African origin of human T-lymphotropic virus type 2 (HTLV-2) supported by a potential new HTLV-2d subtype in Congolese Bambuti Efe Pygmies

Abstract

We identified a potential new subtype within human T-cell lymphotropic virus type 2 (HTLV-2), HTLV-2d, present in members of an isolated Efe Bambuti Pygmy tribe. Two of 23 Efe Pygmies were HTLV-2 seropositive, with HTLV-2 Western blot and enzyme-linked immunosorbent assay reactivities. From one of them the entire genome of the HTLV-2 strain Efe2 could be amplified and sequenced. In all gene regions analyzed, this strain was the most divergent HTLV-2 strain, differing by 2.4% (tax/rex) to 10.7% (long terminal repeat) from both subtypes HTLV-2a and HTLV-2b, yet major functional elements are conserved. The similarity between the HTLV-2 Efe2 Gag and Env proteins and the corresponding HTLV-2a and -2b proteins is consistent with the observed serological reactivity. In the proximal pX region, one of the two alternative splice acceptor sites is abolished in HTLV-2 Efe2. Another interesting feature of this potential new subtype is that it has a Tax protein of 344 amino acids (aa), which is intermediate in length between the HTLV-2a Tax protein (331 aa) and the HTLV-2b and -2c Tax proteins (356 aa) and similar to the simian T-cell lymphotropic virus type 2 (STLV-2) PP1664 Tax protein. Together these two findings suggest a different phenotype for the HTLV-2 Efe2 strain. Phylogenetic analyses confirmed that the Pygmy Efe2 strain potentially belonged to a new and quite divergent subtype, HTLV-2d. When the STLV-2 bonobo viruses PP1664 and PanP were used as an outgroup, it was clear that the Pygmy HTLV-2 Efe2 strain had the longest independent evolution and that HTLV-2 evolution is consistent with an African origin.

FIG. 1

Western blots of the eluted antibodies from the 23 Efe Pygmy blood spots. MTA1, HTLV-1-specific recombinant gp41 peptide; K55, HTLV-2-specific recombinant gp41 peptide. Lanes: HTLV-1 and HTLV-2, positive controls; 1 to 23, Pygmy blood spot eluates.

FIG. 2

(A) LTR nucleotide sequence alignment of the prototype strains of HTLV-2a (Mo, accession no. M1006) (68), HTLV-2b (NRA, accession no. L20734) (45), and the potential new subtype HTLV-2d (Efe2). The functional elements are indicated. (B) Tax amino acid alignment of the prototype strains of HTLV-1 (ATK1, accession no. J02029) (67), STLV-L (PH969, accession no. Y07616) (79), STLV-2 (PP1664, accession no. Y14570 [80], and PanP, accession no. U90557 [9]), HTLV-2a (Mo), HTLV-2b (NRA), HTLV-2c (KAY1, accession no. U32875) (13), and the potential new subtype HTLV-2d (Efe2, accession no. Y14365).

FIG. 2

(A) LTR nucleotide sequence alignment of the prototype strains of HTLV-2a (Mo, accession no. M1006) (68), HTLV-2b (NRA, accession no. L20734) (45), and the potential new subtype HTLV-2d (Efe2). The functional elements are indicated. (B) Tax amino acid alignment of the prototype strains of HTLV-1 (ATK1, accession no. J02029) (67), STLV-L (PH969, accession no. Y07616) (79), STLV-2 (PP1664, accession no. Y14570 [80], and PanP, accession no. U90557 [9]), HTLV-2a (Mo), HTLV-2b (NRA), HTLV-2c (KAY1, accession no. U32875) (13), and the potential new subtype HTLV-2d (Efe2, accession no. Y14365).

FIG. 3

Phylogenetic analysis of the entire LTR sequence (homologous to nt 1 to 769 of HTLV-2 Efe2) by the NJ approach described in Materials and Methods. The values on the branches represent the percentages of trees for which the sequences at one end of the branch are a monophyletic group. The main figure represents the tree obtained with STLV-2 as an outgroup, and the inset represents the tree without an outgroup. Details on the different HTLV and STLV strains are given in Materials and Methods. The main clusters contain the following strains: HTLV-2a U.S. and European IDUs, Mo, ED, LA8A, PUEB.RB, NOR2N, and ATL18; HTLV-2a Brazilian and Amerindian, BRAZ.A21, KAY73, and KAY139; HTLV-2b U.S. and European IDUs, ITA47A, NY185, JA, 324, RVP, SPAN130, RC, SPAN129, 130, ITA50A, Gu, I-EA, I-EC, I-GI, I-AM, I-OG, I-OV, I-IT, JG, JL, BF, DP, AA, and JAN; HTLV-2b U.S., Cameroon Pygmy, and Amerindian, WYU2, PYGCAM-1, SEM1051, SEM1050, NRA, PUEB.AG, and PENN7A.

FIG. 4

Phylogenetic analysis of the entire gp21 gene in the env region (homologous to nt 6119 to 6655 of HTLV-2 Efe2) by the NJ approach described in Materials and Methods. The values on the branches represent the percentages of trees for which the sequences at one end of the branch are a monophyletic group. The main figure represents the tree obtained with STLV-2 as an outgroup, and the inset represents the tree without an outgroup. Details on the different HTLV and STLV strains are given in Materials and Methods. The main clusters contain the following strains: HTLV-2a U.S. and European IDUs, Pueblo, and Navajo, Mo, FLW, 408N, Bo, Md, MSA1bp, and DOG; HTLV-2a Brazilian, SP1, SP2, SP3, SP4, and SP5; HTLV-2a Kayapo, KAY1 and KAY2; HTLV-2b Africans, Amerindians, and U.S., European, and Asian IDUs, GAR, WH6, WH7, PAR, 72969N, Gu, 130P, NRA, JPS, PYGCAM-1, VIET13, VIET19, VIET22, VIET32, and VIET35.

FIG. 5

Phylogenetic analysis of the RT protein (homologous to nt 2534 to 4293 of HTLV-2 Efe2) by the NJ approach described in Materials and Methods. The values on the branches represent the percentages of trees for which the sequences at one end of the branch are a monophyletic group. Details on the different HTLV and STLV strains are given in Materials and Methods. The Cosmopolitan and African HTLV-1 strains include ATK1, MT2, TSP-1, ATL-YS, RD-1, BOI, EL, and HS-35.