Detection and complete genome characterisation of bat coronaviruses from Ghana

Alphacoronaviruses comprise human, livestock, and wildlife coronaviruses of importance, particularly to the livestock industry worldwide. Domestic livestock have the potential to serve as intermediate hosts of coronaviruses, given the contact they have with both humans and wildlife. With the aim of detection and subsequent complete genome sequencing of novel and known but previously undetected alphacoronaviruses, a screening RT-PCR assay was established. In vitro transcripts of two human alphacoronaviruses were generated to evaluate assay performance, and a collection of archived bat and livestock viral RNA samples was tested as part of assay validation. Confirmation of RT-PCR-positive outcomes was done by Sanger and high-throughput sequencing, before phylogenetic analysis. The assay can detect both human alphacoronaviruses with a detection limit of 46 copies/reaction of Alphacoronavirus amsterdamense and 504 copies/reaction of Alphacoronavirus chicagoense in vitro transcripts, as estimated by a probit regression analysis. Validation screening of domestic and peri-domestic livestock species, including cattle, sheep, goats, donkeys, pigs, rabbits, and grasscutters, did not yield any positive outcomes. Confirmed detections were made in bats (3.11%), comprising 1.84% alphacoronaviruses and 1.27% betacoronaviruses. Two complete genomes were obtained for an Alphacoronavirus chicagoense-related bat coronavirus and a putative member of the subgenus Hibecovirus. A near-complete genome was obtained for a Chaerephon spp. bat alphacoronavirus, which was found to be most closely related to a similar virus from Nigeria. This study presents a sensitive assay for coronavirus surveillance and provides additional complete genomes, thereby enhancing our understanding of coronavirus diversity.