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  1. NEWAI (RAJASTHAN) 1st Assignment Of Pharmaceutical Microbiology Submitted To : Submitted By : Dr.Omveer Singh. Ankit Kumar Mishra Assistant Professor B.Pharma 3rd Semester ERP ID – 02109BPH010 Roll No. – 2021BPHR010
  2. •Ultra Structure of Bacteria Introduction: • Ultra Structure of Bacteria components: • Capsule: • Cell wall: • Flagella: • Fimbriae and Pili: • Spinae: • Cell Membrane: • Inclusion bodies: • Ribosomes: • Nucleoid: • Mesosome: • Differences Between Gram-Positive and Gram-Negative Bacteria
  3. Under an electron microscope, the structure of the bacterial cell looks like a capsule. Ultra Structure of Bacteria has the following components like outside the cell membrane and internal to the cell membrane. Capsule: Capsule: It is the outer layer of the bacteria cell. Depending on the chemical nature capsules are thick or thin, and rigid or flexible. It is the most important virulence factor of bacteria which is visualized by the negative staining technique. Generally, it is composed of 98% water and 2% polysaccharide or glycoprotein/polypeptide or both. Only some bacterial species possess capsule. The capsule is usually made of polysaccharides occasionally polypeptide, and hyaluronic acid. Most bacterial capsules are composed of polysaccharides especially homopolysaccharides which are usually synthesized outside the cell from disaccharides by an exocellular enzyme. The capsule of acetic acid bacteria cell is composed of homopolysaccharide (hemicellulose).
  4. There are two types of capsules: 1. Macrocapsule: Thickness of 0.2 µm or more, visible under a light microscope. 2. Microcapsule: Thickness is less than 0.2 µm, visible under Electron microscope. iFunctions: ons: 1. Preventive nature: Capsular polysaccharide bind significant amount of water making cell resistant to drying and also prevent attachment of bacteriophage on the cell surface. 2. Protection: It protects from mechanical injury, temperature, drying, etc. 3. Attachment: Capsule helps in attachment on the surface. For example, Streptococcus mutants that cause dental carries attach on teeth surface by its capsule. 4. Anti-phagocytic: Capsule resists phagocytosis by WBCs. 5. Source of nutrition: It acts as a resource organ of nutrition during insufficient nutrient supply. 6. Repulsion: Same charge capsulated bacteria repel each other.
  5. Cell wall: Cell wall is the rigid wall situated outside of the plasma membrane. It provides shape of the bacteria as well as provides protection from osmotic lysis. Cell wall is made up of peptidoglycan. The gram positive bacteria contains thick homogeneous peptidoglycan or murein layer (20 to 80 nm thickness) outside the plasma membrane whereas gram negative bacteria has 2-7 nm peptidoglycan layer surrounded by 7-8 nm thick outer membrane. Flagella: Flagella are slender and rigid hair like structures. They are about 20 nm cross and up-to 15 to 20 µm long. The eukaryotic flagellum is a long, rod-like structure, surrounded by an extension of the cell membrane like a sheath. The bulk of the structure is a filament called an axoneme. Flagellar Ultra-Structure: Transmission electron microscope studies revealed three regions of flagella which are: (1) The outermost region is filament which is extended from the cell surface to tips. (2) Basal bodies consist of small central rods inserted into cells (3) A short curved segment, the flagellar hook which links the filament to basal bodies and acts as flexible coupling.
  6. The filament is a hollow, rigid cylinder made up of protein subunits flagellin. The molecular weight is varies from 30,000 dalton to 6,000 dalton depends on the bacterial species. Cell Membrane: Cell or plasma membrane is a thin structure that completely surrounds the cell. It is about 8 nm thick. This structure is critical barrier separating the inside of cell from environment. The cell membrane is also highly selective barrier enabling the cell to concentrate a specific metabolite and excrete waste material. Mostly biological membrane is composed primarily of Phospholipids (about 20 to 30 per cent) and proteins (about 60 to 70 per cent). Ribosomes: Ribisomes are loosely attached to the plasma membrane (70S ribosomes). They are made up of both protein and ribonucleic acid. They are the site of protein synthesis. Plasma membrane ribosomes make proteins for transport to the outside. Nucleoid: Prokaryotic chromosome is located in an irregularly shaped region called nucleoid. Nucleoid is composed of 60% DNA, 30% RNA and 10% protein by weight. Prokaryotes contain single circle of double stranded DNA but some are have linear DNA chrosome.
