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Later on, a few convalescent cases of amoebic dysentery, and also "cyst- carriers" giving no history whatever of dysentery, were admitted into the hospital, and I was able to describe, also for the first time, the occurrence of definitely small cysts of E. histolytica, with a diameter not exceeding 10 u., instead of the average of 12-14 u., shown by the ordinary cysts. I regarded (and still regard) these as indicating a distinct "strain" of the parasite, for which I adopted James's name minuta," as being the best, although there is considerable doubt whether the form so described by James was in reality this distinctively small type. In the same paper, which was under the joint authorship of Penfold and myself; (2), we discussed the question of the commonly occurring infections with the Flagellate parasite Lamblia. We had ample evidence (which has been confirmed by further work on my own part as well as by others) that these infections may be a cause of very troublesome chronic diarrhoea. We indicated certain methods of effecting a cure, which we had reason to consider promising; and, though there is no doubt that these infections are often extremely intractable and difficult to cure, subse- quent work has shown that bismuth salicylate, which we were the first to recom- mend, is, when given in large doses, perhaps the most useful drug in this connexion. Late in November I was asked by the War Office, at the same time as were Messrs. Ledingham and Wenyon, if I were willing to give service abroad. I at once expressed my willingness to do so, but found that the War Office refused to give me a commission because I possessed no medical qualification, although my As, however, I was services were asked for purely for protozoological work! anxious to be of what service I could during the War, I eventually agreed to go as a civilian. I will take this opportunity of saying that, in spite of my anomalous position, particularly while in the field, and notwithstanding that I had no official authority and no official recognition of my work, I shall always be glad that I gave this service and did not stay at home. I learnt for myself of what vital assistance a man with an expert knowledge of malarial parasites was to troops in a dangerous malarious country such as Palestine.
Owing, however, to delay in getting the necessary sanction from the Treasury
Up to then for my pay, it was not until May, 1916, that I left England. remained at the King George Hospital, under the conditions referred to at the beginning of this report. During the month or so before I left, owing to the routine work being lighter, Dr. Penfold and myself, with some assistance from Mr. Drew, began experiments in an endeavour to cultivate E. histolytica artificially. In the first place we had to find a means of inducing the Amoebae to excyst. By repeated centrifugalization and washings we managed to obtain cysts fairly free from bacteria. After many attempts with various combinations of bile, dilute hydrochloric acid, and pancreatic juice, we found a reliable method of inducing excystation. This was by leaving the cysts in a medium consisting of nutrient broth to which a certain proportion of Liquor pancreaticus Benger was added, in the incubator, at 37° C., for from four to six hours. Somewhat to our surprise this method did not succeed either with the cysts of E. coli or of Lamblia. We were unable to obtain the excystation of either of the two latter parasites.
The mode of excystation is most interesting. As the ripe cysts always possess (typically) four nuclei (the corresponding number in the case of E. coli-cysts being eight), we thought it quite probable that this nuclear division would be followed by cytoplasmic division, and that four little amoebulæ would emerge from the cyst This is, however, never the case. A small pore appears at one point in the cyst wall, and through this the entire contents of the cyst creep out as one small Amœba. For some while previous to the actual emergence the cyst-contents show slight amoeboid movements. The immediate subsequent development we were, unfor- tunately, not able to ascertain. After moving about a little the Amoebae in our We endeavoured observation-preparations became languid and tended to break
up.
to cultivate these excysted Amoebae by putting the contents of tubes which we knew to contain such into blood-broth tubes. Undoubtedly multiplication took place in these tubes; but whether it was preceded by conjugation of small amœbulae we were unable to say. After twenty-four hours there were certainly more active Amoebae in the drops examined than there would have been of merely excysted forms. (It may be remarked that only a small proportion of the cysts present ever excysted; by the above method we could always count on obtaining some excysta tion, but, of course, a considerable number of cysts remained unaltered. the condi- tions naturally being less favourable than the normal environment at the time of excystation). Some of the Amoebae, in fact, were fairly large and contained
37
ingested red blood-cells. Unfortunately, we could not keep these cultures going. By the end of forty-eight hours the few bacteria unavoidably left in the original material had multiplied to so great an extent in the rich medium that the latter was no longer suitable for the Amoeba and they ceased to thrive. At this period We had intended to very few were found, and these were obviously moribund. pursue this work, but I had then to leave for Egypt. This is the first recorded observation of the excystation and artificial cultivation of E. histolytica. A note on our results (4) was published shortly after my departure.
I returned home, after two years and nine months' absence, about the middle of January, 1919. Although never having had any leave to England during this time I was not granted any furlough on my return, being demobilized straight away. I felt myself entitled, however, to a month's leave before resuming work. In the middle of February I resumed my former position in connexion with the University Department of Protozoology, pending a decision as to what was to be its future. I found that other workers were in full possession of our old labora tories and rooms at the Lister Institute, and were likely to remain for some time longer. Just at this time my friend Dr. J. D. Thomson, who was formerly co-Assistant with myself to the late Prof. Minchin, was in charge of the Laboratory at the King George Hospital; and on account of the pressure of the work there at that time he asked me if I could assist him. In view of the above-mentioned fact, it was impossible to do any work at the Lister, so the University authorities very kindly sanctioned my taking up this post, for which I now received payment. I remained at the hospital until it closed down at the end of June.
At first our time was much occupied with routine work, a large number of nialarial examinations having to be måde. During the second part of this period Dr. Thomson and myself were asked to write the article on the malarial parasites for Byam and Archibald's forthcoming treatise on the "Practice of Medicine in the Tropics," which it is hoped to make the standard work on the subject. As malarial parasites are by far the most important disease-causing Protozoa, and malaria itself is one of the most formidable of tropical diseases, I felt my time could not be better employed than by attempting to repeat, in some degree, for one section of applied Protozoology, the valuable work performed by the late Prof. Minchin for pure Protozoology, when he wrote his book on the Protozoa generally. The experience of the War has shown that pure Protozoology is to a very large extent intimately bound up with applied Protozoology; and there can be no doubt that the practical value of the study of Protozoa is largely due to the fact that so many of the parasitic forms are associated with disease in man and animals.
The work involved a considerably greater expenditure of time and labour than We resolved to have a really good repre- we anticipated when we undertook it. sentative series of illustrations; and notwithstanding that we had plenty of our own material, our endeavour to find the best, most suitable specimens for our figures, which number over 130. entailed far more searching of films than we had realized would be necessary. The need for true, life-like illustrations of malarial parasites is apparent from a comparison of existing text-books. The best hitherto published "Handb. d. Tropenkrank- are in a German work (Ziemann's article in Mense's heiten "); and we are of the opinion that our figures are a distinct improvement on these. It has been our aim-I do not know how far we have succeeded to produce an account which may be regarded as a standard one for some time to come. 18 hoped that the volume on Malaria will appear early in the new year.
After the closing down of the hospital and our completion of the above work. I compiled, as in former years, the annual Record of Protozoa for the Zoological Society (5).
1t
From my
I was then desirous of clearing off certain work remaining over, a result of observations connected with my experiences whilst on active service. recorded notes, and the views which I had formed in the course of my work, 1 wrote a paper on the relative proportions of amoebic to bacillary dysentery among the troops of the Egyptian Expeditionary Force, and discussed the important question of the value of the cellular exudate as a means of distinguishing between these two types of dysentery. I came to the conclusion that while in certain well- defined cases the nature of the cellular-exudate may of itself suffice to enable a correct diagnosis to be made, nevertheless, in a large number of cases the most reliable character, and the one upon which most stress should be laid in routine work, is the determination of the presence or absence of E. histolytica. This paper
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