Microbiology

Microbiology

by Daniel V Lim

Introduction, Development of Microbiology

The study of microorganisms would not be possible, and microbiology would not have developed as a discipline, without the aid of the light microscope, an instrument that makes it possible to magnify minute specimens up to two thousandfold. Some of the earliest microscopes were made by Zacharias Janssen (1580–c. 1638), a Dutch optician who, as a young boy, combined two lenses to construct crude microscopes. These early instruments subsequently were improved and by the late 1600s microscopes were capable of magnifying objects by 50–300 diameters. Anton van Leeuwenhoek (1632–1723), a Dutch linen draper, made hundreds of these microscopes during his lifetime and observed small, living particles, which he called ‘animalcules’, through them. Leeuwenhoek is considered to be the ‘father of microbiology’ because of his work with these early microscopes.

Louis Pasteur (1822–1895), a French chemist, played a significant role in the development of microbiology with his experiments to disprove the theory of spontaneous generation. This theory, which became popular during the eighteenth century, proposed that spoilage organisms arose spontaneously in putrefied food. Pasteur captured airborne microorganisms on guncotton filters to show that these airborne microbes were responsible for food spoilage and did not arise spontaneously.

In 1876 a German country physician named Robert Koch (1843–1910) discovered that the lethal, contagious cattle disease anthrax could be transmitted from animal to animal by injecting blood from an infected cow into a healthy cow. His experiments were formulated into a set of criteria, known as Koch's postulates, that have become the cornerstone for associating specific microorganisms with specific infectious diseases. Koch's postulates initiated an era known as the Golden Age of Microbiology (1876–1906), during which the causes of many microbial diseases were discovered.

Through the accomplishments of these and many other individuals, microbiology developed into a distinct discipline. Today, microbiology continues to grow as a discipline and has entered into a new golden age closely affiliated with molecular biology and biotechnology.

Aspects of Prokaryology, Bacteriology, Mycology, Protozoology, Phycology and Virology

The five agents (bacteria, fungi, algae, protozoa and viruses) associated with microbiology each have their own unique characteristics. Bacteria, which do not have a well-defined membrane-enclosed nucleus, are considered prokaryotes (Latin pro, before, Greek karyon, nucleus), whereas fungi, algae and protozoa have a well-defined membrane-enclosed nucleus and, therefore, are eukaryotes. Viruses, which are not cellular but are intracellular parasites containing either deoxyribonucleic acid (DNA) or ribonucleic acid (RNA), are not considered either prokaryotes or eukaryotes.

Classification of microorganisms

The separation between prokaryotes and eukaryotes has become less distinct in recent years with the emergence of molecular techniques and the discovery in the 1980s that some prokaryotes have features that more closely resemble the eukaryotes. In 1981, Carl Woese proposed that all living organisms be separated into three domains: Archaea (Greek Archaios, ancient, as in the Archaean era, a geological period approximately 3.9–2.6 billion years ago), Bacteria and Eukarya (Figure 1). This separation is based on the unique ribosomal RNA sequences for members of each domain, as well as other characteristics such as cell wall and plasma membrane composition, complexity of RNA polymerases and mechanism of protein synthesis. Ribosomal RNA sequences have especially been important in determining the evolutionary, or phylogenetic, relationships among organisms in the three domains and indicate that the Archaea are the most primitive organisms.



The Archaea are a diverse group of prokaryotes that inhabit extreme environments. These organisms include methanogens (organisms found in anaerobic swamps, marshes and animal intestinal tracts that produce methane as a product of metabolism), sulfate reducers (organisms occurring in marine hydrothermal vents and that use sulfate as an electron acceptor), extreme halophiles (organisms found in high-salt environments such as the Great Salt Lake and the Dead Sea), extremely thermophilic sulfur metabolizers (organisms that exist in high-temperature environments and are capable of metabolizing sulfur), and cell wall-less thermophiles (organisms without walls that live in high-temperature environments).

The remaining prokaryotes are classified under the domain Bacteria. The domain Bacteria is an extensive group of microorganisms, consisting of hundred of thousands of species that are ubiquitous and found in various environments, including soil, water, plants and animals. Both the Archaea and the Bacteria typically are grouped into taxonomic families, genera and species, primarily on the basis of structural and morphological characteristics, such as cell shape, size and appendages, and secondarily on the basis of biochemical and physiological traits, such as growth factor requirements, range of carbohydrates used as carbon and energy sources, and end products of metabolism. This classical approach to taxonomy is used to organize prokaryotes in a reference manual called Bergey's Manual of Systematic Bacteriology. This classification of prokaryotes is rapidly undergoing revision and modification as newer molecular techniques provide more accurate information on phylogenetic relationships of living organisms through ribosomal RNA and DNA sequence analyses. Molecular analyses are also important in characterizing the vast majority of prokaryotes that are nonculturable and can only be classified by RNA or DNA composition.
 

