In this article we have complied various notes on bacteria. After reading this article you will learn about:- 1. Meaning of Bacteria 2. General Characteristics of Bacteria 3. Economic Importance 4. Distribution 5. General Morphology 6. Size 7. Structure 8. Cell Division 9. Staining 10. Nutrition 11. Respiration 12. Reproduction 13. Genetic Recombination 14. Host-Bacterial Interactions 15. Bacterial Culture and Others.
- Notes on the Meaning of Bacteria
- Notes on the General Characteristics of Bacteria
- Notes on the Economic Importance of Bacteria
- Notes on the Distribution of Bacteria
- Notes on the General Morphology of Bacteria
- Notes on the Size of Bacteria
- Notes on the Structure of Bacteria
- Notes on the Cell Division of Bacteria
- Notes on the Staining of Bacteria
- Notes on the Nutrition of Bacteria
- Notes on the Respiration of Bacteria
- Notes on the Reproduction of Bacteria
- Notes on the Genetic Recombination of Bacteria
- Notes on Host-Bacterial Interactions
- Notes on Bacterial Culture
- Notes on the Growth of Bacteria
- Notes on the Variability in Bacteria
- Notes on Bacteriophage
- Notes on the Relationships of Bacteria
- Notes on the Classification of Bacteria
- Notes on Bacterial Nomenclature
Note # 1. Meaning of Bacteria:
Bacteria are microscopic organisms, which are often known as ‘germs’ and ‘microbes’. They are among the simplest forms of life known, and, hence show the characteristics of both plants and animals. Their relationship to other living organism is very obscure.
Though they are placed in the plant kingdom, this does not mean that they are more closely related to plants than to animals. Their inclusion under the plant kingdom is entirely for the sake of convenience.
Since the discovery of bacteria in 1676 by Anton van Leeuwenhoek, a Dutchman, they were objects of curiosity until middle of the nineteenth century, when Louis Pasteur and Robert Kock demonstrated that these organisms are responsible for some of the most important diseases of human beings and animals.
Their brilliant work revolutionized medicine and pioneered the field of antiseptic surgery paving the way for the development of the study of bacteria into independent science of bacteriology. It was Pasteur who disproved the theory of spontaneous generation by furnishing irrefutable evidence that microorganisms arise from pre-existing living entities.
He also demonstrated that fermentation is a biological phenomenon rather than a purely chemical one.
Koch’s experimental methods made possible the formulation of rules or postulates, which are followed whenever possible, before a suspected organism could be finally accepted as the cause of a specific disease. These are known as Koch’s Postulates or Rules of Proof.
The Koch’s postulates are as follows:
1. The organism must be consistently associated with the disease in question.
2. The organism must be isolated from the diseased plant or animal and grown in pure culture and accurately described.
3. The organism of the pure culture, when inoculated back into healthy plant or animal, must be capable of reproducing the characteristic symptoms of the disease.
4. The organism must be re-isolated from the diseased plant or animal tissue, grown in pure culture and must be identical with the original inoculated organism.
All these postulates have been accepted as working guides by workers of diseases of human beings and other animals, and also of plants. But they are not applicable in case of viruses, insects, and fungi which cannot be grown on artificial media.
Note # 2. General Characteristics of Bacteria:
Bacteria are among the smallest of living organisms. They are unicellular (some nulticellular) or thalloid living organisms, sometimes with some differentiation of cells, but without differentiated tissues. Bacterial cells are procaryotic possessing datively simple nucleus without any nucleous and nuclear membrane.
Some species of bacteria are parasites. They attack the living cells of other plants or of animals and secure their food from that source. But most bacteria grow as saprophytes on dead remains or the products of plant and animal life without a direct relationship with living cells.
Parasitic bacteria are responsible for some of the diseases of plants and animals, whereas, the saprophytic kinds may be beneficial in one way or the other.
While most of the bacteria are heterotrophic, a few are photosynthetic and chemosynthetic autotrophs and are capable of synthesizing carbohydrates out of carbon dioxide and water. Photosynthetic autotrophs contain photosynthetic pigment known as bacteriochlorophyll and can carry on a type of photosynthesis — photoautotrophic.
While others, lack such pigment, but can manufacture carbohydrates by chemosynthesis— chemoautotrophic.
More than 3,000 species of bacteria have been described in Bergey’s Manual. It is estimated that each person carries some 1014 bacteria, and that the total human population excretes from its collective gut 1022— 1023 bacteria per day.
Bacterial physical environments range from hot springs at 80°G to refrigerated foods, and from distilled water with trace contaminants to the Dead Sea.
Bacteria reproduce by binary fission. Some forms produce spores. Sexuality in bacteria is a very intriguing problem which is being thoroughly investigated. Transfer of genetic material from one cell to the other has, however, been reported in some bacteria. It has also been reported that bacteria maintain certain specific, inheritable attributes through countless series of cell generations.
They also possess some nuclear mechanism of inheritance. Both DNA and RNA are present in bacterial cells.
Note # 3. Economic Importance of Bacteria:
Bacteria are of major importance to main kind for they include some of his most destructive enemies and also some of his greater benefactors. Injurious species include those responsible for disease of human beings and other animals and those that destroy food and reduce crop production by inducing various diseases of crop plants.
Beneficial species liberate fertilizer elements for growing crops, destroy sewage and other wastes. Activities of many bacteria have been harnessed for various industrial purposes to produce valuable chemicals, medicines and various other products essential for human society.
I. Beneficial Activities.
Some of the beneficial activities of bacteria are:
1. Bacteria and soil fertility:
Of all the living organisms found in soils, bacteria are among the most active. They are especially abundant in the ‘surface layers of soil, decreasing in numbers with depth of soil. These bacteria, along with other soil organisms, play a dominant role in soil fertility.
In general, they succeed in converting insoluble or unavailable materials into forms that can be used by higher plants. Of the essential elements secured by the green plants from the soil, nitrogen is, next to hydrogen and oxygen, the one which is needed in the largest quantities. This element makes up a large part of the molecule of the different proteins of which protoplasm largely consists.
Large quantities of the nitrogen from the soil which has been built into plant proteins of crop plants are removed from the field in the grain or other useful parts.
Although the waste products from domestic animals containing much of the nitrogen of the food which they have eaten and utilized in their metabolism, are in part returned to the soil as fertilizer, much of the nitrogen of the crops removed from the soil is never returned to it. Maintenance of the supply of soil nitrogen is thus one of the principal problems of soil fertility.
The element nitrogen passes repeatedly through a cycle which is known as nitrogen cycle, in which it is first converted from simple to complex compounds and then returned to the simple form in a cyclic order by the activity of bacteria and fungi in collaboration with higher plants and animals. Most plants absorb nitrogen from the soil principally in the form of nitrate ions, although they can also absorb ammonium ions.
Since nitrate ions are continually being removed from the soil by green plants the supply of nitrates would eventually be exhausted if there were no way of replenishing it. Neither the elaborated nitrogenous wastes excreted by animals are directly available for reuse by most green plants.
The conversion of nitrogen from proteins and other nitrogenous compounds into the form of nitrates involves several steps and is due in large part to the activities of bacteria, although some of the earlier stages are also carried on by fungi.
Certain groups of bacteria are involved in the decomposition of nitrogenous organic compounds of both plant and animal bodies and the nitrogenous waste excreted by animals and nitrogen transformation in the soil and thereby play an important role in the maintenance of soil fertility. They are ammonifying bacteria, nitrifying bacteria, and nitrogen fixing bacteria.
The bacterial decomposition of nitrogenous organic compounds in the absence of abundant oxygen usually results in the formation of materials of offensive odour chiefly of sulphur compounds.
Such anaerobic decomposition is termed putrefaction. The decomposition of organic compounds in the presence of oxygen and without the development of odoriferous substance is called decay. Actually there is no sharp line of differentiation between putrefaction and decay.
Both the processes are carried on by bacteria in the soil. The decomposition of organic compounds is important to human society from at least two standpoints. It prevents the accumulation of organic matter, both plant and animal, on the earth, and it results in the formation of simple compounds or set free elements that are returned to the soil to be used again by plants.
The ammonifying bacteria transform various proteinaceous substances into ammonia in the soil. The process is known as ammonification. In the first step in the process, the proteins of the soil organic matter are broken down to amino acids. This occurs during ordinary decay of organic matter and is brought about chiefly by bacteria and fungi.
In the next step, the ammonifying bacteria, together with certain fungi, convert the amino-nitrogen to ammonia. Some of the ammonia escape into the air and is lost.
But most of it usually reacts with water to form ammonium hydroxide without the intervention of bacteria. Ammonium ions can be absorbed directly by most plants and used as a source of nitrogen. The ammonia of the ammonium salts is oxidized to stable nitrates by the nitrifying bacteria and the process is known as nitrification.
In the first step, the ammonia is oxidized to nitrite by bacteria known as the Nitrosomonas group. Finally, the oxidation of nitrite to nitrate is due chiefly to Nitrobacter. Nitrate is most readily utilized by green plants.
Ammonifying and nitrifying bacteria are directly concerned in the transformation of protein compounds of plant and animal dead bodies and the animal wastes into nitrates, and thus are essential in maintaining a supply of nitrates in the soil.
Nitrogen-fixing bacteria assimilate atmospheric nitrogen (which green plants cannot utilize) by converting into organic nitrogen compounds, which are subsequently decomposed by other bacteria and transformed through, a series of stages into nitrates. The process by which they assimilate free nitrogen is called nitrogen-fixation.
There are two types of nitrogen-fixing bacteria:
(i) Non-symbiotic nitrogen-fixing bacteria live independently in the soil. Two prominent genera of this group are Azotobacter and Clostridium. These free-living bacteria fix independently atmospheric nitrogen in their body and convert to organic nitrogen compounds. When they die, the organic nitrogen compounds of their body are made available to higher plants through the activities of other bacteria.
(ii) Symbiotic nitrogen-fixing bacteria, consisting of species of Rhizobium, live symbiotically in small, swellings or nodules on the roots of various seed plants, chiefly leguminous plants. These bacteria fix atmospheric nitrogen to synthesize organic nitrogen compounds. When the leguminous plants are harvested, the roots with nodules containing bacteria are left in the soil.
When they decay, the organic nitrogen compounds made by bacteria become available as nitrates through the process of nitrification. A leguminous crop in a rotation thus possesses an advantage besides that of the crop itself, it increases the amount of nitrogen in the soil.
Each type of leguminous plant requires a particular strain of nodule-forming bacteria in its roots. If this strain is not present in the soil, it can be added artificially by inoculating the seeds before they are sown. The relationship between nodule- forming bacteria and their leguminous plants is one of symbiosis.
The bacteria secure food from the tissues of the leguminous plant, and the leguminous plant obtains nitrogen which is fixed by the bacteria.
The nodule-forming bacteria are thus also known as symbiotic nitrogen-fixing bacteria. These bacteria are widely distributed in soils, so that even in the absence of seed inoculation with these bacteria, leguminous seedling roots sooner or later become infected with symbiotic nitrogen- fixing bacteria.
2. Bacterial Metabolism—its Commercial Importance:
The metabolic processes and their products of bacteria have been utilized in many industries. In their metabolic activities bacteria excrete waste products some of which have exceedingly important commercial uses. The large-scale production of valuable substances from bacterial metabolism is a relatively new field in the economic utilization of bacteria, and, as such, is one in which new discoveries are frequent and often very startling.
(i) Source of Antibiotics:
Soil is perhaps the most important source of microorganisms which produce antibiotic substances. These include filamentous bacteria (actinomycetes). Some of the antibiotic substances are secreted outside the cells and into the environment; others are retained largely within cells and must be separated by extraction.
The modern period of antibiotics began in 1939 with the finding of tyrothricin, produced by a soil bacterium.
Prominent among the antibiotics from actinomycetes are:
streptomycin, discovered in 1944 and obtained from Streptomyces griseus, strtptothricin from S. lavendulae; Chloromycetin, in 1947 obtained from S. venezuelae; aureomycin, in 1948 obtained from S. aureofaciens; neomycin, in 1949 obtained from S. fradiae and terramycin, in 1950 obtained from S. rimow.
Besides these, bacitracin and polymyxin are antibiotics produced by Bacillus subtilis and Bacillus sp. respectively were reported long ago.
