GENERAL MICROBIOLOGY
Microbial World, History and Development of Microbiology, Scope of
Microbiology
Dr. (Mrs) S. Sharma
Professor
Department of Microbiology
CCS Haryana Agricultural University
Hisar-125001
and
Dr. Neeraj Dilbaghi
Reader
Department of Biotechnology
Guru Jambheshwar University of Science & Technology
Hisar-125001
01-May-2006 (Revised 12-Dec-2006)
CONTENTS
Introduction
Historical Developments
Leeuwenhoek’s Findings
Biogenesis versus Abiogenesis
Contributions of Pasteur
Contributions of Robert Koch and Germ Theory of Disease
Rise of Medical Microbiology
Microorganisms as Geochemical Agents
Microbiology in the Twentieth Century
Activities of microorganisms important in our daily life
Scope of Microbiology
Some recent applications of bacteria
Keywords
Microorganisms, microscope, fermentation, disease
Introduction
Microbiology, the study of microscopic organisms, derived its name from three Greek words:
mikros (“small”), bios (“life”), and logos (“science”). Taken together they mean the study of
microorganisms which are very small and cannot be seen by unaided eye. If an object has a
diameter 0.1 mm or less, eye cannot see it and very little details can be seen in an object having
diameter 1 mm. So roughly speaking organisms having diameter 1 mm or less are called
microorganisms and are studied in Microbiology.
Although microorganisms are ancient by many standards, microbiology itself is a comparatively
new science. The existence of microorganisms was unknown until the discovery of Microscope.
Microscope is an optical instrument which can magnify small objects which cannot be seen by
naked eye. Microscopes were invented in the beginning of 17th century. Early Microscopes were
of two types; Simple Microscope, with a single lens of very short focal length and Compound
Microscope, with two double convex lens system including ocular and objective lens with higher
magnifying power. Most of the epoch making original discoveries about microorganisms was all
made using simple and compound microscopes.
Characteristics of microorganisms
1. Their size is very small.
2. There is no cellular differentiation. They are unicellular and one cell is capable of
performing all the functions. Some microorganisms are multicellular with little or no
cellular differentiation.
3. Microorganisms are present everywhere on the bodies of animals and humans, on plant
surfaces, in the air, water, dust, soil, and even inside the intestinal canal of all insects,
birds, animals and human beings.
Taxonomic Groups
Microorganisms have wide taxonomic distribution and include organisms such as protozoa,
algae, fungi, bacteria and virus.The schematic illustrations of different microorganisms are
shown in Figure S.1.
Protozoa are unicellular eukaryotic organisms, motile having cilia, flagella and pseudopodia,
saprophytic or parasitic. They are generally present in soil, water and marshy places and their
size varies from 5-200 µm. They are animal-like in that they ingest particulate food, lack a rigid
cell wall, do not contain chlorophyll. The study of protozoa is known as protozoalogy. They are
differentiated on the basis of morphological, nutritional and physiological characteristics. Their
role in nature is varied, but the best known protozoa are the few that cause disease in human
beings and animals, such as malaria in humans. Some protozoa are beneficial, such as those
found in stomach of cattle, sheep and termites that help digest food.
Algae are relatively simple organisms, their size varies from 1 µm to several feet. They are
considered plant-like because they contain the green pigment chlorophyll, carry out
photosynthesis, and have rigid cell walls. They are unicellular to multicellular and either motile
or nonmotile. The study of algae is known as Algology or Phycology. These organisms are
autotrophic and are found most commonly in aquatic environments or in damp soil. They cause
problems by clogging water pipes, releasing toxic chemicals into water bodies, or growing in
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swimming pools. But extracts of some species have commercial uses: as emulsifiers for foods
such as ice-creams; as a source of agar used as solidifying agent in microbial medias and as antiinflammatory drugs for ulcer treatment.
A
B
C
D
E
F
G
H
I
J
Figure S. 1. Schemetic illustrations of different microorganisms. [A] Bacillus cereus, [B]
Staphylococcus aureus, [C] Saccharomyces cerevisiae, [D] E.coli, [E] Listeria monocytogenes,
[F] Red algae, [G] Blue green algae Oscillatoria, [H] Amoeba, [I] Hepatitis virus, [J]
Euglena
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Fungi are either saprophytes or parasites. They have eukaryotic cell structure which, like algae,
have rigid cell walls. They form characteristic hyphae called mycelium which may be septate,
nonseptate or coenocytic. They form fruiting structures called conidia or exospores and
endospores. Spores of fungi are always present in air, dust and soil. Multicellular fungi are also
called molds while yeast is an important unicellular fungus. Size range of molds is 2.0-10 µm
and yeast has size varying in the range of 5-10 µm. Molds have considerable value; they are used
to produce antibiotics- penicillin, cephalosporin etc, fermented products like soy sauce, tempeh,
miso, Roquefort and Camembert cheeses, and many other products. But they are also implicated
in various human, animal and plant diseases including athlete’s foot and the moldy spoilage of
grains and peanuts. The unicellular yeasts are widely used in Baking industry and for the
production of all alcoholic beverages like wine, beer etc. On the other hand, some yeasts cause
food spoilage and diseases such as vaginitis and thrush (an oral infection).
