COVENANT UNIVERSITY

Department of biological sciences

PROGRAM: MICROBIOLOGY

COURSE: MICROBIAL TAXONOMY, NOMENCLATURE AND IDENTIFICATION

CODE: MCB 122

TOPIC: CLASSIFICATION OF BACTERIA AND VIRUSES

INTRODUCTION TO THE BACTERIAL CELL (EUBACTERIA)

Bacteria (singular: bacterium) constitute a large domain of prokaryotic microorganisms. Typically a few micrometres in length, bacteria have a wide range of shapes, ranging from spheres to rods and spirals. Bacteria were among the first life forms to appear on Earth, and are present in most habitats on the planet, growing in soil, water, acidic hot springs, radioactive waste, and deep in the Earth's crust, as well as in organic matter and the live bodies of plants and animals, providing outstanding examples of mutualism in the digestive tracts of humans, termites and cockroaches. Bacteria are microscopic living things that were among the first creatures to inhabit the earth. They can be found almost anywhere, and they come in all forms and shapes, some being harmful to other living things like man, while some are beneficial. People are often scared of being contaminated or infected by bacteria but the types of bacteria which are actually useful in maintaining health and preventing disease.

CLASSIFICATION OF BACTERIA

The classification of the invisible bacterial life. Bacterial classification is based on several major properties, CLASSIFICATION OF BACTERIA BASED ON GRAM STAINING

This method of classification of bacteria was discovered by Hans Joachim Christian Gram 1883, a Danish Physician, while attempting to differentiate bacteria from human tissue by different staining methods. He noted differences in stain retention by bacteria due to the complex differences in the

component and structure of their cell wall. Gram’s Stain, widely used method of staining bacteria as an aid to their identification.

In Gram's method, bacteria are first stained with gentian violet (a dye consisting of a methyl derivative of pararosaniline) and then treated with Gram's solution, consisting of 1 part iodine, 2 parts potassium iodide, and 300 parts water. After being washed with ethyl alcohol, the bacteria will either retain the strong blue color of gentian violet or be completely decolorized. Sometimes a counterstain such as fuchsine or eosin is applied to give the decolorized bacteria a reddish color to make them more visible.

Bacteria that retain the blue stain are known as gram-positive; those that do not are known as gram-negative. Organisms that sometimes retain the blue color and sometimes do not are known as gram-variable. Typical gram-positive bacteria are those staphylococci that produce boils; typical gram-negative bacteria are the bacilli that cause whooping cough; typical gram-variable bacteria are the

bacilli that cause tuberculosis.

The ultrastructure and chemical composition of the cell walls of Gram-positive and Gram negative bacteria are quite different. Inprofile,the cell wall of Gram- positive bacteria reveals a single thick and more or less homogeneous layer, whereas Gram-negative bacteria have a thinner,distinctly layered cell wall with an outer membrane resembling the typical trilaminar cytoplasmic membrane. The polymers found in the cell walls of these two groups of bacteria are chemically quite different. The walls of Gram-negative cells are mainly composed of lipopolysaccharide, phospholipid ,protein, lipoprotein, and relatively little peptidoglycan(usuallylessthan10% of the total cell wall).The Gram-positive cells contain peptidoglycan(usuallymorethan30%ofthetotalcell wall),polysaccharides or teichoicacid(orboth),or teichuronicacid, as major components. Thus, in contrast to the Gram-negative bacteria, the Gram-positive bacteria contain hardly any lipids in their cell walls. There is, however, one exception: acid-fast bacteria. They are resistant to decolorization with acidic ethanol after staining with fuchsin (Ziehl-Nielsen staining). These acid-fastbacteria (Mycobacterium, Nocardia, and Corynebacterium sensustricto) are Gram-positive bacteria which contain large amounts of lipids in their cell walls; inparticular, mycolicacids(high-molecular- weight,3-hydroxyacids with a long alkyl branch

Gram staining classification of bacteria The Gram staining technique attempts to classify bacteria into two wide classes based on their reaction to the stain when viewed under a bright field microscope, it is important to note that this system of classification is based on bacterial cell wall content ,natural classification scheme that reflects major differences in cell wall structure and to some extent the mechanisms involved in disease.

Gram positive: The Gram positive bacteria appear purple in colour under the bright field microscope when viewed with appropriate magnification. They have a single membrane consisting of a thick peptidoglycan layer – no lipopolysaccharide. Lipoteichoic acids and wall teichoic acids make the surface of Gram-positive bacteria negatively charged.

