of the cases from infections with the bovine type, and tuber- culosis of the mensenteric glands may be traced to the same type in 40 to 50% of the cases. A portion of these affections, as has al- ready been mentioned, may terminate fatally. Among the fatal forms of tuberculosis in children 76% are caused by the human type and 24% by the bovine form. The meningitis type of the Tuberculosis. disease is brought on in only about 11% of the cases by the bacil- lus of animal origin and in 89% by the human type of the organism. In tuberculosis of the bones and joints the figures are 5% and 95% respectively. Gaffky, Kothe and Ungermann found in 400 bodies of children, 76 infections with tuberculosis, in which they succeeded in estab- lishing the variety of the bacillus. In one case they found the bovine type (1.32%), and among 171 other autopsies on children, of which 39 were tuberculous, two (5.1%) cases of bovine infection were observed. The results of tuberculous infections among children of the population of Berlin were therefore 95 to 96% of human origin, while only 4 to 5 % were of bovine origin, iii spite of the fact that during infancy the danger of bovine infection is the greatest (Kossel). [According to figures compiled by Park of the New York City Board of Health, the frequency of bovine tuberculosis in man as collected by various investigators is as follows : In adults, 955 cases have been examined of which 940 showed human infection and 15 bovine infection. In children from five to sixteen years of age, out of 177 cases investigated, 131 were human infections and 46 bovine infections. Among children under five years old there were 368 cases of which 292 were found in- fected with the human type and 76 with the bovine type of tuber- culosis. Furthermore Park mentions the very suggestive results obtained from nine children under 6 years of age who were fed exclusively on cow's milk at the Foundlings' Hospital. Five of these children died of bovine infection and four of human infec- tion. On the other hand in the Babies' Hospital where the infants are nursed or fed on prescription milk, out of 63 children dying of tuberculosis, 59 proved to be human infection and 4 bovine infection. The figures taken from clinical work in England indicate that from 23 to 25% of the fatal cases of tuberculosis in children are due to bovine infections. Stiles of Edinburgh has presented in- teresting statistics to illustrate how bovine tuberculosis particular- ly affects young children. Of 67 consecutive tuberculous bone and joint cases, the bovine bacillus was present in 41, the human bacil- lus in 23, while in 3 cases both types were present. In those af- fected children under 12 months old, only the bovine bacillus was found. Of the 12 children between 1 and 2 years of age, 8 owed their disease to bovine infection, 2 to human infection and 2 to both bovine and human infection. There were 15 cases in 2 to 3 year old children, 11 of which were bovine, 3 human and 1 both infections. The 10 cases from the 3 to 4 year period were 6 bovine and 4 human infections, while the 4 to 5 year period included 3 cases of each type of infection. Stiles further reports on 72 cases of tuberculous cervical glands operated on at the Children's Hospi- Control of Bovine Tuberculosis. tal in Edinburgh, in which the disease was due to the bovine bacil- lus in 65 cases, while in only 7 patients was the disease caused by the human bacillus. — Trans.] Conclusions. If we compile the results of this chapter the following conclu- sions may be established: Although tuberculosis of cattle is less dangerous for man than tuberculosis of man, the danger from the enormous spread of the disease in our herds, and especially among the dairy cows, should in no way be under-estimated. Theoretically the possibility of infection is afforded in all cases in which the ingestion of living tubercle bacilli with the milk takes place ; from a practical stand- point however this possibility of infection comes into consideration only when the bacilli enter the individual in great quantities, and the resistance (of a local or general nature) of the body is not equal to this quantitative attack. This disposition, or these rela- tive conditions between the injurious agents and resistance, appear to be especially unfavorable in children ; therefore the requirement of the elimination from dairy herds of all tuberculous animals which pass tubercle bacilli writh their milk, appears to follow as a matter of course. According to the experience at the tuberculosis eradication stations only those animals must be considered as eliminators of tubercle bacilli wrhich are affected with open tuber- culosis, and expel the tubercle bacilli with their secretions and excretions, especially animals affected with tuberculosis of the udder, open pulmonary tuberculosis, tuberculosis of the uterus, intestinal tuberculosis, and furthermore animals with tuberculosis of the liver, kidneys, skin, eyes, and larynx. Measures Against the Danger. The elimination of animals passing tubercle bacilli should also be energetically encouraged on general economic grounds. For this work three methods may be followed : 1. Treatment of the disease and curative attempts. 2. The immunization of healthy herds. 3. Energetic sanitary police eradication measures, reduction of the possibilities of infection, and protection of young animals from infection, together with favorable conditions for bringing up young stock as a preventive measure against their accidental infection, toward which we are powerless. The curative measures in affected animals may be left out of consideration as measures of control, since — excepting the uni- formly bad or only slightly favorable results — the methods of treatment for veterinary practice are too complicated, and are not practicable in consideration of the value of the animal. For the sake of completeness the experiments with iodipin should be mentioned here (Hauptmann). Creosote has also been employed. Tuberculosis. Of the specific remedies, tuberculin, tulase, tulase-lactin, tulon and tuberculase could be considered in the treatment of affected animals. These bacterial preparations, however, accord- ing to Romer and Arloing, are ineffective, since the results were negative. Better results were promised at the onset, from the specific immunization methods, which aimed at a systematic preliminary treatment with slightly virulent strains, or with attenuated bovine tubercle bacilli, to increase artificially the resistance of the im- munized animals, that is, to protect them against a later accidental natural infection. As a matter of fact cattle immunized with tubercle bacilli prove for a time to be immune, or at least manifest a considerable resistance against a subsequent artificial infection with bovine tubercle bacilli, when compared with non-immunized control animals. For immunization purposes there have been used: 1. Dry tubercle bacilli of the human type (bovo-vaccine, Von Behring's method). The injection is made into the blood circula- tion and is repeated. Animals treated in this way after 3 to 4 months, resist an intravenous injection of bovine tubercle bacilli, to which untreated animals invariably succumb. This increased resistance however lasts only a short time. According to the in- vestigations of Rossignol and Vallee and Hutyra it diminishes to- wards the end of a year, and after another six months it practically disappears. Against the slight practical success of this method the disagreeable fact should be considered that the injected tubercle bacilli of man are retained alive in the body of the cattle for years, and may even produce in the udder local tuberculous processes, from which the bacteria of human tuberculosis may enter into the milk (Lignieres, Weber and Titze). Titze found that following an intravenous injection of human tubercle bacilli, they were eliminated from the udder even 16 months after the injection. In this regard the various individuals manifest an entirely different behavior. In three other cases bacteria were eliminated after a single injection, from the fourth week up to the 144th day. In a second cow which received an injection of tubercle bacilli of human and bovine type the elimination commenced after the third injection, and in a third cow as early as 24 hours after the injection. All three animals eliminated the bacilli from only one quarter, without this showing tuberculous changes. Bongert found in 186 bovo-vaccinated cattle, 36 which passed tubercle bacilli with their milk. The protective vaccination of Von Bearing therefore is not only of little practical value, but grave dangers must be considered in connection with it, since the vaccinated animal may eliminate tubercle bacilli with the milk for 2y2 years and longer. Koch and Schiitz, Neufeld and Miessner recommended for the immunization of cattle a single injection of 0.01 gm. tubercle bacilli in suspension, which vaccine they termed ' ' tauruman. ' ' The above statement applies equally for tauruman as it does for bovo-vaccine. Similar results to the immunizing value of the in- travenous injections, according to Baumgarten, Lignieres and Tuberculosis Vaccination. Klimmer may be derived from the single subcutaneous admin- istration of human tubercle bacilli. According to Lignieres even in such cases the bacteria may remain alive for as long as two years. According to Von Behring, Calmette and Guerin, Roux and Vallee, cattle may become immunized by feeding with slight quantities of bacilli from tuberculosis of the horse (or bovine tuberculosis). Arloing attempted to immunize with homogenized cultures of strains which had been cultivated in 6% glycerine bouillon (human type and bovine type). Better results were obtained from the intravenous than from the subcutaneous applications and this again proved superior to administration per os. Klimmer eliminated the danger of the vaccination for man by heating the human tubercle bacilli to 52-53 deg. C., or by rendering them avirulent by continuous passages through the crested newt. Both these vaccines are no longer pathogenic for guinea pigs, and they cannot regain their virulence by means of passages through animals. The results of immunizations are supposed to be favora- ble (Klimmer on 10,000 cattle), especially if the vaccination is carried out together with general protective measures, such as raising calves on milk free of tuberculosis, and the elimination of animals with open tuberculosis. Glockner even believes that vac- cination has a favorable action on the curing of animals which were already affected with bovine tuberculosis prior to the vaccination, whereas Eber attributes the improvement of the vaccinated herds to the simultaneously executed prophylactic and hygienic meas- ures. Friedmann aimed to produce immunization with his tuber- cle bacillus from cold blooded animals (turtle). Other authors however failed in producing an effective immunization with such strains (Libbertz and Euppel, Weber and Titze, Orth). Heymanns attempted to immunize cattle by the introduction under the skin of cattle of a closed sack of vegetable fiber, contain- ing living tubercle bacilli (human or bovine in origin). The sup- position is that these vegetable sacks will confine the bacilli at the seat of inoculation, and that the treated animal will be immunized by protective metabolic products, that continuously form in small quantities within the sack and pass outward from it into the animal's system generally, by an osmotic process. The vaccination, which is carried out with the aid of a tro- car to insert the capsule under the skin of the back, must be re- peated annually, since the bacilli may die. Heymann's method has been successfully used by its discov- erer on more than 20,000 cattle, and the percentage of reactors to the tuberculin test diminished from 45 to 21 (18 herds with 188 animals). Animals which have formerly reacted may appear free at the subsequent test. . Good results were obtained by Vallee from passive immuniza- tion. He inoculated young cattle with 100 to 200 c. c. of a protec- tive serum, which he obtained from a horse treated with slightly virulent strains from horses, and then with strains from men. 118 Tuberculosis. With this method he succeeded in rendering the animals resistant to artificial infection with bovine tubercle bacilli. Since immunization methods have not offered uniformly satisfactory results, and since they must be prohibited on the ground of milk hygiene, therefore results may be expected only from proved sanitary police measures. The methods which must be followed in the eradication of bovine tuberculosis are: 1. Diminution or elimination of the sources of infection, (a) By removal of the animals passing bacilli, (b) By separation of healthy and suspected or diseased animals, (c) By bringing up tuberculosis-free young animals. 2. Improvement of the general methods in the care of young stock, by introducing conditions which approach the natural mode of living: (a) Proper care and feeding in well ventilated and lighted stables, (b) Dividing the pastures so that the animals may be sepa- rated (according to whether they are suspected or healthy) and kept in accordance with their age and with the use for which they are later intended. Measures for eradication must be applied in accordance with the rules here outlined. The most effective method of eradication was worked out by Bang, and consists in the elimination of clinically recognizable dis- eased animals, the separation of reacting animals, and the bringing up of calves on milk free of tubercle bacilli. The remarkable value of Bang's methods has been proven fully in practice by the results obtained since 1892. It is important for the results to separate completely the animals which fail to react to tuberculin, that is the healthy cattle, from those which harbor the disease and which react to the tuber- culin test. This should be done in such a way that the healthy animals are placed in a freshly disinfected stable or in a portion of a stable provided with a separate entrance, and separated with a board wall, from that part in which the reacting cattle are housed. The attendants of the healthy herd should not come in contact with those of the diseased herd. Animals of the reacting group which after a time become affected so that they may be clinically recog- nized, should be slaughtered as soon as possible. Young stock which react should not be permitted to breed, or at least should be immediately placed with the reacting group, providing their breeding value is such that this procedure is deemed advisable. All reacting animals under six months of age should be slaughtered, that is they should be utilized for meat. Young stock and work oxen should also be included in the Methods of Eradication. segregation, and the healthy ones must be kept from contact with reacting animals. Of the calves which are born after the separation, those from non-reacting cows remain with their mothers; the calves from reacting cows, after receiving the colostrum from their mother on the first day after birth, should be placed in the stable of healthy animals, and should be fed with the milk of healthy cows or should be brought up on sterilized milk, or they may be allowed to suck from healthy nurse cows. As soon as possible after weaning the calves should also be subjected to the tuberculin test, and those giv- ing a reaction should be immediately removed. From 1 to "2% of these calves react. It is proper to place the healthy calves in a stable of healthy young stock, and they may pasture with them, or if this is not pos- sible they should be placed with the older non-reacting group of animals. Before the first breeding the heifers again should be subjected to the tuberculin test, in order to place them in the prop- er group of cows. The tuberculin test is annually repeated in the healthy herd, in order to eliminate the animals which in the course of the year have had a possible opportunity of becoming affected with tuberculosis. Newly purchased animals are clinically examined and tested with tuberculin, and are added to the healthy herd only when the results are entirely satisfactory. The male animals which are to be used for breeding purposes should not react to the tuberculin test. Under unavoidable cir- cumstances, a reacting bull may be reserved for breeding pur- poses but only under special precautionary measures. The results of Bang's eradication method, if carefully carried out, are remarkably satisfactory. It has been adopted to the greatest extent in Denmark, Sweden and Norway, and it has also been successfully carried out in Hun- gary and Finland. The report of Regner, in 1911, affords a good review of the results of Bang's method, and in it are described the results of the governmental eradication of tuberculosis in Sweden. Regner di- vides the eradication work into an offensive one in herds in which the disease prevails, and into a defensive procedure whose purpose is the prevention of the introduction of diseased animals into herds free of tuberculosis. Of the groups into which Regner separates the herds and the animals, the first group includes those which originally (on the first tuberculin test which in some instances was applied years previously) were found tuberculous. At that time 16,852 animals had been tested with a percentage of 30 . 2 reactors. In 1908, 18,719 animals in 457 herds proved to be entirely free from tuberculosis. % The herds of the second group, which proved to be tuberculous 120 Tuberculosis. at the time of the inauguration of the method and which continued to contain reacting animals, included 375 herds of 21,899 animals, with 41.5% of reactors. At the end of 1908 the number of cattle had increased to 26,181, of which only 1,496, or 5.7% reacted. The results were not so pronounced when the reacting animals were retained with the healthy animals, when cattle without the necessary precautionary measures were placed in herds free of tuberculosis, when animals which had not reacted in the old herd were removed into the free herd without being previously tested, or when an opportunity was given for the transmission of the in- fection by a reacting bull causing the infection in the herd to ap- pear to be renewed. Also in cases when the milk used for the feed- ing of calves was not free from tubercle bacilli, the results were unsatisfactory. In the interest of systematic eradication, it is necessary, espe- cially at the commencement of the eradication work, to subject the animals to the tuberculin test quite frequently, with short intervals. As a third group Regner included 436 herds containing 7,835 animals at the beginning of the work and 9,114 cattle in 1908, which at the first examination, and again in 1908 were free from reacting animals. The fourth group contains the herds which originally were free from tuberculosis but were not so at the test in 1908. The 98 herds included at first 2,526 and in 1908, 3,720 animals, of which 265 or 7.1% reacted. Regner concludes from his tabulations : that on the first tuber- culin test in 1366 herds, out of 49,112 animals tested, 14,175 or 28.9% reacted; that in 1909 the same herds contained 57,734 ani- mals, of which 1761, or 3.1% reacted; that Bang's method is the strongest factor in the general promotion of breeding, and of stable and milk hygiene. In other countries the results were similarly favorable. Bang succeeded in Denmark, from 1893 to 1908, in gradually reducing the percentage of reacting animals from 40 to 8 . 5. Malm in Norway from 1896 to 1903 reduced the disease from 8.4 to 4.9%. Hojer in Finland in 1894 to 1900 caused the infection to drop from 24 to 10.1%. Hutyra reports on experiments carried out on the government farm of Mezohegyes. In this herd the first tuberculin test in 1898 showed 44.8% of reactors out of 329 cows or 26.6% of the entire herd (647 animals), whereas in the fall of 1903 out of 502 cows only 2.8%, and out of the total of 1,132 animals only 1.8% reacted to the tuberculin test. The herd had been increased in this period by 75%, without purchasing additions to it, and the percentage of reactions had dropped 88%. The stringent measures of Bang have been somewhat modi- fied in certain cases for economic reasons, or when the strict execu- tion of Bang's method has presented peculiar difficulties. On the Ostertag's Method. other hand the requirements have been accentuated in cases where favorable considerations prevailed. Thus for instance in a herd in which only a few animals react it would be advisable to dispose of them without further consideration, and after a thorough disinfec- tion of the stable the defensive work against tuberculosis may be instituted, through the introduction of only non-reacting cattle, and by the disposing of all animals which prove tuberculous on the following tuberculin tests. The Siedamgrotzky-Ostertag method consists of immediate disposition of all animals with open tuberculosis (by this means the animals eliminating tubercle bacilli are excluded), and in bringing up the calves free of tuberculosis by feeding them with pasteurized milk or with milk from healthy cows. The calves are subjected to the tuberculin test after they are weaned, and the re- acting animals are not bred. The herds which are included in this method of eradication are subjected semi-annually to a clinical examination, and the clinically suspected animals are removed and disposed of. Further than this, the mixed milk of the herd, as well as the suspicious secretions and excretions are examined bacteriologically. The results of the Ostertag eradication method of course can not be compared with that of Bang. Since there are retained in the herd all tuberculous animals which show no clinical form of tuberculosis, or in which there is a suspicion of open tuberculosis but whose secretions and excretions fail to reveal the presence of tubercle bacilli. Therefore a constant danger of infection for the animals free of the disease prevails, as tuberculosis may at any time develop into an open form. But since it is required that the calves should be brought up free of tuberculosis, and that the elim- inators of tubercle bacilli should be determined by periodical clin- ical examinations as well as by the testing of the entire mixed milk of the herd and the individual secretions and excretions of sus- pected animals, Ostertag has obtained relatively very good results, where his requirements have been conscientiously carried out. This method has an advantage in that the stock owners who offer great objections to radical methods of eradication on ac- count of the immediate economic losses which they entail, are will- ing to work intelligently and with pleasure with a system of eradi- cation such as is offered by Ostertag's method. This assertion is best proven by the tabulation of Bautmann, which shows the in- creasing popularity of this method. The method was voluntarily adopted in the fol- lowing cases: 1903-1904, 1,457 animals; 1904-1905, 1,372; 1905-1906, 5,333; 1906-1907, 5,395; 1907-1908, 5.193; 1908-1909, 8,839; 1909-1910, 18,822; and 1910-1911, 19,828 animals. The following data illustrate the results obtained with the method: Open tuberculosis was present in the province of: East Prussia n 1900 in 2.7 % out of 10900 examinations East Prussia ; • n 1904 in 1.3 % out of 17500 examinations Pommerania n 1902 in 2.93% out of 8808 examinations Pommerania n 1906 in 0.6 % out of 22356 examinations Brandenburg m 1903 in 3.46% out of 5200 examinations 122 Tuberculosis. Brandenburg in 1907 in 1.5 % out of 5810 examinations Schleswig-Holstein in 1903 in 2.8.