The genetic drift of human papillomavirus type 16 is a means of reconstructing prehistoric viral spread and the movement of ancient human populations.

We have investigated the diversity of a hypervariable segment of the human papillomavirus type 16 (HPV-16) genome among 301 virus isolates that were collected from 25 different ethnic groups and geographic locations. Altogether, we distinguished 48 different variants that had diversified from one another along five phylogenetic branches. Variants from two of these branches were nearly completely confined to Africa. Variants from a third branch were the only variants identified in Europeans but occurred at lower frequency in all other ethnic groups. A fourth branch was specific for Japanese and Chinese isolates. A small fraction of all isolates from Asia and from indigenous as well as immigrant populations in the Americas formed a fifth branch. Important patterns of HPV-16 phylogeny suggested coevolution of the virus with people of the three major human races, namely, Africans, Caucasians, and East Asians. But several minor patterns are indicative of smaller bottlenecks of viral evolution and spread, which may correlate with the migration of ethnic groups in prehistoric times. The colonization of the Americas by Europeans and Africans is reflected in the composition of their HPV-16 variants. We discuss arguments that today's HPV-16 genomes represent a degree of diversity that evolved over a large time span, probably exceeding 200,000 years, from a precursor genome that may have originated in Africa. The identification of molecular variants is a powerful epidemiological and phylogenetic tool for revealing the ancient spread of papillomaviruses, whose trace through the world has not yet been completely lost.     

Intrauterine transmission of human T-cell leukemia virus type I in rats.

To analyze intrauterine transmission, MT-2 cells, a human T-cell line producing human T-cell leukemia virus type I (HTLV-I), were injected into eight pregnant F344 rats, and cesarean section was performed at day 23 of pregnancy. HTLV-I provirus was detected by PCR in the liver and spleen taken from one of the eight fetuses. Moreover, 71 offspring were delivered by cesarean section from the remaining seven dams and fostered by seven normal rats. HTLV-I provirus was detected in peripheral blood mononuclear cells in 2 of the 71 offspring 4 weeks after cesarean section. These results indicate for the first time the intrauterine transmission of HTLV-I. To confirm the postnatal transmission, MT-2 cells were injected into a dam within 24 h after delivery, and six offspring were fostered by this dam. HTLV-I provirus was detected in peripheral blood mononuclear cells of all six offspring. This animal model may be useful for analysis and prevention of mother-to-child transmission of HTLV-I.     

Genetic heterogeneity in human T-cell leukemia/lymphoma virus type II.

DNA from the peripheral blood mononuclear cells of 17 different individuals infected with human T-cell lymphoma/leukemia virus type II (HTLV-II) was successfully amplified by the polymerase chain reaction (PCR) with the primer pair SK110/SK111. This primer pair is conserved among the pol genes of all primate T-cell lymphoma viruses (PTLV) and flanks a 140-bp fragment of DNA which, when used in comparative analyses, reflects the relative degree of diversity among PTLV genomes. Cloning, sequencing, and phylogenetic comparisons of these amplified 140-bp pol fragments indicated that there are at least two distinct genetic substrains of HTLV-II in the Western Hemisphere. These data were confirmed for selected isolates by performing PCR, cloning, and sequencing with to 10 additional primer pair-probe sets specific for different regions throughout the PTLV genome. HTLV-II isolates from Seminole, Guaymi, and Tobas Indians belong in the new substrain of HTLV-II, while the prototype MoT isolate defines the original substrain. There was greater diversity among HTLV-II New World strains than among HTLV-I New World strains. In fact, the heterogeneity among HTLV-II strains from the Western Hemisphere was similar to that observed in HTLV-I and simian T-cell lymphoma/leukemia virus type I isolates from around the world, including Japan, Africa, and Papua New Guinea. Given these geographic and anthropological considerations and assuming similar mutation rates and selective forces among the PTLV, these data suggest either that HTLV-II has existed for a long time in the indigenous Amerindian population or that HTLV-II isolates introduced into the New World were more heterogeneous than the HTLV-I strains introduced into the New World.     

