In autumn and early winter certain ditches around Cambridge became offensive, and their water turned milky. This led to a grave suspicion of sewage contamination, which, however, was satisfactorily disproved. On the other hand certain bacilli with single flagella were isolated from the water of the ditches and found capable of giving off from their artificial cultures an odour comparable to that of the ditches. And to these bacilli, acting on the dead leaves which found their way into the water, and, probably, in conjunction with other anaerobic bacteria which were generating hydrogen disulphide, the sewage-like odour is attributed.

Certain patchy changes of colour in the leaves and wood immersed in the water are attributed to a chromogenic flagellate which was found growing on them.

1. The bacterial content of the air of the Underground Railways, when the average of all results of the bacteriological investigations is taken, does not numerically compare unfavourably with the outside air of London.

2. The ratio of the number of organisms growing at room temperature appears to be about 14 for railway air to 10 outside air. For those growing at body temperature the ratio is considerably higher, namely 2 to 1 respectively. The mean per litre, for room temperature organisms, is about 9 in railway air, 6·3 in the outside air; for body temperature organisms 4·6 for railway air, 2·2 for outside air.

3. The bacterial content of platform air, except on the City and South London Railway, would appear to be higher than that of carriage air; the total mean for platform air being 52 and for carriage air 42·8 organisms per 5 litres, or a ratio of 16·4 and 13·5 respectively to 10 of the open air. The higher proportion in platform air is generally speaking to be accounted for by the greater amount of draught and dust disturbance.

4. The ratios of the total bacterial content of railway carriage air and carriage and platform air on the six lines to open air are estimated in the following proportions:

Arthur William Bacot was born in London on April 28th, 1866, the third son and fourth child of Edmund Alexander and Harriet Bacot.

Much work has already been done in connection with the differentiation of true diphtheria bacilli from diphthomorphic organisms. Some observers have maintained that all disease producing strains could be distinguished from non-pathogenic types by their sugar reactions. Others have laid much stress on some particular forms of virulence test. More recently serological classification has been attempted in order to help to solve this problem.

With regard to sugar reactions Graham-Smith (1908) found that certain strains produced acid in media containing saccharose, while Hine (1913), on the other hand, maintained that no true diphtheria bacillus could ferment this sugar. Eagleton and Baxter (1922) have shown that diphtheria bacilli, virulent and non-virulent, ferment glucose but do not produce any change in saccharose. Jordan and his collaborators (1922) have found that virulent and non-virulent bacilli fermented glucose and maltose and produced no change in saccharose and that their action on dextrin was variable.

Work on the serological classification of these organisms has also shown some considerable variation. Havens (1920) isolated 206 strains of diphtheria bacilli. These represented cultures from acute cases, release cultures and cultures from healthy carriers. He found that these 206 strains could be divided into two serological groups, one group containing 169 strains, the other 37. There was no evidence of any cross agglutination. Durand (1920) isolated 255 strains of typical diptheria bacilli and after excluding those strains which would not form stable suspensions divided them into five groups, A. B. C. D. and E. containing 16, 8, 25, 61 and 40 strains respectively. Eighty-seven strains were found to be inagglutinable by these five sera but nevertheless could absorb agglutinins from such sera. His final figures for the five groups of bacilli are 18, 8, 31, 76 and 51. Durand and Guerin (1921) in a further paper showed that small isolated outbreaks were always due to the same type of bacillus. They also found that healthy carriers gave the same type of bacillus as was found in the cases arising from these sources of infection. Bell (1922) isolated 133 strains of B. diphtheriae indifferently collected from cases and carriers. He was able to agglutinate 80 per cent. of these strains by three monovalent sera. Group I contained 17 strains, Group II 8 strains, Group III 80 strains.

In view of the results which have just been described it has appeared important to investigate further a considerable number of strains as encountered in routine work, not only as regards the serological characteristics of the organisms but also in regard to their morphology, their biochemical reactions, and their virulence for guinea-pigs.

In a paper on “The Spread of Bacterial Infection” (Journal of Hygiene, xix. No. 4, March 22, 1921) Dr W. W. C. Topley has shown the results of many months' observation of the progress of an epidemic in a population of mice. The author thus describes the experiment: “This experiment lasted from May 21, 1919, until June 11, 1920.…The epidemic started about the middle of October, 1919, and continued in the manner to be described until the conclusion of the experiment.…The period May 21 to September 17 is omitted.…During this time there was no indication of the active spread of infection.…The whole experiment may be divided roughly into four periods. The first extends from May 21 to Sept. 17, and has been referred to already. From Sept. 18 and onwards, only normal mice were added to the cage. From this date until Jan. 5, 1920, the mice were added in such a way as to keep the total number in the cage roughly constant. From Jan. 6 to April 27, 1920, three normal mice were added each day, except on two occasions when none was added. The number of mice in the cage during this period varied from day to day, reaching a maximum immediately before each considerable wave of mortality, and falling to a minimum just before its cessation. During the final period from April 28 to June 11, 1920, two normal mice were added daily instead of three.”