% out of 2435 examinations Schleswig-Holstein in 1905-6 in 1.93% out of 11000 examinations Saxony in 1903 in 3.6 % out of 1457 examinations Saxony in 1906-7 in 2.41% out of 5395 examinations In these statistics it should be considered that every year new, unexamined herds have been included, and further that the experts continually gained more skill in making the examination. A proof of the reduction of tlie dangerous forms of tuber- culosis is first of all indicated by the above figures, and also by the marked reduction of tuberculosis of hogs. Thus for instance ac- cording to Stier the percentage of tuberculous creamery hogs which amounted to 40% was reduced to 4% after the elimination of six cattle with tuberculosis of the udder, although none of the skimmed milk fed to the hogs had been sterilized. In 1907 out of 38,454 animals examined in Schleswig, 1.4% were found to be affected with pulmonary and udder tuberculosis. Udder tuberculosis alone was demonstrated in 0.124%. In spite of the great advantages of the method, the results eventually come to a standstill, as may be seen from the more recent reports of the eradication stations. The number of the dangerous forms cannot be reduced below a certain percentage, since latent forms contin- uously change into the dangerous forms, and it is therefore im- possible to eliminate the sources of infection from the herds. The method of Ujhelyi also deserves mention. In this method the cattle are divided into a healthy herd and those which react to tuberculin. The newly born animals of the group giving a positive tuberculin reaction are allowed to remain only in emergency cases with the reacting mothers, but if possible they are nursed by healthy cows. After weaning the calves are tested with tuberculin. This method differs from Bang's method only in that sterilized milk is not used (prevention of calf diarrhea) and further the calves are allowed at times to remain with the reacting cows. According to Ujhelyi 's report this method has given irreproachable results. It has to be considered however that of the weaned calves a greater proportion of animals must be eliminated when this method is employed, than when Bang's method is followed. Before the introduction of Ujhelyi 's method, out of 1,031 adult cattle, 884, or 85.7% reacted. Out of 626 young stock 333, or 53.2% reacted. After a period of eradication for 4% years Ujhelyi succeeded in reducing the infection to 4.1% among the adults, and 2.6% in the young stock. He succeeded in the periods from 1898 to 1902, and from 1904 to 1905, in reducing the number of positive reactions among 1,715 cattle, of eight state farms, from 59% to 3%. The eradication of tuberculosis has been .subjected to official control for several years in Denmark, Sweden, Norway and Fin- land. Thus Denmark in 1893 contributed $13,500.00, later $27,-- 000.00 towards the eradication of tuberculosis, furnished the tuber- culin free of charge at first for young animals, later for adults, and finally since 1898 took upon itself the total expenses of eradication (Hutyra). The skimmed milk is permitted to be returned from the creameries to the stock owners only after being heated to 80 deg. C. Cows affected with tuberculosis of the udcler are destroyed, the owners being reimbursed. There are about 600 such animals paid for annually. Similar results were obtained in Sweden, which adopted legis- lative measures and made a contribution of $225,000.00. In the Results of Control Work. years from 1897 to 1908 in 1,370 herds with 48,576 animals, of which 14,225 or 29.3% reacted, the amount of infection was re- duced by their work of eradication to such an extent that out of 57,660 cattle only 3.1%, that is 1,765 animals, reacted. Although the statistics of individual countries having strict measures of eradication (Denmark, Sweden, etc.) appear to show the splendid effects of carefully executed control work, based on scientific principles, and in spite of the fact that the milder modifi- cations, as for instance that of Ostertag, by no means show the same good results, nevertheless measures of too strict requirements cannot be absolutely approved. Thus for instance Belgium in 1895 required the destruction within a certain time of all clinically affected and all reacting animals, and in 1896 out of 19,004 cattle examined 9,280 were slaughtered. The difficulty of the execution was lessened by the law of 1897 which required that only the visibly affected animals should be destroyed, a measure which resulted in the destruction of 10,269 cattle with reimbursement amounting to $300,000.00, in 1902 (Hutyra and Marek). Theoretically, the most radical eradication measures may pos- sibly be considered as the quickest and most effective, and therefore from an economic standpoint as the best methods for the control of tuberculosis. Owing to the extraordinary spread of this dis- ease in almost all herds, drastic measures however may result in the sudden infliction of such heavy economic losses, not alone through the animals destroyed, but through changes of values for breeding, dairy purposes, meat production, etc., that the stock owners, dealers, consumers, etc., would have good grounds to pro- test against the execution of such methods. Therefore it is advisable to adopt Ostertag's method at the ini- tiation of the general work of eradication, and after the stock own- ers have been convinced that the idea is rational following the favorable practical results obtained, then Bang's method may be introduced, unless it is possible to persuade them to employ, at the beginning, the rational execution of Bang's method. In no instance however should destruction of the reacting animals be required in connection with Bang's method. From the standpoint of milk hygiene it does not seem to be justifiable, according to the present status of the question of the infectiousness of the milk of reacting animals, to require their exclusion from the production of milk, unless they show clinical evidence of the disease. In spite of the separation of the reacting from the non-react- ing animals, the milk of the reacting group could be marketed, from the standpoint of milk hygiene, with the milk of the other group, without interference, as has been previously practiced, so long as there is no substantial proof offered as to the danger of marketing such milk. With the new law on diseases of animals the initiation of eradication, based on uniform legislative measures, has been in- stituted in Germany, and thereby serves as a stimulus to extensive private activity in matters of eradication. 124 Tuberculosis. The law requires that clinically affected tuberculous animals, or those in which tuberculosis probably exists to a great extent, may be ordered destroyed by the police authorities. If this is not carried out, or if the destruction is postponed, sanitary police protective measures should be inaugurated against further spread of the disease, by branding the animals. The police measures against the spread of the disease con- sist in separation, observation of police control of the affected, sus- pected and susceptible animals; if necessary restriction of traffic of both man and animals, and special limitations relative to the use of affected or suspected animals, and their carcasses, and finally the usual requirements of disinfection. For animals which are destroyed by the requirements of the police, and those which after destruction has been ordered, die of the disease on account of which they had been ordered destroyed, the government allows corresponding reimbursement. Of great importance in tuberculosis eradication is the require- ment prohibiting the return of skimmed milk and other milk residue to the milk producers, as food for other animals, unless the same has been heated. This clause is included in the general requirements of the measure. Centrifugal slime, which has formerly caused the development of ingestion tuberculosis in hogs, must be destroyed by burning or burying. The measures differentiate three danger classes in tuberculo- sis: (1) the simple suspicion, (2) the great probability of its presence, and (3) the actual existence of the disease. In the pres- ence of the clinically recognizable classes of tuberculosis, it is required that the milk from such affected animals should not be sold or otherwise utilized without being previously subjected to a required temperature for a certain length of time. The milk from cows affected with tuberculosis of the udder cannot be used for human consumption even after subjecting it to the required heat, nor can it be utilized for the preparation of dairy products. The requirements in Bavaria order the destruction of an ani- mal only when it belongs to a herd in which cattle breeding, or raising of cattle is industrially followed, and an appropriate volun- tary method of eradication of cattle tuberculosis may then be carried out in the herd under veterinary supervision. This ought to result in a considerable improvement of the tu- berculosis question, and with the elimination of animals which prin- cipally enter into consideration as distributors of bacilli, a point is gained which temporarily should thoroughly satisfy even the milk hygienists. With such measures the stock owner is pleased, as the pro- fessional direction of rational breeding in connection with eradica- tion is shown to be for his advantage. This constitutes the basis Bacillus Pyogenes. on which the entire milk production in all its relations may be ele- vated, and will be elevated, since the voluntary intelligent co-opera- tion of the owners constitutes the fundamental principle on which the state bases its allowance of reimbursement. Without the voluntary co-operation of the producers, the elevation of milk hygiene is practically impossible. It will take years before the conditions will markedly improve, but the improvements will surely come, and they will not confine themselves alone to the tuberculosis question. Other Forms of Mastitis. The other forms of chronic mastitis, with the exception of tuberculosis of the udder, are of slight importance for practical purposes when compared with streptococcic mastitis. Thus for instance the mastitis produced by the Bacillus pyog- enes bovis is relatively rare, and the author has had the oppor- tunity on only three occasions to attribute the development of chronic mastitis to the Bacillus pyogenes. Glage, Nielsen, Kuhl- mann, and Sven Wall, however, have observed the infection fre- quently, and even describe an epizootic extension of the infection. Mixed infections of staphylococci and colon bacteria, with the Bacillus pyogenes, appear to be more frequent and in these cases a severe mastitis is produced. It results in abscess formation and necrosis of the affected parts, with an induration of the tissues. The secretion is sanio-purulent, and mostly of an offensive odor. Kiinnemann found the bacillus at first in suppurations of cattle, and Grips in suppurating processes of hogs. They are small, deli- cate rods of the size of the swine erysipelas bacillus, growing bet- ter anaerobically than aerobically, forming dew-drop like colonies on agar, or serum agar. Milk coagulates to a uniform clot. The bacillus does not take the Gram stain, but it may be stained by Weigert's method. Very little is known relative to the behavior of the Bacillus pyogenes bovis towards man. According to the author's observa- tions it appears to belong to the group of pseudo-influenza bacilli (Pfeiffer). Such rods were found in influenza-like pneumonias, in bronchitis (Pfeiffer), in suppuration of the middle ear (Kossel, Hartmann, Pielicke and Cantani), also in whooping cough (Afan- asieff, Szewetschenko, Wendt and others). Friedberger discovered a similar rod in the mucus of the prepuce of a dog. Frank describes it in the pus of a hog. Frosch found it in the blood of geese, and Beck in an infectious pneumonia of rabbits. It belongs to a widely spread bacterial group. The Bacillus pyogenes is non-pathogenic for small, experimen- tal animals and pigeons. Since rods similar to those of the Bacillus pyogenes have been found in man it is not impossible that affections of man may be produced by milk from udders with this form of infection. 126 Mastitis. It has not yet been possible to demonstrate this bacillus in mixed milk, since there occur too many bacteria of similar mor- phology in stable manure, in the air, and under the epithelia of the teats. At any rate the milk must be considered as spoiled when it contains secretion from udders with pyogenic infections, and should be excluded from the market, since its injurious effects upon health seem to have been demonstrated in the sense of the pure food act. Infections with bacteria of the coli-typhus group frequently occur when the cows are kept in filthy condition, with unclean bed- ding, and also when manipulations are undertaken by milkers in order to dilate the milk ducts (penetration with straws, quills, and CHiltures of Bacillus paratyphus. 1 X 800. (After Kitt.) contaminated milking tubes). Representatives of this group of organisms were described by Jensen and Streit, Guillebeau, Kitt, Freudenreich, Lucet, Sven Wall and Weicliel, as the cause of high- ly acute forms of mastitis. The bacteria are short rods with round- ed ends, mostly motile; they do not take Gram's stain. Accord- ing to their biologic characteristics various varieties may be dis- tinguished, which at times approach more closely to the colon group, at other times more to the aerogenes group, and at times even to the enteritidis group, which cause meat poisoning. Milk becomes coagulated with gas formation. The acidification and coagulation occur earlier with some varieties than with others. The colon group always ferments galactose, glucose, laevulose, mannose, lactose, maltose, arabinose, rhamnose, xylose, mannit and sorbit, frequently also sorbose, saccharose, raffinose and dulzit, but not Causes of Mastitis. erythrit and adonit. Their action is different towards saccharose, raffinofe, sorbose and dulzit, and this differentiation is utilized to separate the groups into those which do not attack any of the mentioned bodies, those which ferment all four, those which split up dub it and sorbose, and finally those splitting up saccharose and raffinose. Of course bacteria cannot be strictly separated by their fermentative action, since in the cultivation of colon strains in sugar-containing media they may acquire the faculty of fermenting a kind of sugar towards which they formerly were refractory (Twort, Massini). Through the first group the colon bacteria approach the more dangerous group of Bacillus para- typhus B., Bacillus enteritidis of Gartner and paracolon bacteria with their related organisms, for instance the Bacillus raiin. Bacillus suipesiifer, B. typhi murium, etc. These dangerous groups may be separated by agglutination into three classes, the Bacillus enteriiidis Gartner group, the Bacillus paratyphus B. group, and finally the Paracolon group. Weichel succeeded in isolating from two cases of severe sep- tic mastitis an organism belonging to the group of Bacillus enteri- tidis, and another to the Bacillus paratyplius-B. Excluding the inflammatory products which may also possess disease-producing properties in this group of mastitis forms, it is necessary to exercise special care in the inflammations of the ud- der caused by the Coli-eiiteritidis-paratyplius-paracoli organisms, since among the representatives of this group of bacteria there are those which may produce severe forms of enteritis in man, with symptoms of poisoning, which are known in general as meat poi- soning. True paratyphus bacteria may also enter the milk in other ways than with the secretion of an infected quarter, for instance through bacilli-carriers who are employed for handling the milk, through the rinsing water, and also from other sources. It will be of interest to mention here the results of examinations of market milk for the occurrence of Bacillus paratyphus-B. Uhlenhuth and Hiibener twice found paratyphus in 100 sam- y>les, while Hiibener in 40 samples of market milk noted this bacil- lus 4 times and in 30 other samples of market milk, observed it 3 times. Klein in 39 mixed milk samples found the Bacillus enteritidis 9 times. The occurrence of coli-aerogenes bacteria in milk would be something very ordinary, and would be considered less injurious for the health than the presence of varieties which are known as toxin producers. Nevertheless the coli-aerogenes infections of the udder should be considered with the greatest care ; although in general the enter- itidis and paratyphus varieties produce severe septic inflammations with ichorous secretions and frequently with a fatal termination, the severity of mastitis and the appearance of the secretion are by no means a certain indication of the character of the infection. Mixed milk to which the secretion from animals with acute af- fections of the udder has been added, is spoiled according to the pure food act, and should be considered as capable of injuring human health. The milk of healthy quarters from such infected udders is also 128 Mastitis. suspicious of being contaminated with the infective agents, and therefore should be prohibited from entering the market. According to Weichel reports on paratyphus and enteritidis infections which may be traced to milk are rare, and no publications can be found which absolutely trace affections in man to a coli- paratyphus mastitis. The Dairy Journal of Berlin reported in 1900 that according to "Dag. Nyheter" nine families in Stockholm became affected with symptoms of meat poisoning (fever, depression, fainting spells, nausea, vomiting, diarrhea, muscular cramps). The milk, to the consumption of which the affection was traced, originated from 14 cows, one of which suffered from an inflammation of the udder. In the secretion of the affected udder the same bacteria were found as in the feces of the affected people. Two female attendants of the stable from which the injurious milk was obtained also became affected with similar symptoms. The observation of Moro also belongs here. Moro observed in six persons after the consumption of milk from a goat suffering with a gangrenous inflammation of the udder, chills, nausea, headaches, and 11 hours later colic, vomiting and thirst. The milk was consumed mixed with coffee. Weigmann and Gruber report a case of injurious effect (vomiting), from cream which had been prepared from mastitis milk, and they traced the affection to a bacillus of the colon group (immobilis). Weichel fed a six-weeks old dog with the milk of a goat which was artifically infected in the udder with a paratyphus strain from septic mastitis. The feeding was undertaken after the appearance of the mastitis in the goat. Three hours after the consumption of 200 c. c. of the secretion the dog showed marked symptoms of restlessness and barked frequently; lachrymation and later repeated vomiting appeared. He soon recovered but refused to partake again of this milk. Only after 60 c. c. of this fluid had been mixed with 200 c. c. of good milk would he touch it ; he then took a small quantity but with apparent distaste. Within five minutes he showed pain, and manifested similar symptoms as the day before, but again recovered before the following day. In a second feeding experiment on a seven-weeks old dachshund the affection commenced only on the fifth day of the experiment. The animal became listless, refused food, whined, and in addition lachrymation, nasal discharge, and periodical chills appeared. This dog also recovered on the second day. Weichel also reports a case in which the wife and daughter of a dairyman became affected with a diarrhea after the ingestion of inflammatory products of a cow with coli-mastitis. The dog of the owner also showed similar symptoms after drinking the milk. As milk offers very favorable conditions for the multiplication of bacteria of this group, the danger from milk containing coli- paratyphus bacteria must be considered greater than in the case of meat bearing the same infection. Various data exist relative to the resistance of these bacteria towards influences of heat. According to Fischer heating to 60 desr. C. for a half hour does not suffice to kill all paratyphus germs ; likewise some of the bac- teria remained active after heating the milk for 10 to 35 minutes at 70 deg. or for five minutes at 75 deg. C. Although Kolle states that the typhoid, paratyphoid, and enteritidis bacteria are without exception destroyed when sub- jected to a temperature of 59 deg. C. for 10 minutes, nevertheless it must be remembered that the conditions in milk are markedly dif- ferent than in suspensions of culture, and that some of the varieties are capable of producing a heat-resisting toxin. According to Gart- ner the toxins of the meat-poisoning organisms withstand 100 and Mixed Infections of Udder. even 120 deg. C. These facts were confirmed by Van Ermengem, Drigalski, Fischer, Hoffmann, Peels, Hoist, Dhant, Riemer, and others. In practice therefore it is necessary to consider the mixed milk of the affected cow and all dairy milk to which such milk has been added as injurious to health, whenever it is proven with certainty that it contains secretion from acutely affected quarters. If it is proven with certainty that the secretion contained bac- teria of the paratyphoid or enteritidis group such milk may even destroy human health. Of course the danger which threatens man from such milk must not be overestimated. The changes in the udder and in the milk are pronounced and striking, and usually appear very sudden- ly, especially in the colon inflammations, somewhat less in para- typhoid and enteritidis infections. Nevertheless in the presence of carelessness of the milker such infections may enter the milk. According to Fauss the duration of the elimination of the bacteria from affected udders persists for 12 to 30 days, in fatal cases until death. The number of the eliminated bacteria and the duration of the elimination are proportional to the severity of the case, and they cease when the milk again approaches its normal condition. In other cases of acute mastitis staphylococci have been dem- onstrated. Guillebeau isolated the Stapliylococcus mastitidis, Galactococcus versicolor, Galactococcus fulvus, and Galactococcus albus. Experimentally it is also possible, as proven by Kitt with the Botryococcus ascoformans (a staphylococcus), to produce an acute mastitis, with a tendency to chronic development. The staphylococci infections of the parenchyma of the udder are relatively rare, but occur more frequently as mixed infections with the Bacillus pyogenes. While the course of the staphylomyco- sis of the udder is mostly acute, with a favorable progno- sis, yet in the presence of a mixed infection with the Bacillus pyog- enes it frequently results in abscess formation and sequestration of the udder. The staphylococci are small round microbes, separated into two or four parts by division. They take the Gram staining. They are easily cultivated on all media and are frequently chromogenic. They liquefy gelatin from the surface down, since they grow better aerobically than anaerobically. Staphylococci corresponding to their ubiquitous distribution are present in almost all milk during its first phases of decomposi- tion ; but although they possess pathogenic importance as pus-pro- ducers in man, from the standpoint of market milk hygiene, they are of no special importance under such conditions. However when the secretion of a cow with staphylomycosis of the udder contains staphylococci, the milk may be injurious to health. Karlinski, for instance, reports a case of pyemia in a child in which infection 130 Mastitis. resulted from the milk of the mother containing staphylococci. At least the cocci which Karlinski isolated from the milk, and from f eces and blood from the child were identical. The secretion there- fore must be considered as spoiled food, and must be excluded from consumption. The same judgment as stated for staphylococci infections also applies to botryomycosis of the udder. This represents a chronic form of a staphylomycosis, in which the single cocci that grow in colonies are compressed by swelling of the cocci lying on the out- er borders, forming capsulated spherical colonies. The central cocci continue to grow, burst the capsules, and the process of the swelling of the bordering zone is renewed until mulberry-like fun- goid colonies result. A method of distinguishing Botryococcus ascoformans from staphylococci has not yet been devised. The botryomycotic formations develop mostly in the horse which is probably proof of certain immunity strength of the horse (that is of equidia), against staphylococci infections. In other animals and also in cattle the disease is extremely rare. Mohler, Czokov, Immel- mann, and Eeinhardt have observed botryomycosis in the udder of cows. Botryomycosis in cattle is of no practical inportance in the judgment of milk. Actinomycosis of the udder is also of slight importance from a practical standpoint. The purulent fibroplastic actinomycotic mastitis occurs in cattle with less frequency than the actinomycosis of other organs. It has been described by Kasmussen (four times), Jensen (20 times), Maxwell (once), Bang, and Johne, and represents a chronic suppuration with nodular cicatrization of the udder. After the in- fection, nodules from a bean to a hen's egg in size, with softened centers and fibrous borders develop, or a diffuse inflammation with a tendency toward cicatrization and hardening of the entire udder results. Actinomycosis of the udder may be primary (McPhail, Williamson) and develop from the introduction of barley beards into the