Gram negative: The Gram negative bacteria appear purple in colour under the bright field microscope when viewed with appropriate magnification. they posses inner and outer membranes, with outer membrane having lipopolysaccharide molecules .they are characterised by Relatively thin peptidoglycan layer Outside of the peptidoglycan layer is an outer membrane characterized by unique polymers – Lipopolysaccharide (LPS) extending from the outer surface – Lipoprotein anchoring the outer membrane to the peptidoglycan layer – Porins that facilitate transport of large molecules across the outer membrane.

BUT WHY DOES GRAM STAIN COLOUR DIFFERENTLY ?

In order to understand the action of the gram stain in differentiating the bacterium, we need to have a look at the cell wall composition of the bacteria .In them, the cell wall has a little difference . Cell Wall in Gram positive bacteria -

Has relatively thick layer of peptidoglycan , Techoic acid (polymer of glycerol or ribotol connecterd with phosphate groups,

The various layers of peptidoglycan are linked by covalent linking of techoic acid to muramic acid to form a mesh

Consequence— The crystal violet color is not allowed to decolorize , so the cells appear blue black.

Cell Wall in Gram negative bacteria

In gram negative bacteria, the cell wall is much thinner with low peptidoglycan concentration but high lipid content.

Hence the crystal violet gets decolorized when washed due to restraining activity of the mesh like peptidoglycan in gram positive cells.

Besides, in gram negative bacteria, the outer part of cell wall is high in lipid, polysaccharides and proteins, but there is a space called periplasmic space outside the central membrane external to which is the peptidoglycan followed by the final external membrane

Thus another stain (counter stain ) becomes very important .

Mammals recognize the LPS of Gram-negative– This generates an inflammatory response,Overdone, it can be dangerous. Hence, we know LPS as an endotoxin and Lysozyme catalyzes

MORPHOLOGICAL CLASSIFICATION OF BACTERIA

Most bacteria (singular,bacterium) are very small organisms, on the order of a few micrometers in lenth.it would take about 1,000 bacteria placed end to end to equal one millimetre, which is about the width of a pencil line. In fact, however, bacteria, one um in length, placed end-end to equal one millimetre, which is about the width of a pencil line. Infact, however, bacteria come in a wide variety of shapes and sizes, called the bacillus (plural, bacilli) form, or spherical, called the coccus (plural, cocci) form. The rod forms vary considerable from very short rods that almost look like cocci, to very long filaments thousands of microns in length. Bacteria also form spirals and corkscrews, ovals (coccoid), commas , and elaborately branched structures. The cocci often take on multi-cell forms; as two cocci joined together (diplococci) , as chains of cocci(streptococci), or as tetrads(four cells in a cube).

Bacteria display a wide diversity of shapes and sizes, called morphologies. Bacterial cells are about one tenth the size of eukaryotic cells and are typically 0.5–5.0 micrometres in length. However, a few species — for example, Thiomargarita namibiensis and Epulopiscium fishelsoni — are up to half a millimetre long and are visible to the unaided eye; E. fishelsoni reaches 0.7 mm. Among the smallest bacteria are members of the genus Mycoplasma, which measure only 0.3 micrometres, as small as the largest viruses.[26] Some bacteria may be even smaller, but these ultramicrobacteria are not well-studied.

Most bacterial species are either spherical, called cocci (sing. coccus, from Greek κόκκος-kókkos, grain, seed), or rod-shaped, called bacilli (sing. bacillus, from Latin baculus, stick). Elongation is

associated with swimming. Some rod-shaped bacteria, called vibrio, are slightly curved or comma-shaped; others can be spiral-shaped, called spirilla, or tightly coiled, called spirochaetes. A small number of species even have tetrahedral or cuboidal shapes.More recently, bacteria were discovered deep under the Earth's crust that grow as long rods with a star-shaped cross-section. The large surface area to volume ratio of this morphology may give these bacteria an advantage in nutrient-poor environments. This wide variety of shapes is determined by the bacterial cell wall and cytoskeleton, and is important because it can influence the ability of bacteria to acquire nutrients, attach to surfaces, swim through liquids and escape predators.

