Comprehensive Antiretroviral Restriction Factor Profiling Reveals the Evolutionary Imprint of the ex Vivo and in Vivo IFN-ß Response in HTLV-1-Associated Neuroinflammation.

HTLV-1-Associated Myelopathy (HAM/TSP) is a progressive neuroinflammatory disorder for which no disease-modifying treatment exists. Modest clinical benefit from type I interferons (IFN-α/ß) in HAM/TSP contrasts with its recently identified IFN-inducible gene signature. In addition, IFN-α treatment in vivo decreases proviral load and immune activation in HAM/TSP, whereas IFN-ß therapy decreases tax mRNA and lymphoproliferation. We hypothesize this "IFN paradox" in HAM/TSP might be explained by both cell type- and gene-specific effects of type I IFN in HTLV-1-associated pathogenesis. Therefore, we analyzed ex vivo transcriptomes of CD4+ T cells, PBMCs and whole blood in healthy controls, HTLV-1-infected individuals, and HAM/TSP patients. First, we used a targeted approach, simultaneously quantifying HTLV-1 mRNA (HBZ, Tax), proviral load and 42 host genes with known antiretroviral (anti-HIV) activity in purified CD4+ T cells. This revealed two major clusters ("antiviral/protective" vs. "proviral/deleterious"), as evidenced by significant negative (TRIM5/TRIM22/BST2) vs. positive correlation (ISG15/PAF1/CDKN1A) with HTLV-1 viral markers and clinical status. Surprisingly, we found a significant inversion of antiretroviral activity of host restriction factors, as evidenced by opposite correlation to in vivo HIV-1 vs. HTLV-1 RNA levels. The anti-HTLV-1 effect of antiviral cluster genes was significantly correlated to their adaptive chimp/human evolution score, for both Tax mRNA and PVL. Six genes of the proposed antiviral cluster underwent lentivirus-driven purifying selection during primate evolution (TRIM5/TRIM22/BST2/APOBEC3F-G-H), underscoring the cross-retroviral evolutionary imprint. Secondly, we examined the genome-wide type I IFN response in HAM/TSP patients, following short-term ex vivo culture of PBMCs with either IFN-α or IFN-ß. Microarray analysis evidenced 12 antiretroviral genes (including TRIM5α/TRIM22/BST2) were significantly up-regulated by IFN-ß, but not IFN-α, in HAM/TSP. This was paralleled by a significant decrease in lymphoproliferation by IFN-ß, but not IFN-α treatment. Finally, using published ex vivo whole blood transcriptomic data of independent cohorts, we validated the significant positive correlation between TRIM5, TRIM22, and BST2 in HTLV-1-infected individuals and HAM/TSP patients, which was independent of the HAM/TSP disease signature. In conclusion, our results provide ex vivo mechanistic evidence for the observed immunovirological effect of in vivo IFN-ß treatment in HAM/TSP, reconcile an apparent IFN paradox in HTLV-1 research and identify biomarkers/targets for a precision medicine approach.

Changed epidemiology during intra and interhospital spread of high-risk clones of vanA-containing Enterococcus in Brazilian hospitals.

We report changes in the molecular epidemiology of vanA-containing Enterococcus during the intra and interhospital spread of high-risk clones, in Southeastern Brazil. While VRE faecalis predominated during 1998 to 2006, a reversal has been observed in the last years, where VRE faecium belonging to ST114, ST203, ST412, ST478 and ST858 have become endemic.

MAIT cells are reduced in frequency and functionally impaired in human T lymphotropic virus type 1 infection: Potential clinical implications.

