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Bacteria
Bacteria are microscopic organisms that are less than 3 micrometres (μm) in size. μm) in size. m) in size. Size of cocci range from 0.5 to 3 μm) in size. m, and the size of a rod shaped bacteria range from 0.15 to 2 μm) in size. m (μm) in size. width) to 0.5 to 20 μm) in size. m (μm) in size. length). Bacteria are measured using a calibrated slide under a compound microscope or using an electron microscope.Bacterial morphology deals with size, shape, and arrangement of bacterial cells. The three basic shapes of bacteria are spherical, rod shaped and spiral. Spiral-shaped bacteria can be further categorized depending in part on how much spiraling they show. Not all bacteria are capable of causing disease, but each morphology-based group has at least some disease-causing representatives.
Shape of Bacteria
Coccus Coccus or Cocci are bacterial cells that are spherical, and resemble tiny balls.They may be single bacteria or they may occur in pairs, chains or clusters of bacteria, depending on the bacterium and environmental conditions. Bacillus Bacillus are bacterial cells that are rod shaped, and resemble a pill. Some bacilli have rounded ends while others have squared ends. Spirillum Spiral bacteria have twisted or helical morphology that resembles little corkscrews. They may appear as curved rods, called vibrios, spirilla or spirochetes having pliant bodies. Vibrio Members of the vibrio group represent one of the three types of bacteria that have a spiral shape. Vibrio bacteria are comma-shaped, appearing like curved rods. They typically live in aquatic environments and move in a darting motion using a single flagellum, a whip-like structure. Spirochete
Spirochete are long, thin and flexible corkscrew-shaped bacteria. They typically move in a distinctive rotating manner that allows them to be mobile in mucus-lined tissue or viscous environments.
Shape has selective value
The first issue to get settled is that the shape of a bacterium has biological relevance. One argument favoring this assertion is that even though bacteria have a wide variety of shapes, any one genus typically exhibits a limited subset of morphologies, hinting that, with a universe of shapes to choose from, individual bacteria adopt only those that are adaptive. Another clue is that some bacteria can modify their morphology in response to environmental cues or during the course of pathogenesis, suggesting that shape is important enough to merit regulation.Two evolutionary arguments also support the utility of bacterial shape. First, shape has a vector through evolutionary time – rod-like organisms having arisen first and coccoid forms being derivatives at the ends of evolutionary lines. Progressive
Beneath the external structures is the cell wall. It is very rigid & gives shape to the cell. Its main function is to prevent the cell from expanding & eventually bursting due to water uptake. Cell Wall constitutes a significant portion of the dry weight of the cell and it is essential for bacterial growth & division. The cell wall cannot be seen by direct light microscopy and does not stain with simple stains. It may be demonstrated by microdissection, reaction with specific antibodies, mechanical rupture of the cell, differential staining procedures or by electron microscopy. Chemically the cell wall is composed of peptidoglycan. Mucopeptide (μm) in size. peptidoglycan or murein) formed by N acetyl glucosamine & N acetyl muramic acid alternating in chains, cross linked by peptide chains. Embedded in it are polyalcohol called Teichoic acids. Some are linked to Lipids & called Lipoteichoic acid. Lipotechoic acid link peptidoglycan to cytoplasmic membrane and the peptidoglycan gives rigidity. The functions of Teichoic acid are ● Gives negative charge ● Major antigenic determinant ● Transport ions ● Anchoring ● External permeability barrier characteristics Gram positive Gram negative Thickness Thicker Thinner Variety of amino acids Few Several Lipids Absent Present Theichoic acid Present Absent Fig 3: characteristics of gram positive and gram negative
Nucleus
The Nucleus is not distinct and has no nuclear membrane or nucleolus and the genetic material consist of DNA. The cytoplasmic carriers of genetic information are termed plasmids or episomes
Capsule
Capsule is the outermost layer of the bacteria (μm) in size. extracellular). It is a condensed well defined layer closely surrounding the cell. They are usually polysaccharide and if polysaccharide envelops the whole bacterium it is capsule and their production depends on growth conditions. They are secreted by the cell into the external environment and are highly impermeable. When it forms a loose mesh work of fibrils extending outward from the cell they are described as glycocalyx and when masses of polymer that formed appear to be totally detached from the cell and if the cells are seen entrapped in it are described as slime layer. The Capsule protects against complement and is antiphagocytic. The Slime layer & glycocalyx helps in adherence of bacteria either to themselves forming colonial masses or to surfaces in their environment and they resists phagocytosis and desiccation of bacteria.
