MB 602 — PUBLIC HEALTH MICROBIOLOGY

EXAM-READY MASTER GUIDE

Exam Pattern Reminder: - Group A (Long Answer): Attempt 2 of 3 → ~5 marks each sub-question → Expect: epidemiology + pathogenesis + lab diagnosis + prevention - Group B (Short Answer): Attempt 3 of 4 → 5-6 marks each → Brief but complete - Group C (Very Short): Attempt 5 of 6 → Enlist/Mention/Define → 2-3 points enough

SECTION 1: WATER-BORNE INFECTIONS

SALMONELLA (Typhoid / Salmonellosis)

Organism

  • Salmonella typhi (typhoid fever) / S. typhimurium, S. enteritidis (food-borne salmonellosis)
  • Gram-negative bacillus, family Enterobacteriaceae
  • Non-spore forming, peritrichous flagella (motile), non-capsulated (except Vi antigen)
  • Antigens:
    • O antigen (somatic, LPS): Rises first in Widal, falls first
    • H antigen (flagellar): Rises later, persists longer
    • Vi antigen (virulence/capsular): Inhibits phagocytosis; indicates carrier state in Widal

TYPHOID FEVER (Water-borne / Long Answer Level)

Epidemiology: - Fecal-oral route via contaminated water and food - Endemic in Nepal (monsoon peak) - Incubation: 7–21 days - Source: Chronic carriers (gallbladder reservoir) → shed in feces/urine - Infective dose: 10³–10⁶ organisms

Pathogenesis: 1. Ingestion → reaches small intestine 2. Penetrates epithelium via M cells in Peyer’s patches 3. Taken up by macrophages → survives intracellularly 4. Spreads to mesenteric lymph nodes → bloodstream (bacteremia — Week 1) 5. Multiplies in Peyer’s patches, liver, spleen, gallbladder 6. Week 2–3: Ulceration of Peyer’s patches → risk of intestinal perforation and hemorrhage 7. Endotoxin → systemic toxemia (fever, headache)

Clinical: Stepwise fever, bradycardia relative to fever, rose spots on trunk (10%), diarrhea or constipation, hepatosplenomegaly

Laboratory Diagnosis:

Week Best Test Notes
Week 1 Blood culture (gold standard) Most sensitive; Widal usually negative
Week 2–3 Stool + Urine culture, Widal Bacteria shed in feces
Any time Bone marrow culture Most sensitive (90%); remains positive even after antibiotics
  • Widal Test:
    • O titre ≥1:160 + H titre ≥1:160 = Significant
    • Vi titre = Carrier state
    • Limitations: False positive with other Salmonella, malaria; cross-reactions
  • Typhidot Test: IgM anti-S. typhi dot-ELISA; more specific
  • Culture Media: MacConkey, DCA (Deoxycholate Citrate Agar), SS agar, Selenite F broth (enrichment)
  • Biochemistry: Glucose + (no gas for S. typhi), Lactose −, H₂S +, Urease −, Indole −

Prevention & Control: - Safe water supply, improved sanitation - Vaccines: TAB (killed whole cell), Ty21a (live attenuated oral), Vi polysaccharide (parenteral) - Treatment of carriers: Prolonged antibiotics ± cholecystectomy

NON-TYPHOIDAL SALMONELLOSIS (Food-borne)

Organisms: S. typhimurium, S. enteritidis, S. choleraesuis
Source: Poultry, eggs, meat, dairy products
Incubation: 6–48 hours (shorter than typhoid)
Symptoms: Nausea, vomiting, watery/bloody diarrhea, fever, abdominal cramps; self-limiting 3–7 days
Pathogenesis: Invasion of intestinal epithelium → inflammatory diarrhea → rarely bacteremia
Lab: Stool culture on SS/DCA agar, Selenite F enrichment; same biochemistry as typhoid
Prevention: Proper cooking of poultry/eggs, refrigeration, avoid cross-contamination

CAMPYLOBACTER (Food/Water-borne)

Organism

  • Campylobacter jejuni (most common), C. coli
  • Gram-negative, comma/S-shaped or spiral rod (seagull wing on Gram stain)
  • Microaerophilic (5% O₂, 10% CO₂) — key feature
  • Thermophilic: Grows at 42°C (diagnostic advantage)
  • Single polar flagellum → darting/corkscrew motility on dark field microscopy
  • Oxidase +, Catalase +

Epidemiology

  • Most common cause of bacterial gastroenteritis worldwide
  • Zoonotic: Poultry (main reservoir), cattle, sheep, pets
  • Transmission: Undercooked poultry, raw milk, contaminated water
  • Incubation: 2–5 days
  • Peak: Summer/monsoon months

Pathogenesis

  • Colonizes jejunum and ileum
  • Cytolethal Distending Toxin (CDT): Cell cycle arrest, apoptosis
  • Invasion of epithelium → inflammatory bloody diarrhea
  • Post-infectious complications (IMPORTANT):
    • Guillain-Barré Syndrome (GBS): Molecular mimicry — LOS of C. jejuni resembles gangliosides on peripheral nerve myelin → autoimmune demyelination
    • Reactive arthritis (HLA-B27 individuals)
    • Hemolytic Uremic Syndrome (rare)

