HTLV-1 in mouthwash cells from a TSP/HAM patient and asymptomatic carriers

Summary.  Using in situ hybridization, the presence of T-cell lymphotrophic virus type I (HTLV-I) was shown in blood lymphocytes of one tropical spastic paraparesis (TSP/HAM) patient and in two asymptomatic carriers. HTLV-I was also detected in epithelial cells derived from mouthwash of the TSP/HAM patient. Mouthwash of one of the carriers showed an infected lymphocyte while mouthwash of the other carrier was negative. The infected epithelial cells stained both in the nucleus and in the cytoplasm, which indicated the presence of the virus in both subcellular compartments. Our observations suggest that saliva cells, lymphocytes and epithelial cells, may potentially participate in oral transmission of HTLV-I. Accepted November 18, 1997 Received August 27, 1997     

Anti-HTLV-1 tax antibody and tax-specific cytotoxic T lymphocyte are associated with a reduction in HTLV-1 proviral load in asymptomatic carriers

Previous studies have suggested that higher anti-human T-lymphotropic virus 1 (HTLV-1) antibody titer and lower anti-HTLV-1 Tax antibody reactivity are risk factors for adult T-cell leukemia/lymphoma. In the present study, we analyzed the relationships between these factors and clarified their significance. Forty-five carriers were examined for anti-HTLV-1 and anti-Tax antibody by ELISA. In addition, 43 of the 45 carriers with HLA-A*0201 and/or A*2402 were examined for frequency of Tax-specific cytotoxic T lymphocytes (CTLs) using HTLV-1/HLA tetramers, and 44 were examined for proviral load by real-time PCR. The relationships between these factors were analyzed statistically. The frequencies of Tax11-19 and Tax301-309-specific CTLs were significantly higher in the anti-Tax antibody-positive group as compared with the antibody-negative group (P = 0.002 and 0.033, respectively). Anti-HTLV-1 antibody titer had a positive correlation with proviral load (P = 0.019), whereas anti-Tax antibody did not show a significant correlation. Higher frequencies of both Tax11-19 and Tax301-309-specific CTLs are related to a reduction in proviral load (P = 0.017 and 0.015, respectively). Synergistic interactions of humoral and cellular immunity against Tax protein were demonstrated in HTLV-1 carriers. Tax-specific CTL may reduce HTLV-1 proviral load to prevent asymptomatic carriers from developing adult T-cell leukemia/lymphoma.     

Human T cell lymphotropic virus type 1 (HTLV-1) proviral load of HTLV-associated myelopathy/tropical spastic paraparesis (HAM/TSP) patients according to new diagnostic criteria of HAM/TSP

A high human T‐cell lymphotropic virus type 1 (HTLV‐1) proviral load is described in HTLV‐1‐associated diseases, especially HAM/TSP. However, the cut‐off value to define high levels of HTLV‐1 proviral load is not well established. 281 HTLV‐1‐infected patients from the HTLV reference center in Salvador, Brazil, were followed from 2005 to 2008. Patients were classified as asymptomatic, possible‐, probable‐, and definite‐HAM/TSP, in accordance with diagnostic criteria proposed by De Castro‐Costa et al. (2006): AIDS Res Hum Retroviruses 22:931–935. HTLV‐1 proviral load was determined using real‐time PCR. A receiver operator characteristic (ROC) curve was constructed using only asymptomatic individuals and definite‐HAM/TSP patients. The ROC curve was used to predict the proviral load level that differentiates these two groups. Out of 281 patients, 189 were asymptomatic and 92 were diagnosed with HAM/TSP (22 possible, 23 probable, 47 definite). The mean HTLV‐1 proviral load was higher in possible‐ (89,104 ± 93,006 copies/10⁶ PBMC), ‐probable (175,854 ± 128,083 copies/10⁶ PBMC), and definite‐HAM/TSP patients (150,667 ± 122,320 copies/10⁶ PBMC), when compared to asymptomatic individuals (27,178 ± 41,155 copies/10⁶ PBMC) (P < 0.0001). A comparison of all HAM/TSP groups showed the highest proviral loads in probable‐HAM/TSP patients, yet the differences in mean values were not statistically significant. The ROC curve suggested a value of 49,865 copies/10⁶ PBMC, with 87% sensitivity (95% CI = 74–95) and 81% specificity (95% CI = 75–86), as the best proviral load cut‐off point to differentiate definite HAM/TSP patients from asymptomatic individuals. HTLV‐1 proviral loads are higher in groups of infected patients with neurological symptoms and may represent a relevant biological marker of disease progression. J. Med. Virol. 83:1269–1274, 2011. © 2011 Wiley‐Liss, Inc.     