Many bacterial species exist simply as single cells, others associate in characteristic patterns: Neisseria form diploids (pairs), Streptococcus form chains, and Staphylococcus group together in "bunch of grapes" clusters. Bacteria can also be elongated to form filaments, for example the Actinobacteria. Filamentous bacteria are often surrounded by a sheath that contains many individual cells. Certain types, such as species of the genus Nocardia, even form complex, branched filaments, similar in appearance to fungal mycelia. Bacteria often attach to surfaces and form dense aggregations called biofilms or bacterial mats. These films can range from a few micrometers in thickness to up to half a meter in depth, and may contain multiple species of bacteria, protists and archaea. Bacteria living in biofilms display a complex arrangement of cells and extracellular components, forming secondary structures such as microcolonies, through which there are networks of channels to enable better diffusion of nutrients. In natural environments, such as soil or the surfaces of plants, the majority of bacteria are bound to surfaces in biofilms.Biofilms are also important in medicine, as these structures are often present during chronic bacterial infections or in infections of implanted medical devices, and bacteria protected within biofilms are much harder to kill than individual isolated bacteria.

Even more complex morphological changes are sometimes possible. For example, when starved of amino acids, Myxobacteria detect surrounding cells in a process known as quorum sensing, migrate towards each other, and aggregate to form fruiting bodies up to 500 micrometres long and containing approximately 100,000 bacterial cells.In these fruiting bodies, the bacteria perform separate tasks; this type of cooperation is a simple type of multicellular organisation. For example, about one in 10 cells migrate to the top of these fruiting bodies and differentiate into a specialised dormant state called myxospores, which are more resistant to drying and other adverse environmental conditions than are ordinary cells

Morphological shapes of bacteria includes:

Rod like Bacilli

Spherical Cocci

Comma shaped Vibrio

Spirulla/Spirochaetes

filamentous There may be Some correlation between morphology and disease Spiral bacteria---Treponemes, Borrelias, Leptospiras, Spirillium tend to cause systemic diseases Pathogenic filamentous bacteria Actinomyces, Nocardia, Mycobacteria tend to cause chronic diseases .Cocci, Staphylococcus more likely to cause skin infections, Streptococci skin and pneumonia

CLASSIFICATION OF BACTERIA BASED ON THEIR METABOLIC BEHAVIOUR This method of classification is based on the metabolic properties of bacterial cells in relation to their environmental factors. Bacteria exhibit an extremely wide variety of metabolic types.The distribution of metabolic traits within a group of bacteria has traditionally been used to define their taxonomy, but these traits often do not correspond with modern genetic classifications. Bacterial metabolism is classified into nutritional groups on the basis of three major criteria: the kind of energy used for growth, the source of carbon, and the electron donors used for growth. An additional criterion of respiratory microorganisms is the electron acceptors used for aerobic or anaerobic respiration.

Carbon metabolism in bacteria is either heterotrophic, where organic carbon compounds are used as carbon sources, or autotrophic, meaning that cellular carbon is obtained by fixing carbon dioxide. Heterotrophic bacteria include parasitic types. Typical autotrophic bacteria are phototrophic

cyanobacteria, green sulfur-bacteria and some purple bacteria, but also many chemolithotrophic species, such as nitrifying or sulfur-oxidising bacteria. Energy metabolism of bacteria is either based on phototrophy, the use of light through photosynthesis, or based on chemotrophy, the use of chemical substances for energy, which are mostly oxidised at the expense of oxygen or alternative electron acceptors (aerobic/anaerobic respiration).

Filaments of photosynthetic cyanobacteria

Finally, bacteria are further divided into lithotrophs that use inorganic electron donors and organotrophs that use organic compounds as electron donors. Chemotrophic organisms use the respective electron donors for energy conservation (by aerobic/anaerobic respiration or fermentation) and biosynthetic reactions (e.g. carbon dioxide fixation), whereas phototrophic organisms use them only for biosynthetic purposes. Respiratory organisms use chemical compounds as a source of energy by taking electrons from the reduced substrate and transferring them to a terminal electron acceptor in a redox reaction. This reaction releases energy that can be used to synthesise ATP and drive metabolism. In aerobic organisms, oxygen is used as the electron acceptor. In anaerobic organisms other inorganic compounds, such as nitrate, sulfate or carbon dioxide are used as electron acceptors. This leads to the ecologically important processes of denitrification, sulfate reduction and acetogenesis, respectively. Another way of life of chemotrophs in the absence of possible electron acceptors is fermentation, where the electrons taken from the reduced substrates are transferred to oxidised intermediates to generate reduced fermentation products (e.g. lactate, ethanol, hydrogen, butyric acid). Fermentation is possible, because the energy content of the substrates is higher than that of the products, which allows the organisms to synthesise ATP and drive their metabolism.

