PART 3

THE STREPTOTHRICOSES

The organisms belonging to the genera Streptothrix, Actinomyces, Discomyces and Nocardia as named by various authors have in common the power to produce local chronic inflammation of gradually spreading character and chronic course. Their most conspicuous representative, the ray fungus, is best known as the producer of lumpy jaw in cattle and as an occasional pathogen in man. Other members of the group cause certain lymph-channel disease in domestic animals and pulmonary disease in man. Pathogenic power, it is believed, lies in the ability of these organisms to colonize and irritate, thus producing continuously enlarging tumefactions, no evidence being at hand that any of them produce a toxin either in their surroundings or within their own bodies. Because of their constant irritation, bacterial mixed infection often ensues so that purulent degeneration may occur at the original site of disease and thence may spread via the blood vessels, or by continuity of tissues or, if the mucous membrane of the pharynx be diseased, by the air passages.

The study of the genesis of actinomycosis is by no means a closed one. While it is believed that pastures and fodder carry the organism and that it gains access to the tissues by passing into small wounds that are made by sharp sticks or grain beards, the exact origin of the disease is not understood. The original lesion is certainly trifling and the fully developed one may not be discoverable until it is well under way and causes external deformity. Even when sloughing has occurred, the disease is not very communicable. The method of contraction of lymphatic streptothricosis in cattle is believed to be from other cases _via_ skin wounds or if abrasions be soiled by infective dirt. Just how human beings contract these infections, in the absence of infected cattle is unknown, but for the pulmonary form the route usually followed in tuberculosis is probably taken.

The material of our zoological collection permits few observations of value upon “lumpy jaw” but we have encountered a streptothricosis of kangaroos which may throw some light upon the whole subject and to these cases I shall devote considerable space since no description of it occurs in the literature.

ACTINOMYCOSIS.

This disease has been diagnosed with certainty in two American Tapirs and with reasonable satisfaction in three deer. Two Malayan Tapirs have also had lumpy jaw clinically but the organisms were not found. It is interesting and noteworthy that other zoological collections have observed the disease in this same animal, a fact which suggests the high susceptibility of the tapir to actinomycosis. There are recorded in the protocols a few times sluggish ulcers on the tongue in other ungulates but I am not prepared to label them as actinomycotic since on one occasion smears and sections were studied with great care and nothing found to justify such a diagnosis; nor were there maxillary or pulmonary lesions. Before passing to a discussion of the diagnosis and morbid characters it seems worthy of emphasis that our cases of this disease should appear in one family of Perissodactyla and in one family of Artiodactyla, in the latter not affecting Bovidæ, the family to which domestic cattle belong.

The diagnosis of lumpy jaw depends upon the growth of tumors in the neck and maxillary regions which tend to break down and discharge a thick pus containing “sulphur granules,” little masses of necrotic matter surrounding colonies of the ray fungus. When these conditions are fulfilled, the matter is easily enough settled. This was possible with the tapirs but in the deer the conclusion was not so easily reached and the diagnosis had to be made partly by exclusion. Anatomically the gross and microscopic appearances of lumpy jaw in the tapirs follows the text- book descriptions but our cases in the deer deserve separate comment. Whether or not these differences mean a peculiarity of resistance on the part of the animal or a new variety of streptothrix only further study can settle.

The beginning of the lesion in the deer was in the jaw bone as circumscribed or fusiform swellings appearing on the under surface. Growth usually progressed into the pharyngeal cavity and backward under the ear, but a large tumor stretching down the neck was only observed once. Suppuration and ulceration occurred twice but only once were bacteriological observations possible before death, and then they were negative. The fatal outcome seemed to be due to inanition, possibly because the animal could not eat, for respiratory tract involvement was only present once and then to a trifling degree. At autopsy, actinomyces in ray form were found in one animal only, the diagnosis resting upon histology in the other two. Nor did the degenerated centre of the swelling contain the sulphur granules in any case.

The microscopic characters of the tumors resembled those of giant cell sarcoma and chronic rarefying osteoperiosteitis with areas of round cell infiltration but no granulomata as are occasionally seen in lumpy jaw. I have always felt that a “giant cell sarcoma” with inflammation when seated in the jaw of a lower animal should be looked upon with grave suspicion and be searched diligently for fungi. The organisms could not be found in sections of any of these cases, although present in the pus from a pocket in one. Reference has been made in discussing tumors of the bones in gazelles and opossums to their resemblances to osteofibroma and actinomycosis. The diagnoses were made after long study of the notes and sections. Actinomycosis is usually unilateral while leontiasis ossium is commonly bilateral; the tumors have not broken down nor spread into the neck. In one macerated jaw bone the osteoporosis and hypertrophic periosteitis were comparable to those of the bovine form but the masses were not so extensive as is common for domestic cattle.

Treatment of this disease was attempted in the tapirs but not in the deer; the latter are too nervous to be handled repeatedly with safety to themselves. Following the usual method, potassium iodide was administered in saturated solution on bread, beginning at twenty grains thrice daily and rising in two cases to sixty grains thrice daily. It cannot be stated that any material improvement followed this heavy dosage although in one case the disease was very protracted—some six months, so that it may have modified the progress of the lesion. However, other things were done for the beast so that the effect of any one kind of treatment is difficult to evaluate. It was noted that iodide served to keep the stools quite loose and that its withdrawal was followed by constipation; upon resumption of the drug normal bowel movements appeared. Every soft spot was opened surgically to allow the pus to drain away. A vaccine of _Act. bovis_ was prepared and injected under the hide beginning at 0.5 mg. and rising to 2.5 mg. in five doses after which the animal became so unruly that the injections had to be discontinued. On the whole we are not impressed with the probability of success in the treatment of actinomycosis in tapirs. In the future we propose to try operation and the use of Dakins solution or Dichloramin T.

STREPTOTHRICOSIS OR NOCARDIOSIS OF KANGAROOS.

A fatal disease of Australian marsupials characterized by swellings and ulcerations about the lips, teeth, tongue and cervical tissues is known apparently all over the world by observers of these animals in collections. From commercial shippers of animals, from zoologists and naturalists we have reports that wherever kangaroos and wallabies are exhibited this disease makes its appearance and carries off a considerable percentage of the collection. A fully developed case bears a noteworthy resemblance to lumpy jaw, being called “jaw disease” by non-medical observers. However, it is highly probable that, while the most conspicuous morbid changes occur around the jaw, the agent provocative of the disease is capable of causing different pathological effects and that certain cases of septicemia and gastroenteritis are due to it; Doctor Blair of New York concurs in this opinion. Our study of the problem would inculpate a variety of Nocardia, possibly assisted by certain schizomyces. I have seen in the literature, but unfortunately cannot locate, a reference to an article by a Russian who observed the disease and was convinced that its cause is to be found in a streptothrix[105] obtainable from the necroses in the soft tissues of the jaw, a view entirely in accord with our findings.