tissue, or possibly from pasturing on stubble fields, or again it may develop by metastatic formations from other lesions in the body. McPhail believes that some cases of so-called udder tuberculo- sis are in reality actinomycotic infections of the udder. Should an actinomycotic process soften in the udder and the abscess burst into the secreting tissue, the finding of actinomyces in the milk is possible. The latter appear as colonies of ray-like fungi (streptothrix). The branching threads form a mesh-like mycelium with spherical or club-shaped enlargements on the end of the threads. The fungous threads proliferating in the animal tissue are influenced by the action of the body fluids. The sheaths swell and club-shaped bodies result, arranged in a radiating man- ner, which later become adherent to each other forming rosettes in which the mycelium, protected from the immune bodies and leuco- cytes continues to proliferate or to degenerate and calcify. Actinomycosis. The actinomyces are widely spread forms of the higher bac- teria with true branching, and stand between the lower bacteria and hyphomycetes. They almost invariably occur on grain, hay, straw, fruit, manure, soil, flour and milk. Most of the actinomyces are harmless provided a foreign body does not facilitate their colr onization in the animal body. Splinters of wood, and especially beards of barley are frequently the carriers of the infection. Transmission from man to man or from animal to man is not known up to the present time. The basis of an infection always lies in wound infection either through the above mentioned for- eign bodies, or by the fungi gradually becoming accustomed to exis- tence in necrotic tissue of the animal body (caries of teeth). Johne succeeded in producing actinomycosis of the udder through the injection of actinomycotic cultures into the milk cistern. Although transmission to man through milk from actinomy- cotic udders is not to be feared, prohibition of the sale of such milk is required since it must be considered as spoiled on account of the presence of pus and other associated changes. Contrary to actinomycosis, "actinobacillosis" first described by Lignieres and Spitz in Argentina, and which clinically resem- bles actinomycosis, is of a contagious nature. Therefore although actinobacillosis has not yet been described in man it should be more carefully judged than actinomycosis. In Germany cases of actin- omycosis have been reported which from the bacteriological find- ings, should be classed as actinobacillosis, and these cases occur sometimes in an enzootic form or as stable outbreaks. Thus Imminger in Oberpfalz and Preusse in Western Prussia, described an enzootic extension of actinomycosis, and Schulze mentioned a case in which the disease affected most of the animals in the stable (of 30 steers 27 were affected) . Of 87 newly purchased animals more than half of those placed in the stable became af- fected, while 12, which had been stabled on other premises and which received the same feed, remained healthy. Milk from udders affected with actinobadllosis, and mixed milk which contains such secretion must be considered as spoiled and prohibited from consumption. Mixed infections of the udder with these described bacteria and others, should be similarly judged, and likewise infections with malignant edema bacteria, Bacillus necrophorus, etc. CHAPTER VII. EXTERNAL INFLUENCES WHICH ACT UPON MILK. (a) Their effect upon the body, thereby influencing milk secretion ; (b) Their effect upon milk after its secretion. Although our knowledge, relative to the development of the individual components of milk from the substances in the blood, scarcely extends beyond the border of hypothesis, nevertheless it is established that milk formation is dependent to a certain extent upon the feeding, although only within limits denned by the breed, family, individual, lactation period and age. Through outside conditions, those factors of production are especially influenced, which are themselves subject to variations, especially the quantity of milk and fat content, less so the proteid and sugar content, and only very slightly the salt content. The influence of feeding could be explained by reasoning that the gland increases its activity at the moment in which a larger quantity of nutritive substances circulates in the blood, after the ingestion of large quantities of easily digested food. This sup- position could be even enlarged upon by considering that the activ- ity of the cell is stimulated by specific substances in the food in such a way that it assimilates to better advantage and in increased quantities the necessary constituents which it draws from the blood. From practical experience and scientific experiments it must be considered as established that the milk produced is dependent both in quantity and quality, upon the quantity of digestible food and on the presence of specific substances which stimulate milk formation. This view has been accepted for a long time by practical dairymen,' who for instance have observed that clover hay, in spite of its greater nutritive contents has not come up to the value of good meadow hay; that meadow hay cannot be replaced by a mixture of straw and concentrated food mixed in a way to make its nutritive value equal to the meadow hay; further that sweet hay proves a better milk producer than sour hay with equal nutritive value, etc. The value of individual pastures also shows wide differences in the production of milk, although examination of the grasses of the pastures gives similar results. In these investigations however it was found that beyond certain limits the influence of nutrition was no longer usable, and that with sufficient feeding of wholesome and tasty food no influence could be exerted upon production through increased rations. 132 Effect of Feed on Yield. 133 If animals are allowed to starve, the change in the quality of the milk will result only after the reserve deposits of the body have been utilized to their fullest extent, or completely exhausted. In the state of starvation the milk fat shows an approach in its composi- tion to that of the body fat. If experiments are started with starving animals, or with animals which only receive small rations, the milk yield, according to Kellner, increases with the added increase of feed. Such cows after an increase of rations yielded: With an increase of 1.5 kg. bean bran, the increase of milk amounted to 0.92 and 0.53 kg. With 3 kg. bean bran, the increase was 2 . 40 and 1 . 01 kg. With 1 kg. malt, the increase was 0.84 and 0.3 kg. With 2 kg. malt bran, the increase was 1.31 and 0.40 kg. The increase in yield however was not parallel with the in- crease of the ration, but the closer the quantity of milk produced approached the maximal production of the individual, the slighter became the increase in yield. In attempts to increase the produc- tion of the cow, the last liter of milk is the most expensive. It re- quires for its production the largest addition of rations. In general it may be said that sufficient quantities of digesti- ble proteins are the fundamental requirements for normal milk production, and that although other food substances are present in sufficient quantities the yield of milk diminishes rapidly when the protein content is decreased below the amount necessary for main- taining the body weight. For 1000 kg. of body weight 1.212 kg. of digestible proteins must be figured, together with a sufficient addi- tion of fat and carbohydrates. Fat and carbohydrates and non- protein nitrogenous substances in sufficient quantities act as econo- mizers of proteins. For continuous milk production an excess of about 0.40 to 0.55 to 0.65 kg. of digestible proteins is required for 10 kg. of milk (Schmeck and Kellner). Experiments of Morgen and Fingerling proved that while feeding tasteless non-stimulating food consisting of straw, cut straw, starch and oil, the yield of milk may be increased by the ad- dition of substances which by themselves cannot be utilized in the production of milk, and therefore they are considered stimulating substances which principally stimulate the gland to activity. Increased Increased Addition yield yield in milk in fat Malt 0 gm. 0 gm. Buckhorn seed 10 gm. 0.8 gm. Hay distillate and fennel 2.03 gm. 7.9 gm. 153 gm. 5.6 gm. 312 gm. 9.8 gm. 109 gm. 6.7 gm. The percentage of fat increased 0.25 to 0.32%. 134 Effect of External Influences. In order to produce an increase of milk it was sufficient to introduce into the food small amounts of fennel, or to sprinkle it with distillate of hay. If in the experiments good meadow hay was fed in sufficient quantities with other food-stuffs, the addition of the stimulating substances was without effect. These observations are of special importance in view of the swindles carried on with milk powders, by which money is still extorted from the farmers. Through the addition of salt to tasteless food an increased yield in milk and fat was obtained amounting to from 20.6 to 21.9%. Hansen reports on the influence of concentrated foods on the milk yield, which he investigated extensively for seven years. This author divides the concentrated food into four groups : 1. Foods which increase the milk yield and diminish the percentage of fat, as for instance farina, corn, oats, and possibly also soja beans. 2. Those which do not influence the quantity of milk but increase the fat content, as for instance palm-seed oil and cocoanut oil cakes (the specific action of cotton-seed meal is less pronounced) peanut meal, corn-slop and bread flour. 3. Those which do not change the yield of milk but reduce the fat content, as for instance, poppy seeds, flour of rice and other concentrated food, beneficial for fattening, as for instance cake of sesame (Ramm). 4. Those which have no specific action, as for instance wheat bran, and malt sprouts. Such food substances are especially de- sirable for the use of fattening dairy cattle. From the experiments of Hansen it appears as a matter of fact, that certain food substances possess a specific action. In this regard the composition of the food is of course of importance, since the action of a certain food may be checked by feeding coun- teracting substances. It has long been known from practical ex- perience that the quality of milk may be greatly influenced through the method of feeding, and not only as far as the constituents of the milk are concerned but also its odor, taste, etc. To what extent food bacteria play a part in this, will be discussed in the chapter devoted to that subject. Summer butter, mountain butter, and stable butter, are richer in fatty acids with low molecular weight, than fall butter or butter from cows which have been kept on low land pastures, or pasture butter in general prepared in the same manner. Feed rich in carbohydrates produces a soft milk fat. If abnormal constituents of fats are artificially added in experimental feeding, or if fats are fed which are otherwise not found in the body, such constituents are again found in the milk, for instance sesame oil (Engel), linseed oil, hemp seed oil (Gogitidse), iodin and iodipin (Caspari and Winternitz), Sudan III, a specific fat coloring matter (Gogitidse). According to Schrodt and Hansen pasture milk on account of its greater contents of casein, contains more phosphoric acid than stable milk, which on the other hand is richer in chlorin. Accord- ing to Sanson, Hesse and Schaffer the feeding of phosphate also increases the content of phosphoric acid ; this however according Feed Recommended. to Neumann is not immediate but appears only after weeks and then in insignificant proportions. Jensen succeeded in finding only an insignificant influence on the milk from the feeding of considera- ble amounts of lactates of iron, calcium sulphate, disodium phos- phate, dicalcium phosphate, dimagnesium phosphate, potassium chloride, chloride of sodium, and nitrate of potassium. Nitrates appeared in the milk only after 75 gm. of saltpetre had been fed. The administration of from 30 to 40 gm. of saltpetre failed to result in the^presence of nitrates in the milk. The salt content of milk therefore changes only insignificantly provided normal conditions are present. According to Henseval and Mullie, the health of the animals plays a part when salts pass into the milk. If these authors fed from 5 to 25 gm. of saltpetre to 20 healthy and 8 diseased animals, the milk of the sick animals always contained nitrates, whereas the milk of the healthy animals did so only exceptionally. Definite quantities of sulphuric acid are supposed to occur in milk after the administration of Glauber salts. Of the various foods, meadow grass, green clover, rowen, green alfalfa, and peas in which a large amount of young grain has been sown are recommended, for instance, vetch with oats, barley or rye, plants of the white mustard, rape, sainfoin, Kohlrabi tur- nips, etc., with which oats, barley or rye have been grown. Fod- der or straw should be mixed with the green feed. In the winter instead of green feed, mangels, chopped roots, ensilage, grain, potato slop and corn slop should be fed. Relative to the injurious effects of the various bacteria found in feeds and pastures, see the chapter on milk abnormalities. Good hay and good fodder may be recommended as dry feed. As con- centrated food the substances mentioned by Hansen as indifferent, or those food substances of the first and second group which are recognized as milk and fat producers, will be found satisfactory. Rough fibrous foods cause a loss of energy, and are not well utilized on account of the increased work of mastication and be- cause the intestines are too greatly burdened by this feed. Individ- ual feeding according to the milk yield appears advisable, and the best milkers may be allowed additional rations corresponding to their heavy production. In this regard of course the yield and quality of the milk should be established by sample milkings and examination of the secretion. In cow-fattening dairies the fatten- ing of the animals should commence only in the last three months since fattening foods and fattening of the animals diminish the yield of the milk. Pure drinking water has a great influence on milk produc- tion, and the animals should be enabled to partake of it freely ac- cording to their needs. Heyken mentions a case in which each cow yielded one-half liter of milk per day more, when instead of hard marshy spring water containing iron, good well water was sub- 136 Effect of External Influences. stituted. Backhaus observed an increase of milk and fat con- tent after the introduction of an automatic water supply. Milk of poor quality is known to have resulted from the use of poor drinking water. Stagnant waters give the milk a repulsive taste. Taken as a whole all foods and all food mixtures which are par- taken of and digested by the animals without disturbance in their general condition are adapted to the feeding of milk animals. Food which in continuous feeding causes diarrhea or other intestinal disturbances should be avoided. Intestinal disturbances which quickly subside and which sometimes develop as a result of sudden change of food are of no consequence in the judgment of the food. They may cause considerable fluctuation in the yield of milk and fat content, which however subsides in a few days. From the above statement it will be seen that under certain conditions, espe- cially when a heavy production of cream is necessary, the effects of a change of food must be considered. If, when considering evidence of adulteration, methods of examination are used which take note of the approximately constant factors in milk, that is, such as pertain to the protein-free milk serum, the influence of a feeding method, or a sudden change of feed should be taken into consideration in regard to its effect upon the milk of each indi- vidual animal. Spoiled food injures the taste and odor of the milk and butter, and its effects may last for a long period after the time of feeding such food. The feeding of large quantities of beet or tur- nip tops should be guarded against, likewise over feeding with fresh or sour chopped roots, potato slops, residues from starch factories, brewer's grains, rape seed cake, flaxseed meal and poor straw. The taste is improved by feeding on pasturage, red clover, meadow grass, carrots, oats and rice flour. Firm tallowy butter is derived from grass of acid soil, from grass from fall pastures, late hay made from sour grasses, leaves of sugar-beets, or red beets, chopped roots, potatoes, peas, palm seeds, cocoanut and flaxseed cake, and cotton-seed meal. Soft butter results from the feeding of oat hulls, corn bran, wheat bran, rice flour, rape-seed cake and sunflower-seed cake. Clover pastures are not suitable for the production of milk for cheese making since the cheese becomes permeated with small holes, and has a sharp repulsive odor. This condition is probably brought about by bacteria which vegetate on the clover plants of the pasture. Changes from one feed to another should not be made too sud- denly if it is desired to prevent an effect on the milk production. Newly introduced food substances should not be fed in large quantities at first. In changing from dry food to pastures a dimin- ished milk yield first results, then a period of normal yield and Plants Affecting Milk. 137 finally an increase in the quantity, together with an improvement in quality. Pastures or green cultivated forage containing many buttercups should be avoided, since these plants are supposed to produce red and bitter milk, especially before blooming. Meadows or pastures in which Euphorbia plants are growing exert a bad influence; they may produce enteritis with a fetid diarrhea, also paralysis of the bladder and hematuria, and may even cause abor- tion. The milk turns thin and bluish. Bluish milk may also result after feeding upon plants of the Polygonum species, the ox tongue (Anchusa offic.), the cat's tail (Butomus umbellatus), the euphorbia (Mercurialis), the marsh marigold (Ehinanthus major), the forget-me-not (Myosotis), and after feeding upon poppy-cake and green alfalfa. Red milk is produced by feeding blood root (Galium verum), madder (Rubia tinctorum), species of Karex, Skirpus, Equisetum, Ranunculus, Euphorbia and after the ingestion of young sprouts of both deciduous and coniferous trees. Yellow milk results from the elimination of plant coloring matter after the feeding of carrots, rhubarb, yellow and red man- gels, and crocus. A garlicky taste may also result from feeding large quantities of poor straw, and according to Werenskiold after feeding of flax seed meal which contains large quantities of weed-seeds, penny- royal (Thlaspi arvense). The taste of the milk may also be changed by the ethereal oils of the following plants : Garlic (Alium ursinum), mint (Teukrium), hyssop (Gratiola offic.), true camomile (Matric. chamomilla), and by rape, rape- cake, oil cake, turnip tops, lupins and orchids. Milk may become fishy from feeding fish meal and through pasturing on marshy fields which have been inundated. Milk turns bitter from feeding kale, rutabagas, turnip tops, lupins, pea-straw, lupin straw, and sorrel. A bitter substance from chicory passes into the milk; the milk may coagulate more readily after the ingestion of thistles or sorrel. The ingestion of euphorbia, hellebore, rushes, and hemlock twigs should be prevented on account of the poisonous qualities of these plants. The secretion of the active poisons of these plants has, however, not been proved. Hop leaves, especially those sprayed with copper sulphate, cause a diminution of milk secre- tion, or even a cessation of the flow. Concerning the elimination of medicinal agents with the milk, or the influence of medicinal agents on milk production the follow- ing may be stated : The passage of iodine into the milk after feed- ing potassium iodide has been proved by Peligot and Stumpf ; if however the iodine is fed in alkaline compounds, or combined with proteins and starches, even when fed in large quantities, it does not pass into the milk. In the latter case only the plasma of the milk 138 Effect of External Influences. contains the halogen in the form of a salt, whereas in feeding iodized fats the milk fat contains iodine. According to Rosenhaupt and Bucura the same applies to bromine. According to Stumpf, Baum and Seeliger, in feeding com- pounds of lead small quantities (0.02%) of this substance pass into the milk. The ingestion of such milk was found harmless for ani- mals. The lead was eliminated for a longer period than the time during which it was fed. Milk which contains salts of lead could produce severe injurious effects if taken for a long time (chronic lead poisoning). Feeding of salts of copper results only in the appearance of traces of copper in the milk. The feeding of iron preparations does not to any noteworthy extent influence the contents of the milk. Mercury may pass into the milk (Bucura) ; likewise arsenic when administered per os or injected in any form subcutaneously (Bucura, Ittalie and Jesionek). Substances like aloes, senna leaves, rhubarb and croton may influence the milk in color and taste, and will be partially eliminated with the milk. According to Eost and Wiley boracic acid may pass into human milk; likewise after the ingestion of Glauber salts the S03 content of the milk is supposed to be increased (Hess and Schaffer). According to Baum tartar emetic is not found in the milk of. cows treated with this drug, whereas Harms claims it is eliminated with the milk. The feeding of large quantities of alcohol effects a diminution of the specific gravity and an increase of the fat content of the milk. The quantity of the milk appears somewhat increased (in goats). Elimination of alcohol with the milk does not occur. Wel- ler and Teichert proved that alcohol would pass into the milk of cows after feeding them with large quantities of incompletely ex- tracted distillery slops. Although Horder and Herdegen claim the secretion of salicylic acid with the milk, Richter, Pauli and Stumpf disclaim its elimination in large amounts. Pauli and Stumpf succeeded in de- tecting small quantities of salicylic acid in the milk of nursing mothers treated with this substance, and also in the urine of their babies, as well as in the milk of experimental goats. In this regard individual peculiarities must also be considered since in one nurs- ing mother the presence of salicylic acid was demonstrated, where- as the examination was negative in another case. According to Pinzoni the same applies to antipyrin. Salol does not appear in the milk after its administration. Chloroform and ether are found in considerable quantities in the milk after anesthesia (Nicloux). Drugs affecting Milk. ^39 Landsberg failed to detect morphine in the blood, urine or in the organs, either after subcutaneous or intravenous injections, and Stumpf and Pinzoni do not believe that after therapeutical administration of morphine it will pass into the milk in demon- strable quantities. This was found by Ittalie to be the case with opium. Oil of turpentine is not eliminated with the milk (Ittalie), and the same is true of santonin (Coronedi). Stumpf undertook experiments with pilocarpin without how- ever being able to find the pilocarpin in the milk, although his methods were unsatisfactory. Atropin and fluorescin administered subcutaneously, accord- ing to Fugin and Bonanni, and Ittalie, may be demonstrated in the milk. It shcmld also be mentioned here that according to Ostertag meat of poisoned animals may be eaten without harm to the health. He established the fact that meat from animals which have received medicinal agents for therapeutic purposes may be consumed without any possibility of danger. The harmlessness of the meat of poisoned animals has been established by Frohner and Knudsen for strychnia, eserin, piloearpin and veratrin; by Harms for nux vomica and tartar emetic; by Feser for strychnine and eserin; by Spallanzini and Zappa and Sonnenschein for arsenic; by Gautier for cotton- seed cake ; by Feser for apomorphine ; by Peschel for colchicum ; by Warnke for morphine ; and Albrecht for litharge. Of course milk may contain certain quantities of poison since the udder has a special function as an excretory organ. The question of elimination of medicinal remedies, however, is not of practical importance since the medicinal doses are relatively small and their elimination occurs only in traces. In this entire question milk inspection is powerless. Through educational advice by the consulting veterinarian the producers may be reminded of their duty corresponding to the prohibitive