Tamil Nadu HSE Second Year Microbiology cheat sheet
Tamil Nadu HSE Second Year Microbiology · Keep the key ideas within reach.
History, Scope and Classification of Microbiology
Anton van Leeuwenhoek (1670s) first observed live microorganisms using single-lens microscopes; called them 'animalcules'.
Louis Pasteur disproved spontaneous generation by swan-neck flask experiments (1861) and developed pasteurization and vaccines against anthrax and rabies.
Robert Koch established the germ theory of disease and postulates: organism must be found in all diseased hosts, be isolated in pure culture, cause disease when inoculated, and be re-isolated.
Alexander Fleming discovered penicillin from Penicillium notatum (1928); Florey and Chain developed it for clinical use.
Carl Woese (1977) used 16S rRNA sequencing to divide prokaryotes into Bacteria and Archaea, giving the three-domain system.
Five-kingdom classification (Whittaker, 1969): Monera, Protista, Fungi, Plantae, Animalia.
Binomial nomenclature by Carolus Linnaeus: genus capitalized, species lowercase, both italicized or underlined separately.
Koch's postulates fail for obligate intracellular pathogens, asymptomatic carriers, and organisms not cultivable in vitro.
Microbial Cell Structure
Prokaryotic cell lacks membrane-bound nucleus and organelles; DNA is a single circular chromosome in the nucleoid region.
Bacterial cell wall peptidoglycan: N-acetylglucosamine (NAG) and N-acetylmuramic acid (NAM) cross-linked by peptide bridges.
Gram-positive wall: thick peptidoglycan (20-80 nm) with teichoic acids; Gram-negative wall: thin peptidoglycan plus outer membrane with lipopolysaccharide (LPS).
LPS lipid A is the endotoxin; O-antigen is the somatic antigen used in serotyping.
Bacterial ribosome is 70S (50S + 30S); eukaryotic is 80S - basis for selective antibiotic action.
Flagellar arrangement types: monotrichous, amphitrichous, lophotrichous, peritrichous; flagellin protein is H antigen.
Bacterial endospore is formed by Bacillus and Clostridium; highly resistant to heat and chemicals due to calcium dipicolinate and dehydrated core.
Pili (fimbriae) mediate adhesion; sex pilus (F pilus) mediates conjugation.
Morphology and Staining
Gram stain steps: crystal violet (1 min) → Gram's iodine mordant → decolourize with 95% ethanol/acetone → counterstain safranin.
Gram-positive cells retain crystal violet-iodine complex (purple); Gram-negative cells decolourize and take safranin (pink/red).
Ziehl-Neelsen acid-fast stain: carbol fuchsin with heat → decolourize with 20% H2SO4 or acid-alcohol → counterstain methylene blue; Mycobacteria appear red.
Spore stain (Schaeffer-Fulton): malachite green with steam, counterstain safranin; spores green, vegetative cells red.
Capsule demonstration: negative staining (India ink) or Anthony's method; capsule appears as clear halo.
Common bacterial shapes: coccus, bacillus, spirillum, spirochaete, vibrio; arrangements: diplo-, strepto-, staphylo-, tetrad, sarcina.
Smear preparation: thin smear air-dried and heat-fixed before staining to attach cells to slide without distorting morphology.
Resolution of light microscope ≈ 0.2 µm; oil immersion (100×) objective increases numerical aperture and resolution.
Microbial Nutrition, Growth and Culture Techniques
Nutritional categories by carbon and energy source: photoautotroph, photoheterotroph, chemoautotroph, chemoheterotroph.
Macronutrients: C, N, P, S, K, Mg, Ca, Fe; micronutrients (trace elements): Zn, Mn, Mo, Cu, Co, Ni.
Growth curve phases: lag → log (exponential) → stationary → death (decline).
Generation time = time for population to double; in log phase N = N0 × 2^n where n = number of generations.
Batch culture is a closed system; continuous culture (chemostat) maintains cells in log phase with steady nutrient supply.
Selective media inhibit unwanted organisms (e.g., MacConkey for Gram-negatives); differential media distinguish colonies by biochemical reaction (e.g., blood agar for haemolysis).
Enrichment culture increases the proportion of a desired organism before isolation.
Obligate anaerobes require anaerobic jar or anaerobic glove box; microaerophiles need reduced O2 (5-10%).
Sterilization, Disinfection and Antimicrobial Agents
Sterilization kills or removes all microorganisms including spores; disinfection reduces pathogens on inanimate objects but not necessarily spores.
Autoclave: 121°C at 15 psi (103 kPa) for 15-20 min; principle is moist heat denaturing proteins.
Hot air oven: 160-170°C for 2 h for dry glassware and metal instruments.
Pasteurization: 63°C for 30 min (holder method) or 72°C for 15 s (flash/HTST); kills pathogens but not spores.
