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Comprehensive Study Notes for Microbiology 112

Chapter 17: ID Pathogens and Diagnosing Infections

  • Overview of diagnostic workflow

    • Patient presentation with signs and symptoms guides presumptive diagnosis
    • Multiple diagnostic modalities used: direct microscopy, culture, biochemical tests, molecular tests, immunologic tests, and in vivo methods

  • Specimen collection and handling (Table 17.1 summaries)

    • Key principle: use aseptic technique; minimize contamination; transport promptly; some specimens require anaerobic conditions
    • Common specimen types and collection methods:
    • Abscess or skin/membrane ulcers: debridement and aspirate fluid; transfer to anaerobic transport; avoid air exposure
    • Blood: inoculate blood culture bottles immediately; avoid air
    • Bone marrow: aspirate; sterile handling; transport promptly
    • CSF: collect via lumbar puncture; sterile tubes; do not refrigerate; transport immediately
    • Feces: collect small specimen; transport promptly; use special kits for protozoa/ova
    • Genital/urinary tract: clean external genitalia; midstream urine; catheter samples if indicated
    • Respiratory tract specimens: sputum, throat swabs; induced if needed
    • Others: nasopharynx, nasopharyngeal swabs, etc.
    • Sampling notes and tips
    • Aspirates preferred over swabs for many deep infections
    • Use appropriate transport media; anaerobic if needed
    • Brushing teeth and rinsing mouth before sampling when the oral cavity is involved
    • For bladder infections, avoid contamination from urethra; use catheter when appropriate
    • Avoid normal flora contamination to improve diagnostic accuracy

  • Phenotypic methods (Chapter 17)

    • Cultivation of specimens
    • Requires knowledge of growth requirements and selective media
    • May use media that inhibit normal microbiota to reveal pathogens (example: Listeria monocytogenes selective media with multiple selective agents)
    • Large pathogen load can reveal characteristics by colony appearance, odor, antibiotic resistance, and microscopy
    • Immediate direct examination
    • Microscopy with differential and special stains
    • Gram staining: differential based on cell wall structure
    • Acid-fast staining: for acid-fast organisms resistant to standard Gram stain
    • Stains for structural features: capsule, flagella, spore
    • Direct fluorescent antibody (DFA) testing
    • Example: syphilis spirochete test using specific antibodies; fluorescence indicates presence of target
    • Direct antigen testing (rapid)
    • Do not require growth of pathogen; quick visual confirmation
    • Kits available for Staphylococcus aureus, Streptococcus pyogenes, Neisseria gonorrhoeae, etc.
    • Biochemical testing from isolates (pure colonies)
    • Enzyme presence/absence and substrates converted to products
    • Species identification requires a panel of tests; no single test is definitive
    • Microscopy vs Biochemistry in practice
    • Some organisms are first identified by morphology (microscopy) and then confirmed by biochemical panels
    • Major morphological/biochemical groups (visual aid)
    • Shigella, Klebsiella, Vibrio, Campylobacter, Sporeformers, Non-sporeformers, Acid-fast organisms, Regular vs pleomorphic rods, Motile vs nonmotile
    • Phage typing, animal inoculation, and antimicrobial sensitivity testing
    • Phage typing: bacteriophage specificity to differentiate strains
    • Animal inoculation: grows only in animals or cell culture
    • Antimicrobial susceptibility testing guides therapy
    • Isolated microbe clinical significance (SHEEPPHOTO LIBRARY concept)
    • Significance judged by number of microbes, repeated isolation, multiple specimen types, presence of true pathogen, and clinical correlation

  • Genotypic methods

    • DNA analysis
    • Hybridization using probes complementary to specific DNA sequences
    • PCR (mass production of species-specific DNA/RNA fragments from a specimen)
    • DNA or RNA targets; can work with viruses
    • Ribosomal RNA sequencing
    • Less specific than species-level markers but useful for broad identification and phylogeny

  • Immunologic methods

    • Serology (in vitro diagnostic testing of serum)
    • Antibodies with high specificity for antigens
    • Antigen-antibody interactions drive macroscopic or molecular readouts
    • Titers (serial dilutions) used to infer exposure or vaccination
    • Agglutination testing
    • Antibodies cross-link whole-cell antigens, forming visible clumps
    • Used for blood typing and some bacterial/viral disease serology
    • Precipitation tests
    • Soluble antigen becomes insoluble when antibody is added (immunoprecipitation in gels)
    • Western blot
    • Electrophoretic separation of proteins, followed by antibody detection
    • Highly specific; used as a confirmatory test for certain infections (e.g., HIV)
    • Enzyme-linked immunosorbent assays (ELISA)
    • Indirect ELISA: antigen bound to plate; patient serum; enzyme-linked secondary antibody; colorimetric readout
    • Direct/ Capture ELISA variants: antibody sandwich formats that trap antigen between two antibodies
    • Enzyme capture (antibody sandwich) ELISA
    • In vivo testing (antigen challenge in the body)
    • Tuberculin skin test and allergy testing