(ii) Fermentation—its Industrial Application:
The process of bacterial fermentation and its products have been utilized in various industries. Some of them are: Clostridium acetobutylicum ferments carbohydrates producing acetone, methyl alcohol, and n-butyl alcohol, which have important industrial uses. Very recently, vitamin B2, a commercially important product has been discovered as a product of fermentation of carbohydrates by this species of Clostridium. This vitamin has been one of the more expensive vitamins to obtain in quantity, is valuable as a preventive of certain nervous disorders and other diseases.
The manufacture of vinegar is one of the oldest processes in human history which involves bacterial metabolism. Vinegar production begins with the fermentation of sugars in apple juice to alcohol by yeasts. In the presence of oxygen, the vinegar bacterium, Acetobacter aceli, oxidizes alcohol to acetic acid, which is responsible for the characteristic odour and flavour of vinegar.
Lactic acid as one of the products resulting from the souring of milk, has many uses in the processing of foods, in pharmaceuticals and in the chemical industry. It is named after the milk constituent lactose, or milk sugar. It is probably the oldest known acid, having been discovered by Scheele in 1780.
A group of bacteria designated as ‘lactic acid bacteria’, ferment lactose of milk to lactic acid. All these bacteria are classified in the family Lactobacteriaceae which includes the genera Lactobacillus, Leuconostoc, and Streptococcus.
The dairy industry finds bacteria an essential aid in a number of processes. Butter is sometimes made from cream which has been allowed to undergo ‘ripening’—that is, a lactic acid fermentation, causing it to become sour. The cream is pasteurized, incubated and then churned. This fermentation eventually results in the formation of substances responsible for the characteristic odour of butter.
The process of pasteurization was named after Louis Pasteur, the French scientist who developed it to prevent the spoilage of wines. It is chiefly associated with the treatment of milk to eliminate pathogenic bacteria. Two methods are used to pasteurize milk. In the older of these, the holding process; the milk is heated to 140°F. and held at that temperature for 30 minutes.
Another short time, high temperature method, sometimes termed flash pasteurization, the milk, in thin layers is exposed to a temperature of 160°F. for a minimum of 15 seconds. The milk is then cooled as rapidly as possible to a temperature which retards bacterial growth. Pasteurization destroys pathogenic bacteria without appreciably affecting tie physical and chemical properties of the milk.
The pasteurization of milk is widely recognized as one of the most important public health measures. It is not a process of sterilization. Spores of bacteria are not killed, and since the temperature falls far short of boiling, neither are all the vegetative cells. This is notably true of the lactic acid bacteria although the number of these is greatly reduced.
Pasteurized milk will, therefore, be normally sour, as will raw milk, although more slowly. Besides all these processes, a number of complex processes involving lactic acid or other kinds of bacteria occur during the ripening of cheese.
(iii) Retting of fibres:
Bacteria play an important role in the retting of jute, flax, and hemp fibres. They hydrolyze the pectic substances which act as cement like materials that bind the fibres together. Retting is carried out by immersing the stalks of jute, flax or hemp in water and weighing them down. Water is absorbed by the tissues, causing swelling and the extraction of water soluble substances.
The water becomes highly coloured containing substances that have been extracted from the submerged stalks. This highly coloured water now becomes a good culture medium for the growth of many kinds of organisms. The aerobic organisms reduce the concentration of dissolved oxygen and create an environment suitable for the growth of anaerobes.
The pectic substances are slowly fermented and dissolved by the anaerobes, leaving the fibres intact. During fermentation, various organic acids and gases are produced. The submerged stalks should be removed from water when the reaction has gone to completion; otherwise overrating will result.
They are then thoroughly washed to remove the organic acids, odours, and other undesirable substances and the fibres removed from them are then spread out in the sun or air to dry. The dried material is then ready for dressing. Bacteria responsible for retting of fibres are: Bacillus subtilis, B. polymyxa, Clostridium tertium, and C. felsimium.
(iv) Other fermentation processes:
Certain streptococci and Iactobacilli are used for the preparation of silage for the consumption of cows.
Tobaco is cured and fermented to improve its colour, texture and aroma. Bacteria of the Bacillus megatherium group are used for the curing process. A satisfactory fermentation is associated with a rapid increase in numbers of the Micrococcus candicans and Bacillus subtilis types.
3. Bacteria as human symbionts:
Bacteria promote digestive and possibly other physiological processes in the intestinal tracts of animals. The digestion of cellulose by such herbivorous animals as horses and cattle results in part from cellulose- digesting enzymes excreted by bacteria inhabiting the intestines of these animals.
Bacteria also dwell in large numbers in the human intestinal tract, particularly in the lower part of the small intestine and in the large intestine.
Escherichia coli, formerly called Bacterium coli and Bacillus coli, is a regular inhabitant of the lower intestinal tract of man and other vertebrates. About 40 per cent, of human feces consists bacteria mainly E. coli in milk, water, or food is used as an index of fecal contamination.
E. coli has common been considered chiefly a commensal, a partner of reciprocal parasitism (commensalism). It is also occasionally known as a pathogen. It is now known, however, that E. coli and other intestinal bacteria synthesize considerable quantities of some of the B vitamins and release these into the intestine.
These bacteria may therefore be properly regarded as normally being symbionts rather than mere commensals. The destruction of the bacterial flora of the colon by vigorous dosage with certain antibiotic drugs sometimes causes functional disturbances which suggest that the usefulness of these organisms in human physiology is not limited to the production of known vitamins.
The activity of these bacteria in the human gut and the significance of this activity are not known positively; some physiologists believe that these bacteria carry on certain digestive activities of value of the human body, while others believe that lactic acid and other metabolic products of these normally occurring bacteria inhibit the growth of putrefactive and possibly certain pathogenic bacteria.
Whatever may be their specific physiological significance, it is certain that the maintenance of a normal bacterial flora in the human intestinal tract is essential to the health of the human organism and that any major disturbance in this intestinal bacterial flora results in derangements of health.
II. Harmful Activities by Bacteria:
Among the harmful or wasteful results of bacterial activity arc these:
1. Reduce soil fertility:
Certain soil bacteria reduce soil fertility by depleting the nitrogen content of soil. They are known as denitrifying bacteria. These bacteria are especially abundant and active under anaerobic conditions in wet soil and soil with high organic matter content.
They break down nitrates through intermediate compounds to free nitrogen gas, which escapes into the air and thereby the soil fertility is reduced to a great extent. This process of breaking up nitrates to free nitrogen is designated, as denitrification. Denitrification is favoured by poor aeration of the soil, and nitrates may eventually disappear from water-logged soils.
2. Spoilage of foodstuffs:
Of all agents involved in the spoilage of foodstuffs, the activities of living organisms are undoubtedly the most important. Spoilage is caused principally by bacteria, yeasts and molds. The organisms may be pathogenic or non-pathogenic. Different kinds of organisms produce different types of changes in food. Bacteria are more exacting in their requirements than either the yeasts or the molds.
They require relatively large amounts of moisture, hydrogen-ion concentration usually near the neutral point, and relatively low osmotic pressures. Bacteria cause spoilage of human food in the form of rotting meat and fish, spoiling butter, all kinds of vegetables and fruits. Certain bacteria cause severe types of food poisoning in persons who eat bacteria-contaminated food.
The bacteria found on meat surfaces are usually of Achromobacter and Pseudomonas. These organisms, growing on meat, are probably not danger us to health.
Some of the bacteria concerned with putrefaction of meat are: Bacillus subtilis, B. cereus, Escherichia coli, Proteus vulgaris, and Aerobacter cloacae.
Spoilage of fish is caused largely by bacteria. They belong to the genera Achromobacter, Flavobacterium, Micrococcus, Micro- bacterium and Pseudomonas. Bacteria chiefly responsible for food poisoning are: Micrococcus pyogenes var. aureus, Clostridium botulinum, Salmonella enteritidis and Solmonella typhimurium.
The most common kind of food poisoning is Staphylococcus food poisoning which follows the consumption of food contaminated by staphylococci. A second and less common food-borne disease is caused by bacteria of the genus Salmonella, and is commonly known as Salmonella food infection. Although infection has been traced to several kinds of foods, including milk, meat is usually involved.
Salmonella also infects domestic animals and infections in human beings may result from the consumption of such meat when it is insufficiently cooked. Another type of food poisoning is, Botulism food poisoning which follows the consumption of bacteria contaminated food. It is caused by Clostridium botulinum.
This organism thrives on meat and other high-protein foods like peas and bean in the absence of air, and the spores often survive the temperatures used in home canning. C. botulinum multiplies in food liberating a potent toxin which causes botulism.
3. Cause animal diseases:
That bacteria cause diseases of animals was detected as early as 1850, but the final proof of the bacterial cause of animal disease was obtained in 1876 by Robert Koch.
Bacteria cause tuberculosis of cattle, anthrax of sheep, chicken cholera, pneumonia, glanders in horses, sheep and goats. One of the oldest diseases of animals is anthrax which is caused by Bacillus anthracis.
Species of the genus Rickettsia cause diseases like Rocky Mountain spotted fever, classical typhus fever of man and other animals. They are transmitted to humans via arthropod vector. Strains of Chlamydia trachomatis cause keratoconjunctivitis, trachoma that often results in blindness.
A number of microorganisms isolated from animal mucous membrane and sewage were termed pleuropneumonia-like organisms (PPLO). They were subsequently designated as mycoplasmas. Mycoplasma pneumoniae is the causative agent of primary atypical pneumonia.
4. Cause human diseases:
Many of the serious human diseases are caused by bacteria. Some of them are: tuberculosis (caused by Mycobacterium tuberculosis var. hominis), diphtheria (caused by Corynebacterium diphtheriae), leprosy (caused by Mycobacterium leprae) and tetanus (caused by Clostridium tetani).
5. Cause plant diseases:
Many of the plant diseases are induced by bacteria. Bacteria are usually selective in attacking only particular hosts. It has been seen that bacteria that attack plants do not attack animals.
Comparatively bacterial diseases are more common in animals than plants. Plant pathogenic bacteria are distributed among 8 genera: Pseudomonas, Xanthomonas, Rhizobium (under certain environmental conditions), Agrobacterium, Birwinia, Corynebacterium, Streptomyces, and Mycoplasma.
Most bacteria enter the host through wounds, stomata, hydathodes, lenticels, and nectaries.
Species of Streptomyces are known to penetrate the cuticle directly and the legume nodule bacteria (Rhizobium) penetrate the non-cuticularized root hairs. Since the discovery of Professor T. J. Burrill of the University of Illinois, U.S.A. in 1879, that bacteria are capable of causing plant diseases, many bacterial diseases of plants have been discovered. Bacteria cause various plant diseases.
Some of them are:
(i) Leaf spots:
Invasion and multiplication within the substomatal chamber and the intercellular spaces lead to necrosis of the invaded plant tissue causing spots. The organisms responsible for leaf spot disease produce a vigorous attack on the plant, with the result that the cells become heavily infected and strongly discoloured. The discoloured areas dry up and frequency fall out, leaving holes in the leaves.
Some of the organisms causing leaf spot disease are Pseudomonas angulata, the causal agent of angular leaf spot of tobacco; P. maculicola, causes cauliflower spot; P. meliea, the etiological agent of leaf spot of tobacco; Xanthomonas cucurbitae, causes leaf spot of squash; X. malvacearum, the agent of angular leaf spot of cotton; X. ricinicola, the causal agent of leaf spot of castor bean.
(ii) Extensive blights:
Here the progress of the bacteria is more rapid than leaf spots causing a more extensive and rapid necrosis. Organisms producing blight diseases are capable of penetrating considerable distances between cells, leaving the neighbouring tissue intact. The bacteria grow in the plant juices without producing any digestion of the tissues.
Usually a discolouration of the leaves and branches is produced. Death is due probably to an interference with the flow of the plant sap. Some of the organisms producing blights include Erwinia amylovora, the agent of fire blight or pear blight; E. lathyri, the cause of the streak disease of sweet peas and clover.
(iii) Soft rots:
The major effect is a slimy softening of the tissue by the secretion of an enzyme which diffuses in advance and dissolves the middle lamella of the cells, plasmolysis and death of the cells follow, and the bacteria grow upon the dead plant tissue rather than upon the living cells.
Organisms responsible for soft rots reduce the plant tissue to a soft, very moist, pulpy mass. The organisms producing soft rots differ from the other forms found in the soil in that they have the power to attack healthy plant tissue by the secretion of an extracellular enzyme.
The enzyme dissolves the pectin or cement-like material that binds the plant cells. The action is probably hydrolytic, resulting in the liberation of soluble sugars, which are utilized by the bacteria for food. The result is that the plant tissue is reduced to a mass of separate cells, which become converted later into a slimy, pulpy material.
