HTLV-I Genome Research Articles

Transfusion-Transmitted Human T-Cell Lymphotropic Virus Type I Infection in Taiwan: A True Risk and Occasional Coinfection With HepatitisC Virus Shown in a Prospective Study

To study the incidence of human T-cell lymphotropic virus (HTLV) after blood transfusion in Taiwan, serum samples from 699 patients in a prospective study were examined for seroreactivity of anti-HTLV. By an enzyme immunoassay, 9 of the 699 recipients were repeatedly positive. Serial serum samples of these 9 patients were then confirmed with a Western blot analysis and with a polymerase chain reaction (PCR) assay for HTLV-I genome. Four were already positive for anti-HTLV before transfusion, 1 carried antibodies to HTLV-I transiently after transfusion, and only 4 cases had de nova seroconversions. These patients and their family members were called back and tested for HTLV-I genome in the peripheral blood mononuclear cell (PBMC) and plasma. All the serologically positive patients, except the "transient one," were positive for HTLV sequences in the PBMCs. Viral sequences could also be detected in several serum or plasma samples. In the family members, only the spouse of a pretransfusion-positive patient was infected. These results suggested that approximately 0.6% of the blood recipients were infected by HTLV-I through transfusion in Taiwan, and that the frequency of intrafamilial HTLV-I transmission is low. We also observed the unusual coinfection by both HTLV-I and hepatitis C virus in 2 patients, and superinfection of hepatitis C virus after blood transfusion in 1 HTLV-I carrier. Cases of coinfection suggest a prevalence of both viruses in blood donors and warrant further screening.

Absence of HTLV-I and HTLV-II Proviral Genome in the Brains of Patients with Multiple Sclerosis and Amyotrophic Lateral Sclerosis

Previous studies have failed to provide serological evidence to incriminate a retroviral infection in the cause of multiple sclerosis. Gene amplification techniques have also failed to identify retroviral footprints in DNA from peripheral blood leukocytes. Here we provide evidence that proviral DNA of HTLV-I and HTLV-II is not found in the central nervous system tissues of patients with multiple sclerosis, patients with amyotrophic lateral sclerosis and controls.

No evidence of HTLV-I infection in French patients with multiple sclerosis using the polymerase chain reaction.

The polymerase chain reaction (PCR), using three primer pairs in the pol, tax, and env regions of the HTLV-I genome, was unable to detect HTLV-I in the blood samples of 54 caucasian subjects with multiple sclerosis who were seronegative for HTLV-I/II. Seventeen HTLV-I/II seropositive (by ELISA and Western blot) subjects used as positive controls were positive with the three primer pairs. The PCR was negative in 47 healthy HTLV-I/II seronegative (by ELISA) subjects at low risk of HTLV-I infection used as negative controls. These results suggest that there is no association between the occurrence of HTLV-I sequences and the development of multiple sclerosis.

Absence of highly homologous sequence to HTLV‐I in Japanese multiple sclerosis

We tried to detect HTLV-I-related sequences in Japanese patients with multiple sclerosis with a highly sensitive method that employs the polymerase chain reaction (PCR) of genomic DNA followed by Southern blot hybridization analysis. To amplify HTLV-I sequences, we used primers for LTR, pol, gag, and env coding regions. Fourteen patients with definite MS, 14 disease controls, 12 normal controls, and 3 patients with HTLV-I-associated myelopathy (HAM) were investigated. Results of particle aggregation assay for HTLV-I antibodies were negative in serum from all subjects except for the 3 HAM patients. Neither the 14 MS patients nor the 26 controls showed the presence of any highly homologous sequences to HTLV-I. We did observe faint signals for gag, pol, and env coding regions only at low stringent hybridization in some MS patients as well as some normal controls. The nucleotide sequence analysis of the faint bands was more homologous to major histocompatibility complex molecules than the HTLV-I genome.

Type‐C virus‐like particles in a human B‐cell lymphoma cell line

Type-C virus-like particles (VLPs) were found in an Epstein-Barr (EB) virus-infected human B-cell lymphoma cell line, SP-50B, that was established from a patient with non-Hodgkin lymphoma. The cell line continuously produces a small number of type-C VLPs, 150-200 nm in diameter, over 1 year. SP-50B cells were negative for HTLV-I and HTLV-II antigens and did not contain the HTLV-I genome. In addition, two EB virus nuclear antigen (EBNA)-positive B-cell lines, SP-54-Cord and SP-57-CLL, were established from human cord blood and chronic lymphocytic leukemia (CLL), respectively, by coculture with lethally irradiated SP-50B cells. Type-C VLPs with the same morphology were also found in both cell lines.

Mode of transmission of human T-cell leukemia virus type I (HTLV I) in a human promyelocytic leukemia HL60 cell.

HTLV-I propagated in IMR90 human diploid fibroblasts was transmitted to human myeloid leukemia HL60 cells at a low efficiency. After co-cultivation for 3 months, the viral genome was detected in 14/48 HL60 cell clones. Among the 14 HTLV-I-infected clones, 8 contained subgenomic fragments alone or in addition to the complete HTLV-I genome. The frequency of deleted proviruses (9/24 total proviruses) was unexpectedly high. Hirt's supernatant of some of the clones harboring complete HTLV-I genome(s) in the chromosome contained both linear and circular HTLV-I proviral DNAs. The circular DNAs were composed of one LTR and 2 LTR closed circular proviruses. These clones produced infectious HTLV-I constitutively, which was proved by transmission of the viral genome into fresh IMR90 cells by co-cultivation. However, in these clones, re-integration of extrachromosomal provirus into their own chromosomes was not observed.

