Proc. Natl. Acad. Sci. USA Vol. 88, pp. 7694-7698, September 1991 Medical Sciences Highly divergent molecular variants of human T-lymphotropic virus type I from isolated populations in Papua New Guinea and the Solomon Islands (retrovirus/PCR/Melanesia/evolution) ANTOINE GESSAIN*, RICHARD YANAGIHARAt4, GENOVEFFA FRANCHINI*, RALPH M. GARRUTOt, CAROL L. JENKINS§, ANDREW B. AJDUKIEWICZ¶, ROBERT C. GALLO*, AND D. CARLETON GAJDUSEKt *Laboratory of Tumor Cell Biology, National Cancer Institute, and tLaboratory of Central Nervous System Studies, National Institute of Neurological Disorders and Stroke, National Institutes of Health, Bethesda, MD 20892; §Papua New Guinea Institute of Medical Research, Goroka, Papua New Guinea; and Ministry of Health and Medical Services, Central Hospital, Honiara, Solomon Islands Contributed by D. Carleton Gajdusek, May 28, 1991 ABSTRACT To determine the molecular genetic relation- inhabitants from widely separated regions in the Solomon ship between Melanesian strains of human T-lymphotropic Islands (13). It has been suggested that the serological data virus type I (HTLV-I) and cosmopolitan prototype HTLV-I, we are consistent with the existence of variant viruses, phylo- amplified by PCR, then cloned, and sequenced a 522-base-pair genetically related to, but distinct from, prototype HTLV-I region of the HTLV-I env gene in DNA extracted from uncul- (2, 5, 8, 11, 13). The subsequent identification of a case of tured (fresh) and cultured peripheral blood mononuclear cells HTLV-I myeloneuropathy in a life-long resident of the So- obtained from six seropositive Melanesian Papua New Guine- lomon Islands (14) and the isolation of HTLV-I-like viruses ans and Solomon Islanders, including a Solomon Islander with from a healthy Hagahai man (15, 16) and from unrelated HTLV-I myeloneuropathy. Unlike isolates of HTLV-I from Solomon Islanders (17, 18) establish the existence ofHTLV-I Japan, the West Indies, the Americas, and Africa, which share in Melanesia. We now report on the molecular genetic .97% sequence homology, the Melanesian strains of HTLV-I relationship between cosmopolitan prototype HTLV-I and were only 91.8%-92.5% identical with a prototype Japanese these variant viruses from Melanesia, tentatively designated HTLV-IATK-I. The nucleotide sequence of proviral DNA from HTLVdlMelanesia.lI the Solomon Islander with HTLV-I myeloneuropathy also diverged markedly from that of HTLV-I isolated from Japa- nese patients with HTLV-I-associated myelopathy and from MATERIALS AND METHODS Jamaican patients with tropical spastic paraparesis, suggesting Study Population. Eight individuals from Papua New that these variant viruses are capable of causing disease. The Guinea and the Solomon Islands, all of whom possessed IgG HTLV-I variants from Papua New Guineans, in turn, differed antibodies against HTLV-I gag-encoded proteins p19 and by nearly 4% from the Melanesian variants from Solomon p24 and env gene products gp46 and/or gp2l, were studied. Islanders, indicating the existence of another HTLV-I quasi- Except for one individual with HTLV-I myeloneuropathy species. By contrast, HTLV-I strains from two residents of (patient 6) (13), the others either were in good health or had Bellona Island, a Polynesian Outlier within the Solomon Is- diseases not known to be caused by HTLV-I (Table 1). lands, were closely related to cosmopolitan prototype HTLV-I Among the study participants were a 21-year-old Hagahai (.97% sequence identity), suggesting recent introduction, man from whom PNG-1, a CD8' T-cell line infected with an possibly during this century. These findings are consistent with HTLV-I variant (15, 16), was derived and his 60-year-old a proto-Melanesian HTLV-I strain of archaic presence, which seropositive mother (Table 1). The Hagahai are a small, evolved independently of contemporary cosmopolitan strains, hunter-horticulturalist group living in the fringe highlands of and pose new questions about the origin and global dissem- Papua New Guinea, which made first sustained contact with ination of HTLV-I. government and missionary workers in 1983 (12). Linguisti- cally, the Hagahai have been classified into the Piawi family, Melanesia, the ethnogeographic region that includes New tentatively assigned to the Sepik-Ramu phylum, a non- Guinea, Solomon Islands, Vanuatu, and New Caledonia, Austronesian language group. More recent data provided by contains some of the most remote and inaccessible popula- analyses of HLA-DR antigens suggest that the Hagahai are tions on earth. High prevalences ofantibodies against human distinct from both coastal and highland groups and may T-lymphotropic virus type I (HTLV-I) have been reported for represent descendants of an independent group of migrants several coastal and inland