September 25, 2020

HISTORY OF MICROBIOLOGY

The Theory of Spontaneous Generation

The Greek philosopher Aristotle (384–322 BC) was one of the earliest recorded scholars to articulate the theory of spontaneous generation, the notion that life can arise from nonliving matter. Aristotle proposed that life arose from nonliving material if the material contained pneuma (“vital heat”). As evidence, he noted several instances of the appearance of animals from environments previously devoid of such animals, such as the seemingly sudden appearance of fish in a new puddle of water.

This theory persisted into the seventeenth century, when scientists undertook additional experimentation to support or disprove it. By this time, the proponents of the theory cited how frogs simply seem to appear along the muddy banks of the Nile River in Egypt during the annual flooding. Others observed that mice simply appeared among grain stored in barns with thatched roofs. When the roof leaked and the grain molded, mice appeared. Jan Baptista van Helmont, a seventeenth century Flemish scientist, proposed that mice could arise from rags and wheat kernels left in an open container for 3 weeks. In reality, such habitats provided ideal food sources and shelter for mouse populations to flourish.

However, one of van Helmont’s contemporaries, Italian physician Francesco Redi (1626–1697), performed an experiment in 1668 that was one of the first to refute the idea that maggots (the larvae of flies) spontaneously generate on meat left out in the open air. He predicted that preventing flies from having direct contact with the meat would also prevent the appearance of maggots. Redi left meat in each of six containers (Figure 1). Two were open to the air, two were covered with gauze, and two were tightly sealed. His hypothesis was supported when maggots developed in the uncovered jars, but no maggots appeared in either the gauze-covered or the tightly sealed jars. He concluded that maggots could only form when flies were allowed to lay eggs in the meat, and that the maggots were the offspring of flies, not the product of spontaneous generation.

An open container with meat has flies and the formation of maggots in meat. A cork-sealed container of meat has no flies and no formation of maggots in meat. A gauze covered container of meat has flies and maggots on the surface of the gauze but no maggots in the meat.

Figure 1. Francesco Redi’s experimental setup consisted of an open container, a container sealed with a cork top, and a container covered in mesh that let in air but not flies. Maggots only appeared on the meat in the open container. However, maggots were also found on the gauze of the gauze-covered container.

In 1745, John Needham (1713–1781) published a report of his own experiments, in which he briefly boiled broth infused with plant or animal matter, hoping to kill all preexisting microbes.^[2] He then sealed the flasks. After a few days, Needham observed that the broth had become cloudy and a single drop contained numerous microscopic creatures. He argued that the new microbes must have arisen spontaneously. In reality, however, he likely did not boil the broth enough to kill all preexisting microbes.

Lazzaro Spallanzani (1729–1799) did not agree with Needham’s conclusions, however, and performed hundreds of carefully executed experiments using heated broth.^[3] As in Needham’s experiment, broth in sealed jars and unsealed jars was infused with plant and animal matter. Spallanzani’s results contradicted the findings of Needham: Heated but sealed flasks remained clear, without any signs of spontaneous growth, unless the flasks were subsequently opened to the air. This suggested that microbes were introduced into these flasks from the air. In response to Spallanzani’s findings, Needham argued that life originates from a “life force” that was destroyed during Spallanzani’s extended boiling. Any subsequent sealing of the flasks then prevented new life force from entering and causing spontaneous generation.

Finally it was Louis Pasteur who disproved spontaneous generation theory with his famous Swan — Neck Flask experiment.

Note: Swan — Neck Flask experiment is discussed in detail under Pasteur’s contributions to microbiology

CONTIBUTIONS OF IMPORTANT SCIENTISTS TO MICROBIOLOGY

Robert Hooke

Robert Hooke was born on the Isle of Wight, England on July 28, 1635 Active in the 17th century, Robert Hooke is one of the most important scientists of his generation and contributed in an amazing variety of scientific fields. Among other things, he was the first to discover the cell; Robert Hooke used an improved compound microscope he had built to study the bark of a cork tree. In doing so he discovered and named the cell – the building block of life. However he didn’t know its true biological function. Hooke coined the term cell and published the discovery in his famous 1665 book Micrographia. Hooke’s Micrographia, the first scientific best seller and one of the most important books ever written, demonstrated the tremendous power of the microscope and inspired people to use it for scientific exploration. Robert Hooke was the first person to use a microscope to study fossils and he published his findings in Micrographia. He concluded that fossils had once been living creatures whose cells had become mineralized. He also concluded that some species that had once existed must have become extinct. This was a controversial suggestion as most people at the time found the concept of extinction theologically unacceptable. Thus Hooke was one of the first proponents of a theory of evolution.

