RESUMEN

INTRODUCTION AND OBJECTIVES: The hepatitis B virus (HBV) surface antigen (HBsAg) variations suggested having some effects on infection outcome. Due to some controversial issues, the aim of this study was to compare the pattern of HBsAg variation between asymptomatic carriers and HCC/cirrhosis patients. MATERIALS AND METHODS: In this cross-sectional study, 19 HCC/cirrhotic and 26 asymptomatic patients were enrolled. After viral DNA extraction, HBs gene was amplified using an in-house nested-PCR. Then, PCR products were introduced into bi-directional Sanger sequencing. The retrieved sequences were compared with references, to investigate the variation of immunologic sites, major hydrophilic region (MHR) of HBsAg as well as reverse transcriptase (RT), and also to determine genotype/subtype. RESULTS: The analysis of MHR and epitopes on HBsAg showed dozens of substitution, which occurred more prevalently in I110, P120, Y134, G159, S193, Y206, S207, I208, L213 and P214 positions. However, Y134N/F/L (P=0.04) and P120T/S (P=0.009) were significantly detected in MHR and B-cell epitope of HCC/Cirrhotic group. A number of truncation-related mutations were higher in HCC/Cirrhotic group (P>0.001), albeit only C69* stop codon was statistically significant (P=0.003). In RT, some potentially resistant substitutions such as Q215S, V191I and V214A, were revealed. Phylogenetic analysis showed that all of isolates belonged to genotype D, and the major serotype was ayw1. CONCLUSION: The higher frequency of substitutions in MHR and immune epitopes at positions such as Y134 and P120 as well as stop codons such as C69* in HCC/cirrhotic group might candidate them as predictive factors for infection outcome.

Asunto(s)

Carcinoma Hepatocelular/virología , Portador Sano/virología , Farmacorresistencia Viral/genética , Productos del Gen pol/genética , Antígenos de Superficie de la Hepatitis B/genética , Hepatitis B Crónica/virología , Cirrosis Hepática/virología , Neoplasias Hepáticas/virología , Adulto , Infecciones Asintomáticas , ADN Viral/análisis , Epítopos de Linfocito B/genética , Epítopos de Linfocito T/genética , Femenino , Genotipo , Humanos , Evasión Inmune/genética , Masculino , Persona de Mediana Edad , Mutación , Adulto Joven

RESUMEN

Nef proteins from all primate Lentiviruses, including the simian immunodeficiency virus of chimpanzees (SIVcpz), increase viral progeny infectivity. However, the function of Nef involved with the increase in viral infectivity is still not completely understood. Nonetheless, until now, studies investigating the functions of Nef from SIVcpz have been conducted in the context of the HIV-1 proviruses. In an attempt to investigate the role played by Nef during the replication cycle of an SIVcpz, a Nef-defective derivative was obtained from the SIVcpzWTGab2 clone by introducing a frame shift mutation at a unique restriction site within the nef sequence. This nef-deleted clone expresses an N-terminal 74-amino acid truncated peptide of Nef and was named SIVcpz-tNef. We found that the SIVcpz-tNef does not behave as a classic nef-deleted HIV-1 or simian immunodeficiency virus of macaques SIVmac. Markedly, SIVcpz-tNef progeny from both Hek-293T and Molt producer cells were completely non-infectious. Moreover, the loss in infectivity of SIVcpz-tNef correlated with the inhibition of Gag and GagPol processing. A marked accumulation of Gag and very low levels of reverse transcriptase were detected in viral lysates. Furthermore, these observations were reproduced once the tNef peptide was expressed in trans both in SIVcpzΔNef and HIV-1WT expressing cells, demonstrating that the truncated peptide is a dominant negative for viral processing and infectivity for both SIVcpz and HIV-1. We demonstrated that the truncated Nef peptide binds to GagPol outside the protease region and by doing so probably blocks processing of both GagPol and Gag precursors at a very early stage. This study demonstrates for the first time that naturally-occurring Nef peptides can potently block lentiviral processing and infectivity.