Prokaryotes

The typical prokaryote is 1–2 mum long and 0.2–0.5 mum in diameter, although the smallest prokaryotes (Mycoplasma) have diameters of 125–250 nm and the largest prokaryotes (Epulopiscium) have dimensions of 600 mum by 80 mum. Prokaryotes exist in three basic forms: rod (bacillus), spherical (coccus) and spiral (spirillum) (Figure 2).

 

Prokaryotic DNA is found in a single, circular chromosome, although some prokaryotes have small, extrachromosomal, circular pieces of DNA called plasmids, which carry genes for antibiotic resistance, toxins and resistance to heavy metals. The cytoplasm and nuclear region of prokaryotes is typically surrounded by a phospholipid bilayer plasma membrane and a cell wall consisting of the repeating carbohydrate derivatives N-acetylglucosamine and N-acetylmuramic acid complexed with amino acids in a rigid structure called peptidoglycan. See also Bacterial Chromosome, Bacterial Plasmids, Archaeal Plasmids, Bacterial Cell Wall, Bacterial Cytoplasmic Membrane, Peptidoglycan, and Bacterial Genomes

A common staining procedure used in microbiology, the Gram stain (named after Hans Christian Gram (1853–1938), who first developed this stain in 1884), separates most prokaryotes into two groups: Gram-positive prokaryotes, which have a thick peptidoglycan cell wall layer; and Gram-negative prokaryotes, which have a thin peptidoglycan cell wall layer and an additional external membrane called the outer membrane. Gram-positive prokaryotes, which include bacteria such as Staphylococcus and Streptococcus, stain deep violet with the Gram stain, whereas Gram-negative prokaryotes, which include bacteria such as Escherichia coli and Pseudomonas, stain pink or red.

Many prokaryotes have long, whiplike appendages called flagella which aid in motility. Some prokaryotes, particularly those associated with disease, have outer layers of material called capsules which protect them from phagocytosis. Certain prokaryotes are capable of producing, under unfavourable conditions, a thick, multilayered structure called an endospore, which is resistant to high temperatures, desiccation, many disinfectants and other adverse environmental conditions. The endospore is a dormant form of the bacterium in which there is little metabolism. When conditions become favourable again, the endospore germinates into a normal, vegetative cell. Bacillus and Clostridium are examples of prokaryotes that form endospores.


Prokaryotes generally divide by binary fission, a process in which one cell divides into two equal cells. The most rapid-growing prokaryotes can divide every 15–20 min under ideal growth conditions. Because prokaryotes double in number with every generation, growth occurs rapidly and the number of prokaryotes in a population of cells can increase exponentially from one cell to several billion cells in a few hours. This rapid growth of prokaryotes and their binary fission into equal daughter cells makes these microorganisms ideal tools for genetic and metabolic studies.

Fungi

Fungi are eukaryotic microorganisms that, like plants, have rigid cell walls and are nonmotile but, unlike plants, lack chlorophyll and are nonphotosynthetic. Fungi are as diverse as the prokaryotes and can be found in various environments ranging from arid deserts to tropical forests. Fungi live primarily off dead, decaying organic material and are therefore important in the decomposition and recycling of organic matter. See also Fungi and the History of Mycology, and Fungal Ecology

Fungi can exist as two fundamental morphological forms: moulds, which consist of filaments called hyphae that can branch into masses known as mycelia, and yeasts, which are unicellular, oval forms. Some fungi, such as Candida, are dimorphic and alternate between the mould and yeast forms, depending on environmental conditions. Fungi are capable of sexual and asexual reproduction. Sexual reproduction can involve an alternation of generations between a sporophyte that produces spores and a gametophyte that produces gametes. Examples of asexual reproduction are budding, in which a new daughter cell arises as a bud on the surface of the mother cell, and formation of spores at the tips of hyphae.