Clinical Features

  • Prodrome: Fever, headache, myalgia (1–3 days)
  • Profuse watery/bloody diarrhea with mucus
  • Severe colicky abdominal pain
  • Self-limiting: 5–7 days

Laboratory Diagnosis

  1. Specimen: Fresh stool
  2. Dark-field / Phase contrast microscopy: Corkscrew darting motility (rapid, presumptive)
  3. Gram stain: S-shaped or spiral Gram-negative rods
  4. Culture (key):
    • Selective media: Skirrow’s agar, CCDA (Charcoal Cefoperazone Deoxycholate Agar), Butzler medium
    • Temperature: 42°C (selective for C. jejuni/C. coli)
    • Atmosphere: Microaerophilic (CampyGen/Campypack — 5% O₂, 10% CO₂)
    • Incubation: 48–72 hours
  5. Biochemical ID:
    • Oxidase +, Catalase +
    • Hippurate hydrolysis + (C. jejuni; negative in C. coli — differentiates them)
    • Nalidixic acid sensitive (C. jejuni), resistant (C. coli)
  6. Serology: Not routine
  7. PCR: Reference labs, species confirmation

Prevention & Control

  • Thorough cooking of poultry (internal temp >74°C)
  • Pasteurization of milk
  • Hand hygiene (especially after animal contact)
  • Safe water supply
  • Avoid cross-contamination in kitchen

SECTION 2: VECTOR-BORNE INFECTIONS

VISCERAL LEISHMANIASIS (Kala-azar)

Organism

Leishmania donovani — Obligate intracellular protozoan parasite

Two forms: | Form | Location | Features | |——|———-|———-| | Promastigote | Sandfly / NNN culture | Elongated, anterior flagellum; infective stage | | Amastigote (LD body) | Human macrophages | Oval, non-motile, intracellular; diagnostic stage |

Vector

Phlebotomus argentipes (female sandfly) — Indian subcontinent/Nepal

Epidemiology

  • Nepal: Endemic in Terai districts — Saptari, Siraha, Dhanusha, Mahottari, Sarlahi, Rautahat, Bardia
  • Part of South Asia VL elimination programme (target: <1 case/10,000 population)
  • Incubation: 2–6 months (range: 10 days to years)
  • Reservoir: Humans (anthroponotic in South Asia)

Pathogenesis

  1. Infected sandfly bites → promastigotes injected into skin
  2. Taken up by dermal macrophages → transform into amastigotes
  3. Multiply within macrophages → spread via lymphatics and blood
  4. Invade cells of RES (spleen, liver, bone marrow, lymph nodes)
  5. Result:
    • Massive splenomegaly (most prominent sign)
    • Hepatomegaly
    • Pancytopenia (bone marrow infiltration)
    • Hypoalbuminemia, Hypergammaglobulinemia
    • Immune suppression → susceptibility to secondary infections (pneumonia, TB, dysentery)
    • Dark skin pigmentation (melanin deposition) → kala-azar = black fever (Hindi)

Clinical Features

  • Prolonged fever (double-peak/Pel-Ebstein type)
  • Massive splenomegaly (hallmark)
  • Hepatomegaly
  • Weight loss, cachexia, weakness
  • Pancytopenia (anemia, bleeding, infections)
  • Dark skin discoloration
  • PKDL (Post Kala-azar Dermal Leishmaniasis): Hypopigmented/nodular skin lesions appearing months–years after treatment; important reservoir

Laboratory Diagnosis

Definitive — Demonstration of Amastigotes (LD bodies):

Specimen Sensitivity Safety
Splenic aspirate 95–98% (gold standard) ⚠️ High risk of bleeding
Bone marrow aspirate 70–85% Safer (preferred in practice)
Lymph node aspirate 60–70% Safe, less sensitive
Liver biopsy 70–80% Moderate risk
Buffy coat smear Low Easiest, least sensitive
  • Staining: Leishman stain or Giemsa stain (amastigotes appear as oval bodies with nucleus + kinetoplast)

Culture: - NNN medium (Novy-MacNeal-Nicolle): Promastigotes grow at room temperature; 1–4 weeks

Serology: | Test | Notes | |——|——-| | rK39 ICT (RDT) | Rapid, field diagnosis; highly specific; WHO recommended | | ELISA | Anti-Leishmania antibodies; sensitive | | DAT (Direct Agglutination Test) | Good specificity; needs equipment | | Formol Gel (Aldehyde) Test | Non-specific (hypergammaglobulinemia); no longer recommended | | Complement Fixation Test | Old method |

Leishmanin Skin Test (Montenegro Test): - Negative in active VL (immune suppression) - Positive after recovery (delayed-type hypersensitivity, memory) - Positive in CL and MCL

Molecular: - PCR: Species identification, sensitivity monitoring

Other: - CBC: Pancytopenia - Serum proteins: ↓Albumin, ↑Globulin

Prevention & Control — Five Pillars of VL Elimination Programme

  1. Early case detection and complete treatment (prevents reservoir)
  2. Vector control: Indoor Residual Spraying (IRS) with DDT or synthetic pyrethroids
  3. Social mobilization and community engagement (BCC - behavior change communication)
  4. Clinical and operational research (new diagnostics, drugs)
  5. Monitoring, evaluation and surveillance (track progress toward elimination)