Asunto(s)

Productos del Gen nef/metabolismo , VIH-1/fisiología , Virus de la Inmunodeficiencia de los Simios/fisiología , Replicación Viral , Animales , Línea Celular , Mutación del Sistema de Lectura , Técnicas de Inactivación de Genes , Productos del Gen gag/metabolismo , Productos del Gen nef/genética , Productos del Gen pol/metabolismo , Humanos , Pan troglodytes , Unión Proteica , Virus de la Inmunodeficiencia de los Simios/genética

RESUMEN

BACKGROUND: Nucleoside/nucleotide analogue (NA) treatment causes selection pressure for HBV strains carrying mutations conferring NA resistance. Drug-resistance mutations occur in the reverse transcriptase (RT) region of the HBV polymerase gene and spontaneously arise during viral replication. These mutations can also alter the hepatitis B surface (HBs) protein and in some cases reduce binding to HBs antibodies. The spread of NA-resistant HBV may impact the efficacy of antiviral treatment and hepatitis B immunization programmes. In this study, we used direct sequencing to assess the occurrence of HBV carrying known mutations that confer NA resistance in the largest cohort of treatment-naive patients with chronic hepatitis B (CHB) to date. METHODS: HBV DNA samples isolated from 702 patients were sequenced and the RT region subjected to mutational analysis. RESULTS: There was high genetic variability among the HBV samples analysed: A1 (63.7%), D3 (14.5%), A2 (3.3%), A3 (0.1%), B1 (0.1%), B2 (0.1%), C2 (0.9%), D1 (0.9%), D2 (4.6%), D4 (5.1%), D unclassified subgenotype (0.7%), E (0.6%), F2a (4.6%), F4 (0.4%) and G (0.4%). HBV strains harbouring mutations conferring NA resistance alone or combined with compensatory mutations were identified in 1.6% (11/702) of the patients. CONCLUSIONS: HBV strains harbouring resistance mutations can comprise the major population of HBV quasispecies in treatment-naive patients. In Brazil, there is a very low frequency of untreated patients who are infected with these strains. These findings suggest that the spread and natural selection of drug-resistant HBV is an uncommon event and/or most of these strains remain unstable in the absence of NA selective pressure.

Asunto(s)

Antivirales/farmacología , Farmacorresistencia Viral/genética , Productos del Gen pol/genética , Virus de la Hepatitis B/efectos de los fármacos , Hepatitis B Crónica/virología , Mutación , Adenina/análogos & derivados , Adenina/farmacología , Anticuerpos Antivirales/genética , Anticuerpos Antivirales/inmunología , Brasil , ADN Viral/genética , ADN Viral/inmunología , Productos del Gen pol/antagonistas & inhibidores , Productos del Gen pol/metabolismo , Genotipo , Guanina/análogos & derivados , Guanina/farmacología , Antígenos de Superficie de la Hepatitis B/genética , Antígenos de Superficie de la Hepatitis B/inmunología , Virus de la Hepatitis B/genética , Virus de la Hepatitis B/inmunología , Hepatitis B Crónica/diagnóstico , Hepatitis B Crónica/inmunología , Humanos , Lamivudine/farmacología , Pruebas de Sensibilidad Microbiana , Organofosfonatos/farmacología , Estudios Retrospectivos , Análisis de Secuencia de ADN , Tenofovir/farmacología , Replicación Viral/efectos de los fármacos

RESUMEN

During the 1990s, high prevalences of HIV/human T lymphotropic virus type 1 (HTLV-1) and HIV/human T lymphotropic virus type 2 (HTLV-2) coinfections were detected in São Paulo, Brazil in association with intravenous drug use (IDU). The current prevalences and risk factors for HIV/HTLV-1/-2 were evaluated in 1,608 patients attending the AIDS/STD Reference and Training Center in São Paulo. Blood samples were analyzed for HTLV-1/2-specific antibodies using enzyme immunoassays (EIA Murex HTLV-I+II, Diasorin, and Gold ELISA HTLV-I+II, REM) and immunoblotting (HTLV Blot 2.4, MP Biomedicals and INNO-LIA HTLV-I/II, Innogenetics) and for the pol proviral DNA segments of HTLV-1 and HTLV-2 by "in-house" real-time PCR. These analyses revealed that 50 (3.11%) of the samples were HTLV positive, including 25 (1.55%) that were HTLV-1 positive, 21 (1.31%) that were HTLV-2 positive, and 4 (0.25%) that were HTLV positive (untypeable). The median age of the HIV/HTLV-coinfected individuals was 50 years versus 44 years in the overall population (p=0.000). The risk factors associated with HIV/HTLV-1/-2 coinfections were female gender (OR 3.26, 1.78-5.95), black/pardo color (OR 2.21, 1.21-4.03), infection with hepatitis B virus (HBV) (OR 4.27, 2.32-7.87) or hepatitis C virus (HCV) (OR 24.40, 12.51-48.11), and intravenous drug use (IDU) (OR 30.01, 15.21-59.29). The current low prevalence of HTLV-1/2 in HIV-infected patients in São Paulo could be explained in part by programs providing IDUs with sterile needles and syringes and changes in the drug usage patterns of individuals from injecting cocaine to smoking crack cocaine.

