Proviral Load In Peripheral Blood Research Articles

Role of the CTCF Binding Site in Human T-Cell Leukemia Virus-1 Pathogenesis.

During HTLV-1 infection, the virus integrates into the host cell genome as a provirus with a single CCCTC binding protein (CTCF) binding site (vCTCF-BS), which acts as an insulator between transcriptionally active and inactive regions. Previous studies have shown that the vCTCF-BS is important for maintenance of chromatin structure, regulation of viral expression, and DNA and histone methylation. Here, we show that the vCTCF-BS also regulates viral infection and pathogenesis in vivo in a humanized (Hu) mouse model of adult T-cell leukemia/lymphoma. Three cell lines were used to initiate infection of the Hu-mice, i) HTLV-1-WT which carries an intact HTLV-1 provirus genome, ii) HTLV-1-CTCF, which contains a provirus with a mutated vCTCF-BS which abolishes CTCF binding, and a stop codon immediate upstream of the mutated vCTCF-BS which deletes the last 23 amino acids of p12, and iii) HTLV-1-p12stop that contains the intact vCTCF-BS, but retains the same stop codon in p12 as in the HTLV-1-CTCF cell line. Hu-mice were infected with mitomycin treated or irradiated HTLV-1 producing cell lines. There was a delay in pathogenicity when Hu-mice were infected with the CTCF virus compared to mice infected with either p12 stop or WT virus. Proviral load (PVL), spleen weights, and CD4 T cell counts were significantly lower in HTLV-1-CTCF infected mice compared to HTLV-1-p12stop infected mice. Furthermore, we found a direct correlation between the PVL in peripheral blood and death of HTLV-1-CTCF infected mice. In cell lines, we found that the vCTCF-BS regulates Tax expression in a time-dependent manner. The scRNAseq analysis of splenocytes from infected mice suggests that the vCTCF-BS plays an important role in activation and expansion of T lymphocytes in vivo. Overall, these findings indicate that the vCTCF-BS regulates Tax expression, proviral load, and HTLV pathogenicity in vivo.

HTLV-1 Proviral Load in Vaginal Fluid Correlates with Levels in Peripheral Blood Mononuclear Cells.

The prevalence of human T-lymphotropic virus type-1 (HTLV-1) infection is higher in women, and sexual intercourse has been described as an important route of male-to-female transmission. The present study aimed to quantify HTLV-1 proviral load (PVL) in vaginal fluid, and to investigate correlations with PVL in peripheral blood mononuclear cells (PBMCs). In addition, cytopathological alterations and vaginal microbiota were evaluated. HTLV-1-infected women were consecutively recruited at a multidisciplinary center for HTLV patients in Salvador, Brazil. All women underwent gynecological examinations to obtain cervicovaginal fluid and venipuncture for blood collection. PVL, as measured by real-time quantitative polymerase chain reaction (RT-qPCR), was expressed as the number of copies of HTLV-1/106 cells in blood and vaginal fluid samples. Light microscopy was used to assess cervicovaginal cytopathology and vaginal microbiota. In the 56 included women (43 asymptomatic carriers and 13 diagnosed with HTLV-1-associated myelopathy/tropical spastic paraparesis-HAM/TSP), mean age was 35.9 (SD ± 7.2) years. PVL was higher in PBMCs (median: 23,264 copies/106 cells; IQR: 6776-60,036) than in vaginal fluid (451.9 copies/106 cells; IQR: 0-2490) (p < 0.0001). PVL in PBMCs was observed to correlate directly with PVL in vaginal fluid (R = 0.37, p = 0.006). PVL was detected in the vaginal fluid of 24 of 43 (55.8%) asymptomatic women compared to 12 of 13 (92.3%) HAM/TSP patients, p = 0.02. Cytopathologic analyses revealed no differences between women with detectable or undetectable PVL. HTLV-1 proviral load is detectable in vaginal fluid and correlates directly with proviral load in peripheral blood. This finding suggests that sexual transmission of HTLV-1 from females to males may occur, as well as vertical transmission, particularly in the context of vaginal delivery.

