Evaluation of the Dilution Effect of Respiratory Specimen Pooling on the Sensitivity of SARS-COV 2 RT-PCR Tests

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 C