The Delta variant wave in Tunisia: Genetic diversity, spatio-temporal distribution and evidence of the spread of a divergent AY.122 sub-lineage.

Introduction: The Delta variant posed an increased risk to global public health and rapidly replaced the pre-existent variants worldwide. In this study, the genetic diversity and the spatio-temporal dynamics of 662 SARS-CoV2 genomes obtained during the Delta wave across Tunisia were investigated. Methods: Viral whole genome and partial S-segment sequencing was performed using Illumina and Sanger platforms, respectively and lineage assignemnt was assessed using Pangolin version 1.2.4 and scorpio version 3.4.X. Phylogenetic and phylogeographic analyses were achieved using IQ-Tree and Beast programs. Results: The age distribution of the infected cases showed a large peak between 25 to 50 years. Twelve Delta sub-lineages were detected nation-wide with AY.122 being the predominant variant representing 94.6% of sequences. AY.122 sequences were highly related and shared the amino-acid change ORF1a:A498V, the synonymous mutations 2746T>C, 3037C>T, 8986C>T, 11332A>G in ORF1a and 23683C>T in the S gene with respect to the Wuhan reference genome (NC_045512.2). Spatio-temporal analysis indicates that the larger cities of Nabeul, Tunis and Kairouan constituted epicenters for the AY.122 sub-lineage and subsequent dispersion to the rest of the country. Discussion: This study adds more knowledge about the Delta variant and sub-variants distribution worldwide by documenting genomic and epidemiological data from Tunisia, a North African region. Such results may be helpful to the understanding of future COVID-19 waves and variants.

Pooling Nasopharyngeal Swab Specimens to Improve Testing Capacity for SARS-CoV-2 by Real-Time RT-PCR.

BACKGROUND: The detection of SARS-CoV-2 using qRT-PCR with the pooling of samples can reduce workload and costs especially when the prevalence rate of COVID-19 in a population is low. To analyse the effect of pooling samples on the sensitivity of RT-qPCR for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) detection, we compared the cycle threshold (Ct) values of pools of 5 and 10 that tested positive with Ct values of individual samples that tested positive in that pool. Twenty positive nasopharyngeal (NP) specimens with low to high viral load were selected and pooled individually with four and nine negative NP. RESULTS: In NP specimens, the sensitivity of pools of 5 and 10 were 90 and 85%, compared to individual sample testing, respectively. The RT-qPCR sensitivity of pools of 5 and 10 against individual testing were not significantly different (p > 0.05). Detection of positive samples with low Ct values (< 36) was consistently achieved in pools of 5 and 10. However, there were higher false negatives when samples with high ct values (> 36) were pooled and tested. The mean Ct values obtained with the 5-sample pooled testing exceeded individual sample testing by 1.85 ± 1.09 cycles, while Ct values obtained with the 10-sample pooling exceeded individual sample testing by 3.4 ± 1.65 cycles. CONCLUSIONS: In a low prevalence setting, testing capacity can be increased by pooling 5 or 10 samples, but the risk of additional false negatives needs to be considered.

SARS-CoV-2 infection virological diagnosis.

SARS-CoV-2 infection has to be confirmed by virological diagnosis. Multiple diagnostic tests are available without enough perspective on their reliability. Therefore, it is important to choose the most suitable test according to its sensitivity and specificity but also to the stage of the disease. Currently, the RT-PCR detection of the viral genome in respiratory samples is the most reliable test to confirm the diagnosis of an acute SARS-CoV-2 infection. It has to be done in Class II biological safety laboratory. However, it may lack sensitivity, particularly in the advanced phase of infection, and depends closely on the samples' quality. Rapid PCR by cartridge system reduces response times but is not suitable for laboratories with high throughput of requests. Detection of virus antigens on respiratory samples is a quick and easy to use technique; however it has not good specificity and sensitivity and cannot be used for diagnosis and patient management. The detection of specific antibodies against SARS-CoV-2 is better used for epidemiological analyses. Research should be encouraged to overcome the limits of the currently available diagnostic tests.