HTLV-1 infection is associated with several inflammatory disorders, including the neurodegenerative condition HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP). It is unclear why a minority of infected subjects develop HAM/TSP. The cellular immune response has been implicated in the development of inflammatory alterations in these patients; however the pathogenic mechanisms for disease progression remain unclear. Furthermore, HTLV-1-infected individuals have an increase incidence of Mycobacterium tuberculosis (Mtb) infection, suggesting that immunological defect are associated with HTLV-1 infection. Evidence suggests an important role for Mucosal-associated invariant T (MAIT) cells in the early control of Mtb infection. Chronic viral infections like HIV and HCV have been associated with decreased frequency and functionality of MAIT cells. We hypothesized that HTLV-1 infection is associated with similar perturbations in MAIT cells. We investigated MAIT cell frequency, phenotype, and function by flow cytometry in a cohort of 10 asymptomatic and 10 HAM/TSP HTLV-1 infected patients. We found that MAIT cells from HTLV-1-infected subjects were reduced and showed high co-expression of the activation markers CD38 and HLA-DR but normal levels of CCR6 and CD127. MAIT cells had a lower expression of the transcription factor PLZF in HAM/TSP patients. Unlike Tax-specific CD8+T cells, which are hyperfunctional, MAIT cells from HTLV-1-infected subjects had a poor IFNγ response following antigen stimulation. MAIT cell perturbations in HTLV-1 infection were not associated with HTLV-1 proviral load and MAIT cells were not infected by HTLV-1 in vivo. Rather, MAIT cells loss was associated with immune activation. Overall, our results do not support a role for MAIT cells in HAM/TSP pathogenesis but reduced numbers of MAIT cells, together with their poor functionality, could contribute to the increased susceptibility of HTLV-1-infected individuals to other infectious agents.

The best algorithm to confirm the diagnosis of HTLV-1 and HTLV-2 in at-risk individuals from São Paulo, Brazil.

The ability to confirm the diagnosis of human T-lymphotropic virus types 1 and 2 (HTLV-1 and HTLV-2) in at-risk individuals in São Paulo, Brazil by Western blotting (WB), conventional polymerase chain reaction (tax and pol PCR) and real-time PCR (pol) is compared. Seventy-three blood samples that were reactive in HTLV-1/2 serological screening enzyme immunoassays (EIAs) were evaluated. HTLV-1/2 was confirmed in 53 blood samples: 48 were positive by WB, 41 were positive by PCR and 42 scored positive by real-time PCR assays (37 of 48 WB-positive samples plus five WB-indeterminate samples that were further confirmed by sequencing). Although WB was able to detect more cases of HTLV-1/2 infection, the real-time PCR assay was able to discriminate between these two viruses and confirm an individual HTLV-1/HTLV-2 diagnosis in two HTLV WB-untyped samples and five WB-indeterminate samples. Because of the large number of WB-indeterminate samples and the cost of the WB assay in Brazil, it is proposed an algorithm that employs two EIAs for screening and then real-time PCR to confirm the infection, followed by testing any PCR-negative samples with the WB assay. This strategy reduces costs and improves the accuracy of the diagnosis of HTLV-1/2.

Comparison of signal-to-cutoff values in first, second, and third generation enzyme immunoassays for the diagnosis of HTLV-1/2 infection in "at-risk" individuals from São Paulo, Brazil.

Data obtained during routine diagnosis of human T-cell lymphotropic virus type 1 (HTLV-1) and 2 (HTLV-2) in "at-risk" individuals from São Paulo, Brazil using signal-to-cutoff (S/C) values obtained by first, second, and third generation enzyme immunoassay (EIA) kits, were compared. The highest S/C values were obtained with third generation EIA kits, but no correlation was detected between these values and specific antibody reactivity to HTLV-1, HTLV-2, or untyped HTLV (p=0.302). In addition, use of these third generation kits resulted in HTLV-1/2 false-positive samples. In contrast, first and second generation EIA kits showed high specificity, and the second generation EIA kits showed the highest efficiency, despite lower S/C values. Using first and second generation EIA kits, significant differences in specific antibody detection of HTLV-1, relative to HTLV-2 (p=0.019 for first generation and p<0.001 for second generation EIA kits) and relative to untyped HTLV (p=0.025 for first generation EIA kits), were observed. These results were explained by the composition and format of the assays. In addition, using receiver operating characteristics (ROC) analysis, a slight adjustment in cutoff values for third generation EIA kits improved their specificities and should be used when HTLV "at-risk" populations from this geographic area are to be evaluated.