Human T-cell lymphotropic virus type 1 subtype C molecular variants among indigenous australians: new insights into the molecular epidemiology of HTLV-1 in Australo-Melanesia

Abstract

Background: HTLV-1 infection is endemic among people of Melanesian descent in Papua New Guinea, the Solomon Islands and Vanuatu. Molecular studies reveal that these Melanesian strains belong to the highly divergent HTLV-1c subtype. In Australia, HTLV-1 is also endemic among the Indigenous people of central Australia; however, the molecular epidemiology of HTLV-1 infection in this population remains poorly documented.

Findings: Studying a series of 23 HTLV-1 strains from Indigenous residents of central Australia, we analyzed coding (gag, pol, env, tax) and non-coding (LTR) genomic proviral regions. Four complete HTLV-1 proviral sequences were also characterized. Phylogenetic analyses implemented with both Neighbor-Joining and Maximum Likelihood methods revealed that all proviral strains belong to the HTLV-1c subtype with a high genetic diversity, which varied with the geographic origin of the infected individuals. Two distinct Australians clades were found, the first including strains derived from most patients whose origins are in the North, and the second comprising a majority of those from the South of central Australia. Time divergence estimation suggests that the speciation of these two Australian clades probably occurred 9,120 years ago (38,000-4,500).

Conclusions: The HTLV-1c subtype is endemic to central Australia where the Indigenous population is infected with diverse subtype c variants. At least two Australian clades exist, which cluster according to the geographic origin of the human hosts. These molecular variants are probably of very ancient origin. Further studies could provide new insights into the evolution and modes of dissemination of these retrovirus variants and the associated ancient migration events through which early human settlement of Australia and Melanesia was achieved.

Figure 1. Map of central Australia.

Indicated are the distribution of the three main Aboriginal language groups (Ngarrkic, Western Desert and Arandic) and the place of origin of the samples used in this study based on place of childhood residence and language (Northern, ?; Southern, ?).

Figure 2. PCR strategy for amplifying both the complete HTLV-1c proviral genome (figure 2A) and the entire Long Terminal Repeat (LTR) region (figure 2B).

Both LTR-gag, pro-pol, pol-env and tax-LTR fragments, which were amplified by PCR and sequenced are shown (orange arrows) as well as internal primers pairs (purple arrows) designed for sequencing reactions (see table 2 for oligonucleotide primers sequences). Nucleotide numbering is according to the HTLV-1 ATK-1 prototypic sequence (Genbank: J02029).

Figure 3. Phylogenetic trees generated with both Neighbor-Joining (NJ) (figure 3A) and Maximum Likelihood (ML) (figure 3B) methods performed in the PAUP program, on a 522-bp fragment of the gp21 env gene, for 53 HTLV-1 available sequences including the 5 proviral sequences generated in this work and two previously characterized Australian proviral strains (MSHR-1 and Aus-RDJ).

The selected model was the Tamura Nei. a-f correspond to the major HTLV-1 subtypes. HTLV-1c subtype corresponds to the Australo-Melanesian one including the new Australian proviral sequences: Aus-CS, Aus-DF, Aus-NR, Aus-GN and Aus-GM (Genbank: KF242506, KF242505, JX891479, KF242507 and JX891478 respectively) and 2 previously described Australian sequences: MSHR-1 (M92818) and Aus-RDJ (JX891480); 9 sequences from Vanuatu: VAN54, VAN136, VAN251, VAN335, PE376, EM5, PE376, ESH18 and ESW44 (Genbank: AY549880, AY549882, AY549879, AY549881, EF061856, EF061884, EF061856, EF061885 and EF061886 respectively); 7 sequences from the Solomon Islands: MEL3, MEL4, MEL5, MEL6, MEL8, MEL9, MEL10 (Genbank: M94198, M94199, M94200, M93099, U11578, U11580, U11566 respectively) and 3 sequences from Papua New Guinea: MEL1, MEL2 and MEL7 (Genbank: L02533, M94197 and U11576, respectively).

Figure 4. Phylogenetic trees generated with both Neighbor-Joining (figure 4A) and Maximum Likelihood (figure 4B) methods, performed in the PAUP program, on a 746-bp fragment of the LTR region, for 55 HTLV-1 available sequences including the 27 proviral sequences generated in this work.

The selected model was the General Time-Reversible (GTR). a-g correspond to the major HTLV-1 subtypes. HTLV-1c subtype corresponds to the Australo-Melanesian one including the 23 new Australian proviral sequences (Genbank: KC786899-KC786919 and JX891478-JX891479) and 4 sequences from Vanuatu (Genbank: KC786962-KC786965). ?, HTLV-1c subtype strains from Northern part of central Australia; ?, HTLV-1c subtype strains from Southern part of central Australia.

Figure 5. Phylogenetic trees generated with both Neighbor-Joining (figure 5A) and Maximum Likelihood (figure 5B) methods performed in the PAUP program, on a 2,346-bp fragment of the colinearized gag-tax genes, for 29 HTLV-1 available sequences including the 27 proviral sequences generated in this work.

The selected model was the Tamura Nei. ATK-1 strain was used as an outgroup (Genbank: J02029). The Australo-Melanesian HTLV-1c subtype includes the 23 new Australian sequences characterized in this study (Genbank: KC786920-KC786961 and JX891478-JX891479) and 4 proviral sequences from Vanuatu (Genbank: KC786966-KC786973). ?, HTLV-1c subtype strains from Northern part of central Australia; ?, HTLV-1c subtype strains from Southern part of central Australia.

Figure 6. Phylogenetic trees generated with both Neighbor-Joining (figure 6A) and Maximum Likelihood (figure 6B) methods, performed in the PAUP program, on a 6,000-bp fragment of the colinearized gag-pol-env-tax genes, for 23 HTLV-1/-2/-3 available sequences including the 4 Australian proviral sequences generated in this work.

The selected model was the Tamura Nei. The Australo-Melanesian HTLV-1c subtype includes the 4 complete Australian sequences characterized in this study (Genbank: KC786920-KC786961 and JX891478-JX891479) and the MEL5 proviral sequence from the Solomon Islands (Genbank: L02534). ?, HTLV-1c subtype strains from Northern part of central Australia; ?, HTLV-1c subtype strains from Southern part of central Australia.