Long-term safety and efficacy of mogamulizumab (anti-CCR4) for treating virus-associated myelopathy.

Some carriers of human T-cell leukemia virus type 1 (HTLV-1), a retrovirus that primarily infects CD4+ T cells and causes lifelong infection, develop HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP). Current treatments for HAM/TSP are insufficient with problematic long-term side effects. This study evaluated the long-term safety and efficacy of the anti-CCR4 antibody mogamulizumab in patients with HAM/TSP over a 4-year period. We conducted an open-label, extended long-term study (UMIN trial number: UMIN000019942) of a Phase 1-2a trial with mogamulizumab for HAM/TSP (UMIN000012655). The study participants were patients with corticosteroid-resistant HAM/TSP who could walk 10 m with or without assistive tools. Mogamulizumab was administered at 0.01, 0.03, 0.1, or 0.3 mg/kg at intervals of ≥8 weeks (0.01 and 0.03 mg/kg) or ≥12 weeks (0.1 and 0.3 mg/kg). HTLV-1 proviral load, cerebrospinal fluid inflammatory markers, and clinical symptoms were summarized by descriptive statistics. Missing observations were imputed using the last-observation-carried-forward method. As a post-hoc analysis, we evaluated the therapeutic effect of mogamulizumab on gait function by comparing it with contemporary control data from a HAM/TSP patient registry. Of the 21 participants in the Phase 1-2a, 18 (86%) enrolled in the long-term study and 15 (71%) continued repeated doses of mogamulizumab for 4 years. The median dose was 0.1 mg/kg after 4 years. Seventeen of 21 participants (81%) experienced grade 1-2 skin-related adverse events. Observed grade 3 drug-related adverse effects included three cases of lymphopenia and one case each of microscopic polyangiitis, elevated levels of aspartate aminotransferase, and neutropenia. Four of 21 participants (19%) developed neutralizing antibodies. After 4 years, the peripheral blood proviral load and the number of infected cells in CSF decreased by 60.7% and 66.3%, respectively. Neopterin and CXCL10 CSF concentrations decreased by 37.0% and 31.0%, respectively. Among the 18 participants, spasticity and Osame Motor Disability Score (OMDS) improved in 17 (94%) and 4 (22%), respectively. However, 10 m walking time worsened by 7.3% on average. Comparison with the contemporary control group demonstrated that mogamulizumab inhibited OMDS progression (p = 0.02). The results of the study suggest that mogamulizumab has long-term safety and inhibitory effect on lower limb motor disability progression in corticosteroid-treated patients with HAM/TSP. This will provide a basis for the application of mogamulizumab in HAM/TSP treatment.

Expression-based analysis of genes related to single nucleotide polymorphism hits associated with bovine leukemia virus proviral load in Argentinean dairy cattle

In dairy cattle infected with bovine leukemia virus (BLV), the proviral load (PVL) level is directly related to the viral transmission from infected animals to their healthy herdmates. Two contrasting phenotypic groups can be identified when assessing PVL in peripheral blood of infected cows. A large number of reports point to bovine genetic variants (single nucleotide polymorphisms) as one of the key determinants underlying PVL level. However, biological mechanisms driving BLV PVL profiles and infection progression in cattle have not yet been elucidated. In this study, we evaluated whether a set of candidate genes affecting BLV PVL level according to whole genome association studies are differentially expressed in peripheral blood mononuclear cells derived from phenotypically contrasting groups of BLV-infected cows. During a 10-mo-long sampling scheme, 129 Holstein cows were phenotyped measuring anti-BLV antibody levels, PVL quantification, and white blood cell subpopulation counts. Finally, the expression of 8 genes (BOLA-DRB3, PRRC2A, ABT1, TNF, BAG6, BOLA-A, LY6G5B, and IER3) located within the bovine major histocompatibility complex region harboring whole genome association SNP hits was evaluated in 2 phenotypic groups: high PVL (n = 7) and low PVL (n = 8). The log2 initial fluorescence value (N0) transformed mean expression values for the ABT1 transcription factor were statistically different in high- and low-PVL groups, showing a higher expression of the ABT1 gene in low-PVL cows. The PRRC2A and IER3 genes had a significant positive (correlation coefficient = 0.61) and negative (correlation coefficient = -0.45) correlation with the lymphocyte counts, respectively. Additionally, the relationships between gene expression values and lymphocyte counts were modeled using linear regressions. Lymphocyte levels in infected cows were better explained (coefficient of determination = 0.56) when fitted a multiple linear regression model using both PRRC2A and IER3 expression values as independent variables. The present study showed evidence of differential gene expression between contrasting BLV infection phenotypes. These genes have not been previously related to BLV pathobiology. This valuable information represents a step forward in understanding the BLV biology and the immune response of naturally infected cows under a commercial milk production system. Efforts to elucidate biological mechanisms leading to BLV infection progression in cows are valuable for BLV control programs. Further studies integrating genotypic data, global transcriptome analysis, and BLV progression phenotypes are needed to better understand the BLV-host interaction.

