Marcos Balangero

HTLV Type 1 Genetic Types among Native Descendants in Argentina

The province of San Salvador de Jujuy, located in the northwest of Argentina, is a highly endemic area for HTLV-1 infection and a foci of tropical spastic paraparesis/HTLV-1-associated myelopathy (HAM/TSP). Therefore, to better understand this, we carried out a genetic characterization of a large set of HTLV-1 strains (n = 65) of descendants of Amerindians from this region. The LTR and env regions were analyzed. The genetic analysis showed that all of these new HTLV-1 isolates from Argentina belong to the Transcontinental subgroup A of the HTLV-1a Cosmopolitan subtype, with the exception of three isolates that cluster within the Japanese subgroup B. Interestingly, the majority of the sequences from Jujuy province belonged to a distinct cluster within the Latin America Transcontinental subgroup, referred to here as the Jujuy subcluster, and were characterized by specific signatures in the LTR. Given that the samples analyzed in this study belong to the Amerindian population and the high prevalence of HTLV-1 in Jujuy in contrast to the low prevalence of this virus in the country, it could be that HTLV-1aA was spread in Argentina from the Amerindians to the cosmopolitan population. Moreover, this is the first report of an HTLV-1aB or Japanese subgroup in descendants of non-Japanese people in South America.

Molecular epidemiology of Hepatitis B virus in Córdoba, Argentina

BACKGROUND The analysis of the genomes of hepatitis B virus (HBV) identifies phylogenetic variants called genotypes, which may lead to distinct biological and clinical behaviors. OBJECTIVES The aim of this study was to describe the current molecular epidemiology and genetic diversity of HBV in Córdoba, Argentina. STUDY DESIGN A total of 52 HBV samples, 40 from HBV mono-infected and 12 from human immunodeficiency virus (HIV)-HBV co-infected patients, were sequenced in the S gene and in the basal core promoter-precore (BCP-pC) region. RESULTS Presence of subgenotypes F1b (35%) and F4 (17.5%), subgenotype A2 (37.5%), C (5.0%) (subgenotype could not be defined) and D (5.0%) (subgenotype D2, and the other could not be defined) were observed among mono-infected patients. The co-infected individuals displayed a different genotype distribution: sub-genotype A2 was the most common (75.0%), followed by subgenotype F1b (25.0%). CONCLUSIONS These results showed two epidemiologic scenarios: the mono-infected population may represent the ethnic composition of the current human population of Córdoba, where the Amerindian (genotype F) and European origins (subgenotype A2) account for the 90% of the samples; for the co-infected patients, the high prevalence of subgenotype A2 resemble previous analyses from Buenos Aires. In addition, mutations in hepatitis B surface antigen (HBsAg), polymerase and BCP-pC regions were identified, mainly in chronic or co-infected patients.

Silent dissemination of HTLV-1 in an endemic area of Argentina. Epidemiological and molecular evidence of intrafamilial transmission

Background Molecular and epidemiological studies of transmission routes and risk factors for infection by HTLV-1 are extremely important in order to implement control measures, especially because of the high prevalence of HTLV-1 in several regions of the world. San Salvador de Jujuy, Northwest Argentina, is a highly endemic area for HTLV-1 and foci of tropical spastic paraparesis/HTLV-1-associated myelopathy. Objective To gain further insight into the role of intrafamilial transmission of HTLV-1 in a highly endemic region in Argentina. Method Cross-sectional study in Northwest Argentina. Epidemiological data and blood samples were collected from 28 HTLV-1 infected subjects (index cases) and 92 close relatives/cohabitants. HTLV-1 infection was diagnosed by detection of antibodies and proviral DNA. The LTR region was sequenced and analyzed for genetic distances (VESPA software), in addition to determination and identification of polymorphisms to define HTLV-1 family signatures. Results Fifty seven of the 120 subjects enrolled had antibodies against HTLV-1 and were typified as HTLV-1 by PCR. The prevalence rate of HTLV-1 infection in family members of infected index cases was 31.52% (29/92). The infection was significantly associated with gender, age and prolonged lactation. Identity of LTR sequences and presence of polymorphisms revealed high prevalence of mother-to-child and interspousal transmission of HTLV-1 among these families. Conclusion There is an ongoing and silent transmission of HTLV-1 through vertical and sexual routes within family clusters in Northwest Argentina. This evidence highlights that HTLV-1 infection should be considered as a matter of public health in Argentina, in order to introduce preventive measures as prenatal screening and breastfeeding control.