The important species causing soft rots include the following:
Erwinia aroideae, the cause of soft rot of potato; eggplant, cauliflower, radish, cucumber, cabbage, parsnip, turnip, and tomato; E. atroseptica, responsible for black rot of stem and tuber of potato and other vegetables; E. betivora, the etiological agent of sugar beet rot; E. carotovora, the cause of soft rot in carrot and cabbage.
(iv) Vascular diseases:
In some cases the bacterial invasion in the vascular system becomes systemic, in others, bacteria multiply in the vascular system resulting acute wilt. The bacterial wilts constitute a group of very important and destructive plant diseases. The infecting organisms multiply and accumulate in large numbers in the vascular system, causing an interruption in the flow of sap in the plant.
A complete interruption in the flow of sap results in a rapid wilting of the plant. Some important organisms causing wilt diseases are Bacterium stewartii, the cause of wilt disease of maize; Corynebacterium insidiosum, the agent of vascular disease of alfalfa; Erwinia tracheiphila, the etiological agent of wilt of cucumber, pumpkin, and squash.
(v) Bacterial galls:
Here the primary effect of the bacteria on the host is the stimulation of the cell division hyperplasia leading ultimately to hypertrophy, the combined effect of which results in the gall formation. These abnormal growths are produced by the action of organisms on the meristematic tissue of the plants. In some infections the galls remain small; in others they may assume large proportions. The important organisms producing intumescence diseases include: Agrobacterium rhizogenes the agent of hairy root of apple; Bacterium pseudotsugae, the agent of galls of Douglas fir.
During early part of disease development bacteria are contained primarily in the phloem, rather than the xylem and are more active in the living host tissue at the outer edge of the canker. Example, Cornebacterium michiganense, the cause of canker of tomato.
(vii) Mycoplasmal disorders:
Species of the genus Mycoplasma induce various plant diseases. Some of them are: Little leaf of Brinjal, Greening disease of Citrus, and Grassy shoot disease of sugarcane (GSD).
Note # 4. Distribution of Bacteria:
Bacteria are widely distributed in nature under varied conditions. They vary in numbers from one locality to another, depending upon the environmental conditions. Bacteria are present on and beneath the surface of the earth, in fresh-water and in the sea, on and in other organisms, and on the dust particles which float in the air.
They do not generally occur inside normal, healthy cells of other organisms, but otherwise they are usually found wherever food is available to them.
Some bacteria grow belt at 0°C; others require temperatures above 45°C and may even grow at 80°C. Some require atmospheric oxygen; others are indifferent to or inhibited by it.
Almost all naturally occurring organic compounds can be used as food by one or another kind of bacteria. On or in a substratum suitable for their growth and development they may become extremely numerous. Decaying vegetable and animal materials and solutions rich in organic matter are usually excellent places for the growth of saprophytic species.
Bacteria are useful criteria of water and air pollution and cause many serious diseases of human beings, domesticated animals, and cultivated plants.
These are known as Pathogenic Bacteria. Bacteria constitute a normal part of:
(1) The soil flora,
(2) The intestines of animals promoting digestive and possibly physiological process and are particularly abundant in the tropics. Some can withstand extreme heat (40-75°C) —thermophilic and others extreme cold (0-30°C) psychrophilic. Whereas, the majority of bacteria are intermediate in their temperature requirements (20-46°C)—mesophilic.
Most bacteria are not only harmless but absolutely necessary for the existence of living things—non-pathogenic bacteria. Life could not exist in the complete absence of bacteria. Plants and animals owe their existence to the fertility of the soil, and this in turn depends upon the activity of microorganisms which inhabit the soil.
Bacteria are, involved in nitrogen transformations in the soil and help maintenance of soil fertility.
Note # 5. General Morphology of Bacteria:
Most bacteria are unicellular; there are some mycelioid forms too. Cross-walls are absent except in the mycelioid ones (actinomycetes), where septation does take place previous to sporulation and as a prerequisite to it. Bacterial cells may be motile bearing flagella or non-motile without any flagella.
Shape and Arrangement (Fig. 330):
According to the shape of cells, bacteria may be of four types:
(i) Coccus (pl. Cocci)—cells spherical, (ii) Bacillus (pl. Bacilli)—cells rod-shaped, (iii) Spirillum (pl. Spirilla)—cells cork-screw-like, and (iv) Vibrio or Comma (pl. Commas)—cells ‘comma’-like.
Cocci may be of different forms:
(a) Micrococcus, cells are small and occur singly;
(b) Diplococci, cells divide in one plane and remain attached predominantly in pairs;
(c) Streptococci, cells divide in one plane and remain attached to form chains;
(d) Tetracocci, cells divide in two planes and characteristically form groups of four cells;
(e) Staphylococci, cells divide in three planes in an irregular pattern producing bunches of cocci;
(f) Sarcinae, cells divide in three planes in a regular pattern producing a cuboidal arrangement of cells.
Bacilli are not arranged in patterns as complex as those of cocci, and most occur singly or in pairs—diplobacilli. Some species form chains—streptobacilli, others form trichomes which are similar to chains but have a much larger area of contact between the adjacent cells, and again in others the cells are lined side by side like matchsticks— palisade arrangement.
There are other kinds of bacteria that exhibit a considerably different morphology.
For example, some species of bacteria possess appendages, bacteria of the genus Saprosphira form helical filaments, individual cells of the genus Caulobacter are rod- shaped or fusiform with a stalk sometimes protruding from one pole, species of Streptomyces produce a well-developed branched mold-like mycelium through much narrower than fungal mycelium.
Although most bacterial species have cells that are of a fairly constant characteristic shape, some have cells that are pleomorphic, i.e., that can exhibit a variety of shapes. A bacterial species is generally associated with a definite cell form when grown on standard media under certain specified conditions, such as temperature of incubation of the medium.
Bacteria usually exhibit their characteristic morphology in young cultures and on media possessing favourable conditions of growth.
They are of three forms:
(1) The embryonic,
(2) The mature, and
(3) The senescent.
The embryonic forms correspond to the growth phase and are of long slender uniform cells. The mature form corresponds to the resting phase and is characterized by short cells of small size but more variable in form. The senescent form corresponds to the death phase and show’s great variation in both form and size.
Some morphologists have considered them as definite stages in an orderly life cycle of an organism. Young cells are, in general, larger than old ones of the same species. The changes in age are only temporary; the original forms appear when the old cells are transferred to fresh medium.
Note # 6. Size of Bacteria:
Bacteria vary greatly in size according to the species. Regardless of their size, Hone can be clearly seen without the aid of a microscope. The bacteria most frequently studied in the laboratory measure approximately 0.5 to 10 µm in diameter.
Staphylococci and streptococci may have diameters ranging from 0.75 to 1.25µm. Rod (bacillus) forms, such as typhoid and dysentery bacteria often have a width between 0.5 and 1 µm and a length of 2 to 3 µm. Some filamentous forms may exceed 100 µm in length and diameter between 0 5 and 1.0 µm.
Note # 7. Structure of Bacteria:
A bacterial cell has certain definite structures inside (internal) and outside (external) the cell wall. Some cellular parts, such as the cell wall and cytoplasm are common to almost all cells, again some structures are present in only certain species; still others are more characteristic of certain species than of others.
a. Structures Internal to the Cell Wall:
Immediately beneath the cell wall is a thin membrane or covering called the cytoplasmic membrane (Fig.331 A), also called bacterial cell membrane or simply the plasma membrane. It is semipermeable, selective membrane that controls the passage of nutrients and waste products into and out of the cell.
The bacterial cell membrane is an important centre of metabolic activity where all enzymes for the replication of DNA and other metabolic reactions are located.
It also contains many different kinds of proteins, each of which probably has a specific catalytic function responsible for the transport of many organic and inorganic nutrients into the cell and biosynthesis of membrane lipids and various classes of macromolecules that compose the bacterial cell wall (peptidoglycans, teichoic acid, lipopolysaccharideg and simple polysaccharides).
It often contains important components of the machinery of ATP generation. In some bacteria, the membrane appears to have a simple contour, which closely follows that of the enclosing cell wall.
In others, it is infolded, at one or more points into cytoplasmic region resulting in the increase of surface area of the membrane. The infoldings of the bacterial cell membrane are known as mesosomes. The mesosome performs the functions of DNA replication and septum formation in a bacterial cell. It is a link between cytoplasmic membrane and nuclear material.
Mesosome may be lamellae type, as in the species of Lactobacillus or vesicular type structure found in the species of Bacillus. Bacterial cells are characterized by the absence of mitochondria and Golgi bodies. Hence a large number of enzymes involved in the generation of energy in a bacterial cell are attached to the cytoplasmic membrane (membrane-bounded enzymes).
Mesosomes, like mitochondria, also control the Respiratory activities of the bacterial cells, they are also known as chondrioids.
Cell material, contained within the cytoplasmic membrane may be divided into the cytoplasmic area and chromatinic or nuclear area (nuclear material) (Fig. 331A) which is less dense than the surrounding cytoplasm. The cytoplasmic area is granular and is rich in RNA.
The RNA in combination with protein forms macro- molecular bodies. These RNA-protein particles are called ribosomes which contain enzymes useful in protein biosynthesis.
Ribosomes are small granular structures ranging from 10/µm to 13/µm in size and are about 10,000 to 15,000 in number in a cell. Electron microscope study reveals that each ribosome is composed of two unequal and roughly spherical halves having nearly equal amount of RNA and protein.
The greater the rate of protein synthesis greater is the number of ribosomes. Ribosomes active in protein synthesis occurring groups are the polysomes (polyribosomes). They are held together by messenger RNA. All rapidly growing bacteria contain numerous ribosomes.
The cytoplasm of a bacterial cell also contains volutin granules, also known as metachromatic granules, various kinds of polysaccharides as food reserves; and one or more vacuoles.
Globules of fat also accumulate in bacterial cell cytoplasm. In anoxygenic photosynthetic bacteria there occur extensive intracullar membrane systems which are the site of the photosynthetic apparatus, the infoldings of which accommodate bacteriochlorophyll.
The chromatinic or nuclear area is rich in DNA which can be made visible under the light microscope by Feulgen staining. Bacterial cell does not contain nucleus characteristic as in eucaryotic cell. There is no evidence of nuclear membrane separating the nuclear area from the cytoplasm.
Since the DNA of the bacterial cell does not form a discrete nucleus and is not capable of mitotic and meiotic divisions, it has been suggested that this structure be designated as chromatin body, nucleoid, nuclear equivalent, and even bacterial chromosome.
Besides giving rise to enzymes and other proteins of specific nature in biological systems, DNA molecules are responsible for passing on the necessary information to make a new Cell from one generation to the next.
The basic proteins (histones) are not associated with bacterial DNA.
Electron microscopy reveals that fibrils of extremely long circular molecule of DNA are highly folded to form a compact mass known as chromosome, though it differs from the chromosome of eucaryotic cell as because the DNA molecule does not undergo shortening or thickening. It may assume various shapes, from a sphere to an elongated or dumbell or otherwise.
The bacterial chromosome is about one millimetre long and has a molecular weight of about 2×109. It is covalently closed and occupies about 10 per cent of the volume of the cell. The ends of the polynucleotide threads are covalently joined to make a continuous circular molecule.
DNA in such a configuration is said to be in the covalently closed, circular (CCC) form (Fig. 332a). The chromosome is double stranded having succession of genes (about 10,000 in all) arranged either singly or in groups (operons). The individual genes and operons are controlled by regulatory mechanisms of variable complexity.
Replication of the circular DNA of the bacterial chromosome is unidirectional and autonomous. When replication begins the chromosome is attached to a specific point on the bacterial cell membrane, this is known as replicator site with which the enzyme machinery responsible for DNA replication is associated.
The replication fork is located at the replicator site, and its passage along the chromosome involves movement of the chromosome past the attachment site. Prior to the initiation of replication, a new replicator site adjacent to the old one is formed on the membrane, and the free end of the broken DNA strand is attached to it.
Separation of daughter chromosome is effected by the localized synthesis of the membrane in a region situated between the old and new attachment sites. Membrane growth consequently spreads the attachment sites farther and farther apart (Fig. 332).
Most bacteria carry pieces of DNA, either as extra-chromosomal elements called plasmids or in the form of bacterial viruses (bacteriophages) carried in a quiescent form by the cells. Plasmids are closed circular autonomously replicating double-stranded DNA molecules like the chromosome having molecular weight ranging from 106 and 108 and are 1 per cent of the chromosome in size.