Simultaneous evaluation of diagnostic marker utility for enzootic bovine leukosis

BackgroundEnzootic bovine leukosis (EBL) is a disease of cattle caused by bovine leukemia virus (BLV). More than 60% of BLV-infected cattle remain subclinical and are thus referred to as aleukemic (AL) cattle. Approximately 30% of infected cattle show a relatively stable increase in the number of B lymphocytes; these cattle are termed persistent lymphocytosis (PL) cattle. A small percentage of infected cattle develop BLV-induced B cell lymphoma (EBL) and are called EBL cattle. Due to the increase in the number of BLV-infected cattle, the number of EBL cattle has featured a corresponding increase over recent years in Japan. Several diagnostic criteria for EBL (e.g., enlarged superficial lymph nodes, protrusion of the eye, increased peripheral blood lymphocyte, etc.) are used for on-farm diagnosis and antemortem tests at slaughterhouses. Since the slaughter of EBL cattle for human consumption is not allowed, on-farm detection of EBL cattle is important for reducing the economic loss incurred by farms. Therefore, establishing new diagnostic markers to improve the efficiency and accuracy of the antemortem detection of EBL cattle is a critical, unmet need. To simultaneously evaluate the utility of candidate markers, this study measured the values of each marker using the blood samples of 687 cattle with various clinical statuses of BLV infection (EBL, PL, AL and non-infected cattle).ResultsSensitivity (Se) and specificity (Sp) were highest for the serum thymidine kinase (TK) followed by the serum lactate dehydrogenase (LDH) isozyme 2. The number of peripheral blood lymphocytes and proviral load in peripheral blood had the lowest Se and Sp. The values of all markers other than TK were influenced by the sex of the tested cattle.ConclusionsAlthough tLDH and its isozymes (LDHs) may be influenced by the sex of the tested cattle, the high accuracy of TK and LDH2 as well as accessibility and simplicity of the protocol used to measure these enzymes recommend the utility of TK and LDHs for EBL cattle detection. Using these markers for screening followed by the application of existing diagnostic criteria may improve the efficiency and accuracy of EBL cattle detection on farms, thereby contributing to the reduction of economic losses in farms.

Evaluation of the cervicovaginal environment in asymptomatic Human T-cell lymphotropic virus type 1 infected women