The disease is not very communicable because its appearance in a pen need not be followed by secondaries in the mates of the sick beast. It appears chiefly in newly acquired specimens but may develop sporadically in those exhibited a long time and apparently not associated with recent acquisitions. This suggests two or three possibilities. It may be imported by new arrivals, or newcomers may meet a germ to which they are unaccustomed and therefore less resistant, the strange surroundings reducing their opposition to it. Old specimens may have enough resistance to withstand infection entirely or only succumb to large doses. Lastly one comes to the explanation commonly employed for actinomycosis, the presence of the organisms in fodder or pasture, perhaps all the time, but gaining entrance to the animal’s body _via_ wounds made by sharp sticks or the beards of grain.

While circumstantial evidence offers some support to this general idea, it cannot be accepted as proven. I have not made studies of the feed for the purpose of isolation of the streptothrix but cultures from the lips and gingival margin of healthy and infected animals were made for its cultivation. These attempts were fruitless, and without wonder since the germ when isolated from a fully developed case is quite finical in its manner of growth; there are so many kinds of bacterial life that they may easily overgrow the one in quest. So, too, cultures made directly from subcutaneous necrotic areas may not always give a positive growth although smears from the same material may reveal numerous threads under the microscope.

The idea that sharp grasses are responsible for the origin of Kangaroo disease finds a protagonist in Dr. A. S. LeSouef, Director of the Zoological Garden at Sidney, Australia, a gentleman whose judgment carries weight. He writes: “We have found that it is entirely due to getting spear or barley grass in their food; owing to the formation of the mouth, this grass gets wedged in between the teeth and the cheeks, penetrates the flesh and allows the bacteria to get a footing, this in time heaps up on the inside and forms an abscess that bursts exteriorly. Formerly all the Australian Zoos lost animals through this cause, but now, through being very careful not to give any rough spined grasses, we never have the trouble.” Since receiving this letter we have removed straw bedding, and feed only soft alfalfa which is carefully inspected for foreign substances. During this time we have had two cases but the period of observation is too short for final judgment as to the value of feeding grasses without sharp beards and spines. The appearances of cases sporadically without reference to the arrival of new specimens, the low communicability of the disease, its beginning in the jaw in most cases and the prominence of pulmonary and gastric lesions, all seem to support the thought that the virus is received with the fodder or drink. The anatomy of the kangaroo’s buccal cavity favors the collection of material between the gums and cheeks and between the root of the tongue and the molars, while the “hare lip” also affords a crevice in which food particles or foreign bodies may accumulate. These three places seem to be the starting points of most of the cases.

THE COURSE OF THE ATTACK.

Despite careful watching of the exhibition specimens it is often difficult to detect the beginning stages of the disease. Since our last outbreak it has been the practice to examine all kangaroos thrice yearly by catching them, inspecting the buccal membranes, teeth, tongue and nose and by palpating the jugular and sublingual regions. This procedure succeeded in catching one very early case from which the original changes can be described.

The animal appeared in generally good condition but close inspection revealed a “running nose,” a purplish mottling along the gingival margin of one lower jaw below which was a doughy swelling; no internal ulceration had appeared nor was there a visible change in the external contour of the jaw. Within a few days a small fusiform lump appeared along the body of the lower maxilla which spread gradually backward, the nearby soft parts becoming involved very shortly. This particular animal died without ulceration but with evidences of septicemia. Usually at the time that the lump is noticeable the animal loses appetite, becomes inactive and seems depressed; no especial change in the coat need be perceptible although it may be lusterless or at times ruffled. In the cases with great involvement of the cervical tissues, dyspnœa is an early sign but I lay this more to pulmonary disease than to mechanical obstruction of the upper air passages. The loss of appetite is in large part due no doubt to the discomfort of chewing and swallowing in the presence of an inflammatory mass in the neck. The eyes usually remain normal until quite late. No change in the character of the droppings is recorded.

A slightly different course is followed by the cases that have the primary lesion in the “hare lip” and nose; from these the masses along the jaw may be entirely missing. After an initial stage of “running nose” with or without swelling of the upper lips and alæ of the nose, the animal rapidly goes down hill, with dyspnœa, loss of flesh, perhaps loose stools, lusterless eyes and a “dead” coat, a series of signs indicative of a septicemic state not pronounced in the first variety; any form of this infection may however present course and pathology of a septicemic character. If the beast live long enough ulceration may appear on the upper lips or a large area of necrosis between them may be discovered.

There have been in our series two cases, believed to be due to the same virus, which gave a picture of septicemia with pulmonary localization; they will be discussed in detail later but are of interest here because they were not known to be sick until the day before their death. A similar failure to evince signs of sickness is found in the gastrointestinal cases, those with ulcerations in the stomach and perhaps an accompanying catarrhal intestinal inflammation. At most the report will be that the specimen was “off its feed.”

[Illustration:

FIG. 62.—KANGAROO STREPTOTHRICOSIS. ULCERATIVE AND NECROTIZING PROCESS IN “HARE LIP” AND IN MUCOSA AND BONE OF ANTERIOR PORTION OF HARD PALATE; TEETH HAVE FALLEN OUT ON LEFT SIDE. ]

The signs of Kangaroo disease with exception of those applying directly to the nose and jaw are therefore very vague and one is limited to observation of the contour of the head and of the discharge from the nostrils. Because of the indefinite nature of the earliest changes, the duration of the disease cannot be stated with accuracy but from the time that the swellings are perceptible it is not very protracted if no treatment be given. Some cases die in four or five days while others may last up to three weeks and we believe that two of our cases may have been existent longer than that. It is impossible to estimate the duration of the septicemic and gastric forms although the latter, judging by the appearance of the ulcers, are believed to be chronic. We believe that frequent inspection and the precautions as to the character of fodder are the only special hygienic measures indicated.

THE INCIDENCE OF THE DISEASE.

Not the least puzzling character of the disease is the variability of its appearance. There have been groups of cases in our records; for example the following periods showed several while the intervening years lacked them entirely—1905, 1907–8, 1911–2, and 1920–1. The second and fourth outbreaks were definitely related to a new arrival but the records do not show that such was the case for the other two. Mr. Joseph who supplied us with many specimens, tells us that he has had an experience of fifty-four cases in 200 kangaroos and then failed to encounter the disease for years. Perhaps this irregularity of appearance has something to do with the character of food supplied to the animals.

Among seventy deaths of Macropodidæ we have had thirty-three cases of the varieties which I have included in this infection, made up of the following forms: cases limited to the jaws, pharynx and neck, six; cases of this sort with extension to lungs and stomach, ten; cases of this sort with general spread suggesting septicemia, five; gastrointestinal and hepatic, eight; nasal and sinus infection without necrosis in the jaw and with general spread including the lungs, four. The total incidence in Kangaroos is therefore 47 per cent., the necrotizing forms being 30 per cent., the gastrointestinal 11.4 per cent.

THE PATHOLOGY.

