Koch’s Postulates

1. The bacteria must be present in every case of the disease.
2. The bacteria must be isolated from the host with the disease and grown in pure culture.
3. The specific disease must be reproduced when a pure culture of the bacteria is inoculated into a healthy susceptible host.
4. The bacteria must be recoverable from the experimentally infected host.

 

Prior to 1882, it was generally accepted that tuberculosis (or consumption) arose directly and spontaneously from within the body of the patient. The clinicians of the day referred to tuberculosis by a number of different terms, apart from tuberculosis: The white death, the white plague, the captain of the men of death, the robber of youth, the graveyard cough, the King’s Evil.

At 7:00pm on the 24th of March 1882, a former country General Practitioner named Robert Koch, stood up before an audience at the monthly meeting of the Berlin Physiological Society and gave a lecture titled “On Tuberculosis.” 36 scientists were present. Koch had not previously disclosed his work, which had been carried behind the door of his office.

Koch chose the Physiological Society because he had been barred from giving lectures at the Medical School by Professor Rudolph Virchow, who actively detested Dr. Koch. (In 1882, Virchow was Chair for Pathological Anatomy and Physiology at the Friedrich-Wilhelms University, as well as Director of the newly built Institute for Pathology on the premises of the Charité. Koch was in good company as Virchow also considered Charles Darwin to be a fraud.)

Koch was not a good speaker and he started hesitantly. He painstakingly outlined all the steps in his research, beginning with an acknowledgement of the work of Jean Villeman, who had demonstrated that TB was transmissible (rabbits) but his work had been ignored.  He then mentioned four other researchers who had demonstrated transmissibility via the anterior chamber of the eye (rabbit) and by inhalation (dog), but all had reached the conclusion that “the direct demonstration of the tubercle [agent] is still an unsolved problem.”

Koch then proceeded to describe the discovery of a stain that allowed him to demonstrate the presence of the TB bacillus in tissues and samples from tuberculous patients. He empathised that the bacilli were seen in proximity to pathological changes associated with TB. His scrupulous description of the stain included the fact that the stain took 24 hours to perform.

In his lecture notes, Koch’s description of the tubercle bacillus was meticulous and included descriptions of the appearance of the bacillus in the various stages of TB infection. He also commented on the resemblance to Hansen’s bacillus, which had been discovered in 1873 by Dr. Gerhard Hansen, a Norwegian pathologist who described the presence of a bacillus in lepromatous lesions. Hansen became the first proponent of the theory that a chronic disease could be caused by a microbe and carried out experiments on animal inoculation (unsuccessfully) using leprosy infected tissues.  He additionally attempted to induce leprosy in a human subject, a female patient who had been diagnosed with leprosy previously.  Hansen placed the infectious material in the patient’s eye without seeking permission to do so. She sued him and he lost his job.

Koch, in spotting the resemblance, had no idea M. tuberculosis and M. leprae were of the same species.  They just looked the same.

At the Physiological Society meeting, Koch stressed that the TB bacillus was seen in all cases of TB, and not in controls. He also linked the quantities of bacilli to the stage of the infection. But the evening was not yet half-way through.

Koch then described how he had developed a solid medium which supported the growth of the bacillus. He had been down this path before and had been the first person to grow bacteria deliberately on solid media. The reason for this, he explained, was to obtain samples of the bacilli free of tissue and body fluids. The media was manufactured from heat coagulated serum from cows and sheep. He fully described the appearance of the colonies.

Paul Ehrlich, a 28 year old medical student (and reputedly hard drinker and bon vivant) was in the audience and witnessed the presentation. He later described it as his most gripping experience in science. Ehrlich, then training in dermatology, went straight home and that night started experimenting with stains, using a culture given to him by Robert Koch. Five years later Ehrlich diagnosed himself as having TB. He nevertheless survived to become a great researcher, discovering Salvarsan, the first effective treatment for syphilis.

Koch then moved on to his experiments with guinea pigs. He had infected 217 guinea pigs with TB and described his findings. He also observed that he had never seen a case of TB in uninoculated animals, but in inoculated guinea pigs the appearances had been strikingly similar and TB was isolated from the infected lesions at autopsy in all cases. His description of the infected organs and nodes was clear and precise. He used his experience of animal study to describe what would become  Koch’s postulates:

It was necessary to isolate the bacilli from the body; to grow them in pure culture until they were freed from any disease-product of the animal organism which might adhere to them; and by administering the isolated bacilli to animals, to reproduce the same morbid condition which is obtained by inoculation with spontaneously developed  tuberculous material.

At the end of the lecture the audience sat in an unnerving silence, and there was no applause.

Virchow put on his hat and stormed from the room without saying a word. Koch, stunned by the silence, was probably racked with what we now know as imposter syndrome. Three weeks later he published in the Berlin Clinical Weekly and sent a copy to John Tyndall in London. Koch had used Tyndall’s ideas on the use of serum to make a solid medium. Twelve days later Tyndall sent a letter to The Times. The New York Times received the news several weeks later and erupted editorially on the slow progress of “the news of one of the great scientific discoveries of the age” when the “progress or receding of sundry royal gouts are given the wings of lightning.”

Louis Pasteur, talking about the role luck plays in research said, “Chance favours the prepared mind.” Koch had previously worked on a zoonosis (anthrax) so was primed for the challenge of TB. The rest was hard, lonely, repetitive work.

By the middle of 1882 Paul Ehrlich had come up with an improved stain. In 1883, Ziehl and Neelsen described the ZN stain.

The Stain

The Ziehl-Neesen stain is used worldwide today as the simplest diagnostic test for tuberculosis. The stain has more variations than lotto, but in the classic form is excellent for spotting M. tuberculosis.

The following years saw many variations tried and numerous researchers studying TB. One of the findings they occasionally described was the existence of round, coccoid forms of TB. In 2005, I saw them in the blood culture of a Crohn’s patient. This has led to 15 years of intermittent experimentation to enhance the stain and study the relationship between mycobacteria and Crohn’s disease.

As part of this work, we have found that cell-wall deficient strains of mycobacteria are present in almost all human controls. The explanation for this probably lies in Netea’s work on the trained immune system.

The challenge for us has been to develop media that distinguishes between these commensal stealth forms of mycobacteria, and the organisms more clearly associated with Crohn’s disease.  To this end a new medium was developed, which we call 7DN.

For more information on the research of John Aitken and Otakaro Pathways, see the following resources:

2018 Berkeley Conference presentation:
https://humanpara.org/berkeley-conference-the-development-and-validation-of-a-novel-biomarker-for-diagnosis-of-inflammatory-bowel-disease/

2019 Journal Article:
https://www.nzimls.org.nz/journals-recent,article,,77,336,A+revaluation+of+the+use+of+conventional+Ziehl-Neelsen+stain+for+detection+of+non-tuberculous+mycobacteria.html

Human Para Blog Articles:
https://humanpara.org/news/