Additional measures: - Insecticide-treated bed nets (ITNs) - Personal protection (repellents, long sleeves) - Treatment of PKDL cases (act as reservoir) - Improved housing (sandfly breeding site reduction)

Cutaneous Leishmaniasis (CL) — Brief Notes

  • Nepal: L. major, L. tropica
  • Lesion: Painless ulcerated papule → “volcano crater” appearance; heals with scar
  • Lab: Slit-skin smear from lesion edge, biopsy (amastigotes), PCR
  • Montenegro test: Positive in CL

SECTION 3: HOSPITAL ACQUIRED INFECTIONS (HAI)

NOSOCOMIAL / HOSPITAL ACQUIRED INFECTIONS

Definition

An infection acquired in a healthcare setting that was not present or incubating at the time of admission (develops ≥48 hours after admission OR within 30 days of discharge)

Types (Most Common)

Type Frequency Common Association
Urinary Tract Infection (UTI) ~35% (most common) Urinary catheter (CAUTI)
Surgical Site Infection (SSI) ~20% Surgical procedures
Pneumonia / Respiratory ~15% Mechanical ventilator (VAP)
Bloodstream Infection (BSI) ~15% IV/central venous catheters (CLABSI)

Sources

Endogenous (most common): Patient’s own normal flora (especially gut bacteria)

Exogenous: - Healthcare workers (hands — most important route) - Environment (surfaces, water, air, dust) - Other patients (cross-infection) - Contaminated equipment/devices - Blood products

Routes of Transmission

  1. Contact transmission (most common):
    • Direct: HCW hands → patient
    • Indirect: Contaminated equipment → patient
  2. Droplet transmission: Large droplets (>5 μm) — flu, pertussis
  3. Airborne transmission: Droplet nuclei (<5 μm) — TB, measles, chicken pox
  4. Common vehicle: Contaminated food, water, medications, IV fluids
  5. Vector-borne: Rare in hospital settings

Factors Increasing Susceptibility to HAI

Host factors: - Extremes of age (neonates, elderly) - Underlying diseases (diabetes, cancer, AIDS) - Malnutrition - Immunosuppression (steroids, chemotherapy) - Breaks in skin integrity (burns, wounds)

Healthcare factors: - Invasive devices (catheters, ventilators, IV lines) - Broad-spectrum antibiotic use (disrupts normal flora, selects resistant organisms) - Prolonged hospitalization - Surgical procedures - ICU admission

Major Microorganisms Causing HAI in Nepal

Bacteria: - Escherichia coli (UTI, ESBL producing) - Klebsiella pneumoniae (pneumonia, UTI, ESBL/KPC producing) - Staphylococcus aureus (wound infections, BSI) — especially MRSA - Pseudomonas aeruginosa (VAP, burn wounds) - Acinetobacter baumannii (ICU, carbapenem resistant) - Enterococcus spp. (UTI, VRE) - Coagulase-negative Staphylococci (CONS — catheter-related BSI)

Fungi: Candida spp. (immunocompromised, catheter-related)

Viruses: HBV, HCV (blood transfusion/needlestick), CMV (immunocompromised)

DISPOSAL OF INFECTIVE HOSPITAL WASTE

Categories of Hospital Waste (Color-coded)

Category Color Examples Method
Infectious waste Yellow Cultures, contaminated dressings, pathological waste Autoclaving or incineration
Sharps Red/Yellow (rigid) Needles, syringes, blades, glass Puncture-resistant container → Incineration/autoclaving
Pharmaceutical Blue Expired/unused drugs Return to pharmacy / incineration
Chemical Black Lab chemicals, fixer, developer Chemical treatment, separate disposal
Radioactive Purple/White Radiotherapy waste Specialized disposal (hold for decay)
General non-hazardous Black Paper, packaging Municipal landfill

Methods of Disposal

  1. Autoclaving (Steam Sterilization):

    • 121°C, 15 psi, 20–30 minutes
    • For: Microbiological cultures, contaminated lab materials, liquid waste
    • Then can go to regular landfill

  2. Incineration:

    • High-temperature burning (>800°C for primary chamber, >1000°C secondary)
    • Most effective method; destroys all microorganisms and reduces volume
    • For: Anatomical/pathological waste, sharps, cytotoxic waste, infectious waste
    • By-product: Ash (much smaller volume)

  3. Chemical Disinfection:

    • Sodium hypochlorite (1–10% depending on load), glutaraldehyde, formaldehyde
    • For: Liquid infectious waste (blood, urine, sputa), surfaces
    • Chemical sterilants: Glutaraldehyde 2% (Cidex), ortho-phthalaldehyde, hydrogen peroxide

  4. Microwave Treatment: For non-metallic infectious waste

  5. Sanitary Landfill: For treated/non-hazardous waste only

  6. Sharps Containers:

    • Rigid, puncture-resistant, leak-proof
    • Fill to ¾ only → seal → autoclaved or incinerated

Steps in Disposal of Infectious Waste (Step-by-step for exam)

  1. Segregation at source — MOST CRITICAL STEP (correct color-coded containers at point of generation)
  2. Collection in appropriate containers (label with biohazard symbol)
  3. Labeling (type of waste, ward, date)
  4. Storage in designated secure area (not >24 hours for infectious waste)
  5. Internal transport (designated trolleys, routes, times)
  6. Treatment (autoclaving, incineration as appropriate)
  7. External transport (licensed vehicles)
  8. Final disposal (licensed facility/landfill)

