Detailed phylogenetic analysis of primate T-lymphotropic virus type 1 (PTLV-1) sequences from orangutans (Pongo pygmaeus) reveals new insights into the evolutionary history of PTLV-1 in Asia.

While human T-lymphotropic virus type 1 (HTLV-1) originates from ancient cross-species transmission of simian T-lymphotropic virus type 1 (STLV-1) from infected nonhuman primates, much debate exists on whether the first HTLV-1 occurred in Africa, or in Asia during early human evolution and migration. This topic is complicated by a lack of representative Asian STLV-1 to infer PTLV-1 evolutionary histories. In this study we obtained new STLV-1 LTR and tax sequences from a wild-born Bornean orangutan (Pongo pygmaeus) and performed detailed phylogenetic analyses using both maximum likelihood and Bayesian inference of available Asian PTLV-1 and African STLV-1 sequences. Phylogenies, divergence dates and nucleotide substitution rates were co-inferred and compared using six different molecular clock calibrations in a Bayesian framework, including both archaeological and/or nucleotide substitution rate calibrations. We then combined our molecular results with paleobiogeographical and ecological data to infer the most likely evolutionary history of PTLV-1. Based on the preferred models our analyses robustly inferred an Asian source for PTLV-1 with cross-species transmission of STLV-1 likely from a macaque (Macaca sp.) to an orangutan about 37.9-48.9kya, and to humans between 20.3-25.5kya. An orangutan diversification of STLV-1 commenced approximately 6.4-7.3kya. Our analyses also inferred that HTLV-1 was first introduced into Australia ~3.1-3.7kya, corresponding to both genetic and archaeological changes occurring in Australia at that time. Finally, HTLV-1 appears in Melanesia at ~2.3-2.7kya corresponding to the migration of the Lapita peoples into the region. Our results also provide an important future reference for calibrating information essential for PTLV evolutionary timescale inference. Longer sequence data, or full genomes from a greater representation of Asian primates, including gibbons, leaf monkeys, and Sumatran orangutans are needed to fully elucidate these evolutionary dates and relationships using the model criteria suggested herein.

Maintenance of picobirnavirus (PBV) infection in an adult orangutan (Pongo pygmaeus) and genetic diversity of excreted viral strains during a three-year period.

The present work provide data about the maintenance of picobirnavirus (PBV) infection during adulthood in a mammalian host. For this purpose PBV infection was studied in an adult orangutan (Pongo pygmaeus) by PAGE/SS, RT-PCR and nucleotide sequencing. PBV infection in the animal was asymptomatic and was characterized by interspaced silent and high/ low active viral excretion periods. The PBV strains excreted by the studied individual were identified as genogroup I and revealed a nucleotide identity among them of 64-81%. The results obtained allowed to arrive to a deeper understanding of the natural history of PBV infection, which seems to be characterized by new-born, juvenile and adult asymptomatic hosts which persistently excrete closely related strains in their feces. Consequently, picobirnaviruses could be considered frequent inhabitants of the gastrointestinal tract, leaving the question open about the molecular mechanisms governing persistent and asymptomatic coexistence within the host and the potential host suitability to maintain this relationship.

Characterization of novel polyomaviruses from Bornean and Sumatran orang-utans.

Serological screening of sera from orang-utans demonstrated a high percentage of sera that cross-reacted with antigens of the polyomavirus (PyV) simian virus 40. Analysis of archival DNA samples from 71 Bornean and eight Sumatran orang-utans with a broad-spectrum PCR assay resulted in the detection of PyV infections in 11 animals from both species. Sequence analysis of the amplicons revealed considerable differences between the PyVs from Bornean and Sumatran orang-utans. The genome from two PyVs, one from each species, was therefore amplified and sequenced. Both viral genomes revealed a characteristic PyV architecture, but lacked an obvious agnogene. Neighbour-joining analysis positioned the viruses in a large cluster together with viruses from bats, bovines, rodents and several primate PyVs from chimpanzees, African green monkeys, squirrel monkeys and the human Merkel cell PyV.

Novel cytomegaloviruses in free-ranging and captive great apes: phylogenetic evidence for bidirectional horizontal transmission.