Flagella
Flagella are long hair like helical filaments extending from cytoplasmic membrane to exterior of the cell. Flagellin is highly antigenic and functions in cell motility. The location of the flagella depends on bacterial species as polar situated at one or both ends which swims in back and forth fashion and lateral at along the sides. The parts of flagella are the filament, hook and the basal body. Filament is external to cell wall and is connected to the hook at cell surface, the hook & basal body are embedded in the cell envelope. Hook & filament is composed of protein subunits called as flagellin. Flagellin is synthesized within the cell and passes through the hollow centre of flagella. The arrangement of flagella may be described as (μm) in size. i) Monotrichous – single flagella on one side (μm) in size. ii) Lophotrichous – tuft of flagella on one side (μm) in size. iii) Amphitrichous – single or tuft on both sides (μm) in size. iv) Peritrichous – surrounded by lateral flagella
Some bacteria have the ability to form highly resistant resting stage called spores, which helps them to overcome adverse environmental conditions that are unfavorable for vegetative growth of cell. They are not a reproductive form and not a storage granule. These spores are resistant to bactericidal agents and adverse physical conditions. Each spore can give rise to only one endospore which play a role in heat resistance. Spores consists of three layers namely core, cortex and spore coat.
Gram staining
Gram staining was discovered by Christian Gram in 1884. He found that by using dyes the microorganisms could be more readily seen under a microscope and hence know the identity, the morphology and arrangement of the bacteria.There are 2 types of stains: Gram positive and Gram negative.The procedure is based on the ability of microorganisms to retain color of the stains used during the gram stain reaction. Gram-negative bacteria are decolorized by the alcohol,losing the color of the primary stain, purple. Gram-positive bacteria are not decolorized by alcohol and will remain as purple. After decolorization step, a counterstain is used to impart a pink color to the decolorized gram-negative organisms.
Figure:Bacterial cell wall
Gram Stain Mechanism:
o Gram Positive Cell Wall:
Gram-positive bacteria have a thick mesh-like cell wall which is made up of peptidoglycan (μm) in size. 50-90% of cell wall), which stains purple. As adjacent layers of peptidoglycan are formed, they are cross linked by short chains of peptides by means of a transpeptidase enzyme, resulting in the shape and rigidity of the cell wall.The thick peptidoglycan layer of Gram-positive allows these organisms to retain the crystal violet-iodine complex and stains the cells as purple. Lipoteichoic acid (μm) in size. LTA) is another major constituent of the cell wall of Gram- positive bacteria which is embedded in the peptidoglycan layer.
Iodine (μm) in size. I – or I3 –) acts as a mordant and as a trapping agent. A mordant is a substance that increases the affinity of the cell wall for a stain by binding to the primary stain, thus forming an insoluble complex which gets trapped in the cell wall. In the Gram stain reaction, the crystal violet and iodine form an insoluble complex (μm) in size. CV-I) which serves to turn the smear a dark purple color. At this stage, all cells will turn purple. 3) Decolorization with 95% ethyl alcohol. Alcohol or acetone dissolves the lipid outer membrane of Gram negative bacteria, thus leaving the peptidoglycan layer exposed and increases the porosity of the cell wall. The CV-I complex is then washed away from the thin peptidoglycan layer, leaving Gram negative bacteria colorless. On the other hand, alcohol has a dehydrating effect on the cell walls of Gram positive bacteria which causes the pores of the cell wall to shrink. The CV-I complex gets tightly bound into the multi-layered, highly cross-linked Gram positive cell wall thus staining the cells purple. The decolorization step must be performed carefully, otherwise over- decolorization may occur. This step is critical and must be timed correctly otherwise the crystal violet stain will be removed from the Gram-positive cells. If the decolorizing agent is applied on the cell for too long time , the Gram-positive organisms to appear Gram-negative. Under-decolorization occurs when the alcohol is not left on long enough to wash out the CV-I complex from the Gram-negative cells, resulting in Gram-negative bacteria to appear Gram-positive. 4) Counterstain with Safranin
The decolorized Gram negative cells can be rendered visible with a suitable counterstain, which is usually positively charged safranin, which stains them pink. Pink colour which adheres to the Gram positive bacteria is masked by the purple of the crystal violet. Table: Colour changes that occur at each step in the staining process
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