Asunto(s)

Infecciones por VIH/complicaciones , Infecciones por HTLV-I/epidemiología , Infecciones por HTLV-II/epidemiología , Adulto , Anciano , Animales , Anticuerpos Antivirales/sangre , Brasil/epidemiología , Estudios Transversales , Femenino , Productos del Gen pol/genética , Humanos , Immunoblotting , Técnicas para Inmunoenzimas , Masculino , Persona de Mediana Edad , Reacción en Cadena de la Polimerasa , Prevalencia , Factores de Riesgo

RESUMEN

Human immunodeficiency virus type 1 (HIV-1) genetic diversity is one of the most important features of HIV-1 infections and the result of error accumulation during reverse transcription and of high viral turnover. HIV-1 reverse transcription is influenced by factors such as the level of nucleotides and/or the cellular activation state. HIV-1 diversity was investigated after 48 h of viral propagation in peripheral blood mononuclear cells (PBMCs) obtained from healthy donors in three different cell culture conditions: (1) resting PBMCs, (2) simultaneous infection and PBMC activation, and (3) PBMC activation 72 h before infection. Cellular DNA was extracted and proviruses of each culture condition were amplified. Single-genome PCR clones were obtained and the protease and reverse transcriptase of the pol gene were sequenced. An elevated number of nucleotide substitutions in all three culture conditions were observed. In condition 1, the mutational rate observed ranged from 1.0 × 10(-3) to 2.1 × 10(-2), the genetic diversity was 0.6%, and hypermutation was observed in 7.1% of sequenced clones. In condition 2, the mutational rate ranged from 1.0 × 10(-3) to 1.0 × 10(-2), the genetic diversity was 0.8%, and hypermutation affected 6.7% of clones. In condition 3, the mutational rate ranged from 2.8 × 10(-3) to 1.1 × 10(-2), the genetic diversity was 1%, and 5.9% of clones were hypermutated. Substitutions occurred more frequently in some specific nucleotide stretches, and a common pattern for substitutions in all the different conditions was identified. There was a significant accumulation of mutations during the initial periods of in vitro HIV-1 propagation irrespective of culture conditions. The rapid accumulation of virus diversity might represent a viral strategy when colonizing new hosts. Complementary studies are necessary to allow for a better understanding of the initial periods of infection, which represent a crucial event related to disease progression.

Asunto(s)

Variación Genética , VIH-1/crecimiento & desarrollo , VIH-1/genética , Leucocitos Mononucleares/virología , Mutación , Productos del Gen pol/genética , Proteasa del VIH/genética , Transcriptasa Inversa del VIH/genética , Humanos , Tasa de Mutación , Análisis de Secuencia de ADN , Cultivo de Virus

RESUMEN

BACKGROUND AND AIMS: Patients undergoing hemodialysis are at risk of infection with both hepatitis B virus (HBV) and hepatitis C virus (HCV). Occult HBV infection is usually associated with low levels of HBV and is frequently detected in HCV-infected patients. The aims of the present study were to compare the prevalence of occult HBV infection among anti-HCV-positive and anti-HCV-negative patients undergoing hemodialysis, and characterize the molecular patterns of HBV isolates from patients with occult infection. METHODS: Serum samples from 100 patients negative for hepatitis B surface antigen undergoing hemodialysis, half of whom were positive for anti-HCV antibodies, were tested for the presence of HBV-DNA using semi-nested polymerase chain reaction (PCR). PCR products of the S gene were directly sequenced. RESULTS: HBV-DNA was detected in 15 samples. There were no significant differences in HCV status, sex, age, time of dialysis, alanine aminotransferase levels or HBV serological markers between patients with or without occult HBV infection, with the exception of antibody to hepatitis B core antigen (anti-HBc)-only serological marker (P = 0.003). All six HBV isolates that could be sequenced were of genotype A/subgenotype A1. Four of these six HBV isolates contained mutations associated with lamivudine resistance in the DNA polymerase (two with L180M/M204V and two with rt173V/180M/204V) and a specific substitution (Y100C) in the HBV small surface protein. CONCLUSIONS: HBV isolates with the identified substitutions have the potential to spread silently by nosocomial transmission within the hemodialysis unit. These results have potential implications for the management of patients with occult HBV infection undergoing hemodialysis.