Fungi are economically important. Yeasts (for example, Saccharomyces cerevisiae), which ferment carbohydrates to alcohol and carbon dioxide, are extensively used in the making of bread, wine and beer. Mushrooms are a type of fungus and certain cheeses such as Roquefort (Penicillium roqueforti) and brie (Penicillium camemberti) are produced from fungi. Many antibiotics (for example, penicillin, cephalosporin and griseofulvin) and microbial enzymes (for example, amylases, pectinases and proteases) are produced by fungi. However, fungi can cause serious human diseases, such as histoplasmosis, coccidioidomycosis, cryptococcal meningoencephalitis, dermatophytosis (for example, athlete's foot and ringworm) and sick building syndrome; animal diseases, such as poultry haemorrhagic syndrome and ich (fish dermatitis); and plant diseases, such as apple scab, potato blight and corn brown spot. Fungi also are responsible for bread mould, mildew, rot and other destructive processes.

Protozoa

Protozoa are unicellular, nonphotosynthetic eukaryotic microorganisms that lack cell walls. Some protozoa are large enough to be seen with the unaided eye, although most are microscopic. Protozoa ingest materials by pinocytosis, a process in which liquids are surrounded by invagination of the plasma membrane and brought into the cell, and phagocytosis, a similar process in which larger particulate matter is engulfed. See also Protozoa, and Protozoan Ecology

Protozoa reproduce sexually or asexually. In asexual reproduction, the parent cell mitotically divides either into two equal daughter cells (binary fission) or into several daughter cells (multiple fission). Some protozoa asexually reproduce by budding or by spore formation. Sexual reproduction can occur by several methods, including conjugation, in which two cells join, exchange nuclei and produce progeny by budding or fission.

Protozoa are diverse in their habitats and distribution. Some cause human diseases, such as malaria (Plasmodium), toxoplasmosis (Toxoplasma), amoebic dysentery (Entamoeba), cryptosporidiosis (Cryptosporidium) and trichomoniasis (Trichomonas). Others (for example, Paramecium) are innocuous members of the biosphere, where they exist as important initial links in the food chain.

Algae

Algae are plant-like eukaryotic microorganisms that are distinguished from the fungi and protozoa by their chlorophylls and photosynthetic abilities. They contribute significantly to the aquatic food chain in marine and freshwater environments. Algae range in size from single-celled microscopic organisms to large, complex cell aggregates that attain lengths of more than 30 m. The colourful algal hues often visible in aquatic environments result from different chlorophylls and other photosynthetic pigments. Diatoms, elaborate shells that have two, thin overlapping halves, are among the simplest algae. Certain algae exist in association with microscopic animals as plankton. Other algae exist as multicellular aquatic structures known as seaweed.

Most algae reproduce asexually by mitotic division, whereas others form spores or reproduce by fragmentation of cells from larger aggregates. Some algae reproduce sexually by forming diploid zygotes from haploid gametes.

Over the years, algae have become increasingly important as food and food additives for humans. The red alga Porphyra, known as nori, is popular in sushi and as a toasted soup additive. Other red algae are also eaten as vegetables or as sweet jellies. Seaweed, which has high iodine and vitamin value, is used as a diet supplement. Algae can also produce economic loss when they overgrow, or bloom, because of excess nutrients in the water, which are often the result of human pollution or the release of sewage to bodies of water. Outbreaks of red tide are caused by algal blooms. The algae release toxic products that can kill fish and other marine life.

Viruses

Viruses are considered microorganisms, but occupy a unique place in the microbial world because they possess only a single type of nucleic acid (single-stranded or double-stranded DNA or RNA) contained within a simple protein coat. Furthermore, viruses are intracellular parasites that rely upon an infected host for metabolism and reproduction. Viruses that infect bacteria are called bacteriophages. Most viruses are too small (less than 200 nm in diameter) to be seen with the light microscope and can only be seen with an electron microscope. Because the viral particle is so small, its nucleic acid can code for only a few genes. Consequently, viruses are among the simplest forms of life. They often have repeating chemical building blocks in their structures and assume only a few symmetrical forms.

Viruses replicate within an infected host cell, using host protein-synthesizing machinery such as ribosomes to assist in replication. Following replication, progeny viruses are released as the host cell is lysed and dies. Successful viral infections typically lead to physical cell damage in prokaryotes and eukaryotes. Viruses are responsible for diseases such as acquired immune deficiency syndrome (AIDS), influenza, measles, poliomyelitis, rabies and encephalitis. A unique infectious agent that is simpler in structure than the virus is the prion. Prions (proteinaceous infectious particles), infectious agents that contain only protein and no nucleic acid, cause neurological diseases such as Creutzfeldt–Jakob disease and bovine spongiform encephalopathy (mad cow disease).