These processes are also important in biological responses to pollution; for example, sulfate-reducing bacteria are largely responsible for the production of the highly toxic forms of mercury (methyl- and dimethylmercury) in the environment. Non-respiratory anaerobes use fermentation to generate energy and reducing power, secreting metabolic by-products (such as ethanol in brewing) as waste. Facultative anaerobes can switch between fermentation and different terminal electron acceptors depending on the environmental conditions in which they find themselves. Lithotrophic bacteria can use inorganic compounds as a source of energy. Common inorganic electron donors are hydrogen, carbon monoxide, ammonia (leading to nitrification), ferrous iron and other reduced metal ions, and several reduced sulfur compounds. Unusually, the gas methane can be used by methanotrophic bacteria as both a source of electrons and a substrate for carbon anabolism. Both aerobic phototrophy and chemolithotrophy, oxygen is used as a terminal electron acceptor, while under anaerobic conditions inorganic compounds are used instead. Most lithotrophic organisms are autotrophic, whereas organotrophic organisms are heterotrophic. In addition to fixing carbon dioxide in photosynthesis, some bacteria also fix nitrogen gas (nitrogen fixation) using the enzyme nitrogenase. This environmentally important trait can be found in bacteria of nearly all the metabolic types listed above, but is not universal. Regardless of the type of metabolic process they employ, the

majority of bacteria are only able to take in raw materials in the form of relatively small molecules, which enter the cell by diffusion or through molecular channels in cell membranes. The Planctomycetes are the exception (as they are in possessing membranes around their nuclear material). It has recently been shown that Gemmata obscuriglobus is able to take in large molecules via a process that in some ways resembles endocytosis, the process used by eukaryotic cells to engulf external item. We also have Types of Bacteria Based on Their Environment

1. Osmophiles - live in high sugar osmotic environments 2. Mesophiles - survive in moderate conditions. 3. Extremophiles - can live in extreme conditions. 4. Neutrophiles - can live in moderate conditions. 5. Thermophiles - can live at high temperatures. 6. Alkaliphiles - can live in a high pH environment. 7. Acidophiles - can live in a low pH environment 8. Halophiles - can live in saline environments. 9. Psychrophilic bacteria - can live in extremely cold environments.

Growth Requirements: Microorganisms can be grouped on the basis of their need for oxygen to grow. Facultatively anaerobic bacteria can grow in high oxygen or low oxygen content and are among the more versatile bacteria. In contrast, strictly anaerobic bacteria grow only in conditions where there is minimal or no oxygen present in the environment. Bacteria such as bacteroides found in the large bowel are examples of anaerobes. Strict aerobes only grow in the presence of significant quantities of oxygen. Pseudomonas aeruginosa, an opportunistic pathogen, is an example of a strict aerobe.

Microaerophilic bacteria grow under conditions of reduced oxygen and sometimes also require increased levels of carbon dioxide. Neisseria species (e.g., the cause of gonorrhea) are examples of micraerophilic bacteria.

GENETIC CLASSIFICATION SYSTEMS

Universal Phylogenetic Tree: Woese has developed a “universal phylogenetic tree” for all living organisms that establishes a tripartite division of all living organisms– bacteria, archaea and eucarya. His work is based on a comparison of 16s ribosomal RNA sequences. These sequences are highly conserved and undergo change at a slow, gradual and consistent rate. They are therefore useful for making comparisons among the different living organisms. Ribosomal RNA (rRNA) sequence analysis: This has emerged as a major method for classification. It has been used (as described above) to establish a phylogenetic tree. In addition, it is now also used to rapidly diagnose the pathogen responsible for an infection, to help select appropriate therapy and to identify noncultivatable microorganisms. Molecular subtyping: Sometimes it is necessary to determine whether strains from the same species are the same or different. For example, if there is an outbreak of infections that appear due to the same bacterial species, the hospital epidemiologist will want to know if all of the infections are due to the same strain. Clues can be obtained by examining the biochemical studies or the antibiotic susceptibility profile, but a more reliable method is by molecular analysis. Pulsed Field Gel Electrophoresis (PFGE) is the most frequently used molecular technique. Chromosomal DNA is digested with a restriction enzyme that makes relatively infrequent cuts in the DNA and as a result creates large DNA fragments. The DNA fragments from the different strains are then run on a gel and compared.

