Evaluation of the Dilution Effect of Respiratory Specimen Pooling on the Sensitivity of SARS-COV 2 RT-PCR Tests
Year: 2025
Type: article
Abstract: Introduction: The Covid-19 pandemic affects worldwide health and causes economic disasters. Testing sample in pool is the best strategy for surveillance. Pooling of samples reduces cost and turn-around time of RT-PCR test if done in the setting of low prevalence. But at the same time, it may give false negative results due to dilution effect, especially in higher range of cycle threshold value (Ct value). So, it is recommended that every RT-PCR laboratory should optimize pool size for improving test output utilizing minimum resources and without compromising sensitivity of the test. Aim: Current study was conducted to evaluate the dilution effect of pooling of samples on the sensitivity of RTPCR Test and compare Ct value of positive pool with that of individual positive sample. Materials and Methods: Wide range of positive samples starting from highest Cycle threshold value(Ct) 35 to lowest 15( i.e. 21 sets different Ct value positive specimen) were tested for 7 different sets of pool such as 3,5,10,15,20,25,30 specimen pool for each gene(N and ORF1 ab) separately. After pool formation RNA extraction was done by both manual and automated extraction method followed by detection of COVID-19 target gene by RT-qPCR. Results: All pooled specimens of both genes were tested positive until and below Ct value 24 by both extraction methods. Dilution effect of pooling were noticed from Ct value 25 upwards. Sensitivity of pooled specimen decreases with increasing pool size. 100% sensitivity observed up to 10 pool and 15 pool sample in manual and automated extraction method respectively. Conclusion: Pool testing is the best strategy to carry out population-wide surveillance as new variant of COVID-19 is coming up frequently. Our study indicates we can increase pool size upto15 specimen pool with minimal loss of sensitivity. Current study also observed sensitivity of automated extractor outweighs manual extraction method. COVID-19 no longer constitutes a public health emergency of international concern but being a highly contagious disease, it has the potential to cause rapid outbreak especially in healthcare settings. So periodic surveillance of every ward and every new patient on admission should be incorporated in Hospital infection control programme of every Health care setting.
Authors: Priyanka Saha, Kuhu Pal, Srijita Ghosh, Retina Paul, Satakshi Basu, Arpita Sinha, Joydeep Karmakar
Language: English
Subfield: Infectious Diseases
Field: Medicine
Domain: Health Sciences
Open Access status: bronze
OpenAlex ID: W4415760133