The essential features of the necrotizing variety of this disease are similar to those of actinomycosis—an inflammation giving rise to much fibrous tissue overgrowth enclosing pockets of softening, the whole process causing a deforming tumefaction. While primarily developing in the soft parts, this streptothricosis behaves like the ray fungus in that it spreads not only along clefts of tissue but directly through muscles and organs and even bones. Rarefying osteitis with irregular attempt at repair in the form of productive periostitis may be found in both infections. There is however a greater tendency to ulceration and general disease in the marsupial form, variations which seem referable to secondary invaders. When however the massive tumorous, necrotic and ulcerative characters of this streptothrix disease are insignificant or absent, the pathology is modified to the extent of obvious bacterial mixed infection, there then being catarrhal and fibrinous inflammations with degenerations of the viscera.

BACTERIOLOGY.

Since the pathology varies with the bacteriology as seen at this laboratory, it is well to pause at this time in a discussion of the former subject to introduce a brief statement of our findings in the latter, leaving however a full description thereof for later paragraphs. It is relatively easy to find in smears from necrotic masses threads of streptothrix, straight or curved with heavy blunt, but not bulbous, ends and never branched. Similar forms may be found in the necrotic tissue, both free in the softened area and near the margin of the healthy tissue, as irregular colonies growing in a tangled mass from the edge of which radiating threads may be seen. It has not been possible to find a “ray” growth with anything like the regularity so characteristic of the actinomyces nor do the ends present the bulb distinctive of that organism. Branching has been found once only, it being very uncommon in tissues although beautifully developed in cultures. While not especially sought, cocci and bacilli have not been seen, by Gram stains, within or immediately around the streptothrix colonies. Surrounding the mycelial groups is a necrotic zone about which is a loose connective tissue full of mononuclears and a few polynuclears. The centre of the colony is made up of tangled mycelial threads and necrotic debris. From uncontaminated necrotizing masses we have obtained cultures three times out of very many attempts.

[Illustration:

FIG. 63.—KANGAROO STREPTOTHRICOSIS. ANTEROPOSTERIOR SECTION OF HEAD, SHOWING MASSIVE TUMEFYING PROCESS IN LOWER JAW BONE. ]

When ulceration or suppurative softening has taken place mixed infection with lower bacterial forms naturally occurs and the whole picture changes. Pulmonary complications, with or without evident ulceration in the pharynx, also admit other bacteria. Streptothrical forms are often easy to detect in stained smears and in cultures but the very extensive bacterial flora soon overgrows them and attempts at isolation are fruitless. Under the best of conditions their colonial development is slow and tiny until they are well accustomed to saprophytic life. The complicating bacteria that have been identified are _Streptococcus pyogenes_, pneumococcus, pyocyaneus and colon bacilli to which may be added moulds of the Aspergillus group but these all have been variable in numbers and appearance; the most frequent and therefore probably most important secondary invader is an organism we have not been able to identify.

This germ, a tiny, Gram-negative, non-motile rod with a tendency to bipolar staining, will appear in smears from an ulcerated necrotic mass, from the nasopharyngeal exudate and from pulmonary lesions and may develop upon agar or blood media for the first generation but refuses to grow after that despite our best efforts. At present we hope to have it by growing material a long time in blood broth. Microscopical examination has not revealed it in the tumor-forming variety but on one occasion it was found in the lung; its Gram-negative characters make its detection in tissue very difficult. For obvious reasons the importance of this germ cannot be estimated but it seems from the frequency with which it is encountered that in some manner the streptothrix may be aided by this unidentified bacillus especially in the ulcerative and septicemic varieties of Kangaroo disease.

In so far as the diagnosis of this infection in the uncomplicated form, like lumpy jaw, is concerned the finding of streptothrix by stain seems adequate and its presence in the gastric ulcers and hepatic necroses identifies this variety. The most difficult question to decide is the identity of the cases without one or the other of these distinctive features but with mucocatarrhal or purulent nasosinusitis followed by pneumonia or septicemia, and of cases of primary pulmonary involvement. These instances have been diagnosed as belonging to the same category because of the presence of streptothrix in the exudate at the site of the important lesions and because the type of lesion is similar to that which complicates accepted characteristic cases. Inability to reproduce experimentally any of these infections limits our criteria for judgment in the matter. I am inclined to view these septicemic cases therefore as initiated by the streptothrix, growing in the nose and sinuses or inhaled into the lungs, aided by lower bacteria, an unidentified Gram- negative bacillus being the most important.

Having reviewed briefly the bacteriology of Kangaroo disease, its strict pathology may be discussed more definitely in terms of the type of infection. Reference has already been made to the method of pathogenesis employed by the streptothrix and its congeners. Whether or not a toxin is elaborated by these organisms is an unsettled question, especially for the marsupial variety because as yet it cannot be made to produce lesions in other animals. It is highly probable that all these organisms find colonization easy in the animal’s body once they get well settled, and that they act mechanically, producing necroses by their growth and by attracting leucocytes in such large numbers that digestion of devitalized tissue occurs, to an extent that resembles pus. The inflammatory tissue is not distinctive, except in so far that fibrosis enclosing pus pockets is peculiar to it. In softer tissue, like the liver, fibrosis is not so prominent, whereas diffuse and irregular spread is more pronounced. At the margin with the healthy tissue, reactive, that is resistant, inflammation is no more in evidence than within the tumor growth itself and as a matter of fact the tissues do not seem to put up a good fight against the spread of the inflammation.

[Illustration:

FIG. 64.—KANGAROO STREPTOTHRICOSIS. STOMACH, SHOWING TWO ULCERATIONS AND DEEP INFILTRATIONS OF THE WALLS. ]

Histologically, aside from the finding of the streptothrix colonies, there is nothing distinctive, the peculiar expressions of the disease being most manifest in their gross characters. For the purpose of describing the pathological features, the cases have been divided into the necrotizing form around the jaw, a similar process in the stomach and liver, necrotic cervical cases followed by lung involvement, the nasal variety upon which pneumonia succeeds and a septicemic form arising from any locality. Illustrative cases will be cited for each of these forms, a method of presenting the pathology thought to be superior to a general discussion.

The first illustrative case is one localized in the tongue and pharyngeal wall; it is quoted because of its strict localization.

Great Gray Kangaroo (_Macropus giganteus_). Sick four days, tongue swollen so he could not eat.

