Expression and Purification of a Novel Computationally Designed Antigen for Simultaneously Detection of HTLV-1 and HBV Antibodies.

Computational tools are reliable alternatives to laborious work in chimeric protein design. In this study, a chimeric antigen was designed using computational techniques for simultaneous detection of anti-HTLV-I and anti-HBV in infected sera. Databases were searched for amino acid sequences of HBV/HLV-I diagnostic antigens. The immunodominant fragments were selected based on propensity scales. The diagnostic antigen was designed using these fragments. Secondary and tertiary structures were predicted and the B-cell epitopes were mapped on the surface of built model. The synthetic DNA coding antigen was sub-cloned into pGS21a expression vector. SDS-PAGE analysis showed that glutathione fused antigen was highly expressed in E. coli BL21 (DE3) cells. The recombinant antigen was purified by nickel affinity chromatography. ELISA results showed that soluble antigen could specifically react with the HTLV-I and HBV infected sera. This specific antigen could be used as suitable agent for antibody-antigen based screening tests and can help clinicians in order to perform quick and precise screening of the HBV and HTLV-I infections.

Adult T-cell leukemia/lymphoma in an adolescent presenting with skin lesions.

Adult T-cell leukemia/lymphoma is a T-cell malignancy caused by the human T-cell lymphotropic virus-I. Adult T-cell leukemia/lymphoma is primarily a disease of adults due to the long latency period between initial infection and development of leukemia. We present a case of acute adult T-cell leukemia/lymphoma in an adolescent. Skin lesions had appeared 3 years earlier and were the initial sign of human T-cell lymphotropic virus-I infection and T-cell malignancy. Her disease failed to respond to both intensive chemotherapy and antiviral therapy. Cutaneous lesions are sometimes the initial sign of adult T-cell leukemia/lymphoma and early recognition is imperative.

Increased frequency of CD4+T cells expressing fractalkine receptor CX3CR1 in patients with HTLV-1 associated myelopathy/tropical spastic paraparesis (HAM/TSP), but its AIDS susceptible polymorphisms are not associated with the disease.

To investigate whether fractalkine receptor CX3CR1 polymorphisms that have been associated with rapid progression to AIDS among HIV-1 positive individuals also affects the risk of human T cell lymphotropic virus type 1 (HTLV-1) associated myelopathy/tropical spastic paraparesis (HAM/TSP), we compared the allele frequencies of V249I and T280M between 233 HAM/TSP patients and 213 HTLV-1 seropositive asymptomatic carriers (HCs). Although the frequency and absolute number of peripheral blood CX3CR1+CD4+T cells were significantly increased in HAM/TSP patients compared to HCs and uninfected controls independent of HTLV-1 trans-activator protein Tax, we could not observe any association between the two polymorphisms and the risk of HAM/TSP in our cohort.

A prospective uncontrolled trial of fermented milk drink containing viable Lactobacillus casei strain Shirota in the treatment of HTLV-1 associated myelopathy/tropical spastic paraparesis.

Ten patients with human T-cell lymphotropic virus type-1 (HTLV-1)-associated myelopathy/tropical spastic paraparesis (HAM/TSP) were treated in an uncontrolled preliminary trial by oral administration of viable Lactobacillus casei strain Shirota (LcS) containing fermented milk. HTLV-1 provirus load, motor function, neurological findings, and immunological parameters were evaluated after 4 weeks. Although LcS did not change the frequencies or absolute numbers of all the examined cell surface phenotypes of peripheral blood mononuclear cells, NK cell activity was significantly increased after 4 weeks of oral administration of LcS preparation. Improvements in spasticity (modified Ashworth Scale scores) and urinary symptoms were also seen after LcS treatment. No adverse effect was observed in all the 10 patients throughout the study period. Our results indicated that LcS may be a safe and beneficial agent for the treatment of HAM/TSP; therefore randomized controlled studies are warranted.

Human T-cell lymphotropic virus type 1 (HTLV-1) and lymphoid malignancies in Dominica: a seroprevalence study.