Wild great apes often suffer from diseases of unknown aetiology. This is among the causes of population declines. Because human cytomegalovirus (HCMV) is an important pathogen, especially in immunocompromised individuals, a search for cytomegaloviruses (CMVs) in deceased wild and captive chimpanzees, gorillas and orang-utans was performed. By using a degenerate PCR targeting four conserved genes (UL54-UL57), several distinct, previously unrecognized CMVs were found for each species. Sequences of up to 9 kb were determined for ten novel CMVs, located in the UL54-UL57 block. A phylogenetic tree was inferred for the ten novel CMVs, the previously characterized chimpanzee CMV, HCMV strains and Old World and New World monkey CMVs. The primate CMVs fell into four clades, containing New World monkey, Old World monkey, orang-utan and human CMVs, respectively, plus two clades that each contained both chimpanzee and gorilla isolates (termed CG1 and CG2). The tree loci of the first four clades mirrored those for their respective hosts in the primate tree, suggesting that these CMV lineages arose through cospeciation with host lineages. The CG1 and CG2 loci corresponded to those of the gorilla and chimpanzee hosts, respectively. This was interpreted as indicating that CG1 and CG2 represented CMV lineages that had arisen cospeciationally with the gorilla and chimpanzee lineages, respectively, with subsequent transfer within each clade between the host genera. Divergence dates were estimated and found to be consistent with overall cospeciational development of major primate CMV lineages. However, CMV transmission between chimpanzees and gorillas in both directions has also occurred.

Lineage-specific expansions of retroviral insertions within the genomes of African great apes but not humans and orangutans.

Retroviral infections of the germline have the potential to episodically alter gene function and genome structure during the course of evolution. Horizontal transmissions between species have been proposed, but little evidence exists for such events in the human/great ape lineage of evolution. Based on analysis of finished BAC chimpanzee genome sequence, we characterize a retroviral element (Pan troglodytes endogenous retrovirus 1 [PTERV1]) that has become integrated in the germline of African great ape and Old World monkey species but is absent from humans and Asian ape genomes. We unambiguously map 287 retroviral integration sites and determine that approximately 95.8% of the insertions occur at non-orthologous regions between closely related species. Phylogenetic analysis of the endogenous retrovirus reveals that the gorilla and chimpanzee elements share a monophyletic origin with a subset of the Old World monkey retroviral elements, but that the average sequence divergence exceeds neutral expectation for a strictly nuclear inherited DNA molecule. Within the chimpanzee, there is a significant integration bias against genes, with only 14 of these insertions mapping within intronic regions. Six out of ten of these genes, for which there are expression data, show significant differences in transcript expression between human and chimpanzee. Our data are consistent with a retroviral infection that bombarded the genomes of chimpanzees and gorillas independently and concurrently, 3-4 million years ago. We speculate on the potential impact of such recent events on the evolution of humans and great apes.

The phylogeography of orangutan foamy viruses supports the theory of ancient repopulation of Sumatra.

Phylogenetic analysis of foamy virus sequences obtained from Bornean and Sumatran orangutans showed a distinct clustering pattern. One subcluster was represented by both Bornean and Sumatran orangutan simian foamy viruses (SFV). Combined analysis of host mitochondrial DNA and SFV phylogeny provided evidence for the hypothesis of the repopulation of Sumatra by orangutans from Borneo.

Phylogenetic relationships in the Lymphocryptovirus genus of the Gammaherpesvirinae.

Complete DNA sequences were determined for the glycoprotein B (gB) genes of four viruses from the genus Lymphocryptovirus, whose hosts had been assigned as baboon, orangutan, chimpanzee and gorilla. Together with published sequences for the gB genes of three lymphocryptoviruses, namely the human pathogen Epstein-Barr virus (EBV), a rhesus monkey virus and a marmoset virus, the sequences were used to investigate evolutionary relationships in the genus. The chimpanzee and orangutan viruses' sequences were found to be so close that it is unlikely both represent natural infections in these hosts. Phylogenetic analyses showed that the New World marmoset virus lineage formed a sister clade to that of the Old World viruses, consistent with a cospeciational separation. Within the Old World virus group, resolution of branching pattern was incomplete, and suggestive of a complex history. In particular, it was inferred that separation of the EBV lineage from that of the gorilla virus plus the chimpanzee/orangutan virus may have predated separation of the present day host species.

Structural and evolutionary analysis of an orangutan foamy virus.

The full-length proviral genome of a foamy virus infecting a Bornean orangutan was amplified, and its sequence was analyzed. Although the genome showed a clear resemblance to other published foamy virus genomes from apes and monkeys, phylogenetic analysis revealed that simian foamy virus SFVora was evolutionarily equidistant from foamy viruses from other hominoids and from those from Old World monkeys. This finding suggests an independent evolution within its host over a long period of time.

Primate hepatitis B viruses - genetic diversity, geography and evolution.

There are six well characterised genotypes (A-F) of human hepatitis B virus that have distinct geographic ranges which generally relate to chronic HBV infection. A seventh human genotype (G) has recently been described, but there is limited information on ethnic and geographic distribution. Despite the fact that early studies indicated that HBV antigens were present in other primates, the prevailing dogma that HBV was a human disease precluded alternative explanations. Within the past 5 years, hepatitis B viruses have been characterised from all the Old World great apes (orangutan, gibbons, gorillas and chimpanzees) and from a New World woolly monkey. Each group of non-human primates appears to have a distinct strain of hepatitis B virus that can be distinguished from human sequences based upon the nucleotide sequence and selected amino acid changes in the viral proteins. The woolly monkey HBV is most divergent from other primate and human sequences, while the great ape HBV sequences cluster together with separate branches for each group.