Multiple interspecies transmissions of human and simian T-cell leukemia/lymphoma virus type I sequences

Using two sets of nucleotide sequences of the human and simian T-cell leukemia/lymphoma virus type I (HTLV-I/STLV-I), one consisting of 522 bp of the env gene from 70 viral strains and the other a 140-bp segment from the pol gene of 52 viral strains, I estimated cladograms based on a statistical parsimony procedure that was developed specifically to estimate within-species gene trees. An extension of a nesting procedure is offered for sequence data that forms nested clades used in hypothesis testing. The nested clades were used to test three hypotheses relating to transmission of HTLV/STLV sequences: (1) Have cross-species transmissions occurred and, if so, how many? (2) In what direction have they occurred? (3) What are the geographic relationships of these transmission events? The analyses support a range of 11-16 cross-species transmissions throughout the history of these sequences. Additionally, outgroup weights were assigned to haplotypes using arguments from coalescence theory to infer directionality of transmission events. Conclusions on geographic origins of transmission events and particular viral strains are inconclusive due to small samples and inadequate sampling design. Finally, this approach is compared directly to results obtained from a traditional maximum parsimony approach and found to be superior at establishing relationships and identifying instances of transmission.      

In vivo cellular tropism of human T-cell leukemia virus type 1.

To establish the phenotype of human T-cell leukemia virus type 1 (HTLV-1)-infected cells in peripheral blood, the polymerase chain reaction was used to detect and quantitate viral DNA in subpopulations of leukocytes obtained from patients with tropical spastic paraparesis and asymptomatic carriers. HTLV-1 could not be detected in peripheral blood mononuclear cells thoroughly depleted of T lymphocytes (E- CD3-), nor could it be detected in highly enriched populations of B lymphocytes (E- CD19+), monocytes (E- CD14+), or natural killer cells (E- CD16+). T lymphocytes were strongly positive for HTLV-1, and fractionation of this population revealed that 90 to 99% of the HTLV-1 DNA segregated with the CD4+ CD8- and CD45RO+ subsets. No difference between the cell type distribution of HTLV-1 in the asymptomatic carrier and the subjects with tropical spastic paraparesis was evident. Southern hybridization of genomic DNA prepared from the peripheral blood of HTLV-1 carriers indicated that up to 10% of circulating leukocytes may carry the HTLV-1 provirus.     

The genetic background as a determinant of human T-cell leukemia virus type 1 proviral load

Human T-cell leukemia virus type 1 (HTLV-1) is etiologically linked with HTLV-1-associated diseases. HTLV-1 proviral load is higher in persons with adult T-cell leukemia and HTLV-1-associated myelopathy/tropical spastic paraparesis than in asymptomatic carriers. However there are little data available on the factors controlling HTLV-1 proviral load in carriers. To study the effect of genetic background on HTLV-1 proviral load, we employed a mouse model of HTLV-1 infection that we had established. Here we analyzed nine strains of mice and found there is a great variation of proviral load among mouse strains that is not necessarily dependent on major histocompatibility complex. The antibody response is also different among these strains. To our knowledge, this is the first demonstration of the importance of the genetic background other than major histocompatibility complex controlling the HTLV-1 proviral load.     

Phylogenetic associations of human and simian T-cell leukemia/lymphotropic virus type I strains: evidence for interspecies transmission.

Homologous env sequences from 17 human T-leukemia/lymphotropic virus type I (HTLV-I) strains from throughout the world and from 25 simian T-leukemia/lymphotropic virus type I (STLV-I) strains from 12 simian species in Asia and Africa were analyzed in a phylogenetic context as an approach to resolving the natural history of these related retroviruses. STLV-I exhibited greater overall sequence variation between strains (1 to 18% compared with 0 to 9% for HTLV-I), supporting the simian origin of the modern viruses in all species. Three HTLV-I phylogenetic clusters or clades (cosmopolitan, Zaire, and Melanesia) were resolved with phenetic, parsimony, and likelihood analytical procedures. Seven phylogenetic clusters of STLV-I were resolved with the most primitive (deeply rooted) divergence involving several STLV-I strains from Asian primate species. Combined analysis of HTLV-I and STLV-I revealed that neither STLV-I clusters nor HTLV-I clusters recapitulated host species specificity; rather, multiple clades from the same species were closer to clades from other species than to each other. We interpret these evolutionary associations as support for the occurrence of multiple discrete interspecies transmissions of ancestral viruses between primate species (including human) that led to recognizable phylogenetic clades that persist in modern species. Geographic concordance of divergent host species that harbor closely related viruses reinforces that physical feasibility for hypothesized interspecies virus transmission in the past and in the present.     

Interspecies transmission of macaque simian T-cell leukemia/lymphoma virus type 1 in baboons resulted in an outbreak of malignant lymphoma.