DIAGNOSIS.—Necrotizing process of floor of mouth and pharyngeal wall, dilatation of heart, passive congestion of liver, acute diffuse nephritis, inflammatory edema of lungs. General condition good. Jaws and teeth negative. Floor of mouth firm in places, boggy in others, but generally infiltrated. Anterior two-thirds of tongue purple and green as if gangrenous. Root of tongue and adjacent floor of mouth yellowish, wet as if from recent coagulation necrosis. In the muscle of the tongue a line of demarkation is shown at end of hemorrhagic zone behind which muscle is fairly good. Sides of pharynx, palate, tonsillar region show superficial pseudomembranous inflammation and yellowish gray, wet infiltration of muscles. Epiglottis purple and swollen to twice normal size. Laryngeal mucosa deeply injected, swollen and covered with tenacious gray mucus. Trachea and bronchi deeply injected and slimy. The lungs are uniformly deeply injected and along course of bronchi in lower lobe, lung tissue is distinctly more boggy than elsewhere. On section this area is slightly paler and more granular than the rest of the lung. Lung is everywhere slightly edematous. The bronchial lymphatics are swollen, pale pink and edematous. The heart is dilated acutely judging from the left ventricle wall which is nowhere over 1 cm. The liver is slightly enlarged, surface smooth, edges sharp, color deep purple, section surface very bloody. The spleen is soft, capsule smooth, pulp homogeneous purple, follicles not visible, trabeculæ normal. The kidney is slightly large, capsule smooth, strips easily leaving purple surface. The cut surface swells out, has irregular striæ, congested lines between, glomeruli visible and large. Smear from centre of tongue muscle shows staphylococci in some places in colonies, and long, slender rods.

The following case is one of gastric, intestinal and hepatic involvement, apparently primary, the last possibly arising by a hematogenic or lymphogenic route. Judging by the slides of the gastric wall the process started deeply and broke through the mucosa. This cannot be asserted definitely since kangaroos are susceptible to gastritis so that the streptothrix may have been implanted upon a preëxisting inflammation.

Black Wallaby (_Macropus ualabatus_). Congestion and edema of lungs, abscess of stomach and liver (streptothrix), ulcerative enteritis, necroses of spleen and lymph nodes, congestion of kidney. The animal is thin, hair loose. The mouth and nose seem to be absolutely healthy. The weight of the lungs is increased by congestion, they are solid, homogeneously red, with no air in any lobe except at edges. A piece cut from centre of lung sinks quickly in water. The trachea contains frothy blood. The heart muscle is soft, flabby and lustreless, chambers dilated, valves normal. The liver is of normal size, firm, smooth surface, sharp edges, red-brown color. The small sublobe of the liver which lies between the gall-bladder and the pyloric end of the stomach shows a large abscess 4 × 3 cm., apparently starting in the substance of the liver _via_ the bile ducts. This is certainly not extension from the stomach abscess as the liver lying against the stomach is nearly normal. The abscess is sharply circumscribed with a zone of congestion about it. Aside from congestion the rest of the liver is normal. The common bile duct is large and freely patulous. The capsule of the spleen is thick, consistency firm, pulp deep red, irregularly mottled by pale areas of necrosis. The kidney capsule is smooth, strips easily leaving a smooth, brown surface. The organ is firm. The section surface is glistening, the cortex wide and congested, the medulla normal. The adrenal medulla is deep purple with congested line between it and the pale cortex. Most of the gastric mucosa seems good. At about the middle of the lesser curvature is an ulcer about 4 cm. across. The shelving edges are covered with apparently normal mucosa. The centre contains bloody pus and nodular masses of the submucosa extending in finger-like projections through the pus. At one point on the greater curvature there is a small pocket of pus on the serous side which has not ulcerated through to the mucosa nor broken into the peritoneum. The large intestine is deep red and the follicles appear from the serosa as darker areas. On the mucus side the follicles have ulcerated, having a necrotic centre and shelving edges. The rest of the mucosa in the neighborhood is swollen and deep red. The colon mucosa is dry and the contents are hard, dry “baked” feces. The main pancreatic duct and the common bile duct form a thick, firm, cord-like mass running through the pancreas and enlarging the papilla of Vater into the duodenum. All abdominal lymph nodes are large, firm and on section mottled with red areas. Culture from the liver abscess failed to grow. Histological section of lung shows moderate congestion, collapse of alveoli or their filling by edema, epithelial and small round cells. There seems to be no fibrin. This could be an early stage of pneumonia. Bronchi are for the most part negative, little peribronchial round cell infiltration. No streptothrix in two areas of round cell infiltration or in bronchi. Liver section shows a part of the liver destroyed by hemorrhage, degeneration and necrosis. The abscess consists of necrotic matter surrounded by a zone of about equal numbers of mono- and polynuclears and around this a loose fibrocellular zone. Streptothrix abundant in the abscess. Lymph nodes show chronic inflammation and coagulation necrosis without abscess formation. No streptothrix in areas of necrosis. Kidney is very much congested with little or no damage to secreting parts. Spleen shows enormous congestion, moderate amount of pigmentation, connective tissue both trabeculæ and through pulp increased, no areas of necrosis. In the stomach the mucous membrane shows slight cellular activity and some degeneration—this amounts to a true catarrhal gastritis especially in view of the submucous cellular infiltration and the granulation tissue which has separated the muscularis and involved most of the connective tissue. The edge of the necrotic part begins abruptly, the mass of necrosis lying on an active fibrocellular submucous and muscular layer. Streptothrix can be seen at edge and in necrosis.

Pneumonia originating either by inhalation or _via_ the blood stream, is illustrated in two stages by the succeeding cases. The first history illustrates the pulmonary involvement as secondary to necrotic streptothricosis around the jaw and tongue while the second animal’s disease began in the nose and related sinuses. These two protocols provide material for a discussion of two phases of the subject.

The character of the early bronchopneumonia in the first is peribronchial, and there is distinct indication of a generalized process suggesting a hematogenic origin, whereas there is but one area of bronchopneumonia in the second—a necrotizing lesion beginning in the bronchus. Streptothrices are rare in the first case but reasonably easy to find in the second. This latter is one of the cases which seem to support the idea that nasosinusitis may have a streptothrix as its basis in the absence of the usual picture of necrotizing “lumpy jaw.” These cases also indicate that pneumonia may originate either by inhalation or by the blood stream, and that perhaps the hepatic lesion may have the latter origin. There have been two instances of necrotizing periarthritis, in one of which the threads could be found. This also suggests that spread through the blood stream can occur, possibly in this respect to places where previous injury prepares for the reception of the organisms.

