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official statistics therefore give the percentage of reactors in herds known to be infected, and not of the whole of them. But there can be no doubt that premises which are suspected by the sanitary authorities contain a higher average of tuberculous animals, and that there are herds which are completely free from the disease. The importance of these last, from the point of view of the statistics, remains to be proved, but if they were subjected to the test the effect would be to lower the percentage of positive reactions.

The official figures refer only to premises which are as a rule the most seriously affected. There is not sufficient information to permit of any exact estimate of the incidence of the disease, but when the statistics of 20th December 1921 show that the infected premises contained 144,065 animals, these were" contaminated" animals (of which a certain proportion did not react to tuberculin), and not "tuberculous" animals, as stated in Calmette's report.

Although there is an impression that tuberculosis is on the increase, veterinary practitioners will scarcely admit that a third or a half of the stock are affected. If it were so cancellations of sales would be never-ending, and the cattle trade would be so upset that it would be necessary to make an immediate alteration in the law whereby a single reaction to tuberculin is sufficient to annul a sale.

The author states that the note has been written, not with a view to denying that bovine tuberculosis is increasing, but to put those who may be interested in the subject on their guard against the exaggerated estimate likely to be formed on the strength of statistics that are misleading when not fully explained. (Sausseau, Rev. Vet., June 1922, Vol. LXXIV., No.6, PP·362-3 6s·)

CHARACTERISTICS OF THE BACILLUS OF BLACKLEG.

IN this paper are described methods for the isolation of Bacillus chauvcei, and for its differentiation from other amerobes found in similar infections.

The organism with which Bacillus chauvcei is most frequently confounded is the vibrion septique. The differentiation of these two from most other ancerobes found in similar infections is facilitated by the fact that they do not form proteolytic ferments. The media used by the authors were Hibler medium, 2 per cent. dextrose agar, liver broth, and liver agar. The following are the details of the preparation of these :-

Hibler Medium.-Beef liver 500 grammes and brain 500 grammes are ground, and to the liver are added 1000 cc. of water. Both are steamed for an hour. The liver broth is strained through cheese cloth and cotton, and I per cent. peptone and '5 per cent. salt are added. The broth after titration to PH 8'2 is tubed, and brain is added in the proportion of one part to two of broth. The tubes are autoclaved for an hour at IS lbs.

Dextrose Agar.-Beef infusion (500 grammes per litre) has added to it I per cent. peptone, '5 per cent. salt, and 2 per cent. dextrose. The

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reaction is then corrected to PH 8'2. Agar is added in the proportion of 2 per cent., and the whole is steamed until the agar is melted. The medium is then cleared with egg albumen, filtered, tubed, and autoclaved at 15 lbs. for twenty minutes.

Liver Broth is prepared like that used in the Hibler medium, cleared with egg albumen, filtered, tubed, and autoclaved at 15 lbs. for twenty minutes.

Liver Agar is made in the same way as the liver broth, save that 2 per cent. agar is added and melted by steaming before clarification.

Ffteen strains of Bacillus chauvcei have been examined. Of these fourteen were isolated from cases of blackquarter, and the remaining one was from muscle received from the Bureau of Animal Industry.

The morphology of the strains was as follows :-The organisms occur singly or in pairs. Chain-formation is exceptional.

The length is from 3 to 4 microns, and the breadth '5. Tbey frequently contain granules, and become pleomorphic after incubation for twenty-four to forty-eight hours. Clostridial and navicular forms are the most common. In young cultures the organisms are sluggishly motile, but in local lesions of infected guinea-pigs they exhibit active movements.

The spores which develop in eighteen to twenty-four hours are centrally or terminally placed, and are wider than the parent cell.

In lesions in guinea-pigs Bacillus chauvcei is distinguishable from the Vibrion septique by the fact that the latter is frequently discoverable on the surface of the liver in long chains.

Old cultures are Gram-negative, but young cultures are positive to Stirling's modification of that method.

Dextrose agar is useful in distinguishing the two organisms, since Bacillus chauvcei will not grow in it unless blood or tissue is transferred to it when the tubes are inoculated. Vibrion septique and other anrerobes grow well in this medium.

The addition of 2 cc. of an extract of guinea-pig liver in salt solution (strength not stated) to 25 cc. of glucose agar enables Bacillus chauvcei to grow in the medium. Without the liver extract no growth takes place.