Bacteria are unicellular microorganisms. Their size varies from 1-5 µm and have rod, coccus or
spiral shape. They have prokaryotic cellular organization and cell division is usually by binary
fission. Some bacteria having mycelial morphology are known as Actinomycetes and are very
important in production of antibiotics. Bacteria are important in agriculture and play important
role in cycle of biological nitrogen fixation. With respect to food, they are important in
fermentations, food spoilage, food poisoning and food preservation. The wide range of industrial
products derived from bacteria affect the human society in numerous ways. Their activities are of
enormous importance and some are beneficial while others are harmful. The study of bacteria is
known as Bacteriology.
Viruses are ultra-microscopic, noncellular obligate parasites of plants, animals and bacteria as
well as other protists. Their size varies from 0.015µm -0.2 µm and shapes from spherical, rod,
flexuous to cozohedral. They can be seen only under an electron microscope. Unlike cells,
viruses contain only one type of nucleic acid, either DNA or RNA, which is surrounded by a
protein-coat. They lack the cellular components necessary for metabolism or independent
reproduction, viruses can mutiply only on living cells. The study of viruses is known as virology.
Viruses cause large number of diseases in humans (such as AIDS, common cold, poliomyelitis,
SARS, genital herpes, hepatitis etc), plants (tobacco mosaic disease, papaya ring spot disease
etc) and foot-and-mouth disease of animals. In addition, some retroviruses have also been
implicated in the growth of some malignant tumors.
Historical Developments in Microbiology
Some momentous discoveries in science were made by amateurs, rather than by professional
scientists. One of such major stalwarts in the history of microbiology a natural scientist, owned
his own dry goods store, and was also the official wine taster for the city of Delft in Holland.
Antony van Leeuwenhoek (1632 -1723; Figure S.2A) known as the Father of Microbiology,
was a pioneer in the field of Microscopy and used microscopes of his own design and
manufacture. He was a linen merchant who built microscopes as a hobby, had little formal
education and knew only Dutch language. He made about 500 optical lenses that could magnify
objects 275 times and was an amazing feat. He enjoyed using his microscopes to look at the
various things including river water, pepper infusions, saliva feces and more. He communicated
his findings to the Royal Society of London in the form of long series of letters which were
translated and published in the Proceedings of Royal Society.
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Leeuwenhoek constructed many microscopes with a single lens which consisted of a spherical
lens mounted between two small metal sheets of silver or brass (Figure S.2B). The specimen was
placed on the point of a blunt pin and brought into focus by manipulating two perpendicular and
a linear screws.
Leeuwenhoek’s Findings
Leeuwenhoek had unusual degree of curiosity and observed every object that could be seen
through his microscope. He was one of the greatest Innovators driven by curiosity and infinite
energy.
In his letter of September 17, 1683 with his drawings about ‘animals’ in the scrapings of teeth he
described different types of bacteria and called them ‘animalcules’ (Figure S.2C). He also made
magnificent observations on the microscopic structure of seeds and embryos of plants and some
invertebrate animals. He discovered Spermatozoa, RBC and is therefore known as Father of
Animal Histology. He described characteristic microflora of human mouth, curd, vinegar and
infact discovered all the different types of microorganisms known today including protozoa,
algae, yeast and bacteria. He also emphasized the abundance of these microorganisms besides
their great diversity.
Figure S.2 [A] Antony Von Leeuwenhoek. [B] The Leeuwenhoek’s microscope.
[C] Leeuwenhoek’s sketches of bacteria from the human mouth.
He was elected a fellow of the Royal Society of London (FRS) in 1680.The Royal Netherland
Academy of Arts and Science established a ‘Leeuwenhoek Medal’ in his honor in 1877. The
‘Antony van Leeuwenhoek chair’ established in Delft places a strong emphasis on research and
appointments are made on the basis of achievements.
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Origin of Leeuwenhoek’s microorganisms
Soon after the discovery of microorganisms by Leeuwenhoek, scientists began to study the origin
of these small organisms from the point of view of the two schools of thought; One believed in
the Theory of Abiogenesis or the concept of spontaneous generation i.e. living animalcules are
formed spontaneously from nonliving matter, while the other supported the theory of biogenesis
i.e. they are formed from the ‘seeds’ or ‘germs’ of these animalcules which are always present in
the air. It was also believed at that time that many plants and animals can be generated
spontaneously under special conditions.
