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🔵 IMMUNOLOGY (I)

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🔴 RESPIRATORY SYSTEM (RS)

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🟢 CNS (Central Nervous System)

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🟠 STD (Sexually Transmitted Diseases)

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🟣 ZOONOSIS

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🔶 UTI (Urinary Tract Infection)

Note: The question list does not have dedicated UTI questions explicitly. The above are the closest associated organisms. If your syllabus has a separate UTI chapter covering E. coli UTI, Klebsiella, Proteus, etc., those would fall here - but they aren't listed as standalone questions in this set.

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⚪ REMAINING / GENERAL (R)

Bacteriology - Gram-positive organisms

Bacteriology - Gram-negative / Enteric

Mycobacteriology (non-RS overlap)

Mycology

Miscellaneous Bacteriology

Virology - General

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• Immunology: 12 questions
• Respiratory System: 25 questions
• CNS: 5 questions
• STD: 14 questions
• Zoonosis: 23 questions
• UTI: 3 questions (closely associated)
• Remaining (General Bacteriology, Virology, Mycology): ~94 questions

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RESPIRATORY SYSTEM (RS)

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5 & 55 & 56. Whooping Cough / Bordetella pertussis - Virulence Factors & Vaccines

1. Pertussis Toxin (PT) - the major virulence factor. An A-B toxin that ADP-ribosylates a G protein, disrupting adenylate cyclase regulation. Causes lymphocytosis, sensitization to histamine, and impairs phagocyte function.
2. Filamentous Hemagglutinin (FHA) - primary adhesin; binds ciliated respiratory epithelium via integrin sequences. Also stimulates cytokine release and interferes with TH1 immune responses.
3. Pertactin - outer membrane protein; adhesin.
4. Pili (Fimbriae) - adhesins contributing to attachment.
5. Tracheal Cytotoxin (TCT) - a disaccharide-tetrapeptide fragment of peptidoglycan. Destroys ciliated epithelial cells, producing the characteristic ciliary paralysis and cough.
6. Adenylate Cyclase Toxin - enters host cells, produces supraphysiologic cAMP levels, inhibits phagocyte killing.
7. Dermonecrotic Toxin - causes local tissue necrosis.
8. Lipopolysaccharide (LPS) - structurally distinct from Enterobacteriaceae LPS; contributes to inflammation.

• Catarrhal stage (1-2 weeks): Mild URI symptoms, highly infectious.
• Paroxysmal stage (2-4 weeks): Repeated bursts of coughing (paroxysms) followed by inspiratory "whoop"; post-tussive vomiting; lymphocytosis.
• Convalescent stage (weeks to months): Gradual recovery.

• Nasopharyngeal swab/aspirate - best specimen
• Culture on Bordet-Gengou (potato-glycerol-blood agar) or Regan-Lowe medium
• PCR - most sensitive
• DFA (Direct Fluorescent Antibody)
• Serology (paired IgG/IgA)
• Characteristic lymphocytosis on CBC (unique - lymphocytes are the predominant cell, unlike bacterial infections that show neutrophilia)

1. Whole-cell pertussis vaccine (wP): Part of DPT/DTP. Highly effective but causes more febrile reactions.
2. Acellular pertussis vaccine (aP): Contains purified PT, FHA, pertactin, and fimbriae. Used in DTaP. Fewer side effects but shorter-lived immunity (10-12 years).
3. Schedule: 6 weeks, 10 weeks, 14 weeks (primary), boosters at 18 months, 5 years (DPT). Tdap booster recommended for adolescents and pregnant women.

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20 & 154. Laboratory Diagnosis of Pulmonary TB / Tuberculosis

• Sputum (early morning, 3 consecutive days)
• BAL, bronchial washings
• Gastric lavage (children, who swallow sputum)
• Pleural fluid, CSF, urine, tissue biopsies

• Decontamination with N-acetyl-L-cysteine (NALC) + NaOH (Petroff's method)
• Concentration by centrifugation

• Ziehl-Neelsen (ZN) stain: AFB appear red on blue background
• Auramine-rhodamine fluorescence stain: more sensitive, AFBs fluoresce yellow-green
• Sensitivity: ~40-60%; requires 10,000 bacilli/mL

• Lowenstein-Jensen (LJ) medium - egg-based solid medium; colonies in 4-8 weeks; rough, buff-coloured, cauliflower-like "eugonic" colonies
• Middlebrook 7H10/7H11 - agar-based; 3-4 weeks
• BACTEC MGIT 960 (liquid) - fastest; results in 1-3 weeks; detects fluorescence due to O2 consumption
• MODS (Microscopic Observation Drug Susceptibility) - cord formation visible

• Niacin production test - positive in M. tuberculosis
• Nitrate reduction - positive
• Catalase test - weakly positive; heat-labile (negative at 68°C)
• Slow growth (>7 days), non-pigmented

• GeneXpert MTB/RIF (cartridge-based NAAT) - detects MTB and rifampicin resistance in 2 hours. WHO-endorsed first-line test.
• Line probe assay (LPA/Hain test) - detects resistance mutations
• Whole genome sequencing

• Mantoux (TST): 0.1 mL PPD (5 TU) intradermal; read at 48-72 hrs; induration ≥10 mm = positive (≥5 mm in immunocompromised)
• IGRA (QuantiFERON-TB Gold, T-SPOT.TB) - measures IFN-γ release to ESAT-6 and CFP-10; not affected by BCG vaccination

• Sputum smear microscopy at designated microscopy centres
• GeneXpert as initial test for all presumptive TB patients (current policy)
• CBNAAT (Cartridge-Based NAAT) for drug-resistant TB
• Culture and DST for MDR-TB confirmation

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47. Mantoux Test

• 0.1 mL of PPD (5 tuberculin units) injected intradermally on the volar surface of the left forearm
• A wheal of 6-10 mm should form immediately
• Read at 48-72 hours
• Measure induration (not erythema) in millimetres

• ≥10 mm: Positive (general population)
• ≥5 mm: Positive in HIV-positive, recent TB contacts, immunosuppressed, abnormal CXR
• ≥15 mm: Positive in low-risk persons

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48. Multidrug-Resistant Tuberculosis (MDR-TB)

• Inadequate/interrupted treatment
• Poor compliance
• Subtherapeutic drug levels
• Monotherapy (adding one drug to a failing regimen)
• Poor quality drugs
• Primary transmission of resistant strains

• GeneXpert MTB/RIF: detects rifampicin resistance (proxy for MDR-TB) in 2 hours
• Line Probe Assay (LPA): detects INH and RIF resistance mutations
• Culture + Drug Susceptibility Testing (DST): gold standard
• Whole genome sequencing

• Bedaquiline + Pretomanid + Linezolid (6 months) for XDR-TB and treatment-intolerant MDR-TB
• Shorter MDR-TB regimen: 9-12 months with bedaquiline, moxifloxacin, clofazimine, pyrazinamide, etc.