In deep tubes of beef liver agar a good growth of sman spherical or elliptical colonies is obtained in eighteen to twenty-four hours. Vibrion septique forms fluffy colonies in this medium. When the agar content is lowered to I per cent. Bacillus chauvcei grows in the medium, and the colonies present appearances similar to those of Vibrion septique in the 2 per cent. medium.

Bacillus chauvcei produces little or no gas in deep tubes of beef liver agar, whereas Vibrion septique rapidly produces large quantities of gas. Poured plates of the same medium give good growths of Vibrion septique in twenty­four hours, but Bacillus chauvcei fails to yield visible growth after three days' incubation.

Liver broth is fermented by Bacillus cbauvcei with the production of both acid and gas, and the organism will grow in liver broth to which cubes of cooked liver have been added without any other step to obtain anrerobic conditions.

In liquid media the organism causes a turbidity which soon clears up. In Hibler medium gas is produced, the medium clears up rapidly, and the cultures have no odour save a slight one of butyric acid. The odour is most easily detected if a film of an old culture is spread on a slide and slightly heated.

Vibrion septique will grow in recently boiled milk if a large amount of seed material is used, but Bacillus chauvcei will not.

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In gelatin prepared with liver broth as a base Bacillus chauvcei forms fluffy colonies, the medium is liquefied, and gas is produced by some strains.

Five strains of Bacillus chauvcei were tested for fermentation of a large number of carbohydrates, but no growth occurred in any of the tubes.

For the isolation of the organism from suspected blackleg muscle the following procedure is advised: A piece of the tissue is macerated with salt solution, and a few cubic centimetres of the suspension are drawn into a pipette and heated at 60° C. for forty-five minutes. The suspension is then used for the inoculation of liver agar, glucose agar, and Hibler medium. A guinea-pig is also inoculated subcutaneously.

Colonies having the characteristic appearances are fished out of the liver agar and again sown out on the media mentioned. These alternations should be repeated until purity is assured by obtaining organisms with the morphology of Bacillus chauvcei which will not grow in 2 per cent. glucose agar, and which give positive animal inoculation tests, etc. If guinea-pigs die in less than sixteen hours the cause of death is probably not Bacillus chauvcei. For separating Bacillus chauvcei from Vibrion septique the guinea-pig may be passively immunised against the latter with serum.

The guinea-pig is the animal of choice for experimental work with Bacillus chauvcei, but the results obtained from inocuh,tion, even with successive cultures of the same strain, are not always parallel.

Vibrion septique infections of the guinea-pig are more rapidly fatal than those caused by Bacillus chauvcei. Marked swelling round the seat of inoculation for some hours prior to death is characteristic of such infections. In rapidly fatal cases of blackquarter in guinea-pigs the organism may not be present in the heart blood at the time of death.

The pigeon can be infected with blackquarter; the muscular tissue presents a similar appearance and has an odour similar to the lesions found in cattle.

The authors have never isolated true Bacillus chauvcei from the muscle of sheep alleged to have died from blackleg in the field.

Experiments showed that sheep could be infected with virulent cultures if large doses (5 cc.) were used.

Blackleg occurs less frequently in sheep than in cattle owing to the higher powers of resistance possessed by the sheep. Rabbits and white rats are resistant.

Diverse opinions have been expressed regarding the protection conferred by Vibrion septique against Bacillus chauvcei. The authors of the present paper are in agreement with those who hold that cross immunity tests yield results which indicate a clear-cut distinction between the two infections.

In their detailed experiments they show that a dose of . I cc. of anti­blackleg serum protects a guinea-pig against twenty-five lethal doses of culture, whereas it does not afford protection against a single lethal dose of Vibrion septique.

Various views have been put forward regarding the value of cross agglutination tests for the purpose of distinguishing between Bacillus chauvcei and Vibrion septique. The authors' opinion is that this test, as far as the particular purpose is concerned, is still in the experimental stage.

One of the questions awaiting experimental investigation is the relation­ship between the effects of normal horse sera and specific horse sera with these organisms.

It has been found that the serum of some normal horses cause agglutina­tion of both the bacteria in dilutions up to 1 in 200. (Goss, Barbarin, and Haines,Jl. Infect. Dis., December 1921, Vol XXIX., No.6, pp. 615-629.)