Direct RT-PCR Testing For COVID-19 Without RNA Extraction: A Rapid And Cost-Effective Diagnostic Approach

Abstract

The COVID-19 pandemic created an urgent demand for rapid, scalable, and cost-effective diagnostic methods. Real-time reverse transcription polymerase chain reaction (RT-PCR) is considered the gold standard for SARS-CoV-2 detection but requires RNA extraction—a resource-intensive and time-consuming step. Direct RT-PCR, which bypasses RNA extraction, offers a potential alternative for timely and affordable diagnostics, especially in resource-limite settings.Methods:This experimental study compares the diagnostic performance of direct RT-PCR with traditional RT-PCR (including RNA extraction) using 300 clinical samples. The direct RT-PCR protocol involved sample lysis at 65°C followed by amplification targeting SARS-CoV-2 genes. Diagnostic accuracy was evaluated in terms of sensitivity, specificity, positive predictive value (PPV), negative predictive value (NPV), and overall accuracy. Cost and time analyses were also conducted.Results:In a representative dataset of 10 samples, direct RT-PCR achieved 60% sensitivity, 60% specificity, 60% PPV, and 60% NPV, with an overall diagnostic accuracy of 60% when compared to traditional RT-PCR. The method reduced per-test costs by approximately 65–70% and turnaround time by more than half (1.5–2 hours vs. 4–6 hours). Visualization tools—including confusion matrices and performance bar charts—highlighted areas of agreement and discrepancy between methods.Conclusion:Direct RT-PCR presents a promising, lower-cost, and faster alternative to traditional RT-PCR, with potential applications in large-scale screening and in settings where resources are limited. Although its diagnostic performance is moderate, further optimization and validation could enhance its accuracy and utility. Integration into public health frameworks may facilitate improved diagnostic reach and responsiveness during infectious disease outbreaks.