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46 & 157. Atypical Mycobacteria (Non-Tuberculous Mycobacteria - NTM)

• Pulmonary disease (MAC, M. kansasii)
• Lymphadenitis in children (MAC, M. scrofulaceum)
• Skin/soft tissue (M. marinum, M. ulcerans - Buruli ulcer)
• Disseminated disease in AIDS (MAC)
• Catheter-related infections (M. fortuitum)

• Not transmitted person-to-person
• Environmental organisms (soil, water)
• Less virulent
• Niacin negative, catalase strongly positive
• Often resistant to standard anti-TB drugs

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49. Cultivation of Mycobacterium leprae

1. Armadillo model (Nine-banded armadillo - Dasypus novemcinctus):

• Best animal model; produces massive systemic infection (10^10-10^11 bacilli/gram of tissue)
• Used for antigen preparation and drug testing
• Armadillos have low body temperature (32-35°C), ideal for M. leprae

1. Mouse footpad (Shepard's technique - 1960):

• 5,000-10,000 organisms injected into mouse footpad
• Limited multiplication (max 10^6 bacilli) - no dissemination in normal mice
• Used for drug susceptibility testing and vaccine trials
• Nude (athymic) mice allow greater multiplication

1. Cell cultures:

• Limited growth in macrophages and Schwann cells in vitro

• Lacks many metabolic enzymes (incomplete TCA cycle, no purine synthesis)
• Dependent on host for metabolites
• Optimal temperature: 27-30°C (explains tropism for cooler body parts - skin, peripheral nerves, testes)
• Extremely slow generation time (~12-14 days)

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50 & 3. Lepromin Test

• Dharmendra lepromin: Suspension of killed M. leprae (chloroform-ether treated)
• Mitsuda lepromin: Heat-killed autoclaved M. leprae in whole tissue homogenate

1. Early (Fernandez) reaction: Read at 24-48 hours. Erythema and induration = DTH to antigens. Seen in tuberculoid; negative in lepromatous.
2. Late (Mitsuda) reaction: Read at 3-4 weeks. Nodule or granuloma formation = CMI response. Positive = papule >5 mm.

• Strongly positive (3+): Tuberculoid leprosy (strong CMI)
• Weakly positive (1-2+): Borderline forms
• Negative: Lepromatous leprosy (absent CMI), or in 40% of normal healthy persons without leprosy exposure

• NOT a diagnostic test for leprosy
• Indicates immune status / prognosis
• Helps classify leprosy along the spectrum
• Used in epidemiological studies (population CMI)

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51 & 159. Lab Diagnosis of Mycobacterium leprae / Leprosy

• Modified Ziehl-Neelsen (MZN) or Fite-Faraco stain (uses weaker decolorizer - peanut oil + xylene)
• M. leprae appears as solid-staining, beaded, or granular AFBs (dead bacilli are fragmented/granular)
• Bacteriological Index (BI): Log scale (0-6+) of bacillary load per oil-immersion field
• Morphological Index (MI): % of solid-staining (viable) bacilli; indicates treatment response

• Tuberculoid: Epithelioid granulomas around nerve twigs; few/no bacilli; Langhans giant cells
• Lepromatous: Foamy macrophages (Virchow cells/lepra cells) packed with bacilli; no granulomas; "Grenz zone" (clear subepidermal band)

• Anti-PGL-1 (phenolic glycolipid-1) IgM antibodies: elevated in lepromatous leprosy (high bacillary load); poor sensitivity in paucibacillary disease
• ML Flow test (rapid lateral flow)

• PCR for M. leprae-specific 18-kDa protein gene or RLEP sequence
• Useful in early/paucibacillary cases with low BI

• Paucibacillary (PB): 1-5 patches, BI = 0; treat with Rifampicin + Dapsone x 6 months
• Multibacillary (MB): >5 patches or BI ≥1; treat with Rifampicin + Clofazimine + Dapsone x 12 months

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53 & 143 & 149. Haemophilus influenzae - Lab Diagnosis

• Encapsulated (typeable): Types a-f based on capsular polysaccharide. Type b (Hib) - polyribosylribitol phosphate (PRP) capsule - most virulent; causes meningitis, epiglottitis, cellulitis, septic arthritis, pneumonia.
• Non-typeable (NTHi): No capsule; causes otitis media, sinusitis, conjunctivitis, exacerbations of COPD, community-acquired pneumonia.

1. Specimens: CSF, blood, sputum, throat swab, middle ear fluid
2. Smear: Gram-negative coccobacilli; pleomorphic
3. Culture:

• Chocolate agar (provides both X and V factors)
• Blood agar alone - does NOT support growth
• Satellitism test: H. influenzae colonies grow around S. aureus streaks on blood agar (S. aureus hemolyzes RBCs releasing both factors)

1. Factor requirement test:

• X + V factors disk: both required for growth = H. influenzae

1. Quellung reaction: Capsule swelling with type-specific antisera
2. Latex agglutination / CIE: Rapid antigen detection in CSF/blood
3. PCR: Highly sensitive
4. Optochin resistance: Distinguishes from pneumococci

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Legionella pneumophila - Lab Diagnosis (Q148)

• Legionnaires' disease: Severe pneumonia; incubation 2-10 days; high fever, dry cough, confusion, diarrhoea, hyponatraemia (characteristic).
• Pontiac fever: Self-limiting flu-like illness; no pneumonia.

1. Specimens: Sputum, BAL, pleural fluid, lung biopsy
2. Staining: Gram stain often negative or shows faint Gram-negative rods. Dieterle silver stain or Gimenez stain shows organisms in tissue.
3. Culture: BCYE (Buffered Charcoal Yeast Extract) agar + L-cysteine + alpha-ketoglutarate + iron; takes 3-5 days; blue-grey colonies with ground-glass appearance; growth on BCYE but NOT on blood agar.
4. Urinary Antigen Test (UAT): Detects L. pneumophila serogroup 1 antigen; rapid (2-3 hours); highly specific; positive for weeks even after antibiotics started. Test of choice for rapid diagnosis.
5. Direct Fluorescent Antibody (DFA): On respiratory secretions; rapid but less sensitive.
6. Serology: 4-fold rise in IFA titre ≥1:128; retrospective diagnosis.
7. PCR: Highly sensitive and specific; detects all serogroups.

