Microbiology Fundamentals

Microbiology is the study of microorganisms — bacteria, viruses, fungi, protozoa, and helminths — that are too small to be seen with the naked eye. For health science students, microbiology provides the foundation for understanding infectious disease pathogenesis, antimicrobial therapy, and infection control.

Bacterial Structure

Bacteria are prokaryotes with a cell wall that is the primary target of many antibiotics. The bacterial cell wall is composed of peptidoglycan (murein) — a polymer of N-acetylglucosamine (NAG) and N-acetylmuramic acid (NAM) cross-linked by short peptide chains. The structure and composition of the cell wall classifies bacteria into two major groups by Gram staining:

Gram-positive bacteria have a thick (20–80 nm) peptidoglycan layer outside the plasma membrane. The crystal violet-iodine complex is retained during decolourisation, staining cells purple. Examples: Staphylococcus aureus (skin infections, endocarditis, toxic shock syndrome; MRSA from mecA gene encoding PBP2a with altered β-lactam affinity), Streptococcus pyogenes (pharyngitis, rheumatic fever, necrotising fasciitis), Streptococcus pneumoniae (pneumonia, meningitis, otitis media), Clostridium (C. difficile — pseudomembranous colitis from antibiotic-disrupted microbiome; C. perfringens — gas gangrene; C. botulinum — botulism; C. tetani — tetanus).

Gram-negative bacteria have a thin (2–7 nm) peptidoglycan layer sandwiched between the inner (cytoplasmic) membrane and an outer membrane (OM). The OM contains lipopolysaccharide (LPS) on its outer leaflet. LPS consists of lipid A (the toxic moiety — activates TLR4/MD2 on macrophages → NFκB activation → TNF-α, IL-1, IL-6 release → fever, hypotension, DIC in septic shock), the core polysaccharide, and the O-antigen (variable, species-specific). The alcohol/acetone decolouriser removes lipids from the thin peptidoglycan layer, allowing crystal violet to wash out; subsequent safranin counterstain gives Gram-negative bacteria a pink/red colour. Examples: Escherichia coli (UTI, neonatal meningitis, traveller's diarrhoea — ETEC), Neisseria meningitidis (meningitis, septicaemia — needs capsule for virulence), Klebsiella pneumoniae (hospital-acquired pneumonia, UTI), Pseudomonas aeruginosa (cystic fibrosis lung infections, burns, nosocomial infections), Helicobacter pylori (peptic ulcer disease, gastric cancer — urease produces NH₃ to buffer stomach acid), Haemophilus influenzae, Salmonella, Shigella.

Acid-fast bacteria (Ziehl-Neelsen stain): Mycobacteria (M. tuberculosis, M. leprae, non-tuberculous mycobacteria) have a cell wall with a thick layer of mycolic acids (long-chain fatty acids) that resist decolourisation with acid-alcohol. The Ziehl-Neelsen stain uses carbol-fuchsin (red), decolourises with acid-alcohol, and counterstains with methylene blue — acid-fast organisms appear red.

Other notable bacterial structures: Capsule — polysaccharide coat outside the cell wall; antiphagocytic (resists opsonisation by macrophages); important virulence factor in S. pneumoniae, N. meningitidis, Klebsiella. Flagella — helical protein appendages (flagellin, TLR5 ligand) for motility; H antigen in E. coli serotyping. Pili/fimbriae — adhesins for host cell attachment (e.g., type I pili of E. coli bind mannosylated proteins in the urinary tract). Spores — endospores (Bacillus, Clostridium) are metabolically dormant, resistant to heat, desiccation, and disinfectants; activated by return of favourable conditions.

Bacterial Reproduction and Genetics

Bacteria reproduce asexually by binary fission — each cell divides into two identical daughters. Generation time varies: E. coli ~20 min, M. tuberculosis ~20 hours. Bacterial growth follows four phases: lag (adaptation, no increase in number), exponential/log (rapid doubling), stationary (nutrient depletion, growth = death), and decline/death.