Melanesian populations by using (19). Furthermore, the absence of HLA-A2, a haplotype screening tests such as enzyme immunoassay and gelatin associated with recent Austronesian admixture, indicates particle agglutination (1-9). These reported high prevalences that the Hagahai predate the last Austronesian migration into of antibodies against HTLV-I, however, have been viewed Papua New Guinea, currently dated at -5400 B.P. (20). with skepticism by some investigators because of the failure Also studied were six unrelated Solomon Islanders (in- of such Melanesian sera to neutralize a prototype strain of cluding two residents of Bellona Island) from three of whom HTLV-I (10). Using stringent immunoblot criteria, we re- HTLV-I-infected T-cell lines were derived (17, 18) (Table 1). cently demonstrated an HTLV-I seroprevalence of 14% Bellona (population 650), known also as Mu Ngiki (or "small among the Hagahai (11), a remote, recently contacted hunter- horticulturalist group living in the highland fringe of Papua Abbreviations: HTLV-I, human T-lymphotropic virus type I; New Guinea (12), and seroprevalences of 2%-10%o among HTLV-II, human T-lymphotropic virus type II; STLV-I, simian T-lymphotropic virus type I; PBMC, peripheral blood mononuclear cells. The publication costs of this article were defrayed in part by page charge tTo whom reprint requests should be addressed. payment. This article must therefore be hereby marked "advertisement" lThe sequence reported in this paper has been deposited in the in accordance with 18 U.S.C. §1734 solely to indicate this fact. GenBank data base (accession no. M73745). 7694 Downloaded by guest on September 27, 2021 Medical Sciences: Gessain et al. Proc. Natl. Acad. Sci. USA 88 (1991) 7695 Table 1. Demographic features of six Melanesians and two Polynesians in whom a 522-base-pair (bp) region of the HTLV-I env gene was amplified and sequenced Age and Place or Province or Cultural Patient sex Origin village island affinity Virus Source of provirus 1 21M Papua New Guinea Luyaluya Madang Melanesian HTLV-I Melanesia 1 T-cell line (PNG-1), fresh PBMC 2 60F Luyaluya Madang Melanesian HTLV-I Melanesia 2 Cultured PBMC 3 40F Solomon Islands Marovo New Melanesian HTLV-I Melanesia 3 T-cell line (SI-1) Georgia 4 60F Marau Guadalcanal Melanesian HTLV-I Melanesia 4 Fresh PBMC 5 58M Kolosulu Guadalcanal Melanesian HTLV-I Melanesia 5 T-cell line (SI-5) 6 38M Aola Guadalcanal Melanesian HTLV-I Melanesia 6 Cultured PBMC 7 60F Bellona Central Polynesian HTLV-I Bellona 1 T-cell line (SI-3), fresh PBMC 8 50F Bellona Central Polynesian HTLV-I Bellona 2 Cultured PBMC island"), is, along with Rennell, Tikopia, Anuta (Cherry RESULTS Island), Sikaiana (Stewart Island), and Ontong Java (Lord Genetic Composition of a HTLV-I Quasispecies. Alignment Howe Atoll), a Polynesian-speaking Outlier within the So- and comparison of the nucleotide sequence of each provirus lomon Islands. This island lies 180 km south of Guadalcanal with the published genomic sequence of a prototype (Japa- and is populated by Polynesians. Genetic-distance analysis, nese) HTLV-IATK-1 (23) revealed the existence not only of based on allele frequencies of ABO blood groups, erythro- highly divergent variants of HTLV-I in Melanesia but of a cyte enzymes, and serum proteins, indicate that the inhab- quasispecies (or genetically distinct viral populations) within itants of Bellona are distinct from Melanesians (as they are this HTLV-I variant. A marked divergence of -8% (39-43 linguistically and culturally) despite their close geographical base substitutions in the 522-bp region sequenced) was found proximity (21). in the six Melanesian HTLV-I variants (Table 2). For any Gene Amplification and Sequence Analysis. DNA was iso- individual, the degree of sequence variation was identical (or lated from uncultured (fresh) peripheral blood mononuclear nearly so) whether the DNA was extracted from uncultured cells (PBMC), PBMC in culture for 4 weeks, and HTLV-I- (fresh) PBMC or PBMC cultured for 4 weeks or from T-cell infected T-cell lines derived from the Hagahai (PNG-1) (15, lines derived from Melanesians (PNG-1, SI-1, SI-5), indicat- 16) and Solomon Islanders (SI-1, SI-3, SI-5) (17, 18) by using ing that these variant sequences did not result from selection a nonorganic method (Oncor, Gaithersburg, MD) and was during prolonged maintenance of a few virus-infected cells in subjected to PCR. Oligonucleotide primer pairs derived from culture over many months. near a single base difference) between highly conserved regions of the HTLV-I env gene (sense The identity (only strand, 5'-TTTGAGCGGCCGCTCAAGCTATAGTC- the two Papua New Guinean HTLV-I strains (Melanesia 1 TCCTCCCCCTG-3'; antisense strand, 5'-ACTTAGAAT and 2) was not unexpected because they originated