Biogenesis versus Abiogenesis
It took several clever experiments, which appear too simple today, and more than hundred years
to resolve the controversy. The abiogenesis for plants and animals was disapproved as a result of
the experiments by Italian Physician Fransesco Redi in 1665, who showed that maggots
developing in putrefying meat are the larval stages of flies and will never appear if the meat is
well protected in a vessel with the fine gauge mesh so that flies cannot lay their eggs on meat.
In 1745, John Needham took hot boiling mutton gravy (meat infusion) in a flask and closed this
flask with a cork. He found the spoilage of this infusion and observed animalcules in it. He killed
and destroyed the living matter by boiling and thus concluded that animalcules arose
spontaneously from the meat. In 1769, Lazaro Spallanzani (1729-1799) an Italian naturalist
performed a series of experiments and showed that heating can prevent appearance of
animalcules in infusion although duration and level of heating required is variable. He was not
satisfied with using cork to plug the flask and sealed it hermetically to prevent contact with the
air completely. He found that sealed infusions remained barren for a long time. A tiny crack in
the flask can result in development of animalcules and they will not appear unless new air
entered the flask to come in contact with the infusion. Spallanzani infact took a series of flasks
and gave heat treatments for different interval of times. He could distinguish animalcules of
different types, i.e., Superior or animalcules of higher class which were destroyed by slight
heating undoubtedly protozoa and animalcules of lower class which were very minute, and much
more heat resistant - the bacteria.
Although the experiments Spallanzani conducted were very good but faulty experiments
continued to be performed and evidence gathered in favor of abiogenesis. Moreover, Needham
objected to the observations by Spallanzani that there was no growth in the infusions because air
which is essential for life had been excluded from his flasks.
An interesting practical application of Spallanzani’s observation was done by Francois Appert
(1805) for preservation of foods by enclosing them in airtight containers and then heating the
containers called ‘Appertization’. He was thereby able to preserve highly perishable food and the
process named canning (called later) was widely used much before its scientific principles were
understood. He performed an experiment by first making the air free of microorganisms by
passing it through red hot tube. This air which still contained 19.4 % oxygen, with and without
heating was passed through a set of flasks containing boiled infusions, the former remained
unaffected.
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Proof of Biogenesis
During the period when these experiments were done, a new figure was emerging in science, in
the name of Louis Pasteur (1822-1895) in France (Figure S.3). A chemist by training, he
emerged as one of the greatest biologists of the 19th century. His contributions are the most
significant in the history of science and industry and his work with germs and microorganisms
opened new areas of scientific studies. Pasteur was born on 22nd Dec 1822 in the eastern French
town of Dole and later became the Dean of the new science faculty at Lille University in 1854.
We pay tribute to him as he was a great benefactor of humanity.
Figure S.3: Louis Pasteur in his laboratory
Pasteur first demonstrated through a series of definitive experiments that air contains
microscopically observable ‘organized bodies’. He aspirated large quantities of air through a
tube which contained a plug of cotton to serve as a filter. He removed the cotton plug and
suspended it in a solution of alcohol and ether. When he examined the sediments microscopically
he observed the presence of small oval shaped bodies. He later confirmed that when heated air is
passed through a boiled infusion no microbial development takes place but when cotton plug is
suspended in the heated infusion, microbial growth occurs. Pasteur repeated his experiment
through swan necked and gooseneck flasks (Figure S.4) so that the germs from air cannot ascend
into it. He boiled the broth in it to kill all microorganisms in the neck as well as in the flask. The
infusion remained sterile in this flask until the neck of the flask was broken resulting in the
growth of microorganisms. Thus he established that development of microorganisms in organic
infusions bring about chemical changes. Schroeder and Von Dusch (1850) started the technique
of cotton plugging because cotton acts as a filter and traps microorganisms.
One of the traditional arguments against biogenesis was the claim that heat used to sterilize the
air or specimens was destroying an essential “vital force”. Those supporting abiogenesis said
that, without this force, microorganisms could not spontaneously appear. In response to this
argument, an English Physicist John Tyndall (1820-1883, Figure S.5) conducted experiments in
a specially designed box called ‘Tyndall chamber’ to prove that dust carries the germs. He
demonstrated that if no dust was present, sterile broth remained sterile for indefinite period.
While doing these experiments, Tyndall (1877) also devised a process for complete sterilization
by alternate heating and cooling known as ‘Tyndallization.’ He found that in some cases even
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boiling the infusion for more than 5 hours was not sufficient to sterilize it and concluded that
bacteria have both thermo stable and thermo labile phases. These thermo stable resting bodies
were also observed by Ferdinand Cohn in hay bacteria and were called endospores. The
experiments by Pasteur and Tyndall finally disapproved the theory of spontaneous generation
and promoted the general acceptance of theory of biogenesis.
