Hepatitis B, C, and E infection among HIV-infected patients in Franceville, Gabon: retrospective cross-sectional study.

Access to antiretrovirals has increased the life expectancy of patients living with HIV. However, HIV-viral hepatitis coinfections in countries endemic for these infections make management more difficult. To determine the extent of these coinfections in Gabon, we investigated markers of hepatitis B, C, and E viruses in 762 adults infected with HIV-1 by ELISA. We used real-time PCR to quantify plasma HBV DNA (HBV VL) and amplified HCV and HEV RNA by nested RT-PCR and PCR.The seroprevalence of chronic hepatitis B infection was 9.3 %. Among patients with isolated HBcAc profiles, the prevalence of occult hepatitis B infection was 26.7 %, for a real prevalence (detectable CV-HBV) of 17.3 % of the total population. HCV seroprevalence was 8.8 %. Of the 67 HIV-positive patients, 76.1 % had replicative profiles (detectable HCV RNA), that is, 6.7 % of the total population. For hepatitis E, seroprevalence was 3.5 %. No case of chronic HEV infection was found. In conclusion, this study highlights a high rate of HIV-Hepatitis B, C and E coinfections in Gabon. In addition, we show the interest of looking for chronic infections (replicative profiles) in HIV-infected patients in Gabon. The establishment of technical platforms for this type of research, accessible to middle-income countries, is necessary.

Hepatitis viruses in non-human primates.

BACKGROUND: Previous epidemiological studies of rural human populations in Gabon reveal a high prevalence of human hepatitis A, B, C and D viruses. In order to investigate the prevalence of the blood-born hepatitis viruses in apes and monkeys living in the same area, we performed an epidemiological survey of HBV, HCV and HDV in wild-born non-human primates. METHODS: We tested 441 wild-born non-human primates from Gabon and Congo and 132 imported monkeys for the presence of serological markers of HBV, HCV and HDV infections. RESULTS: None of Cercopithecidae monkeys were reactive against HBV/HDV and HCV. In contrast, 29.2% of wild-born great apes (154 chimpanzees and 14 gorillas) were positive for HBV serological markers. Nine chimpanzees were in the replicative phase of HBV infection. None of these HBV infected chimpanzees exhibited symptoms or significant changes in serum clinical chemistry related to HBV infection. CONCLUSIONS: The negativity to HCV-related viruses and the negativity of the Cercopithecidae species tested against HBV/HDV do not allow us to definitively rule out the presence of an animal counterpart of human hepatitis viruses in non-human primates.

Identification of hepatitis B virus genome in faecal sample from wild living chimpanzee (Pan troglodytes troglodytes) in Gabon.

Non-invasive faecal sampling in the equatorial forest in Gabon allowed the first identification of the hepatitis B virus (HBV-Ch(RC170)) genome in samples collected from wild chimpanzees (Pan troglodytes troglodytes). The HBV-Ch(RCl70)sequence clustered with 100% bootstrap support with previous viral sequences obtained from Pan troglodytes subspecies. This is the first evidence of HBV infection in wild apes and confirms that the HBV-like strains thus far characterized in captive apes are directly related to those circulating in the wild.

Two distinct STLV-1 subtypes infecting Mandrillus sphinx follow the geographic distribution of their hosts.

The mandrill (Mandrillus sphinx) has been shown to be infected with an STLV-1 closely related to HTLV-1. Two distinct STLV-1 subtypes (D and F) infect wild mandrills with high overall prevalence (27.0%) but are different with respect to their phylogenetic relationship and parallel to the mandrills' geographic range. The clustering of these new STLV-1mnd sequences with HTLV-1 subtype D and F suggests first, past simian-to-human transmissions in Central Africa and second, that species barriers are easier to cross over than geographic barriers.

Occurrence of hepatitis viruses in wild-born non-human primates: a 3 year (1998-2001) epidemiological survey in Gabon.