[Illustration:

FIG. 65.—KANGAROO STREPTOTHRICOSIS. ULCERATION IN GASTRIC WALL AND MASSIVE NECROSES IN LIVER. ]

[Illustration:

FIG. 66.—KANGAROO STREPTOTHRICOSIS. SECTION OF LUNG SHOWING EARLY ABSCESSES AND NECROSES, ONE WITHIN A BRONCHUS, ONE IN SEMICONSOLIDATED PULMONARY TISSUE. NOCARDIAL STRANDS COULD BE FOUND IN BOTH AREAS. ]

Thigh striped Wallaby (_Macropus thetidis_). Streptothricosis of soft tissues of jaw. Early bronchopneumonia. Acute fermentative gastritis. Acute general infiltrative enteritis. Cloudy swelling of myocardium. The general condition of coat and of nutrition is good. The jaws are wide and the maxillocervical region full, both due to an indurative inflammation of the gums, tongue, floor of mouth and upper cervical tissues. At either side of the tongue and running around body of maxilla both sides, the inflammatory tissue becomes softer and there is an area about one inch long where it is soft, gray and contains yellow gray bodies in a grumous matrix. The teeth seem sound as do the external buccal tissues. The nasopharynx is free from induration. The bone on the left side shows a periosteitis with involvement of the superficial layers of bone, while on the right side the periosteum is swollen and opaque but the bone is free. The thyroid is imbedded in the edematous infiltration of the lower cervical tissues. The pleuræ are free of fluid and adhesions. Lungs are collapsed, uniformly pink somewhat emphysematous at places but give the impression of being lumpy. On palpation numerous nodular areas are detected. These prove to be peribronchial areas of gray-red solidity which swell out on section. The bronchus contains a gray and bloody thick mucoid matter. There is distention of the mesenteric vessels especially near the enteric insertion. The liver surface is smooth, edges very sharp, consistency firm, tough, resilient, color deep red, the section surface is glistening, moist, opaque, architecture probably normal. The gall-bladder is distended with viscid green bile; the common duct is patulous. The spleen has a rough, thin capsule, consistency tough and resilient, the section surface is mottled red with purple points; on section two small, pale objects seem to be squeezed out. The kidney capsule is smooth, strips easily leaving a smooth, deep red surface, the consistency is soft, the cortex is deep red, then a purple line between it and the red medulla, striæ invisible. The stomach contains frothy grayish mush. The mucosa is finely mammillated, deep pink until the last third when it becomes deep red, deeply injected and somewhat thickened. The pylorus is closed. Externally the gut is congested, in places translucent but for most part seems thickened by reddish swelling of both external layers and mucosa. The mucosa is granular or pebbly with here and there a small bloody suffusion. The histological section of lung shows alveoli open, septa relatively thin but somewhat congested, bronchi mostly open and connective tissue not increased. Some few bronchi, especially the larger, show a slight catarrhal bronchitis but mostly an infiltrative peribronchitis. The nearby veins and arteries show the most striking change, there being in nearly all of them a distinct thrombosis without circumferential pneumonia. In one place a distinct peribronchial pneumonitis was found. The kidney shows very marked congestion of all parts, causing compression, cloudiness and granularity of the epithelium. Glomeruli and connective tissue about normal. The intestinal serosa is negative save for congestion. Submucosa is densely infiltrated with mononuclears, some in definite groups. Section does not show areas mentioned in notes but these could be accumulations of cells with congestion. No streptothrix forms. Section from the infectious focus of face consists of active granulation tissue, densely injected and filled with mononuclears of two types, one the lymphoid cell, the other of the young connective tissue type. Areas of grouping like abscesses are seen and some necroses. Streptothrix in small numbers in the cellular collections.

Nail tailed Wallaby (_Macropus unguifer_). Kangaroo disease of nasal region. Necrotizing bronchopneumonia (Aspergillus fumigatus and Micrococcus albus). Acute diffuse splenitis. Congestion of liver and kidney. The general condition of coat and nutrition is good. The face is wide just below the eyes. About the “hare lip” and the nose the soft tissues are soft, gray, necrotic. All the internal nasal tissues seem swollen, gray-red. There is subcutaneous edema, bloody in places, around the right face, eye and jugular angle. Tissues of nasopharynx swollen, deeply injected and covered by a thick mucus. Pharyngeal and buccal cavities negative. Tonsillar areas pink and flat. Larynx and trachea slightly swollen but pale on mucosa. Salivary glands and cervical glands normal in size and pale pink. Pleuræ pale and empty. Lungs swollen out uniformly, quite cottony except at lower right base where there is a nodule about 3 × 5 cm. firm and doughy. On section it is found to be a peribronchial consolidation of pale reddish gray color and indefinite outline. The bronchus itself is deeply congested and contains a grumous mass. The peribronchial lymph nodes are small, soft, pink, homogeneous. The heart is negative. The liver is large, surface smooth, edges sharp, color deep purple, consistency soft. Section surface is glistening, smooth, moist, very dark purple with obscure markings. The gall-bladder contains fluid brown bile; common duct is patulous. The spleen is soft, tough, capsule pebbly, section surface is mottled, light and deep pink, follicles and trabeculæ not distinguished. The kidney capsule is smooth, strips easily leaving a smooth purple surface, section surface is glistening, deeply congested, striæ obscure but seem normal, glomeruli not visible, organ is soft. The gums and teeth are not involved in the mycosis. The stomach contains mushy digesting food. The mucosa is mottled pink, soft, digesting, at lower half submucosa is deep pink, a few small ecchymoses. From pylorus to ileum, serosa is deeply injected, edematous, mucosa swollen and edematous, deep pink, loosened in places, but translucent. Below this the mucous membrane becomes smooth, flat, pink-yellow. Lower ileum and colon contain rather firm fecal balls. Follicles nowhere prominent. The pancreas is small, soft, yellow pink. The follicles of the mesentery are small, pink gray and homogeneous. Smears from the bronchopneumonia show a threadlike Gram- positive form and a few Gram-negative rods. Cultures from lung show _Aspergillus fumigatus_ and _Micrococcus albus_. Nose too foul for culture. Histological section of lung shows the alveoli mostly open but the septa widened by congestion. Blood vessels are open and contain recent clots; one vessel near lesion below is thrombotic. The two large bronchi in section show catarrhal bronchitis and infiltrative peribronchitis of which the latter is more severe and advanced. Beside the larger is a necrotizing pneumonitis from which nearly all the architecture has disappeared. The exudate is chiefly mononuclear around the edges; centre no cellular identity. Another mononuclear process not connected with bronchus in section is found with an early necrosis. Streptothrix strands may be found in the bronchial exudate and near the margin of the necrotic patch. They do not grow in colonies however. The spleen shows general congestion without pigmentation. Follicles large, solidly lymphoid. Connective tissue about normal. The kidneys show marked congestion everywhere. Capsule and intrarenal fibrous tissues about normal. Very severe congestion which seems to have caused compression and granularity of the epithelium.

The last case, judging by stained smears, is one of pure nasosinusitis from streptococci and streptothrices. Cultures were not tried because of the enormous bacterial flora.