  • Virus diagnosis approaches

    • Signs and symptoms guide initial assessment
    • Direct visualization and antigen detection
    • Fluorescent staining for viral antigens in infected cells
    • Electron microscopy for direct visualization of virus particles (Dane particle for rotavirus, etc.)
    • Serology and confirmatory testing
    • Western blot or ELISA for antibody detection (e.g., HIV testing algorithm)
    • Culture and molecular approaches
    • Embryo cell culture, culture techniques for viral propagation (where possible)
    • PCR fingerprinting for rapid viral detection

Chapter 18: Gram-positive and Gram-negative Cocci

  • General notes
    • Cocci: spherical bacteria; Gram-positive vs Gram-negative distinctions are clinically important
    • Staphylococcus vs Streptococcus: arrangement and biochemical tests differentiate genera

Staphylococcus aureus

  • Morphology and culture

    • Gram-positive cocci in clusters
    • Grows in large, round, opaque colonies; optimum temp ~37°C
    • Facultative anaerobe; tolerates high salt and extreme pH

  • Identification and tests

    • Catalase-positive
    • Coagulase-positive differentiates S. aureus from coagulase-negative staphylococci
    • Enzymes: hyaluronidase, lipases; penicillinase (β-lactamase)
    • Virulence factors: toxins (hemolysins α, β, γ, δ), erythrogenic toxin, TSST (toxic shock toxin), enterotoxins; surface factors include fimbriae and capsule

  • Diseases and clinical spectrum

    • Localized infections: folliculitis, furuncles, carbuncles, impetigo
    • Systemic infections: bacteremia, endocarditis, meningitis, septic arthritis, pneumonia
    • Toxigenic diseases: staphylococcal food poisoning (enterotoxins), Staphylococcal scalded skin syndrome, toxic shock syndrome

  • Epidemiology and resistance

    • Carriage in 20-60% of healthy adults; nasal and skin reservoirs
    • MRSA strains common; resistance to methicillin and often multiple drugs; some strains resistant to many antibiotics except vancomycin
    • In healthcare settings, control relies on hygiene and infection-control practices

  • Coagulase-negative staphylococci

    • S. epidermidis, S. hominis, S. capitis, S. saprophyticus
    • Often associated with device-related infections and nosocomial infections; biofilm formation on catheters and implants

  • Other notes

    • 95% carry penicillinase, leading to penicillin/ampicillin resistance

Genus Streptococcus

  • General traits

    • Gram-positive cocci in chains
    • Can form capsules/slime layers; facultative anaerobes
    • Differentiation often by hemolysis pattern and Lancefield grouping (A, B, C, D, etc.)

  • Streptococcus pyogenes (Group A)

    • Virulence factors: surface antigens (fimbriae), hyaluronic acid capsule, other factors that resist lysozyme and phagocytosis
    • Extracellular toxins: streptolysins (β-hemolysis; tissue injury), erythrogenic toxin (fever and red rash), superantigens (massive immune activation)
    • Diseases: pharyngitis (strep throat); impetigo; erysipelas; necrotizing fasciitis; scarlet fever; rheumatic fever (type II hypersensitivity) and acute glomerulonephritis (type III hypersensitivity)
    • Transmission: human-to-human; contact, droplets, food, fomites; entry usually skin or pharynx
    • Key clinical notes: children most affected; untreated pharyngitis can lead to rheumatic fever or glomerulonephritis; penicillin is standard therapy; alternatives include erythromycin or cephalosporins if penicillin-allergic

  • Streptococcus pneumoniae

    • Alpha-hemolysis on blood agar; encapsulated; group D designation in some categorizations; nasopharyngeal colonization common
    • Diseases: pneumonia, meningitis, otitis media, sinusitis; significant cause of community-acquired infections
    • Tests: optochin sensitivity; bile solubility; vaccine-preventable disease (pneumococcal vaccines)

  • Streptococcus groups B, C, D and viridans group

    • Group B (S. agalactiae): neonatal infections (pneumonia, sepsis, meningitis); screening and intrapartum prophylaxis in pregnant women
    • Group D and viridans streptococci: oral and GI tract inhabitants; viridans group notable for dental plaques and potential subacute endocarditis; enterococci (E. faecalis, E. faecium) cause UTI, wound infections; increasingly multidrug-resistant
    • Group C, Group E, etc. include pathogens like S. equi/zooepidemicus; diversity requires biochemical panels for confirmation

