Unit 1.3  A Brief History of Microbiology

First Observations:

Several ancient civilizations appear to have had some understanding that disease could be transmitted by things they could not see. The ancient Greeks attributed disease to bad air, mal’aria, which they called “miasmatic odors.” They developed hygiene practices that built on this idea. The Romans also believed in the miasma hypothesis and created a complex sanitation infrastructure to deal with sewage. In Rome, they built aqueducts, which brought fresh water into the city, and a giant sewer, the Cloaca Maxima, which carried waste away and into the river Tiber. Some researchers believe that this infrastructure helped protect the Romans from epidemics of waterborne illnesses.

The Greek physician Hippocrates (460–370 BC) is considered the “father of Western medicine” (Figure 1.9). Unlike many of his ancestors and contemporaries, he dismissed the idea that disease was caused by supernatural forces. Instead, he posited that diseases had natural causes from within patients or their environments. Hippocrates and his heirs are believed to have written the Hippocratic Corpus, a collection of texts that make up some of the oldest surviving medical books. Hippocrates is also often credited as the author of the Hippocratic Oath, taken by new physicians to pledge their dedication to diagnosing and treating patients without causing harm.

While Hippocrates is considered the father of Western medicine, the Greek philosopher and historian Thucydides (460–395 BC) is considered the father of scientific history because he advocated for evidence-based analysis of cause-and-effect reasoning (Figure 1.10). Among his most important contributions are his observations regarding the Athenian plague that killed one-third of the population of Athens between 430 and 410 BC. Having survived the epidemic himself, Thucydides made the important observation that survivors did not get re-infected with the disease, even when taking care of actively sick people. This observation shows an early understanding of the concept of immunity.

Marcus Terentius Varro (116–27 BC) was a prolific Roman writer who was one of the first people to propose the concept that things we cannot see (what we now call microorganisms) can cause disease (Figure 1.10). In Res Rusticae (On Farming), published in 36 BC, he said that “precautions must also be taken in neighborhood swamps. . . because certain minute creatures [animalia minuta] grow there which cannot be seen by the eye, which float in the air and enter the body through the mouth and nose and there cause serious diseases.”

While the ancients may have suspected the existence of invisible “minute creatures,” it wasn’t until the invention of the microscope that their existence was definitively confirmed. Anton van Leeuwenhoek, sometimes hailed as “the Father of Microbiology,” is typically credited as the first person to have created microscopes powerful enough to view microbes (Figure 1.11).  In 1674, he described his observations of single-celled organisms, whose existence was previously unknown, in a series of letters to the Royal Society of London. His report was initially met with skepticism, but his claims were soon verified and he became something of a celebrity in the scientific community.

While van Leeuwenhoek is credited with the discovery of microorganisms, others before him had contributed to the nascent field of microbiology. Van Leeuwenhoek’s contemporary, the Englishman Robert Hooke (1635–1703), also made important contributions to microscopy, publishing in his book Micrographia (1665) many observations using compound microscopes. Viewing a thin sample of cork through his microscope, he was the first to observe the structures that we now know as cells (Figure 1.12). Hooke described these structures as resembling “Honey-comb,” and as “small Boxes or Bladders of Air,” noting that each “Cavern, Bubble, or Cell” is distinct from the others (in Latin, “cell” literally means “small room” and is still used in modern English, e.g. jail cell). They likely appeared to Hooke to be filled with air because the cork cells were dead, with only the rigid cell walls providing the structure.

The Debate Over Spontaneous Generation:

With the discovery of this hidden, microscopic world, all around them, scientist turned their focus to where these “animalcules” were coming from. 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 17th 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.

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 set of containers, so of which were open to the air and others that were tightly sealed (Figure 1.13). His hypothesis was supported when maggots developed on the meat in the uncovered jars, but no maggots appeared on the meat in 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. Proponents of spontaneous generation were not convinced. They argued that fresh air was required for spontaneous generation. So Redi redid is experiments, but this time instead of sealing the jars completely, he covered them with a fine net of gauze. This set-up allowed for air to enter the jar, but still prevented the flies from entering. No maggots appeared on the meat in the gauze-covered jars even in the presence of fresh air.

Redi’s results provided strong evidence that organism could not just arise form nonliving matter. Despite this evidence, many scientists believed that while spontaneous generation may not hold true for larger organism, microorganism were simple enough to be able to generate spontaneously.

This argument was strengthened in 1745, when 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.  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.

Lazzaro Spallanzani (1729–1799) did not agree with Needham’s conclusions, and suggested that microorganisms from the air had contaminated Needham’s broth after it had been boiled. To test his hypothesis, he performed hundreds of carefully executed experiments using heated broth.  As in Needham’s experiment, he used infused broth, but boiling the broth before sealing the flasks, he sealed some flasks before boiling them. 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 “vital force” that was destroyed during Spallanzani’s extended boiling. Sealing of the flasks then prevented this force from entering and causing spontaneous generation. Spallanzani’s experiments also drew the same criticism as had Redi’s: by sealing the flasks, he was preventing air from entering, which would prevent spontaneous generation from occurring (Figure 1.14).

The debate over spontaneous generation continued well into the 19th century, with scientists serving as proponents of both sides. To settle the debate, the Paris Academy of Sciences offered a prize for resolution of the problem. Louis Pasteur, a prominent French chemist who had been studying microbial fermentation and the causes of wine spoilage, accepted the challenge. In 1858, Pasteur filtered air through a gun-cotton filter and, upon microscopic examination of the cotton, found it full of microorganisms, suggesting that the exposure of a broth to air was not introducing a “life force” to the broth but rather airborne microorganisms.