Asunto(s)

Antivirales/uso terapéutico , Infección Hospitalaria/tratamiento farmacológico , Farmacorresistencia Viral/genética , Virus de la Hepatitis B/genética , Hepatitis B/diagnóstico , Lamivudine/uso terapéutico , Mutación , Diálisis Renal , Adulto , Anciano , Brasil/epidemiología , Infección Hospitalaria/diagnóstico , Infección Hospitalaria/epidemiología , Infección Hospitalaria/transmisión , ADN Viral/sangre , Femenino , Productos del Gen pol/genética , Genotipo , Hepatitis B/tratamiento farmacológico , Hepatitis B/epidemiología , Hepatitis B/transmisión , Anticuerpos contra la Hepatitis B/sangre , Antígenos de Superficie de la Hepatitis B/sangre , Virus de la Hepatitis B/inmunología , Anticuerpos contra la Hepatitis C/sangre , Humanos , Masculino , Persona de Mediana Edad , Datos de Secuencia Molecular , Reacción en Cadena de la Polimerasa , Prevalencia , Proteínas del Envoltorio Viral/genética

RESUMEN

Feline immunodeficiency virus (FIV) is the Lentivirus responsible for an immunodeficiency-like disease in domestic cats (Felis catus). FIV is divided into five phylogenetic subtypes (A, B, C, D, and E), based on genetic diversity. Knowledge of the geographical distribution of subtypes is relevant for understanding different disease progressions and for vaccine development. In this study, viral sequences of 26 infected cats from Rio de Janeiro, 8 undergoing treatment with zidovudine (AZT) for at least 5 years, were successfully amplified from blood specimens. gag capsid (CA), pol reverse transcriptase (RT), and env gp120 (V3-V4) regions were analyzed to determine subtypes and to evaluate potential mutations related to antiretroviral drug resistance among treated cats. Subtyping based on phylogenetic analysis was performed by the neighbor-joining and maximum likelihood methods. All of the sequences clustered with subtype B in the three regions, exhibiting low genetic variability. Additionally, we found evidence that the same virus is circulating in animals in close contact. The analysis of FIV RT sequences identified two new putative mutations related to drug resistance located in the RT "finger" domain, which has 60% identity to human immunodeficiency virus (HIV) sequence. Amino acid change K-->R at codons 64 and 69 was found in 25% and 37.5% of the treated animals, respectively. These signatures were comparable to K65R and K70R thymidine-associated mutations found in the HIV-1 HXB2 counterpart. This finding strongly suggests a position correlation between the mutations found in FIV and the K65R and K70R substitutions from drug-resistant HIV-1 strains.

Asunto(s)

Síndrome de Inmunodeficiencia Adquirida del Felino/tratamiento farmacológico , Productos del Gen env/genética , Productos del Gen gag/genética , Productos del Gen pol/genética , Virus de la Inmunodeficiencia Felina/metabolismo , Inhibidores de la Transcriptasa Inversa/farmacología , Animales , Brasil , Gatos , Farmacorresistencia Viral , Síndrome de Inmunodeficiencia Adquirida del Felino/virología , Femenino , Productos del Gen env/metabolismo , Productos del Gen gag/metabolismo , Productos del Gen pol/metabolismo , Técnicas Genéticas , Variación Genética , Masculino , Filogenia

RESUMEN

The in vivo evolution of genotype F HBV variants was recorded in a chronically infected patient throughout a 3-year observation period. Fluctuating levels of HBs Ag and anti-HBs antibodies were recorded, both of them cocirculating in peripheral blood samples at given times. Fifty S gene derived clones were sequenced and phylogenetically analyzed. As expected, some amino acid replacements within the S ORF were also observed within the P ORF while others were silent for the former. Such change was statistically significant for both S and P overlapping genes, which clearly indicates the appearance of a positive selection pressure. Supporting this notion, amino acid replacements were documented at both B and T cell epitopes in samples from 1997 and 1998. Several mutations were documented within and outside the "a" determinant in the major hydrophilic region. Such substitutions might have resulted from the attempt of HBV to evade both humoral and/or cellular immune response. To the best of our knowledge this unusual profile of HBV variants in presence of usually "neutralizing" anti-HBs antibodies was examined in vivo for the first time.