Hepatitis B and C infections are endemic in human population in central Africa, particularly in Gabon. The aim of this study was to determine the prevalence of hepatitis B virus (HBV) and eventual occurrence of hepatitis C virus (HBC)-related strains in a variety of wild-born non-human primates living in Gabon and Congo. Plasma samples were screened for HBV and HCV markers. A non-invasive method of DNA extraction from faeces followed by specific HBV-DNA amplification was developed to study this infection in wild troops of chimpanzees and gorillas. No HCV infection in non-human primates, wild-born or captive, was detected among 596 samples tested. No HBV infection could be detected in samples tested and obtained from Cercopithecidae. In contrast, 14.7 and 42.2% of wild-born chimpanzees in Gabon and Congo were infected with HBV or had evidence of past HBV infection. At Centre International de Recherches Médicales (CIRMF) Primate Centre, 32.1% of chimpanzees and gorillas were HBV positive or had evidence of past infection. In the cases with past infection, 5.9% wild-born and 8.3% at CIRMF harboured HBV-DNA despite the presence of neutralizing HbsAb. Together with previous findings, we confirm the high HBV prevalence not only in humans but also in chimpanzees and gorillas in Gabon and Congo.

Molecular evidence for deep phylogenetic divergence in Mandrillus sphinx.

Mandrills (Mandrillus sphinx) are forest primates indigenous to western central Africa. Phylogenetic analysis of 267 base pairs (bp) of the cytochrome b gene from 53 mandrills of known and 17 of unknown provenance revealed two phylogeographical groups, with haplotypes differentiated by 2.6% comprising seven synonymous transitions. The distribution of the haplotypes suggests that the Ogooué River, Gabon, which bisects their range, separates mandrill populations in Cameroon and northern Gabon from those in southern Gabon. The haplotype distribution is also concordant with that of two known mandrill simian immunodeficiency viruses, suggesting that these two mandrill phylogroups have followed different evolutionary trajectories since separation.

Phylogenetic analysis of 49 newly derived HIV-1 group O strains: high viral diversity but no group M-like subtype structure.

We assess the genetic relationships between 49 HIV-1 group O strains from 24 and 25 patients living in Cameroon and France, respectively. Strains were sequenced in four genomic regions: gag (p24) and three env regions (C2-V3, gp41, and for 22 C2-gp41). In each of the genomic regions analyzed, the genetic diversity among the group O strains was higher than that exhibited by group M. We characterize three major group O phylogenetic clusters (O:A, O:B, and O:C) that comprised the same virus strains in each of the genomic regions analyzed. The majority of strains cluster in O:A, a cluster previously identified by analysis of pol and env sequences. Group O recombinants were also identified. Importantly, the distinction between these three major group O clades was weak compared to the strong clustering apparent in the global group M phylogenetic tree that led to the identification of subtypes. Thus, these clusters of group O viruses should not be considered as equivalent to the group M subtypes. This difference between the pattern of group O and the global group M diversity, both taking into account the pandemic status of the group M subtypes and the comparatively small number of group O-infected individuals (the majority being from Cameroon), indicates that the group O phylogeny primarily represents viral divergence in the Cameroon region, analogous to group M viral diversity present in the Democratic Republic of Congo.

Wild Mandrillus sphinx are carriers of two types of lentivirus.

Mandrillus sphinx, a large primate living in Cameroon and Gabon and belonging to the Papionini tribe, was reported to be infected by a simian immunodeficiency virus (SIV) (SIVmndGB1) as early as 1988. Here, we have identified a second, highly divergent SIVmnd (designated SIVmnd-2). Genomic organization differs between the two viral types; SIVmnd-2 has the additional vpx gene, like other SIVs naturally infecting the Papionini tribe (SIVsm and SIVrcm) and in contrast to the other SIVmnd type (here designated SIVmnd-1), which is more closely related to SIVs infecting l'hoest (Cercopithecus lhoesti lhoesti) and sun-tailed (Cercopithecus lhoesti solatus) monkeys. Importantly, our epidemiological studies indicate a high prevalence of both types of SIVmnd; all 10 sexually mature wild-living monkeys and 3 out of 17 wild-born juveniles tested were infected. The geographic distribution of SIVmnd seems to be distinct for the two types: SIVmnd-1 viruses were exclusively identified in mandrills from central and southern Gabon, whereas SIVmnd-2 viruses were identified in monkeys from northern and western Gabon, as well as in Cameroon. SIVmnd-2 full-length sequence analysis, together with analysis of partial sequences from SIVmnd-1 and SIVmnd-2 from wild-born or wild-living mandrills, shows that the gag and pol regions of SIVmnd-2 are closest to those of SIVrcm, isolated from red-capped mangabeys (Cercocebus torquatus), while the env gene is closest to that of SIVmnd-1. pol and env sequence analyses of SIV from a related Papionini species, the drill (Mandrillus leucophaeus), shows a closer relationship of SIVdrl to SIVmnd-2 than to SIVmnd-1. Epidemiological surveys of human immunodeficiency virus revealed a case in Cameroon of a human infected by a virus serologically related to SIVmnd, raising the possibility that mandrills represent a viral reservoir for humans similar to sooty mangabeys in Western Africa and chimpanzees in Central Africa.