Human T-cell lymphotropic virus type 1 (HTLV-1) is endemic in certain regions of the world where it is associated with lymphoid malignancies. Herein we aim to describe the seroprevalence of HTLV-1 in lymphoid malignancies in Dominica. We carried out a 10-year retrospective study of histologically proven hematologic malignancies and HTLV-1 seropositivity at the Princess Margaret Hospital, Dominica. Ninety-eight cases were reviewed (59% males, 41% females), ranging in age from 3 to 91 years. HTLV-1 was seropositive in 38.6% (31/80) of all hematologic malignancies. Three of 6 cases of Hodgkin disease (50%), 16 of 36 (44.4%) of non-Hodgkin lymphoma, and 3 out of 8 unclassified lymphomas (37.5%) were seropositive; all 6 cases (100%) of acute adult T-cell leukemia/lymphoma (ATLL) were seropositive. One case each of chronic lymphocytic leukemia and myeloproliferative disorder was seropositive. HTLV-1-seropositive lymphomas presented at a younger age than did seronegative cases. Thus, HTLV-1 is significantly associated with lymphoid malignancies in Dominica, and further studies are needed before a causal relationship with Hodgkin disease can be established.

Expression of the bovine leukemia virus envelope glycoprotein (gp51) by recombinant baculovirus and its use in an enzyme-linked immunosorbent assay.

The gene encoding the major envelope glycoprotein (gp51) with its signal sequence, represented by an additional NH2-terminal 33-residue amino acid sequence of bovine leukemia virus (BLV), was inserted into a baculovirus transfer vector. A recombinant virus expressing a secreted gp51 protein in insect cells was isolated. The recombinant gp51 expressed was characterized by using an anti-BLV monoclonal antibody by both Western blotting analysis and enzyme-linked immunosorbent assay (ELISA). The secreted gp51 was used as an antigen, and an ELISA with recombinant gp51 (rgp51) was developed for the detection of BLV antibodies. This new procedure was compared with a previous ELISA method for the detection of BLV antibodies and an agar gel immunodiffusion test performed with an unpurified BLV antigen preparation. The comparative testing of field samples showed that the ELISA with rgp51 is more specific and also suitable for the testing of pooled sera.

Prevalence of antibodies to hepatitis C virus, HIV and human T-cell leukaemia/lymphoma viruses in injecting drug users in Tayside, Scotland, 1993-7.

The prevalence of blood-borne viruses in injecting drug users (IDUs) in Tayside, Scotland was determined by testing serum samples from IDUs who underwent attributable HIV antibody testing during 1993-7. The prevalence of antibodies to HIV was 29/802, (3.6%); to hepatitis C virus (HCV) 451/691, (65.3%); and to human T-cell leukaemia/lymphoma viruses type 1 and 2 (HTLV) 0/679, (0.0%). The prevalence of HIV and HCV antibodies were higher in subjects over the age of 25 (P = 0.03 and P = 0.001, respectively). During 1993-7 the prevalence of HCV fell only in younger female IDUs (P < 0.01). HIV prevalence has declined dramatically since 1985, when a rate of 40% was recorded in similar populations. Harm reduction measures have failed to control HCV the spread of infection among IDUs in Tayside, as indicated by the high proportion of antibody positive IDUs, particularly males under the age of 25. Future studies should address the nature and effective reduction of continuing risk taking among IDUs in Tayside.

Viral markers and use of factor products among Finnish patients with bleeding disorders.

In the seventh national voluntary cross-sectional survey (in 1999) of Finnish patients with haemophilia A or B, type 3 von Willebrand disease or factor XIII deficiency, a plasma sample was received from 193 patients (67%). The samples were tested for hepatitis B and C, human immunodeficiency virus (HIV) and human T-cell leukaemia virus (HTLV) antibodies. Fifty-one percent of the patients were hepatitis C antibody positive and 34% hepatitis B core antibody positive. None of the patients had antibodies against HIV or HTLV. Eighteen percent of the patients had an elevated alanine aminotransferase activity. Abnormal alanine aminotransferase was significantly associated with hepatitis C seropositivity. No new seroconversions were detected among the haemophiliacs or patients with type 3 von Willebrand disease when compared with the last two surveys in 1993 and 1996, and there was no seroconversion in sole users of solvent/detergent-treated factor products. Currently, 32% of the patients use prophylactic factor treatment as their principal mode of therapy, particularly the younger patients with severe forms of the bleeding diseases.

