Detection and characterization of coronaviruses from African bat species

Abstract

The severe acute respiratory disease syndrome or SARS epidemic emerged in Hong Kong, China, in 2002 with a mortality rate of 15%. The etiological agent for SARS was identified to be a previously unrecognized coronavirus (SARS-CoV) which was found to be zoonotic in origin. A possible reservoir for the SARS-CoV was proposed to be the Chinese horseshoe bat species (Rhinolophus spp.) due to the detection of SARS-related bat coronaviruses (BtCoV) within these bat species. Since the SARS-CoV epidemic, new interest regarding the origin and pathogenicity of coronaviruses has been generated. As such, surveillance studies of BtCoV in numerous bat species have been performed in Asia, Europe, North and South America as well as 3 African countries. Recent BtCoV investigations in Kenya, Ghana and Nigeria identified BtCoV from both the Alpha- and Betacoronavirus genera and provided the first evidence for the presence of coronaviruses in African bats. Previously, the presence of antibodies against SARS-related CoV has been reported in two bat species native to South Africa. This study investigated the possible presence of BtCoV in a panel of bat specimens collected from sites in South Africa and Rwanda, and how they are related to previously detected BtCoVs from other parts of the world. Here we report the development of two PCR assays, the PanBtCoV/9 primer nested RT-PCR and PanBat/AB/6 primer hemi-nested RT-PCR assay, which were used in coronavirus detection from alimentary specimens collected from 15 bat genera. The combined assays amplified coronavirus RNA from 5 samples of the 201 analysed samples collected in South Africa (n=113) and Rwanda (n=88). Three alphacoronaviruses were detected in 3 different South African bat species, Miniopterus spp. (Miniopterus-Bat coronavirus/Irene/South Africa/2009), Neoromicia capensis (Neoromicia-Bat coronavirus/167/South Africa/2007), and Mops midas (Mops-Bat coronavirus/1364/South Africa/2011). From Rwanda, a single betacoronavirus, a SARSrCoV was detected within 2 Rhinolophus spp. individuals (Rh-BtCoV/441/Rwanda/08 and Rh- BtCoV/445/Rwanda/08). Phylogenetic analysis of these sequences was performed and showed that the South African Miniopterus alphacoronavirus and the Rwandan betacoronavirus cluster together with previously detected African BtCoV from the same host genera. The South African alphacoronavirus from Mops midas was closely related to an alphacoronavirus identified within another member of the Molossidae family, Chaerephon spp. from Kenya. Being the first BtCoV identified from the Neoromicia genus, no African BtCoV sequences were available for comparison and as such the virus clustered together with European BtCoV from the Nyctalus spp., another member of the Vespertilioninae subfamily. This study has detected the first BtCoV viral RNA from the native bat species of South Africa and Rwanda, providing confirmation to the presence of bat coronaviruses circulating in these countries. From these preliminary results further investigations into the prevalence and infection cycles of bat coronaviruses in specific bat populations can be performed in the future. The possibility of either these alpha- or betacoronaviruses spilling over and eventually adapting to and infecting other species, though unlikely, cannot be excluded since such rare events are hypothetically responsible for the establishment coronaviruses in humans, livestock, poultry and pets. Caution may still be merited when interacting with bats in roosts and caves.