IntroductionHuman T-cell lymphotropic virus type 1 (HTLV-1) is sexually transmitted and causes persistent infection. This virus induces activation of the immune system and production of inflammatory cytokines. This study aimed to assess the cytokine profile and cytopathological findings in the cervicovaginal fluid of asymptomatic HTLV-1-infected women. MethodsHTLV-1-infected and uninfected women were selected at the Centro de Atendimento ao Portador de HTLV in Salvador-Brazil. None of the included HTLV-1-infected women reported any HTLV-1-associated diseases. All volunteers underwent gynecological examination to collect cervicovaginal fluid. Cytokine quantification was performed using the Cytometric Bead Array (CBA) Human Th1/Th2/Th17 kit. Light microscopy was used to evaluate cervicovaginal cytopathology. In addition, proviral load in cervicovaginal fluid and peripheral blood was measured by real-time quantitative polymerase chain reaction. Results112 women (63 HTLV-1-infected and 49 uninfected) were evaluated. No differences were found with respect to cytopathological cervicovaginal findings between the groups. IL-2, TNF, IL-4, IL-10, and IL-17 levels were significantly higher in cervicovaginal fluid of the HTLV-1-infected women than in uninfected women (p<0.05). Conversely, IFN-γ was found to be lower in the HTLV-1-infected women (p<0.001) compared to uninfected individuals. Cervicovaginal proviral load was detectable in 53% of the HTLV-1-infected women and was found to be consistently lower than the proviral load in peripheral blood. ConclusionsHTLV-1 infection induces immune activation in cervicovaginal environment, characterized by elevated concentrations of Th1, Th2, and IL17 in the cervicovaginal fluid.

The effectiveness of colostral antibodies for preventing bovine leukemia virus (BLV) infection in vitro

BackgroundBovine leukemia virus (BLV) is the causative agent of enzootic bovine leukosis (EBL). The incidence of EBL in Japan is increasing annually; and the cases of EBL in cattle younger than 2 years old has been reported. Therefore, it is vital to find a method to control BLV infection, especially in young calves. In this study, to evaluate the protective ability of colostral antibodies against BLV infection, as well as the potential for BLV infection mediated by colostrum/milk, we investigated temporal fluctuations in the anti-BLV antibody titer and BLV proviral load (PVL) in colostrum/milk and peripheral blood of six infected dams during lactation. The association between PVL and antibody titer in colostrum and peripheral blood was then investigated using samples from a further twenty-seven cattle. Antibody concentrations were measured with a Syncytium-induction Inhibition Assay using colostral/milk whey and serum. PVL in peripheral blood and colostrum was measured by real-time PCR.ResultsColostral antibodies showed high inhibitory activity until day 3 of lactation. The antibody titer and PVL in peripheral blood showed lesser changes than those in colostrum/milk throughout lactation. The colostral antibody titer was significantly higher than the serum antibody titer in all samples, whereas the colostrum PVL was significantly lower than the blood PVL. The blood PVL showed a significant correlation with serum antibody titer, colostrum PVL, and colostral antibody titer. However, there were no major correlations between the serum and colostral antibody titers.ConclusionsThis is the first report investigating the temporal changes in colostral antibody titer in terms of inhibiting BLV infection in vitro. The results of antibody detection by Syncytium-induction Inhibition Assay suggested that the protective activity of the colostral antibodies against BLV infection would be conferred by anti-BLV gp51 antibody. The high antibody titer of colostral whey suggests that colostral whey could be a potential source of antibodies with a low risk of infection in neonatal calves.

BOLA-DRB3 gene polymorphisms influence bovine leukaemia virus infection levels in Holstein and Holstein×Jersey crossbreed dairy cattle.

Bovine leukemia virus (BLV) infections, causing persistent lymphocytosis and lethal lymphosarcoma in cattle, have reached high endemicity on dairy farms. We observed extensive inter-individual variation in the level of infection (LI) by assessing differences in proviral load in peripheral blood. This phenotypic variation appears to be determined by host genetics variants, especially those located in the BoLA-DRB3 MHCII molecule. We performed an association study using sequencing-based typed BOLA-DRB3 alleles from over 800 Holstein and Holstein×Jersey cows considering LI invivo and accounting for filial relationships. The DBR3*0902 allele was associated with a low level of infection (LLI) (<1% of circulating infected B-cells), whereas the DRB3*1001 and DRB3*1201 alleles were related to a high level of infection (HLI). We found evidence that 13 polymorphic positions located in the pockets of the peptide-binding cleft of the BOLA-DRB3 alleles were associated with LI. DRB3*0902 had unique haplotypes for each of the pockets: Ser13 -Glu70 -Arg71 -Glu74 (pocket 4), Ser11 -Ser30 (pocket 6), Glu28 -Trp61 -Arg71 (pocket 7) and Asn37 -Asp57 (pocket 9), and all of them were significantly associated with LLI. Conversely, Lys13 -Arg70 -Ala71 -Ala74 and Ser13 -Arg70 -Ala71 -Ala74 , corresponding to the DRB3*1001 and *1201 alleles respectively, were associated with HLI. We showed that the specific amino acid pattern in the DRB3*0902 peptide-binding cleft may be related to the set point of a very low proviral load level in adult cows. Moreover, we identified two BOLA-DRB3 alleles associated with a HLI, which is compatible with a highly contagious profile.