Sylvatic transmission of arboviruses among Bornean orangutans.

Wild populations of nonhuman primates live in regions of sylvatic arbovirus transmission. To assess the status of arbovirus transmission in Bornean forests and the susceptibility of wild orangutans to arboviral infection, blood samples of wild orangutans, semi-captive orangutans, and humans were examined. Samples were tested by plaque reduction neutralization test for antibodies to viruses representing three families (Flaviviridae, Alphaviridae, and Bunyaviridae), including dengue-2, Japanese encephalitis, Zika, Langat, Tembusu, Sindbis, Chikungunya, and Batai viruses. Both wild and semi-captive orangutan groups as well as local human populations showed serologic evidence of arbovirus infection. The presence of neutralizing antibodies among wild orangutans strongly suggests the existence of sylvatic cycles for dengue, Japanese encephalitis, and sindbis viruses in North Borneo. The present study demonstrates that orangutans are susceptible to arboviralinfections in the wild, although the impact of arboviral infections on this endangered ape remain unknown.

Analysis of two genomic variants of orang-utan hepadnavirus and their relationship to other primate hepatitis B-like viruses.

We recently described orang-utan hepadnavirus (OuHV) (Warren et al., Journal of Virology, 73, 7860-7865, 1999). Phylogenetic analyses indicated that the various isolates of OuHV can be divided into two genomic variants. Two representatives from each genomic cluster were analysed both molecularly and phylogenetically. Their genome organization was highly similar to other hepadnaviruses of apes and humans. The complete genome sequences of the two OuHV types had an overall 5% sequence difference. Research on 25 seropositive Bornean orang-utans showed that, of the 19 animals infected with one variant, 12 originated from East Kalimantan. Phylogenetic analysis was performed using the full-length genomes of various primate hepadnaviruses. The tree topology revealed one cluster of Old World hepadnaviruses that is divided into two subclusters, one consisting of the ape viruses, and the other comprising the human genotypes A-E. These data suggest that the great apes and gibbons have been infected with a common ancestor hepadnavirus.

Many human endogenous retrovirus K (HERV-K) proviruses are unique to humans.

BACKGROUND: Endogenous retroviruses contribute to the evolution of the host genome and can be associated with disease. Human endogenous retrovirus K (HERV-K) is related to the mouse mammary tumor virus and is present in the genomes of humans, apes and cercopithecoids (Old World monkeys). It is unknown how long ago in primate evolution the full-length HERV-K proviruses that are in the human genome today were formed. RESULTS: Ten full-length HERV-K proviruses were cloned from the human genome. Using provirus-specific probes, eight of the ten were found to be present in a genetically diverse set of humans but not in other extant hominoids. Intact preintegration sites for each of these eight proviruses were present in the apes. A ninth provirus was detected in the human, chimpanzee, bonobo and gorilla genomes, but not in the orang-utan genome. The tenth was found only in humans, chimpanzees and bonobos. Complete sequencing of six of the human-specific proviruses showed that full-length open reading frames for the retroviral protein precursors Gag-Pro-Pol or Env were each present in multiple proviruses. CONCLUSIONS: At least eight full-length HERV-K genomes that are in the human germline today integrated after humans diverged from chimpanzees. All of the viral open reading frames and cis-acting sequences necessary for HERV-K replication must have been intact during the recent time when these proviruses formed. Multiple full-length open reading frames for all HERV-K proteins are present in the human genome today.

A new group of hepadnaviruses naturally infecting orangutans (Pongo pygmaeus).

A high prevalence (42.6%) of hepatitis B virus (HBV) infection was suspected in 195 formerly captive orangutans due to a large number of serum samples which cross-reacted with human HBV antigens. It was assumed that such viral infections were contracted from humans during captivity. However, two wild orangutans were identified which were HBV surface antigen positive, indicating that HBV or related viruses may be occurring naturally in the orangutan populations. Sequence analyses of seven isolates revealed that orangutans were infected with hepadnaviruses but that these were clearly divergent from the six known human HBV genotypes and those of other nonhuman hepadnaviruses reported. Phylogenetic analyses revealed geographic clustering with Southeast Asian genotype C viruses and gibbon ape HBV. This implies a common origin of infection within this geographic region, with cross-species transmission of hepadnaviruses among hominoids.

Human endogenous retrovirus K homologous sequences and their coding capacity in Old World primates.