Spontaneous agglutination is a not uncommon occurrence, especially in the case of organisms of the enteric and diphtheria group, where it causes some difficulty in serological diagnosis. Many investigations have been made to clear up the principles of this phenomenon. Recently Arkwright (1921) has thrown much light on this problem. He was able to isolate from old pure cultures of dysentery, typhoid, paratyphoid and enteritidis (Gaertner) bacilli, two variants which he named the “R” (rough) and “S” (smooth). The “R” variant agglutinates spontaneously in physiological saline and in broth, while the “S” variant forms good emulsions in both. Specific immune sera prepared for these two variants show only slight cross-agglutination. Arkwright regards the two forms as potentially inherent in most of the individuals in the young culture, which may become segregated in different individuals in old cultures. “Rough” varieties have been observed also by Zoeller (1922) in B. dysenteriae Shiga and by Schütze (1921) in B. paratyphosus B.

Although much work on the epidemiology of plague has been done in many parts of the world since the discovery of the Bacillus pestis in 1894, the origin of the outbreaks in Eastern and Southern Russia has, until a short time ago, remained obscure. Transbaikalia, together with extensive areas of Northern Manchuria and North-east Mongolia that are conterminous with it, and, again, the region in Southern Russia which includes the Kirghese and Kalmuck steppes and especially that portion of it which lies between the lower reaches of the rivers Volga and Ural have long been known to contain endemic foci of plague, and have been the source of considerable outbreaks of pneumonic plague. Thus, in the winter of 1878–79, an outbreak of this type at Vetlianka, a Cossack village on the right bank of the Volga, caused alarm in Western Europe. Competent epidemiologists–British, French and German–visited the village after the event, and examined the circumstances that favoured the spread of the infection. Their observations were brought together and analysed by Netten Radcliffe (1881) in his memorandum on plague, which gives the first adequate description of a pneumonic plague epidemic. The more recent epidemics of pneumonic plague, namely, those of Manchuria in 1910–11 with 50,000 deaths, Middle China in 1917–18 with 15,000 deaths, and Manchuria in 1920–21 with 9000 deaths, owed their origin to ill-defined centres of infection in the immense tract of land which includes Transbaikalia and which is contiguous to the north-west boundary of China.

In the preceding report (Topley and Ayrton, 1923 b) it has been shown that, if various strains of B. aertrycke be fed to mice and the subsequent events observed over a period of 42 days, and if all strains fed to the mice or isolated from their faeces or tissues be examined as regards their type of growth and their agglutination reactions, then the following associations are found to exist between the characters studied. The presence of group antigen is associated with persistent faecal excretion, whether the strain be rough or smooth. Roughness is associated with decreased faecal excretion and a decrease in the percentage mortality, when rough strains are compared with, smooth.

In the course of experiments on the cultivation of anaerobic bacilli, it was found advisable to be able to calculate with some certainty on the production of spores by the strains of anaerobes worked with. Observations were accordingly made of the conditions of cultivation by which sporulation appeared to be influenced. It was evident that in the case of B. sporogenes and other well-known types observed, the proportion of spores produced within a given period varied markedly in response to variations in the same factors that mainly influenced the rate of growth in cultures, viz. concentration of food material, incubation temperature, and reaction of medium. With the exception of cultures in media containing added carbohydrate, it was evidently also the rule that sporulation was most active in cultures commencing under conditions most favourable to rapidity of vegetation.

1. Normal rabbits injected intravenously with diphtheria antitoxin obtained from a horse vary considerably both in regard to the duration of Phase B and the duration and the intensity of Phase C.

2. Phase C, the phase of accelerated loss due to formation of precipitin appears sooner and is more pronounced in older rabbits.

3. The early appearance and more rapid formation of antibody (precipitin) in certain rabbits is indicative of naturally acquired immunity.

4. The highest concentration in the blood of rabbits injected subcutaneously with antitoxic horse serum is seen after 2 to 3 days, when it reaches approximately the same value as that of rabbits injected intravenously, and represents only one-third of the total antitoxin injected. Two hours after subcutaneous injection the antitoxic content is only one two-hundredth of that of a rabbit injected intravenously.

“Health is more affected by atmospheric conditions than by any other influence. From the moment the new born child utters its first cry, until life finally ceases, he is directly and immediately dependent on the atmosphere he breathes for continued existence. Records of sickness and of mortality clearly show that the nearer atmospheric conditions are to those of the open country, the better is the health of the population, and that the more these conditions are modified owing to urbanisation and industrialisation, the more pronounced is the ill-effect upon general health” (1).

The list of bacteria recorded as causative agents in enteric fever and “food-poisoning” increases steadily, and the proved pathogens already number eleven types, of which three have been found in both continued fever and food-poisoning, three in continued fever alone, and five in food-poisoning alone. This list will undoubtedly be added to when cultural methods for the diagnosis of these conditions are more widely employed, especially in the tropics.

A number of methods have been described for anaerobic cultivation in separate plates. The principle offers certain advantages over the cultivation of a number of plates in the same anaerobic container, which has to be opened whenever it is desired to inspect any particular culture; and it is useful where anaerobic plate cultures are needed only occasionally or in small numbers.