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150. Lab Diagnosis of Whooping Cough (see Q5 above for more detail)

1. Culture: Bordet-Gengou medium (potato-glycerol-blood agar) or Regan-Lowe transport/culture medium. Colonies in 3-7 days: small, pearly, "mercury drop" appearance with a zone of beta-hemolysis. Confirmation by agglutination with specific antisera.
2. PCR: Most sensitive; gold standard; particularly useful in vaccinated individuals.
3. DFA: On nasopharyngeal swabs; rapid; requires expertise.
4. CBC: Marked lymphocytosis (absolute lymphocyte count >10,000/μL) - virtually pathognomonic during paroxysmal stage.
5. Serology: Anti-PT IgA or IgG; useful in adults and partially vaccinated. Single high IgA titre is diagnostic in adults.

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165. Mycoplasma pneumoniae

• Smallest self-replicating organism (0.2-0.3 μm)
• NO cell wall - therefore resistant to all beta-lactam antibiotics
• Bounded only by a triple-layered plasma membrane
• Pleomorphic (no fixed shape)
• Facultative anaerobe
• Very slow growth: 1-3 weeks on SP4 or Hayflick's medium
• Ferments glucose; does NOT hydrolyze arginine

• Tip organelle (P1 adhesin protein) attaches to sialic acid receptors on respiratory epithelium
• Produces H2O2 and superoxide - causes ciliostasis and epithelial damage
• Causes "atypical pneumonia" (walking pneumonia)

• Gradual onset; persistent dry hacking cough; headache; fever
• CXR shows patchy interstitial infiltrates (worse than clinical picture)
• Extrapulmonary manifestations: haemolytic anaemia (cold agglutinins), erythema multiforme/Stevens-Johnson syndrome, encephalitis, myocarditis, pericarditis, arthritis

1. Cold agglutinins: Non-specific IgM antibodies against I antigen on RBCs; titre ≥1:64 or 4-fold rise; positive in 50-75%; classic "cold agglutinin" test (4°C agglutination reverts on warming).
2. Culture: On SP4 or Hayflick's agar (horse serum + yeast extract); 1-3 weeks; "fried egg" colonies.
3. Serology: Complement fixation (CF) or ELISA for anti-Mycoplasma IgM/IgG.
4. PCR: Most sensitive and specific; rapid; method of choice.

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88. Severe Acute Respiratory Syndrome (SARS)

• Spike protein (S protein) binds ACE2 (angiotensin-converting enzyme 2) receptor on type II pneumocytes
• Viral replication causes diffuse alveolar damage (DAD)
• Cytokine storm contributes to ARDS

• Incubation: 2-10 days
• Prodrome: Fever >38°C, malaise, myalgia, headache
• Respiratory phase: Dry cough, dyspnoea → ARDS in ~20%
• Diarrhoea in ~25%
• CXR: Interstitial infiltrates, ground-glass opacities

• RT-PCR (gold standard): Nasopharyngeal swab, stool
• Serology: ELISA/IFA for IgM/IgG (retrospective)
• SARS-CoV detection in Vero cell culture (BSL-3 required)
• CBC: Lymphopenia, thrombocytopenia, elevated LDH/ALT

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155. Diagnosis of TB Under RNTCP / NTEP

1. All presumptive TB patients - GeneXpert/CBNAAT as initial test (WHO recommendation)
2. If GeneXpert positive - treat for TB; check rifampicin sensitivity
3. If RIF resistant - perform Line Probe Assay (LPA) for first-line and second-line drug resistance
4. Culture + DST at RNTRL (Revised National Tuberculosis Reference Laboratory) for confirmation of MDR/XDR

• Presumptive TB: Any person with cough ≥2 weeks, fever, weight loss, night sweats, haemoptysis
• Bacteriologically confirmed: Positive by smear, culture, or NAAT
• Clinically diagnosed: Diagnosed by clinician without bacteriological confirmation

• New cases: 2HRZE/4HR (isoniazid, rifampicin, pyrazinamide, ethambutol)
• Previously treated: Culture + DST first; then tailored regimen

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156. Concentration Techniques for Sputum

1. Petroff's method (standard):

• Sputum + equal volume of 4% NaOH
• Mix, stand 20 minutes at room temperature
• Centrifuge 3000 rpm x 20 minutes
• Discard supernatant; neutralise sediment with HCl
• Smear from sediment

1. NALC-NaOH method (preferred):

• N-acetyl-L-cysteine (NALC) - mucolytic agent
• NaOH - decontaminant
• Less toxic to mycobacteria than Petroff's
• Centrifuge; wash with phosphate buffer
• Sediment used for smear, culture, and molecular tests

1. Cetylpyridinium chloride (CPC) + NaCl: Used for transport specimens
2. Bleach (sodium hypochlorite) method: Low-cost field method for resource-limited settings; kills the organisms but allows smear microscopy on killed, concentrated bacilli.

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CNS (Central Nervous System)

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58. Negri Bodies

• Round or oval, eosinophilic (pink) intracytoplasmic inclusions
• 2-10 μm in diameter (may reach up to 27 μm)
• Contain basophilic inner granules (Lyssa bodies)
• Hematoxylin and eosin (H&E) stain: pink inclusions with basophilic inner granules

• Most commonly in hippocampus (Ammon's horn) and Purkinje cells of cerebellum
• Also found in cortical neurons, brainstem neurons

• Found in 70-90% of confirmed rabies cases
• Absence does NOT rule out rabies
• Corneal impression smears (Sellers stain) was used for rapid antemortem diagnosis
• Now replaced by more sensitive tests (immunofluorescence, PCR)

• Intranuclear: Negri bodies are intracytoplasmic; Cowdry A bodies (HSV, CMV) are intranuclear
• Babes-Ernst granules (polar granules): Seen in Corynebacterium diphtheriae - different organism

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60 & 81 & 103. Polio / Prophylaxis / Lab Diagnosis / Pulse Polio Immunization

• Faecal-oral transmission
• Replication in oropharynx and gut (Peyer's patches, tonsils)
• Viraemia → spread to CNS via blood or retrograde axonal transport
• Destruction of anterior horn cells (lower motor neurons) of spinal cord → flaccid paralysis
• 90-95%: Asymptomatic
• 4-8%: Abortive polio (minor illness - fever, sore throat, vomiting)
• 1-2%: Non-paralytic (aseptic meningitis)
• <1%: Paralytic poliomyelitis

1. Specimens: Stool (2 samples 24 hours apart within 14 days of onset), CSF, throat swab
2. Virus isolation: Inoculation onto RD cells (rhabdomyosarcoma), L20B cells (transgenic expressing poliovirus receptor - human CD155); cytopathic effect (CPE) observed
3. Intratypic differentiation (ITD): Real-time RT-PCR to distinguish wild poliovirus (WPV) from vaccine-derived poliovirus (VDPV) and Sabin strains
4. Sequencing: Full VP1 sequencing - gold standard to distinguish WPV1, WPV3, cVDPV
5. Serology: 4-fold rise in neutralising antibody titre (retrospective)

1. OPV (Oral Polio Vaccine / Sabin vaccine): Live attenuated; trivalent (tOPV) or bivalent (bOPV - types 1 and 3). Given orally. Stimulates secretory IgA + humoral immunity. Can spread to contacts (herd immunity). Risk of vaccine-associated paralytic poliomyelitis (VAPP): 1 per 2.5 million doses.
2. IPV (Inactivated Polio Vaccine / Salk vaccine): Formalin-killed; given IM. No risk of VAPP. Does not produce secretory IgA (gut immunity). Used in high-income countries. India introduced IPV in 2015.
3. Schedule in India: OPV at birth, 6, 10, 14 weeks, 16-24 months; IPV at 6 and 14 weeks.