Correlation of HTLV-1 Tax genetic diversity with HTLV-1 associated myelopathy/tropical spastic paraparesis progression and HTLV-1a genotypes in an HTLV-1 endemic region in Argentina

The oncoprotein Tax was characterized genetically from a large cohort of human T‐cell lymphotropic virus type 1 (HTLV‐1) seropositive individuals from the most endemic region of HTLV‐1‐associated myelopathy/tropical spastic paraparesis (HAM/TSP) and HTLV‐1 infection in Argentina, the province of San Salvador de Jujuy. Sixteen HAM/TSP patients and 47 HTLV‐1 healthy carriers were evaluated. Six Tax genetic polymorphisms were identified and observed in 70.8% of healthy carriers and 62.5% of HAM/TSP patients. Tax genetic polymorphisms were not associated with clinical status but A8344C polymorphism statistically provide a borderline protective effect of HAM/TSP outcome. Nucleotide diversity in healthy carriers was 0.00549, whereas HAM/TSP virus population revealed a low diversity of 0.00379, suggests a positive selection for Tax protein conservation in this group. It is concluded that tax genetic polymorphisms do not increase the risk of developing HAM/TSP in this endemic region. However, in spite of the low prevalence of HTLV‐1aB genotype, statistical analysis revealed an important correlation of tax genetic signatures with HTLV‐1aA trans‐continental subgroup. J. Med. Virol. 82:1438–1441, 2010.

Development and validation of a real-time PCR assay for a novel HTLV-1 tax sequence detection and proviral load quantitation

A quantitative real-time PCR (qPCR) assay using SYBR Green dye was established in order to detect and quantify the proviral DNA of HTLV-1 in peripheral blood mononuclear cells (PBMCs). Primers were designed, and the assay was standardized to amplify a novel, conserved HTLV-1 tax region. Proviral load was normalized to the amount of cellular DNA by quantitation of the human albumin gene. Firstly, the qPCR was assessed determining the specificity, sensitivity, dynamic range and intra- and inter-assay reproducibility of the technique. The limit of detection as determined by PROBIT analysis using dilutions of the standard was 2.97 copies. The assay had an excellent dynamic range from 10⁵ to 10¹ copies per reaction and good intra- and inter-assay reproducibility, CVs less than 2%. Secondly, the performance of the qPCR was tested on 40 HTLV-1 seropositive individuals. Proviral load for HTLV-1 carriers ranged from 2.2×10² to more than 8.3×10⁴ copies/10⁶ PBMCs. The high sensitivity and wide dynamic range allowed the determination of a broad range of HTLV-1 proviral loads in infected individuals. This assay is a valuable alternative diagnostic tool when current available serological assays are insufficient. In addition, it will facilitate the study of the relationship between proviral load and pathogenesis.

Diagnóstico serológico de HTLV-1/2: combinación de técnicas de tamizaje para definir el estatus serológico en donantes de sangre

Resumen Con el objetivo de reducir el numero de resultados falsos reactivos al determinar los anticuerpos contra HTLV-1/2, se evaluaron algoritmos alternativos. De 20 210 muestras estudiadas, el 0,37 % (74/20 210) fueron reactivas por ELISA Murex Abbott. De estas, 23 se confirmaron positivas por inmunofluorescencia indirecta y 51 fueron negativas; valor predictivo positivo (VPP) 31,08 %. Al combinar ELISA Murex con aglutinacion de particulas de gelatina (AP) y con ELISA MP, se obtuvieron los siguientes resultados: 26/74 resultaron reactivas por Murex y AP, VPP 88,5 %, y 32/74 reactivas con Murex y ELISA MP, VPP 71,8 %. Del analisis de la curva ROC se determino que para un valor de RP de 4,74 por Murex, la sensibilidad, la especificidad, el VPP y el VPN son 100 %, 98,04 %, 95,8 % y 100 %, respectivamente. Proponemos que las muestras reactivas por ELISA Murex con RP ≤ 4,74 sean retesteadas por duplicado por AP, y que las que resulten concordantemente no reactivas sean definidas como negativas para HTLV-1/2.

Usefulness of nucleic acid testing to reduce risk of hepatitis B virus transfusion-transmitted infection in Argentina: high rate of recent infections.

Results from 10‐year experience using nucleic acid test (NAT) screening in a blood bank of Córdoba are presented, showing the first data on prevalence of recent hepatitis B virus (HBV) infections and occult HBV infections (OBIs) in Argentina.