[Illustration:

FIG. 67.—KANGAROO STREPTOTHRICOSIS. LOW POWER PHOTOMICROGRAPH OF A NOCARDIA COLONY WITH NECROSIS WITHIN AND AROUND IT. THIS WAS FOUND IN A SECTION FROM THE LIP OF THE SPECIMEN SHOWN IN FIG. 62. THE BLACK BORDER CONSISTS OF PARALLEL THREADS SO CLOSELY PLACED THAT THEIR SEPARATION UNDER THE CAMERA IS PRACTICALLY IMPOSSIBLE. THIS TYPE OF COLONY RESEMBLES THE “RAY” COLONY OF ACTINOMYCES. ]

Robust Kangaroo (_Macropus robustus_). Acute purulent ethmoiditis. General acute purulent anterior cranial sinusitis. Acute necrotizing glossitis and pharyngitis. Cloudy swelling of kidney. The face seems a little full and the subcutaneous tissues slightly edematous. The nasopharynx contains a thick tenacious mucopus. Ethmoid and frontal sinuses and turbinate spaces contain a thick purulent matter, the mucosa being densely injected, swollen and velvety. Pharyngeal wall and right half of posterior half of tongue are involved in a dull brown and necrotizing process, quite sharply outlined by zone of congestion. This process is comparable to the necrotizing gingivitis seen in front of jaw in kangaroos. Larynx, trachea and lungs seem uninvolved save for slight generalized congestion. Cervical lymph nodes especially those about the larynx are definitely enlarged, soft, moist and brown. Mediastinal nodes slightly enlarged, soft and pink. The heart is negative. Liver normal. Spleen is soft, homogeneous dull red. The capsule of the kidney is smooth, strips easily leaving a purple surface. The glistening section surface swells slightly, vasa recta are congested, striæ wide and pale, glomeruli not visible; consistency is resilient. The mouth and teeth are not involved in the process mentioned above. There is a small quantity of properly digesting food in the stomach. Stomach and intestines negative. Brain not involved. No extension from anterior cranial sinusitis. Smears from the mucopus confirm the gross appearance and contain short chains of streptococci and large diplococci. Smear from cut surface of tongue shows innumerable small bacilli and diplococci but especially mycelia with rather heavy clubbed ends but without true branching. One group was found arranged like ray fungus. It is noteworthy that there is no aspiration pneumonia and very slight evidences of septicemia.

BIOLOGY OF N. MACROPODIDARUM.

The original discovery of the streptothrical forms was made in stained smears from necrotizing lesions. They were considered as secondary invaders until repeated observations of a similar character aroused the suspicion that they stood in some important relationship to the lesion. Early attempts at their cultivation were made under anaerobic precautions, a method now known to be almost certainly doomed to failure because a strain long under cultivation requires two to three weeks to make an appreciable growth in the absence of air. Finally in 1911 a successful cultivation occurred by searing the surface of an unopened mass in a freshly dead animal and planting bits of the interior upon aerobic blood serum plates. Colonies grew after three or four days and from them the first strain was started. It grew for several generations, long enough for the preparation of a vaccine, which will be described later, when by mischance it was lost. In 1920 another successful cultivation occurred, this time by incising a mass in the soft sublingual tissue and plating in the same manner; upon this culture the biology is described. Smear preparations offer no more than has already been mentioned.

Colonies develop upon blood serum plates as opaque, pale yellow, circular, discrete masses with a slightly depressed uneven centre, but without umbilication. They remain smooth and slightly glistening for several days, then become slightly wrinkled and twisted with a more definitely raised edge and a tendency to an uneven sinking in of the centre. Transfers to agar slants show wrinkled continuous opaque, dull yellow, sharply outlined growths which soon wrinkle, fold, and twist like certain tubercle bacillus cultures. Spreading occurs, but is slow after forty-eight hours. As medium becomes drier it is possible to see a thin, colorless, wrinkled film stretching out from the main growth. If the medium be dry or old or if only a small portion of seed material be used and this scattered over the surface of the slant, discrete colonies arise. These are circular, seldom exceeding 3 mm., dirty yellow-white, distinctly umbilicated and without clear film of spreading around them.

In nearly all quite old cultures, a white chalky efflorescence appears over the surface.

The morphology of the young agar culture is chiefly mycelial or filamentous, whereas from a culture on dried media and those showing efflorescence, the organisms are short, heavy, deeply granular and of the mycobacterial type.

Glycerine agar.—Corresponds to agar.

Blood agar.—Similar to agar but much less luxuriant.

Blood serum.—Limited dirty yellow, raised, dull, wrinkled and granular, tightly adherent to the medium.

Potato.—Spreading, dirty yellow, much wrinkled, friable, tightly adherent.

Gelatine.—Limited growth as a wrinkled, tough scum only on surface.

[Illustration:

FIG. 68.—KANGAROO STREPTOTHRICOSIS. HIGHER MAGNIFICATION OF EDGE OF STREPTOTHRIX COLONY, FIG. 67. IT SHOWS THE DEEPLY STAINING MYCELIA SEPARATING MUSCLE FIBRES WHICH ARE DEGENERATING. ]

Litmus milk.—No change for six days, then beginning slight alkalinity which increases very little, shows digestion of the caseinogen, slight, thin filmy growth on surface.

On media such as litmus lactose agar and old Endo it grows slowly on surface and assumes the color of the medium.

Broth.—Only surface growth appearing during early generations, after 3–6 days as a wrinkled, pale yellow scum very much like the tubercle bacillus growth; later generations grow perceptibly in one to three days. Medium perfectly clear. If a large mass be seeded into neutral broth there is a perceptible increase in the growth after ten days. The medium thereafter tends to a faint turbidity. Titration of broth growth after twelve days shows alkalinity requiring 0.3 cc. decinormal acid, while the control tube incubated same length of time showed an acidity requiring 0.57 cc. of decinormal NaOH.

On the following sugars there is a slight surface growth without change in the color, Andrade indicator—dextrose, lactose, saccharose, maltose, mannite, dextrin, galactose, salicin.

Cultures observed on two per cent. neutral agar.

A.—Stained by Loeffler’s stain.

Twenty-four hours.—Shows threads growing out from a central amorphous mass, but the whole does not retain the regularity or parallelism of actinomyces. Threads are poorly stained and rather disconnected but not jointed. Small number of metachromatic bodies apparently in older individuals, certainly in the better formed ones. No intercalary spores, unless the metachromatic bodies be so considered. Individual threads measure from one-third to one micron in width. Metachromatic bodies measure on the average one micron.

The threads in the forty-eight hour preparation seem distinctly wider, up to one micron and possibly become heavier toward the end, but do not have a distinct bulbous extremity.

In three days the threads are much longer, show distinct branching and a tendency to transverse segmentation. More than one metachromatic body may be present in one segment.

Four days.—Still coarser, short segments have appeared separately. Metachromatic body is coarser and blacker; some of the masses have gone to pieces and show only a diffusely staining smudge of metachromatic bodies. The short segments show a tendency to grow out into threads.

Fifth day.—Condition is much the same plus many young, delicate, poorly staining threads.

Sixth day.—The same but all seem to be somewhat wider and diffusely staining.

Seventh day.—More diffuse staining and decidedly fewer metachromatic bodies.

A.—Stained by Gram’s stain.

Twenty-four hours.—All forms are light purplish. The threads stain much more clearly than by Loeffler’s and show distinct transverse segmentation of rather uniformly long bacilliform shape. Metachromatic bodies not so distinct but seem larger where found. Coarser threads have swellings in some of the areas which are not segmented and this type seems to have more branching and metachromatic bodies; in other words it would seem that this is a form that reproduces by budding or intercalary spore formation.