DISINFECTION OF HOSPITAL ENVIRONMENT

Spaulding Classification of Equipment

Category Definition Method Examples
Critical Enters sterile tissue/bloodstream Sterilization (autoclave, EtO, plasma) Surgical instruments, catheters, implants
Semi-critical Contacts mucous membranes/non-intact skin High-level disinfection (HLD) Endoscopes, laryngoscopes, ET tubes
Non-critical Contacts intact skin Low/intermediate disinfection BP cuffs, stethoscopes, bedpans

Disinfection Methods

Surfaces: - Routine: Sodium hypochlorite 1000 ppm (0.1%) with detergent - Blood/body fluid spills: Sodium hypochlorite 10,000 ppm (1%) — cover, absorb, then disinfect - Quaternary ammonium compounds (quats): surfaces, furniture

Air (OT/ICU): - UV irradiation (germicidal UV-C 254 nm): OT, isolation rooms; inactivates DNA - HEPA filtration: Removes particles ≥0.3 μm (99.97%) - Positive pressure rooms (for immunocompromised) - Negative pressure rooms (for TB, infectious aerosols) - Adequate air changes (12–15 per hour in ICU)

Endoscopes (Semi-critical): - Clean → enzymatic detergent soak → manual brushing → HLD (Glutaraldehyde 2% for 20–45 min, or OPA)

Hands (most important intervention!): - WHO 5 Moments for Hand Hygiene - Alcohol-based hand rub (ABHR) preferred (when hands not visibly soiled) - Soap and water (when visibly soiled, C. diff, spores)

MONITORING OF HOSPITAL SANITATION

Methods

  1. Environmental surface sampling:
    • Swabs from high-touch surfaces
    • Contact plates (RODAC plates)
    • Culture → CFU/cm² or cm³
  2. ATP Bioluminescence Assay:
    • Detects adenosine triphosphate (ATP) as a marker of organic contamination
    • Uses luciferase-luciferin reaction → produces light → measured in Relative Light Units (RLU)
    • Advantages: Rapid (seconds), quantitative, easy to use
    • Disadvantages: Expensive equipment, non-specific (detects ATP from both microbial AND human cells), doesn’t identify organisms
  3. Air sampling:
    • Settle plates (passive sampling — Petri dishes left open 1 hour → count colonies)
    • Active air sampling (impaction: SAS sampler, cascade impactors)
    • Result: CFU/m³
  4. Water testing:
    • Colony counts (Total Viable Count)
    • E. coli / coliform testing (indicator organisms)
  5. Process monitoring (Sterility Assurance):
    • Physical indicators: Temperature, pressure, time charts
    • Chemical indicators (CI): Color change tapes (Class 1–6, Browne’s tubes)
    • Biological indicators (BI): Spore tests — Geobacillus stearothermophilus (autoclave), Bacillus atrophaeus (EtO)
    • Bowie-Dick test: Tests steam penetration in pre-vacuum autoclaves

SECTION 4: UTS TOPICS (Rotavirus, Pneumonia, Hepatitis A, M. tuberculosis)

ROTAVIRUS

Organism

  • Family: Reoviridae, Genus: Rotavirus
  • Double-stranded RNA (dsRNA), 11 segments — key feature
  • Non-enveloped, icosahedral
  • Wheel-like appearance on EM (rota = wheel in Latin)
  • Three-layered capsid:
    • Inner core
    • Inner capsid → VP6 (group antigen — Group A most common in humans)
    • Outer capsid → VP7 (G antigen) + VP4 (P antigen) → serotyping
  • Common types: G1P[8] (most prevalent globally); G2, G3, G4, G9 also important

Epidemiology

  • Leading cause of severe diarrheal disease in children <5 years worldwide
  • ~215,000 child deaths/year globally (mostly developing countries)
  • Nepal: Major cause of childhood diarrheal hospitalization
  • Route: Fecal-oral (highly contagious, very low infective dose ~10–100 particles)
  • Incubation: 1–3 days
  • Season: Winter peak (temperate); year-round with rainy season peak (Nepal)
  • By age 5: virtually all children have been infected (natural immunity builds)

Pathogenesis

  1. Ingestion → infects mature enterocytes at tips of villi (small intestine)
  2. VP4 attaches to sialic acid receptors on enterocyte surface
  3. Viral replication → enterocyte death → villous atrophy
  4. Reduced absorptive surface → malabsorption (osmotic diarrhea)
  5. NSP4 (Non-structural protein 4): Acts as viral enterotoxin → stimulates Cl⁻ secretion via Ca²⁺-dependent pathway → secretory diarrhea
  6. Combined osmotic + secretory diarrhea → profuse watery diarrhea → dehydration

Clinical Features

  • Sudden onset watery diarrhea (profuse, non-bloody)
  • Vomiting often precedes or accompanies diarrhea (distinguishes from bacterial)
  • Fever (low-grade)
  • Rapid dehydration → can be life-threatening in infants
  • Self-limiting: 3–8 days