Asunto(s)

Evolución Molecular , Genes Virales/genética , Anticuerpos contra la Hepatitis B/sangre , Antígenos e de la Hepatitis B/sangre , Hepatitis B Crónica/sangre , Hepatitis B Crónica/virología , Hepatitis B/genética , Secuencia de Aminoácidos , Productos del Gen pol/genética , Humanos , Datos de Secuencia Molecular , Proteínas del Envoltorio Viral/genética

RESUMEN

Worldwide, the distribution of HIV-1 subtypes and intersubtype recombinants is not homogeneous. In Latin America and the Caribbean, HIV-1 subtype B predominates. However, in the south of Brazil and in countries of the Southern cone (Argentina, Chile, Paraguay, and Uruguay) there is a different distribution of viral subtypes and intersubtype recombinants. The aim of this work was to analyze HIV-1 diversity in a cohort of pregnant women (with primarily heterosexual acquisition of the infection) who were diagnosed with HIV-1 infection during their current pregnancy and who received ARVs during pregnancy for perinatal transmission prophylaxis. Analysis of 121 partial pol sequences from subjects enrolled in Argentina, Brazil, the Bahamas, and Mexico was performed by phylogenetic and recombinant characterization. Different prevalences of subtype B were observed (100% for specimens from Mexico and the Bahamas, 61% for Brazil, and 30% for Argentina). Subtypes C and F were found, along with BC, BF, FC, and CBF recombinants in specimens from Brazilians. A high prevalence of BF recombinants was found (70%) in specimens from Argentina. The different patterns of HIV- 1 subtypes and intersubtype recombinants in South America (Argentina and Brazil) compared to those in Central and North America should be considered in the design of future HIV-1 vaccine trials.

Asunto(s)

Variación Genética , Infecciones por VIH/epidemiología , Infecciones por VIH/virología , VIH-1/genética , VIH-1/aislamiento & purificación , Argentina/epidemiología , Bahamas/epidemiología , Brasil/epidemiología , Femenino , Productos del Gen pol/genética , Humanos , México/epidemiología , Filogenia , Embarazo

RESUMEN

Serum hepatitis B virus (HBV) DNA was extracted from a chronically infected patient with cocirculation of hepatitis B surface antigen (HBsAg) and anti-HBs antibodies. Direct PCR and clone-derived sequences of the S and overlapped P genes were obtained. DNA sequences and phylogenetic analysis ascribed this isolate to genotype A (serotype adw2). Five of six HBV DNA clones exhibited point mutations inside and outside the major hydrophilic region, while the sixth clone exhibited a genotype A "wild-type" amino acid sequence. Observed replacements included both humoral and/or cellular (major histocompatibility complex class I [MHC-I] and MHC-II) HBV mutated epitopes, such as S45A, P46H, L49H, C107R, T125A, M133K, I152F, P153T, T161S, G185E, A194T, G202R, and I213L. None of these mutants were individually present within a given clone. The I213L replacement was the only one observed in the five clones carrying nonsynonymous mutations in the S gene. Some of the amino acid substitutions are reportedly known to be responsible for the emergence of immune escape mutants. C107R replacement prevents disulfide bonding, thus disrupting the first loop of the HBsAg. Circulation of some of these mutants may represent a potential risk for the community, since neither current hepatitis B vaccines nor hyperimmune hepatitis B immune globulin are effectively prevent the liver disease thereto associated. Moreover, some of the recorded HBsAg variants may influence the accuracy of the results obtained with currently used diagnostic tests.

Asunto(s)

Epítopos , Anticuerpos contra la Hepatitis B/sangre , Antígenos de Superficie de la Hepatitis B/sangre , Virus de la Hepatitis B/genética , Hepatitis B Crónica/virología , Mutación Puntual , Adulto , Secuencia de Aminoácidos , Sustitución de Aminoácidos , Epítopos/genética , Productos del Gen pol/genética , Antígenos de Superficie de la Hepatitis B/genética , Virus de la Hepatitis B/inmunología , Humanos , Masculino , Datos de Secuencia Molecular , Filogenia , Análisis de Secuencia de ADN