• Strategy launched in India in 1995 under NPSP (National Polio Surveillance Project)
• National Immunization Days (NIDs): Two rounds of OPV given to ALL children <5 years on a single day, regardless of prior vaccination status
• Sub-National Immunization Days (SNIDs): In high-risk states
• Mop-up campaigns: In areas with confirmed or suspected wild poliovirus
• India declared polio-free by WHO on 27 March 2014 (3 years without wild poliovirus)
• Goal: Simultaneously immunize all children - achieves high coverage rapidly, interrupts transmission

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83. Coxsackie Virus

• Group A (Coxsackievirus A, 1-24): Produce diffuse myositis and flaccid paralysis in suckling mice; cause herpangina (A1-10, 22), HFMD (A16), acute haemorrhagic conjunctivitis (A24), aseptic meningitis.
• Group B (Coxsackievirus B, 1-6): Produce focal myositis, focal necrosis of CNS, heart, pancreas, fat in suckling mice; cause pleurodynia (Bornholm disease), myocarditis, pericarditis, meningoencephalitis, neonatal disease.

• Suckling mouse inoculation: Group A causes flaccid paralysis; Group B causes spastic paralysis
• Cell culture: Group B grows readily in Vero, HeLa cells; Group A is more fastidious
• RT-PCR: Gold standard; highly sensitive
• Serology: Neutralization test

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UTI (Urinary Tract Infections)

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118. MRSA (Methicillin-Resistant Staphylococcus aureus)

• mecA gene (carried on Staphylococcal Cassette Chromosome mec - SCCmec) encodes PBP2a
• PBP2a has very low affinity for all beta-lactams
• The cell wall precursor is still cross-linked but is not inhibited by beta-lactams

• Healthcare-associated MRSA (HA-MRSA): Nosocomial; causes wound infections, bacteraemia, UTI in catheterized patients, VAP, CLABSI
• Community-associated MRSA (CA-MRSA): In otherwise healthy persons; often carries Panton-Valentine Leukocidin (PVL); causes necrotizing skin/soft tissue infections, necrotizing pneumonia

• Causes UTI primarily in hospitalized, catheterized patients
• Less common UTI pathogen than Enterobacteriaceae but significant in ICU patients

1. Oxacillin disk (1 μg) on Mueller-Hinton agar: Inhibition zone <13 mm = resistant
2. Cefoxitin disk (30 μg) - surrogate test: Zone <22 mm (S. aureus) = MRSA; more reliable induction of mecA
3. MRSA CHROMagar: Mauve/pink colonies (selective and differential)
4. Latex agglutination: Detects PBP2a
5. PCR for mecA gene: Gold standard

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119. Coagulase-Negative Staphylococcus (CoNS)

• Normal skin flora (most common)
• Major cause of device-related infections (prosthetic valves, joint prostheses, catheters, pacemakers, VP shunts)
• Biofilm formation (slime) on surfaces - key virulence factor; protects from antibiotics and immune responses
• Causes UTI in immunocompromised and catheterized patients

• Second most common cause of UTI in young sexually active women (after E. coli)
• Resistant to novobiocin (distinguishing feature from other CoNS)
• Typically causes uncomplicated cystitis

1. Coagulase negative (tube and slide)
2. Novobiocin susceptibility: S. saprophyticus - resistant; S. epidermidis - sensitive
3. Optochin - no effect (distinguishes from Streptococcus pneumoniae)
4. Urease: S. saprophyticus - positive
5. Biofilm (slime) detection: Congo red agar; tube adherence test

• S. saprophyticus: Genuine UTI pathogen in young women
• S. epidermidis: Usually contaminant in urine culture; significant only with ≥10^5 CFU/mL from catheter specimens in appropriate clinical context

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147. Pseudomonas aeruginosa

• Gram-negative, non-fermenting, aerobic rod
• Motile (polar monotrichous flagellum)
• Oxidase positive; glucose oxidised (not fermented)
• Produces distinctive pigments:
• Pyocyanin (blue-green, diffusible) - inhibits other bacteria; only produced by P. aeruginosa
• Pyoverdin/fluorescein (yellow-green, fluorescent)
• Pyorubrin (red), pyomelanin (brown) - less common
• Produces fruity/grape-like odour (2-aminoacetophenone)
• Grows on MacConkey agar as pale (non-lactose-fermenting) colonies

• Exotoxin A - major toxin; ADP-ribosylates EF-2, inhibiting protein synthesis (same mechanism as diphtheria toxin)
• Elastase (LasB): Destroys IgG, IgA, complement; degrades elastin and collagen
• Alkaline protease: Degrades complement, fibronectin
• Phospholipase C: Destroys lecithin and lung surfactant
• Alginate (mucoid): Biofilm; protects from phagocytosis in cystic fibrosis
• Pili and flagella: Adhesion and motility
• Exoenzymes S, T, U, Y (type III secretion system): Directly injected into host cells
• Pyocyanin: Damages respiratory epithelium, impairs mucociliary function

1. Gram stain: Gram-negative rods
2. Culture: Grows on all media; greenish pigmentation of agar; characteristic smell
3. Oxidase positive (key characteristic)
4. Non-lactose fermenter on MacConkey
5. Growth at 42°C (distinguishes from Pseudomonas fluorescens)
6. Pigment production on King's A medium (pyocyanin) and King's B medium (pyoverdin)
7. Antibiotic susceptibility: Intrinsically resistant to many; test piperacillin-tazobactam, ceftazidime, cefepime, meropenem, amikacin, ciprofloxacin

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STD (Sexually Transmitted Diseases)

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1. LGV (Lymphogranuloma Venereum)

1. Primary stage: Small painless papule, pustule, or shallow ulcer at site of inoculation (genitals, rectum, oropharynx); heals in days without treatment; often unnoticed.
2. Secondary stage (Inguinal/Bubonic stage): Painful inguinal lymphadenopathy (buboes) 2-6 weeks later. "Groove sign" or "sign of the groove" - inguinal ligament separating inguinal and femoral nodes gives a groove-like appearance. Buboes may coalesce, soften, and rupture. Systemic symptoms: fever, chills, myalgia. In MSM: proctocolitis (rectal pain, discharge, bleeding, tenesmus) - "bubonulus" may not develop.
3. Tertiary/Late stage: Genital elephantiasis (esthiomene) - chronic fibrosis, lymphoedema of external genitalia. Rectal strictures and fistulae (especially in women and MSM). Frozen pelvis.