Human T cell leukaemia/lymphoma virus infection in pregnant women in the United Kingdom: population study.

OBJECTIVE: To assess the prevalence of human T cell leukaemia/lymphoma virus (HTLV) infection in pregnant women in the United Kingdom. DESIGN: Population study. SUBJECTS: Guthrie card samples from babies born in 1997-8. Samples were linked to data on mother's age and ethnic status and parents' country of birth and then anonymised. SETTING: North Thames Regional Health Authority. MAIN OUTCOME MEASURES: Presence of antibodies against HTLV in eluates tested by gelatin particle agglutination assay and results confirmed by immunoblot. RESULTS: Of 126 010 samples tested, 67 had confirmed antibodies to HTLV (59 HTLV-I, 2 HTLV-II, 6 untyped) and six had indeterminate results. Seroprevalence was 17.0 per 1000 (95% confidence interval 9.2 to 28.3) in infants whose mothers were born in the Caribbean, 3.2/1000 (1.5 to 5.9) with mothers born in west and central Africa, and 6.8/1000 (3.1 to 12.9) in infants of black Caribbean mothers born in non-endemic regions. In infants with no known risk (both parents born in non-endemic regions and mother not black Caribbean) seroprevalence was 0.06-0.12 per 1000. Mother's country of birth, father's country of birth, and mother's ethnic status were all independently associated with neonatal seroprevalence. An estimated 223 (95% confidence interval 110 to 350) of the 720 000 pregnant women each year in the United Kingdom are infected with HTLV. CONCLUSIONS: The prevalence of HTLV and HIV infections in pregnant women in the United Kingdom are comparable. The cost effectiveness of antenatal HTLV screening should be evaluated, and screening of blood donations should be considered.

Seroindeterminate patterns and seroconversions to human T-lymphotropic virus type I positivity in blood donors from Martinique, French West Indies.

BACKGROUND: Screening for human T-lymphotropic virus type I (HTLV-I) antibodies in volunteer blood donors has been systematic in the French West Indies since 1989. Western blot-indeterminate results are commonly obtained. The significance of these indeterminate serologic patterns in HTLV-I-endemic areas is still unclear. STUDY DESIGN AND METHODS: During a 2-year period, 9759 blood donors were tested for HTLV-I antibodies. The epidemiologic features of HTLV-I-seropositive, -seroindeterminate, and -seronegative donors were compared. A lookback investigation was performed for the HTLV-I-seropositive donors, and the HTLV-I-seroindeterminate individuals were followed up. RESULTS: Thirty-nine donors (0.4%) were HTLV-I seropositive and 49 (0.5%) were seroindeterminate. The age and sex ratio characteristics of the seroindeterminate donors are divergent from those of the HTLV-I-seropositive group and are more like those of the seronegative population. However, during the study period, three cases of seroconversion after an initial seroindeterminate profile were reported. Two cases were detected through follow-up of 38 HTLV-I-seroindeterminate donors over a mean of 8 months (2-24 months). The third seroconverter belonged to the HTLV-I-seropositive group and was identified through lookback investigation. This case is atypical, with p19 reactivity for several months before HTLV-I seropositivity. CONCLUSION: These findings indicate that, although HTLV-I-seroindeterminate donors mainly are HTLV-I-noninfected, the rate of seroconversion in a repeat blood donor population from an endemic region must be taken into consideration. Moreover, the case of delayed seroconversion observed in this study suggests the difficulty of counseling seroindeterminate blood donors in endemic regions.

Human T lymphotropic virus type I (HTLV-I) infection in neurological patients in Salvador, Bahia, Brazil.