Figure S.4: Pasteur’s gooseneck flask
Figure S.5: John Tyndall
Other contributions of Pasteur
Pasteur’s inventions were based on his work on fermentation (1857 -1876) and had a practical
significance. He was called by a winery in Lille in France where they were facing the problem of
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getting a poor product. Careful investigation of the problem by Pasteur led to conclude that
alcoholic fermentation was replaced by another type of fermentation which converted sugar to
lactic acid. Not only did he find the reason for the problem but also the remedies. It was his
ability to apply and relate discoveries to practical world that made his contributions very
significant. The important contributions made by Pasteur are summarized as ; a) Fermentation is
a biological process and is brought about by development and activities of microorganisms, b) A
typical fermentation can be defined by its principal end product, e.g; lactic, alcoholic and acetic
acid fermentation, c) Fermentation is a specific process meaning thereby every fermentation is
accompanied by development of a specific type of microorganism which shows physiological
specificity with respect to fermentation e.g., alcoholic fermentation by yeast and lactic acid
fermentation by lactic acid bacteria.
Pasteur also discovered the process known as pasteurization after his name for preservation of
wine by sudden heating to 60-700C for few minutes and then cooling to destroy the harmful
organisms. This not only saved the wine industry but the process was also applied later to
preserve milk and other liquid foods. Today pasteurization is widely used in fermentation
industries, but we are more familiar with it in the dairy industry.
Germ theory of Disease
Even before Pasteur proved by experiments the possibility of microorganisms as agents of
disease, several careful observers had made strong arguments for the germ theory of disease. A
little earlier to this, John Bassi in1836 and M.J. Berkeley in 1845 had shown that silkworm
disease and the great Potato Blight of Ireland was caused by fungus. Few years later, J.L.
Schonlein showed that certain skin diseases in humans are caused by fungal infections. In 1860 a
disease called ‘Pebrine’ was killing large number of silkworms and destroying the silk industry.
Pasteur described that microbes were killing the silkworms and eliminating the worms will wipe
out the disease. He also demonstrated that by weakening the disease germs in lab and then
infecting the weakend germs into animal or person, the animal developed immunity against that
disease.
Antiseptic surgery
In the 1860’s, an English surgeon named Joseph Lister was searching for a way to prevent
microorganisms infecting wounds, as deaths from post surgery infections were frequent and
accounted to about 45% of the total deaths. Lister used dilute solution of phenol/carbolic acid to
soak surgical dressings and by performing surgery under a spray of disinfectant to prevent
airborne infections (Figure S.6). The incidence of surgical sepsis was greatly reduced and this
became a common practice. This also provided indirect evidence for germ theory of disease. His
experiments were the origin of the present-day aseptic techniques used to prevent infections.
Contributions of Robert Koch and Germ Theory of Disease
In Germany, Robert Koch (1843-1910, Figure S.7) confirmed Pasteur's germ theory and took it
several steps further. His investigations began with a study of Bacillus anthracis, which causes a
disease in cattle. He cultured the anthrax bacillus and later used the same techniques in tracking
and culturing the organism responsible for tuberculosis and cholera. Koch won the Nobel Prize
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for his work on tuberculosis, but is perhaps better remembered for his formulation of four basic
principles or postulates of bacteriology known as Koch’s postulates and they are:
Figure S.6 [A] Joseph Lister, [B] Lister’s carbolic spray in use during
an operation
Figure S.7 Robert Koch (1843-1910)
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1. Microorganism must be present in every case of the disease.
2. Microorganism must be isolated from the diseased host and grown in pure culture.
3. The specific disease must be reproduced when a pure culture of microorganism is injected
into healthy susceptible host.
4. Microorganism must be recovered once again from experimentally inoculated host.
He then carried out a series of experiments to demonstrate biological specificity of disease
causing agent. In the mean time Pasteur also undertook work on Anthrax and reached
conclusions similar to findings by Koch. In 1880, Pasteur used Koch’s technique to isolate and
culture the bacteria that caused chicken cholera. To prove his discovery, he arranged a public
demonstration of the experiment that had been repeated successfully in the laboratory. He
injected healthy chickens with pure culture of cholera bacterium and waited for them to develop
the symptoms and die. But the chickens survived. Pasteur, on reviewing very carefully each step
soon, found an explanation. He discovered that bacteria, if allowed to grow old, could become
avirulent. But these avirulent bacteria could still stimulate something in the host to resist
subsequent infection and immunize the host to that disease. He then applied this principle of
immunization for prevention of anthrax, and again succeeded. He called these avirulent cultures
as vaccines and the process of immunization with such cultures as ‘vaccination’. This recognized
the earlier work of Edward Jenner, who had successfully vaccinated a boy named James Phipps
against smallpox in 1798 (Figure S.8). Jenner had observed that milkmaids exposed to cowpox
never developed the serious smallpox, and thus hypothesized that exposure to cowpox somehow
led to protection against smallpox. In order to prove his point, he inoculated James Phipps first
with cowpox material, and later with smallpox- causing material. The boy did not get smallpox.