• NAATs (PCR): Most sensitive; specimen from genital ulcer, rectal swab, bubo aspirate, urine
• Culture: McCoy cells (intracellular); slow, requires BSL-2
• Microimmunofluorescence (MIF): High LGV titres >1:64 in secondary disease
• Serology: Complement fixation ≥1:64 or 4-fold rise
• Intradermal Frei test: Frei antigen (heat-killed Chlamydia); positive in secondary stage - obsolete

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2 & 40 & 163. VDRL Test / Standard Tests of Syphilis

Non-Treponemal Tests (NTT) - Detect reagin antibody (IgG + IgM against cardiolipin-lecithin-cholesterol antigen):

• Antigen: Cardiolipin (0.03%), lecithin (0.9%), cholesterol (0.5%) in buffered saline
• Patient serum inactivated at 56°C x 30 minutes
• Serum + antigen on slide → rotated → read microscopically
• Positive: Large/medium clumps (4+ to 2+)
• Weakly reactive: Small clumps
• Non-reactive: No clumps

Treponemal Tests (TT) - Detect specific anti-treponemal antibodies:

• Screening: RPR or VDRL
• If reactive: Confirm with FTA-ABS or TPPA
• If treponemal positive: Treat; NTT titres used to monitor treatment response (should fall 4-fold by 6-12 months)
• Treponemal tests remain positive for life (serofast)

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13. Pathogenicity of Gonorrhea

1. Pili (Type IV): Primary adhesins; mediate attachment to non-ciliated columnar epithelium; anti-phagocytic; undergo antigenic variation (pilE gene rearrangement)
2. Opa (opacity) proteins: OMP; mediate firm attachment; undergo phase and antigenic variation
3. PorB (Por/Porin): Outer membrane porin; inserts into phagocyte membrane, inhibiting phagolysosome fusion; allows intracellular survival
4. Rmp (reduction-modifiable protein): Inhibits bactericidal antibodies directed at PorB and LOS
5. LOS (Lipooligosaccharide): Endotoxin; triggers inflammatory cascade; no O-antigen (unlike Gram-negative rods)
6. IgA1 protease: Cleaves secretory IgA1 on mucosal surfaces
7. Transferrin and lactoferrin-binding proteins: Iron acquisition from host

• Attachment to columnar epithelium of urethra, endocervix, rectum, pharynx, conjunctiva via pili and Opa proteins
• Transcytosis through epithelial cells into subepithelial space
• PMN infiltration → purulent exudate
• Complement-mediated and PMN killing; but PorB prevents phagolysosomal killing
• LOS sialylation (acquires host sialic acid) makes it more resistant to complement

• Men: Urethritis (dysuria, profuse purulent discharge), epididymo-orchitis
• Women: Endocervicitis (often asymptomatic), PID, salpingitis, Fitz-Hugh-Curtis syndrome (perihepatitis)
• Both: Proctitis, pharyngitis, conjunctivitis, DGI (disseminated gonococcal infection) - septic arthritis, skin lesions, rarely endocarditis/meningitis

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54 & 144. Haemophilus ducreyi / Chancroid / Soft Sore

• Requires Factor X (hemin) but NOT Factor V
• "School of fish" or "railroad track" arrangement on Gram smear
• Fastidious; grows on GC-agar + 5% fetal calf serum or Mueller-Hinton agar + chocolatised horse blood + vancomycin at 33-35°C in 5% CO2

• Incubation: 3-7 days
• Primary lesion: Starts as erythematous papule → pustule → shallow, painful (soft) ulcer with ragged, undermined, non-indurated edges and dirty, necrotic base (unlike hard, indurated syphilitic chancre)
• "3 Ps": Painful, Purulent, Punched-out
• Multiple ulcers (autoinoculation) in ~50%
• Bubo: Painful inguinal lymphadenopathy in 50%; may suppurate and rupture (unilocular)
• Difference from syphilitic chancre: Chancroid ulcer is SOFT (non-indurated), PAINFUL, MULTIPLE; syphilitic chancre is HARD, PAINLESS, usually SINGLE

1. Smear: Gram smear of ulcer base - "school of fish" pattern (but sensitivity only 50%)
2. Culture: Fastidious; GC or chocolate agar + vancomycin; 3-7 days; small, non-mucoid, yellowish colonies; "gravel-like" texture; can be pushed intact across agar
3. PCR: Most sensitive and specific; multiplex PCR can detect H. ducreyi, T. pallidum, and HSV simultaneously
4. Serology: Not reliable

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44 & 45. Specific Tests of Treponema pallidum / Non-Venereal Treponematosis

Specific tests (Q44):

• Dark-field microscopy: Gold standard for PRIMARY syphilis ulcer (chancre). Motile spirochaetes with characteristic corkscrew motion and flexion. Cannot be used for oral lesions (normal oral treponemes are non-pathogenic but morphologically similar).
• Direct Fluorescent Antibody for T. pallidum (DFA-TP): Uses fluorescent-labelled anti-T. pallidum antibody on smear. More specific than dark-field; can be used on fixed smears; useful for oral lesions.
• Rabbit infectivity test (RIT): Gold standard for confirming viability of T. pallidum; not routine; research use.

Non-Venereal Treponematosis (Q45):

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175 & 176. Chlamydia trachomatis / Lab Diagnosis of Chlamydial Infections

• Elementary body (EB): Infectious, extracellular, metabolically inactive, rigid cell wall; 0.3 μm
• Reticulate body (RB): Replicating, intracellular, metabolically active; 1 μm; binary fission in inclusion; RBs condense into EBs and released by cell lysis

1. NAATs (PCR, TMA, SDA): Method of choice. Highly sensitive and specific. FDA-cleared for genital specimens; also validated for rectal and pharyngeal sites. Urine samples acceptable (especially men). Dacron or rayon swabs with plastic shaft preferred.
2. Cell culture: McCoy cells (treated with cycloheximide); 48-72 hours; inclusions stained with iodine (glycogen) or Giemsa; gold standard but insensitive (60-80%) and time-consuming. Required for medico-legal cases.
3. Direct Fluorescent Antibody (DFA): Fluorescent monoclonal antibody stains EBs in smear; rapid; requires expertise.
4. Antigen detection (EIA): Less sensitive than NAAT; acceptable for screening.
5. Serology: Microimmunofluorescence (MIF) - type-specific antibody titres; used for LGV and PID diagnosis; not for uncomplicated genital infections.
6. Giemsa stain: On conjunctival scrapings (trachoma); intracytoplasmic inclusions around nucleus; contains EBs and RBs.