Applied Microbiology, Agricultural and Food Microbiology, Molecular Microbiology, Biotechnology

It often is believed that most microorganisms are harmful. In reality the vast majority of microorganisms are beneficial. In fact, life as we know it could not exist without microbes. Microorganisms play critical, indispensable roles in the recycling of chemical elements in the biosphere. As animals and plants die, their dead tissues are decomposed by microorganisms, which recycle the tissue components as elements into the biosphere. Organic compounds are degraded into carbon dioxide, which is released into the atmosphere to be fixed once again into organic material by photosynthetic plants, algae and prokaryotes. Proteins, nucleic acids and other organic nitrogenous compounds are decomposed by microorganisms to ammonia, nitrate or nitrogen gas. These products are assimilated into new nitrogenous compounds or, in the case of nitrogen gas, released into the atmosphere. Other chemical elements are recycled in similar fashion by microorganisms.

The decomposition of organic matter by microorganisms is also the basis of sewage treatment. Sewage is the liquid human, animal, plant, industrial and agricultural waste that is carried by a system of pipes and other conduits called sewers to a central discharge point. Sewage must be treated to remove hazardous chemical waste and pathogenic microorganisms before the water can be reused. Chemical wastes in sewage are degraded by microorganisms during a multistage process. The processed sewage is then chlorinated, filtered and/or treated by other means to kill pathogens prior to reuse as water. The effectiveness of such treatment to remove pathogens is generally measured by the presence or absence of indicator organisms. Indicator organisms, although not necessarily pathogenic, provide a relative index of faecal contamination of water. Coliforms and enterococci, which are bacteria that often are associated with human faeces, are two common indicator organisms that are used to monitor water quality. Water that has no indicator organisms and is suitable for drinking is said to be potable.

Although pathogen-free, potable water is not sterile, as evident from the colonization and attachment of microorganisms to the interior surfaces of water pipes. These biofilms, which consist of metabolically active communities of microbes in a matrix of polysaccharides, slow the flow of water and contribute to pipe corrosion. Biofilms are also significant in oral hygiene, where acid-producing bacteria (for example, Streptococcus mutans) embedded in a polysaccharide/glycoprotein matrix develop as dental plaque on teeth.

Applied microbiology

Industry depends heavily on microorganisms for the production of antibiotics, vitamins, enzymes and various other commercial products. Streptokinases produced by Streptococcus are used in medicine to dissolve blood clots. Fungal lactase is taken by individuals who have an intolerance to lactose and need an exogenous source of this enzyme to tolerate milk. Microbial amylases, which hydrolyse starch, are used as desizing agents in the textile industry and as important ingredients in baking, brewing and the manufacture of sugar syrups. See also Antibiotics, and Fungal Fermentation: Industrial

Certain microorganisms are useful in extraction of copper and other metals from low-grade ores in a process called leaching. In leaching, a bacterium such as Thiobacillus ferrooxidans oxidizes the ore, resulting in release of the desired metal. As metal resources are depleted, microbial leaching becomes increasingly important in the mining industry because the process permits recovery of valuable metals from the remaining natural supplies of low-grade ores. Microorganisms can also be used to help break down hydrocarbons from oil spills and synthetic chemical compounds that are not routinely degraded by natural processes. Pseudomonas putida, the first living organism to be patented by the United States Patent and Trademark Office, was used to assist in cleaning up the 1989 oil spill in Prince William Sound, Alaska, by the supertanker Exxon Valdez.

Agricultural and food microbiology

Microorganisms have become increasingly important in agriculture; for example, microbes are being used as biological pesticides. Unlike chemical pesticides that have broad lethal effects, are recalcitrant and persist as harmful chemicals in soil for many years, biological pesticides selectively kill specific insects, are not harmful to other living organisms, and regenerate as the microbe grows. Bacillus thuringiensis is one popular bacterial pesticide that is effective for the control of caterpillars, bollworms, cabbage worms and gypsy moths.

Microorganisms have also been used in agriculture to introduce genetic material into plants. The Ti plasmid of the bacterium Agrobacterium tumefaciens is often used as a vehicle in genetic engineering to transfer desired genes into plants to produce transgenic crops that are more resistant to infection, disease and spoilage.

