1 Unité d'Epidémiologie et Physiopathologie des Virus Oncogènes, Département EEMI, Bâtiment Lwoff, Institut Pasteur, 28 rue du Dr Roux, 75724 Paris cedex 15, France
2 Department of Virology, Biomedical Primate Research Center (BPRC), Lange Kleiweg 139, 2288 GJ Rijswijk, The Netherlands
3 Centre Pasteur du Cameroun, BP 1274, Yaoundé, Cameroon
Correspondence
Antoine Gessain
agessain{at}pasteur.fr
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ABSTRACT |
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The GenBank/EMBL/DDBJ accession numbers for the sequences reported in this paper are AY359406AY359408.
Supplementary tables showing the sequences of oligonucleotide primers and GenBank accession numbers are available in JGV Online.
Present address: Laboratoire de Rétrovirologie, Institut Pasteur de la Guyane, 23 avenue Pasteur, BP 6010, 97306 Cayenne cedex, French Guyana.
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MAIN TEXT |
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Soon after the characterization of HHV-6, chimpanzee lymphocytes were shown to be susceptible to experimental HHV-6 infection (Lusso et al., 1990). In addition, testing of captive chimpanzees by an HHV-6 immunofluorescence assay revealed that 7590 % were seropositive, depending on where they were living, a prevalence similar to that in humans (Higashi et al., 1989
). Furthermore, the existence of HHV-6-related viruses in other non-human primates has also been strongly suggested by serological studies (Blewett et al., 2001
; Higashi et al., 1989
; Jenson et al., 2002
). Nevertheless, such viruses have never been characterized. Thus, to confirm the existence of an HHV-6 homologue indigenous to chimpanzees, we examined blood samples obtained from 77 common chimpanzees.
The larger series of chimpanzees comprised 42 wild-born animals (22 Pan troglodytes vellerosus and 20 P. troglodytes troglodytes) originating from different parts of Cameroon, where they were gathered in wildlife-rescue centres in the south-western province of Cameroon or in Yaoundé, and from the large primate centre of the Centre International de Recherches Médicales de Franceville (CIRMF) in Gabon. The second series (35 chimpanzees, of which 33 were P. troglodytes verus and two were P. troglodytes schweinfurthii) was obtained from a closed breeding colony of chimps housed at the Biomedical Primate Research Centre (BPRC), Rijswijk, the Netherlands (Table 1). DNA was extracted from buffy coats with a QIAamp DNA Blood mini kit (Qiagen), following the manufacturer's instructions. Then, by using consensus degenerate primers targeting the herpesvirus DNA polymerase gene (Rose et al., 1997
), we attempted to amplify herpesvirus sequences from these DNAs as described previously (Lacoste et al., 2001
) (see Supplementary Table S1 in JGV Online). Briefly, DNA samples were initially amplified with the primer pools DFASA and GDTD1B, and an aliquot of these amplification products was then used as a template in a subsequent nested PCR (nPCR) with the VYGA and GDTD1B primer pools. nPCR products were examined by agarose-gel electrophoresis, purified by using a QIAquick gel-extraction kit (Qiagen) and then cloned by TA cloning in the pCR2.1 cloning vector (Invitrogen). Cycle sequencing was performed by Eurogentec (Seraing, Belgium) using BigDye Terminator technology. Initial screening was done on the 25 chimpanzees, mainly from Cameroon, from which we previously identified different rhadinoviruses (Lacoste et al., 2000a
, 2001
). Among these 25 DNAs, 20 scored positive on the ethidium bromide-stained gel. Sequencing of multiple individual clones from each positive chimp allowed us to identify different herpesviruses. Whilst most of the sequences amplified corresponded to PanRHV1a, PanRHV1b, PanRHV2 or PtroLCV1 (Ehlers et al., 2003
; Lacoste et al., 2000a
, 2001
), three of them, from three different chimps, were related closely to HHV-6, as observed by database BLAST searches. To obtain the sequence extending upstream of the VYGA region, a gene-specific, non-degenerate primer (P6as; see Supplementary Table S1 in JGV Online) was derived from the complementary sequence of the HHV-6-like VYGAGDTD1B fragment and used in an nPCR amplification with the DFASA primer pool. The PCR products (DFASAGDTD1B) from the initial PCR were used as template DNAs in these subsequent amplification reactions. The nucleotide sequences of the DFASAGDTD1B fragments yielded 476 bp sequences after exclusion of the primer sequences. Sequence analyses and alignments were performed by using the MacVector 6.0 and AssemblyLIGN software packages (Oxford Molecular Ltd). Phylogenetic analyses of nucleotide and deduced amino acid sequences were performed by using the PHYLIP package (version 3.52c; Felsenstein, 1993
), as described previously (Lacoste et al., 2001
). The phylogenetic analyses based on this fragment further confirmed that the identified sequence was related closely to HHV-6 (Fig. 1
). Lastly, an additional degenerate primer pool (POL6A), upstream of DFASA and derived from a conserved amino acid motif within the DNA polymerase gene of roseoloviruses, as well as two new gene-specific primers (POL6B1 and POL6B2) derived from the complementary sequences of the DFASAGDTD1B fragments, have been designed to finally obtain an 883 bp sequence (see Supplementary Table S1 in JGV Online). This was 6 and 7 % divergent at the nucleotide level and 3 % at the protein level with respect to HHV-6B and -6A sequences, respectively (Table 2
). This novel herpesvirus, related closely to the sixth human herpesvirus, was provisionally named PanHV6 for Pan troglodytes herpesvirus 6. However, among the specialists of the field, a new proposal will certainly be discussed and debated. When a new name is approved, we will modify its name accordingly.