A drawing of Robert Hookes microscope

https://learnodo-newtonic.com/robert-hooke-contribution

Antonie van Leeuwenhoek (1632 — 1723)

Antonie van Leeuwenhoek was born on October 24, 1632, in the small city of Delft in the Dutch Republic . A moderately educated owner of a textile business, he learned how to make his own unique microscopes which offered unparalleled magnification. Using these microscopes he made a number of crucially important scientific discoveries, including single-celled animals and plants, bacteria, and spermatozoa. His microscopy methods were so finely tuned that after he discovered bacteria, this type of organism would not be observed again by any other scientist for over 100 years.

People in the textiles trade had, for hundreds of years, used glass pearls — small spheres of glass — as lenses to examine cloth in fine detail. Leeuwenhoek used glass pearls frequently in his day-to-day business to examine the density of threads and the quality of cloth.

In 1665 the great English scientist Robert Hooke released Micrographia, showcasing drawings he had made of the natural world seen through the lens of his microscope. Micrographia contains a description of how a powerful microscope could be made using a single spherical lens — similar to the glass pearls Leeuwenhoek was already familiar with. It is believed that Leeuwenhooke was inspired by Robert Hooke’s book and began making lenses for observing microscopic life.

Microscopes made from Leeuwenhoek’s tiny spherical lenses — the smallest lenses measured just 1 mm across — were easily capable of magnifying objects by a factor of about 200 — 300, while Hooke’s compound microscope magnified only by a factor of about 40 — 50.

Remarkably, Leeuwenhoek could use his lenses to resolve details as small as 1.35 μm. (This meant that, for example, he could easily see red blood cells, which are typically 6 — 8 μm in diameter.) Leeuwenhoek was a tradesman who had no formal training in science and had never been to college.Nevertheless, the quality of his observations was so high and his discoveries so compelling that his research became well-known through letters he sent to the Royal Society in London. These were translated into English and published in the Society’s journal, Philosophical Transactions.

In 1674, aged 41, Leeuwenhoek made the first of his great discoveries: single-celled life forms. He observed and recorded Red Blood cells, bacteria in water & soil, sperm cells & egg cells, lymphatic capillaries & life cycle of maggots. Antonie van Leeuwenhoek died aged 90 on August 26, 1723.

LOUIS PASTEUR ( 1822-1895)

Louis Pasteur (December 27, 1822 — September 28, 1895) was a French chemist and microbiologist noted for many of his significant findings in microbiology.

Some of the most important contributions made by Pasteur may be enlisted as follows,

Established that microorganism were responsible for fermentation as well as spoilage of wine
Developed the technique of Pasteurization for preservation of milk and other liquids.
Developed vaccines for fowl cholera, rabies
Coined the term ‘vaccine’ to commemorate Edward Jenner’s first developed vaccine against small pox
Disproved spontaneous generation theory by his famous ‘swan neck flask experiment’.
Contributed towards the understanding of causative agents and management of silkworm diseases

Pasteur started his career as a chemist in wine industry where he studied optical isomerism in tartaric acid crystals. It was here that he first established that microorganisms were responsible for the process of fermentation as well as spoilage of wine. He observed that wine could get ‘sick’ & become sour. He found that undesirable microbial growth was the reason behind ‘sick’ wine. Pasteur then developed the technique of pasteurization (patented in 1865) to fight the “diseases” of wine. He realized that these were caused by unwanted microorganisms that could be destroyed by heating wine to a temperature between 60° and 100°C. The process was later extended to all sorts of other spoilable substances, such as milk.

The observation that wine could get sick from bacteria led him to hypothesize that people’s sickness could also be attributed to bacteria. This and the conclusions made by many other contemporary scientists, led Pasteur to propose the ‘Germ Theory Of Disease’. It states that infectious diseases were caused by microorganisms.

He proved that microbes were attacking healthy silkworm eggs, causing an unknown disease, and that the disease would be eliminated if the microbes were eliminated. He eventually developed a method to prevent their contamination and it was soon used by silk producers throughout the world.

FOWL CHOLERA: In 1870’s Pasteur engaged in the studies of disease causing micro organisms . In his experiments on fowl cholera he inoculated the birds with the laboratory cultures and found that no birds could survive cholera.

Months into the experiments, Pasteur let cultures of fowl cholera stand idle while he went on vacation. When he returned and the same procedure was attempted, the chickens did not become diseased as before. Pasteur could easily have deduced that the culture was dead and could not be revived, but instead he was inspired to inoculate the experimental chickens with a virulent culture. Amazingly, the chickens survived and did not become diseased; they were protected by a microbe attenuated over time. Realizing he had discovered a technique that could be extended to other diseases, Pasteur returned to his study of anthrax. Pasteur produced vaccines from weakened anthrax bacilli that could indeed protect sheep and other animals.

