The acute period of the struggle in defence of the phagocyte theory now seemed to have come to an end and Metchnikoff turned his thoughts towards a new field of ideas.

Having elucidated the essence of inflammation, he wished to study the origin of another pathological symptom, i.e. the rise in temperature which constitutes a feverish condition. To that end he undertook a succession of experiments on cold-blooded animals; he injected microbes into crocodiles and serpents, hoping thus to provoke a rise in their temperature. But those experiments did not give the results expected.

In the meanwhile (1892) cholera had made its appearance in France; the specificity of the cholera vibrio was not finally established at that time. The observations made by Pettenkoffer on the immunity of certain regions, despite the presence of the cholera vibrio in the water, and the experiments made upon himself by that scientist, seemed to plead against the specificity of the cholera vibrio; but other facts spoke in its favour. Desirous of solving this question, Metchnikoff went to a cholera centre in Brittany in order to fetch the necessary materials. Having done so, he attempted to produce cholera in divers kinds of animals, but without success.

As he failed to solve the problem of the specificity of the cholera vibrio on animals, he resolved to experiment upon himself and consumed a culture of cholera vibriones. He did not contract cholera, which made him doubt the specificity of the vibrio, and therefore he consented to repeat the experiment on one of his workers (M. Latapie) who offered to submit to it: the result was the same. He then did not hesitate to accept the offer of a second volunteer (M. Jupille). The preceding results having led him to suppose that the cholera vibrio became attenuated in vitro and might perhaps serve as a vaccine against cholera, he gave a culture of long standing to the young volunteer.

To his astonishment and despair, Jupille began to manifest the typical symptoms of cholera, and a doctor who was particularly conversant with the clinical chart of the disease declared the case a severe one because of the nervous symptoms which accompanied it.

Metchnikoff was in mortal anxiety, and even said to himself that he could not survive a fatal issue. Fortunately the patient recovered, and this terrifying experiment proved indisputably the specificity of the cholera vibrio. Yet the irregularity of its action showed that in certain cases conditions existed which prevented the inception of the disease, and Metchnikoff supposed that this might be due to the action of the different intestinal micro-organisms.

In order to simplify the question, he began by making experiments outside the organism. He sowed the cholera vibrio with divers other microbes and saw that some of them facilitated its culture whilst others prevented it. Similar experiments within the organism of animals gave no conclusive results; the simultaneous ingestion of the cholera vibrio and of favourable microbes did not induce cholera.

The flora of the intestines, complex as it is, probably played a part on which it was difficult to throw any light. Yet Metchnikoff did not give up the idea of producing a vaccine against this disease with attenuated microbes, or, if not, to prevent its inception by preventive microbes. His thesis was strengthened when one of his pupils, Dr. Sanarelli, discovered a series of choleriform bacilli in the absence of any cholera epidemic, one of those microbes being found at Versailles, a town which had remained immune during every cholera epidemic.

Metchnikoff thought that this microbe, or some choleriform bacillus, similar though not specific, probably served as a natural vaccine against cholera in those localities which were spared by the epidemic though the cholera vibrio was brought there. This was a question that could only be solved by experiment.

At the time when he had himself absorbed a cholera culture, Metchnikoff admitted the risk of catching the disease; still, his eagerness to solve the problem had silenced in him all other considerations and feelings opposed to his irresistible desire to attempt the experiment. This “psychosis,” as he himself called it later, recurred now, in spite of all the emotions he had gone through on the previous occasion, and he decided once again to experiment on man. It is true that he now only had to deal with choleriform microbes from Versailles which he believed to be quite harmless as they came from the water of a locality free from cholera. He therefore ingested some of the Versailles choleriform vibriones and gave some to several other people. Contrary to expectation, one of the latter, an incurable epileptic, showed some symptoms of cholera, but recovered. But as, a short time later, this patient died from a cause which remained obscure, Metchnikoff thought that possibly the experiment might have had something to do with it, and finally resolved to perform no other experiments on human beings.

How could that unforeseen result be explained? Metchnikoff supposed that the intestine of the subject contained favourable microbes which had exalted the virulence of the bacillus, in itself weak and innocuous. If it were so, then certain intestinal microbes would influence the inception of diseases and the action of the micro-organisms would vary according to the society in which they found themselves. As such problems could only be solved through experiment, he again energetically sought for a means of conferring cholera upon animals. After many failures and difficulties, it occurred to him to try new-born animals whose intestinal flora, not yet developed, could not interfere with the swarming of the ingested bacilli. He chose young suckling rabbits for his experiments and, with the aid of favourable microbes, he succeeded at last in giving them characteristic cholera, through ingestion; thus it became possible to study intestinal cholera on these animals.

However, numerous researches on the prevention of cholera by means of divers microbes gave no results sufficiently conclusive to permit their application to human beings. The problem was rendered extremely complicated and difficult by the many and varied influences of numerous intestinal microbes and the inconstancy of microbian species in the same individual.