Abstract
This study aims at establishing specimens pooling approach for the detection of SARS-CoV-2 using the RT-PCR BGI and Sansure-Biotech kits used in Gabon. To validate this approach, 14 positive samples, stored at -20°C for three to five weeks were analyzed individually (as gold standard) and in pools of five, eight and ten in the same plate. We created 14 pools of 5, 8 and 10 samples using 40 μL from each of the selected positive samples mixed with 4, 7 and 9 confirmed negative counterparts in a total volume of 200 μL, 320 μL and 400 μL for the pools of 5, 8 and 10 respectively. Both individual and pooled samples testing was conducted according to the BGI and Sansure-Biotech RT-PCR protocols used at the Professor Daniel Gahouma Laboratory (PDGL). Furthermore, the pooling method was also tested by comparing results of 470 unselected samples tested in 94 pools and individually. Results of our experiment showed that using a BGI single positive sample with cycle threshold (Ct) value of 28.42, confirmed by individual testing, detection occurred in all the pools. On the contrary samples with Ct >31 were not detected in pools of 10 and for these samples (Ct value as high as 37.17) their detection was possible in pool of 8. Regarding the Sansure-Biotech kit, positive samples were detected in all the pool sizes tested, irrespective of their Ct values. The specificity of the pooling method was 100% for the BGI and Sansure-Biotech RT-PCR assays. The present study found an increase in the Ct values with pool size for the BGI and Sansure-Biotech assays. This trend was statistically significant (Pearson’s r = 0.978; p = 0,022) using the BGI method where the mean Ct values were 24.04±1.1, 26.74±1.3, 27.91±1.1 and 28.32±1.1 for the individual, pool of 5, 8 and 10 respectively. The testing of the 470 samples showed that one of the 94 pools had a positive test similar to the individual test using the BGI and Sansure-Biotech kits. The saving of time and economizing test reagents by using the pooling method were demonstrated in this study. Ultimately, the pooling method could be used for the diagnosis of SARS-CoV-2 without modifying the accuracy of results in Gabon. We recommend a maximum pool size of 8 for the BGI kit. For the Sansure-Biotech kit, a maximum pool size of 10 can be used without affecting its accuracy compared to the individual testing.
Highlights
The outbreak of COVID-19 caused by the novel coronavirus (SARS-CoV-2) has spread globally [1] since it was first reported in Wuhan, China, in December 2019 [2]
Of the 14 positive samples initially selected for the experimental pools, SARS-CoV-2 was confirmed in the retest for the eight samples pool using the BGI assay
For the maximum pool size that is the concern in the pooling method, the results of the present study show that with high viral load (Ct up to 27.20: Table 3), the test efficiency match was recorded between individual and all the pool sizes tested for both kits
Summary
The outbreak of COVID-19 caused by the novel coronavirus (SARS-CoV-2) has spread globally [1] since it was first reported in Wuhan, China, in December 2019 [2]. As in other countries Gabon is affected by COVID-19 and according to the May 2021 report, 11,457 cases and 71 deaths have so far been reported after conducting 456,358 tests [3]. Since the report of the first case in Libreville, the capital city of Gabon, in March 2020, the country has conducted massive SARS-CoV-2 testing to identify infected individuals and to trace cases in order to mitigate its spread. To support this mitigation strategy, > 15 laboratories and > 60 testing centers were created in the country. Since July 2020, Gabon has increased its diagnostic capacity by conducting more than 2,000 tests per day, compared to the 200 tests per week at the start of the pandemic
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