measures, not to include with milk for the market that produced by animals which are under treatment with certain drugs. From a hygienic standpoint only those remedies deserve mention which are eliminated for a long period after their administration, as for instance lead and medicines whose prolonged ingestion may pro- duce disturbances of health even in the smallest doses. Considering the fact that in normal feeding with good feeds of any kind the individual influence is paramount in milk produc- tion, it becomes evident that in establishing regulations for pro- curing children's milk more stress should be laid on the health of the animals, on good attendance and care by healthy milkers, and on thorough cleanliness of the stable, and cleanliness in procuring and handling the milk, than on rigorous regulations for feeding which cannot be satisfactorily carried out by the owner on economic grounds, since he must utilize the by-products or refuse of any industry of his vicinity. There is no reason why pasture milk, or milk obtained after feeding green food should be excluded from the market as certified or children's milk, especially if from a dietetical standpoint the advantages of green feeds for cattle are considered, and the favora- 140 Effect of External Influences. ble influence which the pasturing exerts on milk production and metabolism be regarded. Spoiled feed should be prohibited, and also foods which are readily subject to decomposition (fresh residues of breweries, sugar refineries, etc.). Food which is obtained through fermenta- tion processes (hay, grass, clover, mangels, potatoes, green corn, stored in pits in the ground) should if possible be limited, since substances of odor and taste are eliminated with the milk and especially food bacteria which diminish the keeping qualities of the milk. Although they might not have a direct harmful influence in the human organism nevertheless they may spoil the taste of the milk. The beneficial influence of pasturage cannot alone be attributed to the advantage of change of feeding, but is also the result of the stimulating action of the light and air on metabolism, and of the mild form of exercise. Therefore in the absence of pastures it would be advisable to provide exercising paddocks for the animals. According to Munk moderate exercise increases the yield of milk and its proportion of solid substances. Excessive exercise of cows should be avoided. Although moderate exercise on rich pastures in connection with other factors which increase metabolism, produces more abundant and richer milk, increased exercise reduces the quantity of milk but increases its fat contents. In over-exertion however the quantity and quality of the milk are reduced, and the milk and butter both develop an irritating taste (Dolgich). Exhausting transportation changes the milk production con- siderably, especially when during that period the cows are milked irregularly, or for advantage in selling the cows the udders are allowed to become engorged with milk. Stasis mastitis results, which may be cured only by repeated and thorough milkings. Excitement of any kind, such as fright, taking away the calf, change of surroundings (new purchaser), and change of feed, may for a longer or shorter period cause a diminution of the quantity of milk and a change of its quality. Backhaus observed an increase of over 7% in quantity of milk production and 8% of the fat content after the cows had been curried; in other cases it amounted to 4 and 2y2%, respectively. In pasturing cows, sheds should be provided for shelter from the strong rays of the sun and rain; otherwise according to Schwenk the yield becomes diminished. Kirsten observed a dimin- ution of the production of milk after prolonged rain. According to Ingersoll and Duncanson, marked changes in the weather may even be of significance during the season when the animals are stabled. A rise or a fall in the temperature may cause a reduc- tion in the fat content. In the morning following rainy nights the milk may become richer. The influence of weather and pasture on Method of Stablin milk production has been observed by various German investiga- tors, but the results differed considerably. Some observed a dimin- ution of the fat content, others an increase, while some noted a diminution of the milk yield, and a number of others detected no reaction whatsoever. Following the passage of a heavy thunder shower a diminution of the milk yield and an increase of the fat content were observed which corresponded to the increased activity of the animal in the equalization of the body heat. If herds which are pastured at night are compared with those which are stabled at night, no favorable influence of the stabling at night is observed, neither regarding the quantity of milk nor its fat content. In animals kept uninterruptedly out of doors the fat content increased more rapidly than in those kept in stables. In the former the live weight increased more rapidly than in the latter ; in other experiments, however, the results remained the same. Wychgram in his experiments in East Friesland found the milk yield in stabled animals increased, but the fat content diminished as compared to milk from cows at pasture. The cows which furnish the milk supply of cities as a rule are kept in large stables. The stabling of course should be such that the health of the animals does not suffer, and means should be provided for a pure milk production. It is not so difficult to comply with these two requirements as some farmers believe. They may be attained without great ad- ditional cost, since the increase of expense for proper stabling is amply covered by the increased income from the animals. In equipping so-called model stables, an extraordinary amount is usually expended for luxury in the equipment and furnishings, so that the practical farmer on visiting such stables is frequently disheartened, instead of being encouraged to change his more primitive place of milk production to comply with these models, since a simple calculation of the expense of such buildings for keep- ing cows shows him that a change of his stable conditions to equal those of the models is impossible. A really valuable modern stable however may be built at the present time without considerable additional cost, and may be equipped so that the additional expense of milk production due to the wearing out of the building is not greater than the cost of repairing an unsanitary stable. For the erection of a new stable a dry building site should be chosen if possible, or at least the penetration of dampness from the ground should be prevented through separation and isolation of the ground and walls. Only under such conditions can the require- ment of clean walls be attained. The floor of the stable must be water tight and without cracks and holes, and should permit of ready cleansing and disinfection. It is to be regretted that such water tight floors are frequently too cold for milk cows, and the action of the cold must be diminished 142 Effect of External Influences. by the provision of wooden planks. The floor surface must be rough enough to afford the animals a solid footing. The walls of the stable should be provided to a height of 6 feet with an unpenetrable, washable covering, which however should not be dark as was customary in the past, in order to hide the ac- cumulated dirt, but should be light in order that dirt may be readily seen and removed. The stable ceiling should be separated from the feed loft situated above it, by an air space, and should be whitewashed, the same as the walls. In order that it should be impervious to the sta- ble odors, the ceiling on the inside of the stable should be covered with minerally treated pasteboard. The air space between the ceiling and feed loft should communicate with the outside air. Angles and corners should be rounded off, in order to prevent the accumulation of dust. In large herds the erection of several small separated stables should be given preference to a large single stable for all animals. Each of the stables should be made for 16 to 20 animals. The advantages of the smaller buildings are manifested in better pos- sibilities of ventilation, the easier removal of manure, cheaper con- struction of the roofs, less excitement for the animals, and better possibility of caring for and feeding the individuals. Against these advantages the absence of close supervision, which is afforded by the large stable, is not material (Schuppli). The animals should be placed in rows in such a way that the light may strike them from the side or from the rear. In order to provide a great amount of light, high, broad win- dows and transparent instead of only translucent window glass should be installed, the total lighting surface of which should amount to at least one-twentieth of the floor space of the stable (according to Schlossmann the comparison should not be much less than one-fifth). Placing the animals face to face should be avoided on account of the danger of infection with tuberculosis, or this danger should at least be diminished by broadening the passages. The windows should commence from 5 to 6 feet from the floor. Artificial illumination should be provided for; transoms assist in the airing of the stable by allowing ingress of natural atmosphere. The ventilation should be calculated so that the air of the sta- ble should not contain more than 3:1000 (Marker), or 1:1000 (Schlossmann) carbonic acid. According to Schlossmann a cow weighing 1,100 Ibs. produces 12.71 cubic feet of carbonic acid, which would have to be diluted by 1,000 times its quantity of in- troduced air in order to contain only 1 :1000 of the required quan- tity in the air. This introduction of air is made possible by three changes of air per hour, without permitting a disturbing draught. Therefore, according to Schlossmann, the air space in a stable for cows weighing 1,100 Ibs. must be 12,710 :72=176.5 cubic feet. Gen- Stable Construction. erally however, on account of economic grounds a much smaller air space has to answer the purpose. Schuppli even believes that a reduction of air space below the ordinary 70 to 88 cubic feet of air per animal would be permissible when the ventilation system is working properly, and is satisfied with 42 cubic feet per animal when the ventilation provides for sufficient renewal of air. The supply of air is provided by wide shafts which take the air from the outside at a height of three feet, lead it up through the wall and expel it from the stable ceiling into the stable. The foul air escapes through an opening close to the stable floor of one or more discharge shafts, which are carried to the highest point of the stable ceiling, or sideways from the median line upwards and outwards. The total capacity of the discharge shafts should be somewhat smaller than the capacity of the supply shafts. In intensive ventilation, especially when the air space provided for each cow is considerable, heating of the stable might become necessary, a provision which of course could not be considered for the ordinary, medium-sized or even larger establishments. Well-installed transom ventilators, if sufficient attention is given to their operation, would supply the desired change of air even without heating, and at the same time maintain the desired temperature of 16° to 18° C. If heating is provided the air sup- ply shafts should open over the heaters. In providing stalls, from the standpoint of cleanliness only the so-called Holland type of stable floor should be recommended for dairy stables. The principle on which these are built consists of rather short standing space with broad, deep drainage trough in the rear. The urine and manure falls into this trough, and soiling of the animal is thereby prevented while the contamination of the bedding is minimal. Among objections to the Holland type of stables, it is sometimes claimed that the animals cannot move sufficiently and that such stabling constitutes a cruelty to the animals, etc. The best proofs against these objections without doubt, are the facts that in countries which are in the highest state of agricultural development this method of stabling has been practiced for a long time, and the animals soon get used to this method of stabling without suffering in their general condition or being affected in their milk production. In the Holland method the tails of the animals are tied with a cord in such a way that while the animal stands its tail hangs in a natural position, but when lying down the tail is kept elevated so that it cannot be submerged in the contents of the drain. The cords are tied to a rod which runs near the ceiling, parallel with the row of cows, or the cords, with weights attached, are allowed to hang over this rod. The shortness of the stalls of course requires a low feeding trough, over which the animals may extend their heads while lying down. In order that the animals may not annoy each other, the individuals are separated by means of partitions, which extend 144 Effect of External Influences. upwards to the height of the head or the shoulder and at the same time have fastenings which are used for tying the cows. To prevent the cows from backing into the drain, a moulding of one-half inch is provided along the upper border of the drain trough. This moulding holds the slipping foot and makes possible the placing on the floor of wooden slats when pregnant -animals are about to calve. The fundamental principle against the possibility of slipping is the above-mentioned provision of a sufficiently rough stable floor. The width of the stalls should be about S1/^ feet, the length 51/4 to 53/4 feet. The most satisfactory feeding troughs are those which cor- respond to the conditions of natural feeding in the pasture, and they should be so constructed that they will serve for animals of all ages with the possibility of providing partitions in order to separate the individual rations. For watering the animals it is advisable to provide each stall with automatic water supply. •The accumulated litter in the drain trough should be mechan- ically removed as often as possible into liquid manure pits which terminate in a tunnel with collecting canals, or the litter may be thrown into a chute through a shaft leading to a water-tight liquid manure pit the size of which should be so arranged that 88 to 106 cubic feet of manure space are allowed for each animal. Over the liquid manure pit on wooden lattice frames, or along- side of it, should be placed the manure pile, with 30 square feet of surface for each grown animal. On account of the desired decom- position of the material the first-mentioned arrangement of the manure over the liquid manure pit is most desirable. There should be a separation between the stable and the manure pile of at least 20 feet and the latter should be placed on the side opposite to the principal direction of the wind in that locality. The outlets of the manure drains should be closed from the stable by trap or slid- ing doors. Good straw should be selected for the bedding of animals. The question of straw which is very important in localities where but little is grown is favorably solved by the Holland method of stabling, since by this method a great deal of straw is saved by the short stalls with but slight soiling of the animals. Forest and shade leaves are not recommended, since it is claimed that milk troubles result from their use. Turf straw, shavings and saw- dust should be avoided if possible on account of the formation of dust, but should not be excluded if good straw bedding cannot be obtained. The use of bed straw should be prohibited in milk stables. The feeding of the cattle must be performed after milking, on account of raising the dust. The removal of the manure and the cleaning of the animals should take place at least one hour be- fore the milking. Complaint is frequently made to hygienists that the require- Complete Milking. ments which are made by them relative to stable hygiene must in- crease the cost of keeping the animals, and thereby increase the cost of the milk. This view is only justified to a slight extent. Stable hygiene if satisfactorily adjusted will result in a considera- ble increase in the yield of the dairy animals. The economic losses which are induced by udder affections, which spread with especial rapidity in filthy stables and from unclean milking, have been discussed in a special chapter. Atten- tion should only be directed here to the increased production fol- lowing proper attention to cleanliness of the animals, and to the findings of Bloymeyer and others, according to which cows in well ventilated stables, all other things being equal, yielded from 450 to 480 liters more milk per head each year than cows kept in unven- tilated stables. The favorable influence of exercise and light work has also been discussed above. If possible the animals should be given an opportunity to run out of doors in a paddock for at least one to two hours daily, even in the winter months. Of all outside influences, regular and complete milking con- stitutes the most prominent stimulant for inducing the activity of the udder. It is known that cows which are milked three or four times in each twenty-four hours give more milk than those which are milked only twice (Backhaus). The increased yield from milk- ing three times amounts to from 10 to 15% more than the pro- duction obtained from two milkings ; in four milkings the increase amounts to from 6 to 8% as compared with three milkings. The quantity and composition of the milk at each milking de- pend somewhat upon the time which has elapsed since the last milk- ing. According to general experience the morning milk is of greater quantity with a smaller amount of fat, in comparison with quantity and fat content of the evening milk. During the night absolute rest prevails, whereas during the day the influence of light and motion causes an increase of metabolism which is also mani- fested in the variations of the body temperature shown by the animal in the morning and evening. The differences in milk, which are obvious in irregular or so-called broken milkings, may be explained in a different way ; that is while the milk at the beginning of the milking contains 0.5 to 1.5% fat, in the middle of the milking it shows 2 to 3 to 4%, and again rises towards the end, during the last strokes of milking to 8 and 10% (Melander, Kaull, Gotta, de Vrieze). The fat-free solid substances are subject to slight changes (according to Boussin- gault the fluctuation amounts to from 0.2 to 0.28%). The condi- tions in the milk when the calf sucks are similar. A truly plausible explanation can hardly be given to the supposition of a separation of cream in the udder (Zschokke) ; likewise it hardly seems reasonable to suppose that the thin plasma particles flow towards the cistern, while the fat globules as a result of greater fusion and friction are retained, and are only pressed towards the larger milk ducts and the cistern by the newly formed milk which is secreted during the act of 10 146 Effect of External Influences. milking. The principal cause lies probably in the fact that the separation of the fat represents a greater expenditure of energy than the secretion of the plasma. If the cell is exhausted by previous milking it then secretes milk during the period of rest which is especially rich in plasma and poor in fat. Through this period the alveoli and milk ducts are dilated, and the gland cell becomes flat and is at rest. In this position of rest it recovers and is ready for renewed action when, through renewed milking operations, the fluid is withdrawn and stimulation of the secretion is applied through the teat. If the milk is removed without this stimulation of secretion, with the aid of a milking tube, only milk poor in fat flows from the cistern and the larger milk ducts, and the flow ceases as soon as pressure is no longer exerted on the secretion. If, however, through milking (or other stimulation) new secretion takes place, the rested gland cell engorges with nutritive material, and converts it into fat, which is separated during the milking in increased quantities until the secretion of plasma and the separation of fat cease, which marks the height of these two processes in the secretion of milk. Through an increased stimulation by additional milkings the cell may be further stimulated to a special production, which consists in an increased fat formation (Hegelund). Henkel succeeded by this procedure, in increasing the quantity of milk by 2.4%, and the fat content by 6.2%. As already mentioned Hegelund 's method requires additional work, which may possibly lead to the hiring of additional help and must be considered (Kirchner), when estimating profit and expense. The principal factor in the various methods of milking lies in the thorough milking out of the udder, which will retain its maximum of production only by such practice. Henkel suc- ceeded in demonstrating the extent to which the milk production de- pends on the thoroughness of the milker. The production of a cow when milked by a thorough milker amounted to 8.1 kg. (17.8 Ibs.) of milk, with 4.2% of fat; by a less satisfactory employee to only 5.6 kg. (12.31bs) with 2.7% of fat. At the same time it is immaterial what method of milking is pursued, that is, whether the teats are milked crosswise, or those on one side, or those of opposite quarters, simultaneously. Milk- ing of a single teat at a time, which of course is not customary, yields less milk, and the last milked quarter is the poorest in fat (Lepoutre and Babcock). The influence of special methods of milk- ing has been more fully discussed in the chapter on the procuring of milk. According to Klinkmiiller the milk yield of the right half of the udder is 3. 