Microbes are the only living organisms that are capable of reducing atmospheric nitrogen, which constitutes 80% of the gases in the earth's atmosphere, to ammonia (a process called nitrogen fixation) for utilization by plants and animals. In some instances, prokaryotes fix nitrogen via symbiotic associations with plants; in others, microbial nitrogen fixation occurs nonsymbiotically. It is estimated that prokaryotes are responsible for fixing over 135 million metric tons of nitrogen annually. Were it not for prokaryotic nitrogen fixation, soils would be nitrogen-deficient and much more chemical fertilization would be required.

The food industry relies heavily on microorganisms for the production of fermented milks and milk products, fermented foods and alcoholic beverages. Milk and milk products, such as acidophilus milk (Lactobacillus acidophilus), yogurt (Lactobacillus bulgaricus, Streptococcus thermophilus), butter (Streptococcus) and cheese (for example, Lactobacillus and Streptococcus), are produced by the direct action of microorganisms on milk. Many important foods, including sausage (Pediococcus), sauerkraut (Lactobacillus, Leuconostoc) and olives (Lactobacillus), are made by microbial fermentations. Yeasts (Saccharomyces cerevisiae) are used in the baking industry to raise, or leaven, the dough and in the brewing industry to ferment barley grains. Carbon dioxide produced by these yeasts causes the dough to rise, and the ethanol produced by the yeasts contributes to the alcoholic content of beer.

Molecular microbiology and biotechnology

No other living organism has been studied as extensively as E. coli. This bacterium has been used for so many studies because it is easy to grow and has a small chromosome. Under ideal growth conditions of optimum temperature and ample nutrients, E. coli doubles in number every 15–20 min and within a few hours produces billions of identical cells in a small laboratory flask. The chromosome of E. coli contains 4.7 million base pairs – approximately 0.1% of the amount of DNA found in a eukaryotic cell. Some of the earliest observations of DNA replication, RNA synthesis and protein synthesis were made with E. coli cells. Our knowledge of the control of gene expression is based primarily on landmark experiments performed with prokaryotes. Microorganisms are adept in regulating the types and quantities of proteins formed. Without these precise molecular regulatory mechanisms, microbial cells would needlessly expend their energy synthesizing unnecessary proteins.

Microorganisms have also served as laboratory models to study the transfer of genetic material between microbial cells. Although prokaryotes do not reproduce sexually, they can transfer genetic material by several mechanisms, including transformation (uptake of naked DNA), transduction (transfer of DNA via a bacteriophage from one cell to another cell) and conjugation (direct transfer of DNA between two cells). Genetic exchange among prokaryotes is an important mechanism that has led to genetic diversity and evolution among these microorganisms, and has formed the basis for genetic engineering.

Through genetic engineering, or the deliberate modification of genetic material in a cell or organism, it is now possible to manipulate DNA and develop products for science and commerce. Prokaryotes play an important role in genetic engineering. They produce enzymes called restriction endonucleases that cleave specific areas of DNA. Using restriction enzymes, one can isolate a gene of interest from a prokaryotic or eukaryotic chromosome and insert this gene into a DNA vehicle called a cloning vector. The cloning vector can then be inserted into a host, where the desired gene is replicated and expressed. Insulin, human growth hormone, interferon and streptokinase are examples of products available through cloned genes. These advances in biotechnology have significantly improved human life through purer and less expensive products made possible by genetic engineering.

Medical Microbiology

Infectious microorganisms capable of causing disease live in delicate balance with a host. When such microbes successfully cause damage to the host, disease results. There are two main mechanisms by which microbes damage a host. Pathogens can produce toxins, which are harmful to the host. These toxins can be secreted by the pathogen (exotoxin) or can be part of the outer membranes of Gram-negative bacteria (endotoxin). Exotoxins are among the most potent toxins. For example, 1 mg of botulinum exotoxin could kill 1000 human beings. Microbial pathogens can also invade and harm host tissue by physically damaging the tissue or by depriving it of nutrients. Microbial invasion is assisted by prokaryotic cell appendages (pili) that aid in attachment to host cells, enzymes (leucocidins) that degrade white blood cells, enzymes (collagenases) that degrade collagen in muscle tissue, and thick outer cell coverings (capsules) that protect the invading microbe from phagocytosis. The host responds to microbial invasion and toxins by producing protective antibodies that bind to and inactivate the microbe or toxin, and white blood cells that attack and phagocytose the invaders.