HTLV-I infection represents a major health concern in endemic areas throughout the world, such as Salvador, the main city of Bahia State, with socio-demographic characteristics similar to sub-Saharan African cities, located in the Northeast of Brazil. In order to provide an estimate of the frequency distribution, and range of neurological manifestations potentially related to HTLV-I infection in this city, we conducted a cross-sectional clinical-epidemiological study to determine the prevalence of this infection in patients with neurological diseases. Patients exhibiting vascular diseases, tumoral diseases or trauma were excluded. Over a period of 16 months, we studied 322 consecutive patients with chronic neurological diseases, who attended the neurological clinics of two major hospitals in Salvador. Overall, the prevalence of HTLV-I infection among the patients was 20.9% (67/320). However, the prevalence among the 104 patients with chronic myelopathy was 50.0% (52/104). It was observed that the major prevalence of HTLV-I was between the ages of 40 and 60 years with a female predominance. Our data indicate that, in Salvador city, HTLV-I is associated with chronic myelopathies or myeloneuropathies, which seem to be the only neurological diseases associated with HTLV-I.

Human T cell leukemia virus type I-associated myelopathy in a patient with systemic lupus erythematosus.

A case of human T cell leukemia virus type I (HTLV-1) associated myelopathy (HAM)/tropical spastic paraparesis (TSP) with 14-year history of systemic lupus erythematosus (SLE) is reported. For 9 years, the numbness of the feet and sacral region progressed with occasional urinary incontinence and constipation. She was admitted to hospital due to gait disturbance and aggravation of SLE and the diagnosis of HAM/TSP was confirmed, indicating that HTLV-1 infection is associated with the development of not only HAM/TSP but also SLE.

Is it necessary to test patients with immune thrombocytopenic purpura (ITP) for seropositivity to HTLV-1?

HTLV-111 (HIV-1) has been shown to be associated with thrombocytopenia of a type resembling immune thrombocytopenic purpura (ITP). HTLV-1 is a retrovirus similar to HIV-I (HTLV-III) in a number of features, such as CD4 tropism. It is responsible for several clinical entities, including adult T-cell leukemia/lymphoma. The relationship, if any, of HTLV-1 and thrombocytopenia has not been systematically studied. To determine how frequently ITP patients are commonly infected with HTLV-1, the following study was performed. Frozen serum samples from 123 randomly selected patients with ITP were thawed and tested for antibodies to HTLV-1 by enzyme-linked immunoabsorbent assay. Positives were confirmed by Western blot. Three patients were initially found to be positive for HTLV-1. One was a female of Caribbean ancestry, one was a male HIV-1+ patient, and one was an adolescent female with no known risk factors for HIV-1. The two females later tested negative for HTLV-1. As a screening program for HTLV-1 antibodies was not introduced into blood banks until November 1988, there may have been passive transfer of the virus from intravenous immunoglobulin that these patients had received. This study of a large number of ITP patients shows that it is extremely unlikely that they are infected with HTLV-1, and, therefore, it is unnecessary to screen ITP patients for seropositivity to HTLV-1.

Differential serological diagnosis of HTLV-I and HTLV-II infection by external membrane protein peptide-based enzyme immunoassays.

BACKGROUND: HTLV antibody screening assays detect both antibodies to the etiological agent of adult T-cell leukemia and tropical spastic paraparesis HTLV-I and to the less pathogenic HTLV-II. It is critical to make a differential diagnosis of the two viruses. OBJECTIVES: To design and evaluate synthetic core and envelope-derived peptide enzyme immunoassays (EIA) for serological differential diagnosis. STUDY DESIGN: Peptide EIAs were evaluated with a panel of 202 plasma samples comprised of HTLV antibody positive, serologically classified as confirmed, indeterminate, or non confirmed, characterized as HTLV-I, HTLV-II or neither by genomic amplification. The peptide EIA with the best performance was further used to differentiate between HTLV-I and HTLV-II antibodies in 807 samples from 18 countries in four continents and to provide ratios between the two infections. RESULTS: The gp46 peptide EIA correctly identified 96.5% of HTLV-I and 98.6% of HTLV-II antibody-confirmed samples. HTLV-I was found exclusively in Japan and Caribbean countries; almost exclusively in Africa. HTLV-II represented 10-25% of samples from Canada, Chile and Venezuela and was predominant in the US. CONCLUSIONS: Differential diagnosis between HTLV-I and HTLV-II can be reliably performed using specific peptides from the gp46 envelope protein of each virus.

[Infection with HTLV virus type I-II in patients with cervico-uterine cancer in the Yucatan peninsula, Mexico]. / Infección por virus linfotrópico de células T humanas tipo I/II en pacientes con cáncer cervicouterino de la península de Yucatán, México.