Figure S.8: Edward Jenner vaccinating James Phipps with cowpox material
Pasteur by now was considered as miracle worker in France and was asked to develop vaccine
against rabies- a disease transmitted to humans from the saliva of infected dogs, cats and wolfs.
Pasteur took the challenge and reproduced the disease in rabbits by inoculating them with saliva
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from rabid dogs. He then removed its brain and spinal cord, dried and pulverized them, and
mixed the powder into liquid and successfully vaccinated dogs with a series of shots of this
mixture. In 1885, a boy named Joseph Miester, who was bitten by a rabid wolf, was inoculated
by Pasteur with his vaccine, which saved his life (Figure S.9). When he saved most of the group
of Russian farmers bitten by rabid wolves, the Czar sent him 100,000 francs, which along with
many other donations from around the world, founded the famous Pasteur Institute in Paris.
Figure S.9 Joseph Miester being examined
by Pasteur’s associate, as Pasteur looks up
Rise of Medical Microbiology
The work on Anthrax and rabies started the golden age of medical bacteriology. The Pasteur
Institute in Paris and Institute in Berlin became the world centers of science of Bacteriology.
Robert Koch developed methods for isolation of pure cultures of bacteria and concentrated his
work more on isolation, cultivation and characterization of disease causing agents of major
diseases in man. On the other hand, Pasteur focused his attention on seeking experimental
evidence of how infectious diseases occur in human body and how recovery and immunity
occurs. This was a great medical revolution and within 25 years, methods for prevention by
immunization or hygienic methods were developed against most of the major bacteria causing
human diseases (Table 1).
Discovery of Filterable virus
At Pasteur Institute, filters to retain bacterial cells were developed and were used to get filtrates
free from bacterial cells. In 1892, a Russian scientist Dmitri Iwanovski filtered infectious
extract from tobacco plant infected with mosaic disease. To his surprise he found that filtrate was
still fully infectious. This specific discovery was confirmed soon and within few years, many
other plant and animal diseases were found to be caused by some submicroscopic agents retained
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in the filtrates passed through bacterial filters. A new class of infectious agents was discovered
and these were called ‘viruses’ (from latin word virus, meaning a slimy liquid or a poison). It
was later found that these agents are different from cellular organisms already known in structure
and development. Frederick Loeffler and P Frosch (1898) found that Foot and Mouth disease
(FMDV) is caused by a virus. Bacterial viruses were independently discovered later by F.W.
Twort and F. d’ Herelle (1917). Stanley (1935) crystallized virus and found that it is made up of
protein and nucleic acid.
Table 1: Some Early Discovery of Bacteria Causing Human and Animal Diseases
Date Disease
Causative agent*
Discoverer
1876 Anthrax
Bacillus anthracis
Koch
1880 Typhoid fever
Salmonella typhi
Eberth
1880 Malaria
Plasmodium spp.
Laveran
1882 Tuberculosis
Mycobacterium tuberculosis
Koch
1880 Cholera
Vibrio cholerae
Koch
1883 Diphtheria
Corynebacterium diphtheriae Klebs and Loeffler
1885 Tetanus
Clostridium tetani
Nicolaier
1887 Meningitis
Neisseria meningitidis
Weichselbaum
1894 Plague
Yersenia pestis
Kitasato and Yersin
1896 Botulism
Clostridium botulinum
Van Ermengem
1898 Dysentry
Shigella dysenteriae
Shiga
1905 Syphilis
Treponema pallidum
Schaudinn and Hauffman
1906 Whooping cough Bordetella pertussis
Bordet and Gengou
* Present name and original names of Causative agent, in many instances, was different.
General characteristics of Virus
1. They are obligate parasites and can grow and multiply in living plant, animal or bacterial
cell as host. Such viruses are known as plant, animal and bacterial virus respectively.
2. They are nucleoprotein particles made up of protein and nucleic acid.
3. They have only one type of nucleic acid either DNA or RNA. No virus contains both.
4. They are ultramicroscopic and can only be seen with the help of an electron microscope.
5. They do not have cellular structure and unit of structure is ‘virion’
Chemotherapy
The methods to control and prevent various diseases became very important and the use of
chemicals to kill or inhibit the disease causing organisms was termed chemotherapy.