  • Other notes

    • Streptococci can cause dental caries (mutans group among viridans)
    • Lancefield grouping helps categorize pathogens and guide treatment/toxicity patterns

Neisseria (Gram-negative cocci)

  • Nisseria gonorrhoeae
    • Gram-negative, bean-shaped diplococci; intracellular in neutrophils during infection
    • Transmission: sexually transmitted; also eye, throat, rectum, and other sites
    • Virulence factors: fimbriae (adhesion, slow phagocytosis), IgA protease; no flagella or spores
    • Infectious dose: around 100–1,000 bacteria; low infectious dose
    • ext{Infectious dose} \, ext{range} = 10^2 ext{ to } 10^3
    • Clinical presentation
    • Males: urethritis with discharge; scarring and infertility possible; a portion asymptomatic
    • Females: vaginitis; urethritis; risk of PID if ascends to upper genital tract
    • Diagnostics and resistance
    • Gram stain of male urethral discharge shows Gram-negative intracellular diplococci
    • Penicillinase-producing strains (PPNG) and tetracycline resistance (TRNG) exist; cephalosporins are commonly used
  • Nisseria meningitidis (meningococcus)
    • Gram-negative diplococci; encapsulated; fimbriae; IgA protease; endotoxin
    • Reservoir: human nasopharynx; epidemics in close-quarter populations; high-risk groups include children, adolescents, and college students
    • Disease: meningitis and meningococcemia; rapid onset with fever, neck stiffness, photophobia, severe complications
    • Pathogenesis: bacteria invade nasopharynx, enter blood, cross blood-brain barrier to CSF; endotoxin release contributes to shock
    • Diagnosis and treatment: Gram stain of CSF or blood; culture; rapid tests for capsular polysaccharide; treatment with cephalosporins; vaccination available for prevention

Chapter 19: Gram-positive Bacilli

  • General scheme (Table 19.1)

    • Sporeformers: Bacillus, Clostridium (gram-positive rods; spore-forming)
    • Non-sporeformers: Corynebacterium, Propionibacterium, Mycobacterium (acid-fast), Actinomyces, Nocardia
    • Classification by oxygen requirements: aerobic/facultative vs obligate anaerobes; regular vs irregular shapes; acid-fast status

  • Bacillus anthracis

    • Habitat: lives in soil; spores persist and spread via contaminated animal products
    • Virulence factors: polypeptide capsule; exotoxins that destroy macrophages
    • Disease forms: cutaneous (eschar), pulmonary (inhalation), gastrointestinal (ingestion)
    • Cutaneous: papule → necrotic eschar; least dangerous if treated early
    • Pulmonary: inhalation of spores → germination in lymph nodes → toxins cause macrophage destruction and systemic shock; high fatality if untreated
    • Treatment and prevention: penicillin, tetracycline, or ciprofloxacin; vaccines for livestock and high-risk humans; toxoid-based vaccines with boosters

  • Bacillus cereus

    • Habitat: soil; common in dust; associated with foods
    • Growth: spores survive cooking/reheating; toxin production in food causes vomiting and diarrhea
    • Treatment: supportive care; no specific antimicrobial therapy required in most cases

  • Clostridium perfringens (gas gangrene)

    • Habitat: soil; resident on skin and intestine; spores can be found in vaginal tract
    • Virulence factors: alpha toxin (lecithinase) causes cell lysis and edema; collagenase, hyaluronidase promote tissue destruction
    • Disease: myonecrosis; gas gangrene; associated with wound contamination, especially after trauma or surgical incisions
    • Treatment/prevention: immediate wound debridement, aggressive antibiotics (cephalosporins or penicillin), hyperbaric oxygen therapy

  • Clostridium tetani (tetanus)

    • Habitat: soil; spores enter through puncture wounds or skin breaches
    • Toxin: tetanospasmin blocks neurotransmitter release at neuromuscular junctions, causing sustained muscle contraction
    • Clinical picture: trismus (lockjaw) and severe muscle spasms; potential respiratory failure
    • Management: antitoxin (human tetanus antibody), antibiotics to stop infection, muscle relaxants; vaccination with booster every 10 years

  • Clostridium difficile

    • Normal colon resident in low numbers; antibiotic use disrupts microbiota
    • Toxins cause necrosis of the colonic wall; severe diarrhea and abdominal pain; can lead to pseudomembranous colitis; may cause fever and leukocytosis
    • Management: stool tests; withdrawal of offending antibiotics; vancomycin or metronidazole; fecal microbiota transplant in severe cases; increased infection-control precautions

