Background Corona virus disease 2019 (COVID-19) which initially started as a cluster of pneumonia cases in the Wuhan city of China has now become a full-blown pandemic. Timely diagnosis of COVID-19 is the key in containing the pandemic and breaking the chain of transmission. In low-and middle-income countries availability of testing kits has become the major bottleneck in testing. Novel methods like pooling of samples are the need of the hour. Objective We undertook this study to evaluate a novel protocol of pooling of RNA samples/elutes in performance of PCR for SARS CoV-2 virus. Study design Extracted RNA samples were randomly placed in pools of 8 on a 96 well plate. Both individual RNA (ID) and pooled RNA RT-qPCR for the screening E gene were done in the same plate and the positivity for the E gene was seen. Results The present study demonstrated that pool testing with RNA samples can easily detect even up to a single positive sample with Ct value as high as 38. The present study also showed that the results of pool testing is not affected by number of positive samples in a pool. Conclusion Pooling of RNA samples can reduce the time and expense, and can help expand diagnostic capabilities, especially during constrained supply of reagents and PCR kits for the diagnosis of SARS-CoV-2 infection.

Background and Objectives: Timely diagnosis is essential for the containment of the disease and breaks in the chain of transmission of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2). The present situation demands the countries to scale up their testing and design innovative strategies to conserve diagnostic kits and reagents. The pooling of samples saves time, workforce and most importantly diagnostic kits and reagents. In the present study, we tried to define the pool size that could be applied with acceptable confidence for testing. Materials and Methods: We used repeatedly tested positive clinical sample elutes having different levels of SARS CoV 2 RNA and negative sample elutes to prepare seven series of 11 pools each, having pool sizes ranging from 2 to 48 samples to estimate the optimal pool size. Each pool had one positive sample elute in different compositions. All the pools were tested by SARS CoV 2 reverse transcriptase quantitative polymerase chain reaction. Results: Out of the 77 pools, only 53 (68.8%) were found positive. The sensitivity of pools of 2–48 samples was decreased from 100% (95% confidence interval [CL]; 98.4–100) to 41.41% (95% CL; 34.9–48.1). The maximum size of the pool with acceptable sensitivity (>95%) was found to be of six samples. For the pool size of six samples, the sensitivity was 97.8% and the efficiency of pooling was 0.38. Conclusions: The pooling of samples is a practical way for scaling up testing and ultimately containing the further spread of the CoV disease 2019 pandemic.

22 Background 23 Corona virus disease 2019 (COVID-19) which initially started as a cluster of pneumonia 24 cases in the Wuhan city of China has now become a full blown pandemic. Timely diagnosis 25 of COVID-19 is the key in containing the pandemic and breaking the chain of transmission. 26 In low and middle income countries availability of testing kits has become the major bottle 27 neck in testing. Novel methods like pooling of samples are the need of the hour. 28 Method 29 Extracted RNA samples were randomly placed in pools of 8 on a 96 well plate. Both 30 individual RNA (ID) and pooled RNA RT-qPCR for the screening E gene were done in the 31 same plate and the positivity for the E gene was seen.32

Introduction-Timely diagnosis is essential for the containment of the disease and breaks in the chain of transmission of SARS-CoV-2. The present situation demands countries to scale up their testing and design innovative strategies to conserve diagnostic kits and reagents. The pooling of samples saves time, manpower, and most importantly diagnostic kits and reagents. In the present study, we tried to define the pool size that could be applied with acceptable confidence for testing. Material and methods- We used repeatedly tested positive clinical sample elutes having different levels of SARS CoV 2 RNA and negative sample elutes to prepare seven series of 11 pools each, having pool sizes ranging from 2 to 48 samples to estimate the optimal pool size. Each pool had one positive sample elute in different compositions. All the pools were tested by SARS CoV 2 RT-qPCR. Results- Out of the 77 pools, only 53 (68.8%) were found positive. The sensitivity of pools of 2 to 48 samples was decreased from 100% (95% CL; 98.4-100) to 41.41% (95% CL; 34.9-48.1). The maximum size of the pool with acceptable sensitivity (>95%) was found to be of 6 samples. For the pool size of 6 samples, the sensitivity was 97.8% and the efficiency of pooling was 0.38. Conclusion- The pooling of samples is a practical way for scaling up testing and ultimately containing the further spread of the COVID-19 pandemic.

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