Membrane filtration (0.22 µm) sterilizes heat-labile liquids like sera and antibiotics.
Ethylene oxide gas sterilization is used for heat-sensitive plastics and disposable devices.
Antibiotics act on cell wall (penicillin, cephalosporin), protein synthesis (tetracycline, streptomycin, chloramphenicol), nucleic acid (rifampicin, quinolones), or folate pathway (sulfonamides).
Minimum inhibitory concentration (MIC) is the lowest drug concentration inhibiting visible growth; determined by broth or agar dilution.
Microbial Genetics, Mutation and Genetic Engineering
Bacterial genome is a single circular double-stranded DNA chromosome; plasmids are extrachromosomal, autonomously replicating DNA molecules.
Transformation: uptake of naked DNA by competent cells (Griffith, Avery-MacLeod-McCarty).
Conjugation: transfer of DNA through a sex pilus from F+ to F− cells; Hfr strains transfer chromosomal DNA at high frequency.
Transduction: bacteriophage-mediated DNA transfer; generalized transduction transfers any gene, specialized transfers specific genes.
Mutation types: point (substitution), frameshift (insertion/deletion), silent, missense, nonsense; spontaneous vs induced by mutagens (UV, alkylating agents, base analogues).
Ames test uses his− Salmonella to detect mutagens by reversion to histidine prototrophy.
Restriction endonucleases (e.g., EcoRI) cut DNA at specific palindromic sequences; ligase joins fragments to form recombinant DNA.
PCR steps: denaturation (94-95°C), annealing (50-60°C), extension (72°C) using Taq polymerase.
Immunology: Host Defence, Antigens, Antibodies and Vaccines
Innate immunity: physical barriers, phagocytes, complement, inflammation; non-specific and no memory.
Adaptive immunity: B lymphocytes produce antibodies (humoral); T lymphocytes mediate cell-mediated immunity; both show memory and specificity.
Antibody classes: IgG (most abundant, crosses placenta), IgM (primary response, pentamer), IgA (secretions), IgE (allergy, mast cells), IgD (B-cell receptor).
Antigen: any substance that induces an immune response and reacts specifically with antibody; epitope is the antigenic determinant.
Antibody structure: two heavy and two light chains; Fab region binds antigen, Fc region mediates effector functions.
Primary response is slow with IgM; secondary response is rapid with high IgG due to memory cells.
Vaccine types: live attenuated (BCG, MMR), killed (Salk IPV, pertussis), toxoid (tetanus, diphtheria), subunit/conjugate (Hib, hepatitis B), mRNA.
Hypersensitivity types I (IgE anaphylaxis), II (cytotoxic), III (immune complex), IV (delayed, T-cell mediated).
Medical Microbiology and Communicable Diseases
Koch's postulates guide identification of causative agents; exceptions include carriers and uncultivable organisms.
Mycobacterium tuberculosis: acid-fast bacillus, Ziehl-Neelsen stain, Lowenstein-Jensen medium; causes tuberculosis.
Vibrio cholerae: comma-shaped Gram-negative bacillus, rice-water stools, transmitted by contaminated water; treated with oral rehydration.
Salmonella typhi causes typhoid fever; Widal test detects antibodies; blood culture is definitive in first week.
Streptococcus pyogenes: beta-haemolytic, bacitracin-sensitive; causes pharyngitis, scarlet fever, rheumatic fever.
HIV targets CD4+ T cells; ELISA screening confirmed by Western blot or nucleic acid test.
Malaria caused by Plasmodium species; Anopheles mosquito vector; diagnosed by blood smear.
Nosocomial infections often involve Staphylococcus aureus, Pseudomonas aeruginosa, and E. coli; multidrug resistance is a major concern.
Environmental and Agricultural Microbiology
Nitrogen cycle: nitrogen fixation (Rhizobium, Azotobacter, cyanobacteria), nitrification (Nitrosomonas, Nitrobacter), denitrification (Pseudomonas).
Rhizobium forms root nodules in legumes; symbiotic nitrogen fixation via nitrogenase enzyme.
Biogeochemical cycles: carbon (photosynthesis, respiration, decomposition), nitrogen, sulphur, phosphorus.
BOD (biochemical oxygen demand) measures organic pollution; high BOD indicates heavy organic load.
Indicator organisms of faecal contamination: coliforms such as E. coli; MPN test estimates coliform density.
Bioremediation uses microorganisms to degrade pollutants; activated sludge and trickling filter are sewage treatment methods.
Biofertilizers include Rhizobium, Azotobacter, Azospirillum, and phosphate-solubilizing bacteria.
Mycorrhizae (VAM) enhance phosphorus uptake in plants.
Industrial and Food Microbiology
Fermentation products: ethanol (Saccharomyces cerevisiae), citric acid (Aspergillus niger), penicillin (Penicillium chrysogenum), glutamic acid (Corynebacterium glutamicum).