General Phenotypic Classification of Bacteria

HARMFUL AND BENEFICIAL ASPECTS OF BACTERIA

Helpful Types of Bacteria

Although most people are afraid of getting in contact with bacteria, some types are actually helpful to one's health and their presence may be needed in the body to prevent disease. In general most of these bacteria consist of:

Lactobacilli, which are rod-shaped, Gram-positive bacteria normally found in the human intestine, vagina and mouth. Their presence prevents the overgrowth of harmful bacteria in these body parts by producing lactic acid. They are also found in fermented milk products such as yogurt and may be taken as part of a healthy diet to maintain a normal amount of probiotics in the body.

Bifidobacteria are also rod-shaped (branched) and Gram-positive bacteria,and they are similar to lactobacilliin terms of probiotic benefits, including preventing diarrhea and yeast infection.

Escherichia coli are rod-shaped, Gram negative microorganisms which help breakdown undigested sugars in the intestine, thus aiding digestion. They also produce biotin and vitamin K which are important to many cellular processes.

Other helpful bacteria found in the environment include Streptomyces, Rhyzobia, and Cyanobacteria, which help maintain a healthy habitat for man by preventing the proliferation of harmful bacteria

Beneficial Bacteria:

• Help make essential soil mineral elements, available to the plant Nitrogen Fixation

• Decompose organic matter and improve physical properties of the soil

• Vast numbers of bacteria live in our bodies. One example is found in the intestine. This bacteria and

humans have formed a symbiosis with each other. The bacteria help us with digestion and to

produce vitamins. In exchange, they soak up a little extra food for themselves

• Most dairy products are made by or with the help of bacteria. Some dairy foods are cheese,

buttermilk, yogurt, and sour cream. Some other kinds of foods that involve bacteria in their

production are pickles and high fructose corn syrup.

• Bacteria help in the production of fuel in two major ways. Some bacteria decompose compost,

garbage and sewage and help make methane. Methane is a valuable natural gas. It is used widely

as a fuel. Also, over time, the earth's pressure has changed dead and decomposed animals and

plants into coal, which is also a widespread fuel.

• Bacteria also make, or help to make drugs, hormones, or antibodies.

• Bacteria can even help to break down oil to make clean-up after an oil spill easier.

Harmful Types of Bacteria

Some of the harmful types of bacteria are those which can cause disease or adversely affect one's health, and these include:

Mycobacteria, which are rod -shaped, and neither Gram-positive nor Gram-negative microorganisms that can cause infections of the lungs, skin and other parts of the body. The most common diseases associated with these bacteria are leprosyand tuberculosis.

Clostridium tetani are Gram positive rods that infect the skin and gastrointestinal tract, causing tetanus, which can lead to death.

Yersinia pestis are Gram negative rods which infect the skin and lungs, leading to bubonic and pneumonic plague. It is also nowconsidered as a potential biological weapon.

Helicobacter pylori are a common type of bacteria associated with gastric and peptic ulcers. Although almost half the world's population may harbor these harmful bacteria, some do not manifest symptoms.

Bacillus anthracis are Gram-positive rods which occur in many animals like goats, sheep and cattle but may be transmitted to humans, causing abdominal problems including diarrhea.

Effects of Harmful Types of Bacteria

Bacteria can harm animals, plants, humans and even the environment in different ways. Depending on the type of bacteria, they can cause mild to severe infections which may or may not lead to another organism's death. Many pathogenic bacteria can survive different conditions in the environment which can increase their spread through contact with inanimate objects such as telephones, door knobs, toilet seats, and even through water and

air. Bacteria can cause disease by releasing toxins that can harm the host, causing various conditions and diseases like food poisoning or botulism, urinary tract infection, cholera, typhoid fever, and many more. They have also been associated with chronic diseases such as colon cancer.

In the soil bacteria can cause denitrification by breaking down nitrites into ammonia and free nitrogen. This results in reduced soil fertility and productivity. Some bacteria can also damage crops, kill livestock and other farm animals, resulting in direct and indirect harm to man.

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