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By defining other sets of degenerate primers (see Supplementary Table S1 in JGV Online), we amplified two additional fragments of 490 and 640 bp, corresponding to the U12 and U42 genes of HHV-6, respectively. Whilst U42 is a member of the only family of genes involved in transactivation that is conserved among all of the herpesviruses (Dominguez et al., 1999; Isegawa et al., 1999
), the U12 gene of HHV-6 encodes a functional
-chemokine receptor (Isegawa et al., 1998
), representative of a group of similar G-coupled receptor (GCR) homologues conserved between human and animal members of the Betaherpesvirinae (Murphy, 2001
). It has been shown that the individual viral homologues within this GCR group show greater identity to each other than to the closest known cellular homologue (Raftery et al., 2000
). Moreover, as the PanHV6 U12 homologue shows the same degree of similarity to its HHV-6 counterpart as the DNA polymerase gene, all of these data suggest that these viruses acquired this gene from their cellular host before viral speciation (Table 2
). Sequencing of these two other gene fragments from the same three chimpanzees as mentioned above (each belonging to one subspecies) showed no significant sequence variation (12 nt, with no amino acid change). These results further confirm that there is no distinct PanHV6 genotype infecting the different chimpanzee subspecies. Nevertheless, as our sequencing studies focused on highly conserved regions of the viral genome, another HHV-6 genotype might be identified if regions of greater divergence, such as those between HHV-6A and HHV-6B, are scrutinized.
These studies represent a definitive identification of roseolovirus infection in non-human primates. Indeed, although we previously identified a -herpesvirus from mandrill and drill monkeys that is related to the human roseoloviruses, this virus, named MndHV
, is only distantly related to them (Lacoste et al., 2000b
). Similar to the other chimpanzee herpesviruses, PanHV6 is related very closely to its human counterpart. It is worth noting that the PanHV6 DNA polymerase gene sequence obtained shows the same degree of similarity to its human counterpart compared with the chimpanzee EpsteinBarr virus (EBV), human cytomegalovirus and Kaposi's sarcoma-associated herpesvirus homologues (9597 %) (Davison et al., 2003
; Ehlers et al., 2003
; Lacoste et al., 2000a
). We assume that, when more data become available from other simian herpesvirus 6 isolates, they will support the theory of co-evolution of herpesviruses with their host species (McGeoch et al., 1995
). Interestingly, the sequences obtained from the buffy-coat DNA of these animals are related more closely to HHV-6B than to HHV-6A. This suggests that, if a PanHV6-A variant exists, it it is more likely to be detected in the central nervous system of chimpanzees, due to the apparently different tropism of the two human viral variants.
Since its discovery in 1986, HHV-6 is increasingly recognized as an important pathogen in immunocompromised patients (especially transplant recipients) and is currently hypothesized as a strong suspect in the origin of multiple sclerosis (Dockrell, 2003; Soldan et al., 1997
). Due to the high prevalence of latently infected individuals in the healthy population, its precise role in the formerly mentioned conditions is not well understood (Agut, 1993
; Fillet et al., 1998
; Le Cleach et al., 1998
). Therefore, it will be of interest to determine whether any pathology due to PanHV6 infection occurs in the common chimpanzees. Further characterization of the PanHV6 genome will enable us to make meaningful comparisons between the human and chimpanzee herpesvirus 6 sequences, yielding numerous insights into evolutionary paths within the genus Roseolovirus, and to decipher different biological information encoded in these two genomes.