Laboratory Diagnosis

Method Details
ELISA (VP6 antigen) Most common routine test; detects Group A rotavirus antigen in stool
Rapid ICT Immunochromatographic strip test; field diagnosis
Electron Microscopy (EM) Gold standard; characteristic wheel-shaped particles (70 nm); expensive
RT-PCR Genotyping of G and P types; research/surveillance
PAGE 11-band pattern of dsRNA segments on polyacrylamide gel; characteristic, now replaced by PCR
  • Specimen: Fresh stool (no transport medium needed; refrigerate)

Prevention & Control

  • Vaccines (most important!):
    • Rotarix (RV1): GSK; live attenuated; 2 doses at 6 and 10 weeks; human G1P[8]
    • RotaTeq (RV5): Merck; live attenuated; 3 doses; pentavalent (G1, G2, G3, G4 + P[8])
    • Efficacy: 85–95% against severe disease
    • Nepal: Rotarix included in National Immunization Programme
  • ORS (Oral Rehydration Salts): Management of dehydration — cornerstone of treatment
  • Zinc supplementation: Reduces severity and duration
  • WASH (Water, Sanitation, Hygiene)
  • Breastfeeding (protective)
  • Vitamin A supplementation

PNEUMONIA

Definition

Inflammation of the lung parenchyma (alveoli and interstitium) caused by infectious agents

Classification

By Etiology: - Typical bacterial pneumonia: Streptococcus pneumoniae, H. influenzae, S. aureus, Klebsiella - Atypical pneumonia: Mycoplasma pneumoniae, Chlamydophila pneumoniae, Legionella pneumophila (cannot grow on standard media; interstitial pattern on CXR) - Viral: Influenza, RSV, SARS-CoV-2, Adenovirus, Parainfluenza - Fungal: Pneumocystis jirovecii (PCP — in HIV/immunocompromised)

By Setting: - Community-Acquired Pneumonia (CAP) — S. pneumoniae most common - Hospital-Acquired Pneumonia (HAP) — Gram-negatives (Pseudomonas, Klebsiella, Acinetobacter), MRSA - Ventilator-Associated Pneumonia (VAP) — same as HAP; most severe

By Anatomy: - Lobar pneumonia (S. pneumoniae) - Bronchopneumonia (S. aureus, H. influenzae, aspiration) - Interstitial pneumonia (atypical — viral, Mycoplasma)

PNEUMOCOCCAL PNEUMONIA (S. pneumoniae)

Organism: - Gram-positive, lancet-shaped diplococci - Alpha-hemolytic (green hemolysis on blood agar) - Optochin sensitive (inhibition zone >14 mm) — key identification feature - Bile soluble (positive) — colonies dissolve in bile/sodium deoxycholate - Capsule: 90+ serotypes; major anti-phagocytic virulence factor - Other virulence factors: Pneumolysin, IgA protease, neuraminidase, teichoic acid

Pathogenesis (4 Stages of Lobar Pneumonia):

Stage Timing Features
Congestion Day 1–2 Alveolar edema; bacteria multiply; congested red lung
Red Hepatization Day 2–3 Alveoli filled with RBC, fibrin, PMNs; lung solid/red (liver-like)
Grey Hepatization Day 4–8 RBC lyse; WBC + fibrin remain; lung grey
Resolution Day 8+ Enzymatic digestion; macrophages clear debris; lung returns to normal

Capsule → resists phagocytosis → bacteria proliferate → induce intense inflammation

Signs & Symptoms: - Sudden onset: Rigors, high fever (39–40°C) - Productive cough: Rusty/blood-tinged sputum (classic) - Pleuritic chest pain (sharp, worse with breathing) - Dyspnea, tachypnea - Signs of consolidation: Dullness on percussion, bronchial breathing, increased fremitus - Possible herpes labialis (reactivation)

Laboratory Diagnosis:

  1. Specimen: Purulent sputum (early morning); BAL; blood; pleural fluid
  2. Gram stain of sputum:
    • Valid sample: >25 PMNs/LPF + <10 squamous epithelial cells/LPF
    • Findings: Gram-positive lancet-shaped diplococci surrounded by PMNs
  3. Culture:
    • Blood agar (5% sheep blood): Alpha-hemolysis → draughtsman (dimpled) colonies
    • Optochin disc → inhibition = S. pneumoniae
    • Bile solubility test (positive)
    • MacConkey: No growth (Gram-positive)
  4. Blood culture: Positive in 20–30% bacteremic cases
  5. Quellung (capsular swelling) reaction: Type-specific antisera → capsule swells → serotyping
  6. Urinary antigen test (Binax NOW): Rapid, for S. pneumoniae (serotype 1+) and Legionella; can be positive even after antibiotics
  7. CXR: Lobar consolidation (typical); Ground glass/interstitial (atypical)
  8. Inflammatory markers: ↑CRP, ↑Procalcitonin, ↑WBC (neutrophilia)

Atypical Pneumonia — Quick Notes:

Organism Key Feature Test
Mycoplasma pneumoniae Cold agglutinins positive (IgM vs RBC) Cold agglutinin titer ≥1:64; PCR; serology
Legionella pneumophila Water cooling towers, air conditioning; Pontiac fever Urinary antigen (serogroup 1); BCYE agar (culture)
Chlamydophila pneumoniae Mild community pneumonia Serology, PCR