97 kg. (8.7 Ibs.), the left 3.65 kg. (8.03 Ibs.), with fat contents of 3.65 and 3.31% respectively. The cause of this increased production of the right half of the udder is, according to Klinkmiiller, the result of the practice of milking the right half first, and therefore it is advisable to practice alternation in milking, from right and left. If milking stools are used care should be taken that the milkers do not take hold of the seat with their hands. The most recommendable stools have only a single foot, and are secured around the body by a strap. Switching~of the animal's tail must be prevented during milking by tying it up, or by other effective contrivances. Conditions which prevail in the handling of milk after it has been procured are of special importance in providing milk of Hie best quality. The changes which milk undergoes have been dis- Milk Pails. cussed sufficiently for the purpose of milk hygiene in special chap- ters. Those points principally should be emphasized which are to be followed during the drawing and preparation of the milk, in order to check or prevent undesirable and early decomposition of the product. This relates primarily to cleanliness. The requirements of milk hygiene go hand in hand in this respect with the purely economic requirements of the dairy industry. If it is considered how much milk spoils prematurely on ac- count of improper care and the amount of loss which is sustained when the creameries have to discard hundreds of pounds of cheese on account of improper handling of milk, then the economic value of cleanliness in the stable becomes obvious. The Holland method of stabling, cleaning of the cows and especially the udders are quite simple but important factors in such cleanliness. In keeping the udder clean special attention should be given towards preventing its contact with filth. Dry cleaning with suitable straw, or rough towel is prefera- ble to moist washing which often consists in spreading the softened dirt over the entire udder. If the dry method of cleaning the ud- der is used such milking pails should be provided which will pre- vent the milk from becoming contaminated by the dust originating from the cleaned udder. If the udders are washed it should be done with lukewarm water without soap. Subsequently the udders should be rubbed dry and slightly lubricated with paraffin salve. Even with these simple operations milk may be obtained containing only very small num- bers of bacteria, and would suffice for all practical purposes. Cov- ering the animals with linen sheets, disinfection of the udder in water-tight bags, and washing of the entire animal represent some- what exaggerated procedures, and besides they require additional expense, which can be afforded only through a special increase of the price of milk. The milk pails should be so constructed that they will pre- vent dust and dirt from falling into the milk. This is accomplished by using covered pails, which possess a special receiving tube sup- plied with a funnel for taking in the milk. Between the receiving tube and the funnel an arrangement for filtering through cotton may be placed. The so-called Algauer milk pails are provided with such arrangements ; likewise the Konigsf order milk pails and the sanitary pails of Gurler and North. The funnel should be rinsed and provided with a fresh piece of cotton after the milking of each cow. After milking is finished the milk should be immediately taken from the stable. This is frequently accomplished by pouring it into a funnel arrangement fastened to the wall through which the milk passes into a suitable tin lined tube to the milk room. This tube should be removable in order that it may be properly }48 Effect of External Influences. cleaned. In the milk room the milk is further treated by another straining and cooled by simultaneous aeration after which it is either directly filled into bottles, cans or collected in a vat in order that it may be thoroughly mixed. The milk should be handled as little as possible, since each manipulation not absolutely necessary, means a poorer condition of the product from a hygienic standpoint. The producer there- fore after straining the milk through cotton strainers should cool it and fill it into clean bottles or well-galvanized and properly cleaned cans. The straining of milk through straining cloths which have been carelessly cleansed by rinsing in cold water, and which in most instances fail to answer the purpose on account of their large meshes, is, it is to be regretted, in most instances merely a pre- tense, which only tends to further spoil the dirty milk. Milk which is obtained in an unclean way cannot be deprived of its poor qualities by any mechanical means, since the filth dissolves and the bacteria pass through the straining cloths and the cotton filter. If the accidentally contaminating bacteria are removed immediately during the milking (cotton filter in the funnel of milk pail), a con- siderable improvement of the milk results. The value of artificial cleaning, however, will continue to decrease in proportion to the length of time elapsing between the time of milking and cleaning. If the cleaning of dirty milk is accomplished only hours afterwards at the collecting places and creameries it should be considered as direct fraud, which gives the product the appearance of good qual- ity without however improving it in any way. In such cases filtra- tion and centrifugalization only serve as means of deception. Filthy milk which has been subsequently cleaned, must in spite of its cleaned condition be considered as spoiled in the sense of the pure food law, even if no changes are yet apparent in it. In milk control work there are frequent opportunities for confiscating dirty market milk, and not infrequently the examina- tion reveals that the contamination of the milk consists in dust- sized particles and cow manure, all of the same caliber, which in- dicates that the milk has been subjected after milking, to a straining process which permitted the manure particles which had been dis- integrated during the process of milking to pass through the strainer. Unstrained milk obtained under filthy conditions usual- ly shows the presence of course straw particles, manure, bits of fod- der and cow hair. After straining, the milk is allowed to flow down over the out- side of a double corrugated surface, or a series of parallel horizon- tal pipes for the purpose of cooling and simultaneous aeration; these surfaces are kept cool through pipes containing running water, ice water or brine. Especially practicable and serviceable are the so-called round coolers which are provided with spiral pipes, covered with tinned-copper sheets, over which the milk runs in a thin layer. Aeration. It will be proper to describe here very briefly the changes which milk undergoes through freezing. The freezing of milk oc- curs with remarkable frequency in the winter time, when the milk is subject to long transportation. There is no change in the number of bacteria which were present at the moment of freezing until after the thawing of the milk. There is neither diminution nor marked increase. According to the data of Vieth, Kaiser and Schmieder, Hen- zold, Bordas and Raczkowski, Fritzmann and Mai it may be seen that in the freezing of milk a marked separation takes place. Mai found that such milk under certain conditions may appear at the first glance like ordinary milk, although it is really frozen. Crystal needles of ice make their appearance in such milk. If the freezing continues layers of ice appear at the sides of the milk cans and on the surface, thus enclosing a central fluid portion. The upper part of the milk containing the cream layer freezes more loosely, in a spongy leaf-like manner. After thawing, the milk has its original consistence and its original odor and taste. The peroxi- dase content also remains unchanged. The milk inspector must consider the separation of milk during freezing. In taking a sample, special care should be taken to determine whether the milk cans or other vessels already contain ice. Frozen milk should not be sold to customers until thoroughly thawed. The aeration of milk permits the escape of carbonic acid, hydrogen and sulphide of hydrogen, and supplies the milk with air, so that in all probability the development of certain bacteria is checked, which otherwise, if the milk had been filled into con- tainers in a warm and un-aerated condition, would have imparted to the milk a sharp, disagreeable animal taste and odor ; the milk would have been "suffocated." The corrugated surface coolers are especially suitable for use in small dairies. The cans into which the milk is filled after cooling should be tinned in a satisfactory manner. It is to be regretted, however, that this is the case only with new cans. The tin covering espe- cially on the places where the outside strengthening bands are placed, is very imperfect, and after a shorter or longer time defects in the lining develop, which soon result in an extensive formation of rust. The oxidation of the iron finally results in tears and holes which produce deep, sharply circumscribed depres- sions in the side of the can in which rust, decomposed milk and slime accumulates. The transportation of milk in rusty cans, or those in which the lining has become damaged, gives it a disagreeable tallowy taste. Milk should be protected from bright light. Sun rays and indirect daylight may give the milk a tallowy rancid odor and Effect of External Influences. taste, in the same manner as is the case with the prolonged action of ultra-violet rays. It is important during transportation that the vessels be closed in an air tight manner, and with a cover consisting of non-porous material. All milk utensils should be cleaned with hot soda solution, with subsequent rinsing in fresh pure water, and if possible com- bined with steam sterilization and rapid drying in places protected from dust. The transportation of milk should be rapid, and where pos- sible it should be shipped after each milking. In creameries the treatment of milk after its receipt should be principally confined to cooling. This cooling is carried out in deep cooling appliances or double coolers in which the abstraction of the heat takes place through water at the point at which the milk flows into the cooler, and the lower part is further cooled with ice water or with brine. All further manipulation and at- tempted improvements of milk for drinking purposes are of no use. Spoiled milk should be excluded from the market and not be subjected to renovating processes. Thus in some creameries it is customary to clean the milk not only by renewed filtration, but also by centrifugalization, which is frequently done on the supposition that the bacterial content of milk becomes reduced through such treatment. This, however, is impossible ; on the contrary, such milk often becomes contaminated again by bacteria from the non-sterilized centrifuges, and even if the milk is centrifuged in a sterilized apparatus only those bacteria will be eliminated which adhere to the courser bodies having a higher specific gravity (pus, fibrin, filth, casein coagulum, etc.). The separator slime therefore contains principally fodder and manure bacteria, lactic acid bacteria, species of milk moulds, and bacteria of those fermentation processes which take place in the residue of milk cans and transportation vessels, and further the specific causative agent of mastitis occurring in pus. Therefore, although a great number of bacteria are removed, the bacterial count through plating of the centrifuged milk dis- closes a considerably larger number of bacterial colonies than was the case in the milk prior to centrifulgalization, although the short time of the centrifuging process does not permit of an actual increase of the bacteria. Severin observed an apparent increase in bacteria up to 70%. This may be explained by the fact that through centrifugalization, bacterial clumps and colonies floating in the milk, and the clumps of pus and fatty leucocytes which have embodied bacteria, are broken apart, and the bacteria are thereby distributed in the milk. Therefore, in spite of the removal of considerable numbers, there is an apparent increase. This distribution is such that the separator slime and cream are considerably richer in bacteria than the skim milk. The Bacteria in Cream. richness of the cream in bacteria may on one hand be explained by the fact that large quantities of bacteria are dragged upward with the fat globules, and on the other hand by the fact that leucocytes containing bacteria, inflammatory products, etc., which possess a lower specific gravity, through fatty changes or fat enclosures, are taken up with the fat globules into the cream. The most important factor in the spoiling of milk in cream- eries may usually be found in its being kept for too long a period before it is marketed. CHAPTER VIII. BACTERIA IN MARKET MILK; THEIR ORIGIN AND ACTION. Before milking is commenced the udder should be cleansed of all adhering dirt. Cleanliness in milking is one of the most important factors in giving the milk good keeping qualities. Sub- sequent cleansing through straining, nitration, centrifugaliza- tion, etc., is of little purpose after the dirt particles (straw, manure, dirt) have once imparted to the milk their soluble con- stituents, and an actual inoculation has been accomplished with the bacteria of filth. The tail of the cow should be tied, in order to prevent bacteria from the skin being thrown into the milk by its switching. If left free it may even subject the milk to contamination with coarser substances. The importance of the effect on human health of bacteria which fall into the milk, and which multiply therein when milk is improperly procured, is not known, but the thought is at least repulsive when it is considered that milk consists of a manure sus- pension of a bacterial culture, and on this ground alone absolute cleanliness in milking should be insisted upon. In order to attain this it is again necessary to provide a properly ventilated and well kept stable, as well as milk room. The veterinarians can in no way obtain a better recognition of the milk problem than by always pointing out to the farmer the necessity of keeping healthy cows in properly constructed and well-kept stables, and in impressing upon him the fact that the procuring of pure milk and its proper handling constitute the fundamental principles of a prosperous development of the milk industry in general, and not for the milk supply of the city alone. Many farmers, espe- cially the small ones, can only be convinced by practical demon- strations of the advantages to be derived from proper stabling and care, and therefore it is our duty to win over reasonable and progressive farmers to the erection of model dairies, and to offer to the smaller farmer the aid and advice by which he can improve his condition with the least expenditure. Even if nothing more than diligence, attention and a feeling of responsibility are aroused these alone would mean a tremendous improvement over the con- ditions prevailing at the present time. It is evident that if no special milk rooms are provided every- 152 Procuring Sterile Milk. 153 thing should be avoided during milking which would cause stirring up of the dust, such as removal of manure and feeding. As long as the milk gland is in a healthy condition the cells secrete a sterile product, which becomes contaminated with bacteria only in the lowest part of the teats, in their ducts, or during the process of milking, etc. Bacteria are always present in the lowest parts of the ducts of the cistern, as a result of contamination from the litter. These bacteria and also those which fall into the milk during milking, and the massaging of the quarter, render the procuring of sterile milk practically impossible, even when the strictest care is taken to prevent as far as possible the subsequent contamination of the milk. In spite of opposing views, it may be considered proved at the present time that the milk in the udder is sterile as long as the animal is not affected with diseases of the udder or severe general affections. Lister, Miessner, Escherich, Kitt, Tromms- dorff, Eullmann, Seibold and others succeeded in procuring abso- lutely sterile milk. This, of course, was only in small quanti- ties, and drawn with special care, such as washing the udder, disinfection, protective covers, etc. The first streams of milk are of course always contaminated with bacteria (Schulz, Luz, d'Heil) ; the subsequent ones may be sterile, but frequently they also may contain bacteria, as confirmed by the works of Boekhout, Ott de Vries, Ward, Koning, and Freudenreich. In practice the procuring of milk with a moderate number of bacteria must be considered satisfactory. Schulz, for instance, found that the first milk procured contained 55,566 up to 97,240 bacteria per c. c., while during the middle of the milking it contained only 2,070 to 9,985, and in the last from 0 to 500 bacteria. In an interrupted milking, that is, when the milk was obtained in four portions, Backhaus and Appel counted in the first part 170 to 950, in the second 60 to 255, in the third 10 to 70, in the fourth 0 to 45 bacteria per c. c. For procuring sterile milk the following measures are recommended: Washing the udder with soap and water, disinfection with alcohol (Kitt, Kolle) ; disinfection with mercuric chloride solution (Fauss, Klimmer) ; mercuric chloride solution and rinsing with boracic water and formalin (Boekhout and de Vries) ; washing with a 2% mercuryoxycyanide soap (Freudenreich), followed with rubbing with sterile cotton (Ostertag) ; salicylcotton (Eichert); sterile cloths (Freudenreich); and then after cleaning and thoroughly disinfecting the hands of the milker with soap, water and the same disinfecting agent which has been used for disinfection of the udder, the milk may be carefully drawn by the usual method, which is termed ' ' fisting. ' ' Backhaus after a coarse cleaning of the udder, covers it with a bag which con- tains disinfecting fluids. After a short action of the disinfectant the fluid is allowed to escape through a stop-cock attached at the lower part of the bag and the udder is rinsed with previously boiled warm water. Other authors cover the body of the animal with sheets, leaving only the udder exposed. Boekhout, Ott de Vries and Trommsdorff used sterile milking tubes for taking samples. In this operation it is to be regretted that even in the most careful manipulation with sterile milking tubes, infections of the udder sometimes result. Eullmann therefore prefers direct milking. He also rubs white paraffin salve into the skin of the surround- ing part of the udder. 154 Bacteria in Market Milk. Through such protective measures individual authors obtained the following results: Freudenreich : 200 to 300 bacteria per c. c. Szasz: 2 sterile, 11 with an average of about 2,700 bacteria. Hesse : 1,600 bacteria per c. c. Marshall : 295 bacteria per c. c. Lux : 0 to 97 to 6,800 bacteria per c. c. Kolle: 80 to 15,000; in 33% of the experiments the counts wrere below 300 bacteria per c. c., 50% below 500, others up to 800 per c. c. Only 4.7% yielded 700 to 800 bacteria. Willem and Minne : 1 to 5 bacteria per c. c. Willem and Miele : 0 to 37, 4 to 218 bacteria, respectively. Seibold studied the bacterial content of the milk under the most varied experimental methods, and especially under condi- tions which correspond most nearly to those prevailing in practice. 1. Without protective measures. 2. After soaping the udder. 3. After soaping and disinfecting with alcohol. 4. After repeated disinfection with alcohol, and procuring through sterile milking tubes. The poorest results were obtained, as would be expected, by the first method, and the best results by the fourth method, with which it was frequently possible to obtain completely sterile samples. The number of bacteria by the fourth method fluctuated between 0 and 12, by the third between 0 and 85, and by the second between 0 and 434 per c. c. The first method produced samples of milk with less than 10 up to several thousand bacteria. Trommsdorff and Rullmann observed in samples which had been procured without special precautionary measures, such as cleaning of the udder and hands, on an average (96 samples) 6,700 bacteria per c. c., but only 1,500 bacteria when a thorough cleaning of the udder and of the hands of the milker had been undertaken. Seibold, Trommsdorff and Eullmann found in individual cases an enormously high bacterial content even in freshly pro- cured milk, the colonies on the plates containing mostly strepto- cocci. These samples were obtained from cases of inflammation of the udder, and the milk was already contaminated with streptococci before leaving the udder. These organisms would not otherwise be present in aseptically procured milk (Seibold). ^As it is difficult, even under the strictest conditions, to procure sterile milk, or milk with a very low bacterial content, therefore in the wholesale production of milk such results are still more difficult, and in fact impossible. The milk, immediately after leaving the milk canal, becomes contaminated by bacteria which have colonized there. Among the bacteria which may be found Reduction of Bacterial Content. 155 in the milk from animals free from udder affections, and which has been drawn under aseptic conditions, the groups of staphy- lococci, colon bacteria, Bacillus subtilis and B. mesentencus should be especially mentioned. Seibold also demonstrated acid-fast rods. Eullmann and Trommsdorff found no representative of the colon group, but they isolated staphylococci, a few representa- tives of spore-bearing species, and especially the anthracoides (mycoides) species. In ordinary milk production there also come into consider- ation an army of air and stable bacteria, which adhere to the food, manure and litter, as well as those which vegetate, as saprophytes, on the skin of cattle, especially on the skin of the teats, and on the hands of the milker, besides those groups of bacteria which colonize with special predilection on milking utensils and in the cans. The number and kind of bacteria found by the different authors vary to a considerable extent, depending upon the degree of cleanliness used in obtaining the several samples. Dean found in milk — From filthy cows 9,845 to 17,155 bacteria per c. c. From clean dry cows 8,295 to 9,426 bacteria per c. c. From cows with dampened skins 640 to 2,350 bacteria per c. c. The same work also throws light on the influence of the milk vessels. If the milk was collected in sterile milk vessels, it contained 355 to 1,702 bacteria per c. c. In well cleansed milk vessels. 13,080 to 93,420 bacteria per c. c. In dirty milk cans 215,400 to 806,320 bacteria per c. c. Russell, in using sterile milk vessels, found 165 bacteria per c. c. in freshly drawn milk, while in case of only ordinary cleansing there were 4,625 bacteria per c. c. in such freshly procured milk. Grotenfeld counted in the milk from well-kept animals, in clean stables, only 106 bacteria as compared with 670,000 per c. c. in milk from dirty stables. The kind of milk can also has an influence. Backhaus con- siders enamel cans as the best; tin vessels were found to be almost as good, while milk vessels constructed of wood were unsatisfactory. The work of Koning shows the influence of the bacterial flora of the air on the bacterial content of the milk. The author counted 500,000 to 700,000 bacteria in the stable air, whereas the outside air contained only 90,000 bacteria. He found that the volume of air between the cows was especially rich in bacteria. Milk which is procured in the pasture contains fewer bacteria than stable milk. If it is customary to change the straw and also feed shortly before or during the milking time, these factors tend greatly to increase the bacterial content of the milk. If the milk is subjected to the so-called " improving methods" of the most varied kinds, and has to be transported for long dis- 156 Bacteria in Market Milk. tances, it is obvious that when it finally reaches the consumer it must contain tremendous numbers of microbes of various kinds. The author counted in the market milk of Munich from 13,000 upwards to several millions of bacteria per c. c. Milk offers to most bacteria which may contaminate it a splendid culture medium, their multiplication in it depending on the character of the container (cans, flat or open bowls), temperature and subsequent treatment. Freudenreich, B. Meyer, Cnopf and others conducted experi- ments on the influence of cooling on the number of bacteria, and established definite proof for the statement made in practice that immediate cooling constitutes the best preserving agent for milk. According to Cnopf the multiplication at 0 deg. C. was re- markably low, at 12.5 deg. it was 4 to 935 times greater, and at 35 deg. 2,200 to 3,800 times greater than at 0 deg. C. Freudenreich proved that in milk which at the beginning of experiments contained 10,000 (accurately 9,300) bacteria, they scarcely multiplied when kept for three hours at 15 deg., wThereas at 25 deg. they doubled, and at 35 deg. they tripled in quantity. After six hours at 15 deg. they numbered 2.7 times, at 25 deg. 18.5 times, at 35 deg. about 1,300 times more than the original number, while after nine hours the number when kept at 15 deg. was 5 times, at 25 deg. 108 times, at 35 deg. 3,800 times as numerous as in the original count ; and in 24 hours at — 15 deg. C. the count was 5,700,000, or 613 times 25 deg. C. the count was 50,000,000, or 5,380 times 35 deg. C. the count was 570,500,000, or 61,344 times The author desires at this place to comment especially on the slight, and -somewhat problematical value of bacterial counts, not alone because the results of the different sowing and counting methods show such enormous differences, but because the entire system also depends on a supposition of the development of a colony from a single bacterium which was previously present, a premise which is open to very serious objections. If it is considered how many bacteria attach to tenaciously adhering threads (sarcina, streptococci'), and how many bacteria possess a tendency to proliferate in cultural combinations, and to remain together in the relatively sticky material of milk, then it becomes apparent that the counted bacterial number represents but a small part of the number of bacteria which are actually present in the milk. Of course in general the number of colonies developing on the plate represent a certain initial point for deducing whether and in what degree a bacterial growth has taken place in the milk, but it does not indicate more than the relative age of the milk, since fresh milk may also be rich in bacteria, and besides luxuriantly growing as well as slowly multiplying bacteria may be present in the milk. A better method for the establishment of the actual number of bacteria in milk is the one suggested by Skar, which consists of a direct count of the bacteria in a smear (see technique). Through plating a certain impression is obtained of the kind of bacteria occurring in the milk, and corresponding to the growth of the colonies and the morphology of the bacteria it is possible to draw certain conclusions as to the groups under which the bacteria that are present may probably be classified. Further deductions as to whether the microbes should be con- sidered pathogenic, and whether bacteria are present which confer Thermal Limits of Bacteria. 