An outbreak of malignant lymphoma has been observed in one of the baboon (Papio hamadryas) stocks of Sukhumi Primate Center. More than 300 cases in this "high-lymphoma stock" have been registered since 1967. Human T-cell lymphotropic virus type 1 (HTLV-1)-related virus was implicated as the etiologic agent of Sukhumi baboon lymphoma. The origin of this virus remained unclear. Two possibilities were originally considered: the origin could be baboon simian T-cell leukemia/lymphoma virus type 1 (STLV-1) or HTLV-1 (before the outbreak started, some Sukhumi baboons were inoculated with human leukemic material). The third possibility entered recently: interspecies transmission of rhesus macaque STLV-1 to baboons. It was prompted by the finding of very close similarity between STLV-1 991-1cc (the strain isolated from a non-Sukhumi baboon inoculated with material from a Sukhumi lymphomatous baboon) and rhesus STLV-1. To test this hypothesis, we investigated 37 Sukhumi STLV-1 isolates from baboons of high-lymphoma stock by PCR discriminating rhesus type and baboon type STLV-1 isolates. All of them were proved to be rhesus type STLV-1. In contrast, all six STLV-1 isolates from baboons belonging to other stocks or populations were of baboon type. The PCR results were fully confirmed by DNA sequence data. The partial env gene gene sequences of all four STLV-1 isolates from Sukhumi lymphomatous baboons were 97 to 100% similar to the sequence of known rhesus STLV-1 and only 85% homologous with the sequence of conventional baboon STLV-1. Thus, interspecies transmission of STLV-1 from rhesus macaques (or closely related species) to baboons occurred at Sukhumi Primate Center. Most probably this event initiated the outbreak of lymphoma in Sukhumi baboons.     

Expression of human T-cell leukemia virus type I protease in Escherichia coli.

Human T-cell leukemia virus type I (HTLV-I) genome is believed to encode its own protease, although the protease has not yet been detected. To identify the HTLV-I protease, an in-frame gag (3' portion)-prt region was expressed in Escherichia coli. The 14-kDa product was detected using antisera against a synthetic peptide mimicking the fragment of HTLV-I protease, although the molecular weight of the primary translational product was 27,000. A cell extract had a proteolytic activity to cleave a synthetic peptide substrate containing the cleavage site of gag p19/p24 at the correct site in vitro. Replacement of the putative active site Asp-64 with Gly abolished both in vivo processing activity and in vitro proteolytic activity. These results suggest that the 14-kDa product is the mature enzymatically active HTLV-I protease generated through posttranslational autoprocessing in E. coli.     

Low CD4/CD8 T-cell ratio associated with inflammatory arthropathy in human T-cell leukemia virus type I Tax transgenic mice

Background

Human T-cell leukemia virus type I (HTLV-1) can cause an aggressive malignancy known as adult T-cell leukemia/lymphoma (ATL) as well as inflammatory diseases such as HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP). A transgenic mouse that expresses HTLV-1 Tax also develops T-cell leukemia/lymphoma and an inflammatory arthropathy that resembles rheumatoid arthritis. The aim of this study was to identify the primary T-cell subsets involved in the development of arthropathy in Tax transgenic mice.

Principal Findings

By 24 months of age, Tax transgenic mice developed severe arthropathy with a cumulative incidence of 22.8%. The pathological findings of arthropathy in Tax transgenic mice were similar to those seen in human rheumatoid arthritis or mouse models of rheumatoid arthritis, with synovial proliferation and a positive rheumatoid factor. Before the onset of spontaneous arthropathy, young and old Tax transgenic mice were not sensitive to collagen and did not develop arthritis after immunization with type II collagen. The arthropathic Tax transgenic mice showed a significantly decreased proportion of splenic CD4⁺ T cells, whereas the proportion of splenic CD8⁺ T cells was increased. Regulatory T cells (CD4⁺CD25⁺Foxp3⁺) were significantly decreased and CD8⁺ T cells that expressed the chemokine receptor CCR4 (CD8⁺CCR4⁺) were significantly increased in arthropathic Tax transgenic mice. The expression of tax mRNA was strong in the spleen and joints of arthropathic mice, with a 40-fold increase compared with healthy transgenic mice.

Conclusions

Our findings reveal that Tax transgenic mice develop rheumatoid-like arthritis with proliferating synovial cells in the joints; however, the proportion of different splenic T-cell subsets in these mice was completely different from other commonly used animal models of rheumatoid arthritis. The crucial T-cell subsets in arthropathic Tax transgenic mice appear to resemble those in HAM/TSP patients rather than those in rheumatoid arthritis patients.     