Prevention in Nepal: - PCV13 (Pneumococcal Conjugate Vaccine — 13 valent): In national EPI schedule for children (at 6, 10, 14 weeks) - PPSV23 (Polysaccharide, 23-valent): Adults ≥65 years, high-risk groups - Exclusive breastfeeding - Reduction of indoor air pollution (cooking fires — significant in Nepal) - Vitamin A and zinc supplementation - Management of malnutrition - Early and appropriate antibiotic treatment

HEPATITIS A

Organism

  • Family: Picornaviridae, Genus: Hepatovirus
  • Single-stranded, positive-sense RNA virus
  • Non-enveloped, icosahedral, 27–32 nm
  • Single serotype (one vaccine protects against all strains)
  • Stable in environment: Resists acid, mild heat, detergents → survives in water/food
  • Inactivated by: 85°C for 1 min, chlorine, UV, formalin

Epidemiology

  • Most common cause of acute viral hepatitis worldwide
  • Route: Fecal-oral exclusively (no parenteral/sexual transmission unlike HBV/HCV)
    • Contaminated water (major route in Nepal/developing countries)
    • Contaminated food (raw shellfish, unwashed vegetables, food handlers)
    • Person-to-person (close contact)
  • Nepal: High endemicity area — most children infected early → asymptomatic → lifelong immunity
  • Incubation period: 15–50 days (average 28 days)
  • Travelers to endemic areas at high risk
  • Outbreaks: Linked to contaminated water supply or infected food handler

Pathogenesis

  1. Ingestion → absorbed in small intestine → portal blood → liver
  2. Replicates in hepatocytes
  3. Peak fecal shedding: 2 weeks BEFORE symptoms (spreads even before illness apparent)
  4. Fecal shedding continues until ~2 weeks after jaundice onset
  5. Liver damage: Immune-mediated (T-cell cytolysis of infected hepatocytes + cytokines)
  6. NEVER becomes chronic (unlike HBV, HCV) → complete recovery or rarely fulminant

Clinical Features

Three Phases:

Phase Features
Pre-icteric (Prodrome): 1–2 weeks Fever, malaise, anorexia, nausea/vomiting, RUQ pain, distaste for cigarettes/alcohol
Icteric phase: 2–8 weeks Jaundice (scleral icterus first), dark urine (bilirubin), pale stools (bilirubin not reaching gut), hepatomegaly, pruritus
Convalescent phase Gradual resolution

Complications (rare): - Cholestatic hepatitis (prolonged jaundice) - Relapsing hepatitis - Fulminant hepatic failure (rare, <1%; higher risk if pre-existing liver disease, HBV co-infection)

NO CHRONICITY — Complete recovery in >99%

Laboratory Diagnosis

Serology (most important):

Marker Interpretation
Anti-HAV IgM (+) Acute/recent infection (appears at symptom onset, positive for 3–6 months)
Anti-HAV IgG (+) only Past infection OR vaccinated (immune; lifelong)
Both negative Susceptible (never infected, never vaccinated)

Liver Function Tests (LFT): - ALT/AST: Markedly elevated (>10× normal in acute hepatitis) - Bilirubin: Both conjugated and unconjugated elevated - Alkaline phosphatase: Mildly elevated - PT/INR: Elevated in severe disease

Viral detection (not routine): - HAV RNA by RT-PCR (stool/blood — research, outbreak investigation) - Electron microscopy (stool — classical 27 nm particles)

Prevention & Control

Active Immunization (Primary Prevention): - Inactivated HAV vaccine (Havrix, VAQTA) - 2 doses: 0 and 6–12 months - Efficacy: >94% after 2 doses - Recommended for: Travelers to endemic areas, food handlers, children in endemic areas, MSM, chronic liver disease patients, healthcare workers

Passive Immunization: - Normal Human Immunoglobulin (IG/HNIG): Post-exposure prophylaxis if given within 2 weeks of exposure - Also used for pre-travel short-term protection

Environmental Control: - Safe water supply (chlorination — inactivates HAV) - Proper sewage disposal and sanitation - Food hygiene: Hand washing, cooking shellfish thoroughly - WASH improvements (key in Nepal context)

MYCOBACTERIUM TUBERCULOSIS

Organism

  • Acid-Fast Bacillus (AFB) — key characteristic
  • Why acid-fast? Mycolic acids in cell wall → retain carbol fuchsin dye after acid-alcohol decolorization
  • Gram-positive (weakly) but Gram stain not used routinely
  • Strictly aerobic, non-spore forming, non-motile, non-capsulated
  • Extremely slow growing — doubling time ~20 hours; colonies visible in 4–6 weeks on LJ medium
  • Cell wall: Peptidoglycan + mycolic acids + arabinogalactan → thick waxy coat → resistant to drying, chemicals, many disinfectants

Virulence Factors

  • Cord factor (Trehalose 6,6’-dimycolate): Most important; inhibits macrophage functions, toxic to cells; causes cords (parallel arrangement of bacilli in culture)
  • Sulfatides: Inhibit phagosome-lysosome fusion (survival in macrophage)
  • Cell wall lipids (Mycolic acids): Resist killing by lysosomal enzymes
  • LAM (Lipoarabinomannan): Inhibits macrophage activation, scavenges ROS

