Carolina Torres

Hepatitis B virus genetic diversity in Argentina: Dissimilar genotype distribution in two different geographical regions; description of hepatitis B surface antigen variants

BACKGROUND The hepatitis B virus (HBV) molecular epidemiological data of Argentina are still scarce, since most of the previous analyses have been performed in the Metropolitan Region. OBJECTIVES To deepen the current molecular and epidemiological information about the geographical distribution of HBV genotypes and subgenotypes, and to describe the hepatitis B surface antigen (HBsAg) variants circulating in Argentina. STUDY DESIGN Eighty-eight Argentine partial HBsAg sequences from both the Northern and the Metropolitan Regions of the country were analyzed along with 67 Argentine HBV sequences existing in GenBank. RESULTS Phylogenetic and amino acid sequence analysis grouped the 88 samples as genotypes A (14.8%), D (21.6%) and F (63.6%). In the Northern Region, 44 out of the 48 sequences analyzed (91.7%) grouped as genotype F. Differently, in the Metropolitan Region, the 40 samples grouped as genotype F (30.0%), genotype D (42.5%), and genotype A (27.5%). An elevated proportion (14.8%) of the genomes presented mutations in the major hydrophilic region (MHR). CONCLUSIONS The different genotype distribution in both Argentine regions indicates that the epidemiological landscape of HBV infection appears to be the result of the diverse human migratory movements that have given shape to the present population. Our findings show that the prevalence of HBsAg variants is quite significant among the Argentine population.

Hepatitis B virus in Buenos Aires, Argentina: genotypes, virological characteristics and clinical outcomes

Hepatitis B virus (HBV) is classified into eight major genotypes, A-H, which are geographically distributed worldwide. The aim of this work was to describe the clinical characteristics associated with the HBV genotypes circulating in Buenos Aires city. The study included 139 patients infected with HBV, whose clinical courses were classified as acute symptomatic self-limiting hepatitis, inactive carrier state and chronic active hepatitis (HBV e-antigen (HBeAg)-positive and HBeAg-negative). The HBV genotypes were determined in 128 patients by PCR-restriction fragment length polymorphism and phylogenetic analysis. Biochemical, virological, clinical and histological features were analysed. A differential distribution of genotypes between acute symptomatic and chronic infections was found. Among the acute cases, genotype F was predominant (65.2%, 30/46) and genotype D was rare (4.3%, 2/46), whereas among the chronic infections, a homogeneous distribution of genotypes A (26.8%, 22/82), D (31.7%, 26/82) and F (36.6%, 30/82), with an unusual presence of genotypes B (1.2%, 1/82) and C (3.7%, 3/82), was observed. Regarding the liver histology of chronically infected patients, genotype F tended to display higher histological activity indexes. Mutations related to HBV surface antigen immunoreactivity, antiviral resistance and HBeAg-negative status were studied. This work constitutes, to our knowledge, the first description of the clinical characteristics related to HBV genotypes in Argentina, where the distribution of genotypes in patients with acute infection has not been reported previously. Finally, it was established that genotype F is the prevalent genotype among the acute symptomatic infections in Buenos Aires city, and that it shows a tendency to cause an adverse disease outcome among the chronic cases.

Environmental surveillance of norovirus in Argentina revealed distinct viral diversity patterns, seasonality and spatio-temporal diffusion processes.

Norovirus (NoV) contamination was evaluated in five rivers of Argentina between 2005 and 2011. NoV was present in all sampled rivers, with distinct NoV patterns in waters impacted by different-sized communities. In rivers affected by medium-sized populations (Salta and Córdoba cities) only one or two genotypes were present, GII.4 being the main one, with winter seasonality. In contrast, in the much more heavily populated area of Buenos Aires city the prevalent GII.4 was accompanied by several additional genotypes (GII.4, GII.b, GII.2, GII.7, GII.17, GII.e and GII.g) and one ungenotyped GII NoV, with no clear seasonality. GII.4 2006b was the main variant detected (60.9%). Phylogeographic and phylodynamic analyses performed in region D of the VP1 gene showed a most recent common ancestor in 2002 and a substitution rate of 3.7×10(-3) substitutions per site per year (HPD95%=2.3×10(-3)-5.2×10(-3)) for this variant still involving a significant population size with a slight decrease since 2008. The spatio-temporal diffusion analysis proposed Europe as an intermediate path between the American Continent and the rest of the World for NoV dissemination. Given the importance of NoV as a cause of epidemic gastroenteritis and the likelihood of its environmental transmission, the results of this work should increase public and institutional awareness of the health risk involved in sewage discharges into the environment. Environmental surveillance of enteric viruses could be a very useful tool not only to prevent waterborne outbreaks, but also to describe the epidemiology of the viruses. The detailed analysis of the viral genomes disposed into the environment contributed to the characterization of the dissemination, diversity and seasonality of NoV in its natural host population. In future studies, environmental surveillance and molecular analysis should be complemented with a quantitative viral risk assessment for estimating the disease burden from viruses in the environment.