About a year ago the specific opportunity which our laboratories had been seeking presented itself. We were brought into touch with a farm on the outskirts of Vancouver engaged in the production of milk and the distributing of the same in the city. The proprietors of the farm, with at that time a herd of twenty-five grade cows, were endeavouring to supply a high grade milk, a milk having a low bacterial content, and secured from cattle giving a negative reaction to the tuberculin test. We approached the proprietors, Mr and Mrs Alexander Hill, and found that they were desirous of co-operating with us. At the same time their milk was obtaining a premium over the usual price obtained for market milk sold in the city. We made it clear that we were anxious to secure data which would be of use to milk producing farmers generally with respect to the possibility of producing a high quality milk; and they—Mr and Mrs Hill—assured us that they desired to have advice and help if the same could be forthcoming. Finally we made the following arrangement: (a) we were to be permitted to obtain samples of milk at any time deemed most suitable to ourselves, (b) these samples were to be examined for bacterial flora to the end that we might secure the data we needed, (c) no report on the examinations was to be sent from our laboratories until it became possible for us to publish the data as a whole, (d) the proprietors of the diary were to be given any information that was available from the laboratory data, but, under no circumstances, was the same to be used for publicity purposes, (e) we were to check up any of the procedures in vogue on the farm and dairy as far as proved to be possible without unduly interfering with the progress of our main project. The arrangement was agreed to mutually, and it is a matter of gratification to us that throughout the entire investigation the conditions detailed above were subscribed to loyally.

1. The course of disappearance of passive immunity in rabbits injected with diphtheria antitoxin obtained from goats, men, guinea-pigs and cows, consists of the same three phases that follow the injection of horse serum.

2. The rabbits examined were more responsive to goat, human and guinea-pig serum than to horse and cow serum.

3. The course of disappearance of passive immunity in rabbits, horses and guinea-pigs injected with homologous antitoxin, consists of Phases A and B only, and Phase B is far slower than when heterologous serum is injected into rabbits.

4. Sheep and goats eliminate antitoxin obtained from a horse at a very slow rate, and Phase C is hardly detectable.

5. Natural immunity of horses to diphtheria toxin is gradually acquired by a number of increasing responses to external stimuli.

“Virulence” is an ill-defined term and its use frequently gives rise to disputes, which remain unresolved because the disputants have no common basis other than that the term applies to organisms which cause disease. Thus it is imperative that the writer clearly defines the meaning of the term virulence as used in this paper, that the issue may not be confused by the reader approaching the subject from a different point of view.

The method most commonly employed for the separation of antitoxic globulins is that by means of Ammonium Sulphate, and this was the process used in these laboratories for several years until, at the end of 1917, the supply of the necessary salt was cut off. We were then thrown back on Sodium Sulphate, a salt we had used before the War but which had not given us satisfactory results. A few fresh trials, however, showed us the faults of our previous technique and proved that Na2SO4 could satisfactorily replace Am2SO4 and so, since February 1918, we have used only the former salt. Changes in detail have been made from time to time and we have now established a routine method which has for some time given us satisfactory results.

The experimental work on which the paper is based was undertaken with the object of investigating the factors which influence the growth of bacteria in various culture media. The B. influenzae was selected as the best organism because of the comparative difficulty of obtaining growth on ordinary media, and the feebleness of the growth when obtained. The importance of food accessory substances, or “Vitamins” for the proper growth and nourishment of animals is generally recognised, and the influence of the same on the nutrition of bacteria has been occupying the attention of various investigators during the last few years.

Towards the end of May, 1923, we received information of two fatal cases of bubonic plague in man, one near Yakoshih (a village on the Chinese Eastern Railway, 168 miles east of Manchouli), and the other at Haranor (in Transbaikalia, 52 miles west of Manchouli); both victims were Russians who had hunted tarbagans and afterwards developed axillary buboes.

1. There is some evidence that an injection of 0·00001 c.c. of horse serum causes an appreciable degree of active immunity to horse serum in rabbits.

2. Rabbits actively immune to horse serum receiving a series of injections of antitoxic horse serum at intervals of three to ten weeks eliminated the antitoxin at rapidly increasing rates.

3. An intravenous injection of antitoxic horse serum given six weeks after the second of a series of 0·5 c.c. in a rabbit already actively immune to horse serum was eliminated so rapidly owing to excess precipitin that less than 1/1000th remained 24 hours after injection.

4. Another rabbit which, six days after a previous injection, was given

serum intravenously, lost over 99 per cent. of the antitoxin within 15 minutes of the injection.

5. The maximum of circulating antitoxin detected after the second subcutaneous injection into a sensitised rabbit was only 1/15,000th of the amount injected.

6. Antitoxic horse serum injected intravenously into rabbits at weekly intervals was rapidly eliminated and less than 2 per cent. could be detected 24 hours after each injection.

7. After 12 daily intravenous injections of antitoxic horse serum into a rabbit, the antitoxic content was only one-half of that after the first injection; later injections were not eliminated so quickly and precipitin formation gradually ceased.

8. The rate of elimination of antitoxic horse serum in a rabbit may be greatly delayed by daily injections of normal horse serum.