Epidemiology in Nepal

  • Nepal = High-burden TB country
  • Incidence: ~117/100,000 population
  • Risk factors: Poverty, overcrowding, malnutrition, HIV co-infection, indoor air pollution (cooking fires), smoking
  • MDR-TB (Multi-Drug Resistant TB): Resistance to ≥ rifampicin + isoniazid
    • Causes: Incomplete treatment, drug supply interruption, poor adherence, substandard drugs
    • Nepal: Emerging problem, significant public health concern
  • XDR-TB: MDR-TB + resistance to fluoroquinolones + injectable agents

Mode of Transmission

  • Airborne droplet nuclei (1–5 μm) expelled during coughing, sneezing, singing, talking
  • Single cough: ~3,000 droplet nuclei released
  • Droplet nuclei remain suspended in air for hours
  • Risk: Close, prolonged contact; crowded, poorly ventilated spaces
  • NOT primarily from fomites

Pathogenesis

PRIMARY TUBERCULOSIS: 1. Inhaled droplet nuclei → alveoli → engulfed by alveolar macrophages 2. If immunity sufficient: Contained → Ghon focus (small granuloma in lung periphery) + hilar lymph node enlargement = Ghon complex (Primary complex) 3. Dormant state → Latent TB (LTBI): TST positive, no symptoms, not infectious; 90% stay latent 4. If immunity insufficient: Primary progressive TB → spread

REACTIVATION (SECONDARY) TUBERCULOSIS: - Triggered by: HIV, diabetes, malnutrition, steroid use, aging - Predilection for upper lobes (high O₂ tension) - Caseous necrosis → cavity formation → liquefaction → infectious (AFB in sputum) - Bronchogenic spread → other lung areas - Hematogenous spread → miliary TB (millet-seed-like lesions throughout body), TB meningitis, skeletal TB

Immune Response: - CMI (Cell-Mediated Immunity): Critical; CD4+ T cells activate macrophages → macrophages kill mycobacteria → granuloma formation - Granuloma structure: Central caseous necrosis ← surrounded by macrophages (epithelioid cells) + Langhans giant cells + lymphocytes + fibroblasts

Clinical Features of PTB (Pulmonary TB)

  • Chronic cough >2 weeks (key indicator in Nepal)
  • Hemoptysis (blood-stained sputum — classic)
  • Fever (low-grade, evening rise)
  • Night sweats
  • Weight loss, anorexia (cachexia)
  • Fatigue, weakness
  • Chest pain
  • (Advanced: dyspnea, clubbing)

Laboratory Diagnosis of PTB

Specimens: Early morning sputum × 3 (spot → overnight → spot), BAL, gastric lavage (children), pleural fluid

DIRECT MICROSCOPY (Quickest):

Method Technique Sensitivity
Ziehl-Neelsen (ZN) stain Carbol fuchsin → acid-alcohol decolorize → methylene blue counter; AFB = red rods on blue background 40–60%
Auramine-rhodamine (Fluorescent) AFB appear bright yellow-green fluorescent on dark background; more sensitive, faster screening 60–80%

ZN Grading: - 3+ = >10 AFB/oil immersion field - 2+ = 1–10 AFB/field - 1+ = 10–99 AFB/100 fields - Scanty = <10 AFB/100 fields (report exact number) - Negative = 0 AFB/300 fields

CULTURE (Gold standard for diagnosis + DST):

Medium Type Time Notes
Löwenstein-Jensen (LJ) Solid (egg-based) 4–6 weeks Buff/cream, rough, dry breadcrumb colonies; cheap
MGIT (Mycobacteria Growth Indicator Tube) Liquid (Middlebrook 7H9 + O₂-sensitive fluorescent indicator) 1–3 weeks More sensitive and faster than LJ; automated (BACTEC 960)

DRUG SUSCEPTIBILITY TESTING (DST): - Proportion method on LJ (slower) - MGIT-based DST (faster, automated) - For MDR-TB detection (rifampicin + isoniazid resistance)

MOLECULAR METHODS:

Test What it detects Time Where used
GeneXpert MTB/RIF (Xpert) M. tb DNA + rifampicin resistance (rpoB gene mutation) ~2 hours WHO-recommended 1st test; peripheral Nepal
Line Probe Assay (Hain MTBDRplus) RIF resistance (rpoB) + INH resistance (katG, inhA) 1–2 days MDR screening
Whole Genome Sequencing (WGS) Full resistance profile, phylogeny Days Reference labs

IMMUNOLOGICAL TESTS:

Tuberculin Skin Test (TST / Mantoux): - 5 TU PPD injected intradermally (volar forearm) - Read at 48–72 hours (measure induration, NOT erythema) - Positive cutoffs: - ≥5 mm: HIV+, recent close contact, CXR evidence - ≥10 mm: Most populations (healthcare workers, immigrants, high-risk groups) - ≥15 mm: Low-risk individuals - False positive: BCG vaccination, NTM infection - False negative: Immunosuppressed, miliary TB, very early infection

IGRA (Interferon Gamma Release Assay): - QuantiFERON-TB Gold Plus, T-SPOT.TB - Measures IFN-γ release by T-cells in response to TB-specific antigens (ESAT-6, CFP-10) - More specific than TST (not affected by BCG) - Cannot distinguish active vs latent TB - Preferred in BCG-vaccinated populations