In vitro cytoskeleton changes of mouse neurons induced by purified HTLV-1, and PBMC from HAM/TSP patients and HTLV-1 carriers

Summary. HTLV-1 is the causative agent of HAM/TSP. This neurological disease affects the CNS producing damage of the motor tracts at the spinal cord. The HAM/TSP pathogenesis remains undefined. It could include direct and indirect actions of HTLV-1. We studied the effect of purified HTLV-1 and the PBMC of 22 Chilean patients co-cultivated with fetal neurons of mouse (CNh cells): 8 HAM/TSP, 8 HTLV-1 carriers, and 6 non-infected controls. The viral antigens and provirus in CNh cells was evaluated with monoclonal and polyclonal antibodies reacting with HTLV-1 by immunofluorescence assay and PCR at 0, 7 and 15 days of co-cultures, respectively. Viral antigens were detected in 0.1–0.5%, and 0–0.3% of the neurons incubated with lymphocytes of HAM/TSP patients and HTLV-1 carriers, respectively. Neurons incubated with cells of 7 HAM/TSP patients, and 3 HTLV-1 carriers showed the presence of nucleotide sequences of tax gene. These results would be showing that CNh cells would express viral antigens and provirus. The HTLV-1 or their proteins were capable in vitro to produce structural and growth changes in the cytoskeleton of CNh neurons. In this series, the purified HTLV-1 was more effective in the neural changes than PBMC of HAM/TSP or HTLV-1 carriers.     

Detection of proviral human T-cell lymphotrophic virus type I DNA in mouthwash samples of HAM/TSP patients and HTLV-I carriers

Summary Human T-cell lymphotrophic virus type I (HTLV-I), is a member of the oncogenic retroviruses family endemic in several parts of the world and also recently identified in the Jewish Mashhadi population who immigrated from Iran to Israel. The virus is the causative agent of adult T-cell leukemia (ATL) and a chronic myelopathy known both as tropical spastic paraparesis (TSP) or HTLV-I associated myelopathy (HAM). The known modes of HTLV-I transmission are by sexual intercourse, from mother to child in breast milk, via blood transfusion, and by sharing of needles by parenteral drug users. In the present study we examined the presence of HTLV-I provirus genomic DNA by nested polymerase chain reaction (PCR) and by DNA hybridization in mouthwash samples obtained from 13 Mashhadi-born Iranian Jews with spastic paraparesis associated with HTLV-I, 4 Mashhadi-born Iranian Jews asymptomatic carriers for HTLV-I and 21 healthy controls. Proviral HTLV-I DNA was detected by mouthwash PCR in 12 of 17 HTLV-I infected subjects (71%) but in none of 21 controls. Proviral DNA was also detected in mouthwash samples using HTLV-I probe by dot blot hybridization assay. The presence of HTLV-I proviral DNA in whole saliva may suggest a possible transmission of the virus via saliva and explain the increased rate of infection in elderly Mashhadi-Jewish population.     

Oral health profile in patients infected with HTLV-1: clinical findings, proviral load, and molecular analysis from HTLV-1 in saliva

Human T‐lymphotropic virus type 1 (HTLV‐1) is associated with adult T‐cell leukemia (ATL) and HTLV‐1 associated myelopathy/tropical spastic paraparesis (HAM/TSP) and has also been implicated in several disorders, including periodontal disease. The proviral load is an important biological marker for understanding HTLV‐1 pathogenesis and elucidating whether or not the virus is related to the clinical manifestation of the disease. This study describes the oral health profile of HTLV‐1 carriers and HAM/TSP patients in order to investigate the association between the proviral load in saliva and the severity of the periodontal disease and to examine virus intra‐host variations from peripheral blood mononuclear cells and saliva cells. It is a cross‐sectional analytical study of 90 individuals carried out from November 2006 to May 2008. Of the patients, 60 were HTLV‐1 positive and 30 were negative. Individuals from the HTLV‐1 positive and negative groups had similar mean age and social‐economic status. Data were analyzed using two available statistical software packages, STATA 8.0 and SPSS 11.0 to conduct frequency analysis. Differences of P < 0.05 were considered statistically significant. HTLV‐1 patients had poorer oral health status when compared to seronegative individuals. A weak positive correlation between blood and saliva proviral loads was observed. The mean values of proviral load in blood and saliva in patients with HAM/TSP was greater than those in HTLV‐1 carriers. The HTLV‐1 molecular analysis from PBMC and saliva specimens suggests that HTLV‐1 in saliva is due to lymphocyte infiltration from peripheral blood. A direct relationship between the proviral load in saliva and oral manifestations was observed. J. Med. Virol. 84:1428–1436, 2012. © 2012 Wiley Periodicals, Inc.     

