This study provides evidence for infection of chimpanzees (P. troglodytes subsp. verus) born in the wild. Despite the different geographical origins and handling histories of the two study chimpanzees, the nucleotide sequences of HBV amplified from Chimps 2 and 4 were similar to each other but phylogenetically distinct from those of humans or other primate species. HBV infection appeared to be relatively common among chimpanzees in West Africa; in the refuge center in Cameroon, HBsAg screening of other wild-born chimpanzees has identified a further 8 positive samples among 26 tested (J.C.M.L., unpublished data). Further PCR testing and sequence analysis are required to confirm the serology results and to investigate further the distribution of the chimpanzee genotype reported in this study.
The existence of a chimpanzee-specific genotype of HBV is supported by the close similarity of this genotype to that of HBV recovered from a captive chimpanzee (HPBVCG [strain LSH]; references 15
), a finding which clarifies the origin of HBV infection in the London Zoo outbreak. The original investigators found HBV infection in three offspring of a male and a female that had been captured in Africa. However, neither in the original description nor in the subsequent report of the sequence of strain LSH were the authors able to determine whether HBV infection was acquired by either the parents or the offspring in the London Zoo, by the parents after capture in Africa (possibly by inoculation of human immunoglobulin preparations), or in the wild from other chimpanzees. The genetic relatedness of strain LSH to human genotypes has led both these (15
) and subsequent investigators (3
) to conclude that the chimpanzee infection arose from human contact. The discovery of the same HBV genotype in chimpanzees sampled independently in this study provides convincing evidence against this hypothesis.
Whether other primate species are infected with HBV in the wild has been controversial. Norder et al. (12
) obtained the sequence of an HBV variant (HBU46935) infecting a chimpanzee after inoculation with serum from a captive white-handed gibbon (Hybolates lar
), although whether the original source of HBV was from the wild has remained controversial. However, the recent unpublished description of six complete genome sequences of HBV that originated from gibbons (S. Grethe et al., accession no. AJ131568
) and group closely with HBU46935 (data not shown) suggests infection of gibbons in the wild, although clarification of this hypothesis awaits publication of the data.
HBV has also been recovered from a captive woolly monkey (Lagothrix lagotricha
; reference 9
). The sequence of WMHBV is the most divergent of all primate HBV variants, and unlike other primate HBVs, there was some evidence for a restriction in host range; an inoculum from an infected woolly monkey failed to efficiently infect a chimpanzee (9
). Very recently, HBV infection was detected in a number of originally wild-caught orangutans (16
). Although sequences from the surface gene appeared to show some relationship to the HBU46935 gibbon virus, that all seven grouped together on the tree also implies that orangutans have a specific variant of HBV. In summary, there is growing evidence from this and previous studies for widespread infection of primates with HBV and for the existence of species-specific genotypes. Furthermore, the three genotypes recovered from Old World primate species show a level of sequence divergence from each other (9 to 10%) similar to that among human HBV genotypes A to E.
The finding of HBV infecting native chimpanzees has a number of implications for theories of the origins of human HBV infection. A hypothesis for a relatively recent origin proposed that HBV spread from the Americas into Europe and elsewhere in the Old World after contact between Europeans and indigenous peoples around 400 years ago (3
). However, the finding that chimpanzees, as well as orangutans (16
), can be infected with HBV in the wild makes the proposed recent spread of HBV from the New World extremely unlikely.
It has been alternatively proposed that HBV infection was present in anatomically modern humans as they migrated from Africa approximately 150,000 to 100,000 years ago (10
), and the different genotypes infecting humans evolved since this dispersal. The problem with the hypothesis is that it also does not explain the origin of the various nonhuman primate viruses which, with the exception of the variant found in woolly monkeys, are interspersed among the human genotypes in the phylogenetic tree.
Finally, it could be argued that the variants found in chimpanzees in this study and previously in gibbons, orangutans, and New World primate woolly monkeys are viruses that coevolved with their primate hosts over periods of 10 to 35 million years. The numerous genotypes found in humans would therefore originate through multiple zoonotic transmission episodes from several nonhuman primate species infected with different species-specific genotypes. Such a scenario is not unprecedented; human immunodeficiency virus type 1 infection in humans originated through at least three separate cross-species transmissions from different subspecies of chimpanzees (6
), while human infection with human immunodeficiency virus type 2 in West Africa arose independently several times through contact with sooty mangabeys (5
). The first difficulty here is that the phylogenetic tree of the various primate HBV variants in no way reflects the phylogeny of the host species, as would be expected for cospeciation. Second, if human genotypes A to F originated in primates, then the actual species involved in transmission to humans remain unidentified.
At this stage, the problems associated with each of three hypotheses for the origin of HBV prevent a definitive conclusion. First, as indicated previously (2
), resolution of this issue requires more extensive HBsAg screening and sequence analysis of HBV infecting wild-caught primates from each of their three principal geographical ranges. Second, indigenous human populations in areas of high endemicity for HBV infection (such as sub-Saharan Africa) are poorly sampled and evidence for the existence of other genotypes should be sought. Combined human and primate studies may succeed in identifying the immediate sources of each of the six human genotypes and the circumstances under which HBV subsequently came to be globally distributed.