History of Microbiology Lecture #2 Bio3124 University of Ottawa

Old times… Humans knew how to deal with germs before even knowing about their existence Storing food in cooler temperatures Salting, drying, smoking Use of spices Cremation of dead concealing and burying dead beyond a nearest passing river or in a distant area Explanation: traditional, mystical and superstitious

Discovery of Microorganisms Antony van Leeuwenhoek (1632-1723), the father of microbiology a hobbyist microscopist Dutchman first to observe and describe microorganisms accurately

Leeuwenhoek’s microscope Composed of one lens Light shines objects at a 45˚ angle Worked like a dark field microscope Magnification: 50-300 fold bright microorganisms could be seen in a dark bkg

Leeuwenhoek’s Observations Spirillum as seen by Leeuwenhoek’s microscope Tripartite structure of a human sperm PMN RBC Original blood smear image by Leeuwenhoek Note the polymorphonuclear cell and RBC Reported to the Royal Society of London (1673) Accurate shape, detailed movement Subjects were most possibly bacteria and protozoa and called them “animalcules“ also reported spermatozoa, blood cells

Don’t forget to check out the paper by Brian J Don’t forget to check out the paper by Brian J. Ford about Leeuwenhoek posted under literature button in your website

17th century Robert Hooke and Antony Van Leeuwenhoek Using the microscope to observe the fruit organs of mushrooms and mouth bacteria.

Where do microorganism originate from? Few days of being exposed to air. Spontaneous generation??

The Conflict over Spontaneous Generation living organisms can develop from nonliving or decomposing matter publicly a common sense vision with societal and cultural roots (almost 2000 years ago) Examples: flies from rotten meat and animal carcasses Mice from fungus infested grain barns Scientific methodology was not established What was the proof?

Jan Baptista Van Helmont (1577-1644) A Flemish noble man, alchemist and physician discovered carbon dioxide, introduced the term gas in its present scientific sense believed in spontaneous generation

Don’t forget to check out the paper by Louis Rosenfeld about Van Helmont posted under literature button in your website

Jan Baptista Van Helmont The Origin of life (17th Centuray) 1579-1644 Adult mice

Spontaneous Generation

But could spontaneous generation be true for microorganisms?

Francesco Redi (1626-1697) First blow to spontaneous generation Laying eggs is required for maggots and flies to come to existence Supporters of SG: Life is necessary in order to bring about life in certain cases!

John Needham (1713-1781) Conclusion: Organic matter had vital force that confers properties of life to non-living matter BUT: he had left the flasks unsealed after bioling

Lazzaro Spallanzani (1729-1799) Air carries germs to broth and boiling kills the existing ones Suppoters of SG: Compounds essential with the generation of life were destroyed by heating!! sealing prevents air that is necessary for spontaneous generation

Louis Pasteur (1822-1895) Observations: John Tyndall: Organisms retained on cotton filters resemble microorganisms in contaminated foods Microorganisms are found in the air Microorganism in air settle on the surfaces

Pateur’s Swan neck flasks could supply air but could also trap ambient germs from entering broth

Pasteur Refutes the Spontaneous Generation

Pasteur Refutes the Spontaneous Generation ambient germs are necessary for promoting growth principle of sterility and aseptic work is important

The Role of Microorganisms in Disease For over 17 centuries the Galenism (Greek physician 129-199 B.C.) was predominant view for diseases according to which, Diseases are due to imbalances in 4 humors: blood, phlegm, yellow bile and black bile)

Recognition of the Relationship between Microorganisms and Disease Agostino Bassi (1773-1856) showed that a disease of silkworms was caused by a fungus

More evidence… M. J. Berkeley (ca. 1845) Heinrich de Bary (1853) demonstrated that the great Potato Blight of Ireland was caused by a water mold Heinrich de Bary (1853) showed that fungi caused cereal crop diseases Louis Pasteur showed that the pébrine disease of silkworms was caused by a protozoan Nosema bombycis

Other evidence… Joseph Lister (1827-1912) provided indirect evidence used diluted carbolic acid (phenol) to wash surgical devices and wash wounds developed the principles of aseptic surgery his patients had fewer postoperative infections Disease frequency dropped in his hospital