157 disease-producing properties to the milk through products of de- composition, splitting up of proteids, etc., can only be possible after an accurate determination of all properties which would allow the recognition of the colony as a certain species belonging to a large group. This determination of the representatives of a group is not simple, and requires study and continued experimental work of days and weeks relative to fermentation qualities, requirements of growth, pathogenic properties on test animals, ferment-like characteristics, etc. These experiments are only of an optional value in practice on account of the easy decomposition of milk as a food substance. Nevertheless through continued experiments on these problems valuable data and results have been obtained increasing our knowledge of the spread of typhoid fever and the methods for combating this and other diseases. Therefore, it should be aimed to prevent the entrance into the milk of directly or indirectly injurious bacteria by procuring the milk in a clean and careful manner. Once such bacteria gain entrance into the milk and multiply, their recognition and isola- tion are too difficult for the practical inspector of milk to consider. Milk hygiene can produce practical results only if it is inau- gurated at the place of production. The pathogenic bacteria of diseases of animals and man will not be considered here, and only brief consideration will be given to the army of saprophytes which gain entrance to the milk from the air, straw, manure and the milk vessels. Although from the numerous possibilities of infection of milk a definite bacterial flora can hardly be expected, nevertheless, cor- responding with the nutritive material of the milk, and the methods employed in its storage and transportation, as well as the subse- quent treatment, conditions are created which are favorable to some varieties of bacteria, while for others they are less favorable or even harmful. Through the growth of a certain kind of bacteria the conditions may be changed in such a way that the require- ments of propagation for other groups are produced. Likewise through symbiosis conditions may be developed which are re- quired by certain species of bacteria, or under which certain species may be destroyed, whereas without symbiosis probably neither of the species could exist, since they are dependent upon each other. The growth of certain species is therefore dependent on numerous influences. According to the thermal limits in which bacteria can live, the species may be separated into those which thrive at 0 deg. C. (up to 15-20 deg.) : psychrophile ; those which thrive at 10-15-40 deg.: mesophiles ; and finally into thermophiles, whose thermal optimum ranges between 40 to 70 deg. C., or even higher. The species of psychrophile propagate even in well cooled milk and at low temperature, and at times change its taste. Ref- erence should be made here to the Bacillus lactis saponacei and the Bacillus sapolacticum, which give a soapy taste to the milk. This defect of flavor is principally observed in cool weather, and at the beginning or end of the winter. 158 Bacteria in Market Milk. Subtilis varieties, mycoid varieties, vinegar bacteria, yeasts, Penicillium glaucum, mucor varieties and aspergilli also grow from 0 to 8 deg. C., as do soil bacteria, fluorescence varieties and bacilli which split up proteids (bitter taste of milk). According to Kniisel, psychrophile bacteria may be demonstrated in sterilized milk, while Bischoff found them in the market milk of Leipsic. Bischoff found that in milk which had been cooled to about 0 deg. C., the bacterial number gradually diminished from the third to the seventh day; it then multiplied rapidly, without showing a considerable increase in the degree of acidity. A bacterial rennet formed, however, and the milk coagulated on boiling. This appeared as early as the fourth to seventh day, when the milk was kept between 6 and 8 deg. C. Frozen milk on the other hand keeps for a remarkably long time. Kniisel found peptonizing bacteria in sterilized market milk which had been kept at 8 deg. C. ; as a result of their growth the milk had the appearance of soapy water, and possessed a bitter taste. Therefore, all milk cannot be protected from spoiling by being kept cool. The milk must be procured at the start with as small a number of bacteria as possible. The opposite of these psychrophile species are the thermo- philes, which may be actually isolated from the army of accom- panying bacteria by a high degree of heat. They continue to grow even in temperatures of 70 deg. C., and over (Zettnow), a tem- perature at which most of the vegetative bacteria and to some extent also spores of the mesophiles and psychrophiles are de- stroyed. Such bacteria were found not only in hot springs by Cert'es, Garrigon, Karlinski, Teich, Tsiklinsky, but also in river water (Miquel, Tieghem, F. Colin, MacFadyan and Blaxall, Michaelis and others) ; finally they were found almost everywhere by Globig, and in the intestinal content of animals, feces, manure, liquid manure, in the soil and upon fodder by Rabinowitsch. The thermophile species are not directly pathogenic. This group, however, contains several toxin producers, and peptonizers of milk. Sporulating bacteria which form spores that resist a heat of 100 deg. C. and over should not be confused with the thermophiles. (Peptonizing species, as mycoides, anthracoides, subtilis, mesen- tericus and the butyric acid bacilli). Between the psychrophiles and the thermophile bacteria lie the large army of mesophiles, to which belong most of the ordinary species of bacteria found in market milk. Corresponding to their requirements for oxygen they are divided into obligatory aerobes, which propagate only in the presence of oxygen, facultative anaerobes, which can vegetate without oxygen, and obligatory anaerobes, which can grow only in the absence of oxygen. They may also be divided, according to the substances which they attack, into those which split sugar, proteids and fats, or, according to the products which they form during their growth in certain media, into acid producers (lactic acid producers, butyric acid producers, etc.), or into alkaligenic species and gas producers, Period of Incubation. alcohol producers, bacteria with rennet-like action, pigment pro- ducers, slime-forming bacteria, etc. The varieties of bacteria which are found in milk under general conditions of production, even when conducted under special provisions for obtaining clean milk with unusual precau- tionary measures (provided that the milk originates from healthy animals and is drawn by healthy milkers), are of special interest to the milk hygienist. These bacteria split sugar and proteids, and attack fat. Ac- cording to Fliigge, they are separated into : 1. Aerobic lactic acid bacteria, which cause spontaneous souring and do not form spores ; 2. The anaerobic butyric acid bacilli, and 3. The aerobic peptonizing bacteria, with remarkably resistant spores. These bacteria, however, do not propagate uniformly well in milk, but they are subject to influences of the medium, which really constitutes an elective culture medium for some of the species, whereas it is destructive for others. The time during which no increase of bacteria can be noted in milk is known as the period of incubation (Soxhlet). In fact there may be not only no multi- plication of bacteria in the milk, but under certain conditions dur- ing the beginning of the incubation there may even be a diminu- tion of the bacterial number which is first found; the bacteria present in the milk are subject to the injurious influences of the animal secretion; the milk is in the germicidal stage (Koning). 'Fokker in 1890 was the first to assert that raw milk (he used for his experiments goat's milk) must have germicidal properties. He proved that raw milk when inoculated with lactic acid bacteria resists spoiling for a longer period than was the case with milk that had been boiled. Prior to his investigations however Wolffhiigel and Eiedel found in 1886 that cholera vibrios readily multiply in boiled or sterilized milk, whereas in raw milk their growth is rapidly checked. As a result of these findings the question as to whether milk possesses germicidal properties became the subject of dispute. While Freudenreich, Hesse, Park, Cozzolino, Conn, Schenk, Behring, Eullmann and Trommsdorff, Eosenau and McCoy, Sassenhagen and Bab claim that milk possesses inhibiting, or even destructive properties for bacteria, Richet, Hueppe, Heim, Friedrich, Kitasato, Uffelmann, Weigmann and Zirn, Basenau, Schrank, Schottelius, Moro, Heinemann, Rubinstein, Stocking, Sommerfeld, Klimmer, Knox and Schorer, and Kuntze express their belief against this power in the sense of the bactericidal action of blood serum, and think that the germicidal properties exist only towards certain species of bacteria. They also believe that- the composition of the milk creates favorable conditions for the propagation of some of the bacteria, while for others this is not the case, just as with elective media, some of the less favored species become injured or destroyed by the multiplication of lactic acid bacteria and their products. The presence of specific germicidal substances (alexins, ambo- ceptors, leucins) in special kinds of milk, such as colostral and mastitis milk, has been proven by the work of Bauer, Sassenhagen, Rullmann and Trommsdorff, whereas the question of the occur- rence of these in normal milk has not yet been sufficiently demon- strated, although the fact of a diminution of bacteria in freshly drawn normal milk has been established by our methods of counting. Bactei'ia in Market Milk. Colostrum : 1. 2. 77,000 82,900 71,000 76,400 In order to furnish a few examples, several experimental results will be cited from the work of Grimmer. Koning found in various samples of freshly drawn milk the following numbers of bacteria per c. c. : Milk: 1 2. 3. immediately 107,000 143,000 18,510 after 6 hours 96,000 142,000 16,000 after 12 hours 74,000 after 18 hours 120,000 155,000 14,700 56,000 after 24 hours 13,200 115,000 after 30 hours 145,000 470,000 13,800 115,000 after 36 hours 2,050,000 after 42 hours 490,000 800,000 106,000 > 596,000 The slight diminution of the bacterial content of this experi- mental series could of course not be attributed to the existence of a specific germicidal action, but rather to errors in our methods, or to the elective action of media, since the differences are relatively slight. The results are more apparent in the tables of Eullmann and Trommsdorff : Keeping at room temperature: Teat Milking Imme- diately 1 hour 2 hrs. 3 hrs. 4 hrs. 5 hrs. 19 hrs. 43 hrs. fr. rgt. beginning 2,400 2,100 2,300 1,900 2,100 2,500 2,100 1,200,000 middle 1,400 900 1,300 1,600 1,800 1,700 1,300 1,400,000 end 700 800 600 900 700 1,100 700 500,000 fr. 1ft. beginning 12,000 900 9,000 7,000 7,000 9,000 6,000 1,000,000 middle 16,000 14,000 1,800 1,400 1,100 1,600 1,600 450,000 end 3,000 1,000 2,400 2,400 3,000 2,700 2,200 1,100,000 hd. rgt. beginning 200,000 118,000 118,000 10,700 98,000 71,000 33,000 420,000 middle 35,000 56,000 37,000 23,000 38,000 28,000 17,000 250,000 end 13,000 11,000 10,000 5,000 2,600 2,400 900 1,400,000 hd. 1ft. beginning 7,000 4,400 4,600 7,200 5,400 5,600 7,200 20,000.000 middle 60 400 160 240 180 240 160 10,000 end 3,400 3,000 3,600 2,600 5,000 2.200 2,100 230,000 In special samples with small bacterial count the comparison was still more striking: Cow No. Teat At room temperature Imme- diately After one c'ay After two days After three days 8 fr. rgt. beginning | 160 40 16,000 2,000,000 middle 80 40 18,000 16,000,000 end 40 80 17,000,000 20,000,000 fr. 1ft. beginning 11.000 9,000 8,000 40,000 middle 1,700 720 40 1,200 end 1,300 600 800 18,000 hd. rgt. beginning 120 400 2,500,000 5,000,000 middle 40 400 18,000,000 60,000 end 180 160 64,000 5,000,000 hd. 1ft. beginning 200 40 78,000 5,000,000 middle 80 500 300,000 6,500,000 end 240 80 50,000 2,500,000 9 f r. rgt. 600 360 2,500,000 25,000,000 fr. 1ft. 40 80 43,000 4,000,000 hd. rgt 450 500 2,500,000 25,000,000 hd. 1ft. 40 240 9,000,000 innumerable Germicidal Properties. Trommsdorff and Rullmann conclude from these and other experiments that the bacterial content of milk does not increase at room temperature in the first period following the milking. "On the contrary in some of the samples inside of the first 5 to 7 hours a pronounced diminution of the bacterial number was observed, which was still more pronounced in the following period so that in a great number of cases the bacterial number, after 1, 2, and even 3, and in one case even after 5 days, was found lower than directly after the milking. Where there occurred no diminution in bacteria the bacterial content remained the same as that found after the milking, during 1 to 3 days. ' ' Milk which has been contaminated with numerous bacteria in the earliest periods after milking (dirty milking, filthy ves- sels), shows only to a very slight extent the germicidal phase. At 37 deg. C. the germicidal substances act more rapidly, but the dura- tion of the germicidal phase is shortened (Koning, Eullmann and Trommsdorff). Heating the milk to over 70 deg. C. destroys its germicidal properties. Bauer and Sassenhagen established the absence of complement in ripe milk which gives the impression that in ripe milk the action is elective in the sense that the contained substances constitute food for one microbe and poison for another. Of course, it has been established for even ordinary kinds of bouillon that, depending on their composition, the growth of certain bacterial species upon them has been checked for a time (Basenau). The lecithin contents of raw milk must not be left out of consideration; in certain concentrations lecithin exerts a strong inhibitory action on bacterial growth. Finally the diminution of the bacterial content may be only apparent, as the bacteria may multiply through their sticking together in agglutinated masses, thereby simulating a diminution, a view which is supported also by Bab for colostral milk. After the germicidal action of the milk has worn off the various phases of decomposition of milk set in, beginning sooner or later, depending on the original contamination of the milk. Koning distinguishes seven such phases. The fight of the microbes, their harmonious, or again anta- gonistic relation to each other, results in a predominance of cer- tain species of bacteria in the various phases. First (second phase according to Koning, the first being the germicidal) the proteolytes split up the proteid bodies of the milk, and thereby prepare the soil for the acid producers, which domi- nate the further decomposing phase (third phase according to Koning) ; the milk coagulates. In the fourth phase the alkali pro- ducers partly neutralize the acid again by further splitting up the albumoses of the acid milk with the formation of ammonia. Pep- tonized casein is also attacked. The principal representative of this decomposing phase is the Bacillus fcecalis alkaligenes (Gram negative, no gas formation, no indol, no spores, colors litmus milk blue). Through neutralization of the lactic acid, certain lactic acid bacteria, Bacillus acidi paralactici, Bacillus acidi Ifsvolactici and Micrococcus acidi paralactici liquefaciens, again regain predomi- nance (fifth phase). Up to this stage the higher fungi have played a subordinate part, although they may have multiplied ; now they appear in great masses. The degree of acidity does not hinder their growth. The 11 162 Bacteria in Market Milk Oidium lactis participates principally in the sixth phase of decom- position, although other varieties of moulds, penicillia, and mucors may also play a part. The degree of acidity being diminished by the oidium, bacteria of the fifth phase again commence to multiply, until the seventh phase is inaugurated through the growth of the anaerobic butyric acid bacilli. According to Koning this phase is reached at room temperature on about the eighth day. Grasberger and Schattenfroh designate ;the principal representative of this bacterial group, the Bacillus saccharobutyricus immobilis llquefaciens, a bacillus which is large, thick and stubby, stains after Gram, and forms spores which are located either centrally or at the end of the bacillus. According to Burri the bacillus may be most readily isolated by boiling the milk for several minutes, and allowing it to ferment at 37 deg. C. Besides this butyric acid ba-cillus, other motile forms or bacteria related to the first group may be found. The seventh phase finally passes into the eighth, in which the milk changes into a stinking putrid fluid, in which the decomposi- tion of the food material is completed by the proteus, subtilis, Bacillus fluorescens, and Bacillus mesentericus, besides mould fungi. Of these phases of decomposition of milk the first three, espe- cially the second and possibly the beginning of the third phase, are the most important. In these two phases the proteolytes and the rennet formers are the first to multiply, causing a partly visible precipitation of the casein, but subsequently again dissolving it; further, the precipi- tated protein is immediately partly dissolved when the dissolving tryptic ferment is present to excess. Weigmann collects these bacteria under the name of casease bacteria. On account of their properties of producing peptones from proteid bodies they are called peptonizing- bacteria. Most of them liquefy gelatin/ The group of casease bacteria includes a great number of forms of bacteria, for instance : Staphylococci, small spherical bacteria which occur ubiquitously and in smears from cultures appear as grape-like conglomerates. They stain by Gram 's method. They grow from 0 deg. to about 40 deg. C., and frequently form at 15 to 20 deg. and over, yellowish orange, or lemon-yellow colored colonies. At a low temperature the casease enzymes are especially active. In gelatin stab cultures Staphylococci first develop a nail-like growth; the liquefied gelatin may then dry and at the point of the stab a bell-shaped air vesicle forms, or the liquefaction may progress rapidly and a cloudy layer of liquid with a sediment of Staphylococci stands above the solid gelatin. In gelatin plates the small round colony drops into a cup-shaped depression which also results from the drying of the liquefied gelatin. Besides the formation of albumose and peptones the milk sugar is split up ; Lohnis therefore classifies these organisms with the lactic acid bacteria. The growth of sarcina, which collect into two, four or eight members, etc., is similar, thereby forming bacterial clumps of certain forms. They are also Gram-positive. On account of their occurrence even in milk drawn in the most cleanly manner, their presence in the udder was accepted, and as a matter of fact pathogenic organisms do occur in the group of Staphylococci, which produce inflammations of the udder. Ordinarily however, they only inhabit the milk duct. The putrefactive representatives of the proteus varieties should be classified among the non-spore bearing casease bacteria. They manifest many forms of growth, and include the following representatives: Proteus sopfii, zerikeri, vulgaris, mirabilis, and fluorescens. Putrefactive Bacteria. 163 They are rod-shaped bacteria, motile, non-spore bearing, and Gram-negative. The forms of colonies and characteristics are quite variable, sometimes showing root-like extensions, and at other times branching outshoots. The most important spore-forming proteolytes originate from forage, hay, stable dust and the stable air, and are collected, under the name of potato bacilli, hay bacilli and root bacilli. They are small rods up to the size of the anthrax bacillus, with central or terminal spores, and are Gram-positive. They grow best under aerobic conditions. Gelatin is liquefied, milk is precipitated as if by rennet, and is then dissolved. In these instances the rennet action is at times more prominent, at other times the action, of the caaease appears more prominently. On solid media the colonies have either a wrinkled, slimy appearance, or they are dry, with a fine map-like drawing on their surface; again they may appear like dull glass, gray, grayish-white, yellowish to brownish, delicate and profuse. On the borders of the culture branch-like shoots or ' ' forms of Medusa heads ' ' similar to the cc lonies of anthrax (anthracoid varieties) may be seen. The more important forms are: The Bacillus mesentcricus vulgatus; the potato bacillus (fuscus, graveolens, ruber) ; Bacillus liodermes; Bacillus subtills (hay bacillus) ; the mycoid varieties; the anthracoid varieties, Bacillus ramosus, implexus, radicosus, tumescens, megatherium, tyrotJirix, etc. In fluid media they generally form prolific wrinkled membranes; at times they cloud the bouillon or they may grow in long, stringy filamentous masses. Some of the varieties also form butyric acid ; thus for instance the Bacillus mesentericus changes lactic acid into butyric acid. To this class some of the mycoid varieties belong, for instance the Bacillus butyricus Hueppe and the Clostridium polymyxa. Through the growth of these and similar forms, the formation of albumose and peptones develops, and the further decomposi- tion of the proteid substances is carried out, if possible, with the production of end products such as leucin, tyrosin, ammonia, car- bonic acid, indol, skatol, methylmerkaptan, sulphureted hydrogen, toxic toxalbumins, and ptomaines. For the judgment of milk which is considerably contaminated with bacteria from litter and forage the increased presence of such bacteria is of special importance, since most of them form spores which are not always destroyed at the temperature of 100 deg. C. and higher. The Bacillus prodigiosus, Bacillus fluorescens liquefaciens, Bacillus amylobacter, and Bacillus putrificus Bienstock, may also split up proteins. Their products vary. The bacterial substances which dissolve proteins and split them up, exert their action especially in neutral and alkaline ma- terial, and they are therefore hindered in their action by the pro- ducts of the third phase, the second decomposing phase. The lactic acid producers, however, proliferate only after the necessary requirements for their propagation have been created by the activity of the peptonizing bacteria. Through the activity of the lactic acid bacteria the milk sugar and other varieties of sugar are fermented to dextro-rotary (ro- tates plane of polarization to right) lactic acid, inactive lactic acid, and levo-rotary (rotates plane of polarization to left) lactic acid, depending on the species of bacteria or the conditions under which the special species prevail. The splitting of lactose C^H^O,, is accomplished by an inverting bacterial enzyme, the lactase, through the introduction of water whereby it is converted into 2C6H12O6 known as D-glucose and D-galaetose, which by further splitting break up into 4C3H6O3. 164 Bacteria in Market Milk. The lactic acid yield, however, corresponds even in the extreme cases only to about 98% of the contained milk sugar, as in the mean- time, depending on the variety of the lactic acid producers, the lactic acid itself is again broken up into simpler acids, acetic acid, valerianic acid, succinic acid, and carbonic acid; alcohol, aldehyde and possibly hydrogen result besides the lactic acid. In spontaneously coagulated milk mostly inactive lactic acid is found, or a mixture of inactive and dextro-rotary lactic acid ; but only in exceptional cases is pure dextro-rotary lactic acid found (Gunther and Tierf elder). Kozai mostly found only the dextro- rotary polarizing form. The lactic acid bacteria may also be considered as ubiquitous micro-organisms ; they have been found in straw, hay, fodder, dust, feces and in the air. Proliferating in milk they soon adapt them- selves to the nutritive medium which at first is not quite suitable for their propagation, and finally they form standard varieties for which the milk is especially adapted. Through continued growing a weakening of the acid forming qualities may develop, and the coagulation of the milk may not take place in spite of their growth, the bacteria having become "milk tired." Under these conditions they may show other properties, the bacteria rendering the milk slimy instead of sour, which is known to be the case with some of the lactic acid streptococci. The specific lactic acid bacteria are aerobes, or facultative anaerobes. Lohnis separates the specific lactic acid bacteria into four col- lective groups: 1. Streptococci. 2. Plump short rods. 3. Slender, long, lactic acid bacilli. 