Chemotherapy has been practiced for hundreds of years. Mercury was used toto treat syphilis as
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early as 1495, and cinchona bark was (Quinine) was used in South America since 17th century to
treat malaria. Paul Ehrlich (1909, Figure S.10) discovered a synthetic arsenic compound (606th
compound) named Salvarsan that proved effective against syphilis bacterium. The discovery of
antibiotic (penicillin) by Alexander Fleming (1929, Figure S.11) opened another era for
chemotherapy to control infectious diseases as they were found to be most effective
chemotherapeutic agents against bacterial diseases. The discovery of penicillin was an accidental
one. One day Alexander Fleming noticed that bacteria were not growing near the mold in some
of the mold contaminated culture plates. This was identified and established to be Penicillium
notatum. He correctly guessed that the mold was producing some substance that was inhibiting
bacterial growth. His original report went unnoticed for almost ten years, when a group from
Oxford University, led by Howard. W. Florey and E. Chain, conducted clinical trials with
penicillin to prove it as a ‘miracle drug’. Florey, Chain and Fleming later were awarded Noble
Prize in 1945 for their work.
Another major discovery in chemotherapy occurred in 1932, when, G. Domagk, a German
physician, discovered a group of chemicals called sulfonamides, or sulfa drugs, which were
observed to be effective against various kinds of bacterial infections. Domagk won the Noble
Prize for his discovery which helped to launch a second wave of research on chemotherapeutic
agents. Now hundreds of new antibiotics have been discovered for the treatment of several
infectious diseases. Development of vaccines since then has become a continued process and an
industry by itself.
Figure S.10: Paul Ehrlich
Figure S.11: Alexender Fleming
Development of pure culture methods
Around 1870 it was realized that pure cultures must be used for proper understanding of form
and function of microorganisms.
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What is a pure culture? A pure culture of an organism is the culture which contains large
population of only one type of microorganism generally developed from a single cell. Brefeld
introduced the practice of single cell isolation and cultivation of fungi on solid medium
containing gelatin as solidifying agent. Joseph Lister developed serial dilution technique for
pure culture isolation. He devised a small syringe prototype of the modern micropipettes to
dispense small aliquots of liquid in different tubes containing milk so that final dilution
contained one or none of the organism and isolated pure culture of bacteria which was confirmed
by microscopic examination. Robert Koch was experimenting with solid media and used sterile
cut surfaces of potato placed on sterile covered plates to grow bacteria. Since the surface was
opaque it was difficult to examine cultures of bacteria. Richard. J. Petri introduced Petri dish
as a suitable medium container for the culture of bacteria. Pour plate and streak methods for the
isolation of pure cultures were also developed by Koch. The use of gelatin as solidifying agent
had following disadvantages; viz., It is a protein which is susceptible to microbial digestion and
changes from gel to liquid at 280C while the optimum temperature for growth of wide range of
bacteria is between 30 – 370C. Fran Hesse introduced agar- a complex polysaccharide extracted
from red algae as the solidifying agent, which was found to be a suitable solidifying agent
because of following properties; a) It is not digested by bacteria easily and its melting point is
1000C and remains solidified below 44 0C, besides producing a transparent stiff gel and growth
of bacterial/fungal colonies can be seen easily on their surface.
Microorganisms as Geochemical Agents
The pioneering work by Sergei Winogradsky and M. W. Beijerinck (1851-1931) showed that
microorganisms exhibit wide range of physiological diversity and carry out chemical
transformations that can’t be performed at all by plants and animals. They play important role in
cycles of C, N and S etc and are responsible for turnover of matter on earth. They discovered a
unique class of bacteria called chemoautotrophic bacteria which can grow in complete inorganic
environments getting energy by oxidation of reduced inorganic compounds and carbon from
carbon dioxide. Important bacteria in this category are sulfur bacteria which oxidize inorganic S
compounds and nitrifying bacteria which oxidize inorganic N compounds. They also developed
enrichment techniques to culture specific bacteria and their isolation.
Nitrogen fixing Bacteria
Microorganisms play an important role in the fixation of atmospheric nitrogen which cannot be
used directly as source of N2 by most living organisms. Symbiotic N2 fixing bacteria
(Rhizobium) and asymbiotic N2 fixing bacteria (Azotobacter) which use gaseous N2 for synthesis
of their cell constituents help to maintain supply of combined N2 on which all other forms of life
are dependent. These kind of bacteria were also discovered by Winogradsky and Beijerinck.
Microbiology in the Twentieth Century
In the 20th century, studies on microorganisms have contributed towards development of other
disciplines such as industrial microbiology, biochemistry and molecular genetics etc. The
discovery of cell free alcoholic fermentation by H. Buchner in 1897 laid down the foundation
for the beginning of Biochemistry. The discovery that vitamins used by animals are similar to
growth factors required by bacteria led to the finding that there is similarity of metabolism in all
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living systems and hence microorganisms were used as models to understand basic fundamental
metabolic processes. Escherichia coli has been extensively used in this category to understand
biochemistry and genetics of various cellular processes. George Beadle and Edward Tatum
(1941, Figure S.12) obtained mutants of bread mold Neurospora and studied the consequences of
permanent genetic changes in biochemical terms. Oswald Avery, Colin Macleod and Maclyn
Mc Carty (1944) proved that DNA is the basic genetic material and a model was proposed for
the molecular structure of DNA by James Watson and Francis Crick (1953, Figure S.13).