Identification of cattle carrying alleles associated with resistance and susceptibility to the Bovine Leukemia Virus progression by real-time PCR

Previous studies have shown a significant association between polymorphisms of the BoLA DRB3 gene and Bovine Leukemia Virus (BLV) infection profile. The presence of allele ∗1501 has been associated with high proviral load in peripheral blood while allele ∗0902 has been associated with low proviral load. The purpose of this study was to develop allele-specific real-time PCRs to identify cattle carrying alleles associated with resistance (BoLA DRB3∗0902) or susceptibility (BoLA DRB3∗1501) to the BLV progression. Specific primers were designed and differential amplification was carried out by real-time PCR and monitored by SYBR® Green dye in DNA samples from peripheral blood. Conditions were also adjusted for traditional PCR amplification (end point amplification). These methods are rapid, simple and suitable for high throughput screening, and could aid in marker-assisted selection of BLV-resistant and susceptible cattle.

Maternal Proviral Load and Vertical Transmission of Human T Cell Lymphotropic Virus Type 1 in Guinea-Bissau

The relative importance of routes of transmission of human T cell lymphotropic virus type 1 (HTLV-1) in Guinea-Bissau is largely unknown; vertical transmission is thought to be important, but there are very few existing data. We aimed to examine factors associated with transmission in mothers and children in Guinea-Bissau, where HTLV-1 is endemic (prevalence of 5% in the adult population). A cross-sectional survey was performed among mothers and their children (aged <15 years) in a rural community in Guinea-Bissau. A questionnaire to identify risk factors for infection and a blood sample were obtained. HTLV-1 proviral load in peripheral blood was determined and PCR was performed to compare long terminal repeat (LTR) sequences in mother-child pairs. Fourteen out of 55 children (25%) of 31 HTLV-1-infected mothers were infected versus none of 70 children of 30 uninfected mothers. The only factor significantly associated with HTLV-1 infection in the child was the proviral load of the mother; the risk of infection increased significantly with the log(10) proviral load in the mother's peripheral blood (OR 5.5, 95% CI 2.1-14.6, per quartile), adjusted for weaning age and maternal income. HTLV-1 sequences of the LTR region obtained from mother-child pairs were identical within pairs but differed between the pairs. Vertical transmission plays an important role in HTLV-1 transmission in this community in Guinea-Bissau. The risk of transmission increases with the mother's proviral load in the peripheral blood. Identical sequences in mother-child pairs give additional support to the maternal source of the children's infection.

Bovine leukemia virus proviral load in naturally-infected animals

Due to the high prevalence of Bovine Leukemia Virus (BLV) in argentinean dairy herds, classical control based on the individual level of infection do not constitute a feasible strategy. An approach based on selective segregation of animals according to their peripheral-blood proviral load (PVL) could be a practical alternative. We conducted this work to get information about the frequency distribution of PVL under natural conditions with the purpose to describe the proportion of animals that could be selected as potential low transmitters. We analyzed the PVL on 332 cattle from four infected farms with different levels of seroprevalence (Farm A: 30%, Farm B: 66%, Farm C: 87% and Farm D: 92%) together with the correlation of PVL with BLV antibody levels. In all farms, more than 50% of the seropositive animals showed a proviral load lower than our internal calibrator sample which corresponds to 1 infected cell in 100 non-infected cells according to: Farm A: 78%, Farm B: 79%, Farm C: 55% and Farm D: 79%. The analysis of antibodies showed that low p24-ELISA reactivity is associated with low PVL in the vast majority of animals. The present study provides biological data that could contribute with the design of a management program that includes a selective segregation policy for BLV control based on the level of proviral load, which represents a measure of integrated viral genome in host cells and a surrogate marker of the level of BLV transmission under natural conditions.