4. Micrococci and staphylococci. The most frequent of these are the representatives of the streptococcus group. Arranged into wreath-like, shorter or longer bodies, the individual members are characterized by coccus or oval- shaped bodies. On artificial media they frequently manifest vacuo- lar degenerative forms which change the individual microbes to the size of bacilli. Frequently pure diplococcic forms may be found which at their ends are mostly pointed in a lancet shape. Strepto- cocci grown in milk are composed of individual members mostly in the shape of a figure "8" which lie with their long axis in the direction of the chain proving them to be streptococci that entered the milk after its secretion as compared with the forms from the udder. Some of the representatives form capsules but only under special cultural conditions, as for instance in blood medium, while in other media capsule formation appears to be a constant charac- teristic and occurs especially in old milk cultures. Most of the streptococci are Gram-positive. They possess no motility and form no spores. On solid media the colonies usually remain delicate and small; in fluid media Streptococci. 165 Fig. 24. ^ <*"" v'-" $^ '-"•£$ '•^.' A / ~X>^ *'.. .-•K ../. ., -^ v- -¥&* the growth takes place either with general clouding, or with the formation of flakes and tufts and the bouillon then remains clear. The optimum temperature of the growth lies between 10 and 42 deg. C. Most of the varieties are facultative anaerobes. Coagulation of milk may take place with acid formation, or in spite of acid formation it may not coagulate, or again coagulation may take place with only slight acid formation, which in all probability depends on the formation of substances having the properties of rennet. Gas formation has never been observed by the author, but it is supposed to occur; slime formation is typical of some of the species. Lohnis classifies the varieties of streptococci found in milk as: 1. Those which coagulate milk and form gas: Micrococcus sornthali (Adametz), Streptococcus "a" from Kefir (Freudenreich), Streptococcus memelcnsis (Leichmann), Streptococcus caucasicus (Migula). 2. Those which coagulate milk but form no gas: Streptococcus guntlieri, Bacillus lacticus Kruse, Bacillus lactis Lister, Lactococcus 'beijerink, Bacillus acidi lactici Groten- feld, and the streptococci from Armenian buttermilk. 3. Those which neither co- agulate milk nor form gas : Streptococcus Jcefir, Streptococcus "~b" Freudenreich, Streptococcus soya. 4. Those which neither co- agulate milk nor form gas: some streptococci of cheese, Strepto- coccus inocuus. 5. Those which form slime ; some varieties of the Streptococcus pi/ogenes, leuconos- toc varieties, Streptococcus liol- landicus, Bacterium lactis longi, "sticky milk" producers, Micro- coccus mucilaginosus, Bacterium lactis acidi. 6. Those which in culture form vine or tongue-like shoots. 7. Those which liquefy gelatine. It is well known that the cultural characteristics of the streptococci may readily change : a strong acid forming variety may lose this characteristic by long cultivation in milk, and may become a slime producing variety, so that the distinguishing feature is not absolute. There are transi- tory forms between one and the other type, and one type at some time may change into another type by changing its characteristics. Some varieties also belong to those streptococci (collective name streptococcus or Bacterium lactis acidi Leichmann), which produce volatile substances and alcohol from milk sugar and which at the moment of their development unite into a fragrant substance, a so-called fruit ester, which reminds one of the odor of a certain fruit or fruits; other representatives of this variety produce other substances with odor and taste, which may be described as straw-like, sorrel-like and especially malt-like. The author never succeeded in producing striking odoriferous substances in sterile milk with the streptococci cultures at his command. He, however, does not wish to refute from his few experiments with about 20 strains from various provinces, the possibility of the production of special odoriferous substances by the Streptococcus lacticus. Under certain conditions tremendous quantities of streptococci may occur in market milk from cows which are affected with strep- tococcic mastitis. Special reference has been made in the chapter on affections of the udder, regarding the similarity of these patho- - :•< - - " r Slreptococcits lacticus. 1 X 1000. 166 Bacteria in Market Milk. [Fig. 25. genie varieties, to the probably harmless varieties of lactic acid producers. The collective group of the Bacterium acidi lactici is the second of importance in the lactic acid group and is also invariably represented in milk. While the streptococci produce dextro-rotary lactic acid, among the representatives of Bacterium acidi lactici there are those which produce levo-rotary lactic acid. Since the growth of both species of bacteria depends on the temperature and the method of keeping the milk (in a shallow bowl or in a deep vessel), therefore under certain con- ditions whereby the Bacterium acidi lactici has better chances for vegetating (aerobe, optimum at about 37 deg.), it produces levo-rotary lactic acid and in soiling in deep vessels (the Streptococcus lactis acidi is a facultative anaerobe and grows well at 20 deg.) it produces dextro-rotary lactic acid. These results have been confirmed by the observa- tions of Conn and Esten. The findings of Heinemann, Thiele and Holling that at incuba- tor and room temperature, in milk drawn under specially clean conditions, ' ' d-lactic acid ' ' is formed only at the beginning, are of interest. Against the practical utilization of these observations, namely the conclusion that the ex- clusive presence of dextro-rotary lactic acid is dependent on the specially clean procurance of the milk, are the investigations of Pere and Harden, who claim that the nature of the acid formed depends not only on the producer but also in the case of the same producer on the character of experimental procedure; thus for instance one and the same strain of Bac- terium coli produced lactic acid showing opti- cally different properties when cultivated under different conditions, aerobically or anaerobi- cally, etc. The Bacillus acidi lactici (Hueppe) group also does not rep- resent constantly uniform species, but it is a collective name which unites all bacteria with especially Representatives of the coli-aerogenes Strong acid forming properties that group, from a culture. 1 X 800 (Bac- Ipovi toward <3 tllP Polon and P^TIP- tcrium plilcgmasiac ubcris, after Kitt.) -ledll L clHU cially to the aerogenes species. The Bacterium acidi lactici Hueppe and the Bacillus pneumonia? Friedlander are similar to the coli-aerogenes species. They are plump, mostly Gram-negative, from cocci to short rods in appearance, forming individually longer rods and short thread-like filaments; they grow luxuriantly, forming moist or almost dry indented colonies with a slimy or jelly-like consistence. In dextrose media usually a strong acid formation takes place. On potatoes the growth is either luxuriant with gas bubbles, or brownish and thin, or transparent. The odor varies, being either disagreeable or pleasant, or at times even odorless. From this description it may be seen that a great number of bacteria are united in this group which are classed by Lohnis as follows: 1. Type of the Bacillus acidi lactici Hueppe. Gas formation with milk coagulation. To these belong the Bacillus aerogenes, Bacillus "a" Guillebean, Bacillus "b" Freudenreich, Bacillus laevolacticus, Bacillus acidi lactici Gortenfeld, also the lactic acid bacilli of Fokker and others which possess differing characteristics, as for instance the formation of esterlike odors, cheesy odors, etc. 2. Milk coagulation without gas formation. Bacillus Umbatum (acidi lactici) Marpmann, without special tendency to deep growth. Bacillus acidi aromaticus, Bacillus granulosum, crenatum, spirans and ramificans (Weiss) and others. 3. Gas formation without coagulation of milk, producing white, circumscribed, hemispherical colonies, as for instance the Bacillus pneumoniae Friedlander. 4. Neither gas formation nor coagulation. Cheese Bacteria. 5. Neither gas formation nor coagulation, but slime formation is present, for instance the Bacillus capsulatus, Bacillus viscosus, Bacillus capsulatus mucosus, Bacillus lactis pituitosi, Bacterium ozacna. 6. The tendril-shaped colony forming Bacillus aerogenes capsulatus. 7. Without gas formation, but with liquefaction, for instance the Pneumobacillus liquefaciens bovis. A decisive separation of these types cannot be strictly ac- complished anywhere in the entire group. Each type has indi- vidual or several representatives which show transitory tendencies towards one or the other group, and the entire group is closely re- lated to the colon aerogenes group, but in the latter, there are no such pronounced lactic acid formations. The colon group is motile, the aerogenes group non-motile. While the colon bacilli form indol and split up proteids, these properties are absent in the aerogenes group. Milk is coagulated with gas and acid formation, in which the casein usually remains on the surface of the pressed-out serum in the form of a spongy coagulum. The milk receives at the same time an unpleasant, slightly offensive odor, and a salty, bitter taste. Some varieties change the milk in this manner without the special changes being perceptible. The several groups adapt themselves very rapidly to their surrounding conditions, for instance to the cultivation on cabbage or turnip media, and when transferred to milk they impart to these the well-known changes of taste. Both bacteria are found in the intestines of sucklings (Escherich). It is an important fact that most bacteria of this group are destroyed in a short time at 65 deg. C., so that their occurrence in pasteurized milk (in bottle pasteurization) may be an indication that the heating to which the milk has been subjected was insufficient. According to De Jong and Graaf some varieties of the colon group resist heating to 70 deg. C. for a short time. The slender "cheese bacteria" of the third group of the lactic acid producers are again divided into: 1. Those coagulating milk with gas formation, for instance Bacterium casei (Freudenreich), Bacillus caucasicus of Kefir (Freudenreich) ; 2. Those coagulating milk but with no gas formation; 3. Those producing gas but no coagulation ; 4. Those showing neither of these characteristics; 5. Those producing slime; 6. Those growing in tendril-shaped colonies. The bacteria of this third group are almost invariably non- motile, and sporeless, mostly without capsules. They are Gram- positive. The fermentation of sugar varies. In milk they mostly form levo-rotary lactic acid, while the other two varieties are rarely produced. Some peptonize proteids ; their growth is favored more by aerobic than by anaerobic conditions. Bacteria in Market Milk. The bacteria do not grow well in milk but they are found in cheese, in the oriental varieties of sour milk and in sour food. These bacteria are of only slight importance in the ordinary lac- tic acid fermentation. They prefer higher temperatures and pro- duce fermentation only in the absence of oxygen, although their growth is prolific even in the presence of oxygen. As representatives of this group the Bacillus panis fermentati occurring on sour bread should be mentioned, and the Bacillus delbrucki found on sour food. Some representatives of these "cheese bacteria" are capable at high temperature (40-50 cleg. C.) of producing and withstanding large quantities of lactic acid, up to 1.5%, and even to 2 to 2.5%. Lohnis classifies the staphylococci as the fourth most widely distributed bacterial group of the lactic acid producers, but on ac- count of their peptonizing characteristics they might better be considered with the casease bacteria of Weigmann. Their proper- ties have already been described during the discussion of bacteria of the first decomposing phase. The staphylococci may also be separated into : (a) Those coagulating milk and liquefying gelatin. (b) Those which only liquefy gelatin. (c) Those which only coagulate milk. (d) Those which posses neither of these properties, nor form slime, produce gas, nor form tendrils. The species mentioned by no means exhaust the number of species and groups which are capable of producing lactic acid. Thus the anthrax bacillus splits up sugar and starch into lactic acid, and also forms acetic acid (Napias), and formic acid (Iwanoff). Feinberg demonstrated for the diphtheria bacillus the capability of splitting up milk sugar with the formation of alcohol, aldehyde and volatile as well as non-volatile acids. The bacillus of malignant edema, according to Kerry and Frankel, in the anaerobic fermentation of grape sugar, produces ethyl alcohol, formic acid, butyric acid and lactic acid ; cholera vibrios and related organisms form lactic acid ; for instance the vibrio of Asiatic cholera, the Vibrio proteus (Finkler and Prior), Vibrio ma-ssauah, vibrio danubicvs and others form levo-rotary lactic acid, the vibrio of Deneke forms dextro-rotary and Vibrio berolinensis produces an inactive lactic acid. The formation of these lactic acids, however, does not depend on the bacillus alone. The Oidium lactis, a milk mould, and others, are also capable of producing lactic acid from milk sugar. Some of the lactic acid forming varieties are rare in milk, others may accustom themselves to milk so rapidly that they form the typical acidifying flora, the presence of which under certain conditions may be desirable since by their multiplication the vegetation of the harmful peptonizing bacteria and of the producers of butyric acid is inhibited. With this we leave the most important varieties of bacteria which are responsible for the normal spoiling of milk, and will briefly discuss those varieties of microbes belonging to the bacte- riology of milk and milk production which always occur in market milk. Milk also invariably contains butyric acid bacilli. Their pre- dominance is inhibited by the lactic acid fermentation. Butyric Acid Bacilli. 169 The specific butyric acid bacilli are obligatory anaerobic or facultatively anaerobic, that is they thrive best in the absence of oxygen ; there are, however, aerobic bacteria which are capable of forming butyric acid, for instance several of the above-mentioned peptonizing varieties, the hay and potato bacilli, which without specially attacking the milk sugar, form butyric acid from the products of the split proteids. The individual varieties show varying properties toward the different kinds of sugars, as well as toward the formed by-products, as butyl alcohol, isobutyl alcohol, formic acid, acetic acid, propionic acid, valerianic acid, carbonic acid, hydrogen, etc. The obligatory butyric acid producers are rods, chains or threads, with either a plump or a slender form; they possess oval or roundish polar or central spores. The bacteria Fig. 26. Blackleg bacilli. 1 X 1200. (After Friedberger & Frbhner.) frequently form so-called clostridium forms, and especially in starch-containing media they take up granulose; therefore certain parts of their body or even the entire bacillus may be stained blue with iodide of potassium. The representatives of this group are known to pathologists as producers of blackleg, gaseous phlegmons, malignant edema, bradsot, botulism, and tetanus. These bacilli at times are capable of energetically forming butyric acid, and at other times less in- tensively; at the same time peptonizing ferments (tryptic) are formed, which become active in the absence of acid. Generally motile and non-motile forms of butyric acid bacilli are distinguished in milk. The latter form, of the granular Bacillus saccliarobutyricus is considered by Grasberger and Schattenfroh as a developing form of the motile, spore-forming variety which takes up granulose, forms toxins, and attacks lactic acid. According to their properties they may be divided into those which form butyric acid principally from certain carbohydrates, Bacteria in Market Milk. as for instance the blackleg bacillus, the non-motile butyric acid bacillus (Grasberger and Schattenfroh), and the bacillus of gaseous phlegmons (Frankel). The others are producers of putrefaction and split up the proteids into forms from which volatile fatty acids develop. Obligatory fat-splitting bacteria may also, although less fre- quently, be found in milk, as for instance the Bactridium lipolyti- cum (Huss), through the growth of which the milk acquires a rancid taste. The Bacillus fluorescens, Bacillus prodigiosus and others, for instance certain mould fungi, may also produce fat-splitting enzymes. Actinomyces form the transition organisms which lead from bacilli to higher fungi. These fungi form long threads with true branchings. Widely distributed on grasses and especially grain, as well as in the soil, they are of course also contained in manure and litter, and may occur in milk and milk products, butter or cheese, and multiply therein. As is the case with all milk bacteria, among the actinomyces there may occur forms which under certain conditions such as wound infection, produce diseases (chronic suppurations). Some varieties of bacteria classified by Lohnis as lacto-bacilli, as for instance a microbe isolated by Chatter jee from "Dadhi," (Indian sour milk) Streptothrix dadhi and several bacteria which were found in Mazun, Yoghurt and in the Montenegrin "Grusa- vina" and "Kysla varenika," appear from their morphological properties, to belong to the streptothrix (actinomyces). Finally it will be advisable to discuss the higher fungi, yeasts and moulds which occur in milk. They cause a slight alcoholic fer- mentation of the milk; not all varieties however attack the milk sugar, although a great number of the most varied fungi and yeast are found in milk, for instance penicilia, mucors, aspergilli. By far the most frequent fungus in milk is the Oidium lactis, tinder which name are collected all mycelial forms, whose radiating mycelia carry hyphae, that break up into small, rectangular, cylin- drical members, the so-called oidia, which in proper media again grow out into a mycelium. The growth of the oidium varieties gives the surface of the cream layer a yellowish-white, velvety, fre- quently wrinkled appearance, which later may take up a glassy transparent appearance. Oidium lactis causes fermentation in sugar-containing media, and develops carbonic acid and a slight amount of alcohol. A pleasant aroma results from cultures in dextrose but in the split- ting up of saccharose, lactose and maltose, an intensive cheesy odor develops. Besides sugar, proteids if present are split up. Therefore in the zones of growth of the oidium varieties a pepton- ization is manifested in the milk. Lactic acid is also produced and later again disappears. Besides the oidia there may also be found the closely related Buttermilk. moniliar varieties wliicli at times grow like the oidia with a typical mycelium, at other times it is a sporulating fungus (Mon. varia- bilis; candicans, etc.) ; also varieties of mycoderma, which always multiply in a longitudinal direction, by the protrusion of daughter cells which continuously bud out new daughter cells and these con- tinue to grow in the already established direction. In the preparation of certain fermented forms of milk which are frequently desired in certain sour milk preparations, the sporu- lating fungi which multiply in all directions of space through sporulations are of importance. Through their activity, that is through the formed enzymes, the milk sugar is split up into dextrose and D-galactose, and ulti- mately the dextrose is split up into alcohol and carbonic acid. Milk may contain saccharomyces varieties, which form endospores and torula varieties, whose daughter cells no longer separate in all directions but arranging themselves into rows form mostly spheri- cal shaped or sausage-shaped buds, and have no endospores. Milk Preparations, Buttermilk, Etc., Produced By Special Fermentation. Many varieties of foreign buttermilk or sour milk have recent- ly become known in this country. Especial dietetic value is attri- buted to them ; as to whether they possess advantages over our own buttermilk or not is not yet known. The author believes that our native buttermilk possesses the same advantages provided it is prepared with the same care as the buttermilk known as Yoghurt, Mazun, Leben-raib, Gioddu (Sardinia), etc., besides many of the foreign milk preparations which are marketed under various names frequently contain nothing more than native varieties of our lactic acid streptococci and certain cheese bacteria. Yoghurt is the buttermilk of Bulgaria. It is prepared by adding to the milk the ferment maya after the milk has been boiled down to half of its volume, and cooled to about 50 deg. C. The mass is then kept at 40-50 deg. and after 10-14 hours the Yoghurt is finished. The necessary fermentation temperature is obtained through cooking boxes, or covering the hot vessels with non-con- ducting material (woolens). Weigmann in his "Mycology of Milk" quotes the verbal information of KostoiT from which it may be seen that the concentration of the boiled milk is not always car- ried out in Bulgaria, but a ferment (Maya, in Bulgarian Podkwas- sa) is stirred up with a small amount of boiled milk, which is added to the milk and kept at 45-48 deg. C. If a sufficient amount of fer- ment is added the Yoghurt is finished in from 3%-4 hours. It is cooled for 1-2 hours, and may then be consumed. According to information obtained by the author there is an- other method of preparation in Bulgaria which is carried out by the dairymen, and produces a primary Yoghurt. ^ According to the description of Marcoff, to whose kindness I am indebted for this 172 Milk Preparations. information, the dairymen take a widely grown herb (name was unknown to Marcoff), which they crush up in small linen sacks. A small quantity of the juice is then squeezed out and added to the raw milk, whereupon without further treatment the coagulation of the milk takes place within a few hours. From this preparation the Yoghurt then may be prepared. According to this description the primary juice action may be attributed to a vegetable rennet. With the plant juice other bacteria also enter into the milk, the product of which is represent- ed by their elective cultivation. As effective bacteria in the production of Bulgarian butter- milk are considered: 1. A lactic acid long rod, which belongs to the acidophilic bacilli of the intestinal tract : the Bacillus bulgari- cus. This is the aroma producer of Yoghurt. The Yoghurt also contains streptococci of lactic acid; yeasts are not desirable (see Table IV). The same conditions exist in the Armenian Mazun, a very aromatic preparation of buttermilk, which is prepared from boiled cow milk, buffalo, sheep or goat milk. Diiggeli demonstrated that satisfactory Mazun (Tartaric Katych) contains principally three varieties of microbes, a streptococcus, a long rod-shaped lactic acid bacterium, and yeasts, the activity of which produces the aromatic bodies, besides a slight amount of alcohol and carbonic acid. Leben-raib, according to Bist and Khoury contains five micro- organisms, among them being two varieties of yeast and two lactic acid producers. The buttermilk is prepared in a similar way to Kefir, by using cow, buffalo or goat milk. Kefir has been used for a much longer time and therefore is more generally known. It contains alcohol and is very rich in car- bonic acid ; it has a pleasant, slightly acid odor and taste. It con- tains the milk proteins split up to some extent (Hueppe) in ad- dition to alcohol and carbonic acid and a slight amount of glycerin, succinic acid, butyric acid and acetic acid. Kefir is best prepared from skimmed milk, since in full milk, cream clumps may readily result and the Kefir thereby becomes rancid, which diminishes the consuming value. According to Freu- denreich the fermentation is principally produced by four varieties of organisms which include yeasts, two streptococci varieties, and one microbe described as the Bacillus caucasicus. The yeasts are the Torula kefir and Saccliaromyces fragilis, both of which ferment lactose. Investigations which have been re- cently conducted by Kuntze showed that the bacteria of Kefir con- sist of an aroma-forming rod which produces casease and alcohol, and a lactic acid long rod which at first acidifies the milk and then renders it alkaline. They are the Bacillus esterificans Maassen and the Bacillus kefir classified by Kuntze as belonging to the group of butyric acid bacilli (cited by Weigmann). These bacilli inoculated Kefir. 