  • Clostridium botulinum

    • Habitat: soil and water; spores form in environmental conditions
    • Botulinum toxin: inhibits acetylcholine release at neuromuscular junctions → flaccid paralysis
    • Disease: foodborne botulism from improper canning; infant botulism from infant ingestion of spores (honey); wound botulism
    • Diagnosis and treatment: antitoxin; supportive care; antibiotics for infectious botulism; prevention via proper food preservation

  • Listeria monocytogenes

    • Genome: not detailed in the provided notes; described as a New Pathogen entry
    • Notable: often associated with foodborne illness and high-risk populations (pregnant women, immunocompromised)

  • Corynebacterium diphtheriae

    • Morphology: Gram-positive irregular bacilli; pleomorphic with granules; club-shaped
    • Transmission: respiratory droplets from carriers or active cases
    • Disease: diphtheria; local pharyngitis with pseudomembrane; toxemia affecting heart and nerves
    • Treatment: antitoxin plus antibiotics (penicillin or erythromycin); vaccine-preventable via toxoid series

  • Mycobacteria (Genus)

    • Characteristics: Gram-positive irregular bacilli; acid-fast staining; strict aerobes; waxy mycolic acids; do not form spores
    • Notable features: cord factor linked to virulence and granuloma formation; slow growth; complex treatment regimens
    • Major species and diseases (highlights):
    • M. tuberculosis: tuberculosis in humans; granulomatous lesions (tubercles); latency and reactivation possible; Mantoux tuberculin test; chest X-ray; acid-fast bacilli in specimens; multi-drug therapy with isoniazid, rifampin, pyrazinamide, and others (e.g., Rifater combination)
    • M. leprae: Hansen’s disease (leprosy); slow-growing; cannot be grown in artificial media
    • Other species (M. avium complex, M. kansasii, M. marinum, M. intracellulare, M. fortuitum, M. chelonae, etc.): opportunistic infections in immunocompromised individuals (notably AIDS); varied habitats; treatment often involves multi-drug regimens
    • Diagnosis/Testing: Mantoux test (PPD) for in vivo testing; imaging and bacteriological confirmation; acid-fast staining; culture with biochemical testing

  • Other pathogens highlighted in the notes

    • Mycobacteria leprae: two disease forms (tuberculoid and lepromatous); nerve involvement common; long incubation; multidrug therapy
    • M. avium complex and atypical mycobacteria: opportunistic infections in AIDS and immunocompromised patients
    • Crohn’s disease connections mentioned in context of M. paratuberculosis in raw cow’s milk (noted as exploratory in the slide deck)

  • Quick reference to key diagnostic concepts from the slides

    • Direct visualization and staining (Gram, acid-fast, DFA) provide rapid presumptive identification
    • Culture and biochemical tests refine identification; genotype-based methods offer high specificity
    • Immunologic methods (serology, ELISA, Western blot, precipitation, agglutination) provide serodiagnosis and exposure history
    • In vivo tests (e.g., Tuberculin skin test) evaluate host response to pathogens
    • Specimen collection guidelines (Table 17.1) emphasize asepsis, specimen type appropriateness, and timely transport
    • Understanding of virulence factors (toxins, capsules, enzymes, adhesins) is essential for predicting disease presentation and treatment strategies

  • Key equations / quantitative references (converted to LaTeX)

    • Infectious dose examples:
    • ext{Infectious dose for Neisseria gonorrhoeae}
      ightarrow 10^2 ext{ to } 10^3 ext{ organisms}
    • Global burden reference:
    • frac{1}{3} ext{ of the world population are carriers of Mycobacterium tuberculosis}

  • Connections to foundational concepts

    • Morphology, staining, and culture form the backbone of clinical microbiology diagnostics
    • Virulence factors explain clinical manifestations and guide therapeutic targets (e.g., toxins, capsules, adhesion factors)
    • Antibiotic resistance patterns (e.g., MRSA, PPNG) shape treatment strategies and infection-control measures
    • Host-pathogen interactions (immunity, latency, and hypersensitivity) underpin sequelae like rheumatic fever, glomerulonephritis, and post-infectious syndromes

  • Relevance to real-world practice

    • Accurate specimen collection, rapid diagnostic tests, and appropriate antibiotic selection reduce morbidity and transmission
    • Vaccination (where applicable) and public health surveillance (e.g., meningitis vaccination, TB skin testing) prevent outbreaks
    • Understanding organism ecology (habitats, reservoirs, vectors) informs prevention and outbreak response

  • Ethical and practical implications

    • Handling of infectious samples requires biosafety precautions to protect patients and health workers
    • Antibiotic stewardship is critical to prevent resistance; overuse and misuse can worsen outcomes
    • Vaccination programs raise considerations about access, equity, and informed consent