RESUMEN

Se estudiaron pacientes seropositivos para el virus de inmunodeficiencia humana tipo 1 (VIH-1) con y sin tratamiento, con el fin de determinar el polimorfismo y la prevalencia de mutaciones de resistencia a la terapia antirretroviral. El material genético viral fue extraído a partir de células mononucleares de sangre periféricas (ADN) y del plasma (ARN) de 30 pacientes. Se amplificaron 2 regiones del gen Pol, Transcriptasa Reversa (TR) y Proteasa (Pr) y el gen de envoltura (Env) por medio de la técnica de PCR y se obtuvo la secuencia genómica de los productos. Todos los aislados analizados pertenecieron al subtipo B. No se observaron mutaciones primarias asociadas a resistencia a inhibidores de Pr pero sí un alto porcentaje (86 por ciento, 19/22) de mutaciones no asociadas con resistencia sino a restitución de la capacidad replicativa de cepas mutantes (mutaciones secundarias). Se observó la presencia de mutaciones asociadas a resistencia a inhibidores nucleósidos de la TR (INTR) en 35 por ciento (6/17) de los pacientes sometidos a tratamiento, mientras que 12 por ciento (2/17) de ellos presentaron mutaciones de resistencia a inhibidores no nucleósidos de la TR (INNTR). Interesantemente, un paciente no tratado estaba infectado con una cepa que presentaba mutaciones primarias (7,7 por ciento); este resultado sugiere que podría ser importante plantearse el estudio local de determinación de resistencia genotípica en pacientes antes del tratamiento, con miras a minimizar fallas terapéuticas. Se requieren estudios adicionales para evaluar el rol de las mutaciones secundarias en el éxito de la infección viral

Asunto(s)

Humanos , Productos del Gen pol , VIH-1 , Medicina , Venezuela

RESUMEN

Partial sequences of HIV-1 polymerase from 185 patients, 141 ARV experienced and 44 naive, of gag (p24) and env (C2V3) from a subset of naive cases were evaluated in São Paulo, Brazil. Antiretroviral resistance mutations were detected in 4% of 26 recently (<2 years) infected patients. Polymorphisms at the protease gene were common both in contemporary and pre-HAART era isolates, some significantly associated with the viral clade. HIV-1 clade B was preponderant, in 79%, with 11% clade F and one case of HIV-1 C. Recently infected women had a significantly higher proportion of non-B clade HIV-1. A mosaic pol was observed in 9%, all B/F except for one G mosaic. A CRF-12-BF structure, observed in 20% of B/F pol mosaics, provides evidence for an epidemic relationship in the major South American metropolitan areas.

Asunto(s)

Infecciones por VIH/virología , VIH-1/clasificación , Secuencia de Aminoácidos , Secuencia de Bases , Brasil , Cartilla de ADN , Femenino , Productos del Gen env/genética , Productos del Gen gag/genética , Productos del Gen pol/genética , Humanos , Masculino , Datos de Secuencia Molecular , Filogenia , Homología de Secuencia de Aminoácido

RESUMEN

In order to investigate the existing notion that the human immunodeficiency virus type 1 (HIV-1) infection in Puerto Rico (PR) was imported from the continental United States of America (USA) we sequenced and analyzed 900 bases of the HIV-1 pol sequence from individuals in PR for comparison with pol sequences from the USA mainland. The sequences were derived by direct sequencing of reverse transcription-polymerase chain reaction products generated from plasma virus. The products were sequenced in both directions and the complementary strands were compared prior to analysis. These processed sequences and GenBank sequences from the continental USA were subjected to phylogenetic analyses. The PR and USA sequences did not form independent clusters, indicating a shared HIV-1 infection. This may be due to the continuous human traffic or, less likely, may indicate a similar evolution of a common source virus. Analysis of drug resistance mutations, fairly similar in frequency in the PR and USA sequences analyzed here, supports human traffic as a rationale for the common infection. This work indicates that an efficacious vaccine developed for use in the USA mainland will also be effective in prevention in PR and perhaps the other countries of the Caribbean region.

