Novel highly divergent sapoviruses detected by metagenomics analysis in straw-colored fruit bats in Cameroon

Abstract

Sapoviruses (SaVs) belong to the Sapovirus genus, in the family Caliciviridae. They have been associated with gastroenteritis in humans and in pigs but not in other animals. In addition, some strains from pigs, chimpanzees and rodents show close sequence identity with human SaVs thereby suggesting the possibility of interspecies transmissions. Bats are known to be a major reservoir of zoonotic viruses, however, very little is known about the genetic diversity of SaVs in bats. To explore the genetic diversity of bat SaVs, fecal samples of Eidolon helvum and Epomophorus gambianus were treated according to the NetoVIR protocol and sequenced by Illumina technology. Nearly complete genome sequences of six highly divergent SaVs and one partial SaV (only VP1 region) were identified in Eidolon helvum and based on sequence identities and phylogenetic analysis, they potentially represent two novel genogroups, only distantly related to known SaVs. Furthermore, comparing these sequences with currently used screening primers and probes indicated that the novel SaVs would not be detected in routine epidemiological screening studies in humans in case an interspecies transmission would occur. Therefore, we designed and validated new primers that can detect both human and bat SaVs. In this study, we identified multiple novel bat SaVs, however, further epidemiological studies in humans are needed to unravel their potential role in gastroenteritis.

Figure 1

Genome organization of novel sapovirus strains from straw-colored fruit bats. (A, B) SaV strains Limbe65 and Limbe899b with non-overlapping ORF1 and ORF2 in different reading frames; (D) SaV strain Lysoka36 with an overlapping ORF1 and ORF2; (C, E, F) SaV strains Limbe25, Limbe899a and Limbe900 with non-overlapping ORF1 and ORF2 in the same reading frame.

Figure 2

Maximum-likelihood phylogenetic tree, based on a VP1 amino acid sequence alignment of Limbe65, Limbe899a, Limbe899b, Limbe25, Limbe900, Limbe894 and Lysoka36, and 55 other representative SaVs strains. Previously known bat SaVs are indicated with open triangles, whereas those described in this paper are indicated with filled triangle. Filled circles are strains with the potential history of cross between species. The numbers at the internal nodes represent the bootstrap probabilities (in percent), as determined for 1000 iterations. Only bootstrap values greater than 70% are shown. The scale bar indicates the genetic distance (amino acid substitutions per site).

Figure 3

Polyacrylamide gel electrophoresis of the PCR product amplified with novel primers (A) and currently used primers (B). Wells 1 and 14 or 15: DNA Molecular Weight Marker VIII; wells 2–9: human samples positive for SaV (genogroups GI.1, GI.3, GII.3, GII.5, GII.6, GIV and GV; wells 10–13: bat samples positive for SaV (novel putative genogroups GXVIII and GXIX).