Evaluation of three viral concentration methods for detection and quantification of SARS-CoV-2 in wastewater.

Wastewater-based epidemiology (WBE) could be useful as an early warning system for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic spread. Viruses are highly diluted in wastewater. Therefore, a virus concentration step is needed for SARS-CoV-2 wastewater detection. We tested the efficiency of three wastewater viral concentration methods: ultrafiltration (UF), electronegative membrane filtration and aluminum hydroxide adsorption-elution. We spiked wastewater with inactivated SARS-CoV-2 and we collected 20 other wastewater samples from five sites in Tunisia. Samples were concentrated by the three methods and SARS-CoV-2 was quantified by reverse transcription digital PCR (RT-dPCR). The most efficient method was UF with a mean SARS-CoV-2 recovery of 54.03 ± 8.25. Moreover, this method provided significantly greater mean concentration and virus detection ability (95%) than the two other methods. The second-most efficient method used electronegative membrane filtration with a mean SARS-CoV-2 recovery of 25.59 ± 5.04% and the least efficient method was aluminum hydroxide adsorption-elution. This study suggests that the UF method provides rapid and straightforward recovery of SARS-CoV-2 in wastewater.

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.