They usually contain three to four genes. Plasmids are attached somewhere on the inner face of their host’s cell membrane.
Some of the plasmids are non-transferable and are lost, others have the ability to transfer themselves from one cell to another. Plasmids transfer genetic material between bacteria to spread resistance to antibiotics and genes for the degeneration of complex organic compounds in geochemical cycle. Like bacteriophages, plasmids are employed as cloning vehicles or vectors.
Both chromosome and plasmids carry genetic information to daughter cells. But plasmids are independently replicating DNA pieces which may sometimes fail to replicate or be distributed to daughter cells leading to loss from the bacterial cell.
Besides plasmids, bacterial cells also have attached to the cell membrane pieces of DNA called episomes which are one-hundredth times the size of the bacterial chromosome. Episomes sometimes replicate autonomously in the cytoplasm and at other times become integrated into the chromosome DNA and replicate with it.
This behaviour distinguishes an episome from a plasmid because the latter does not integrate into the chromosome.
b. Structure of Cell Wall:
External to the cytoplasmic membrane and immediate contact with the cytoplasm is the cell wall which provides a rigid framework to the bacterial cell and also supports and protects the more labile protoplasmic entities from osmotic damage. Thickness of cell wall ranges from 10 to 25 µm. Bacterial cell wall is essential for bacterial cell growth and division.
It is a mucocomplex structure and is composed mainly of diaminopimelic acid (DPA), muramic add, and teichoic acid, in addition to which are present amino acids, aminosugars, carbohydrates, and lipids which form a polymeric substance known as peptidoglycan (mucopeptide murein).
The mucopeptides form the rigid part of the bacterial cell wall. Two different groups of bacteria are commonly differentiated on the basis of their response to a differential stain (Gram stain) named after Danish Bacteriologist Christian Gram.
Those species of bacteria which retain the crystal violet stain are designated as Gram-positive, while others which do not are Gram-negative. The cell wall of Gram-positive bacteria is thicker, more rigid and chemically less complex than that of Gram-negative ones. It is composed of homogeneous layer of 10-50 nm wide having chemical composition—peptidoglycan, teichoic acids, and polysaccharides.
Whereas in Gram-negative bacteria the cell wall is more complex than that of Gram-positive ones. It is thin being composed of inner layer 2-3 nm wide and outer layer 7-8 nm wide with chemical composition—peptidoglycan, phospholipids, proteins and lipopolysaccharides. The lipopolysaccharide which is also known, as endotoxin material determines antigenicity, toxigenicity, and sensitivity to phase infection.
c. Structures External to the Cell Wall:
Structures that may be present external to the bacterial cell wall are: flagella, pili, prosthecae, and capsule. Flagella are extremely thin hair-like helical appendages that protrude through the cell wall and are responsible for swimming motility. They are 0.01 to 0.02 µm in diameter and upto 70 µm in length.
Each flagellum has three parts:
a basal body associated with the cytoplasmic membrane and cell wall, a short hook, and a helical filament visually several times as long as the cell.
A bacterial flagellum usually consists of a single proteinaceous fibril —monofibrillar, which in some species has been d mionstrated to be composed of a sheath and a core. Flagella are composed of protein subunits called fiagellin which has the properties of fibrous proteins as kerasin -and myosin. The number and arrangement of flagella vary with different bacteria but they are generally constant for each species.
Motile bacteria exhibit their motion with the help of flagella which commonly have a rotary motion. Certain helical bacteria exhibit swimming motility, particularly in highly viscous media, yet they lack flagella. They have flagella-like structures located within the cell, just beneath the outer cell envelope.
These are called periplasmic flagella or axial flagella or endoflagella. Some other bacteria are motile only when they are in contact with a solid surface. As they glide they exhibit a sinuous, flexing motion, known as gliding motility.
Bacteria without flagella are known as atrichous (as in the genus Streptococcus) and those with a single polar flagellum—monotrichous (Fig. 333A) (as in the genera Vibrio, Spirillum), with a tuft of flagella at one pole—lophotrichous (Fig. 333 B & E) (as in the genus Pseudomonas), a tuft of flagella at each pole—amphitrichous (Fig. 333C) (as in Alkaligenes faecalis), or several flagella originating at various points over the surface of cell—peritrichous (Fig. 333D) (as in Escherichia coli and genus Salmonella).
Majority of the spherical bacteria are non-motile flagella being rare- among bacilli, there are many motile as well as non-motile species; and most spiral forms are motile.
Bacteria may possess very fine, hollow, nonhelical, filamentous appendages much smaller, shorter and more numerous than flagella which do not form regular waves as flagella do. These appendages are called pili (sing, pilus) or fimbriae (Fig. 331B). They measure less than 10nm in diameter and usually less than one pm long and can be seen only by electron microscopy.
Pili have no Junction for motility. There are many morphological types of pili, at least ten have been recognized.
They are named according to their function. Pili have several functions. One kind of pili known as F-pilus (or sex pilus) serves as the port of entry of genetic material (DNA) between donor and recipient cells during bacterial mating. The sex pili are little hollow protein tubules which serve as conjugation tubes in which there is just enough room for the DNA to transfer itself.
Some pili are concerned with attaching two bacterial cells together prior to the transfer of DNA from one cell to the other. Other types of pili cause bacteria to adhere to one another and to foreign cells, such as red blood cells, epithelial cells, etc. Whereas others serve as attachment sites for bacterial viruses- and pathogenic bacteria.
Again others may also serve to keep bacteria near the surface of liquid or where oxygen is most available.
Some bacteria produce semirigid extensions of the cell wall and cytoplasmic membrane known as prosthecae (sing, prostheca). Prosthecae increase surface area of cells for nutrient absorption and may also serve as aids in attachment to surfaces when they have adhesive substance at the end. Certain non-living ribbon-like or tubular appendage excreted by the cell is termed stalk.
Many bacteria produce a nonliving secretion of viscid material around the external surface of their cell wall.
According to the amount of material produced and the degree of its association with the cell, one of the three terms may be applied: capsules, slime layers and sheaths.
If the material is closely adherent to the cell and detectable only with difficulty it is known as a microcapsule; if equally sharply defined but extensive and readily visible it is called a macrocapsule; if copious in quantity and only relatively loosely associated with the cell and parts freely from it, it is slime layer or free slime or gum and it is designated as sheath ‘ when the amount of material is very insignificant.
The above viscous substance is composed of polysaccharides of several types—dextan, levan, and cellulose. It is believed that capsule and free slime are morphologically and biochemically distinct— the capsule is a part of the cell, whereas the free slime is a secretion.
Capsule formation depends upon the composition of the culture medium, but slime layer may be modified layer of the cell wall formed as an extracellular material of slimy or gelatinous nature.
Capsules are not essential structures because:
(i) They are not synthesized under all environmental conditions,
(ii) mutants exist which have lost the ability to produce them
(iii) cells from which they have been removed by enzymic digestion remain viable.
They may be 10µ thick and are stainable by copper salt. The presence of a capsule is often associated with the virulence of pathogenic bacteria as they are able to resist phagocytosis by the white cells of the blood. Capsules may act as ion exchangers.
Gapsule formation may be responsible for considerable economic loss in dairy and other food industries. Encapsuled bacteria are better protected against unfavourable conditions. They are pathogenic to humans. A number of bacteria often adhere together with their gelatinous material forming the zoogloea stage. The capsule may often function in the storage of food substances or waste materials.
In some aerobic bacteria the capsules form a raft in which are found the actively growing cells of other organisms. Bacterial capsules are species-specific and can therefore be used for immunological distinction of closely related species, for example, Acetobacter xylinum and Pseudomonas aeruginosa.
Note # 8. Cell Division of Bacteria:
Bacterial cells are haploid. In rapidly growing bacterial cells nuclear division proceeds ahead of cell division. The bacterial cell division is completed by the doubling of all the cell constituents followed by partitioning of the cell to produce two daughter cells.
The sequence of events is:
(i) Duplication of DNA,
(ii) Separation of two strands of DNA, and
(iii) Cross-wall formation leading to separation of two daughter cells.
The duplicated DNA separates into two strands and replicates individually. Each of the DNA strands are distributed in two daughter cells. During cross-wall formation, transverse plasma membrane is laid down which is followed by the centripetal growth of cell wall and the membrane is splitted into two halves. Sometimes chain of cells are formed by continued division.
Note # 9. Staining of Bacteria:
Due to smallness, bacteria are invisible to the naked eye. This necessiciates one to study the size, shape and structural characteristics of bacteria with the help of micros- scope which permits a wide range of magnifications. Many techniques have been developed by which specimens of bacteria can be prepared for examination in detail under the microscope.
Depending upon the principle of magnification, microscopes are of two categories:
light (or optical) and electron. Optical microscopy includes: bright-field, dark-field, ultraviolet, fluorescence and phase-contrast. Bright-field microscope is the most widely used instrument to study bacteria.
For this, two general techniques are employed to make microscopic preparations:
(i) Hanging-drop of suspension of organisms in a liquid; and
(ii) Dried, fixed, and stained films or smears.
Hanging-drop preparations are made by placing a drop of the bacterial suspension on a cover slip and inverting it over the concave area of a hollow-ground slide.
Fixed, stained preparations are more widely used for the observation of the morphological characteristics of bacteria. The essential steps in the preparation of a fixed, stained smear are: preparation of the film or smear, fixation and application of one or more staining solutions.
A large number of coloured organic compounds (dyes) are available for staining bacteria. These dyes may be acid, basic, or neutral. Acid dyes generally stain basic cell components, and basic dyes generally stain acidic cell components. A neutral dye is a complex salt of a dye acid with a dye base.
The staining process is dependent upon the nature of the bacterium. Some forms strain easily, others do not. Spores and flagella are difficult to stain.
Simple Staining Method:
A dried fixed smear of bacteria is flooded with a dye solution (say, methylene blue) for a specified period of time, after which this solution is washed off with water and the slide blotted dry. The cells of bacteria usually stain uniformly. This may not be true with some bacteria where the interior part of the cell may be deeply stained than the rest of the cell.
Other Staining Methods:
Several bacterial staining methods are used for studying bacteria.
Some of the commonly used staining methods are given below in brief:
(i) Gram staining:
In this process the fixed bacterial smear is subjected to the solutions of: Crystal Violet, Iodine solution, 95 per cent alcohol (decolourizing agent), and Safranin or some other suitable counter stain.
Bacteria stained by the Gram staining method fall into two groups: Gram-positive bacteria retain crystal violet and appear deep violet; and Gram-negative bacteria lose the crystal violet but appear red as they are stained by the Safranin. The Gram stain has its greatest use in characterizing bacteria.
Earlier workers used Gram-staining technique to classify bacteria into two groups: Gram-positive and Gram-negative. Even now Gram staining is widely used in bacteriological study. In addition to Gram-positive and Gram-negative bacteria, some bacteria are Gram-variable being Gram-positive at sometimes, and Gram-negative at others.
(ii) Acid-fast staining:
In this technique the fixed bacterial smear is subjected to the solutions—Garbolfuchsin (heated), acid-alcohol, and methylene blue in regular sequence. Bacteria retaining the Garbolfuchsin appear red are classified as acid-fast bacteria, and those decolourized by the acid-alcohol and counterstained by the methylene blue are designated as non-acid-fast bacteria.
(iii) Ziehl’s Carbol-Fuchsin:
This is a good spore stain. It is prepared by adding 10 c.c. saturated alcoholic solution of basic fuchsin to 100 c.c. of 5 per cent aqueous carbolic acid.
Flame fixed bacterial smear on a coverslip is to be treated for 5 to 10 seconds with 5 per cent aqueous chromic acid, washed in water, then stained in Ziehl’s Garbol-Fuchsin for one minute heating the solution until it steams, destained in 5 per cent aqueous sulphuric acid, washed in water, counterstained with methylene blue, to be washed dried in the air, and finally mounted in Canada balsam.
Staining combination of 5 per cent aqueous malachite green counterstained with 0 5 per cent aqueous safranin also gives good results.
Ziehl’s Garbol-Fuchsin stain with some procedural modifications is also used for staining bacterial flagella.
To demonstrate bacteria in the host tissue, stain combinations which give excellent result are: Garbol-Fuchsin and light green, Giemsa and safranin.
(iv) Endospore staining:
The endospore smear is allowed to remain in contact with crystal violet or methylene blue stain for a long time by gentle heating and then steaming for 30 to 60 seconds. Heating results in loosening of the vegetative cells from the smear and ultimately only the endospores remain attached to the slide.