Asunto(s)

Productos del Gen pol/genética , VIH-1/genética , Filogenia , Farmacorresistencia Viral/genética , Evolución Molecular , Infecciones por VIH/epidemiología , Infecciones por VIH/transmisión , Humanos , Puerto Rico/epidemiología , Alineación de Secuencia , Análisis de Secuencia de ARN , Estados Unidos/epidemiología

RESUMEN

The emergence of antiretroviral (ARV) drug-resistant human immunodeficiency virus type 1 (HIV-1) quasispecies is a major cause of treatment failure. These variants are usually replaced by drug-sensitive ones when the selective pressure of the drugs is removed, as the former have reduced fitness in a drug-free environment. This was the rationale for the design of structured ARV treatment interruption (STI) studies for the management of HIV-1 patients with treatment failure. We have studied the origin of drug-sensitive HIV-1 quasispecies emerging after STI in patients with treatment failure due to ARV drug resistance. Plasma and peripheral blood mononuclear cell samples were obtained the day of treatment interruption (day 0) and 30 and 60 days afterwards. HIV-1 pol and env were partially amplified, cloned, and sequenced. At day 60 drug-resistant variants were replaced by completely or partially sensitive quasispecies. Phylogenetic analyses of pol revealed that drug-sensitive variants emerging after STI were not related to their immediate temporal ancestors but formed a separate cluster, demonstrating that STI leads to the recrudescence and reemergence of a sequestrated viral population rather than leading to the back mutation of drug-resistant forms. No evidence for concomitant changes in viral tropism was seen, as deduced from env sequences. This study demonstrates the important role that the reemergence of quasispecies plays in HIV-1 population dynamics and points out the difficulties that may be found when recycling ARV therapies with patients with treatment failure.

Asunto(s)

Fármacos Anti-VIH/farmacología , Evolución Molecular , Infecciones por VIH/tratamiento farmacológico , VIH-1/clasificación , VIH-1/efectos de los fármacos , Inhibidores de la Transcriptasa Inversa/farmacología , Adulto , Secuencia de Aminoácidos , Fármacos Anti-VIH/uso terapéutico , Esquema de Medicación , Farmacorresistencia Viral/genética , Femenino , Productos del Gen env/genética , Productos del Gen pol/genética , Infecciones por VIH/virología , Proteasa del VIH/genética , Transcriptasa Inversa del VIH/genética , VIH-1/enzimología , Humanos , Masculino , Datos de Secuencia Molecular , Filogenia , Inhibidores de la Transcriptasa Inversa/uso terapéutico , Análisis de Secuencia de ADN , Insuficiencia del Tratamiento

RESUMEN

The first lentivirus isolated from sheep in Brazil was analysed phylogenetically. Evolutionary trees of the proviral 597 nucleotide gag and 432 nucleotide pol sequences obtained by the maximum likelihood method demonstrated that the sheep isolate clustered with prototype Maedi Visna virus whereas three lentiviruses isolated from goats in the same geographic region were close to caprine arthritis encephalitis prototypes. A subsequent comparison of sequence data of these viruses with those contained in the EMBL sequence database revealed that, in contrast to caprine prototypic viruses, all prototypic Maedi Visna viruses contain a deletion of six nucleotides in the gag gene resulting in the deletion of two residues in the central region of capsid protein. This deletion may be a useful marker in the analysis of small ruminant lentiviruses, especially when considering possible transmission of lentiviruses between sheep and goats.

Asunto(s)

Productos del Gen gag/genética , Productos del Gen pol/genética , Cabras/virología , Lentivirus/genética , Ovinos/virología , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Brasil , ADN Viral , Lentivirus/clasificación , Lentivirus/aislamiento & purificación , Datos de Secuencia Molecular , Filogenia , Homología de Secuencia de Aminoácido

RESUMEN

Among the major circulating HIV-1 subtypes, subtype C is the most prevalent. To generate full-length subtype C clones and sequences, we selected 13 primary (PBMC-derived) isolates from Zambia, India, Tanzania, South Africa, Brazil, and China, which were identified as subtype C by partial sequence analysis. Near full-length viral genomes were amplified by using a long PCR technique, sequenced in their entirety, and phylogenetically analyzed. Amino acid sequence analysis revealed 10.2, 6.3, and 17.3% diversity in predicted Gag, Pol, and Env protein sequences. Ten of 13 viruses were nonmosaic subtype C genomes, while all three isolates from China represented B/C recombinants. One of them was composed primarily of subtype C sequences with three small subtype B portions in gag, pol, and nef genes. Two others exhibited these same mosaic regions, but contained two additional subtype B portions at the gag/pol overlap and in the accessory gene region, suggesting ongoing B/C recombination in China. All subtype C genomes contained a prematurely truncated second exon of rev, but other previously proposed subtype C signatures, including three potential NF-kappa B-binding sites in the viral promoter-enhancer regions, were found in only a subset of these genomes.