Monitoring the HTLV-1 proviral load in the peripheral blood of asymptomatic carriers and patients with HTLV-associated myelopathy/tropical spastic paraparesis from a Brazilian cohort: ROC curve analysis to establish the threshold for risk disease

Human T‐lymphotropic virus 1 (HTLV‐1) infection is associated with HTLV‐associated myelopathy/tropical spastic paraparesis (HAM/TSP), which affects approximately 5% of carriers. High proviral load is a risk marker for HAM/TSP, although there is an overlap of proviral load levels in peripheral blood between asymptomatic carriers and HAM/TSP patients. In this study, receiver operating characteristic curve analysis was used to define a set point of HTLV‐1 proviral load that better indicates an increased risk for HAM/TSP. Proviral load was quantified in 75 asymptomatic carriers and 78 HAM/TSP patients in a Brazilian cohort. The cut‐off of proviral load was defined as 114 copies/10⁴ cells, with 78.2% sensitivity to identify true HAM/TSP patients. The mean proviral load levels were not significantly different between males and females with the same clinical status, and there was no significant correlation between proviral load and age at blood sampling, age at the onset of illness, or duration of disease. In HAM/TSP patients, proviral load was significantly higher in wheelchair‐bound patients than in individuals able to walk without support and in those with the worst spinal cord injuries. Follow‐up of HTLV‐1‐infected individuals showed that proviral load was more stable in asymptomatic carriers than in HAM/TSP patients. In a cohort study, periodically quantifying proviral load in asymptomatic carriers is necessary to identify those at risk for developing neurological disease, and it is necessary for HAM/TSP patients to monitor spinal injury and progression to walking disability. The measure of proviral load in clinical practice implicates the definition of the cut‐off of proviral load and its validation during follow‐up. J. Med. Virol. 84:664–671, 2012. © 2011 Wiley Periodicals, Inc.     

Proviral load and immune markers associated with human T-lymphotropic virus type 1 (HTLV-1)-associated myelopathy/tropical spastic paraparesis (HAM/TSP) in Peru

Human T-lymphotropic virus type 1 (HTLV-1) is the aetiological agent of HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP). The objective of this study is to identify which ex vivo and in vivo markers are associated independently with HAM/TSP in a Peruvian population. Eighty-one subjects (33 men/48 women) were enrolled: 35 presented with HAM/TSP, 33 were asymptomatic HTLV-1 carriers (ACs) and 13 were HTLV-1-seronegative controls (SCs). Ex vivo markers included T cell proliferation and Th1 [interferon (IFN)-gamma], Th2 [interleukin (IL)-4, IL-5], proinflammatory [tumour necrosis factor (TNF)-alpha] and anti-inflammatory (IL-10) cytokine production in non-stimulated peripheral blood mononuclear cell (PBMC) cultures. In vivo CD4(+) T cell count, markers of Th1 [interferon-inducible protein (IP)-10] and Th2 (sCD30) activity in plasma and HTLV-1 proviral load in PBMCs were also evaluated. In univariate analysis, several markers, including T cell proliferation, IFN-gamma, IP-10, sCD30 and proviral load were associated with HAM/TSP, but in a multiple logistic regression analysis only the proviral load remained associated significantly with disease manifestation [adjusted OR 9.10 (1.24-66.91)]. Our findings suggest that HAM/TSP is associated primarily with proviral load, whereas the observed association with some immune markers seems secondary.     