Endospores are-coloured green, after washing they may be counterstained with safranin. After rinsing and drying the preparation is ready for observation.
(v) Nuclear staining:
Bacterial smear is fixed in osmic acid vapour, hydrolysed with NHG1 for 8 minutes at 58°C, then stained in Fuchsin, washed, dried and mounted in canada balsam.
(vi) Negative staining:
By this technique bacterial cells are made readily visible without staining in a dark-field preparation. Bacterial suspension is mixed with India-ink or nigrosin, which is then spread in a thin film across a slide and allowed to dry. Under the microscope the bacterial cells appear transparent and outlined by the dark background.
Bacterial cells do not receive any physical or chemical treatment. This technique is advantageous for studying capsuled bacteria.
Note # 10. Nutrition of Bacteria:
Depending on their mode of nutrition, bacteria may be autotrophic (autotrophs) or heterotrophic (heterotrophs).
Most bacteria are heterotrophic. They are unable to synthesize organic substances from carbon dioxide and live at the expense of organic compounds manufactured by other organisms. They are either parasites or saprophytes.
Among the parasitic bacteria there are also gradations of parasitism. For example, obligate parasites, they can survive only at the cost of living protoplasm and facultative parasites usually live upon the living tissues of a host, but may, under certain circumstances, derive nourishment from non-living organic matter. Similarly saprophytic bacteria may be obligate saprophytes and facultative saprophytes.
The strict or obligate heterotrophic bacteria cannot synthesize their complex protoplasm from simple inorganic salts but must have organic compounds, such as proteins, peptones, amino acids, and vitamins for growth. Whereas, the facultative heterotrophic ones show characteristics intermediate between the two autotrophic and heterotrophic, being able to utilize both inorganic and organic compounds.
This group comprises the great majority of bacteria that have been studied and classified.
Some species of bacteria are very selective in the types of organic compounds which they utilize, deriving their nourishment from certain specific substances, whereas, other species can use a great variety of organic substances. Many bacteria live largely upon carbohydrates, while others are primarily feeders upon proteins, fats, amino acids, and other organic substances.
In general, the common types of saprophytic bacteria which are responsible for processes of decomposition in nature are rather cosmopolitan in their nutritional requirements, obtaining nourishment from a variety of organic substances.
Many heterotrophic bacteria take in simple organic molecules and treat them in two ways some molecules are used as building blocks to make the necessary macromolecules for cell growth, while the others are degraded to provide the necessary energy.
A few molecules have a fate which depends on the metabolic state of the organism at the time and the external environment in which it finds itself. Many parasitic, disease-producing bacteria are very exacting in their food demands and thus can utilize only a few specific types of organic compounds.
Thus, certain parasitic bacteria can live only in blood, different species requiring different types of blood, or on certain specific tissues of host plants or animals.
Like other kinds of plants, bacteria manufacture enzymes, with the aid of which they digest foods, converting complex or water-insoluble foods into simpler or water- soluble foods. A number of enzymes have been isolated and prepared from bacteria. The enzymes are classified on the basis of what they do rather than what they are. For example, lactase hydrolyzes lactose to glucose and galactose.
There are also general names for groups of enzymes, such as esterases, carbohydrases, proteases, dehydrogenases, and oxidases. It is customary to name the enzyme after the substance acted upon and to add the suffixase.
Enzymes of bacteria are of two groups, those secreted into the surrounding medium are the extracellular enzymes or exoenzymes and others remaining confined within the cell, the intracellular enzymes or endoenzymes.
The extracellular enzymes usually dissolve or destroy the organic compounds into simpler substances. Whereas, the intracellular enzymes are concerned with the respiratory activities of bacteria. The enzymes produced by bacteria act upon a tremendous diversity of organic compounds.
Some of the enzymes of bacteria include amylase, which converts starch to malt sugar; lipases, which digest fats into fatty acids and glycerol; invertase, which transforms cane sugar into glucose and fructose; maltase, which digests maltose into glucose; and proteolytic enzymes which digest proteins and protein-like compounds into simpler substances such as amino acids.
A minority of bacterial species is autotrophic, that is, capable of synthesizing organic compounds from carbon dioxide and other simple, inorganic substances.
Such autotrophic species thus resemble green plants which are also autotrophic. Autotrophic bacteria may be separated into two groups chemo- synthetic or chemoautotrophic species, which obtain the energy required for their synthesis of organic compounds from the oxidation of certain chemicals; and photosynthetic or photoautotrophic species, which utilize light energy in promoting their food-making processes.
Among the common chemosynthctic bacteria are these:
1. Sulphur bacteria, which inhabit sulphurous waters, soils, and sewage and which oxidize hydrogen sulphide to free sulphur and then to sulphuric acid thereby obtain energy which they utilize in synthesizing organic compounds from carbon dioxide and other inorganic substances.
2. Iron bacteria, which live in iron-bearing waters and oxidize ferrous compounds, thus securing energy for the synthesis of organic compounds.
3. Hydrogen bacteria, which oxidize molecular hydrogen in soils to form water, with the release of energy.
4. Nitrifying bacteria, of which there are two types, those oxidizing ammonia to nitrites, and those oxidizing nitrites to nitrates; both types occur widely and abundantly in soils and secure energy from their oxidative activities.
There are two principal groups of photosynthetic bacteria. They are known as purple bacteria and green bacteria, which can be sharply distinguished from one another by the nature of their photosynthetic pigments. In purple bacteria, there is always one kind of chlorophyll, bacteriochlorophyll, accompanied by a variety of carotinoids, all of which are aliphatic (open chain compounds).
The green bacteria contain either one or two chlorophylls, known as chlorobium chlorophylls, accompanied by alicyclic carotinoids (that is, molecules which contain a closed ring). As a consequence of the chemical differences between their respective pigment systems, purple bacteria and green bacteria which perform photosynthesis have completely different absorption spectra.
Bacterial photosynthesis is always an anaerobic process, since it results neither in the production nor in the consumption of oxygen. Most photosynthetic bacteria are, in fact, strict anaerobes. Consequently, the presence of light and the absence of oxygen are both essential factors for their development in nature.
The photosynthetic bacteria are aquatic organisms and can develop only in the deeper layers where they can find anaerobic conditions. It is very common for them to receive light which has passed through a biological filter composed of some overlying algae.
However, since the wave-lengths of light that are particularly effective for bacterial photosynthesis are ones that are not absorbed by any of the overlying algae, the bacteria can still photo- synthesize effectively under such conditions. The unique pigment systems of the photosynthetic bacteria are thus essential for their survival in competition with algae in natural environments.
Photosynthetic bacteria like green plants use light as a source of energy.
As a consequence all the polymerized carbon in the bacteria can be built up from CO2 by a reaction which is basically:
CO2 + 2H2X Light (CH2O) + H2O + 2X
where (CH2O) denotes cell material and a source of reducing power (H2X) reduces the CO2 to a form that can be ultimately used for biosynthesis.
In green plants the source of reducing power is water, and the general equation is in the form:
CO2 + 2H2O Light (CH2O) + H2O + O2
oxygen gas is evolved. Distinction between bacterial photosynthetic systems and those of green plants is that bacteria produce no oxygen while green plants do.
Photosynthetic bacteria use light energy for the formation of ATP and the accumulation of reducing power by trapping the light in the pigments. The ATP and the reducing power are then re-used to reduce CO2 which do not involve light.
The photosynthetic bacteria can be conveniently grouped in the following manner:
1. Purple sulphur bacteria, containing a purplish pigment which apparently functions in a manner comparable with the activity of chlorophyll in green plants in the synthesis of organic compounds.
2. Purple non-sulphur bacteria, which utilize molecular hydrogen in the presence of light, reducing CO2 in the process and synthesizing organic compounds.
3. Green bacteria, which oxidize H2S in the presence of light reducing carbon dioxide and synthesizing organic compounds.
These chemosynthetic and photosynthetic bacteria comprise only a small number of species and thus form a distinct minority of all bacterial species.
They are widely distributed and their importance in nature is tremendous. Sulphur bacteria, for example, are responsible for the conversion of hydrogen sulphide, a common product of protein decay, to sulphur and then to sulphuric acid, which undergoes chemical reactions in soils to form sulphates, the principal source of sulphur for green plants.
The sulphur bacteria (Beggiatoa) thus play a very important role in nature, since, without their activity, most of the sulphur in nature might be locked up as n-hydro- gen sulphide, and the supply of sulphates, necessary for the nutrition of higher plants, might disappear.
This important activity of sulphur bacteria proceeds not only on land, but also in the fresh-water and salt-water bodies, with the result that hydrogen sulphide in constantly being transformed into sulphur compounds which both land and sea plants can utilize.
Similarly, the nitrifying bacteria maintain in soils a continuing supply of nitrates, the most readily available nitrogen compounds for most green plants. Many biologists believe that the world’s iron-ore deposits are products of iron-oxidizing bacteria. Hence iron bacteria (Leptothrix) of past ages have been important in determining the chemical nature of portions of the earth’s surface.
These bacteria fix (utilize) atmospheric free nitrogen by converting into organic nitrogen compounds which are again decomposed and transformed into nitrates through a series of stages. Thereby soil fertility is increased.
The nitrogen-fixing bacteria are of two types:
(i) Non-symbiotic and
The non-symbiotic nitrogen-fixing bacteria live freely in the soil and fix atmospheric free nitrogen and convert into organic nitrogen compounds. There are many non-symbiotic nitrogen-fixing bacteria of which most common are: Azotobacter and Clostridium. Azotobacter is an extremely active nitrogen fixer in vitro, to fix 5-20 pounds of nitrogen per acre it would utilize 454 kg. of carbohydrate.
There are several genera of symbiotic nitrogen-fixing bacteria of which Rhizobium is widely known. Rhizobium is a Gram-negative rod which forms symbiotic association with the roots of angiospermic plants mainly belonging to the family Leguminosae. Strains of Rhizobium are wide spread as free living bacteria in the soil. In this condition they cannot fix atmospheric nitrogen.
When they enter into the roots of leguminous plants, they induce the roots to produce nodules where they live securing food from the tissues of the nodules.
They use food as a source of energy for the fixation of atmospheric nitrogen. During infection of a root, free-living. Rhizobia aggregate around root hairs in response to some unidentified attractant exuded by the root and gradually work their way inward by means of ‘infection thread’.
Coincident with the passage of the thread, cortical cell division is stimulated to form the bulk of the eventual nodule tissue of polyploid cells. The infection thread usually terminates in a polyploid host cell. It may branch and spread through other tetraploid cells as they are formed.
Many small groups of bacteroids are also formed in the nodule cells. The bacteria utilize the carbohydrates and other food of the leguminous plant and undergo a change in form, considered a type of degeneration, resulting V and Y-shaped organisms. In this condition they utilize atmospheric nitrogen to synthesize organic nitrogen compounds.
Some of these nitrogen compounds become available to the leguminous plant in which the bacteria are growing.
A pigment leghemoglobin is formed by host tissue and colours the nodule pink. Only under this condition there is fixation of nitrogen in the nodules. At the same time bacteria produce nitrogen fixing enzyme nitrogenase which is very sensitive to oxygen.
It is believed that the pigment leghemoglobin combines with molecular oxygen in such a manner that the function of the enzyme nitrogenase is not inhibited, but the combined oxygen can become available for the oxidative metabolism in the host cytoplasm near the bacteroids.
The original source of energy for fixation of nitrogen is host photosynthesis, which also provides the carbon skeletons necessary to combine the ammonia which is the first stable product of nitrogen fixation.
In the entire process the bacteria provide the keyenzyme (nitrogenase) and the host plant root provides the optimum environment in which there is an abundant supply of energy coupled with protection of enzyme against molecular oxygen. Structural organization of the nodule also promotes rapid removal of the products of nitrogen fixation (the accumulation of which inhibits fixation).
Certain Rhizobium strains form effective association with certain legume species, whereas others can infect more than one group of legumes. It was previously believed that Rhizobia could form nodules only on the roots of leguminous plants, but there is recent report of their occurrence in nodules on the rootlets of Trema aspera, a member of the family Ulmaceae.
A number of non-leguminous plants (e.g., Alnus, Ceanothus, and others) in association with the Actinomycetes also form root nodules capable of nitrogen fixation. Non-leguminous root nodules do not contain any haemoglobin-like pigments, though in many other aspects the process of nodule formation resembles that of leguminous one.
The leaves of Pavetta, a member of the family Rubiaceae produce nodules which contain Mycobacterium rubiacearum.