The sequences reported in this paper have been deposited in GenBank under accession numbers AY359406AY359408. Due to the quasi-identity of the sequences obtained from the three chimpanzee subspecies, only PanHV6 sequences obtained from P. troglodytes verus have been submitted to GenBank.
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ACKNOWLEDGEMENTS |
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REFERENCES |
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---|
Agut, H. (1993). Puzzles concerning the pathogenicity of human herpesvirus 6. N Engl J Med 329, 203204.
Blewett, E. L., White, G., Saliki, J. T. & Eberle, R. (2001). Isolation and characterization of an endogenous cytomegalovirus (BaCMV) from baboons. Arch Virol 146, 17231738.[CrossRef][Medline]
Challoner, P. B., Smith, K. T., Parker, J. D. & 12 other authors (1995). Plaque-associated expression of human herpesvirus 6 in multiple sclerosis. Proc Natl Acad Sci U S A 92, 74407444.
Clark, D. A. (2000). Human herpesvirus 6. Rev Med Virol 10, 155173.[CrossRef][Medline]
Davison, A. J., Dolan, A., Akter, P., Addison, C., Dargan, D. J., Alcendor, D. J., McGeoch, D. J. & Hayward, G. S. (2003). The human cytomegalovirus genome revisited: comparison with the chimpanzee cytomegalovirus genome. J Gen Virol 84, 1728.
Dockrell, D. H. (2003). Human herpesvirus 6: molecular biology and clinical features. J Med Microbiol 52, 518.
Dominguez, G., Dambaugh, T. R., Stamey, F. R., Dewhurst, S., Inoue, N. & Pellett, P. E. (1999). Human herpesvirus 6B genome sequence: coding content and comparison with human herpesvirus 6A. J Virol 73, 80408052.
Ehlers, B., Ochs, A., Leendertz, F., Goltz, M., Boesch, C. & Mätz-Rensing, K. (2003). Novel simian homologues of Epstein-Barr virus. J Virol 77, 1069510699.
Felsenstein, J. (1993). PHYLIP (phylogeny inference package), version 3.52c. Department of Genome Sciences, University of Washington, Seattle, USA.
Fillet, A.-M., Lozeron, P., Agut, H., Lyon-Caen, O. & Liblau, R. (1998). HHV-6 and multiple sclerosis. Nat Med 4, 537.[Medline]
Hall, C. B., Long, C. E., Schnabel, K. C. & 7 other authors (1994). Human herpesvirus-6 infection in children a prospective study of complications and reactivation. N Engl J Med 331, 432438.
Hall, C. B., Caserta, M. T., Schnabel, K. C., Long, C., Epstein, L. G., Insel, R. A. & Dewhurst, S. (1998). Persistence of human herpesvirus 6 according to site and variant: possible greater neurotropism of variant A. Clin Infect Dis 26, 132137.[Medline]
Higashi, K., Asada, H., Kurata, T., Ishikawa, K., Hayami, M., Spriatna, Y., Sutarman & Yamanishi, K. (1989). Presence of antibody to human herpesvirus 6 in monkeys. J Gen Virol 70, 31713176.[Abstract]
Isegawa, Y., Ping, Z., Nakano, K., Sugimoto, N. & Yamanishi, K. (1998). Human herpesvirus 6 open reading frame U12 encodes a functional -chemokine receptor. J Virol 72, 61046112.
Isegawa, Y., Mukai, T., Nakano, K. & 10 other authors (1999). Comparison of the complete DNA sequences of human herpesvirus 6 variants A and B. J Virol 73, 80538063.
Jenson, H. B., Ench, Y., Zhang, Y., Gao, S.-J., Arrand, J. R. & Mackett, M. (2002). Characterization of an EpsteinBarr virus-related gammaherpesvirus from common marmoset (Callithrix jacchus). J Gen Virol 83, 16211633.