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ZOONOSIS

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41 & 164. Weil's Disease / Leptospirosis

• Entry through skin abrasions → leptospiraemia → haematogenous spread
• Vasculitis of small blood vessels is the fundamental lesion
• Liver: Hepatocellular damage and intrahepatic cholestasis
• Kidney: Tubulointerstitial nephritis
• Lung: Pulmonary haemorrhage (ARDS - increasing recognition)
• Meninges: Aseptic meningitis

1. Mild (anicteric) form (90%): Abrupt fever, headache, myalgia (severe calf muscle pain - pathognomonic), conjunctival suffusion (not injection), nausea, vomiting; resolves in 1-2 weeks.
2. Severe (icteric/Weil's disease, 5-10%): Jaundice + renal failure + haemorrhage triad. High fever, profound jaundice (conjugated), AKI (non-oliguric initially), thrombocytopenia, haemorrhages (petechiae, ecchymoses, pulmonary haemorrhage), uveitis (late). CFR: 5-40%.
3. Pulmonary leptospirosis: Massive pulmonary haemorrhage + ARDS (ARDS from Leptospirosis - ARDS-L); high mortality.

1. Direct detection:

• Dark-field microscopy of blood/CSF/urine (first 10 days): Very low sensitivity, many false positives
• Culture: EMJH (Ellinghausen-McCullough-Johnson-Harris) medium; semi-solid; up to 13 weeks; BSL-2

1. Serology (main method):

• MAT (Microscopic Agglutination Test): Gold standard. Live leptospires + patient serum; titre ≥1:100 early, ≥1:400 confirmed; paired sera 4-fold rise. Serovar-specific.
• ELISA IgM: Most practical; positive from day 5-7; used for diagnosis and screening
• Rapid dipstick (Panbio Leptocheck): Point of care

1. PCR: Highly sensitive in leptospiraemic phase (first 5-7 days); blood, urine
2. CBC: Leucocytosis, thrombocytopenia
3. LFTs: Elevated bilirubin (predominantly conjugated), mildly elevated transaminases (AST/ALT usually <200 IU/L - lower than in viral hepatitis)
4. Urine: Haematuria, proteinuria, granular casts

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42 & 160. Lyme Disease

• USA: Ixodes scapularis (black-legged tick)
• Europe: Ixodes ricinus

• Erythema migrans (EM): Pathognomonic; bull's-eye/target lesion expanding from site of bite; ≥5 cm diameter; central clearing with expanding red border; painless; may have fever, fatigue, myalgia.

• Neurological (Lyme neuroborreliosis/Bannwarth syndrome): Cranial nerve palsy (facial nerve palsy, bilateral), lymphocytic meningitis, radiculopathy, encephalitis.
• Cardiac: AV block (varying degree), myocarditis, pericarditis.
• Multiple secondary EM lesions.

• Lyme arthritis: Migratory; large joints (knee >90%); episodes of swelling and pain; can become chronic.
• Chronic Lyme neuroborreliosis: Encephalopathy, polyneuropathy.
• Acrodermatitis chronica atrophicans (ACA): Mainly B. afzelii; skin atrophy on extremities.

1. Clinical diagnosis for early Lyme (EM rash in endemic area + tick bite history)
2. Serology (2-tier testing, CDC recommendation):

• First tier: ELISA for IgM/IgG to B. burgdorferi
• Second tier: Western blot (IgM: 2 of 3 bands; IgG: 5 of 10 specific bands)
• IgM peaks at 3-6 weeks; IgG persists for years

1. PCR: Synovial fluid for Lyme arthritis (most useful); CSF in neuroborreliosis
2. Culture: BSK-II medium; weeks; not practical

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43 & 161. Relapsing Fever

• Epidemic (LBRF): Human body louse (Pediculus humanus corporis); only B. recurrentis; man-to-man (no animal reservoir)
• Endemic (TBRF): Soft ticks (Ornithodoros species); multiple Borrelia species; reservoir in small rodents

• Borrelia undergoes programmed rearrangement of the surface lipoprotein (VMP - Variable Major Protein; also called Vsp and Vlp)
• The bacterium expresses a new antigen type during each relapse that escapes prior immunity
• Each febrile episode is caused by a new serotype
• 2-10 relapses may occur

• Sudden high fever (39-40°C), chills, headache, myalgia, nausea, hepatosplenomegaly
• Fever lasts 3-7 days → crisis (Jarisch-Herxheimer reaction when treated) → afebrile interval of 5-9 days → relapse
• Petechiae, jaundice in severe cases
• LBRF: More severe, higher mortality (up to 40% untreated), fewer relapses
• TBRF: Milder, more relapses

1. Blood smear (thick and thin): During febrile episode; Giemsa or Wright stain; extracellular spirochaetes visible; only reliable during febrile paroxysm (bacteraemia).
2. Dark-field microscopy: Blood; motile spirochaetes
3. Mouse inoculation: Blood inoculated IP into mice; spirochaetes detectable in tail blood in 3-5 days
4. PCR: Most sensitive; detects Borrelia DNA even in afebrile periods
5. Serology: Not specific; cross-reacts with other Borrelia species; not routinely useful

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6. Bubonic Plague

• F1 capsule antigen: Anti-phagocytic; expressed at 37°C
• V antigen (LcrV) and Yop proteins (Yersinia outer proteins): Type III secretion system; inject cytotoxic Yops into phagocytes
• Coagulase: Blocks flea proventriculus (flea becomes "blocked" - bites frantically, regurgitating bacilli)
• Plasminogen activator (Pla): Degrades complement; fibrinolysis; promotes dissemination
• Pesticin, purine synthesis: Iron acquisition systems
• Grows at both 28°C (ambient) and 37°C (mammalian body)

1. Bubonic plague (most common): Enlarged, painful, matted lymph nodes (buboes) - inguinal, axillary, or cervical; high fever; may progress to septicaemia if untreated. Mortality: 40-60% untreated; <5% treated.
2. Septicaemic plague: Bacteraemia without bubo; DIC; ecchymoses → "Black Death" (skin necrosis from DIC); very high mortality.
3. Pneumonic plague: Inhalation of infectious droplets (person-to-person); most dangerous form; bloody frothy sputum; CFR nearly 100% if untreated.

