The present paper contains the results of an enquiry into the differentiation of certain organisms of varying source, which present characters more or less closely allied to those of the “mannite” or “Flexner” type of the dysentery bacillus.

On June 14th, 1910, the attention of one of us was directed by the House Physician of the Hampstead General Hospital, to an outbreak in Paddington of what was thought to be “food-poisoning.” Two of the patients were at the time under his care.

(1) B. typhosus ferments dulcite-peptone-water in about 10 days. By sub-culture this is readily shortenedto one day. It produces nonfermenting colonies on dulcite-agar plates. The colonies show after five days and upwards, secondary colonies as papillae, which may or may not have an acid reaction.

Growth of B. typhosus on dulcite media readily gives rise to bacilli forming, from the commencement, acid colonies on dulcite plates. The dulcite character has a tendency to atavism in the early stage which further growth on dulcite removes.

(2) B. typhosus can be trained to ferment lactose, only with very great difficulty. Definite and conclusive results have been obtained by meonly with the white atavists of Twort's trained lactose strain. Other strains (20) have not yielded to such training methods, although many of them have been trained for over a year. The lactose character when acquired is very unstable, and is soon lost when growth takes place in a lactose-free medium. A fermenting colony on replating gives rise to a mixture of fermenting and non-fermenting colonies.

(3) All strains of B. typhosus which I have examined show the secondary colonies on isodulcite described by Reiner Müller. Growth on isodulcite yields directly a strain without this papillae-forming power. So-called “acid” papillae as described by Reiner Müller have not been observed in the strains (20) investigated by me. This new strain does not regain the power to form isodulcite papillae by frequent sub-culture apart from isodulcite. Apparently the acquired character has arisen by mutation. Fermentation of isodulcite requires a considerably longer process of training.

(4) The above three characters vary in permanency. The permanency varies inversely as the time required for the selection or training.

(5) Papillae-formation arising on carbohydrate media on colonies of members of the typhoid-coli group may indicate variation by slow degrees just as well as by mutation. I have observed it in normal typhoid varying towards dulcite and lactose as well as isodulcite. In the case of Twort's strain which fermented sorbite-agar.

Formation of papillae simply indicates apparently that the variation only affects very few individuals of the colony to a great extent, and does not guarantee permanency in the character.

If the papillae arise early and without sub-culture, the evidence adduced so far suggests that the character will have considerble permanency. In the case of dulcite the results show that the formation of papillae on certain carbohydrate media cannot, as Reiner Müller stated, be regarded as definitely specific for the variety of microorganism exhibiting them.

The variations detailed in the above communication do not in our view invalidate the sugar tests as aids to differentiation, since these chemical functions are subject to variation probably to no greater extent than other functions of bacteria. Further, the ability of an organism to vary in any particular direction may be of considerable value in differential diagnosis.

This instability makes it imperative to have as large a number of differentiating characters as possible for every species, and also to take care in laboratory culture that we do not grow bacteria under conditions likely to alter their characters. It would be inadvisable for example to propagate the B. typhosus from a dulcite culture a few days old if one desired it to conform to the normal typhoid type in respect of dulcite.

In sending this paper to the Press I would like to acknowledge my indebtedness to Dr Martin, who has supervised my work on behalf of the British Medical Association, and to Dr Ledingham, who suggested the work, from both of whom I have received much valuable help and criticism. Many other colleagues at the Lister Institute have helped me from time to time in points of difficulty, and I take this opportunity of thanking them.

1. The term “paratyphoid group of bacilli” has been confined in the present enquiry to those strains which in their cultural characters and agglutination reactions are indistinguishable from B. suipestifer and B. paratyphosus (B). B. enteritidis Gaertner and B. paratyphosus (A) are excluded.

2. The strains of bacilli belonging to the “paratyphoid” group can be separated into two classes by means of the absorption method, namely one group identical with recognised, standard strains of B. suipestifer and one identical with standard strains of B. paratyphosus (B); and we regard these two bacilli as separate organisms.

3. In our experience, B. suipestifer has been found only in food or in outbreaks of acute illness attributable to food-poisoning, whereas B. paratyphosus (B) has been found in cases of paratyphoid fever or in persistent paratyphoid “carriers.”.

4. We are disposed to put forward the suggestion indicated by these observations, that these two organisms have a different distribution in nature, the normal habitat of B. suipestifer being the alimentary canal of the pig (and other animals) and of food derived from such animals, whereas the normal habitat of B. paratyphosus (B) is the human alimentary tract (including the gall-bladder).

5. The examination of cultures from the faeces and urine of 300 typhoid convalescents was completely negative, neither B. suipestifer nor B. paratyphosus (B) being obtained. These observations confirm those of other writers, and, apart from carrier cases, the occurrence of B. paratyphosus (B) in healthy human beings appears to be unknown in this country.