The Chemical diversity of bacterial metabolism shows how extensively bacteria have evolved. In some cases the need for metabolic enzymes has been sacrificed at the expense of having to take up molecules preformed from the environment, and in some pathogenic bacteria this specialization has reached the point that they cannot grow outside the organism that they are infecting.
At the other extreme a totipotent synthetic apparatus within the cell allows bacteria to grow in chemical environments that are far too spartan for organisms which do not derive their energy requirements from light or simple inorganic reactions.
Note # 11. Respiration of Bacteria:
Like all living organisms, bacteria carry on respiration, release energy which is necessary for growth, reproduction and other cellular processes. The respiratory process is controlled by enzymes which are synthesized by bacteria. The activities of these enzymes in relation to respiration are very complex.
Depending upon the conditions under which the respiratory process is carried out the bacteria may be aerobic, anaerobic and facultative anaerobic.
Most aerobic species of bacteria, which live only in an environment containing free oxygen, complete the process of respiration through the aerobic phases, liberating carbon dioxide, water, and energy. Whereas, anaerobic bacteria, which thrive in the absence of free, gaseous oxygen, carry.
On anaerobic respiration or fermentation, in which under the control of certain oxidizing enzymes, foods are broken down, with the formation of carbon dioxide and intermediate organic compounds, such as alcohols and organic acids, and with the release of energy.
Again some facultative anaerobes, when they grow under anaerobic conditions, Win carbon dioxide and intermediate organic compounds, given sufficient free oxygen, these organisms may complete the oxidation of these intermediate compounds to water and carbon dioxide.
As a result of then various metabolic activities, chiefly respiration, different species of bacteria form different types of products.
Some of them are:
lactic, butyric, and acetic acids; gases including carbon dioxide, methane, hydrogen sulphide, hydrogen, and others; and red, yellow, orange, and blue pigments; and other substance. These products are frequently important in distinguishing among different species of bacteria, and are of great importance in industry.
Note # 12. Reproduction of Bacteria:
As a rule bacteria reproduce very rapidly. The common method of reproduction in bacteria is by the process of cell division, commonly known as fission, binary fission, or simple fission, in which a bacterium simply divides into two new one-called bacteria. The individual cells may divide at times repeatedly and multiply in number in one plane or in two or three planes.
Cell division is by in growth and constriction, and may result in semi-permanent cross-walls, dividing the organism into one to many cells.
The daughter cells may separate immediately, or they may remain attached and produce chains or colonies of cells. The fission may be slowed down or often entirely stopped due to shortage of food supply, by the accumulation of poisonous waste products of metabolism, presence or absence of oxygen, or variation of optimum temperature.
Nuclear division in a bacterial cell, as already indicated, is quite different from that of eucaryotic cell.
In the genera Bacillus and Clostridium due to dehydration of the cell protoplast and accumulation of Calcium and dipicolinic acid (DPA), the vegetative cell is transformed into a thick-walled structure known as endospore (Fig. 331G). The endospore is resistant to high temperatures, disinfectants, and other unfavourable factors of the environment.
The spore germinates in a suitable environment by one of several methods, depending upon the species. Only one bacterium comes from each spore. Opinions differ with regard to the functions of spores and as to the factors involved in the process of sporulation.
Spore formation in bacteria is not to be considered a method of reproduction, since the germination of a spore results merely in the reappearance of the protoplasm of the original bacterial cell. It is not a process to increase bacterial numbers, because a cell rarely produces more than one spore. It may be regarded as a stage for the survival of unfavourable conditions.
Bacterial spores thus differ from the reproductive spores of the algae, fungi, and other spore- bearing plants. Bacteria having mold-like mycelial body do not produce endospores, but produce chain of spores, or conidia with which they reproduce asexually (Fig. 338).
Reproduction by budding is not uncommon. But reproduction by more complex methods than budding or fission is more common in bacteria. Many bacteria produce tiny motile gonidia, and almost all have a resting or distributive stage which may be more or less spore-like. Again some aquatic bacteria show alternate nation of generations between motile and non-motile stages.
Typical sexual processes involving fusion of gametes and their nuclei to form a zygote, followed eventually by meiosis, have not yet been clearly demonstrated in bacteria. It is, however, clear that in some bacteria transfer of genetic material takes place from one cell to the other.
Note # 13. Genetic Recombination of Bacteria:
Many microbiologists refused to believe that bacteria had stable hereditary systems which could undergo permanent changes. But in 1943 Max Delbruck and Salvador Luria through a classical experiment demonstrated that bacteria have stable hereditary mechanisms.
The establishment of the similarity between the genetic systems of bacteria and higher forms of life resulted in the experimental use of Escherichia coli. The first demonstration of genetic recombination in bacteria was achieved by Lederberg and Tatum in 1946.
They combined two different auxotrophic strains (strains that use only inorganic materials as a source of nutrients) of E. coli and allowed them to mate.
They found prototrophic (nutritionally independent) colonies growing there, these must have been the result of transfer of genetic material (DNA) from cell to cell and a recombination between the auxotroph’s by conjugation and is indeed a true sexual fusion. But genetic recombination in bacteria is basically different from that of higher plants and animals.
In genetic recombination there is the occurrence of progeny whose combinations of genes are different from those that are present in the parents. In higher plants and animals genetic recombination is the fusion of two haploid gametic nuclei resisting in the development of a diploid nucleus.
The complete genomes of both gametes are involved and recombination comes about from the independent assortment of chromosomes or by the process of crossing-over between homologous chromosomes.
Whereas bacteria are free-living organisms and are not differentiated into gametic and somatic cells. Each cell is a potential gamete.
Bacteria displaying sexuality have cells which behave as gametes and are differentiated into two functional types:
Donors (or males) and recipients (or females).
As a rule, in bacteria only a fraction of the genetic constitution of the donor cell is transferred to recipient cell which contributes its entire genome and cytoplasm resulting in the development of a merozygote, a zygote that is incompletely diploid. Genetic recombination is limited to the diploid portion of the genome and occurs when a new, recombinant chromosome is formed.
The recombinant chromosome is formed from DNA contributed by two different organisms.
The transfer of DNA from donor cells to recipient cells and recombinant forms can be accomplished by three distinct mechanisms:
Cell conjugation (mating), transformation (uptake of naked soluble DNA from the environment), or transduction (bacteriophage infection).
In conjugation there is an actual cell-to-cell contact between the mating bacteria (Fig. 334).
The double-stranded closed DNA opens and part of it is transferred through pili called sex pili which behave as bridges through which DNA is transferred from donors to recipient cells. The amount of DNA transferred is directly related with the duration of conjugation between the mating bacterial cells. In 1952 Hayes first presented evidence for sexual differentiation in E. coli.
It is now known that mating types exist and all pairs are not fertile. Donors (male cells) contain a small circular piece of DNA called the fertility, sex, or F factor and labelled as F+. Recipient (female cells) are labelled as F– and are devoid of F factor. Crosses between two F~ strains are sterile. Only F+xF– crosses yield recombinants. Donor strains are of two kinds: F+ and Hfr (high frequency of recombination).
Donor F+ strain donates only a small portion of its genome, whereas donor Hfr strain donates large amount of genome to the recipients. Sex-factor-DNA is an episome which sometimes replicates autonomously in the bacterial cytoplasm and at other times integrated into the bacterial chromosome DNA and replicates with it.
In addition to Escherichia, conjugation has also been reported to occur in the genera: Salmonella, Pseudomonas, and Serratia.
In transformation, the DNA passes between the bacteria as a naked molecule, and nucleic acid can be isolated in active form from the medium separating the bacteria (Fig. 334). This phenomenon was discovered by Fred Griffith (1928) working on Dip- lococcus pneumoniae.
During this process, lysis (disruption) of donor cell takes place and its DNA becomes fragmented and the fragments are thrown out in the culture medium. The fragments of DNA are taken up by the recipient bacterial cells. The introduced genetic fragment, called an exogenote, undergoes recombination with the recipient bacterial chromosome replacing a homologous segment.
This takes place during late log phase when the recipient bacteria are said to be competent to take up and incorporate donor DNA and its characteristics. The process of recombination which basically involves the replacement of one region in the molecule by different pieces of DNA has given rise to the exciting prospects of ‘Genetic Engineering’.
In 1944 Avery, Macleod, and McCarthy implicated DNA as the transforming principle.
Transduction involves the transfer of DNA from a donor bacterial cell into a recipient bacterial cell through the agency of a virus (bacteriophage) which acts as a vector (Fig. 334).
This phenomenon was discovered by N. D. Zinder and J. Lederberg in 1952 when they searched for sexual conjugation among Salmonella species. During transduction bacteriophage undergoes a rapid lytic growth cycle in its host bacterial cell in which it injects its DNA. The DNA replicates rapidly and directs the synthesis of new phage proteins.
The new DNA combines with new proteins to make phage particles which are released by the destruction of cell wall and lysis of the host bacterial cell.
The new phage particles carry the genes of bacterium in which they are produced. There are some phages known as temperate phages which ordinarily do not lyse the host bacterial cell, carry DNA that can behave as a kind of episome in bacteria. These viral genomes can become integrated into the host bacterial genome. They are then known as pro-phages.
There are several types of transduction:
(i) When a temperate phage transfers any gene on a bacterial chromosome it is called generalized transduction, and
(i) When a temperate phage transfers any gene on a bacterial chromosome it is called generalized transduction, and
(iii) When the phages transduce only those bacterial genes adjacent to the pro-phage in the bacterial chromosome it is known as specialized or restricted transduction. Transduction has been demonstrated in several bacterial species. This technique is used for making new bacterial strains.
Again in lysogenization, the agency is a temperate phage whose DNA on entering into the bacterial cell remains attached to the DNA of the host bacterial cell, reproduces along with it, and is transferred to the progeny without causing lysis of the host bacterial cell. The phage DNA is called the pro-phage.
The phage DNA may be carried along several generations of the host bacteria. Sometimes, when the balance of the host bacterial cell is disturbed, the phage DNA detaches from the bacterial DNA and goes into a virulent state. Because of this latent potential for lysis, cultures containing pro-phage are called lysogenic and the phenomenon is lysogeny.
Note # 14. Host-Bacterial Interactions:
Entire human life is spent with bacteria. Many kinds of bacteria colonize human body surfaces, alimentary tract, and orifices. Human health and well-being are influenced by the presence or absence of bacteria in the surrounding environment. Depending on their nature and activities, bacteria may be non-pathogenic and pathogenic. Non-pathogenic bacteria are harmless.
They often inhabit in various parts of human body giving protection to it against the attack of various pathogens. The activities of pathogenic bacteria are varied. The virulence, or the disease-producing capacity of pathogenic bacteria is determined by genetic properties which may be fully expressed only under certain environmental conditions including physical conditions and host susceptibility.
To initiate infection, pathogenic bacteria must first survive the competition with other microorganisms on the surface of the host and then penetrate into the tissues. Once within tissues, the invading bacteria grow and multiply to produce disease symptoms either by increasing in a local lesion or by spreading throughout the host via the lymphatic’s and blood.
Some bacteria produce poisonous substances of high molecular weight known as toxins. The capacity of bacteria to produce toxin and the potency of the toxin are important factors in the ability of bacteria to cause disease. Some bacteria do not produce toxins that can be demonstrated in vitro. Bacterial toxins when excreted into the surrounding medium are known as exotoxins or retained within the cell are endotoxins.
Exotoxins are proteins and are readily separable from the living producer cells. They are produced mainly by Gram-positive bacteria, highly toxic to the host, heat sensitive, and are unstable to chemicals. Exotoxins produce symptoms of food poisoning within 6 hours when ingested in foods. Since these toxins are absorbed by the gut wall they are called enterotoxins.
Common examples are tetanus and diphtheria toxins. Exotoxins when heated or treated with acid lose their toxicity and the resulting substances are called toxoids. Toxins and toxoids have the ability to stimulate the production of antitoxins, which neutralize toxins in the body of the host. This is important in the protection of susceptible hosts from disease caused by bacterial toxins.
Endotoxins are produced mainly by a Gram-negative bacteria. They are liberated when the bacterial cells disintegrate, relatively heat-stable, do not form toxoids, and are less toxic than the exotoxins.
Plant pathogenic bacteria can be grown with varying degrees of ease on fairly simple culture media such as nutrient agar. Many appear to be rather unspecialized parasites, some seem to have a rather wide host range, whereas others, often considered as parasitically more highly specialized, may attack only a few closely related species of plants.
