Full Genome Nobecovirus Sequences From Malagasy Fruit Bats Define a Unique Evolutionary History for This Coronavirus Clade

Abstract

Bats are natural reservoirs for both Alpha- and Betacoronaviruses and the hypothesized original hosts of five of seven known zoonotic coronaviruses. To date, the vast majority of bat coronavirus research has been concentrated in Asia, though coronaviruses are globally distributed; indeed, SARS-CoV and SARS-CoV-2-related Betacoronaviruses in the subgenus Sarbecovirus have been identified circulating in Rhinolophid bats in both Africa and Europe, despite the relative dearth of surveillance in these regions. As part of a long-term study examining the dynamics of potentially zoonotic viruses in three species of endemic Madagascar fruit bat (Pteropus rufus, Eidolon dupreanum, Rousettus madagascariensis), we carried out metagenomic Next Generation Sequencing (mNGS) on urine, throat, and fecal samples obtained from wild-caught individuals. We report detection of RNA derived from Betacoronavirus subgenus Nobecovirus in fecal samples from all three species and describe full genome sequences of novel Nobecoviruses in P. rufus and R. madagascariensis. Phylogenetic analysis indicates the existence of five distinct Nobecovirus clades, one of which is defined by the highly divergent ancestral sequence reported here from P. rufus bats. Madagascar Nobecoviruses derived from P. rufus and R. madagascariensis demonstrate, respectively, Asian and African phylogeographic origins, mirroring those of their fruit bat hosts. Bootscan recombination analysis indicates significant selection has taken place in the spike, nucleocapsid, and NS7 accessory protein regions of the genome for viruses derived from both bat hosts. Madagascar offers a unique phylogeographic nexus of bats and viruses with both Asian and African phylogeographic origins, providing opportunities for unprecedented mixing of viral groups and, potentially, recombination. As fruit bats are handled and consumed widely across Madagascar for subsistence, understanding the landscape of potentially zoonotic coronavirus circulation is essential for mitigation of future zoonotic threats.

Discovery and Genomic Characterization of a Novel Henipavirus, Angavokely virus, from fruit bats in Madagascar

“…This novel HNV is estimated to have diverged approximately 9,800 years ago, prior to the currently known African and Asian bat-borne HNV lineages but considerably more recently than the estimated mid-to late-Miocene divergence of E. dupreanum from its sister species, E. helvum , on the African continent (53). Recent characterization of Betacoronaviruses in Madagascar fruit bats demonstrates surprising identity to lineages circulating in West Africa (22), suggesting that, despite their endemism, Malagasy fruit bats likely experience some form of contact with the African continent. Of the 49 bat species that inhabit the island nation of Madagascar, nine species are widely distributed across Africa, Asia, and/or Europe, presenting opportunities for inter-species viral transmission via island-hopping.…”

“…Animal capture and handling and subsequent collection of biological samples were conducted in strict accordance with the Madagascar Ministry of Forest and the Environment (permit numbers 019/18, 170/18, 007/19) and guidelines posted by the American Veterinary Medical Association. Field protocols were approved by the UC Berkeley Animal Care and Use Committee (ACUC Protocol # AUP-2017-10-10393), as previously described (22).…”

“…Animal capture and handling and subsequent collection of biological samples were conducted in strict accordance with the Madagascar Ministry of Forest and the Environment (permit numbers 019/18, 170/18, 007/19) and guidelines posted by the American Veterinary Medical Association. Field protocols were approved by the UC Berkeley Animal Care and Use Committee (ACUC Protocol # AUP-2017-10-10393), as previously described (22). Animal capture, sample collection, and RNA extraction Fruit bats were captured and processed in part with a long-term study investigating the seasonal dynamics of potentially zoonotic viruses in Madagascar, as has been previously described (21)(22)(23)(24)(25).…”

“…Field protocols were approved by the UC Berkeley Animal Care and Use Committee (ACUC Protocol # AUP-2017-10-10393), as previously described (22). Animal capture, sample collection, and RNA extraction Fruit bats were captured and processed in part with a long-term study investigating the seasonal dynamics of potentially zoonotic viruses in Madagascar, as has been previously described (21)(22)(23)(24)(25). Animals were identified morphologically by species, sex, and age class (juvenile vs. adult), and urine swabs were collected into viral transport medium from any individual that urinated during handling.…”

“…Association. Field protocols were approved by the UC Berkeley Animal Care and Use Committee (ACUC Protocol # AUP-2017-10-10393), as previously described (22). Animal capture, sample collection, and RNA extraction Fruit bats were captured and processed in part with a long-term study investigating the seasonal dynamics of potentially zoonotic viruses in Madagascar, as has been previously described (21)(22)(23)(24)(25).…”