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Chapter: Microbiology and Immunology: General Microbiology: History of Microbiology

Microorganisms as a Cause of Disease

Among various causes, the causes suggested for the occurrence of disease were the effect of supernatural phenomena like plan-etary alignments and effect of bad bodily humors; the faulty environment was also implicated.

Microorganisms as a Cause of Disease

Among various causes, the causes suggested for the occurrence of disease were the effect of supernatural phenomena like plan-etary alignments and effect of bad bodily humors; the faulty environment was also implicated.

     Even before microorganisms were seen, some investigators suspected their existence and responsibility for disease. Among others, the Roman philosopher Lucretius (about 98–55 BC) and the physician GirolamoFracastoro (1478–1553 AD) suggested that disease was caused by invisible living creatures. Fracastoro was much more than an author of the popular poem on syphi-lis. In his book “De contagione, contagiosismorbisetcuratione (On Contagion, Contagious Diseases, and their Treatment),” pub-lished in 1546, he proposed the revolutionary theory that infec-tious diseases are transmitted from person to person by minute invisible particles. He further suggested that infections spread from person to person by minute invisible seeds, or seminaria, that are self-replicating and act on the humors of the body to cause disease. His theories were ahead of their time, and it took about 200 years for the microscope to be invented and his theo-ries to be proved.

Antony van Leeuwenhoek: The Microscopist

The first person to observe and describe microorgan-isms accurately was an amateur microscopist Antony van Leeuwenhoek (1632–1723) of Delft, Holland. Leeuwenhoek earned his living as a draper and haberdasher (a dealer in men’s clothing and accessories), but spent much of his spare time constructing simple microscopes composed of double convex glass lenses held between two silver plates. His micro-scopes could magnify around 50–300 times. It is believed that he may have illuminated his liquid specimens by placing them between two pieces of glass and shining light on them at 45-degree angle to the specimen plane. This would have provided a form of dark-field illumination and made bacte-ria clearly visible. In 1673, Leeuwenhoek sent detailed letters describing his discoveries to the Royal Society of London. It is clear from his descriptions that he saw both bacteria and protozoa. But he did not evaluate these organisms as agents of disease.

Theory of Spontaneous Generation

There was a considerable controversy surrounding the origin of microbial pathogens. Some proposed that microorganisms originated from nonliving things by spontaneous generation even though larger organisms did not (theory of spontaneousgeneration). They pointed out that boiled extracts of hayor meat would give rise to microorganisms after sometime. Needham (1713–1781) on the basis of his experiments pro-posed that all organic matter contained a vital force that could confer the property of life to nonliving matter.

Louis Pasteur: Father of Microbiology

Louis Pasteur, French Microbiologist, is known as the father of medical microbiology for his immense contributions to the field of medical microbiology. He first coined the term “micro-biology” for the study of organisms of microscopic size. Many of his important contributions are discussed below.

 Germ theory of disease

Many other scientists have contributed to the theory of spon-taneous generation with their experiments, but it was Louis Pasteur (1822–1895) who settled it once for all. Pasteur first filtered air through cotton and found that objects resem-bling plant spores had been trapped. If a piece of cotton was placed in a sterile medium after air had been filtered through it, microbial growth appeared. Next he placed nutrient solu-tions in flasks, heated their necks in a flame, and drew them out into a variety of curves, while keeping the ends of the necks open to the atmosphere. Pasteur then boiled the solutions for a few minutes and allowed them to cool. No growth took place even though the contents of the flasks were exposed to the air. Pasteur pointed out that no growth occurred because dust and germs had been trapped on the walls of the curved necks. If the necks were broken, growth commenced immediately. By this Pasteur proved that all life even microbes arose only from their like and not de novogerm theory of disease). Pasteur had not only resolved the controversy by 1861 but also had shown how to keep solutions sterile.

    Support for the germ theory of disease began to accumulate in the early nineteenth century. AgostinoBassi (1773–1856) first showed that a microorganism could cause disease when he demonstrated in 1835 that the silkworm disease was due to a fungal infection. He also suggested that many diseases were due to microbial infections. In 1845, MJ Berkeley proved that the great potato blight of Ireland was caused by a fungus.


Pasteur for the first time demonstrated that he could kill many microorganisms in wine by heating and then rapidly cooling the wine, a process now called pasteurization. While developing methods for culturing microorganisms in special liquid broths, Pasteur discovered that some microorganisms require air, specifically oxygen, while others are active only in the absence of oxygen. He called these organisms as aerobic and anaerobic organisms, respectively.


In 1877, Pasteur studied anthrax, a disease mainly of cattle and sheep. He developed a vaccine using a weakened strain of the anthrax bacillus, Bacillus anthracis. He attenuated the culture of anthrax bacillus by incubation at high temperature of 42–43°C and inoculated the attenuated bacilli in the animals. He dem-onstrated that animals receiving inoculation of such attenu-ated strains developed specific protection against anthrax. The success of this concept of immunization was demonstrated by a public experiment on a farm at Pouilly-le-Fort in the year 1881. In that public demonstration, he vaccinated sheeps, goats, and cows with anthrax bacillus attenuated strains, but equal numbers of these animals were nonvaccinated. All the vaccinated as well as nonvaccinated animals were subsequently challenged with a virulent anthrax bacillus culture, after which only the vaccinated animals survived whereas nonvaccinated group of animals died of anthrax.