Forty-eight hours.—Much the same, more long threads with transverse division, somewhat more delicate, generally fewer coarse threads with swellings and spores. Still pale purple and not distinctly Gram- positive.

Three days.—Condition much the same.

Four days.—Two forms present—definitely Gram-negative delicate slender threads, nearly Gram-positive, and heavier, curved and twisted long bacillary forms, some streptococcoid threads and a few bulbous short threads. Very few metachromatic bodies.

Five days.—Condition much the same except that the delicate threads are inconspicuous and the darker purple bacilli have increased. Metachromatic bodies increased as have swellings in coarser threads.

Six days.—Much the same but for the appearance of young, delicate definitely Gram-negative threads. There are fewer metachromatic bodies and internal spores.

Seven days.—The same.

B.—Grown on Loeffler’s blood serum.—Loeffler’s stain.

Twenty-four hours.—Delicate, poorly stained short threads, few tiny metachromatic bodies.

Two days.—Not well stained, relatively short threads show numerous metachromatic bodies varying from exceedingly tiny dots to coarse granules wider than the thread. These may be numerous in the same segment and form a row from six to ten. Many short bacillary forms.

Three days.—Poorly stained, metachromatic bodies apparently more numerous but much smaller.

Four days.—Almost entirely short, heavy bacillary forms, some of which are very like diphtheria bacillus in the irregularity of width; many metachromatic bodies, distinct branching, some of the small heavy ones have fusiform swellings; practically no long, heavy threads.

Five days.—Essentially the same, individual elements slightly larger, fewer but coarser metachromatic bodies, more numerous round forms suggesting large pale cocci.

Six days.—Much the same but elements shorter, smaller and some more segmented.

Seven days.—More long forms of uniform staining but still a majority of coccoid or short bacillary forms with irregular staining and metachromatic bodies; no long threads.

B.—Gram’s stain.

Twenty-four hours.—Pale purple, almost Gram-negative, long, slender but well outlined threads, a few coccoid forms, practically no granules.

[Illustration:

FIG. 69.—KANGAROO STREPTOTHRICOSIS. PHOTOMICROGRAPH SHOWING THE SEPARATE THREADS OF NOCARDIA IN A SOFT NECROTIC LESION. ]

Two days.—Very pale, almost Gram-negative threads, very many coccoid forms and short rods, considerable segmentation of the longer threads.

Three days.—Increase in short, heavy bacillary forms with bulbous ends, deeply stained ones and the granules being lightly Gram- positive; long, slender threads are disappearing.

Four days.—Almost exclusively short, heavy forms with bulbous ends with coccoid forms, heavier forms almost definitely Gram-positive, granules Gram-positive.

Five days.—Much the same but more segmentation in the bacillary forms, coccoid forms become more numerous.

Six days.—Individuals are somewhat longer but there are many rods with fusiform swellings containing granules; coccoid forms present in chains sometimes.

Seven days.—More long rods or short threads, pure coccoid and bacillary forms.

The morphology upon bouillon depends somewhat on age and upon the location. Upon the surface the long branching mycelial type appears early and persists until the whole surface is covered whereupon the segments divide into coccoid elements with metachromatic bodies. If heaping-up develop the coarse grains on the mass consist of granular or coccoid rods. When growing in the depth the coccoid form is the predominant one, only a few delicate mycelia, usually Gram-negative, being found.

The Gram character of the organism should be emphasized. The young, delicate mycelia are negative or take a very feeble blue stain. The heavy bacillary forms are Gram-positive. Like the ray fungus the heavy ends are sharply Gram-positive, but unlike it, there has never been seen a Gram-negative bulbous capsule around this end.

The determination of this organism was undertaken from the classifications of Petruschky (Kolle-Wassermann), of Castellani in Castellani and Chalmers’ _Tropical Medicine_, and of the Society of American Bacteriologists. In the first classification it corresponds in some ways with _Streptothrix hominis_, and in some ways with _Streptothrix capræ_. As for the second authority it falls into the Nocardiaceæ, section parasitica, subsection I, in that a distinct earthy odor is absent and that there is no liquefaction of coagulated protein. It resembles several of the species given in this subsection, but does not correspond exactly with any of them. Consultation of the classification of the American Bacteriologists would place it among Mycobacteriaceæ. The facts that it is strongly aerobic, produces whitish efflorescence which may possibly be aerial hyphæ and breaks up into short segments, place it in the genus Nocardia. It seems, however, to belong to a division of Nocardia which is close to the Mycobacterium since the short elements are swollen, cuneate and usually heavy, which is unusual in the more typical Nocardia. It is not, however, acid fast and therefore cannot be classified among the Mycobacteria. This culture seems to be a variety not heretofore described, and since its association with the disease is so definite, whether or not it be the cause, the name NOCARDIA MACROPODIDARUM is proposed, because the kangaroos belong to the order Marsupialia, family Macropodidæ.

The discovery of these organisms within tissues is by no means easy even though the larger colonies may be located by staining. If Loeffler’s method be used the central mass stains quite diffusely and the spreading mycelia around the edge stain faintly. For study purposes this stain is preferable to Gram-Weigert, since despite the positivity of the cultures, the blue dye can be removed very easily from sections and only with great care will enough remain to permit tracing of the separate threads; with Gram stain no detail can be made out in the centre of the colony, it being a diffuse blue. Careful search near the edge of these necroses will usually succeed in the discovery of a few mycelia stretching in between the mono- and polynuclears of the low grade inflammation. This is best seen in the margin of gastric ulcers, but may also be found in the cervical masses. When searching in the pulmonary tissues the organisms are to be found in the bronchial exudate or at the edge of pneumonias. In one nasal mucosa the mycelia were dispersed, not growing in colonies as in localized inflammations.

EXPERIMENTS AT THE REPRODUCTION OF THE DISEASE.

When the first culture was isolated it was injected into guinea-pigs; its loss stopped further work because it could not be regained from the animals. The present culture had been injected into guinea-pigs, rabbits, opossums—all with negative results; such an experience is not unknown for actinomyces. Intraperitoneal, intravenous methods having failed, inoculation was made into the gums of rabbits and of opossums with no result, even after traumatizing the mucous membrane. The injection of about 5. mg. of a twenty-four-hour agar culture was made directly into the masseter muscle of an opossum without producing even a lump at the site. Atomizing a culture into the nose and throat of an opossum seemed also without effect. Injection of cultures into the nose, gums and labial tissues of a wallaby have been negative; nor has any perceptible effect followed the atomizing of a heavy nocardial suspension in broth into the trachea of this animal.

The results of these experiments are in accord with those of many similar attempts to reproduce actinomycosis. Perhaps in Kangaroo disease the small Gram-negative bacillus is a necessary factor.

SPECIFIC PREVENTION AND TREATMENT.