Collapse of the boundaries between the subjects such as Microbiology, Genetics, and
Biochemistry has lead to deeper understanding of biology at molecular levels under new
discipline called Molecular Biology and Genetic Engineering. The application of Molecular
biology has revolutionized the use of genetically engineered microorganisms for technological
purposes. Now microorganisms are being used to produce non-microbial products at commercial
scale for the welfare of human beings. Production of injectable insulin by genetically engineered
E. coli has opened the possibilities for search and development of other suitable organisms for
production of useful products on industrial scale. The exploitation of microorganisms, their
systems or their processes for technological purposes is studied under Microbial biotechnology
or industrial microbiology.
Figure S.12: George Beadle and Edward Tatum
Activities of microorganisms important in our daily life
Microorganisms play an important role in sustaining life on this planet and in our daily life
through the following activities:
1. Transformation of matter: Microorganisms degrade dead organic matter and return to
the atmosphere in inorganic form. They complete the cycle of matter and are responsible
for transformation of C, N and S and other important elements which are essential for
life.
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Figure S.13: Watson and Crick
2. Biological nitrogen fixation: They fix nitrogen from atmosphere and make it available
to the plants in usable form. Important microorganisms under this category include,
Rhizobium, Azotobacter, Azospirillum etc.
3. Mycorrhiza: Association of roots of many plants with fungi forms a composite structure
called mycorrhiza. Fungus helps in absorption of mineral salts from soil and plant in turn
provides carbohydrates for the growth of fungus.
4. Silage: This method is used to preserve feed with its characteristic flavor, taste and
nutritive value. Leaves of green plants are compacted in size and some molasses is added.
Lactic acid bacteria develop and produce lactic acid which helps to conserve the cattle
feed.
5. Cellulose degradation in Rumen: Ruminants feed on straw and grass which contains
about 50 % cellulose. There is symbiotic association of microorganisms with rumen for
degradation of cellulose and about 1010 – 10 11 cells/ml of different bacteria are usually
present in the rumen. Most important of these include Ruminococcus and Clostridium.
6. Biogas: Animal waste products and cellulose containing waste is fermented by
microorganisms (Methanogens). N2 of the animal excreta is preserved in rotting sediment
and methane gas so formed is used as a fuel.
7. Composting: Decomposition of organic matter by microorganisms to convert it into
nutrient rich manure is known as composting. Bacillus, Aspergillus and
Thermoactinomyces are important in this process.
8. Industrial uses: Different microorganisms are used for the production of wide range of
products at industrial scale. These include alcoholic beverages, antibiotics, enzymes,
pharmaceuticals etc.
Scope of Microbiology
Depending on their applications in different fields, the major areas of applied microbiology are:
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Agricultural Microbiology/ soil Microbiology: Microorganisms related to soil fertility, plant
diseases, transformation of matter, biological nitrogen fixations etc are studied.
Food Microbiology: Microorganisms important with respect to food viz., food fermentations,
food spoilage, food poisoning and food preservation are studied in this area.
Industrial Microbiology: Microbial production of useful products like antibiotics, fermented
beverages, alcohols, industrial chemicals, organic acids, enzymes, hormones etc are studied in
this area.
Medical Microbiology: Besides their usefulness, microorganisms are casual agents of several
diseases of plants, animals and human beings. Many diseases are caused by viruses also. Medical
Microbiology deals with studies on causative agents of disease, diagnostic procedures,
identification of disease causing organisms, development of effective vaccines and preventive
measures etc.
Exo-Microbiology: It deals with exploration of existence of biomolecules and microbial life in
outer space.
Geochemical Microbiology: prospects for deposits of coal, mineral and gas, recovery of
minerals from low grade ores, sea water mining operations, coal, mineral and gas formation and
exploration are studied in this area.
Molecular Biology: is the program of interpreting the specific structure and function of
organisms in terms of their molecular structures. Microorganisms have been used as a tool to
explore fundamental life processes because of many advantages; their fast rate of reproduction,
their growth can be easily manipulated, and lysed cells can be studied in terms of specific
chemical reactions, specific products and specific structures involved.
Genetic engineering and Biotechnology: This is an important development in applied
Molecular Biology which refers to the human capability to alter the genetic make up of an
organism. It has been possible because of the detailed knowledge of structure and function of
DNA and discovery of the restriction enzymes which can cleave or cut the DNA at specific sites
along the chain length. Use of genetically engineered microorganisms has opened great potential
for production of drugs, vaccines, improvement of agricultural crops etc.
