Detection of SARS-CoV-2 RNA in bivalve mollusks and marine sediments

Abstract

The presence of SARS-CoV-2 in wastewater pose the question of whether this new pandemic virus could be released into watercourses and potentially continue to finally reach coastal waters. In this study, we employed two bivalve molluscan species from the genus Ruditapes as sentinel organisms to investigate the presence of SARS-CoV-2 signals in the marine coastal environment. Estuarine sediments from the natural clam banks were also analyzed. Viral RNA was detected by RT-qPCR, targeting IP4, E and N1 genomic regions. Positive samples were also subjected to a PMAxx-triton viability RT-qPCR assay in order to discriminate between intact and altered capsids, obtaining indirect information about the viability of the virus. SARS-CoV-2 RNA traces were detected in 9/12 clam samples by RT-qPCR, from which 4 were positive for two different target regions. Viral quantification ranged from <LoQ to 4.48 Log genomic copies/g of digestive tissue. Regarding the sediment samples, 3/12 were positive by RT-qPCR, but only IP4 region was successfully amplificated. Quantification values for sediment samples ranged from <LoQ to 3.60 Log genomic copies/g of sediment. RNA signals disappeared in the PMAxx-triton viability RT-qPCR assay, indicating non-infectious potential. In addition, the recently discovered human-specific gut associated bacteriophage crAssphage was also quantified as a biomarker for the presence of human-derived wastewater contamination on the study area. CrAssphage was detected in 100% of both types of samples with quantification values ranging from <LoQ to 5.94 Log gc/g digestive tissue and from <LoQ to 4.71 Log gc/g sediment. Statistical analysis also showed that quantification levels for the crAssphage in clams are significantly higher than in sediments. These findings represent the first detection of SARS-CoV-2 RNA in the marine environment, demonstrating that it can reach these habitats and make contact with the marine life.

Keywords: Bivalve mollusks; COVID-19; Marine environment; SARS-CoV-2; Sediments; Wastewater.