CXR Findings: - Primary TB: Hilar lymphadenopathy, peripheral consolidation (Ghon focus) - Secondary/Reactivation: Upper lobe infiltrates, cavitation (pathognomonic), fibrosis - Miliary TB: Diffuse bilateral nodular pattern (millet seeds) - Pleural TB: Pleural effusion (unilateral)

MDR-TB

Definition: Resistance to at least rifampicin AND isoniazid (the two most important first-line drugs)

Causes: - Incomplete/irregular treatment (most common) - Poor drug supply and quality - Non-adherence (patient) - Inadequate supervision - Transmission of already-resistant strains

Diagnosis: - Culture + phenotypic DST - Xpert MTB/RIF (detects RIF resistance) - LPA (detects RIF + INH resistance)

Treatment: Longer (18–24 months), second-line drugs (fluoroquinolones, bedaquiline, linezolid, clofazimine)

Prevention & Control of TB

Vaccination: - BCG (Bacille Calmette-Guérin): Given at birth; protects against severe forms (miliary TB, TB meningitis) in children; variable efficacy against pulmonary TB

DOTS (Directly Observed Treatment, Short-course): - WHO End TB Strategy framework - 6-month regimen: 2HRZE/4HR - H = Isoniazid, R = Rifampicin, Z = Pyrazinamide, E = Ethambutol - Patient-centered care; treatment supporters

DOTS-Plus: For MDR-TB

Infection Control: - Respiratory isolation of infectious cases - N95 respirators for healthcare workers - Adequate ventilation (natural/mechanical), UV germicidal irradiation - Negative pressure isolation rooms

Contact Tracing: - Screen all household contacts - TST/IGRA for contacts - Preventive therapy (LTBI treatment) for high-risk contacts

LTBI Treatment (Chemoprophylaxis): - Isoniazid 6–9 months (INH preventive therapy — IPT) - For: HIV+, children <5 years in contact with PTB, TST/IGRA+ with risk factors

Notification and Surveillance: All TB cases must be notified

QUICK EXAM CHEAT SHEET

Definitive Diagnostic Tests by Disease

Disease Gold Standard / Definitive Test
Typhoid Blood culture (Week 1); Bone marrow culture (most sensitive overall)
Campylobacter Culture on Skirrow’s agar at 42°C (microaerophilic)
Kala-azar Splenic aspirate → amastigotes (95–98%); rK39 RDT (field)
Pneumococcal pneumonia Sputum culture + optochin sensitivity; Quellung reaction
Hepatitis A Anti-HAV IgM (acute); EM (virus particles)
Rotavirus ELISA (VP6 antigen); EM (wheel morphology)
Tuberculosis Culture (LJ/MGIT); Xpert MTB/RIF (rapid + RIF resistance)

Selective Media — KEY for Exams

Organism Selective Medium
Vibrio cholerae TCBS agar, Alkaline peptone water (APW)
Salmonella/Shigella SS agar, DCA, Selenite F (enrichment)
Campylobacter Skirrow’s agar / CCDA (42°C, microaerophilic)
S. pneumoniae Blood agar (5% sheep blood)
Mycobacterium LJ medium (solid); MGIT (liquid)
Legionella BCYE agar
N. gonorrhoeae Modified Thayer-Martin (MTM)
Leishmania NNN medium

Vaccines Summary

Disease Vaccine Type Schedule
Typhoid Vi polysaccharide / Ty21a Killed/Live attenuated Single dose / 3 oral doses
Rotavirus Rotarix / RotaTeq Live attenuated 2 doses / 3 doses (infants)
Pneumonia PCV13 / PPSV23 Conjugate / Polysaccharide EPI (children) / adults
Hepatitis A Inactivated HAV Killed 2 doses (0, 6–12 months)
TB BCG Live attenuated Birth (single dose)
Kala-azar None (no licensed vaccine) — —

Very Short Answer Bank (Group C ready)

Question Quick Answer
Vectors of filariasis in Nepal Culex quinquefasciatus, C. tritaeniorhynchus
Vector of Kala-azar Phlebotomus argentipes
Mode of transmission of HAV Fecal-oral (contaminated water/food)
Mode of transmission of JE Culex mosquito bite
Cold agglutinins associated with Mycoplasma pneumoniae pneumonia
Selective medium for Campylobacter Skirrow’s agar / CCDA at 42°C
NSP4 in Rotavirus Viral enterotoxin causing secretory diarrhea
MDR-TB definition Resistance to rifampicin AND isoniazid
Definitive medium for Leishmania NNN medium
Anti-HAV IgM = Acute HAV infection
Anti-HAV IgG only = Past infection or vaccinated
PKDL = Post Kala-azar Dermal Leishmaniasis
Ghon complex = Primary focus + hilar lymph node in primary TB
Optochin sensitivity = Streptococcus pneumoniae
Quellung reaction used for Capsular serotyping of S. pneumoniae
Caseous necrosis seen in Tuberculosis granuloma
ATP bioluminescence measures Relative Light Units (RLU) — organic contamination
Rotavirus dsRNA segments 11 segments
RDT for VL rK39 ICT

ALL THE BEST, KALOKAJI. GO SMOKE THAT EXAM! 🎤