Environmental Microbiology: deals with use of microorganisms to protect the environment
from the toxic pollutants, reduction of microbial load in the sewage and industrial wastes,
pesticides, insecticides, heavy metals etc. and to develop suitable methods for treatment of these
wastes and their recycled use.
Some recent applications of bacteria
Biosensors: Bacteria and their components are being used to detect toxic pollutants and
continuous monitoring of nutrients/other parameters during food processing and fermentation
processes. Technically a Biosensor is a miniaturized analytical tool comprising of highly specific
biological sensing element i.e. either integrated within or associated with transducer which
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convert physiochemical interaction into a discrete or continuous digital electronic signals, which
are proportional to single or related groups of analytes. The detection of toxic pollutants in soil,
water and environment is important for the protection of human and animals. Biosensors using
components from bacteria have been constructed which can detect biologically active toxic
pollutants. Such bacterial sensor requires both a receptor which is activated in the presence of
pollutant and a reporter which will make this change apparent. Biosensors use lux operon from
bacteria Vibrio or Photobacterium. This operon contains inducer and structural gene for enzyme
luciferase. In the presence of coenzyme FMNH2, luciferase reacts with molecule to form enzyme
substrate complex which emits blue green light changing FMNH2 to FMN. Hence a bacterium
containing lux gene will emit light when the receptor is activated. This operon has been
transferred to bacteria like:
Xanthomonas…to monitor progression of infection in plants.
Bradyrhizobium...to monitor root nodule formation and development.
Lactococcus…to detect the presence of antibiotics in milk.
Photo bacterium... to detect toxic pollutants.
E.coli……………to detect the presence of mercury in soil samples.
Biosensors only help to detect the presence of pollutants and separate processes are required to
remove these pollutants. They also find application in medical or diagnostics field or in food and
fermentation operations for measurement of specific parameters. In food industry, biosensors can
be incorporated into food packages to monitor temperature abuse, loss of shelf life, microbial
contamination, and to provide visual indicator to consumers of the state of the product at the time
of purchase.
Bioremediation: The concept of using microorganisms to remove pollutants is known as
bioremediation. We can use either indigenous microorganisms or genetically engineered
microorganisms. Pseudomonas and Bacillus are two most important and most commonly used
bacteria for bioremedical purposes. These bacteria can either turn pollutants into energy source
that they consume or alternately they produce enzymes that break down these pollutants into less
harmful molecular products. Studies are being carried out to develop efficient microbial
inoculants for bioremediation of effluents from distillery, tanneries, textile and food processing
industries etc.
Greenhouse Gas bioremediation: There is great concern about the greenhouse effect. Water,
methane and carbon dioxide in earth atmosphere are important as they absorb infra-red
radiations. This process is important for maintaining the temperature of the earth’s surface. In the
recent years because of increased burning of the fossil fuels, carbon dioxide content in the
atmosphere has increased many folds and has caused rise in the temperature on the earth as a
result of the greenhouse effect. Global warming leading to melting of the icy polar caps, rise in
sea level and flooding of major coastal areas, besides warmer air is causing chronic draughts in
major crop growing areas as a result of modified global air circulation pattern.
Some scientists believe that nature automatically takes care of the problem and they emphasize
that extra carbon dioxide stimulates plant growth. They also believe that increase in crop
productivity after the industrial revolution may infact be due to the presence of CO2 in
abundance in the atmosphere and may not be due to the use of fertilizers and pesticides. This
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increased plant growth may be the cause of increased methane in the atmosphere as methane is
five times more effective as a greenhouse effect gas as compared to CO2.
Concentration of methane has increased alarmingly in the atmosphere and has infact doubled in
100 years which may be due to more plant growth resulting in more decaying plant material.
When dead plants decay under anaerobic conditions bacteria break down organic matter and
release CO2 and H2 which is combined to form CH4 and H2O by the activities of anaerobic
methanogenic bacteria.
The use of aerobic methanotrophic bacteria which oxidize methane is important and is being
studied for greenhouse gas bioremediation. The activities of methanogens and methanotrophic
bacteria should be balanced under ideal conditions and the use of chemical fertilizers and
pesticides may have inhibited the growth of methanotrophs. There is a need to determine the
optimum growth conditions for these bacteria for use in greenhouse gas bioremediation.
Suggested Reading
1.
2.
3.
Pelczar, M.J, Chan, E.C.S & N. R. Krieg. Microbiology- Concepts and Applications (International Edition),
McGraw- Hill Inc.
Stanier, R.Y., Ingraham, J.L., Wheelis, M.L. & P.R.Painter. General Microbiology. Fifth Edition, MacMillan.
Prescott, L. M, Harley, J.P & D.A. Klein. Microbiology. Third Edition, WCB Publishers.
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