Encouragement that we were upon the right track was, however, found in another direction. Improvement in human and bovine actinomycosis having followed the use of vaccines, it occurred to me to try this method as treatment and prophylaxis. The first culture to be isolated was just at hand, so that it could be used at once. Five injections were given under the skin of the thigh to a recently developed case of the ulcerative gingival variety, a noticeable improvement occurring almost at once, and at death there was an apparent cure of the local lesion. However, the accompanying protocol made at the time tells the whole story, no adequate explanation being at hand.

Red Kangaroo (_Macropus rufus_). Disease of the mouth first noticed March 31, 1912, died September 13, 1912. Necrotizing osteitis, arthritis and periarthritis of left ankle, subacute fibrinous right pleuritis, hemorrhagic bronchitis with atelectasis in right middle lobe, abscess of right middle lobe; passive congestion of lungs, liver, kidney, chronic splenitis, chronic general lymphadenitis. The animal is in general good condition except for a fusiform swelling about the left heel with evidence of fracture. The necrotic process in the hare lip, nose and palate has entirely disappeared. One front incisor has gone and the other is loose. There is a scar on the under part of the soft palate in a small healed channel between palate and floor of nose. There is no evidence of pyorrhœa. Cervical and axillary lymph nodes are much enlarged, pale yellow, firm and of the appearance like early stages of Hodgkin’s disease. Fascias of cavities congested. The lungs are mottled purple, air content decreased, section surface purple, exuding frothy blood. The whole right lung is covered with a thick fibrinous exudate, most intense over middle lobe at site of atelectasis. There are light scattered adhesions. The anterior margin of the lung is adherent to the pericardium which is covered in the front by exudate. Upper and lower lobes show hypostatic congestion. Middle lobe has separate bronchus filled with necrotizing blood clot extending into a smaller bronchus with complete occlusion. The alveoli supplied by the last show atelectasis like hemorrhagic infarct. There is a small subpleural abscess near the margin of this atelectatic area. The bronchial lymph nodes are slightly enlarged, mottled yellow and pink, firm with large, diffuse follicles. The pericardium contains 2–3 cc. clear fluid. The heart muscle is pale, purple and soft. All the vessels are full of currant jelly clot. On the posterior surface of the aorta internally about an inch above the valves there is a patch of roughening with a suggestion of thickening and opacity. It is comparable to the early stages of syphilitic aortitis. The liver is normal in size, surface smooth, edges sharp, consistency firm and friable, color purple. The section surface is glistening, smooth, moist, and shows passive congestion. The gall-bladder contains fluid brown bile and the common duct is patulous. The spleen is slightly enlarged, firm and tough, capsule wrinkled. Section surface is mottled red and purple with irregular gray trabeculæ and faint scattered follicles with diffuse margins. The kidney capsule is smooth, strips easily leaving a smooth brown surface. Organ is firm. The section surface is glistening with a line of passive congestion with distended vessels between the cortex and medulla which are of normal widths. Intestines seem normal throughout. The pancreas is firm, pale pink, slightly edematous. The mesenteric lymph glands are moderately enlarged, yellow, firm, homogeneous with congested centres. About the left ankle joint there is a necrotizing infection which has involved the bone causing a pathological fracture of the lower end of the tibia. Smears from the periarthritis, pleuritis and blood clot in the bronchus show streptothrix, a short colon-like rod and a coccus in fours—a picture precisely like that obtained from the jaw bone cases. In addition to the above there is a very distinct encapsulated pneumococcus form in smears from the blood clot in the bronchus. This is the animal which was vaccinated with a culture made from the depths of a necrotic mass, upon which treatment she rapidly improved and as seen from the above notes recovered from the palate condition. Why she should have a second infection apparently with the same organism is difficult to determine. Possibly the second batch of vaccine was not sterile, it not having been controlled because the first batch of vaccine was sterile after one hour at 60° C. Possibly the animal was sensitized and a few bacteria settled in the leg. It was along this leg that the inoculations were made.

We permit ourselves the facetious observation that that vaccine prevented the labial and cervical variety for five years, because during that period it stood in the icebox, and there was no case of that particular form to which to give it, although a few of the nasal and gastric varieties occurred. It was recontrolled and did not show living organisms. That it should have cured the disease in the jaw and apparently later permitted a lighting up of a septicemic and pulmonary form with necroses in the leg is difficult to explain.

Just recently we have used a vaccine from the current culture upon another case beginning in the gums and jaw bones. This case was detected early and was treated with ascending doses beginning at 0.3 mg. and running up to 10. mg. At first there was some improvement, but the animal finally died from pulmonary complications. The course of the disease, however, instead of being three weeks, as is the customary duration, lasted two months, an extension of the course which has made us hopeful. These two experiments, indefinite though they be, have offered encouragement and seem to supply a little additional support to the idea that the organisms stand in etiological relationship to the disease.

The employment of the vaccine has been extended to its use as a prophylactic in animals exposed to the disease or specimens that have slight reddenings or erosions on the buccal mucosæ suggesting possible early stages of streptothricosis. Five animals have now had a course of vaccine injections, ranging in number from 5 to 10 and in quantity from 0.3 to 2.4 mg. over a period of a month. Fourteen months have elapsed at the time of writing and only one case has developed, but this of course cannot settle the efficacy of the method; perhaps it would be safer to demand that no case should ever appear in a treated animal, while the disease did appear in the untreated.

The preparation of the vaccine is by no means a simple matter, since the surface growth upon solid media is so tenacious. Methods such as are employed for the tubercle bacillus have to be used. The first two vaccines were made by scraping off surface colonies from agar and grinding with glass balls. One successful batch was made recently by simply triturating the colony directly on the agar slant, but the latest method seems to offer the simplest and most generally satisfactory way. Neutral broth is placed in flasks containing glass beads and sterilized in the incubator. This is seeded with the Nocardia, incubated at 37° C. until the surface is covered, heated to 60° C. in a steam sterilizer and tested for sterility. If growth occur it is reheated until dead, whereupon the broth is syphoned off, the growth emulsified by whirling the flask, thus grinding the bacterial mass by the glass beads. Sufficient saline is added to make a workable emulsion, and the fluid then poured into bottles. Control by reculturing is again done, and if the fluid be found sterile, 0.5 per cent. trikresol is added to keep it so. These organisms cannot be counted accurately because of the variation in length, their budding and coccoid forms. Standardization is done by weight. A definite equal quantity of the suspension and of the saline used to make it are evaporated to dryness in weighed vessels and the whole then weighed. The difference is the weight of the organisms suspended in the saline. Such a fluid can be diluted so that a given bulk will contain a convenient weight of germs. The one now in use contains 8. mg. per cubic centimetre. Dosage as indicated above usually begins at 0.5 mg., a quantity which does not produce any local inflammation at the site of injection. It is perhaps well to adopt a quantity of 0.1 mg. per kilo as the initial quantity.

The Garden has encountered no case of the remaining important chronic infections, glanders, lymphangitis, and infectious abortion.

SECTION XVII—