Final proof… Robert Koch (1843-1910) established the relationship between Bacillus anthracis and anthrax (Black Death plague) used criteria developed by his teacher Jacob Henle (1809-1895) these criteria now known as Koch’s postulates Read more about R. Koch. Check literature section

Microorganisms and Diseases Robert Koch Observations : The microorganism (Bacillus anthracis) is found only in the infected animals and not in healthy animals. Infected animals pass the disease to healthy ones The microorganism found outside of the animal could infect healthy animals

Koch’s experiment Anthrax: Injected mice with B.anthracis, mice got sick Took spleen from infected mice and grew the anthrax bacillus in a rich medium Injected healthy mice with spores or the cells Mice developed anthrax Independently, similar experiments done by Pasteur

Koch’s postulates The microorganism must be present in every case of the disease but absent from healthy individuals The suspected microorganism must be isolated from infected individuals and grown in a pure culture The same disease must result when the isolated microorganism is inoculated into a healthy host The same microorganism must be isolated again from the diseased host

Questions: Can Koch’s Postulates be applied to all microorganisms that cause disease? How can you apply Koch’s postulates to viruses and viral diseases? How can one apply Koch’s postulates to non-cultivable microorganisms?

The Development of Techniques for Studying Microbial Pathogens Koch’s work led to discovery or development of: agar Petri dish nutrient broth and nutrient agar methods for isolating microorganisms in pure culture

The Pure Cultures and Isolation of Single Colonies Koch : Observation: Slices of potato when exposed to air, will generate large number of bacteria (colonies) of different sizes, colours and forms Colonies

Problem: Not all bacteria are able to grow on potatoes! Solution : Koch used gelatin as a means for solidification of rich culture media. He formulated various culture media

Disadvantages of Gelatin Gelatin liquefaction: It is digested by various microorganisms eg. gelatinase positive enterococci and streptomyces Low melting temperature: It is in the liquid state above the temperature of 28oC.

Agar Gracilaria (red seaweeds) Polysaccharide derived from the cell walls of a red algae (Rhodophyta) A polysaccharide polymer that contains 3,6-anhydrogalactose, 2-O-methyl-α-l-galactopyranose and 6-O-methyl agarobiose Solid at >37oC. Melts at 100oC. Not digested by most bacteria

Conclusion: A unique cells of different bacteria can grow on agar medium to form a single colony. A single colony will generate identical colonies (pure colony) Cultures grown from pure colonies are called pure cultures

The golden age of microbiology (1857-1914) Many disease producing organisms discovered Microbial metabolism studies undertaken Microbiological techniques refined A better understanding of the role of immunity and ways to control and prevent infection by microbes

Other developments… Charles Chamberland (1851-1908) developed porcelain bacterial filters used by Ivanoski and Beijerinck to study tobacco mosaic disease determined that extracts from diseased plants had infectious agents present which were smaller than bacteria and passed through the filters Infectious agents were eventually shown to be viruses

Immunological Studies Edward Jenner (1749-1823) used a vaccination procedure to protect individuals from smallpox NOTE: this preceded the work establishing the role of microorganisms in disease

Other developments… Pasteur and Roux Pasteur and his coworkers discovered that incubation of cultures for long intervals between transfers caused pathogens to lose their ability to cause disease (attenuation) Pasteur and his coworkers developed vaccines for chicken cholera, anthrax, and rabies

More developments… Emil von Behring (1854-1917) and Shibasaburo Kitasato (1852-1931) developed antitoxins for diphtheria and tetanus evidence for humoral immunity Elie Metchnikoff (1845-1916) discovered bacteria-engulfing, phagocytic cells in the blood (phagocytosis) evidence for cellular immunity (CMI)

The Development of Industrial Microbiology and Microbial Ecology Louis Pasteur demonstrated that alcohol fermentations and other fermentations were the result of microbial activity developed the process of pasteurization to preserve wine during storage

Additional Developments… Sergei Winogradsky (1856-1953) and Martinus Beijerinck (1851-1931) studied soil microorganisms and discovered numerous interesting metabolic processes (e.g., nitrogen fixation) pioneered the use of enrichment cultures and selective media

Summary of Keynotes Advances in Microbiology needed Technological advances, new observation Resolution against SG Establishment of “Germ theory” Furthering techniques: Culturing and isolation of pure colonies Application