1. Specimens: Bubo aspirate (fluid), blood, sputum, CSF
2. Smear: Bipolar staining (Wayson/methylene blue stain) - "closed safety pin" appearance
3. Culture: On blood agar, MacConkey, BHIA; 48-72 hours; small, grey, "hammered metal" colonies; "stalactite growth" in broth
4. Fraction 1 (F1) antigen detection: ELISA, immunochromatographic strip (rapid diagnostic test)
5. PCR: Targets F1 antigen gene, pla, caf1 genes
6. Serology: HI test for anti-F1 antibodies (retrospective)
7. Biosafety: BSL-3 precautions required

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11. Wool Sorter's Disease (Anthrax)

• McFadyean's reaction: When blood/tissue smear from anthrax case stained with polychrome methylene blue, capsule appears as blue/pink amorphous material surrounding the dark-blue bacilli. The background stains blue - characteristic "purple-blue" haze. Used for rapid presumptive diagnosis.
• Malignant pustule: The cutaneous form of anthrax. Starts as painless papule → vesicle → central black necrotic eschar surrounded by non-pitting oedema and small satellite vesicles. Despite the name, it is neither malignant (a tumour) nor a pustule (contains clear fluid, not pus).

• Poly-D-glutamate capsule: Anti-phagocytic
• Anthrax toxin (encoded on pXO1 plasmid):
• Protective antigen (PA): Binds receptor; forms pore for toxin entry
• Lethal factor (LF): Zinc metalloprotease; cleaves MAPKK; causes macrophage/neutrophil lysis; PA + LF = Lethal toxin
• Oedema factor (EF): Adenylate cyclase; elevates cAMP; causes massive oedema; PA + EF = Oedema toxin

1. Cutaneous anthrax (most common, 95%): Wool sorter's disease-associated; painless malignant pustule; 20% mortality if untreated; responds well to antibiotics.
2. Inhalation anthrax (Wool sorter's disease, strictly): Inhalation of spores from infected animal hides/wool; mediastinal widening on CXR; haemorrhagic mediastinitis; meningitis; >80% mortality even with treatment.
3. GI anthrax: Ingestion of undercooked infected meat; oropharyngeal or intestinal form; haemorrhagic ascites.
4. Injection anthrax: IV drug users; reported in Europe.

1. Gram stain of blood/lesion: Large bamboo-jointed Gram-positive rods in chains; encapsulated in vivo (shown by India ink, M'Fadyean's stain)
2. Culture: Blood agar - dry, rough, non-haemolytic "Medusa head" colonies (comma-shaped projections from colony margin due to chains of bacteria); "beaten egg white" consistency
3. Capsule staining: India ink / M'Fadyean's polychrome methylene blue
4. Gammaphage lysis: Y-phage specific for B. anthracis (important identification test)
5. String of pearls test: B. anthracis grown in presence of low-dose penicillin → forms characteristic spheroplasts arranged in chains
6. Motility: Non-motile (unlike B. cereus, B. thuringiensis which are motile)
7. Haemolysis: Non-haemolytic (B. cereus is haemolytic)
8. PCR for pXO1/pXO2 plasmid genes
9. ELISA for anti-PA antibodies

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52 & 141 & 142. Melioidosis / Burkholderia pseudomallei

1. Pulmonary melioidosis (most common): Pneumonia - from mild to severe cavitating disease resembling TB; apical involvement; can mimic TB clinically and radiologically.
2. Septicaemic melioidosis: Bacteraemia; high mortality; multiorgan failure.
3. Localised infection: Skin ulcers, parotid abscesses (characteristic in children), prostatic abscesses.
4. Neurological melioidosis: Rare; encephalomyelitis.
5. Latent/recrudescent: Reactivation years later (in immunosuppressed).

1. Culture: Blood agar, Ashdown's medium (glycerol-crystal violet-gentamicin agar - selective); B. pseudomallei produces wrinkled, metallic, cream-coloured colonies ("creamy-yellowish, wrinkled"); distinctive earthy smell. Gram: Bipolar-staining Gram-negative rods.
2. Biochemistry: Oxidase positive; arginine dihydrolase positive; grows at 42°C; resistant to colistin (polymyxin B).
3. Serology: IHA (indirect haemagglutination), ELISA - useful in non-endemic areas; cross-reactions with Burkholderia mallei.
4. PCR: Highly sensitive; targets TTS1 (type III secretion system gene) or TTSS1.
5. Latex agglutination: Rapid antigen detection.

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151 & 152 & 153. Brucellosis / Lab Diagnosis / Undulant Fever

• Entry through mucous membranes or skin
• Ingested by macrophages in regional lymph nodes → bacteraemia
• Localises in reticuloendothelial organs (liver, spleen, lymph nodes, bone marrow)
• Lipopolysaccharide (smooth LPS) resists killing by reactive oxygen species
• Non-caseating granulomas form in organs

• Incubation: 2-4 weeks
• Acute brucellosis: Undulating fever (evening peaks), drenching night sweats, profound malaise, myalgia, arthralgia, headache, anorexia, weight loss
• Hepatosplenomegaly, lymphadenopathy
• Focal complications: Spondylitis (most common focal complication), sacroiliitis, epididymo-orchitis, neurobrucellosis, endocarditis (rare but most common cause of death)

1. Culture (definitive): Blood culture (BACTEC or lysis-centrifugation); hold for 4-6 weeks; BSL-3. Bone marrow culture - highest yield. Brucella grows slowly (3-5 days to 6 weeks).
2. Serum Agglutination Test (SAT): Standard tube agglutination (Wright's test); titre ≥1:80 significant; ≥1:160 or 4-fold rise diagnostic. Detects anti-smooth LPS antibodies (IgM, IgG, IgA). Used for B. abortus and B. melitensis (not B. canis which has rough LPS).
3. 2-Mercaptoethanol (2-ME) test: SAT + 2-ME breaks IgM (false-positive in early acute phase); remaining titre = IgG; ≥1:80 = chronic active infection.
4. Rose Bengal plate test: Rapid screening test; buffered antigen + serum on plate; positive = agglutination.
5. ELISA: IgM, IgG, IgA - sensitive; useful for seroepidemiological studies.
6. Brucellacapt (FELISA): For detecting blocking antibodies in prozone phenomenon.
7. PCR: Highly sensitive; targets IS711 or bcsp31 gene; useful in culture-negative cases.

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146. Lab Diagnosis of Plague (see Q6 above)

• Woolsorter's disease (anthrax) vs plague: Both have animal reservoir and vector; plague has insect vector; anthrax has environmental spore. Plague - flea bite; anthrax - skin contact/inhalation.
• Laboratory should be notified in advance - BSL-3 laboratory required for culture.
• Rapid dipstick test (RDT) for F1 antigen widely used in field settings.

