Viral diagnostics

Direct Detection - Seeing is Believing

• Microscopy:

  • Light Microscopy: Identifies viral inclusion bodies.
    • Intranuclear: Owl's eye (CMV), Cowdry type A (HSV, VZV).
    • Intracytoplasmic: Negri bodies (Rabies), Guarnieri bodies (Poxvirus).
  • Electron Microscopy (EM): Directly visualizes virions. Useful for viruses that cannot be cultured (e.g., Norwalk virus).

• Antigen Detection (ELISA, Immunofluorescence): Rapidly detects viral proteins using specific antibodies. Widely used for respiratory viruses (e.g., RSV, Influenza).

• Nucleic Acid Amplification Tests (NAATs):

  • PCR: Gold standard for diagnosis and management (e.g., HIV viral load). Offers highest sensitivity & specificity.

⭐ Owl's eye inclusions are pathognomonic for Cytomegalovirus (CMV) infection. They are large, eosinophilic, basophilic intranuclear inclusions surrounded by a clear halo.

Viral Culture - Growing the Enemy

• Gold standard for proving infectivity, but often slow & labor-intensive.
• Process: Live virus is grown in a monolayer of cultured cells.
• Detection: Observed for cytopathic effect (CPE) - visible structural changes in host cells.
  • Examples: Cell rounding, lysis, syncytia (RSV, measles), inclusion bodies (CMV, HSV).
• Limitation: Many viruses are difficult or impossible to culture (e.g., HCV, HPV).

⭐ Shell Vial Culture: A rapid modification using centrifugation and immunofluorescence to detect viral antigens within 24-48 hours, long before CPE appears.

NAATs - Copy & Conquer

Nucleic Acid Amplification Tests (NAATs) are molecular assays that amplify viral genetic material (DNA or RNA) to detectable levels. They offer high sensitivity and specificity for diagnosis.

• Core Principle: Exponentially copy a specific genetic sequence.
• Common Types:
  • PCR (Polymerase Chain Reaction): Amplifies DNA targets.
  • RT-PCR (Reverse Transcriptase PCR): For RNA viruses; first converts RNA to complementary DNA (cDNA).
  • TMA (Transcription-Mediated Amplification): Isothermal RNA amplification.

⭐ NAATs are crucial for diagnosing early HIV (window period) and for quantifying viral load in HIV/HCV to monitor treatment response.

Serology - The Immune Footprint

• Detects host antibodies (IgM, IgG) against viral antigens, not the virus itself. It signals an immune response, not necessarily active infection.
• IgM: Appears first; indicates acute or recent infection. Its presence suggests a primary infection.
• IgG: Appears later and persists, often for life; indicates past infection or immunity.
• Seroconversion: A ≥4-fold increase in IgG titers between acute and convalescent-phase serum samples is the gold standard for diagnosing a primary infection.
• Window Period: Time from infection until antibodies are detectable. Serology is negative.

⭐ For HIV, serology is the standard screening test (ELISA), but it's negative during the window period. Nucleic Acid Testing (NAT) is required for diagnosis during this early phase.

High‑Yield Points - ⚡ Biggest Takeaways

• NAATs (PCR) are the most sensitive and specific method for viral diagnosis.
• Serology showing IgM or a ≥4-fold rise in IgG confirms acute infection.
• Antigen detection (ELISA) provides rapid results but is less sensitive than NAATs.
• Viral culture is the traditional gold standard but is slow; observe for cytopathic effect (CPE).
• Microscopy for inclusion bodies (e.g., Owl's eye in CMV, Negri bodies in Rabies).
• Tzanck smear for HSV/VZV reveals multinucleated giant cells.