173 into milk, together with ordinary lactic acid producers, living in symbiosis with yeast, form granules which grow to raspberry sized clumps and nodes. Sponge-like masses of ruffle-like appearance, the so-called Kefir kernels result, in which are included the neces- sary varieties of microbes. In dried condition these kernels are of the size of millet seeds, but after treating with warm water or warm milk they swell and proliferate in the milk up to the size of a fist. The small young kernels are the best, as the larger readily degen- erate, become slimy and crumbling as compared with the elastic granules of more recent development. In order that they may again produce good Kefir they must be subjected to treatment by washing, drying in the sun, etc. The Kefir kernels may be purchased. The origin of the Kefir kernels, that is the microbe colonies which are clumped in the kernels, is not known, but the primary de- velopment may have some connection with the method of ferment- ing milk in containers made out of goat skins. If it is desired to prepare Kefir it is necessary to first obtain the kernels which may be purchased. The Kefir kernels are allowed to soak in boiled or lukewarm water, and then they are transferred several times (3 to 5 times) from one warm milk into another, the milk being poured off every 3 or 4 hours. The utilizable Kefir kernels increase in size during this time through further swelling, and as a result of becoming lighter in weight through absorption of carbonic acid they rise to the sur- face of the milk, whereas kernels in which one of the varieties of microbes for some reason or other became destroyed and degener- ated remain on the bottom of the vessel. If the degenerated variety of microbes recover through longer treatment with raw milk, and if the proper relation of symbiosis again appears, then these ker- nels are satisfactory for the production of Kefir. This condition is manifested by the kernels rising to the surface of the milk after some days. If a tablespoonful of these kernels is added to about one-half liter of milk and this is allowed to stand for from 8 to 12 hours at 14 to 18 deg. C., with frequent shaking, then a primary or mother Kefir is obtained, from which through further fermentation in corked bottles the Kefir may be prepared ready for use. The ''millets of the prophet," the Kefir kernels, are strained through a sieve, and the homogeneous fermented milk is filled into bottles, or from the strained fluid a considerable quantity is poured into a bottle, to which boiled milk cooled to about 20 deg. C. is added. The bottle is then closed and allowed to continue to fer- ment for from 24 to 28 hours, at from 12-15 deg. C. In this process the casein and serum separate but may be readily homo- genized by shaking. The Kefir is then ready for consumption, and represents a thick, sour, aromatic fluid of a pungent taste, with a remarkable nutritive value. A similar product is prepared by the nomadic population of 174 Milk Preparations. Southern Bussia, Siberia, and Central Asia, which represents a milk wine made from the milk of mares and asses, and which is known by the name of Kumys. In the preparation of Kumys, alcohol and carbonic acid fermentations are the principal processes. After long fermentation Kumys contains up to 2 % of alcohol and \% or more carbonic acid. Bacteriologically Kumys is of similar composition to Kefir containing yeasts, lactic acid, streptococci, and the Bacillus kumys (Schipin), which is a facultative anaerobic microbe which splits up milk sugar with the formation of lactic acid and alcohol and peptonization of the proteids. These act together and form after several days a delicious drink of white color, and creamy con- sistence. Special varieties of milk, containing much sugar, are best adapted for the preparation of Kumys but cow's milk is the least desirable. Gioddu, the buttermilk of Sardinia, is prepared from boiled milk cooled to about 35 deg. C. To four parts of milk one part of old Gioddu is mixed, the Gioddu being added to cow, sheep or goat milk. The fermentation is produced by the Bacillus sardous in symbiosis with the Saccharomyces sardous. According to Grisconi the Bacillus sardous belongs to the streptobacilli. The preparation of buttermilk constitutes an important branch of the utilization of milk in all countries. In northern Bavaria the milk is usually set in large earthen pots and allowed to undergo voluntary fermentation. In southern Bavaria and in the Bavarian forests the "fall milk" is utilized for the preparation of the "sour soup." By keeping buttermilk and continually adding sour skimmed milk to it a fermenting product is obtained which is thickened by the removal of the whey (Herz). In Sweden and Norway a milk product is known under the name of "thick milk" (Tatmjolk), which is produced by slime and lactic acid producing bacteria which vegetate on the leaves of the butterwort (Pinguicula vulgaris). The leaves of this plant are placed on the bottom of the milk vessel and milk poured over them, whereupon the milk becomes so thick in several hours that it must be cut with a spoon or knife in order to be taken into the mouth (Weigmann). New milk may be inoculated with the residue of old milk. The necessary preparations of bacteria for the making of special forms of popular buttermilk may at the present time be purchased in the market. In using any of these ' ' ferments ' ' the directions for use should be carefully followed, since at a tempera- ture either too high or too low an overproduction of undesirable bacteria may readily take place which would make good results im- possible. Even if the directions are carried out most accurately, the propagation from milk to milk may be a failure since the biolog- ical properties of the bacteria are not absolutely constant; the microbes "grow wild" and their pleasant qualities are lost, or they may change, assuming undesirable properties. Table IV. Butyric acid bacilli in boiled milk, kept for two days at 37°C. 1X1200. n. Film of Yoghurt. Bacillus bulgaricus and lactic acid streptococci. Gram-safranin stain. 1 X 1200. Ernst, Milk Hygiene. Bitter Milk. Porcelain vessels and bottles made from glass free of lead are most suitable for the preparation of buttermilk, since the butter- milk may extract lead from enameled earthenware and from pots whose glazing contains lead in its composition. According to Chlopin 0 . 84 mg. of lead was extracted from 100 gm. lactobacillin-buttermilk ; in a second portion (300 gm. butter- milk) which was five days old, the amount reached 7 . 86 mg. Briick- mann obtained similar results : 300 gm. of ordinary buttermilk con- tained after four days 4.2 mg., and after six days 5.7 mg. of lead, when this product had been prepared in pots with lead-containing glazing. Defects of Milk. Bacteria produce certain modifications in milk which partly on account of their frequency are designated as normal processes, or again others appear which are less frequently observed, occur- ring only under special conditions and therefore are known as milk defects. The modification, as has been seen, may be even desirable, as for instance in cream souring and cream ripening for butter making, or in the preparation of Kefir, Yoghurt, and buttermilk, or it may be undesirable and injurious, spoiling the milk, and having a disturbing influence on milk utilization, espe- cially in its use for drinking purposes. Among the changes in milk there are those which appear fre- quently, and others which are very rare. Under conditions which favor propagation of peptonizing bacteria (staphylococci, sarcina, anthracoides, mycoides, mesen- tericus varieties, fluorescens, pyocyaneus, etc.), the milk attains a bitter taste. For instance if uncooled milk is filled into cans which are immediately closed it " suffocates," acquiring a strong stable odor which may even reach a putrid character, causing a solution of the casein by reason of which- the milk no longer coagulates ; or the appearance of a bacterial rennet produces a rennet-like pre- cipitation of the casein, and the milk coagulates without turning sour. It is "sweet-coagulating." By the action of peptonizing micrococci, which in part are psychrophilic the development of a bitter taste may occur in thoroughly cooled, and even in excessive- ly cooled milk. The bacteria of the colon group when the condi- tions of their propagation are favorable may produce an odor in milk ranging from aromatic to rancid, or some varieties of this group which have grown on mangels may confer the odor of man- gels to the milk. A bitter taste in milk may also occur from the feeding of foods containing bitter substances, thus for instance from the feeding of lupins, vetches, mangels, camomile, beet leaves, wood-fern, raw potatoes, mouldy or spoiled hay, straw, etc. It may however be accepted that the development of a bitter taste in milk usually 17G Defects of Milk. results from its contamination with varieties of bacteria vegetating on food substances, which enter into the milk directly from the stable air or indirectly with the manure and litter. They then con- vey to the milk this altered taste. Experiments to confirm these views have been undertaken quite recently by Weigmann and Wolf (Kiel). Defective flavors are frequently present in milk. Of 1,000 retentions made during 1909 in Munich, 90.50% were on account of souring, 14.6% on account of soapy taste, 18.25% rancid, 2.19% fecal contamination, 8.76% oily, 1.46% bitter, 2.92% granular, 2.19% sweet-coagulating, and 1.46% on account of slimy conditions. Eepresentatives of the colon-aerogenes group may actually be cultivated until they become aroma producers if they are allowed to grow for instance upon media made from rape leaves. If an adaptation of these and other bacteria to the ingesta within the gastro-intestinal canal is admitted, then an acquisition of certain other properties, depending on the consumed feed, is readily conceivable. Weigmann and Ritland and Jensen demonstrated such "rape bacteria" in milk having a rape-leaf taste; the milk at the same time had a stale taste and an odor of dish-water. The author ob- served a distinct phosphorus taste in cases in which the milk was placed without being cooled into unclean or poorly cleaned covered cans. Animal and fecal odors result when freshly drawn milk is placed into covered cans without airing and cooling. In these cases the vegetation of anaerobic and facultative anaerobic bacteria may play a part, and the temperature may have an effect on the bacterial elective conditions. The milk attains a taste like animal viscera if it contains bacteria of the mycoid, megatherium or fluor- escens group. Fishy taste of milk may result from pasturing cows on marshy meadows which have been inundated. In these instances the pres- ence of various other varieties of bacteria should be taken into consideration. The multiplication of the Bacillus lactis saponacei (Weig- mann) and the Bacillus sapolacticum (Eichholz), produces a soapy condition of the milk. The milk attains a sharp, rancid, soap-like taste, and when cold it reminds one of valerian ; in a warm state it has a sharp, soapy odor. In shaking such milk a fine, vesicular, persistent, tenacious foam results. The change appears in thor- oughly cooled and excessively cooled milk, and in the cold season of the year and in cold rainy summers the bacteria are psychro- philic, originating from the feed and straw. The Bacillus lacticus saponacei grows well at room tempera- ture, liquefies gelatin, and produces a slight yellowish shimmering coloring matter; the growth is aerobic. The Bacterium sapolacti- Blue Milk. 177 cum grows similarly. It is not supposed to liquefy gelatin. The nutritive media become fluorescent. The propagation of butyric acid bacteria causes rancidity of milk, as does likewise the multiplication of bacteria which spilt up fats, for instance the Bacterium lipolyticum. The appearance of the milk defects here mentioned may sometimes be confined to the product of a single individual in the stable. The milk of one or of several co\vs may manifest these defects which may be retained persistently in spite of changing the feed and disinfecting the stable. Weigmann mentions a case in which, with uniform feeding and care of the animals, the milk of only one cow developed a fishy odor, and to such a marked extent that the milk of the entire herd became fishy (possibly the udder of this cow was diseased). The same author mentions another case which occurred on an estate in northern Germany. In that instance the milk of the Montavania cows in the stable was constantly rancid, whereas the milk of the Holstein cows was faultless, although the animals were all kept under the same conditions. The Bacillus lipolyticum was found to be the dis- turbing bacterium. It is noteworthy that the milk of the Montavania cows was frequently bloody at the same time. Therefore it is possible that the elimination of the aroma bacteria took place from the affected udders, that is, the same bacterium was also the cause of the inflammation of the udder. However, it is more likely that through the secretion of the affected udders conditions were established in the milk from the Mon- tavania animals which favored the propagation of the Bacterium lipolyticum in the milk, or probably the bacteria were present in the milk cisterns of these cows as harmless saprophytes, and the blood content of the milk may be attributed to some affection of the udder (yellow garget), which had no connection with the cause of the rancid milk. It has also been proved that other changes in milk may per- sistently occur in the secretion of certain individuals so that it appears as if the causative agents of the changes in the milk may at times exist as saprophytes in the cistern (Schultze), or that they have at least multiplied in the excretory duct of the cistern. Thus Schultze proved that in the appearance of "blue milk" the defect can only be removed by a thorough cleaning of the sta- ble, animals, milk vessels, and all creamery utensils, and the milk cisterns of the animals must also be treated by antiseptic infusions of the udder. The "blue milk" is produced by the Bacillus cyanogenes, a Gram-negative, aerobic, actively motile, unipolar, flagellated bacillus, with rounded ends, about 0.4/ci thick and 2.4/i long. It is also known as the Bacterium syncyaneum Hueppe (Heim). Growing in sour milk the bacillus produces sky-blue to indigo-blue spots which gradually become confluent. The bacillus attacks the casein, and produces alkali besides a coloring substance, the triphenylrosanilin (Erdmann), which, depending on the reaction of the nutritive media, appears greenish or pale blue, violet or indigo-blue, or blackish-brown. The Bacillus cyanogenes in itself is colorless. The coloring is less typical in sterile milk; a dirty bluish-gray discoloration with a reddish hue of the cream occurs, the color gradually diminishing in the deeper parts. Indigo-blue spots develop only in sour milk (Heim). The changes which occur in milk appear to be especially fre- quent in certain localities ; in others they are more rare and appear to have a connection with the properties of the soil. Pastures rich . in clover are supposed to favor the appearance of the defects while in woodland pastures they have not been observed, or at least only exceptionally. This would explain why the defects occur in cer- 12 178 Defects of Milk. tain periods during the feeding of the incriminated feeds, or while the cows are feeding in certain pastures. According to observa- tions they are observed more frequently in the fall, and during wet, foggy weather than during other periods. These defects persist tenaciously in creameries and dairies and can only be eradicated after a thorough determination of their origin. Disinfection of the milk room and utensils with milk of lime and hot soda solution, and extending this disinfection to the stable in association with cleaning of the animal and possibly an- tiseptic infusions of the milk cisterns may yield the desired results. Another organism causing "blue milk" is the Bacterium cyan- eofliiorescens (Zangemeister). It is actively motile, bipolar, flag- ellated, grows on gelatin in the form of whitish colonies with in- dented borders and produces a fluorescent coloring matter in the nutritive media. The culture has an odor of trimethylamin and putrid fish. The bacteria produce dark blue spots in milk which change to a sky-blue color after coagulation of the milk. Other blue bacteria are those which occur on hay dust, in water, and in sewage in the vicinity of cheese factories, in ditch-water, and also the bacteria cultivated by Voges, Claessen, and Beijerinck which have been described under the names of B. cceruleum, B. indigan- aceum, B. cyaneofuscum. According to the observation of Weigmann and the descrip- tion of Hallier certain hyphomycetes may also possess the faculty of producing a blue coloration; this is accomplished by the ac- tion of the blue coloring matter which they harbor. In the zone of the milk supply of Munich ordinary milk de- fects occur very rarely; the author observed them in only one dairy, and was able to trace the trouble to a certain farm. An- other defect of milk occurs much more frequently in the vicinity of Munich, the cause of which, according to the author, has not yet been described. It concerns the production of brownish milk. The bacterium of brown milk appears to be closely related in all its characteristics to the producer of blue milk; it is 2.4/x. long, 0.5/u, broad, unipolar, flagellated, actively motile, Gram-positive and remarkably resistant to drying. In gelatin it grows especially well aerobically as a fine, iridescent deposit, which later becomes somewhat thicker, turning to a chestnut brown color. The oxygen zone of the lactose gelatin retains a saturated brown to deep brownish red discoloration, the nutritive media becoming alkaline to litmus. A culture of the brown milk organism may be readily produced in milk by rubbing traces of the culture of milk having such a defect upon the bottom of a large Petri-dish, and pouring over it fresh (not sour) milk. In most instances after 15 to 20 hours ocher-colored" to sepia-brown spots develop in the cream layer, which enlarge and coalesce, conveying to the milk a milk and coffee-like appearance. After coagulation the superficial layer of milk again liquefies ; whether this is brought on by the bacteria of brown milk alone or by peptonizing bacteria which multiply especially well when mixed with the bacteria of brown milk, which render the media alkaline, has not yet been established. The skimmed milk is not discolored by the Bacillus fuscogenes. The brownish color gradually diminishes from the surface down and at a depth of 5 mm. it disappears. If the milk is allowed to sour the appearance of gray, orange- red, red, yellow, green-fluorescent and violet spots may frequently Red Milk. be observed, which cause a glassy, transparent thickening of the wrinkled yellowish-white velvety layer of the oidium covering, or they penetrate into the depth of the jelly-like layer of the milk. Thus under certain conditions the Bacillus violaceus, Bacter- ium janthinum, Bacillus lividus, and Bacterium amethystinus, a water organism, may appear in violet spots (Schroeder, Zopf, Maze, Fliigge and others). Greenish-yellow spots and discoloration of the entire sour milk are produced by the Bacillus fluorescens which varies greatly in its characteristics, at times liquefying gelatin, again only dis- coloring it. It is a short rod with motility, but without spore formation. A red coloring matter is produced by the Bacillus erytliro- genes Hueppe, which coagulates milk, but liquefies it later through peptonization, coloring the whey red. According to Gruber a flagellated short rod, the Bacillus lac- torubefaciens is supposed to produce a slimy condition of milk with the formation of a red coloring matter. Other bacteria such as the Micrococcus cerasinum (Keferstein), the Sarcina rosea, the Bacillus prodigiosus and others, form red spots. Red varieties of yeasts have also been found. The author demonstrated through the examination of a dirty and dry milk pail that the layers of color which adhered to dif- ferent parts somewhat like red varnish consisted of blue-red yeasts which had grown on the dried milk residue. The accumulation of color was present in the yeast cells proper, which on examination showed a reddish transparency. Their attempted cultivation was unsuccessful. The discolorations of milk may vary from red, and pink, to rust-color and orange. Yellow coloration sometimes only of the cream, at other times of the entire milk is caused by the Bacillus synxantlvus (Schroter), the Sarcina lutea, the Sarcina flava, and Bacterium fulvum and others. Wild yeasts and moulds, which have been observed by the author may also cause a yellow coloration of sour milk. The Bacillus fluorescens, may at times cause a yellowish-green discoloration. Other bacteria again show the action of their vegetation by the development of a tenacious slimy consistency of the milk. Strains and varieties of the peptonizing bacteria in which acid for- mation is dissipated and the peptonizing action of which retracts against the properties of producing rennet-like substances, may in a few hours cause a casein coagulation, and thereby convey to the milk a granular consistence. This defect is relatively rare, and on the contrary the milk may become non-coagulable, slimy and bitter. More frequently, especially by keeping the milk in a warm place, a change of the milk to a slimy consistence may be observed. The action of the slime-forming bacteria may appear in two forms, 180 Defects of Milk. and render slimy either the entire milk, or the casein is precipitat- ed and only the whey develops a strong tenacious, stringy consistency. The cause of the slimy condition may be produced either by a slimy change of the sugar, which is accomplished with the form- ation of a high molecular weight body, the galactan or the viscose, or by the swelling of the bacterial capsules which form a mucin- like substance. The best known producer of slimy or stringy milk is the Strep- tococcus hollandicus, the cause of the "long whey," which is con- sidered by Weigmann as a degenerated streptococcus of lactic acid fermentation. If cultivation of the producer of the "long whey" is continued at 35 deg. C. it loses the property of producing slime, and changes into a lactic acid producer. From various groups of bacteria the following have been proved to be slime producers: Bacterium lactis longi — a strep- tococcus— in Swedish thick milk (Troili Petersson), Micrococcus (streptococcus) viscosus (Schmidt-Muhlheim), Micrococcus mu- cilaginosus from slimy cream (Ratz), and Streptococcus burri from stringy whey. Slime is further known to be produced by the colon-aerogenes group (Emmerling, Schardinger), the Bacillus guillebeau, as well as the Bacillus lactorubefaciens. Adametz, Duclaux, Gruber, Ward, Eckles, and Marshall have also isolated slime producers from milk, whey, food substances, straw, stable air, and spring water. Other defects of milk which are associated with change of consistency (and color changes), are produced by milk drawn from affected udders, which subject has been discussed in the section on "Diseases of the Udder." Considering the living requirements of the special varieties of bacteria, the defects of milk appear to be especially frequent under certain weather conditions and in certain periods of the year. Thus the milk dealers of Munich complained of the appearance of defects of taste, especially in the cool and cold period of the year, and at the time of changing the animals from stable to pasture feeding and vice versa. The cause may lie in the fact that with the beginning of the dry, that is stable feeding, the microbian flora of the intestinal canal and of the forage and the stable air is different from that existing during the period of pasture feeding, and thereby other species of bacteria, aroma producers, contaminate the milk; likewise in certain cold climates and in certain methods of keeping milk the bacteria, excepting the lactic acid producers, find just the requirements which aid them in their propagation. In 1909, the following defects of milk were found among 1,000 samples examined monthly: Various Defects. 181 CM Oi CO' CO • • -CO' • • • -CO -CO' Oi CM • • • l>- • • • . l>- • CO ^H CO 1O TH TH TH Oi "^ CO TH TH TH CM CM CM CTi ' * t>* .... 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