Later, Pasteur made a series of flasks with long, twisted necks (“swan-neck” flasks), in which he boiled broth to sterilize it (Figure 1.15). His design allowed air inside the flasks to be exchanged with air from the outside, but prevented the introduction of any airborne microorganisms, which would get caught in the twists and bends of the flasks’ necks. If a life force besides the airborne microorganisms were responsible for microbial growth within the sterilized flasks, it would have access to the broth, whereas the microorganisms would not. He correctly predicted that sterilized broth in his swan-neck flasks would remain sterile as long as the swan necks remained intact. However, should the necks be broken, microorganisms would be introduced, contaminating the flasks and allowing microbial growth within the broth.

Pasteur’s set of experiments irrefutably disproved the theory of spontaneous generation and earned him the prestigious Alhumbert Prize from the Paris Academy of Sciences in 1862. In a subsequent lecture in 1864, Pasteur articulated “Omne vivum ex vivo” (“Life only comes from life”). In this lecture, Pasteur recounted his famous swan-neck flask experiment, stating that “…life is a germ and a germ is life. Never will the doctrine of spontaneous generation recover from the mortal blow of this simple experiment.”  To Pasteur’s credit, it never has.

Modern Cell Theory:

While some scientists were arguing over the theory of spontaneous generation, other scientists were making discoveries leading to a better understanding of what we now call the cell theory. Modern cell theory has two basic tenets:

•         All cells only come from other cells.

•         Cells are the fundamental units of organisms.

Today, these tenets are fundamental to our understanding of life on earth. However, modern cell theory grew out of the collective work of many scientists.

Recall that the English scientist Robert Hooke first used the term “cells” in 1665 to describe the small chambers within cork that he observed under a microscope of his own design (See Figure 1.11). At the time, Hooke was not aware that the cork cells were long dead and, therefore, lacked the internal structures found within living cells.

Despite Hooke’s early description of cells, their significance as the fundamental unit of life was not yet recognized. Nearly 200 years later, in 1838, Matthias Schleiden (1804–1881), a German botanist who made extensive microscopic observations of plant tissues, described them as being composed of cells. Visualizing plant cells was relatively easy because plant cells are clearly separated by their thick cell walls. Schleiden believed that cells formed through crystallization, rather than cell division.

Theodor Schwann (1810–1882), a noted German physiologist, made similar microscopic observations of animal tissue. In 1839, after a conversation with Schleiden, Schwann realized that similarities existed between plant and animal tissues. This laid the foundation for the first tenet of cell theory, that cells are the fundamental components of plants and animals.

In the 1850s, two Polish scientists living in Germany pushed this idea further. In 1852, Robert Remak (1815–1865), a prominent neurologist and embryologist, published convincing evidence that cells are derived from other cells as a result of cell division. However, this idea was questioned by many in the scientific community. Three years later, Rudolf Virchow (1821–1902), a well-respected pathologist, published an editorial essay entitled “Cellular Pathology,” which popularized the concept of biogenesis, using the Latin phrase omnis cellula a cellula (“all cells arise from cells”), which is essentially the second tenet of modern cell theory. Given the similarity of Virchow’s work to Remak’s, there is some controversy as to which scientist should receive credit for articulating biogenesis (Figure 1.16).

The Germ Theory of Disease:

Prior to the discovery of microbes during the 17th century, other theories circulated about the origins of disease. For example, the ancient Greeks proposed the miasma theory, which held that disease originated from particles emanating from decomposing matter, such as that in sewage or cesspits. Such particles infected humans in close proximity to the rotting material. Diseases including the Black Death, which ravaged Europe’s population during the Middle Ages, were thought to have originated in this way.

In 1546, Italian physician Girolamo Fracastoro proposed, in his essay De Contagione et Contagiosis Morbis, that seed-like spores may be transferred between individuals through direct contact, exposure to contaminated clothing, or through the air. We now recognize Fracastoro as an early proponent of the germ theory of disease, which states that diseases may result from microbial infection. However, in the 16th century, Fracastoro’s ideas were not widely accepted and would be largely forgotten until the 19th century.

In 1847, Hungarian obstetrician Ignaz Semmelweis (Figure 1.17) observed that mothers who gave birth in hospital wards staffed by physicians and medical students were more likely to suffer and die from puerperal fever after childbirth (10%–20% mortality rate) than were mothers in wards staffed by midwives (1% mortality rate). Semmelweis observed medical students performing autopsies and then subsequently carrying out vaginal examinations on living patients without washing their hands in between. He suspected that the students carried disease from the autopsies to the patients they examined. His suspicions were supported by the untimely death of a friend, a physician who contracted a fatal wound infection after a postmortem examination of a woman who had died of a puerperal infection. The dead physician’s wound had been caused by a scalpel used during the examination, and his subsequent illness and death closely paralleled that of the dead patient.

Although Semmelweis did not know the true cause of puerperal fever, he proposed that physicians were somehow transferring the causative agent to their patients. He suggested that the number of puerperal fever cases could be reduced if physicians and medical students simply washed their hands with chlorinated lime water before and after examining every patient. When this practice was implemented, the maternal mortality rate in mothers cared for by physicians dropped to the same 1% mortality rate observed among mothers cared for by midwives. This demonstrated that handwashing was a very effective method for preventing disease transmission. Despite this great success, many discounted Semmelweis’s work at the time, and physicians were slow to adopt the simple procedure of handwashing to prevent infections in their patients because it contradicted established norms for that time period.

 While studying the causes of beer and wine spoilage in 1856, Pasteur discovered properties of fermentation by microorganisms. He had demonstrated with his swan-neck flask experiments (Figure 1.15) that airborne microbes, not spontaneous generation, were the cause of food spoilage, and he suggested that if microbes were responsible for food spoilage and fermentation, they could also be responsible for causing infection. This was the foundation for the germ theory of disease.