Efficient Transformation of Previously Activated and Dividing T Lymphocytes by Human T Cell Leukemia-Lymphoma Virus

Modifying previously reported techniques, we attempted to increase the efficiency of human T cell leukemia-lymphoma virus (HTLV) transformation of human T lymphocytes. Lethally irradiated donor cells (DCs) were cultured with target mononuclear cells (TMCs). DCs included ten HTLV+ T cell lines with varying degrees of virus expression or seven cell lines that do not express HTLV. TMCs were prepared from 20 cord and 16 adult peripheral blood samples, including eight patients with acquired immunodeficiency syndrome (AIDS). TMCs were either added directly to the DCs or were first stimulated with phytohemagglutinin (PHA) (5 micrograms/mL) and grown in T cell growth factor (TCGF) prior to exposure to DCs. The presence of integrated HTLV proviral DNA in the transformed cells was determined by dot blot hybridization, utilizing a cloned probe to the HTLV-I genome. HTLV production by transformed TMCs was assessed for HTLV p19, reverse transcriptase, and virus particles. No transformation occurred with T cell donor lines that do not express HTLV. Low virus expressor DCs could only, with rare exception, transform preactivated TMCs. High-titer virus-producing DCs could transform activated and nonactivated cord blood cells and activated adult TMCs. Only MT-2 could routinely transform nonactivated normal adult and activated AIDS TMCs. HUT 102 B2 could transform only one activated AIDS sample, the cells of which initially expressed HTLV-like proteins and virions. Transformed cell lines contained subsets of mature T lymphocytes with variable HTLV expression. Prior activation and culture of the T lymphocytes increases the probability and rate of transformation by HTLV, allowing for biologic detection of low HTLV-producing cells and for in vitro expansion of T lymphocyte subsets from selected patients.

Efficient transformation of previously activated and dividing T lymphocytes by human T cell leukemia-lymphoma virus

Modifying previously reported techniques, we attempted to increase the efficiency of human T cell leukemia-lymphoma virus (HTLV) transformation of human T lymphocytes. Lethally irradiated donor cells (DCs) were cultured with target mononuclear cells (TMCs). DCs included ten HTLV+ T cell lines with varying degrees of virus expression or seven cell lines that do not express HTLV. TMCs were prepared from 20 cord and 16 adult peripheral blood samples, including eight patients with acquired immunodeficiency syndrome (AIDS). TMCs were either added directly to the DCs or were first stimulated with phytohemagglutinin (PHA) (5 micrograms/mL) and grown in T cell growth factor (TCGF) prior to exposure to DCs. The presence of integrated HTLV proviral DNA in the transformed cells was determined by dot blot hybridization, utilizing a cloned probe to the HTLV-I genome. HTLV production by transformed TMCs was assessed for HTLV p19, reverse transcriptase, and virus particles. No transformation occurred with T cell donor lines that do not express HTLV. Low virus expressor DCs could only, with rare exception, transform preactivated TMCs. High-titer virus-producing DCs could transform activated and nonactivated cord blood cells and activated adult TMCs. Only MT-2 could routinely transform nonactivated normal adult and activated AIDS TMCs. HUT 102 B2 could transform only one activated AIDS sample, the cells of which initially expressed HTLV-like proteins and virions. Transformed cell lines contained subsets of mature T lymphocytes with variable HTLV expression. Prior activation and culture of the T lymphocytes increases the probability and rate of transformation by HTLV, allowing for biologic detection of low HTLV- producing cells and for in vitro expansion of T lymphocyte subsets from selected patients.

Characterization of a Novel HTLV-Infected Cell Line

A man from Chile developed an aggressive mature T cell leukemia associated with marked eosinophilia. The neoplastic lymphocytes were of T helper surface phenotype, and they expressed the p24 and p19 antigens of human T cell leukemia virus (HTLV). A cell line (ME) was established from the patient's peripheral blood cells that was initially composed of eosinophils and T and B lymphocytes. The B lymphocytes of the cell line are polyclonal and contain Epstein-Barr virus (EBV) DNA. Many of the T lymphocytes, a few of the B lymphocytes, and none of the eosinophils express HTLV p19 and p24 antigens. By 6 months of culture, the ME line no longer contained eosinophils. A variant line of ME was established; this variant (ME-2) is notable because the cells (greater than 80%) adhere tightly to the bottom of the culture flask; they do not express T lymphocyte markers, but 30% of the cells contain cytoplasmic mu heavy immunoglobulin chains. These pre-B and null lymphocytes contain p19 and p24 antigens (80% of cells), have the HTLV-I genome, and are able to transform normal T lymphocytes in vitro. We isolated a B lymphocyte clone (11A) from ME that expresses cytoplasmic immunoglobulin (70% of cells) and p19 and p24 antigens (75% of cells), contains the EBV and HTLV genomes, and can transform T lymphocytes from normal volunteers. These data show that B lymphocytes can be infected with HTLV, although no disease of HTLV-infected B lymphocytes has been reported. The T lymphocytes from normal adult peripheral blood were easily immortalized (about 70% efficiency) by cocultivation with lethally irradiated ME cells. Twenty-five of 27 of the transformant lines were composed of T lymphocytes with helper antigens, and two of the lines were of T suppressor antigen phenotype. All the cell lines that were tested constitutively produce lymphokines, including colony-stimulating factor (CSF), erythroid-potentiating activity (EPA), macrophage migration-inhibitory factory (MIF), neutrophil-inhibitory factor (NIF), and differentiation-inducing factor (DIF).