For example, Xanthomonas malvacearum is mainly restricted to Gossypium. Bacteria with a wide host range probably comprise a number of strains each specialized to a limited range of plants. The problem is complicated by the difficulty of defining a bacterial species satisfactorily. In the past, some so-called species have been based on the host plant attacked.
Some bacteria at present described as distinct species are perhaps better regarded as formae speciales of a larger species comprising a number of parasitically specialized strains. The application of computer techniques which assess the overall simplicity of organisms as based on numerous characteristics may shed light on this problem.
There are a few bacteria which attack fungi. These include Erwinia carotovora on Agaricus, Xanthomonas uredovorus on uredospores of rusts.
Bacteria, including those which attack plants, are very variable organisms, new biotypes arising through mutation, recombination and in other ways. Acquisition and loss of pathogenicity are not uncommon. Some bacterial plant pathogens tend to become non-pathogenic in culture but can often be rejuvenated by passage through their host plants.
Pathogenicity can often be increased by repeated passage through resistant plants and decreased by passage through susceptible ones, presumably due partly to selective survival of races best adapted to the plant used. There is also evidence that some bacteria can be ‘trained’ to attack new hosts.
For example, Xanthomonas malvacearum which infects Gossypium can be made pathogenic to bean plants after four successive passages through beans. The pathogenicity of bacteria tends to be rather unstable and is perhaps affected by the plant with which the bacterium is associated; possibly the associating plant donates genetical material to the bacterium, thereby modifying its pathogenicity.
Note # 15. Bacterial Culture:
The mode of occurrence of bacteria in our environment is very complex. Hence study of bacteria requires techniques for unravelling the complex mixed population or mixed culture of bacteria and other microorganisms into separate, distinct species as pure cultures.
A pure culture is thus a population of cells all derived from a single parent cell. It actually represents an artificial condition for the growth of bacteria imposed on them by laboratory manipulation.
To determine the characteristics, it is imperative that bacteria be isolated and grown in the laboratory as a pure culture.
There are a variety of methods whereby different bacteria can be isolated and grown as pure cultures: the streak-plate and spreading techniques, the pour-plate technique, enrichment-culture technique, the serial-dilution technique, the single-cell isolation technique by the use of micromanipulator.
In most bacteriology laboratories a large collection of identified pure cultures of bacteria, referred to as the stock-culture collection is maintained for screening of new, potentially effective bacteria.
One of the major features of bacteria is their growth characteristics on various media.
Such characteristics are: colour, abundance of growth, and even the odour of the culture provide useful clues for identification.
To determine the growth characteristics of a pure culture it is customary to observe growth of bacteria on: agar media (colonies on plate cultures), agar slants, in nutrient broth and in gelatin stabs.
After inoculation on the medium and after incubation, the cultural characteristics of the organism are determined.
The features of growth on each of the above media to be studied are:
on agar plate cultures—size, margin or edge, elevation, pigmentation, and optical features; on agar slant—amount, margin or edge of growth, consistency of mass of growth, and pigmentation; in nutrient broth—amount of growth, distribution of growth throughout broth, and odour; and in gelatin stabs—growth (no liquefaction) along line of inoculation and liquefaction of gelatin.
Note # 16. Growth of Bacteria:
Bacteria are capable of growing over a wide range of physical conditions and are capable of utilizing many different nutrients, but optimal growth requires certain specific conditions for a given species. Bacterial cells inoculated into a fresh medium under optimum conditions, selectively take up nutrients from their environment.
A major sequence of biochemical synthesis follows. The nutrients that enter the cell from the medium are converted into cell structures —RNA, DNA, protein, enzymes, and other macromolecules.
Cell mass and cell size increase and new cell wall substance is synthesized. Subsequently, the process of binary fission is initiated, ultimately resulting in two new cells. The increase in population is by geometric progression. The time interval required for the cell to divide or for the population to double is known as the generation time.
Not all bacteria have the same generation time which is strongly dependent upon the nutrients in the medium and on prevailing physical conditions.
During this generation time bacteria pass through different phases of growth:
i. The Lag Phase:
It is called the phase of adjustment. The individual cells increase in size -beyond their normal dimensions. Physiologically they are very active and are synthesizing new protoplasm. The bacterial cells are metabolizing, but there is a lag in cell division. As such the population remains temporarily unchanged. At the end of the lag phase the cells divide with gradual increase in population.
ii. The Logarithmic, or Exponential or Log Phase:
During this phase the cells divide steadily at a constant rate and the log of the number of cells plotted against time results in a straight line. Growth rate is maximum during this phase.
iii. The Stationary Phase:
The population remains constant for a time, perhaps as a result of complete cessation of division or the balancing of reproduction rate by an equivalent death rate. This may be due to exhaustion of some nutrients or production of toxic products during growth.
iv. The Phase of Decline or Death:
A variety of conditions contributes to bacterial death, the most important are the depletion of essential nutrients and the accumulation of inhibitory products e.g., acids. The number of viable cells decreases essentially the inverse of growth during the log phase. Bacteria die at different rates, just as they grow at different rates. Some species of Gram-negative cocci die very rapidly, so that there may be very few viable cells left in a culture after 72 hours or less. Other species die so slowly that viable cells may persist for months or ever, years.
v. The Transitional Period:
Between each of the above phases there is a transitional period required, before all bacterial cells enter the new phase.
Physical conditions affecting Bacterial Growth:
Principal factors that affect the physical conditions of bacterial growth are: temperature, moisture, light, and hydrogen-ion concentration.
Bacteria can survive at a temperature range of 0°C to 85 °G or even higher. Again bacteria die slowly when exposed to a low temperature range of 0°C to 5°C, but death rate is very high when the temperature of the medium in which bacteria are growing is quickly lowered.
Bacterial categories based on temperate range of growth:
Spore forming bacteria survive even the boiling temperature. The greater the percentage of water in a medium, the lower will be the temperature required to kill bacteria. Moist heat is a more reliable agent of sterilization than dry heat. Dry bacterial cells are more tolerant to heat than young actively growing cells having more moisture.
Bacteria are more aquatic than terrestrial. They thrive in water of high temperature range and are destroyed in water-logged soils under anaerobic condition. Bacterial cells when dried may temporarily cease activity, but regain normal active condition when moist condition is restored. Again prolonged drying may be lethal to bacterial cells.
The visible light rays are not injurious to the bacterial cells, whereas invisible ones like ultra-violet and infra-red rays affect bacterial cells adversely.
Range of hydrogen-ion concentration suitable for bacterial growth is 5.0 to 9.0.
Since bacterial growth involves the synthesis of complex molecules from more simple precursors, a positive input of energy is needed for bacteria to grow and consequently the bacteria themselves will have to degrade a sufficient number of energy- yielding bonds to provide the necessary biosynthetic potential, and thus anaerobic activity of heterotrophic bacteria must be balanced by an equivalent amount of catabolism.
The source of this energy is the medium, in which bacteria grow and £ break down molecules of the substrate to receive necessary energy.
In many bacterial species these molecules are commonly monosaccharides—notably glucose, but with others amino acids and fatty acids are used for this purpose. The molecular changes that either require or liberate energy are numerous in bacteria. Bacteria use primarily ATP for this purpose.
ATP generated by metabolic inter-conversions in bacterial cells is known as substrate-level phosphorylation reactions. But by far the majority of the ATP generated comes from the conversion of hydrogen atom to H2O. This process is known as oxidative phosphorylation, and approximately three ATP are generated for every pair of hydrogens oxidized to form water.
The conversion of these hydrogen atoms to water is achieved by the cytochrome system of the bacteria working in connection with other hydrogen carriers.
The most efficient alternative source is light energy—photosynthesis, but other bacteria use the chemical inter-conversion of small inorganic molecules—the process known as chemoautotrophy.
Note # 17. Variability in Bacteria:
As a living organism, any bacterium is subject to a temporary or permanent changes. Temporary changes (variations) are physiological or morphological and are induced by such things as changes in either the physical or chemical environment of the bacterium. Reversion to the normal form occurs quickly upon its return to usual conditions.
On the other hand, permanent changes (variations) are brought about by changes in the DNA of the bacterium and are accomplished in any number of ways. In such cases reversion to the normal form occurs very rarely.
The off-springs resulting from binary fission in bacteria are duplicates in morphology and physiology of the parent cell.
In a low percentage a part of the DNA of the nucleoid of the parent cell may change or mutate. If this occurs a sister cell is produced that may give rise- to a culture with characteristics that differ from the parent cell. Mutations can occur spontaneously in nature- spontaneous mutation, and can be induced experimentally— induced mutation.
Mutations express themselves as changes in colony morphology, antigenicity, colour, cell morphology, virulence, phage resistance, bactericide resistance, etc.
Mutations can occur in any part of the bacterial DNA and in some areas of the molecules. As long as the base sequence of the DNA does not change, bacteria will ‘breed true’ from generation to generation. Any change in base sequence, however, will alter the informational content of the DNA and this is likely to produce heritable changes. Such change in base sequence is mutation in DNA.
When the change in the nature of a single base occurs it is point mutation, removal of a section of the DNA is deletion; and removal of a piece of DNA from one position on a replicon to another position on the same replicon, or to a position on another replicon in the same cell is translocation. Again when the sequence of the DNA is altered either by adding or removing a single base pair it is known as a frame-shift mutation.
Point mutation may be caused by a variety of agents known as mutagens which may be chemical compounds. Point mutation can revert by a further change in base sequence when DNA returns to its original structure, this is back mutation to parental type. In deletion a piece of DNA is removed from a replicon and the gap is closed by joining together the ends generated by the process.
The effect of deletions is to remove certain genes completely from the genome and to damage the two genes that span the gap. Deletions are caused by chemical agents and by irradiation. Changes due to translocation are commonly noticed in bacteria where antibiotic resistance markers are present. Such changes are lethal as they destroy essential genes in the cell.
Note # 18. Bacteriophage:
Bacteriophages or bacteria-eating agents or bacterial viruses or just ‘phages’, as they are commonly called, are widely distributed in nature. They are dependent upon living bacteria for their existence, as such, they occur in substrata where bacteria are found. Bacteriophages were independently worked out by F. W. Twort (1915) in England and by F. d’Herelle (1917) in Paris.
Structurally, they resemble other viruses being composed of a nucleic acid (DNA) core surrounded by a protein coat. Morphologically, bacteriophage resembles a tadpole in having a head and a tail-like structure (Fig. 335A) through which it injects the host bacterial cell with nucleic acid through a puncture hole in the cell wall (Fig. 335B).
The nucleic acid then takes control of the cell metabolism and directs the bacterium in the synthesis of more bacteriophage nucleic acid and other materials resulting in the development of new bacteriophages. The newly formed bacteriophages are released by a sudden rupture of the bacterial wall-lysis (Fig. 336).
These bacteriophages are free to inject other susceptible bacteria. There are two types of phage: virulent and temperate. The virulent bacteriophage causes the lysis of the bacterial cell very quickly. Whereas, the temperate bacteriophage replicates in step with its host bacterium being transmitted through cell divisions without necessarily causing lysis.
Note # 19. Relationships of Bacteria:
Little is known positively concerning the origin of bacteria upon the earth’s surface. It is generally agreed that bacteria were among the living organisms that appeared on the surface of the earth, constitute a group of great antiquity.
A large group Of Biologists are of opinion that chemosynthetic organisms probably appeared earlier than the green plants and iron bacteria are responsible for the earth’s tremendous ore deposits.
The simplicity of bacterial cell and mode of reproduction indicate close relationship with blue-green algae, and some fungi. Some Biologists regarded bacteria as a primitive group of plants from which both chlorophyllous and non-chlorophyllous plants may have evolved. Whereas, others prefer to accept bacteria as degenerate organisms and have developed by the loss of chlorophyll.
Since the bacteria have flagellate representatives, there is a possibility that all the bacteria have flagellate ancestry. These ancestors formed a group which had flagellate representatives capable of a photosynthetic, chemosynthetic, and saprophytic mode of existence, and thus had much in common with the smaller flagellate protista.
Accordingly, it is not unlikely that the bacteria arose independently from the flagellate protista just as all other living organisms except the blue-green algae which have branched off much earlier than the flagellate protista having a common source of origin which may be designated as ‘proto cell’ (Fig. 337).
As to the relationships, many authorities hold that the bacteria are closely related to the blue-green algae in various aspects.