Informe breveSensibilidad del equipo Cobas AmpliScreen™ HIV-1 Test, v1.5, para la detección de HIV-1Sensitivity of the COBAS AmpliScreen™ HIV-1 Test v1.5 for HIV-1 detection

The introduction of nucleic acid amplification techniques (NAT) in blood banks was intended to reduce the residual risk of transfusion-transmitted infections. Co-circulation of a great diversity of HIV-1 variants in Argentina portrays the need to assess the sensitivity of serological and molecular assays available for their detection. In this study, we evaluated the sensitivity of the COBAS AmpliScreen™ HIV-1 Test, version 1.5 (Roche) for the detection of HIV-1 RNA in plasma samples of infected individuals from Argentina. The results of this study reveal that this technique has high sensitivity for the detection of HIV-1 RNA under assay conditions: using mini-pool testing, pools ≥ 50 RNA copies per ml achieved ≥ 92 % sensitivity, whereas in the standard procedure, samples ≥ 207 RNA copies/ml achieved 100 % sensitivity. Moreover, the COBAS AmpliScreen™ HIV-1 Test, version 1.5 (Roche) is suitable for detecting prevailing HIV-1 variants.Resumen Las tecnicas de amplificacion de acidos nucleicos (NAT) se incorporaron en los bancos de sangre para reducir el riesgo residual de transmision de infecciones por via transfusional. La cocirculacion de distintas variantes del HIV-1 en Argentina indica la necesidad de evaluar la sensibilidad de los ensayos serologicos y moleculares disponibles para su deteccion. En este trabajo se evaluo la sensibilidad del equipo COBAS AmpliScreen™ HIV-1 Test, version 1.5 (Roche), para detectar ARN viral en plasmas de individuos infectados con HIV-1 de Argentina. Los resultados demuestran que esta tecnica tiene una alta sensibilidad para detectar ARN de HIV-1 en las condiciones ensayadas: para ensayo de mini-pooles (pooles ≥ 50 copias de ARN/ml), la sensibilidad fue ≥ 92 %, y para procedimiento estandar (plasmas ≥ 207 copias de ARN/ml), la sensibilidad fue 100 %. Ademas, la tecnica COBAS AmpliScreen™ HIV-1 Test, version 1.5 (Roche), es adecuada para la deteccion de las variantes de HIV-1 prevalentes.

Sensibilidad del equipo Cobas AmpliScreen™ HIV-1 Test, v1.5, para la detección de HIV-1

The introduction of nucleic acid amplification techniques (NAT) in blood banks was intended to reduce the residual risk of transfusion-transmitted infections. Co-circulation of a great diversity of HIV-1 variants in Argentina portrays the need to assess the sensitivity of serological and molecular assays available for their detection. In this study, we evaluated the sensitivity of the COBAS AmpliScreen™ HIV-1 Test, version 1.5 (Roche) for the detection of HIV-1 RNA in plasma samples of infected individuals from Argentina. The results of this study reveal that this technique has high sensitivity for the detection of HIV-1 RNA under assay conditions: using mini-pool testing, pools ≥ 50 RNA copies per ml achieved ≥ 92 % sensitivity, whereas in the standard procedure, samples ≥ 207 RNA copies/ml achieved 100 % sensitivity. Moreover, the COBAS AmpliScreen™ HIV-1 Test, version 1.5 (Roche) is suitable for detecting prevailing HIV-1 variants.Resumen Las tecnicas de amplificacion de acidos nucleicos (NAT) se incorporaron en los bancos de sangre para reducir el riesgo residual de transmision de infecciones por via transfusional. La cocirculacion de distintas variantes del HIV-1 en Argentina indica la necesidad de evaluar la sensibilidad de los ensayos serologicos y moleculares disponibles para su deteccion. En este trabajo se evaluo la sensibilidad del equipo COBAS AmpliScreen™ HIV-1 Test, version 1.5 (Roche), para detectar ARN viral en plasmas de individuos infectados con HIV-1 de Argentina. Los resultados demuestran que esta tecnica tiene una alta sensibilidad para detectar ARN de HIV-1 en las condiciones ensayadas: para ensayo de mini-pooles (pooles ≥ 50 copias de ARN/ml), la sensibilidad fue ≥ 92 %, y para procedimiento estandar (plasmas ≥ 207 copias de ARN/ml), la sensibilidad fue 100 %. Ademas, la tecnica COBAS AmpliScreen™ HIV-1 Test, version 1.5 (Roche), es adecuada para la deteccion de las variantes de HIV-1 prevalentes.