Maternal proviral load and vertical transmission of Human T cell Lymphotropic Virus type 1 in Guinea-Bissau

Abstract The relative importance of routes of transmission of human T cell lymphotropic virus type 1 (HTLV-1) in Guinea-Bissau is largely unknown; vertical transmission is thought to be important, but there are very few existing data. We aimed to examine factors associated with transmission in mothers and children in Guinea-Bissau, where HTLV-1 is endemic (prevalence of 5% in the adult population). A cross-sectional survey was performed among mothers and their children (aged <15 years) in a rural community in Guinea-Bissau. A questionnaire to identify risk factors for infection and a blood sample were obtained. HTLV-1 proviral load in peripheral blood was determined and PCR was performed to compare long terminal repeat (LTR) sequences in mother–child pairs. Fourteen out of 55 children (25%) of 31 HTLV-1-infected mothers were infected versus none of 70 children of 30 uninfected mothers. The only factor significantly associated with HTLV-1 infection in the child was the proviral load of the mother; the risk...

Long-Term Outcome of ATL Patients Who Underwent Reduced-Intensity Stem Cell Transplantation (RIST): Suggested Potent Graft-Versus-ATL and HTLV-1 Effects.

Long-Term Outcome of ATL Patients Who Underwent Reduced-Intensity Stem Cell Transplantation (RIST): Suggested Potent Graft-Versus-ATL and HTLV-1 Effects.

Quantitation of feline leukaemia virus viral and proviral loads by TaqMan ® real-time polymerase chain reaction

Feline leukaemia virus (FeLV) infection in cats is not only of veterinary importance but also a well-acknowledged animal model for studying the pathogenesis of retroviral disease. After virus exposure, different courses and outcomes of FeLV infection may prevail; they have been associated with cellular and humoral immune responses and the FeLV proviral load in peripheral blood. We hypothesized that the plasma viral RNA load might be an additional relevant indicator for the infection outcome. To quantify these loads, a real-time reverse transcriptase (RT) polymerase chain reaction (PCR) assay was developed. The assay amplifies FeLV-A, -B, and -C as some naturally infected cats could not be identified with a FeLV-A-based assay previously. The assay was applied to determine plasma FeLV RNA loads in cats infected both naturally and experimentally with FeLV. In addition, an improved real-time PCR assay for quantitation of FeLV proviral loads is described. The assays developed were more sensitive than ELISA and virus isolation in the early phase of infection. In addition, PCR allows the identification of provirus carriers that have overcome antigenaemia. Thus, for most effective detection of FeLV exposure and characterization of the infection in a cat, PCR assays are recommended as diagnostic tools.

Phenotypic Analysis of Peripheral Blood γδ T Lymphocytes and Their Targeting by Human Immunodeficiency Virus Type 1 in Vivo

There is increasing evidence that a wider range of lymphoid cell types other than CD4+ T helper lymphocytes are infected with HIV-1 in vivo, including CD8 lymphocytes, natural killer cells, and reticulodendritic cells. Each potentially contributes to the reservoir of infected cells that resist antiviral treatment and to the impairment of immune responses in AIDS. By quantitative PCR for HIV proviral sequences we have now obtained evidence for substantial infection of γδ lymphocytes, contributing 3–45% of the proviral load in peripheral blood. A large proportion of γδ lymphocytes constitutively expressed the chemokine receptors CCR5 and CXCR4, with evidence for marked up-regulation of CD8 in samples from HIV-infected individuals, corresponding to an activated phenotype. That γδ lymphocytes might be susceptible to HIV infection was investigated using in vitro infectivity assays of recombinant HIV-expressing green fluorescent protein, followed by flow cytometry. γδ, CD4, and CD8 lymphocytes were each productively infected, with γδ lymphocytes showing the greatest susceptibility. For each cell type, blocking assays with an anti-CD4 monoclonal antibody indicated that entry was CD4-dependent.