New perspectives on the evolutionary history of hepatitis B virus genotype F.

Hepatitis B virus (HBV) is a globally distributed human pathogen. The aim of this work was to analyze the evolutionary history of HBV genotype F, emphasizing on the study of subgenotypes prevalent in the Southern area of South America. Complete genomes of HBV genotype F from 36 samples from Argentina and Chile were sequenced and analyzed by phylogenetic and Bayesian coalescent methods along with sequences obtained from GenBank database. The phylogeography separated not only Central American from South American isolates but also revealed that different subgenotypes are distributed in constrained although not exclusive areas of the continent. The result obtained with time-stamped complete genomes failed to explain the wide geographical distribution and the clustering observed in this genotype. Conversely, the use of Bayesian coalescent analyses with substitution rates as priors, instead of the co-estimation of tMRCA and substitution rate, allowed us to propose a far origin for the HBV genotype F based on the phylogeographical and epidemiological data.

Genetic and evolutionary characterization of norovirus from sewage and surface waters in Córdoba City, Argentina

Noroviruses (NoVs) are among the most common viral agents that cause gastroenteritis in humans of all ages worldwide. They are excreted in the feces and introduced into environmental waters as raw or treated sewage. In this work, sewage and water samples collected from the Suquía River in the city of Córdoba, Argentina, were evaluated for the presence of NoV. Phylogenetic analysis demonstrated that the main genotype detected was GII.4, belonging to the widely-distributed 2006b variant, followed by strains related to the putative recombinant GII.g virus. Detected NoVs were more phylogenetically related with recent viruses from other countries than with previous local sequences, suggesting a rapid and wide spread of viral strains that prevents a geographically structured phylogeny. A Bayesian coalescent analysis demonstrated that variants isolated in this work have a most recent common ancestor placed in 2007-2008 with estimated substitution rates of 3.7-5.8×10(-3)s/s/y. Environmental samples showed a mixture of both viral types, pointing up to the co-circulation and the risk of mixed infections and recombination. This is the first report on the detection and characterization of NoV in sewage and river water in Argentina.

Analysis of the circulation of hepatitis A virus in Argentina since vaccine introduction

Hepatitis A virus (HAV) has shown intermediate endemicity in Argentina, but its incidence has decreased since vaccine introduction in 2005. Environmental surveillance was conducted in five rivers from Argentina from 2005 to 2012, complementing clinical information. HAV detection decreased since 2005, although its circulation continues, maintaining viral diversity but not undergoing antigenic drift. Most sequences belonged to subgenotype IA, closely related to Argentinean clinical sequences, but one belonged to proposed subgenotype IC, previously undetected in the country. Environmental surveillance might contribute to monitoring the single-dose vaccination schedule, representing not only strains causing disease but also the circulating population and the viral introductions.

Influence of overlapping genes on the evolution of human hepatitis B virus

The aim of this work was to analyse the influence of overlapping genes on the evolution of hepatitis B virus (HBV). A differential evolutionary behaviour among genetic regions and clinical status was found. Dissimilar levels of conservation of the different protein regions could derive from alternative mechanisms to maintain functionality. We propose that, in overlapping regions, selective constraints on one of the genes could drive the substitution process. This would allow protein conservation in one gene by synonymous substitutions while mechanisms of tolerance to the change operate in the overlapping gene (e.g. usage of amino acids with high-degeneracy codons, differential codon usage and replacement by physicochemically similar amino acids). In addition, differential selection pressure according to the HBeAg status was found in all genes, suggesting that the immune response could be one of the factors that would constrain viral replication by interacting with different HBV proteins during the HBeAg(-) stage.