RABIES VACCINE : Rabies had presented a new obstacle for Pasteur in the development of a successful vaccine. Unlike chicken cholera and anthrax, both caused by bacterium, the microorganism causing the disease could not be specifically identified, meaning Pasteur would not be able to develop the vaccine in vitro (in the laboratory).

Pasteur did not know this at the time, but the reason he could not find the microorganism is because rabies is a viral disease. Viruses are small infectious agents that replicate quickly and have a high mutation rate. These rapid mutations can be used to the benefit of researchers in the development of an attenuated vaccine. By serial passage of a virus through a different species, the virus becomes more adapted to that species, and less adapted to its original host, deceasing virulence with respect to the original host (e.g. it is “attenuated”). By passing the virus through rabbits, Pasteur made the virus less dangerous to human hosts, while still giving the body enough information to recognize the antigen and develop immunity to the “wild” version of the disease.

After successfully protecting dogs from the disease, Pasteur agreed to treat his first human patient, a nine-year-old boy who had been so severely attacked by feral dogs there was little doubt he would die if nothing was done. Pasteur injected the boy with a daily series of progressively more virulent doses of the vaccine from the rabies-infected rabbits. The boy never developed symptoms and Pasteur became an international hero.

Until Louis Pasteur developed the rabies vaccine, “vaccines” had referred only to the cowpox inoculation for smallpox. His procedure was originally called “Pasteur’s treatment”, but Pasteur decided to honor the 18thcentury virology pioneer Edward Jenner, who publicized the cure for smallpox, and give these artificially weakened diseases the generic name of “vaccines”. Thus, we largely have Pasteur to thank for today’s definition of a vaccine as a “suspension of live (usually attenuated) or inactivated microorganisms (e.g., bacteria or viruses) or fractions thereof administered to induce immunity and prevent infectious disease or its sequelae.”

Swan Neck flask experiment: In his famous experiment, Louis Pasteur used a special flask whose neck was shaped like an S or the neck of a swan, hence the name “Swan Neck Flask.” He put a nutrient rich broth in the flask, which he called the “infusion.” He then boiled the infusion killing any microorganisms which were already present. Then he allowed the infusion to sit. Because of the shape of the flask, the infusion was exposed to air. However, dust particles and other things in the air never made it into the infusion. Because they were trapped in the curve of the Swan Neck Flask. No matter how long he allowed the flask to sit, microorganisms never appeared in the infusion. However, if he tipped the flask and allowed the things trapped in the neck to get into the infusion then microorgranisms began to appear in the infusion and multiply rapidly. This demonstrates that microorganisms do not appear as a result of Spontaneous Generation. Instead, they are introduced into food through dust particles and other things that happen to land on the food.

Image result for pasteur swan neck flask experiment

ROBERT KOCH

Robert Koch was a German physician who is widely credited as one of the founders of bacteriology and microbiology. He investigated the anthrax disease cycle in 1876, and studied the bacteria that causes tuberculosis in 1882, and cholera in 1883. He also formulated Koch’s postulates. Koch won the 1905 Nobel Prize in Physiology or Medicine.

Major contributions of Koch may be listed out as following:

Identification of causative organism of anthrax as the bacterium Bacillus anthraces
Developed a technique of isolating bacteria as pure culture on solid media
An outstanding contribution to microbiology was proposal of Koch’s postulates
Development of tuberculin and studies on Tuberculosis
Contributions to the study of cholera

Koch also found a way to grow bacteria in pure cultures—cultures that contained only one kind of organism. He tried streaking bacterial suspensions on potato slices and then on solidified gelatin. But gelatin melts at incubator (body) temperature; even at room temperature. Finally, Angelina Hesse the American wife of one of Koch’s colleagues, suggested that Koch add agar (a thickener used in cooking) to his bacteriological media. This created a firm surface over which microorganisms could be spread very thinly—so thinly that some individual organisms were separated from all others. Each individual organism then multiplied to make a colony of thousands of descendants. Koch’s technique of preparing pure cultures is still used today.

Koch’s outstanding achievement was the formulation of four postulates to associate a particular organism with a specific disease. Koch’s Postulates are as follows,

The specific causative agent must be found in every case of the disease.
The disease organism must be isolated in pure culture.
Inoculation of a sample of the culture into a healthy, susceptible animal must produce the same disease.
The disease organism must be recovered from the inoculated animal.

Implied in Koch’s postulates is his one organism–one disease concept. The postulates assume that an infectious disease is caused by a single organism, and they are directed toward establishing that fact. This concept also was an important advance in the development of the germ theory of disease.