Asunto(s)

Genoma Viral , Infecciones por VIH/virología , VIH-1/genética , Adulto , Secuencia de Bases , Brasil , China , Femenino , Productos del Gen env , Productos del Gen gag/genética , Productos del Gen pol/genética , Productos del Gen rev , Duplicado del Terminal Largo de VIH/genética , VIH-1/clasificación , Humanos , India , Masculino , Persona de Mediana Edad , Datos de Secuencia Molecular , Filogenia , Alineación de Secuencia , Sudáfrica , Tanzanía , Productos del Gen rev del Virus de la Inmunodeficiencia Humana

RESUMEN

This paper describes genetic subtypes of HIV-1 found in blood samples from 31 HIV-1-infected people who visited the Counseling and Testing AIDS Center of Instituto de Medicina Tropical in Manaus, Brazil. Manaus, the main city in Brazil's Amazon Basin, is also the closest urban connection for more than 100,000 Indians living in the rain forests of this region. Although to date there is no evidence of increased incidence of HIV-1 infection among the indigenous population, our understanding of both the prevalence and nature of the epidemic in the region as a whole is limited. From the 31 samples analyzed by C2V3 sequencing, we found almost equal proportions of HIV-1 strains belonging to subtype B (n = 16; 51.6%) and subtype F (n = 15; 48.4%), a finding that differs from results from previous studies conducted in urban areas of southeastern Brazil. We also observed the presence of the GWGR amino-acid sequence in the critical tetra-peptide crown of the env V3 loop in the HIV-1 subtype B samples analyzed. Among these samples, we also found 14 mosaic genomes (45.16%) in which different combinations of subtypes B, C, and F were identified between the p24 gag, pro, and env regions. Our data support the hypothesis that the Amazonian HIV-1 infections linked to the urban epidemic in southeastern Brazil. The genetic diversity and the prevalence of mosaic genomes among the isolates in our study confirm an integral role of recombination in the complex Brazilian epidemic.

Asunto(s)

Brotes de Enfermedades , Infecciones por VIH/epidemiología , VIH-1/clasificación , VIH-1/genética , Recombinación Genética , Adulto , Brasil/epidemiología , ADN Viral/análisis , Femenino , Productos del Gen pol/genética , Proteína p24 del Núcleo del VIH/genética , Proteína gp120 de Envoltorio del VIH/genética , Infecciones por VIH/virología , Proteasa del VIH/genética , VIH-1/aislamiento & purificación , Humanos , Indígenas Sudamericanos , Masculino , Persona de Mediana Edad , Datos de Secuencia Molecular , Fragmentos de Péptidos/genética , Filogenia , Reacción en Cadena de la Polimerasa , Análisis de Secuencia de ADN

RESUMEN

For reliable classification of HIV-1 strains appropriate reference sequences are needed. The HIV-1 genetic subtype F has a wide geographic spread, causing significant epidemics in South America, Africa, and some regions of Europe. Previously only two full-length sequences of each of the HIV-1 subtype F subclusters F1 and F2 have been described. To extend the knowledge of subtype F variation on a complete genome level, three new virtually full-length F1 sequences were cloned and sequenced, two from Africa and one from South America. Comparison of the new and previously described sequences showed that monophyletic clustering of the subcluster F1 of subtype F is consistent and highly supported in all genome regions. Two additional full-length strains were shown to be mosaics of subtypes F and D. These epidemiologically unrelated F/D sequences showed similar chimeric structure, suggesting that they may represent a previously undescribed circulating recombinant form (CRF). This was supported by partial sequences from three additional unlinked F/D recombinants. Genetic distances in the phylogenetic trees suggest that the recombination event leading to the putative CRF occurred relatively long ago, close to the divergence of the F1 and F2 subclusters. Furthermore, all five F/D recombinants are linked to the Democratic Republic of Congo, suggesting that the original recombination event took place in central Africa.

Asunto(s)

Genoma Viral , VIH-1/clasificación , VIH-1/genética , Filogenia , África , Clonación Molecular , Evolución Molecular , Femenino , Productos del Gen env/genética , Productos del Gen gag/genética , Productos del Gen pol/genética , Variación Genética/genética , Infecciones por VIH/virología , Humanos , Masculino , Datos de Secuencia Molecular , Proteínas Recombinantes de Fusión/genética , Recombinación Genética/genética , Homología de Secuencia de Ácido Nucleico , América del Sur