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169. Rat Bite Fever

1. Streptobacillary rat bite fever (Haverhill fever)

• Transmission: Rat bite or scratch, OR ingestion of contaminated food/milk/water (Haverhill fever).
• Clinical: 3-10 day incubation; abrupt fever, chills, vomiting → within 2-4 days: migratory polyarthritis (large joints; characteristic), maculopapular rash (trunk, extremities, palms, soles). Can cause septic arthritis, endocarditis, meningitis, abscess.
• Diagnosis: Blood culture in broth (requires media without SPS anticoagulant which is inhibitory); culture on blood agar + 5-10% CO2; "puffball colonies." Serology: agglutinins.

2. Spirillar rat bite fever (Sodoku)

• Transmission: Rat or mouse bite ONLY (not from ingestion).
• Clinical: 1-4 week incubation; site of bite heals then re-ulcerates at relapse; relapsing fever pattern; regional lymphadenopathy; violaceous indurated rash. Arthritis is RARE (contrast with Streptobacillary form).
• Diagnosis: Dark-field microscopy of wound exudate or blood during febrile episode; mouse inoculation; Giemsa stain; PCR; VDRL often positive (BFP).

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171. Lab Diagnosis of Rickettsial Diseases

1. Weil-Felix test (Q.Tx.):

• Patient antibodies cross-react with Proteus antigens (OX-2, OX-19, OX-K)
• Non-specific but widely used historically

1. Specific serology:

• IFA (Indirect Immunofluorescence Assay): Gold standard; detects IgM and IgG to specific rickettsial antigens; single high titre or 4-fold rise confirms diagnosis
• ELISA: Used for large-scale screening
• Western Blot: Confirmatory for cross-reactive IFA results

1. Skin biopsy:

• Immunohistochemistry (IHC) stains rickettsia in endothelial cells of biopsy from eschar or rash
• Very specific; positive early in disease before serology

1. PCR:

• Blood, skin biopsy, eschar swab
• Most sensitive early in disease
• Targets 17-kDa protein gene, gltA (citrate synthase), ompA, ompB

1. Cell culture: BSL-3; Vero cells; slow (days to weeks); not routine

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172. Typhus Fevers

Epidemic (Louse-Borne) Typhus:

• Agent: Rickettsia prowazekii
• Vector: Human body louse (Pediculus humanus corporis) - human-to-human (no animal reservoir)
• Louse defaecates while feeding; rickettsiae in faeces enter through bite wound or mucous membranes
• Clinical: Abrupt fever, severe headache, myalgia, maculopapular rash (starts trunk → periphery, spares face, palms, soles); neurological symptoms (encephalitis, coma); typhus nodules; vasculitis; gangrene; Brill-Zinsser disease (recrudescence years later)
• CFR: 10-40% untreated
• Associated with war, famine, displacement, crowding

Murine (Endemic) Typhus:

• Agent: Rickettsia typhi (and R. felis)
• Vector: Rat flea (Xenopsylla cheopis) + cat flea (Ctenocephalides felis)
• Reservoir: Rats
• Clinical: Milder than epidemic; fever, headache, rash (more subtle, shorter duration); CFR <1%

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173. Scrub Typhus

• Incubation: 6-21 days
• Abrupt fever, severe headache, myalgia, malaise
• Eschar + regional lymphadenopathy
• Maculopapular rash (day 5-7)
• Hepatosplenomegaly
• Severe cases: Pneumonitis, myocarditis, meningoencephalitis, ARDS, MOF; CFR up to 30% untreated

• Weil-Felix: OX-K positive; OX-19 and OX-2 negative
• IFA: Gold standard; anti-Orientia IgM/IgG; serovar heterogeneity complicates testing
• PCR: Blood, eschar swab - most sensitive
• Immunohistochemistry on eschar biopsy
• Culture: BSL-3 (Vero cells)

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174. Q Fever

• ONLY rickettsial disease NOT transmitted by arthropod bite (no vector needed)
• Transmission: Inhalation of contaminated dust/aerosols from infected animals (parturient material, placenta, urine, faeces, milk of cattle, sheep, goats)
• Most contagious infectious agent known (single organism may cause infection)
• Weil-Felix test: NEGATIVE (no Proteus cross-reaction)
• No rash (unlike other rickettsial diseases)

• Phase I: Virulent; LPS with full-length O-antigen; found in nature/animals; antibodies to Phase I = chronic infection or past exposure
• Phase II: Avirulent; truncated LPS; grown in cell culture; antibodies to Phase II appear first in acute infection

1. Acute Q fever: Incubation 2-3 weeks. Self-limiting febrile illness; atypical pneumonia (dry cough, fever, minimal findings on auscultation but extensive radiological changes - "multiple rounded opacities"); hepatitis (granulomatous); rarely endocarditis.
2. Chronic Q fever: Primarily endocarditis (most common manifestation in pre-existing valve disease/prosthetic valves/immunosuppressed); osteomyelitis; hepatitis. Phase I IgG titre >1:800 = chronic Q fever.

1. Serology (IFA) - method of choice:

• Phase II IgM/IgG: Acute Q fever (Phase II IgG >1:200 or IgM >1:50 at 4 weeks)
• Phase I IgG >1:800: Chronic Q fever / endocarditis

1. PCR (real-time): Most sensitive in acute phase (first 2 weeks); blood, tissue
2. Culture: BSL-3 required (extremely hazardous); Shell vial culture, Vero cells
3. Histopathology: Liver biopsy - fibrin-ring granulomas ("doughnut granulomas") - pathognomonic

• Acute: Doxycycline 100 mg BD x 2-3 weeks
• Chronic/endocarditis: Doxycycline + hydroxychloroquine x 18-36 months (hydroxychloroquine alkalinises the phagolysosome, enhancing doxycycline activity); surgical valve replacement often needed

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107. Kyasanur Forest Disease (KFD)

• Incubation: 3-8 days
• Phase 1 (1-2 weeks): Abrupt fever, chills, headache, myalgia, bleeding manifestations (epistaxis, gingival bleeding, haematemesis, melaena, haematuria), conjunctival suffusion, papulovesicular lesions on soft palate
• **Remission (1-2 weeks)
• Phase 2 (neurological phase, in 30%): Fever returns, meningoencephalitis, tremors, severe headache; haemorrhagic features
• CFR: 2-10%

1. RT-PCR: Blood during febrile phase; most sensitive and specific
2. Virus isolation: Vero cells; BSL-4 (extremely hazardous)
3. Serology: ELISA for IgM/IgG; IgM detectable from day 5; plaque reduction neutralisation test (PRNT) - gold standard serological test
4. Antigen detection: ELISA for NS1 antigen

• KFD vaccine (inactivated): Formalin-inactivated KFDV; given to people in endemic areas; 2-dose primary series + annual booster
• Personal protection: Protective clothing, acaricides, tick checks

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Summary Table