Industrial fermentation types: batch, fed-batch, continuous; stirred-tank bioreactor is most common.
Food spoilage organisms: Pseudomonas, Clostridium, Bacillus, moulds; refrigeration slows but does not eliminate growth.
Food preservation methods: canning (commercial sterility), drying, salting, freezing, irradiation, addition of preservatives.
Food-borne pathogens: Salmonella, Staphylococcus aureus, Clostridium botulinum, Listeria monocytogenes, E. coli O157:H7.
Yoghurt production uses Lactobacillus bulgaricus and Streptococcus thermophilus; cheese uses rennet and starter cultures.
Single-cell protein (SCP) from microorganisms such as Spirulina and yeast serves as protein supplement.
Antibiotic production uses submerged fermentation with strain improvement and downstream processing.
Laboratory Practice, Safety and Applied Skills
Biosafety levels: BSL-1 (low risk), BSL-2 (moderate, most clinical labs), BSL-3 (serious, TB), BSL-4 (dangerous, Ebola).
Personal protective equipment: lab coat, gloves, goggles; no eating, drinking, or mouth pipetting in the lab.
Autoclave validation uses biological indicator Geobacillus stearothermophilus spores.
Biological safety cabinet (Class II) provides personnel, product, and environmental protection for infectious aerosols.
Standard precautions treat all clinical specimens as potentially infectious.
Sharps disposal in puncture-resistant containers; spills decontaminated with 0.5-1% sodium hypochlorite.
Media preparation: weigh ingredients, dissolve, adjust pH, dispense, autoclave; store at 4°C.
Record keeping: label plates with date, specimen, medium; incubate at appropriate temperature and atmosphere.
Immunology: Host Defence, Antigens, Antibodies and Vaccines
Innate immunity includes physical barriers, phagocytes, complement, and inflammation; it is rapid and non-specific.
Adaptive immunity involves B and T lymphocytes with specificity and memory.
IgG is the most abundant antibody and crosses the placenta; IgM is the first antibody in primary response.
Antigen-antibody reactions: agglutination, precipitation, complement fixation, ELISA, immunofluorescence.
Vaccines: live attenuated, killed, toxoid, subunit, conjugate, and mRNA.
Herd immunity requires a sufficient proportion of the population to be immune to interrupt transmission.
Hypersensitivity: type I IgE-mediated, type II cytotoxic, type III immune complex, type IV delayed T-cell mediated.
Adjuvants enhance immune response to antigens in vaccines.
Environmental and Agricultural Microbiology
Nitrogen fixation by Rhizobium, Azotobacter, and cyanobacteria converts atmospheric N2 to ammonia.
Nitrification by Nitrosomonas and Nitrobacter converts ammonia to nitrate.
Denitrification by Pseudomonas returns nitrogen to the atmosphere as N2.
BOD measures organic pollution; high BOD indicates heavy organic load.
Coliforms are indicator organisms for faecal contamination; MPN test estimates their density.
Bioremediation uses microorganisms to degrade pollutants.
Biofertilizers include Rhizobium, Azotobacter, Azospirillum, and phosphate-solubilizing bacteria.
Mycorrhizae enhance phosphorus uptake in plants.
Industrial and Food Microbiology
Fermentation products: ethanol (Saccharomyces cerevisiae), citric acid (Aspergillus niger), penicillin (Penicillium chrysogenum), glutamic acid (Corynebacterium glutamicum).
Industrial fermentation types: batch, fed-batch, continuous; stirred-tank bioreactor is most common.
Food spoilage organisms: Pseudomonas, Clostridium, Bacillus, moulds.
Food preservation methods: canning, drying, salting, freezing, irradiation, preservatives.
Food-borne pathogens: Salmonella, Staphylococcus aureus, Clostridium botulinum, Listeria monocytogenes, E. coli O157:H7.
Yoghurt production uses Lactobacillus bulgaricus and Streptococcus thermophilus.
Single-cell protein (SCP) from Spirulina and yeast serves as protein supplement.
Antibiotic production uses submerged fermentation with strain improvement and downstream processing.
Laboratory Practice, Safety and Applied Skills
Biosafety levels: BSL-1 low risk, BSL-2 moderate, BSL-3 serious, BSL-4 dangerous.
PPE: lab coat, gloves, goggles; no eating, drinking, or mouth pipetting in the lab.
Autoclave validation uses biological indicator Geobacillus stearothermophilus spores.
Class II biological safety cabinet provides personnel, product, and environmental protection.
Standard precautions treat all clinical specimens as potentially infectious.
Sharps disposal in puncture-resistant containers; spills decontaminated with 0.5-1% sodium hypochlorite.
Media preparation: weigh ingredients, dissolve, adjust pH, dispense, autoclave; store at 4°C.
Record keeping: label plates with date, specimen, medium; incubate at appropriate temperature and atmosphere.