Transient increment of HTLV-2 proviral load in HIV-1-co-infected patients during treatment intensification with raltegravir

BackgroundNumerous studies have analyzed the effects of raltegravir intensification on HIV-1 viral replication in infected individuals receiving suppressive combined antiretroviral treatment (cART). Nevertheless, there are only two studies on the effect of raltegravir in HTLV-1 infection, and none in HTLV-2.ObjectiveTo study the effect of raltegravir on HTLV-2 infection in HIV-1-co-infected individuals.Study designThis retrospective longitudinal study included four HTLV-2-HIV-1-co-infected individuals who received raltegravir-based cART during 48 weeks and 11 HTLV-2-HIV-1-co-infected individuals under cART without raltegravir during 48 weeks. HTLV-2 proviral load, CD4 and CD8 count and frequency were analyzed.ResultsHTLV-2 proviral load significantly increased at week 24 compared to baseline among all the patients who received raltegravir (p = 0.003), while no significant increases were found in the control group. No significant variation in either CD8 or CD4 counts was found during the follow up in both groups.ConclusionsRaltegravir induced a transient increment on total HTLV-2 DNA proviral load in HTLV-2/HIV-1-coinfected individuals on suppressive cART after 24 weeks.     

Keratoconjunctivitis sicca of human T cell lymphotropic virus type 1 (HTLV-1) infected individuals is associated with high levels of HTLV-1 proviral load

BackgroundA high HTLV-1 proviral load is found in HTLV-1-associated diseases, mainly HAM/TSP. However, the association between proviral load and keratoconjunctivitis sicca (KCS) has not been well established.AimTo verify the association between KCS and HTLV-1 proviral load.Study design104 HTLV-1 infected patients (51 asymptomatic and 52 with HAM/TSP) from the HTLV reference center in Salvador, Brazil were followed from June 2008 to May 2010. Evaluation of tear secretion was performed by BUT (break-up time), Rose Bengal and Schirmer I tests. The diagnosis of KCS was based upon the presence of symptoms and when at least two of three tests were positive. HTLV-1 proviral load was determined using real-time PCR.ResultsThe prevalence of KCS was 44.2%. KCS was more frequent among HAM/TSP patients (p = 0.022). Patients with KCS had higher proviral load (mean 134,672 ± 150,393 copies/10⁶ PBMC) than patients without the disease (mean 66,880 ± 109,525 copies/10⁶ PBMC) (p = 0.001). HTLV-1 proviral load > 100,000 copies/10⁶ PBMC increased significantly the risk of developing KCS (OR = 4.05 and 95% CI = 1.40–11.76). After age > 45 years and HAM/TSP status were excluded in stepway reward analysis, the variables PVL > 100,000 (OR = 4.77 and 95% CI = 1.83–12.44) still remained statistically significant.ConclusionHTLV-1 proviral loads are higher in patients with KCS and may represent a relevant biological marker of disease.     

Levels of serum chemokines discriminate clinical myelopathy associated with human T lymphotropic virus type 1 (HTLV-1)/tropical spastic paraparesis (HAM/TSP) disease from HTLV-1 carrier state

Approximately 5% of people infected with human T lymphotropic virus type 1 (HTLV-1) develop clinical myelopathy or tropical spastic paraparesis (HAM/TSP) that is associated with high-levels of Th1 cytokines, interferon (IFN)-gamma and tumour necrosis factor (TNF)-alpha. Chemokines are known to induce cytokine secretion and direct the trafficking of immune cells to sites of disease. The present study measured serum chemokines correlated with autonomously released IFN-gamma in cell cultures. HTLV-1 infection was defined by enzyme-linked immunosorbent assay (ELISA) and confirmed by Western blot. Subjects included HTLV-1 carriers (n = 56), patients with HAM/TSP (n = 31) and healthy HTLV-1 seronegative volunteer controls (n = 20). Serum chemokines and IFN-gamma autonomously released by mononuclear cells in culture were quantified by ELISA. Compared to HTLV-1 carriers, serum chemokines in HAM/TSP patients showed significantly increased levels of CXCL9 and CXCL10, significantly diminished levels of CCL2 and similar amounts of CCL11 and CCL24. In contrast, CCL11 and CCL24 were significantly lower in serum of HAM/TSP patients than either control. IFN-gamma was positively correlated with CXCL9 and CXCL10 when HAM/TSP and HTLV-1 carriers were used as a combined group. However, despite a large proportion of HTLV-1 carriers having high IFN-gamma levels, these chemokines were not increased in carriers. This study showed that high levels of CXCL9 and CXCL10 in the systemic circulation and low serum CCL2 levels are features of HAM/TSP. HTLV-1 infection and Tax and/or additional viral encoded factor-mediated pathological processes triggering T cell activation with autogenous IFN-gamma release are probably involved in regulating chemokine release.     