Synthetic peptide strategy for the detection of and discrimination among highly divergent primate lentiviruses.

We developed a simple, rapid, inexpensive, and highly sensitive and specific strategy for the detection and lineage differentiation of primate lentiviruses (PIV-ELISA). It is based on the use of two indirect ELISA methods using synthetic peptides mapping the gp41/36 region (detection component) and the V3 region (differentiation component) of four lentivirus lineages, namely SIVcpz/HIV-1 (groups M, O, N, and SIVcpz-gab), SIVmnd, SIVagm, and SIVsm/SIVmac/HIV-2. This strategy was evaluated with panels of sera originating from both humans and nonhuman primates. The human reference panel consisted of 144 HIV Western blot (WB)-positive sera in which the corresponding virus had been genotyped (HIV-1: 72 group M, 28 group O, and 6 group N; HIV-2: 21 subtype A and 10 subtype B; and 7 HIV-1+2) and 105 HIV WB-negative samples. The nonhuman primate reference panel consisted of 24 sera from monkeys infected by viruses belonging to the four lineages included in the PIV-ELISA strategy (5 chimpanzees, 5 macaques, 8 mandrills, and 6 vervets) and 42 samples from seronegative animals. Additional field evaluation panels consisted of 815 human sera from Gabon, Cameroon, and France and 537 samples from 25 nonhuman primate species. All the samples from the two reference panels were correctly detected and discriminated by PIV-ELISA. In the human field evaluation panel, the gp41/36 component correctly identified all the test samples, with 98% specificity. The V3 component discriminated 206 HIV-1 group M, 98 group O, 12 group M+O, and 128 HIV-2 sera. In the primate field evaluation panel, both gp41/36 and V3 detected and discriminated all the WB-positive samples originating from monkeys infected with SIVcpz, SIVagm-ver, SIVmnd-1, SIVmnd-2, SIVdrl, or SIVsun. These results were confirmed by genotyping in every case. Four SIV-infected red-capped mangabeys (confirmed by PCR) were correctly identified by gp41/36, but only two reacted with the V3 peptides in the absence of a specific SIVrcm V3 peptide. Addition of a V3 SIVrcm peptide discriminated all the SIVrcm-positive samples. Fourteen Papio papio samples were positive for SIVsm gp 36 and by WB, but negative by PCR, whereas three Papio cynocephalus samples were positive by gp41/36 but indeterminate by WB and negative by PCR. This combined ELISA system is thus highly sensitive and specific for antibodies directed against HIV and SIV. In addition, the V3-based serotyping results always agreed with genotyping results. This method should prove useful for studies of lentivirus prevalence and diversity in human and nonhuman primates, and may also have the potential to detect previously undescribed SIVs.

Variability of human immunodeficiency virus type 2 (hiv-2) infecting patients living in france.