Human T-cell lymphotropic virus type I and adult T-cell leukemia/lymphoma outside Japan and the Caribbean Basin

Ninety-six patients with the diagnosis of adult T-cell leukemia/lymphoma (ATLL) were identified in countries outside Japan and the Caribbean Basin. Seventy-four of these patients were initially diagnosed in the United States; 25 of 52 patients whose places of birth were known had been born in the United States. The detection of 14 patients born in the southeastern United States, all black, indicates a group deserving particular attention for studies of human T-cell lymphotropic virus type I (HTLV-I), a suspected etiologic agent in most cases of ATLL. Although geographic clustering of ATLL in areas endemic for HTLV-I, particularly southwest Japan and the Caribbean Basin, is a dramatic feature of this disease, a review of the literature indicates that HTLV-I-associated ATLL probably occurs sporadically in a much wider distribution, the disease being diagnosed in native-born African, Chinese, European, and Latin American patients. A registry for ATLL cases is suggested, to assist in the identification of risk factors for this disease and, at the same time, improve case definitions and early diagnosis.     

Pseudotyping with human T-cell leukemia virus type I broadens the human immunodeficiency virus host range.

Several epidemiologic and clinical studies suggest that patients coinfected with human immunodeficiency virus (HIV), the primary etiologic agent in AIDS, and other viruses, such as cytomegalovirus or human T-cell leukemia virus (HTLV), have a more severe clinical course than those infected with HIV alone. Cells infected with two viruses can, in some cases, give rise to phenotypically mixed virions with altered or broadened cell tropism and could therefore account for some of these findings. Such pseudotypes could alter the course of disease by infecting more tissues than are normally infected by HIV. We show here that HIV type 1 (HIV-1) efficiently incorporates the HTLV type I (HTLV-I) envelope glycoprotein and that both HIV-1 and HTLV-II accept other widely divergent envelope glycoproteins to form infectious pseudotype viruses whose cellular tropisms and relative abilities to be transmitted by cell-free virions or by cell contact are determined by the heterologous envelope. We also show that the mechanism by which virions incorporate heterologous envelope glycoproteins is independent of the presence of the homologous glycoprotein or heterologous gag proteins. These results may have important implications for the mechanism of HIV pathogenesis.     

Transcomplementation of simian immunodeficiency virus Rev with human T-cell leukemia virus type I Rex.

A molecular clone of the simian immunodeficiency virus SIVSMM isolate PBj14, lacking the ATG initiation codon for Rev protein (PBj-1.5), did not produce virus or large unspliced or singly spliced viral RNA upon transfection of HeLa cells. Low but significant levels of virus and large viral RNA production were observed upon transfection of PBj-1.5 into HeLa Rev cells expressing the rev gene of human immunodeficiency virus type 1. Furthermore, abundant virus and large viral RNA production occurred upon transfection of PBj-1.5 into HeLa Rex cells expressing the rex gene of human T-cell leukemia virus type I. Virus produced from HeLa Rex and HeLa Rev transfections was infectious, produced large amounts of virus, and was cytopathic for Rex-producing MT-4 cells. In contrast, no or only low levels of virus production were observed upon infection of H9 cells. These studies show that a defective SIV rev gene can be transcomplemented with human immunodeficiency virus type 1 Rev and with high efficiency by human T-cell leukemia virus type I Rex, and they suggest that rev-defective viruses could serve as a source for production of a live attenuated SIV vaccine.     

In vivo genomic variability of human T-cell leukemia virus type I depends more upon geography than upon pathologies.

To investigate the geography- and disease-associated genomic variation of human T-cell leukemia virus type I (HTLV-I), we studied ex vivo DNA from peripheral blood lymphocytes from nine patients by polymerase chain reaction and direct DNA sequencing. For each viral strain, 1,917 bp was sequenced, including parts of the long terminal repeat, the env gene, and the px II, px III, and px IV coding frames of the px region. The number of genomic variations observed in the U3 region of the long terminal repeat was higher than that seen in the env and px genes. Very few mutations were present in the px II and px III genes. In contrast, the px IV open reading frame exhibited numerous single point mutations. While no specific mutation could be linked to any pathology (adult T-cell leukemia/lymphoma or tropical spastic paraparesis/HTLV-I-associated myelopathy), variations among HTLV-I isolates from different geographic areas (Ivory Coast, Caribbean, and Japan) existed. The Ivory Coast HTLV-I appeared to represent a group by itself.