VEGF levels in asthmatic airways do not correlate with plasma extravasation

BACKGROUND: Vascular permeability/vascular endothelial growth factor (VEGF) is a multifunctional cytokine which plays a role in chronic inflammation and angiogenesis. Its expression in bronchoalveolar lavage (BAL) has not been determined although VEGF may be relevant to the pathophysiology of asthma in which oedema is an important feature. METHODS: We studied VEGF, albumin and IgA immunoreactive levels in the BAL fluids obtained from 27 chronic stable asthmatics, nine untreated chronic bronchitis patients and 15 control subjects. RESULTS: BAL fluid levels of VEGF and VEGF normalized to IgA were not significantly different in any patient group. Both asthmatic steroid- and non-steroid-treated groups had significantly lower albumin levels in their BAL fluids explaining most of the 179% increased VEGF normalized to albumin ratios in non-steroid treated asthmatics. Moreover, VEGF and albumin markers correlated in control subjects (r = 0.73, P = 0.006) and in chronic bronchitics (r = 0.75, P = 0.03, Spearman test), but not in asthmatics. VEGF was inversely correlated with asthma severity (GINA/NHLBI scores) in non-steroid treated asthmatics (tau = - 0.52, P = 0.009, Kendall test). CONCLUSIONS: Thus, the potential role of VEGF in asthma requires further studies on bronchial biopsies and induced sputum.     

Intrathecal IgG synthesis and specificity of oligoclonal IgG in patients infected with HIV-1 do not correlate with CNS disease.

The CSF/serum immune response to HIV 1 was studied in 24 patients admitted for investigation. The level of antibody to HIV-1 and specificity of oligoclonal IgG were determined in blood and cerebrospinal fluid (CSF). The majority of patients demonstrated elevated levels of intrathecal IgG synthesis, with levels of HIV-1-specific antibody frequently being significantly higher in CSF than in serum. In 16 of 21 patients the CSF/serum antibody ratio indicated active intrathecal synthesis. Oligoclonal banding was present in CSF from all 24 patients. Immunoprinting of serum and CSF demonstrated antigenic specificity (p24, gp 160, RT) of the clonal antibodies in all of 12 patients though the patterns of reactivity in CSF did not necessarily correspond with that of serum. Although a specific association of particular patterns with HIV CNS disease was not found we feel that these markers should be included in longitudinal studies of HIV-related diseases of the CNS. The specificity of oligoclonal antibodies, both in CSF and in serum was demonstrated, and this specificity may be a useful marker for longitudinal studies in HIV-1 antibody-positive asymptomatic patients.     

High pneumococcal DNA load,procalcitonin and suPAR levels correlate to severe disease development in patients with pneumococcal pneumonia

Community-acquired pneumonia (CAP) is mostly caused by Streptococcus pneumoniae. Identification of the pathogen causing CAP can be achieved by conventional culture techniques of sputum and/or blood, antigen detection from urine or molecular analysis. However, it remains difficult to determine patients who are at risk of severe disease development (intensive care unit [ICU] admittance and/or death). In this retrospective study, 121 patients admitted to the emergency department with pneumonia symptoms were included. Several markers of infection (pneumococcal DNA load in blood (real-time LytA PCR), white blood cell (WBC) count, C-reactive protein (CRP), procalcitonin (PCT) and soluble urokinase plasminogen activator receptor (suPAR) levels) were assessed for their ability to predict severe disease development. Of 121 patients, 6 were excluded from the study because of an alternative diagnosis, whereas 8 were excluded from biomarker analysis because of the presence of co-morbidities. Of the 115 patients analysed by the LytA PCR, 23 were positive. PCR detected S. pneumoniae DNA in 82% of patients with positive blood culture for S. pneumoniae. PCR missed three samples from patients in which S. pneumoniae was recovered by blood cultures. However, eight additional LytA PCR-positive samples were detected from patients whose blood cultures remained negative. Pneumococcal DNA load was also monitored in time for 31 patients, of whom 11 had positive PCR results. For 10 out of 11 (91%) positive PCR patients, a clear increase in Ct-values was observed, indicating a lower pneumococcal DNA load in the blood as a result of antibiotic therapy. Biomarker analysis was performed in 107 patients, of whom 29 showed severe disease development. Pneumococcal DNA load (p = 0.026), PCT (p = 0.046) and suPAR (p = 0.001) levels most reliably predicted severe disease development. In conclusion, in patients with CAP, higher pneumococcal DNA load, PCT and suPAR values are associated with severe disease development (ICU admission and/or death). These biomarkers may be useful tools for triage of patients suspected of having CAP in the emergency department.