High diversity of human polyomaviruses in environmental and clinical samples in Argentina: Detection of JC, BK, Merkel-cell, Malawi, and human 6 and 7 polyomaviruses

New human polyomaviruses have been recently described. The aim of this work was to detect and characterize human polyomaviruses circulating in Argentina by recovering viruses from environmental and sewage samples and evaluating their potential role as viral indicators of human waste contamination. Analysis was performed in a wider context including viruses from clinical samples from an immunocompromised population. River water and sewage samples were analyzed as a strategy to study the molecular epidemiology of viruses excreted by millions of people. Samples belonged to the Matanza-Riachuelo River (2005-2006: n=25 and 2012: n=20) and sewage from Buenos Aires city and suburbs (2011 and 2013: n=24). Viral detection was performed by PCR and the amplified viral genomes were characterized by phylogenetic analysis. Polyomaviruses were detected in 95.8% of sewage samples, identifying BKPyV (87.5%), JCPyV (83.3%), MCPyV (8.3%) and HPyV6 (8.3%). Besides, one sample collected in 2009 resulted positive for HPyV7. In 2005-2006, polyomaviruses were detected in 84.0% of river water samples, with the highest detection for MCPyV (52.0%), followed by BKPyV (44.0%), JCPyV (20.0%) and MWPyV (4.0%). In 2012, polyomaviruses were detected in 85.0% of river samples, finding JCPyV (85.0%), BKPyV (75.0%), MCPyV (25.0%) and HPyV6 (25.0%). Also, polyomaviruses, including JCPyV, BKPyV and MCPyV, were detected in 63.2% of urine samples from patients infected with HIV (n=19). Characterization indicated the coexistence of different genotypes and variants for each virus, particularly in sewage. MCPyV sequences (the only sequences from Argentina) formed a monophyletic group with the single sequence available for South America (French Guiana). The high level of detection and viral diversity found by environmental surveillance, which involved the characterization of viruses not previously described in South America, reinforces the usefulness of this approach to monitor viral contamination and describe the viral epidemiology in the general population.

Comparative analysis of hepatitis B virus genotype a molecular evolution in patients infected with HBV and in patients co-infected with HBV and HIV.

HIV infection has a significant impact on the natural progression of liver disease caused by infection with hepatitis B virus (HBV), but its role in the molecular evolution of HBV is unknown. It is difficult to study the molecular evolution of HBV longitudinally considering its genomic complexity, which implies the analysis of paired samples. This study aimed to analyze the difference in the evolutionary dynamics of HBV among patients with HIV and uninfected individuals. In this study, 17 patients infected chronically with HBV were recruited, 9 of them were co‐infected with HIV. Patients were HBe antigen‐positive and infected with HBV genotype A. Paired plasma samples were collected from each patient 3 years apart, and they were compared subsequently to each other. The HBV phylogenetic inference among isolates from patients infected with HBV and co‐infected with HBV and HIV tends to cluster separately. Likewise, when comparing the HBV evolutionary rate and genetic distances, values were higher in the former in both preC/C and S genomic regions. Intra‐host analyses of HBV isolates revealed high diversity and complexity of quasispecies among patients infected with HBV exhibiting high numbers of viral variants and genetic distance. In summary, after studying the HBV molecular evolution among isolates ascribed to genotype A at inter‐ and intra‐host levels, HBV exhibited low quasispecies complexity and diversity as well as low evolutionary rates in the presence of HIV co‐infection, suggesting that the co‐infection may have an impact on the HBV molecular evolution most likely from the weakened cellular immune response. J. Med. Virol. 84:562–569, 2012.

Identification of a new clade of hepatitis B virus genotype F.

Hepatitis B virus (HBV) is classified into eight main genotypes (A-H) and several subgenotypes. Here, three new genotype F complete genome sequences isolated from patients from Buenos Aires city are reported. The new sequences form a separate monophyletic group from the previously known subgenotype F4 strains. Based on results of phylogenetic, genetic distance and evolutionary analyses, the name F4b is proposed for these isolates and F4a for the formerly known as F4. The identification of new clusters allows deepening the knowledge about the diversification process and evolutionary history of HBV.