HTLV-I specific IFN-γ+ CD8+ lymphocytes correlate with the proviral load in peripheral blood of infected individuals

Human T lymphotropic virus type I (HTLV-I)-associated myelopathy/tropical spastic paraparesis (HAM/TSP) is an inflammatory neurological disease caused by HTLV-I infection. It has been shown that HAM/TSP patients have high proviral loads and an extraordinarily high frequency of circulating CD8+ cytotoxic T lymphocytes specific for HTLV-I in their peripheral blood when compared to asymptomatic HTLV-I carriers (AC). We have previously described an intracellular cytokine detection assay, in which interferon-γ (IFN-γ)+ CD8+ lymphocytes are specific for HTLV-I in infected individuals. Here, we have established a competitive polymerase chain reaction assay to measure the proviral load of patients and investigate a potential relationship between proviral load and virus-specific CD8+ lymphocytes. Genomic DNA was extracted from peripheral blood lymphocytes (PBL) from eight HAM/TSP patients and seven AC for the measurement of HTLV-I measuring proviral loads. The same PBL were analyzed for intracellular IFN-γ expression by flow cytometry. In the HAM/TSP patients and AC, the average proviral loads were 34,482 and 9784 copy/μg DNA ( P=0.021), and the average of IFN-γ+ CD8+ lymphocytes in total PBL were 1.47 and 0.08% ( P=0.001), respectively. It was confirmed that HAM/TSP patients have both high proviral loads and increased HTLV-I-specific CD8+ lymphocytes. Furthermore, we found a positive correlation between both factors in the patients with HAM/TSP ( P=0.044) but not in the AC ( P=0.508). These findings suggest that the high number of HTLV-I-specific lymphocytes may result from the increased proviral load in HAM/TSP patients.

Peripheral blood mononuclear cell human immunodeficiency virus type 1 proviral DNA quantification by polymerase chain reaction: relationship to immunodeficiency and drug effect

Human immunodeficiency virus type 1 (HIV-1) proviral DNA from peripheral blood mononuclear cells (PBMCs) was quantitated in 61 HIV-1-seropositive individuals by a nonisotopic polymerase chain reaction assay. Primers from the gag region (SK38, SK39) were used to determine the log10 HIV-1 proviral copy number per 10(6) CD4+ T lymphocytes (peripheral blood proviral load). A standard curve was generated for each assay by using ACH-2 cell DNA. The peripheral blood proviral load was followed in 15 individuals in a longitudinal study and was measured in 45 individuals in a cross-sectional analysis. Three of four untreated patients who were followed for 14 months had stable PBMC proviral loads and CD4+ T lymphocyte counts; one untreated patient had a sustained increase in PBMC proviral load followed 5 months later by a significant decline in the CD4+ T lymphocyte count. Eleven previously untreated individuals were monitored for 1 year following initiation of zidovudine and/or 2',3'-dideoxyinosine therapy. The mean log10 number of proviral HIV-1 copies per 10(6) CD4+ T cells decreased from 4.3 +/- 0.4 at the baseline to 3.5 +/- 0.6 after 2 to 4 months of therapy (P < 0.01). This initial 0.8 log10 fall in the PBMC proviral load after the initiation of therapy was followed by a rise in the PBMC proviral load by the sixth month of therapy. The PBMC proviral load in 45 subjects, both treated (n = 25) and untreated (n = 20), correlated inversely with the CD4+ T lymphocyte count (P < 0.01, R = 0.49). PBMC proviral DNA quantification by a nonisotopic polymerase chain reaction assay correlates with HIV-1 disease progression and could be used to monitor the effect of antiretroviral therapy.