Our thanks are due to Drs Fisher, Trautmann, Uhlenhuth, Williams, Rommeler, Prigge, Schern, Friedrichs, Savage and Fowler, who have very kindly supplied us with strains of bacilli for investigation.

The outbreak of food infection which we are about to describe is of interest from more than one point of view. Not only was it possible to determine the organism which caused the outbreak and to trace its source to a paratyphoid carrier, but the bacteriological results yielded an important contribution to the difficult question of the differentiation of Bacillus paratyphosus B. and B. enteritidis, which for some years has been much discussed among bacteriologists.

The general consideration of these results only tends to confirm the conclusions already arrived at. A uniform type of life evidently tends to a fairly uniform excretion of tissue cells from the udder. The case of Goat II also emphasises the effect of outside causes in increasing temporarily this excretion, while some of the samples of human milk show plainly that very high cell counts are not by any means necessarily connected with any diseased or disordered condition of the mammary gland.

We again emphasise the view we have already stated, viz. that in the cow the udder must be looked upon as an organ which has by breeding and selection been brought to an artificial condition of milk secretion and that this has been accompanied by a stimulation of the tissues to cell proliferation and that this proliferation may quite easily be caused to become abnormally great, leading to the appearance of an increased number of cells in the secretion. In support of this view we lay great stress on the fact constantly noticed by us, viz. that when the cell count is high for any reason, the cells themselves are always well-defined, showing little signs of degeneration, and also stain in a much more characteristic and definite manner, a fact which is diffcult to explain if they are to be considered to be blood elements.

The recognition of latent tuberculosis, difficult as it often is in the human subject, is of necessity more difficult in the case of cattle. It is especially in early disease that the serum reactions prove often most useful as an aid to diagnosis, but in cattle a proof is also often desired that no pronounced disease is present, because it is by no means of rare occurrence that an apparently fat and healthy animal must be rejected after slaughter.

During my work in Yucatán (tropical Mexico) I observed that we nearly always found by the current analysis of urine, a well pronounced indican reaction, certainly in more than 80% of the cases examined; and frequently the blue colour obtained by the Obermeyer or Jaffé reaction was so strong that one would have been inclined, if in Europe, to postulate the presence of intestinal obstruction or some serious intestinal disorder. Experience soon taught us however that the reaction had not the same significance in Yucatán and probably in other tropical countries as it possesses elsewhere. With the object of studying the underlying causes I suggested to one of my assistants an investigation of the indole-producing power of B. coli isolated from the faeces of different individuals. The results which have been published in a dissertation (Hernandez, 1908) were naturally very incomplete but the work led me to realise the importance of a quantitative indole-determination, and, as I did not find described in the literature any method suitable for our purpose, I proceeded to work out a method for myself. I used the Salkowski test, with nitrite of potassium, sulphuric acid, and extraction with amyl-alcohol, preparing a series of tubes containing determined quantities of indole in progressive concentrations. The indole I distilled from bacterial cultures. The scale kept well, having been preserved for three years in the dark and only brought into the light when in use.

(1) As far as is at present known, the great majority of the fleas infesting Mus rattus and Mus decumanus in different parts of the world, belong to either the species Xenopsylla cheopis, Ceratophyllus fasciatus, Ceratophyllus anisus, Ctenopsylla musculi or Ctenophthalmus agyrtes or are comprised of some admixture of these five species.

(2) Xenopsylla cheopis is the most prevalent in the tropics and sub-tropical regions and often occurs there to the almost complete exclusion of other species. It is common during summer and autumn in some of the warmer parts of the temperate zone, more especially in ports which have maritime intercourse with the tropics.

(3) In the cooler regions Ceratophyllus fasciatus is the most universally distributed flea and is associated with more or less of Ctenopsylla musculi and Ctenophthalmus agyrtes according to the locality and the habitat of the particular rats.

(4) In Japan Ceratophyllus fasciatus is replaced by Ceratophyllus anisus, a closely allied species.

(5) The numerous other fleas which have been captured off rats are only occasional visitors.

(6) Ceratophyllus fasciatus, like Xenopsylla cheopis, readily bites man. Out of 517 experiments 308 fed, or 59% were positive. In 101 experiments, under identical circumstances with a rat, 59, or 58.4% of the fleas fed.

(7) The experiments with Ceratophyllus fasciatus were made upon eight persons and evidence was obtained of preference on the part of the insects for particular individuals.

(8) 111 experiments were made with 46 specimens of Ctenopsylla musculi; only 4 fed=3.6% whereas 9 out of 11 fed on a mouse.

(9) 68 specimens of Ctenophthalmus agyrtes were tried, in some cases upon three persons. None fed, whereas 11 out of 19 of the same fleas fed on a rat under identical conditions.