  7. Mesosome: Mesosomes are present in the cell membrane of bacterial cells that fold inward. They play a major role in cellular respiration.
  8. Gram-Positive Gram-Negative 1. The cell wall is made up of a thick peptidoglycan layer 1. They have a thin lipopolysaccharide exterior cell wall. 2. Effect on dyes: Retain the crystal violet dye, and change it into purple during staining identification. 2. Effect on dyes: Do not retain the crystal violet dye, and react only with a counter-stain, generally stain pink. 3. Effect on antibiotics: Susceptible to the enzyme lysozyme and penicillin 3. Effect on antibiotics: Resistant to penicillin and contain an endotoxin called Lipopolysaccharide. 4. The flagellum has two supporting rings, in the peptidoglycan layer, and the plasma membrane. 4. If present, the flagellum has four supporting rings, namely the ‘L’ ring, ‘P’ ring, ‘M’ ring, and ‘S’ ring. 5. Teichoic acids are present. 5. Teichoic acids are absent. 6. Lipoproteins are absent. 6. Lipoproteins are attached to the polysaccharide backbone. 7. Periplasmic space is absent. 7. Periplasmic space is present.
  9. Morphological Classification of Bacteria Morphological Classification of Bacteria: As per the morphological basis, bacteria are classified into six major groups which are depicted as follows: True bacteria: They are a major group of classification which is further divided into two sub-groups namely Cocci and Bacilli. Cocci are of various types. Like Mono cocci (single cocci): Monococcus species, Diplococci (Cocci in pairs): Streptococcus pneumonia, Staphylococci
  10. They are rigid organisms but look like fungi. They exhibit branching and tend to form filaments. They are Gram-positive, but several species have complex cell wall structures that make the Gram staining unsuitable (e.g. Mycobacteriaceae). They are widely distributed in soil, compost, etc. Actinomycetes ::
  11. Spirochaetes: They are double-membrane bacteria, most of which have long, non-branched, helically coiled cells. They are Gram-negative and are chemoheterotrophic in nature. The length varies between 3 and 500 µm and diameters around 0.09 to 3 µm. Rickettsia: They are non-motile, Gram-negative, non-spore-forming, small intracellular, and pleomorphic bacteria. They are also present as cocci (0.1 µm in diameter), rods (1-4 µm long), or thread-like (10 µm long) forms. Chlamydiae: They are ovoid and Gram-negative bacteria. Their cell wall is consisting of peptidoglycan and other proteins. They are very small, obligate intracellular parasites. They multiply in the cytoplasm of their host cell by a distinctive developmental cycle. Culture Media Culture media are the base where microorganisms can grow with the help of nutrients and under the influence of physical growth parameters. All microorganisms cannot grow in a single culture medium and many cannot grow in any known culture medium. Some organisms that cannot grow in the artificial culture medium are known as obligate parasites.
  12. Types of Cultural Media:
  13. Raw materials for culture Media Raw materials play an important role in media preparation. The quality of media depends on the quality of raw materials. The most important raw materials are used for preparations of media are water, agar, peptone, casein hydrolysate, meat extract, yeast extract, and malt extract. Water as solvent: Water plays a vital role in the preparation of media. Water should be free from copper ion because copper ion inhibits the growth of microorganisms. Petri dishes: Petri dishes are generally sterilized with ethylene oxide (EtO) or gamma-irradiated. Energy source: The most common substance added to culture media is glucose that acts as a source of energy and also increases the rate of growth of organisms.
  14. Nutrients: The nutrients of culture media are selected to recover the required spectrum of organisms in the sample e.g. coliforms or anaerobes. Proper nutrients help for bacterial growth. Indicator Substances: The addition of colored indicator substances in the medium is very effective in detecting the fermentation of specific carbohydrates. They change color distinctly and rapidly at critical pH values. Nutritional Requirements for Culture Media Most of the microorganisms are grown in specific culture media in laboratory conditions and for proper growth, nutrition plays an important role in the structural and functional properties of the cells. Nutrition is generally composed of: •Major macronutrients: C, H, O, N, S, P, K, Mg, Fe, Co, and Mn. •Major micronutrients: Zn, Co, Cu, and Mo. •Carbon and energy sources. •Growth factors. •Vitamins.