He also developed tuberculin, which he hoped would be a vaccine against tuberculosis. Because he underestimated the difficulty of killing the organism that causes tuberculosis, the use of tuberculin resulted in several deaths from that disease. Although tuberculin was unacceptable as a vaccine, its development laid the groundwork for a skin test to diagnose tuberculosis. After the vaccine disaster, Koch left Germany. He made several visits to Africa, at least two visits to Asia, and one to the United States. In the remaining 15 years of his life, his accomplishments were many and varied. He conducted research on malaria, typhoid fever, sleeping sickness, and several other diseases. His studies of tuberculosis won him the Nobel Prize for Physiology and Medicine in 1905, and his work in Africa and Asia won him great respect on those continents.

EDWARD JENNER

Also known as the “Father of Immunology”, Edward Anthony Jenner was an English scientist and is famous for his discovery of the smallpox vaccine. This was the first successful vaccine ever to be developed and remains the only effective preventive treatment for the fatal smallpox disease. His discovery was an enormous medical breakthrough and has saved countless lives. In 1980, the World Health Organization declared smallpox an eliminated disease.

Smallpox was unknown in Europe until the Crusaders carried it back from the Near East in the twelfth century. By the seventeenth century, it was widespread. In 1717 Lady Mary Ashley Montagu, wife of the British ambassador to Turkey, introduced a kind of immunization to England. A thread was soaked in fluid from a smallpox vesicle (blister) and drawn through a small incision in the arm. This technique, called variolation, was used at first by only a few prominent people, but eventually it became widespread. In the late eighteenth century, Edward Jenner realized that milkmaids who got cowpox did not get smallpox, and he inoculated his own son with fluid from a cowpox blister. He later similarly inoculated an 8-year-old and subsequently inoculated the same child with smallpox. The child remained healthy. The word vaccinia (vacca, the Latin name for “cow”) gave rise both to the name of the virus that causes cowpox and to the word vaccine.

Sergei Winogradsky

Sergei Nikolaievich Winogradsky was born on September 1, 1856 in Kiev, which was then in the Russian Empire . Sergei Winogradsky founded microbial ecology and he was a founding father of microbiology. The contributions of Winogradsky may be enlisted as follows,

Discovered chemosynthesis – an entirely new mode of life, in which the energy to build organic molecules comes from chemical reactions rather than from sunlight in the more familiar photosynthesis.
Invented the Winogradsky column.
Discovered and isolated nitrogen fixing bacteria in soil that make nitrates available to green plants.
Founded microbial ecology, where the interactions of microbes in cycles with their natural environments are studied holistically.

Winogradsky’s study on Beggiatoa, a puzzling bacterial organism led to ground breaking discovery of chemosynthesisin organisms. Beggiatoa was puzzling because it did not grow in the nutrient mixtures normally used in laboratories, yet in sulfurous spring waters it could develop into huge colonies, forming large mats.By 1887, the 31 year old Winogradsky had discovered that Beggiatoa did not obtain energy by any previously known method. Beggiatoa’s energy came from a chemical reaction between hydrogen sulfide, present in sulfurous waters, and oxygen. The reaction also produced sulfur and water.Beggiatoa stored the sulfur it produced within its own cells and, if needed, could react this sulfur with oxygen to produce even more energy.It was the first time anyone had discovered a living organism that survived using inorganic compounds/minerals as a source of energy. This type of organism is now called a lithotroph.

Winogradsky column

During his time in Strasbourg, Winogradsky invented the Winogradsky Column, which is still used today to learn which communities of bacteria are present in a sample. It consists of a glass cylinder half-filled with mud from a river or lake, topped up with water. A number of ingredients, such as sodium sulfate, calcium carbonate, and cellulose (newspaper) are added to provide sources of nutrition for bacteria.Within the column, scientists can control the amount of nutrients and light available to bacteria.

Construction of a winogradsky column:

The columns are easy to set up with a glass or perspex tube, about 30 cm tall and 5 cm diameter. Mud from the bottom of a lake or river is supplemented with cellulose (e.g. newspaper), sodium sulphate and calcium carbonate, then added to the lower one-third of the tube. The rest of the tube is filled with water from the lake or river, and the tube is capped and placed near a window with supplementary strip lights.

All the organisms are present initially in low numbers, but when the tubes are incubated for 2 to 3 months the different types of microorganism proliferate and occupy distinct zones where the environmental conditions favour their specific activities. The large amount of cellulose added initially promotes rapid microbial growth which soon depletes the oxygen in the sediment and in the water column. Different layers of the water column generates different zones of organisms that occupy that particular niche in the column.

Winogradsky enjoyed an unusually long and active life. He was 93 years old when he completed a 900 page book: Soil Microbiology: Problems and Methods, Fifty Years of Investigations. Sergei Winogradsky died in his sleep at the age of 96 in Brie-Comte-Robert, France, on February 25, 1953.