High Prevalences and a Wide Genetic Diversity of Simian Retroviruses in Non-human Primate Bushmeat in Rural Areas of the Democratic Republic of Congo

Abstract

Like the majority of emerging infectious diseases, HIV and HTLV are of zoonotic origin. Here we assess the risk of cross-species transmissions of their simian counterparts, SIV and STLV, from non-human primates (NHP) to humans in the Democratic Republic of Congo (DRC). A total of 331 samples, derived from NHP bushmeat, were collected as dried blood spots (DBS, n = 283) or as tissue samples (n = 36) at remote forest sites mainly in northern and eastern DRC. SIV antibody prevalences in DBS were estimated with a novel high throughput immunoassay with antigens representing the actual known diversity of HIV/SIV lineages. Antibody-positive samples were confirmed by PCR and sequence analysis. Screening for STLV infection was done with universal primers in tax, and new strains were further characterized in LTR. SIV and STLV infection in tissue samples was done by PCR only. Overall, 5 and 15.4% of NHP bushmeat was infected with SIV and STLV, respectively. A new SIV lineage was identified in Allen's swamp monkeys (Allenopithecus nigroviridis). Three new STLV-1 subtypes were identified in Allen's swamp monkeys (Allenopithecus nigroviridis), blue monkeys (Cercopithecus mitis), red-tailed guenons (Cercopithecus ascanius schmidti) and agile mangabeys (Cercocebus agilis). SIV and STLV prevalences varied according to species and geographic region. Our study illustrates clearly, even on a small sample size from a limited number of geographic areas, that our knowledge on the genetic diversity and geographic distribution of simian retroviruses is still limited and that humans continue to be exposed to relative high proportions on infected NHP bushmeat.

Keywords: Bushmeat; Democratic Republic of Congo; Non-human primate; Retrovirus; SIV; STLV.

Figure 1

Sites where dried blood spots of nonhuman primates were collected are highlighted with circles on the map; in red for sites from this survey, in green for sites from our previous survey (Ahuka-Mundeke et al. 2011) and in green with red in sites included in both studies (DBS in the previous and tissue samples in this study)

Figure 2

Phylogenetic relationships of the newly derived SIV sequences in pol to representatives of the other SIV lineages. The newly identified strains in this study are highlighted in red, sequences from our previous survey in DRC are in blue (Ahuka-Mundeke et al., 2011) and reference strains are in black. The trees were inferred from 391 bp nucleotides. The analyses were performed as described in methods. One hundred bootstrap replications were performed to assess confidence in topology, only those above 70% are shown with an asterisk. The scale bar represents 0.1 replacements per site. SIV lineages are generally referred to with a three letter code of the common name from the host species: SIVgsn/mus/mon, greater spot nosed, mustached and mona monkeys; SIVtal, talapoin monkeys; SIVasc, red tailed guenon subspecies (C.ascanius schmidti) from Central DRC; SIVrtg, red tailed guenons from Uganda; SIVagm, african green monkeys; SIVasm, Allen's swamp monkeys, SIVmnd-2/drl, mandrills and drills from Cameroon and northern Gabon; SIVrcm, red cammped magabeys; SIVcpz/gor, chimpanzees and gorillas; SIVsmm, sooty mangabeys; SIVwrc, western red colobus; SIVolc, olive colobus; SIVlho/sun, l'Hoest and suntailed monkeys; SIVdeb, de Brazza monkeys; SIVsyk, Syke's monkeys; SIVcol, black and white colobus or mantled guereza's.

Figure 3

Phylogenetic relationships of the newly derived STLV sequences in LTR (373 bp) to representatives of the other HTLV-1/STLV-1 lineages. The analyses were performed as described in methods. One hundred bootstrap replications were performed to assess confidence in topology, asteriks at nodes are from bootstrap values above 70% of the replicates. Scale bar represents the number of nucleotide substitution per site. Sequences from this study are highlighted in red and those from our previous survey in DRC (Ahuka-Mundeke et al. 2012) in blue. Sequences derived from humans are in grey. Nonhuman primates from our studies are coded using the first letter of the genus followed by the first two letters of the species name: Ang, Allenopithecus nigroviridis; Cmi, Cercopithecus mitis; Cas, Cercopithecus ascanius (all samples from this study in red are from the C.a.whitesidei subspecies, in blue from the C.a.schmidti subspecies); Cag, Cercocebus agilis; Cwo, Cercopithecus wolfi; Cne, Cercopithecus neglectus; and Pth, Piliocolobus tholloni. INNOLIA HTLVI/II antibody tests were performed 12 STLV positive samples higlighted with a black circle: Cas463 and Cas160 were negative; Ang496 and Ang502 were HTLV indeterminate and the 8 remaining had serological profiles corresponding to HTLV-1 infections.

Figure 4

Nonhuman primate species frequently sold at bushmeat markets in DRC and prevalence of SIV and STLV infection per sampling site (A) and per species (B). The data used for the figures are from this study and our previous surveys on SIV (Ahuka-Mundeke et al., 2011) and STLV (Ahuka-Mundeke et al., 2012) in DRC.