Southwestern Japan is an endemic zone with high prevalence of HTLV-I infection. In addition, a relation between cancer of the cervix and this retrovirus has been described. A recent study has demonstrated a low prevalence of HTLV-I/II infection in Yucatan, Mexico. However, cancer of the cervix is the most frequent oncological disease in this region. The objective of this study was to determine the relationship between cancer of the cervix and the HTLV-I/II infection. Sera from 123 patients with cancer of the cervix and 662 healthy women were screened for antibodies against HTLV-I/II by ELISA and agglutination test (PA). Results were confirmed by Western blot (WB). In the confirmed cases the differentiation between HTLV-I and HTLV-II was made by synthetic peptides. Only one woman (0.8%) had positive results in the patients group and two women (0.3%) had reactivity in the control group. In all these cases the antibodies were positive for HTLV-II. The prevalence in the group of patients with cancer of the cervix was similar to that of the control group. We therefore concluded that in Yucatan, Mexico there is no relation between HTLV-I/II infection and cancer of the cervix.

Infección por virus linfotrópico de células T humanas tipo I/ii en pacientes con cáncer cervicouterino de la península de Yucatán, México / Infection by lymphotropic virus of human T cell type I/II in patients with cervico-uterine cancer at Yucatan, Mexico

En el suroeste de Japón, zona endémica de alta prevalencia para el virus linfotrópico de células T humanas tipo I (HTLV-I), se ha descrito la asociación entre este retrovirus y el cáncer cérvicouterino (CaCU). La Península de Yucatán, México es una zona de baja prevalancia para el HTLV-I y el CaCU ocupa el primer lugar entre las enfermedades oncológicas en esta región. El objetivo del estudio fue definir la posible asociación entre la infección del HTLV.I y el CaCU en un grupo de pacientes en la Península de Yucatán, México. En 123 mujeres con diagnóstico establecido de CaCU y en 662 mujeres sanas, se realizó la detección de anticuerpos contra el HTLV-I/II (AcHTLV-I/II) mediante aglutinación de partículas sensibilizadas y ensayo inmunoenzimático. Los casos positivos fueron confirmados por inmunoelectrotransferencia y la diferenciación entre el HTLV-I y el HTLV-II se hizo a través de péptidos sintéticos. Sólo una paciente (mujer de la etnia maya) (1/23, 0.8 por ciento) y en dos mujeres del grupo testigo (2/662, 0.3 por ciento) se encontró evidencia de Ac-HTLV-I/II. En todos los casos los Ac-HTLV-I/II fueron selectivos para HTLV-II. Los resultados son concordantes con el concepto de que la infección del HTLV-I en la Península de Yucatán no guarda relación con el CaCU

Anti-HTLV-1 antibody positive cutaneous T-cell lymphoma.

BACKGROUND: Adult T-cell leukemia/lymphoma (ATLL) is a neoplasm of the mature helper T-lymphocyte. Human T-lymphotropic virus type 1 (HTLV-1) has been shown to be the cause of this neoplasm. Recently, however, the HTLV-1 genome has been found in some patients with cutaneous T-cell lymphoma (CTCL), which suggests a causal relation of HTLV-1 to CTCL. Thus, the relation between the HTLV-1 genome and CTCL, as well as the difference between ATLL and CTCL, have come into question. METHODS: The authors examined two patients with CTCL whose serum anti-HTLV-1 antibodies were constantly positive. The Southern blot technique, inverse polymerase chain reaction (IPCR), and polymerase chain reaction (PCR) with four sets of primers for gag, pol, env, and pX regions of HTLV-1 were used to clarify the distinctions between ATLL and CTCL. RESULTS: Clinically, one patient presented with multiple subcutaneous nodules with involvements of the internal organ, and the other patient was typical for mycosis fungoides. No integration of HTLV-1 DNA was detected by IPCR or the Southern blot technique in either patient. PCRs with the four sets of primers were all found to be positive for HTLV-1 except one. CONCLUSIONS: The authors conclude that ATLL should be differentiated from CTCL in view of the responsibility of HTLV-1 for promoting or maintaining CTCL.