The coding capacity for retroviral Gag and Env proteins has been maintained in human endogenous retroviruses of the HERV-K family. HERV-K homologous sequences have been found in all Old World primates. Here, we examined Old World primate species for the presence of full-length HERV-K gag and env genes and the presence of gag and env open reading frames as determined by the protein truncation test. Full-length HERV-K env genes were found in DNAs of all Old World primate species, whereas open reading frames for Env protein were found solely in human, chimpanzee, and gorilla DNAs. The mutational event leading to two HERV-K types was found to have occurred after the separation of hominids from lower Old World primates and before the expansion of hominids. Full-length HERV-K gag genes in hominids displayed a 96-bp deletion compared to those in lower Old World primates. The ancient gag variant has not been maintained during hominid evolution. Open reading frames for HERV-K Gag have been found in all Old World primates except chimpanzees. Our study of the HERV-K family during Old World primate evolution contributes to the understanding of their possible biological functions in the host genomes.

Characterization of a simian T-lymphotropic virus from a wild-caught orang-utan (Pongo pygmaeus) from Kalimantan, Indonesia.

In a recent serological survey among 143 ex-captive orang-utans two individuals were found that reacted positive in an ELISA detecting antibodies which cross-react with human T-lymphotropic virus type I (HTLV-I) antigens. Infection of both animals with an HTLV-I or simian T-lymphotropic virus (STLV)-like virus was confirmed by Western blot analysis. A third wild-caught animal, which was not part of the original serological survey, was also found to be infected with an HTLV-related virus in a diagnostic PCR assay and Western blot assay. Nucleotide sequence analysis of the 709 bp PCR fragment from the tax/rex region of the HTLV/STLV genome confirmed infection of orang-utans with an STLV similar to but clearly distinct from other Asian STLVs.

Isolation of STLV-I from orangutan, a great ape species in Southeast Asia, and its relation to other HTLV-Is/STLV-Is.

To study the evolutionary origin of human T-lymphotropic virus type I/simian T-lymphotropic virus type I (HTLV-I/STLV-I), we isolated and characterized STLV-I from orangutans (Pongo pygmaeus). Plasma samples from 3 out of 41 animals examined were reactive by particle agglutination and immunofluorescence, and one of these three was confirmed to be anti-HTLV-I antibody-positive by western blotting (WB). Cultured peripheral blood mononuclear cells from the WB-positive orangutan were reactive to anti-STLV-I-positive rhesus monkey plasma. The proviral long terminal repeat region was amplified by polymerase chain reaction and sequenced. A phylogenetic analysis indicated that orangutan STLV-I is related to the Melanesian group of HTLV-Is and other Asian STLV-Is, but the degree of divergence is considerable.

Identification of a novel family of human endogenous retroviruses and characterization of one family member, HERV-K(C4), located in the complement C4 gene cluster.

We have identified a novel family of about 10-50 human endogenous retrovirus elements (HERVs) and have characterized one family member (HERV-KC4). This retrovirus element is integrated within intron 9 of and complement C4A genes and also in some C4B genes, and is a principal contribution to interlocus and interallelic length heterogeneity of C4 genes. The HERV-K(C4) sequence has a typical retrovirus structure with elements of gag, pol and env domains, flanked by two long terminal repeats (LTRs) and is similar to type A, B and D retroviruses. Multiple termination codons preclude the existence of long open reading frames, suggesting that the HERV-K(C4) sequence is no longer functional. Zoo blot hybridization reveals that New World monkeys appear to lack sequences similar to HERV-K(C4), suggesting that integration has occurred after the divergence of Old and New World monkeys. Retrotransposition of prototype viruses is presumed to have led to the amplification and integration of the members of the family in different loci, which in humans, appear to be dispersed over several chromosomes. The absence of the HERV-K(C4) element in some C4B genes in both humans and orangutangs indicate that the retrovirus inserted into the C4A gene after the duplication of the cluster. Subsequent spread of the HERV-K(C4) sequence to C4B genes presumably occurred by interlocus sequence exchange mechanisms, such as unequal crossover and gene conversion-like mechanisms.

Isolation of a new foamy retrovirus from orangutans.

We have isolated a new foamy virus from blood samples taken from two apparently healthy orangutans (Pongo pygmaeus). The older orangutan has since died with encephalopathy after a brief acute illness, while the younger one, his grandson, remains well. These animals and 12 other orangutans had specific antibodies to foamy virus as measured by immunofluorescence. The new foamy virus and the antisera showed strong and specific neutralization, with only weak cross-reaction with other simian foamy virus strains. Southern blotting with gag and env probes of human foamy virus and PCR amplification showed that the new foamy virus, designated SFV-11, is related to, yet distinct from, previously characterized strains from humans, chimpanzees, and monkeys.