But the important and fundamental points which distinguish the bacteria from the blue-green algae are that the latter are unique among major groups of living organisms in showing no sign of even possessing any flagellated structure or sexuality. Whereas bacteria, in common with other plants and animals, have representatives which possess flagella or sexuality.
Note # 20. Classification of Bacteria:
The classification and identification of bacteria involve certain difficulties not found in other groups of plants. These difficulties arise in part from the very small size of bacteria and consequent difficulty of recognizing clearly their structural characteristics in part from the relatively restricted range of morphological differences among different species.
In most other groups of plants, morphological differences among species are rather apparent.
In bacteria, however, there is relatively little variation in body structure, and thus different species of bacteria frequently appear similar or identical in structure when they are examined microscopically. There are, of course some morphological differences among bacteria, such as in numbers, lengths, and positions of flagella, in body form, and size.
These differences are used, of course in distinguishing among certain groups of bacteria, but they are often inadequate taxonomically, since several species of bacteria may resemble each other in their flagellum characteristics, their forms, and their body sizes.
Thus, bacteriologists must often depend upon physiological differences to identify and classify different species of bacteria and must therefore investigate the types of organic compounds which bacteria utilize, the kinds of organic acids, gases, pigments, and other compounds produced in their metabolism, their growth in relation to oxygen supply, the shapes and structure of the colonies which they form and their reactions to various biological stains.
The complete classification and identification of bacteria thus are intricate procedures, founded upon a knowledge of both their morphological and physiological characteristics.
No one system for the classification of bacteria is generally accepted. The three most widely used classifications are those of Lehmann and Neumann, Migula, and Bergey.
A comprehensive classification was published by Lehmann and Neumann in 1896. Almost the same time Erwin F.
Smith proposed a classification of bacteria, more or less a modification of Lehmann and Neumann. A fresh system of classification was suggested by Migula in 1900. Later Bergey in 1923 in Manual of Determinative Bacteriology introduced a new system of classification of bacteria. Bergey’s system of classification attracted attention of a large number of workers.
Subsequently, in 1948 an abridged classification was prepared by a committee appointed by the Society of American Bacteriologists. This committee, known as the Board of Editor-Trustees, has the co-operation of a group of approximately 65 bacteriologists interested in developing the systematic relationship of the various groups of bacteria.
The work of the committee was published under the title of ‘Bergey’s Manual of Determinative Bacteriology’ in honour of Dr. D. H. Bergey who was responsible for developing the first edition of the manual.
In Bergey’s manual, bacteria were separated into five orders which may be briefly characterized as follows:
Simple, undifferentiated forms; spherical; rods may be short or long, straight, curved, or spiral; motile or non-motile; endospores sometimes produced. This is the largest order of bacteria. Most of the bacterial species which are economically important and which have been most intensively studied are members of this order.
Organisms forming elongated cells which have a definite tendency to branch. Their vegetative structures resemble fungal hyphae. These hyphae do not exceed 15 µm and are about 1.5µm or less in diameter. Some are soil saprophytes, others are pathogens of animals (actinomycosis of man, ‘lumpy-jaw disease’ of cattle) and of plants (potato scab). Many of these bacteria are very important in processes of decay in soils.
Filamentous water forms which may show false branching. May or may not be ensheathed. In the ensheathed forms, the sheath is composed of, or is impregnated with ferric hydroxide; in others the sheath may contain sulphur particles.
The relatively long, slender, flexible, non-flagellate cells produce a thin spreading colony. They frequently exhibit a creeping type of movement, and they are embedded in large masses of slime which they excrete. They are widely distributed in soil.
Slender, flexuous cell; body in the form of a spiral with at least one complete turn, 6 to 500 µm in length. These organisms resemble both bacteria and protozoa. They are free-living, saprophytes and parasites of animals causing syphilis, chicken septicemia, and many other diseases.
But in course of time bacteria were further grouped into ten orders based on the morphology of vegetative cells and nature of flagellation:
Rods and Cocci not in groups; motile forms with pertrichous flagella.
Rods and Cocci in some groups; motile form with polar flagella.
Cells in groups, with peritrichous flagella.
Fungus-like mycelioid body, but with very narrow cells.
Motile forms in groups.
Non-flagellate slime producing cells (rods, coccoid) with gliding movement.
Cells without flagella forming groups with gliding movement.
Cells not in groups, with polar flagella.
Spiral forms without flagella but with rotary motions.
Cells with filterable stages.
In the 8th edition of Bergey’s Manual of Determinative Bacteriology published in 1974, there has been a radical departure from all previous editions for the classification of bacteria. Instead of allocating to orders and families, bacteria were separated into 19 major Groups (Parts) based on a few readily determinable criteria.
Phototrophic Bacteria (18 genera)—predominantly aquatic organisms, photosynthesize by a method different from that of blue-green algae and green plants’ most of them fix atmospheric nitrogen; morphology spiral, spheres to ovoid; Gram- negative; flagellate to nonflagellate; absence of spore development.
Some genera: Rhodospirillum, Chromatium, Chlorobium.
Gliding Bacteria (27 genera)—Myxobacteriales—mostly soil -organisms; characterized by the ability to glide on a solid or liquid surface and by the tough slime layer that surrounds the cells; cells aggregate to behave as a single colonial organism.
Some genera: Beggiatoa, Stigmatella, Leucothrix.
Sheathed Bacteria (7 genera)- found in both clean and polluted waters and in activated sewage sludge; cells grouped together in gelatinous sheaths.
Some genera: Leptothrix, Sphaerotilus, Streptothrix.
Budding and/or Appendaged Bacteria (17 genera)—mainly soils or aquatic organisms; the products of binary fission are usually unequal and may have specialized functions, e.g., stalks, holdfasts, swarming cells, or may resemble fungal hyphae.
Some genera: Hyphomicrobium, Caulobacter, Gallionella.
Spirochaetes (5 genera)—flexuous helical organisms; largely aquatic;
Medically important genera: Treponema, Leptospira, and Spirochaeta.
Spiral and Curved Bacteria (6 genera)—rigidly helical bacteria having single or grouped flagella which impel the cells with characteristic corkscrew motility; free-living aquatic.
Some genera: Spirillum, Campylobacter, Bdellovibrio.
Gram-negative Aerobic Rods and Cocci (20 genera)—soil and fresh and marine water organisms; cause decomposition of organic matter.
Some genera: Pseudomonas, Azotobacter, Rhizobium.
Gram-negative Facultative Anaerobic Rods (26 genera)— organisms of medical importance.
Some genera: Salmonella, Shigella, Yersinia, Vibrio, Escherichia, Erwima.
Gram-negative Anaerobic Bacteria. (9 genera)—irregularly shaped cells non-sporing rods found among the normal flora of the alimentary tract of man and animals; some constitute sulphur-reducing ones found in brackish waters.
Some genera: Bacteroides, Desulphovibrio.
Cram-negative Cocci and Goccobacilli (6 genera) —most of these organisms are parasitic on man and many are pathogenic.
Some genera: Neisseria, Branhamella, Moraxella, Acinetobacter.
Gram-negative Anaerobic Cocci (3 general — these are parasitic bacteria found in the alimentary tract of warm-blooded animals.
Gram-negative Chemoautotrophic Bacteria (17 genera)—-organisms important in the nitrogen and sulphur cycles in soils and water.
Some genera: Nitrobacter, Nitrosomonas, Thiobacillus.
Methane producing Bacteria(3 genera)— found in anaerobic sediments of muddy waters and in anaerobic sewage digestors as well as in the gastrointestinal tract of animals.
Some genera: Methanobacterium, Methanosarcina, Methanococcus.
Gram-positive Cocci (12 genera) — aerobic bacteria common in soil and freshwater (Micrococcus), facultative anaerobes commensal or pathogenic or warm blooded animals (Staphylococcus), and anaerobic cocci include normal flora of man and animals.
Endospore forming Rods and Cocci (P genera)—these include aerobic genus Bacillus and the anaerobic genus Clostridium, both of which contain organisms essential to soil fertility and well-known pathogens such as B. anthracis, CI. tetani, and CI. botulinum.
Gram-positive Non-sporing Rods (4 genera) —bacteria belonging to this part under Lactobacillaceae are associated mainly with milk and milk products.
Actinomycetes and Related Organisms (39 genera)—(Actinomycetales and Coryneforms)—a tendency to form mycelial growth; most soil organisms, some of which are used in the production of therapeutic antibiotics.
The Rickettsias (18 genera) (Rickettsiales and Chlamydiales)— intracellular parasites that can be cultured in the laboratory only by virological methods. Many cause disease in man or other vertebrate or invertebrate hosts and are often transmitted by arthropod vectors, e.g., Rickettsia prowazekii causing house-borne typhus fever, Chlamydia trachomatis causing trachoma. They multiply only inside host cells.
The Mycoplasmas (4 genera)—these procaryotic organisms lack a cell wall and have probably been derived from a number of different origins. Some resemble wall-less forms of known bacteria, but others seem to be less closely related and their relation to bacteria is unclear. Cells are bounded by a triple-layered membrane and can pass through a bacteriological filter.
Some genera: Mycoplasma, Spiroplasma.
In 1984, major change occurred. The scope of the Manual was greatly broadened by incorporating information dealing with ecology, enrichment, isolation, preservation, and characteristics of bacteria. All these aspects were concerned with bacterial classification and identification. A new name—Bergey’s Manual of Systematic Bacteriology was given to the Manual.
The most widely used reference for bacterial classification is Bergey’s Manual of Systematic Bacteriology. It is now published in four volumes which are briefly outlined below. Each volume is divided into a number of major sections. Within each section the bacteria are divided into formal taxa at various levels, most attention being given to families, genera, and species.
It includes the familiar Gram-negative chemoheterotrophic eubacteria having a relatively simple morphology. Some are saprophytes, others are parasites. They do not form prosthecae, sheaths, endospores and cysts. The mode of reproduction is by transverse binary fission.
The major sections of this Volume are:
The Spirochetes; Aerobic/Microaerophilic, Motile, Helical/Vibrioid, Gram-negative bacteria; Non-motile (or Rarely Motile), Gram-negative curved bacteria; Aerobic Gram-negative Rods and Cocci; Facultatively anaerobic Gram-negative Rods; Anaerobic Gram-negative straight, curved, and Helical Rods; Dissimilatory Sulphate- or Sulphur-Reducing bacteria; Anaerobic Gram-negative Cocci; The Rickettsias and Chlamydias; The Mycoplasmas; and Endosymbionts.
It includes Gram-positive chemoheterotrophic eubacteria having a simple morphology without forming prosthecae, sheaths or cysts, some form endospores. Some are saprophytes, and others are parasites. Reproduction is by transverse binary fission.
The major sections are:
Gram-positive Cocci; Endospore-Forming Gram- positive bacteria; Non-spore forming Gram-positive Rods of regular shape; Non-spore forming Gram-positive Rods of irregular shape; Mycobacteria; and Nocardioforms.
Most of the organisms are Gram-negative eubacteria, but some produce methane gas and may be Gram-negative or Gram-positive belonging to the group Archaeobacteria. Some of these organisms are phototrophic, and others are chemolithotrophic. Gliding motility is more common than motility by flagella. They may form sheaths, pros-thecae and stalks. Reproduction is by budding.
The major sections are:
Anoxygenic Phototrophic bacteria; Oxygenic Phototrophic bacteria; Gliding fruiting bacteria; Gliding non-fruiting bacteria; the sheathed bacteria; Budding and/or Appendaged bacteria; Cemolithotrophic bacteria; and Archaeobacteria.
It includes Gram-positive bacteria possessing branched filamentous hyphae (approximately 1 µm in diameter) resembling fungal hyphae. They are saprophytic and chemoorganotrophic soil organisms which degrade plant or animal residues and some produce a wide range of antibiotics.
One genus (Frankia) fixes N2 symbiotically in woody plants. These organisms reproduce by asexual spores which are known as conidia if they are naked, or sporangiospores if they are enclosed in a sporangium.
The major sections are:
Flamentous bacteria that divide in more than one plane; Filamentous bacteria that form true sporangia; Streptomyces and similar genera; and additional filamentous bacteria having uncertain taxonomic placement.
Note # 21. Bacterial Nomenclature:
The 1947 International Microbiological Congress adopted a tentative International Bacteriological Code of Nomenclature which is, in general, similar to the botanical code.
It has the following points:
1. The starting point for the nomenclature of bacteria is 1753.
2. Latin diagnosis is never required.
3. Generic names which are later homonyms of genera of bacteria, plants, or protozoa are illegitimate.
4. Avoid the introduction of generic names that are in use in Zoology.