Genetic variation in bacteria arises from mutation, transformation (uptake of naked DNA from the environment; basis of pneumococcal competence; Griffith's experiment), transduction (bacteriophage-mediated DNA transfer), and conjugation (direct cell-to-cell DNA transfer via pili and plasmids — major mechanism of horizontal antibiotic resistance gene spread; plasmids carrying β-lactamases, aminoglycoside-modifying enzymes, ESBL genes). Integrons, transposons, and pathogenicity islands facilitate acquisition and transfer of virulence and resistance genes.

Viral Structure and Replication

Viruses are obligate intracellular parasites — they lack ribosomes, cannot synthesise their own proteins or generate ATP, and depend entirely on host cell machinery for replication. A virus particle (virion) consists of a nucleic acid genome (DNA or RNA, single- or double-stranded, segmented or unsegmented), enclosed in a protein coat called the capsid (made of protomeric subunits called capsomers arranged with icosahedral or helical symmetry). Some viruses have a lipid envelope derived from the host cell membrane during budding; enveloped viruses bear viral glycoproteins (e.g., HIV gp120/gp41, influenza haemagglutinin/neuraminidase, SARS-CoV-2 spike) that mediate receptor binding and membrane fusion. Non-enveloped (naked) viruses are generally more resistant to desiccation and lipid solvents (e.g., norovirus, adenovirus, poliovirus).

Viral replication cycle (Baltimore classification provides a framework): (1) Attachment: viral surface protein binds specific host cell receptor — tropism is determined here. HIV gp120 binds CD4 + CCR5/CXCR4 (T cells, macrophages); influenza HA binds sialic acid (respiratory epithelium); SARS-CoV-2 spike binds ACE2 (respiratory, GI epithelium). (2) Entry: receptor-mediated endocytosis or direct membrane fusion (enveloped) or receptor-mediated pore formation (non-enveloped). (3) Uncoating: release of viral nucleic acid into the cytoplasm or nucleus. (4) Replication and transcription: varies by genome type. DNA viruses (except poxviruses) replicate in the nucleus using host DNA polymerase (herpesviruses also encode their own); RNA viruses replicate in the cytoplasm using virally encoded RNA-dependent RNA polymerase (RdRp) — target of remdesivir (COVID-19), sofosbuvir (hepatitis C). Retroviruses (HIV) reverse-transcribe their RNA genome to DNA (via reverse transcriptase — targeted by NRTIs, NNRTIs), then integrate into the host genome as a provirus (integrase — targeted by raltegravir, dolutegravir). (5) Assembly: viral components synthesised by host ribosomes are assembled into new virions. (6) Release: by lysis (non-enveloped) or budding (enveloped — influenza neuraminidase cleaves sialic acid to allow release; targeted by oseltamivir/zanamivir).

Fungal Pathogens

Fungi are eukaryotes with ergosterol in their cell membranes (instead of cholesterol) — the basis of antifungal drug selectivity. Azoles (fluconazole, itraconazole, voriconazole) inhibit CYP51 (lanosterol 14α-demethylase), blocking ergosterol synthesis. Polyenes (amphotericin B, nystatin) bind ergosterol, forming membrane pores → cell lysis. Echinocandins (caspofungin, micafungin) inhibit β-1,3-glucan synthase → weakened cell wall.

Clinically important fungi: Candida albicans (oral thrush, oesophagitis in immunocompromised; vaginal candidiasis; invasive candidiasis in ICU — biofilm on central lines); Aspergillus fumigatus (invasive pulmonary aspergillosis in neutropenic patients/stem cell transplant recipients — classically lung cavities with "halo sign" on CT; treated with voriconazole); Cryptococcus neoformans (meningitis in HIV/AIDS patients — diagnosed by India ink CSF preparation showing encapsulated yeast; antigen detected in CSF/blood); Pneumocystis jirovecii (PCP pneumonia in HIV patients with CD4 <200 — bilateral perihilar ground-glass opacification on CXR; treated with co-trimoxazole); Histoplasma capsulatum, Coccidioides immitis (dimorphic fungi — endemic mycoses in specific geographic regions).