Kasolo, F. C., Mpabalwani, E. & Gompels, U. A. (1997). Infection with AIDS-related herpesviruses in human immunodeficiency virus-negative infants and endemic childhood Kaposi's sarcoma in Africa. J Gen Virol 78, 847855.[Abstract]
Lacoste, V., Mauclère, P., Dubreuil, G., Lewis, J., Georges-Courbot, M.-C. & Gessain, A. (2000a). KSHV-like herpesviruses in chimps and gorillas. Nature 407, 151152.[CrossRef][Medline]
Lacoste, V., Mauclère, P., Dubreuil, G., Lewis, J., Georges-Courbot, M.-C., Rigoulet, J., Petit, T. & Gessain, A. (2000b). Simian homologues of human gamma-2 and betaherpesviruses in mandrill and drill monkeys. J Virol 74, 1199311999.
Lacoste, V., Mauclère, P., Dubreuil, G., Lewis, J., Georges-Courbot, M.-C. & Gessain, A. (2001). A novel 2-herpesvirus of the rhadinovirus 2 lineage in chimpanzees. Genome Res 11, 15111519.
Le Cleach, L., Fillet, A. M., Agut, H. & Chosidow, O. (1998). Human herpesviruses 6 and 7: new roles yet to be discovered? Arch Dermatol 134, 11551157.
Lusso, P., Markham, P. D., DeRocco, S. E. & Gallo, R. C. (1990). In vitro susceptibility of T lymphocytes from chimpanzees (Pan troglodytes) to human herpesvirus 6 (HHV-6): a potential animal model to study the interaction between HHV-6 and human immunodeficiency virus type 1 in vivo. J Virol 64, 27512758.[Medline]
McGeoch, D. J., Cook, S., Dolan, A., Jamieson, F. E. & Telford, E. A. R. (1995). Molecular phylogeny and evolutionary timescale for the family of mammalian herpesviruses. J Mol Biol 247, 443458.[CrossRef][Medline]
Murphy, P. M. (2001). Viral exploitation and subversion of the immune system through chemokine mimicry. Nat Immunol 2, 116122.[CrossRef][Medline]
Niphuis, H., Verschoor, E. J., Bontjer, I., Peeters, M. & Heeney, J. L. (2003). Reduced transmission and prevalence of simian T-cell lymphotropic virus in a closed breeding colony of chimpanzees (Pan troglodytes verus). J Gen Virol 84, 615620.
Raftery, M., Müller, A. & Schönrich, G. (2000). Herpesvirus homologues of cellular genes. Virus Genes 21, 6575.[CrossRef][Medline]
Rose, T. M., Strand, K. B., Schultz, E. R., Schaefer, G., Rankin, G. W., Jr, Thouless, M. E., Tsai, C.-C. & Bosch, M. L. (1997). Identification of two homologs of the Kaposi's sarcoma-associated herpesvirus (human herpesvirus 8) in retroperitoneal fibromatosis of different macaque species. J Virol 71, 41384144.[Abstract]
Salahuddin, S. Z., Ablashi, D. V., Markham, P. D. & 8 other authors (1986). Isolation of a new virus, HBLV, in patients with lymphoproliferative disorders. Science 234, 596601.[Medline]
Schirmer, E. C., Wyatt, L. S., Yamanishi, K., Rodriguez, W. J. & Frenkel, N. (1991). Differentiation between two distinct classes of viruses now classified as human herpesvirus 6. Proc Natl Acad Sci U S A 88, 59225926.
Soldan, S. S., Berti, R., Salem, N. & 9 other authors (1997). Association of human herpes virus 6 (HHV-6) with multiple sclerosis: increased IgM response to HHV-6 early antigen and detection of serum HHV-6 DNA. Nat Med 3, 13941397.[CrossRef][Medline]
Tuke, P. W., Hawke, S., Griffiths, P. D. & Clark, D. A. (2004). Distribution and quantification of human herpesvirus 6 in multiple sclerosis and control brains. Mult Scler 10, 355359.[CrossRef][Medline]
Yamanishi, K., Okuno, T., Shiraki, K., Takahashi, M., Kondo, T., Asano, Y. & Kurata, T. (1988). Identification of human herpesvirus-6 as a causal agent for exanthem subitum. Lancet i, 10651067.[CrossRef]
Zerr, D. M., Meier, A. S., Selke, S. S. & 8 other authors (2005). A population-based study of primary human herpesvirus 6 infection. N Engl J Med 352, 768776.
Received 15 March 2005;
accepted 13 May 2005.
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