To determine the prevalence of human immunodeficiency virus type 2 (HIV-2) subtypes circulating in France and to identify possible relationships between these subtypes and pathogenesis, we studied 33 HIV-2-infected patients living in France. HIV-2 DNA was directly amplified from peripheral blood mononuclear cells by nested PCR with specific HIV-2 env primers, and the env gene was sequenced. The serological consequences of antigenic variability were studied by using a panel of peptides and by Western blotting. Phylogenetic analysis classified the 33 HIV-2 strains as subtype A (n = 23) or B (n = 10). There were no significant clinical or epidemiological differences between patients infected with either of these two subtypes. There was some evidence for geographical clustering. Subtype A strains from patients originating from the Cape Verde Islands and Guinea Bissau clustered together. The majority of patients infected with subtype B strains originated from the Ivory Coast or Mali. Strains from patients originating in Mali also clustered in subtype A but distinctly from the Cape Verde or Guinea Bissau strains. The subtype B strains showed greater diversity and included some highly divergent strains relative to those previously characterized. The V3 loop of HIV-2 subtypes A and B was found to be quite conserved in comparison with HIV-1. A strong HIV-2 subtype B serological cross-reactivity was found on HIV-1 env antigen by Western blot mostly in the gp41 transmembrane glycoprotein. This could partly explain the double HIV-1 and HIV-2 reactive profiles found in countries where HIV-2 subtype B is prevalent.

SIVcpz from a naturally infected Cameroonian chimpanzee: biological and genetic comparison with HIV-1 N.

Thus far, simian immunodeficiency virus from chimpanzees (SIVcpz) genomes have been characterized as Pan troglodytes troglodytes and show a strong relation with human immunodeficiency virus (HIV)-1 N in their env genes. We fully characterized another SIVcpz from P. t. troglodytes. This chimpanzee (Cam5) was, as was also the host of SIVcpz-cam3, wild born in Cameroon, a region where all three groups of HIV-1 (M, N and O) co-occur. In contrast to other SIVcpz, SIVcpz-cam5 was isolated immediately after the rescue of the animal. Our data demonstrate that SIVcpz-cam5, like SIVcpz-cam3, grows easily on human peripheral blood mononuclear cells (PBMCs) and uses CCR5 as a co-receptor similar to HIV-1 N YBF30. Phylogenetic analysis based on the entire env gene shows that SIVcpz-cam5 falls into the same unique subcluster as HIV-1 N YBF30, SIVcpz-cam3 and SIVcpz-US. A phylogenetic relationship was also found with the vif gene of HIV-1 N. This study provides proof that HIV-1 N related viruses circulate in wild P. t. troglodytes.

Highly sensitive method for amplification of human immunodeficiency virus type 2 DNA.

We evaluated a new human immunodeficiency virus type 2 (HIV-2) DNA amplification strategy based on peripheral blood mononuclear cell long PCR (XL PCR) followed by nested PCR amplification. The primers used were located in the highly conserved long terminal repeat and in the pol regions of the genome. Five primer pairs corresponding to different regions of the HIV-2 env gene were used in the nested step. Samples from 42 patients were tested, which yielded positive amplification with at least two primer pairs in 40 (95%) samples. A primer pair (EB2/EB5) located on the V3 region succeeded in amplifying proviral DNA in 40 samples.

Synthetic peptide ELISAs for detection of and discrimination between group M and group O HIV type 1 infection.

We developed and evaluated two peptide-based immunoassays to confirm and discriminate between group M and group O HIV-1 infection. These assays are based on in vitro competition for antibody binding between M and O peptides. The first EIA is based on competition between group M and group O gp41 immunodominant domains and the second on competition between group O and group M V3 regions of gp120. Two panels of sera were used: the first consisted of 109 sera collected from 27 group O- and 92 group M-infected patients in whom the HIV isolates had been genotyped by sequencing or heteroduplex mobility assay. In this panel, the combination of the two assays correctly discriminated 106 samples (100% group O and 96.7% group M samples). The second panel, used for the field evaluation of the two assays, consisted of 157 samples from HIV-1-infected Cameroonian patients, 33 strains having been genotyped. The combination of the two techniques in a serogrouping algorithm discriminated 147 of these samples, 74 being HIV-1 group O and 73 group M. These results always correlated with genotyping results. The 10 sera that were not successfully classified by these assays were from early seroconverters. Altogether, the two assays clearly differentiated 263 of 276 (94.9%) samples in the two panels. On the basis of the genotyping results, the positive predictive value for group discrimination in the two panels was 100% for both GSEIA assays. Our peptide-blocking group-specific EIAs for differentiation and confirmation of HIV-1 group M and group O infection are complementary tools for epidemiological studies and surveillance of HIV-1 group O strain trafficking.