Q80. The figure shows the profiles of quantitative real–time PCR (qRT–PCR) tests
for SARS–CoV–2 conducted on the throat swab samples of three individuals
(X, Y and Z). Tests were carried out under identical conditions. Dotted line
represents the threshold fluorescent value. Identify the correct statement on
the status of the COVID–19 tests of the individuals based on their qRT–PCR
profiles.
(A) X and Y are negative; Z is positive
(B) X and Y are positive; There is no apparent difference in their viral load
(C) X and Y are positive; X has the highest viral load
(D) X and Y are positive; Y has the highest viral load

Interpreting qRT-PCR Profiles for SARS-CoV-2: Correct Answer and Detailed Analysis

The figure displays qRT-PCR amplification curves (dotted lines) for three individuals’ COVID-19 tests, with solid lines as threshold fluorescence levels. Option C correctly identifies X and Y as positive with X having the highest viral load based on curve steepness and earlier threshold crossing.​

qRT-PCR Basics

Quantitative real-time PCR (qRT-PCR) detects SARS-CoV-2 RNA by measuring fluorescence from probes during amplification cycles. The cycle threshold (Ct) marks where fluorescence exceeds the baseline threshold, indicating viral presence; lower Ct means higher initial viral load due to exponential amplification.​
Positive tests show sigmoid curves crossing the threshold; negatives stay flat below it.​

Option Analysis

• A: X and Y negative; Z positive; no apparent difference in viral load: Incorrect. X and Y cross the threshold (positive), while Z likely does not (negative). Viral load differs by crossing timing.​

• B: X and Y positive; Z negative; no apparent difference: Incorrect. X and Y are positive with differing curve slopes—X steeper (lower Ct, higher load); Y shallower.​

• C: X and Y positive; Z negative; X has highest viral load: Correct. Both X and Y cross threshold (positive), Z does not (negative). X’s rapid rise indicates earliest/lowest Ct and highest viral load; Y slower rise means moderate load.​

• D: X and Y negative; Z positive; X highest viral load: Incorrect. X and Y cross threshold (positive), Z flat (negative).​

qRT-PCR profiles for SARS-CoV-2 revolutionized COVID-19 testing by quantifying viral RNA through real-time fluorescence monitoring. These profiles reveal test status and viral load, crucial for diagnosis and management.​

Understanding Amplification Curves

Sigmoid curves in qRT-PCR plots show fluorescence buildup over cycles. Curves crossing the threshold indicate positives; flat lines below signify negatives. Steeper curves with earlier threshold crossing denote higher viral loads via lower Ct values.​

• Ct inversely correlates with viral RNA amount—low Ct (e.g., <25) signals high infectivity risk.​

• Threshold crossing timing differentiates loads among positives.​

Viral Load and Clinical Implications

Highest viral load (steepest curve, lowest Ct) links to greater transmissibility, unlike moderate or low loads (later crossing).​
Factors like sample quality affect curves, but patterns reliably distinguish statuses.

This qRT-PCR profiles SARS-CoV-2 analysis aids CSIR NET aspirants in mastering molecular diagnostics.​