A human host acquires infectious diseases by different routes, including the respiratory tract, the oral cavity and digestive system, and the skin and genitourinary system. In certain instances, the microbial pathogen may be transmitted directly to the host, as in the cases of sexually-transmitted diseases, such as gonorrhoea (Neisseria gonorrhoeae) and syphilis (Treponema pallidum), or respiratory diseases, such as tuberculosis (Mycobacterium tuberculosis) and influenza (influenza virus). In other instances, the pathogen may be transmitted indirectly by inanimate objects, such as towels, or by other living organisms, such as insects or rodents. Rocky Mountain spotted fever (Rickettsia rickettsii) and malaria (Plasmodium) are examples of diseases that are transmitted indirectly by an insect vector to the human host. After a pathogen successfully infects a host, there usually is an incubation period during which the pathogen multiplies and establishes itself before visible symptoms and disease occur.

Antibiotics are often administered to the host to help in the fight against infectious prokaryotic and eukaryotic microorganisms. Antibiotics kill or inhibit pathogens by inhibiting nucleic acid or protein synthesis, damaging the plasma membrane, preventing cell wall synthesis or interfering with cell metabolism. Viruses, which are unable to reproduce independently and replicate only inside a host cell, are not affected by antibiotics and must therefore be treated by other, limited therapies, such as chemical compounds that specifically block viral enzymes or viral nucleic acid synthesis. The indiscriminate use of antibiotics in recent years has led to the development of microorganisms that are resistant to many common drugs. These antibiotic-resistant microbes represent a challenge to conventional chemotherapy and are of major concern to the medical profession. Microbial infections can also be prevented by the use of vaccines. Vaccines, which prompt the host to produce protective antibodies against infectious agents and other foreign antigens, have especially been helpful in protection against childhood diseases such as diphtheria (Corynebacterium diphtheriae), whooping cough (Bordetella pertussis), poliomyelitis (poliovirus) and measles (rubeola virus).

Importance of Microbiology as a Scientific Discipline

Microorganisms are intimately and intricately interwoven into our daily lives. These ubiquitous microscopic forms of life are found wherever other life forms occur and, in some instances, where no other forms of life exist. These unseen organisms, first observed over three centuries ago by Leeuwenhoek with his crude microscopes, are the basis of all life and play indispensable roles in our biosphere. Life and all that it encompasses could not exist without microbes. The study and understanding of these microorganisms would not be possible without the development of microbiology as a scientific discipline.

Glossary

Biofilm
    Microorganisms attached to a surface in an adhesive matrix of polysaccharides and other compounds produced by the microbial cells.
Conjugation
    The direct transfer of DNA between two adjoining prokaryotic cells.
Genetic engineering
    The deliberate modification of genetic material in a cell or organism.
Gram stain
    A fundamental bacteriological staining technique to differentiate prokaryotes that stain deep violet (Gram-positive prokaryotes) from prokaryotes that stain pink or red (Gram-negative prokaryotes).
Microbiology
    The study of organisms too small to be seen with the unaided eye, specifically bacteria, fungi, algae, protozoa and viruses.
Nitrogen fixation
    The reduction of nitrogen gas to ammonia.
Phylogeny
    The comparative study of evolutionary relationships among organisms.
Transduction
    The transfer of DNA via a bacteriophage from one prokaryote to another prokaryote.
Transformation
    The insertion of naked DNA from one prokaryote into another prokaryote.

Further Reading

  • Atlas RM (1997) Principles of Microbiology, 2nd edn. Dubuque, IA: McGraw-Hill.
  • Black JG (1999) Microbiology: Principles and Explorations, 4th edn. Upper Saddle River, NJ: Prentice Hall.
  • Brock T (1961) Milestones in Microbiology. Upper Saddle River, NJ: Prentice Hall.
  • Lim D (1998) Microbiology, 2nd edn. Dubuque, IA: McGraw-Hill.
  • Madigan MT, Martinko JM and Parker J (2000) Brock Biology of Microorganisms, 9th edn. Upper Saddle River, NJ: Prentice Hall.
  • Olsen GJ, Woese CR and Overbeek R (1994) "The winds of (evolutionary) change: breathing new life into microbiology." Journal of Bacteriology 176: 1–6.
  • Prescott LM, Harley JP and Klein DA (1999) Microbiology, 4rd edn. Dubuque, IA: McGraw-Hill.
  • Tortora GJ, Funke BR and Case CL (2001) Microbiology: An Introduction, 7th edn. Menlo Park, CA: Benjamin Cummings.
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