     In 1885, he also developed the first vaccine against rabies in humans that saved millions of human life worldwide. Pasteur coined the term “vaccine” to commemorate Edward Jenner who used such preparations for protection against smallpox. The Pasteur Institute, Paris and subsequently sim-ilar institutions were established in many countries of the world for the preparation of vaccines and for the study of infectious diseases.

 Control of silkworm disease

Following his successes with the study of fermentation, Pasteur was asked by the French government to investigate the cause of pébrine disease of silkworms that was disrupt-ing the silk industry. After several years of work, he showed that the disease was due to a protozoan parasite. The disease was controlled by raising caterpillars from eggs produced by healthy moths.

Joseph Lister: The Pioneer of Antiseptics

Indirect evidence that microorganisms are the agents of human disease came from the work of an English surgeon Joseph Lister (1827–1912) on the prevention of wound infections. Lister, impressed with Pasteur’s studies on the involvement of microorganisms in fermentation and putrefaction, developed a system of antiseptic surgery designed to prevent microorgan-isms from entering wounds. Instruments were heat sterilized and phenol was used on surgical dressings and at times sprayed over the surgical area. The approach was remarkably successful and transformed surgery after Lister published his findings in 1867. It also provided strong indirect evidence for the role of microorganisms in disease because phenol, which killed bacte-ria, also prevented wound infections.

Robert Koch: The Founder of Koch’s Postulates

The first direct demonstration of the role of bacteria in causing disease came from the study of anthrax by the German physi-cian Robert Koch (1843–1910). Koch used the criteria proposed by his former teacher, Jacob Henle (1809–1885), to establish the relationship between B. anthracis and anthrax, and he published his findings in 1876 briefly describing the scientific method he followed. In this experiment, Koch injected healthy mice with a material from diseased animals, and the mice became ill. After transferring anthrax by inoculation through a series of 20 mice, he incubated a piece of spleen containing the anthrax bacil-lus in beef serum. The bacilli grew, reproduced, and produced spores. When the isolated bacilli or spores were injected into mice, anthrax developed.

     During Koch’s studies on bacterial diseases, it became nec-essary to isolate suspected bacterial pathogens. His criteria for proving the causal relationship between a microorganism and a specific disease are known as Koch’s postulates.

 Koch’s postulates

Koch’s postulates (criteria) were useful to prove the claim that a microorganism isolated from a disease was indeed causally related to it. A microorganism was accepted as the causative agent of infectious disease, only when it satisfied all the follow-ing criteria (Fig. 1-1):

1.     The microorganism must be present in every case of the disease but absent from healthy host.

2.     The suspected microorganism must be isolated and grown in a pure culture from lesions of the disease.

3.     The isolated organism, in pure culture, when inoculated in suitable laboratory animals should produce a similar disease.

4.     The same microorganism must be isolated again in pure culture from the lesions produced in experimental animals.

The specific antibodies to the bacterium should be demonstra-ble in the serum of patient suffering from the disease. This was an additional criterion that was introduced subsequently.

     Most of the human bacterial pathogens satisfy Koch’s postulates except for those of Mycobacterium leprae andTreponemapallidum, the causative agent of leprosy and syphilis,respectively. Both these bacteria are yet to be grown in cell-free culture media.

 Solid medium for culture of bacteria

Koch pioneered the use of agar as a base for culture media. He developed the pour plate method and was the first to use solid culture media for culture of bacteria. This development made possible the isolation of pure cultures that contained only one type of bacterium and directly stimulated progress in all areas of bacteriology. Koch also developed media suitable for grow-ing bacteria isolated from the body. Because of their similarity to body fluids, meat extracts and protein digests were used as nutrient sources. The result was the development of nutrient broth and nutrient agar media that are still in wide use today. By 1882, Koch had used these techniques to isolate the bacil-lus that caused tuberculosis in humans. Koch also discovered that cholera was caused by Vibrio cholerae. He invented the hot air oven and steam sterilizer, and also introduced methods to find out the efficacy of antiseptics. There followed a golden age of about 30–40 years in which most of the major bacterial pathogens were isolated.

 Koch’s phenomenon

Koch’s phenomenon is a hypersensitivity reaction against tuberculosis bacilli demonstrated in guinea pigs. This was first demonstrated by Koch, who showed that guinea pigs already infected with tubercle bacillus, on challenge with tubercle bacillus or its protein, developed an exaggerated inflammatory response.

Self-Experimentation Studies

To study diseases in a more elaborate and controlled fashion, there were a few dedicated researchers who went to the extremes of self-experimentation. The discovery that hookworm infes-tation spread by fecal–oral route was first demonstrated by Arthur Loos in 1898. This was known during his attempts at studying Strongyloidesstercoralis by swallowing its larvae and accidentally swallowing a fecal inoculum with hookworm eggs instead!

These attempts did not always have happy and produc-tive endings as is illustrated by the case of Daniel Carrion (1858–1895), a medical student in Lima, Peru. He man-aged to prove that the same organism (later identified to be Bartonellabacilliformis) caused a chronic skin lesion calledver-rugaperuanaand another serious disease calledOroya fever.This he did by inoculating himself with material from the warts of the skin lesion. He did develop Oroya fever as he had hypothesized, but the experiment costed him his life when he succumbed to the disease. In the subsequent 50 years, numer-ous microorganisms were identified as the causative agents of important human diseases and their discovery elucidated (Table 1-1).

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