[Show abstract][Hide abstract] ABSTRACT: Background:
Chikungunya virus (CHIKV), an alphavirus and member of the Togaviridae family, is capable of causing severe febrile disease in humans. In December of 2013 the Asian Lineage of CHIKV spread from the Old World to the Americas, spreading rapidly throughout the New World. Given this new emergence in naïve populations we studied the viral genetic diversity present in infected individuals to understand how CHIKV may have evolved during this continuing outbreak.
We used deep-sequencing technologies coupled with well-established bioinformatics pipelines to characterize the minority variants and diversity present in CHIKV infected individuals from Guadeloupe and Martinique, two islands in the center of the epidemic. We observed changes in the consensus sequence as well as a diverse range of minority variants present at various levels in the population. Furthermore, we found that overall diversity was dramatically reduced after single passages in cell lines. Finally, we constructed an infectious clone from this outbreak and identified a novel 3' untranslated region (UTR) structure, not previously found in nature, that led to increased replication in insect cells.
Here we preformed an intrahost quasispecies analysis of the new CHIKV outbreak in the Caribbean. We identified novel variants present in infected individuals, as well as a new 3'UTR structure, suggesting that CHIKV has rapidly evolved in a short period of time once it entered this naïve population. These studies highlight the need to continue viral diversity surveillance over time as this epidemic evolves in order to understand the evolutionary potential of CHIKV.
[Show abstract][Hide abstract] ABSTRACT: Importance:
Replication, adaptation and evolution of RNA viruses rely in large part on their low-fidelity RNA-dependent RNA polymerase. Viruses artificially modified in their polymerase to decrease fidelity (mutator viruses) are attenuated in vivo, demonstrating the important role of fidelity in viral fitness. However, attenuation was solely attributed to the modification of the viral mutation rate and the accumulation of detrimental point mutations. In this work, we described an additional phenotype of mutator viruses: an increased recombination rate leading to defective interfering particle (DI) overproduction. Because DIs are known for their inhibitory effect on viral replication, our work suggests that fidelity variants may be attenuated in vivo via several mechanisms. This has important implications in the development of fidelity variants as live-attenuated vaccine strains.
Preview · Article · Dec 2015 · Journal of Virology
[Show abstract][Hide abstract] ABSTRACT: Author Summary
When RNA viruses replicate, they do so with a high rate of error; hence, their populations are not composed of a single genotype, but of a swarm of different, yet related, genomes. This mutant spectrum has been described as the viral quasispecies, and its composition has important consequences for evolution, adaptation and emergence. In this study, we analysed adaptation in fine detail thanks to the use of the deep sequencing, and we determined the adaptative pathway of a model RNA virus, Coxsackievirus B3, to a new environment, A549 cells. Our results demonstrate that adaptation occurred in response to a differential expression of the virus receptors in the new cellular environment, compared to the former. Our experiments and mathematical analyses established that the corresponding increase in fitness resulted from the selection and contribution of a group of genotypes, including low frequency variants, and not to the effect of a single, dominant genome. Our work underscores the importance of considering group effects when studying RNA virus biology and evolution.
[Show abstract][Hide abstract] ABSTRACT: Hepatitis C virus (HCV) has infected over 170 million people worldwide and creates a huge disease burden
due to chronic, progressive liver disease. HCV is a singlestranded, positive sense, RNA virus, member of the
Flaviviridae family. The high error rate of RNA-dependent RNA polymerase and the pressure exerted by the host immune system, has driven the evolution of HCV into 7 different genotypes and more than 67 subtypes. HCV evolves by means of different mechanisms of genetic variation. On the one hand, its high mutation rates
generate the production of a large number of different but closely related viral variants during infection, usually referred to as a quasispecies. The great quasispecies variability of HCV has also therapeutic implications since the continuous generation and selection of resistant or fitter variants within the quasispecies spectrum might allow viruses to escape control by antiviral drugs. On the other hand HCV exploits recombination to ensure its survival. This enormous viral diversity together with some host factors has made it difficult to control viral dispersal. Current treatment options involve pegylated interferon-α and ribavirin as dual therapy or in combination with a direct-acting antiviral drug, depending on the country. Despite all the efforts put into antiviral therapy studies, eradication of the virus or the development of a preventive vaccine has been unsuccessful so far. This review focuses on current available data reported to date on the genetic
mechanisms driving the molecular evolution of HCV populations and its relation with the antiviral therapies
designed to control HCV infection.
Full-text · Article · Apr 2015 · World Journal of Hepatology
[Show abstract][Hide abstract] ABSTRACT: Molecular characterization of circulating influenza A viruses (IAV) in all regions of the world is essential to detect mutations potentially involved in increased virulence, anti-viral resistance and immune escape. In order to gain insight into these matters, a phylogenetic analysis of the neuraminidase (NA) gene of 146 pandemic H1N1 (H1N1pdm) influenza A virus strains isolated in Argentina, Brazil, Chile, Paraguay, Peru and Uruguay from 2009 to 2013 was performed. Comparison of vaccine strain A/California/7/2009 included in the influenza vaccine recommended for the Southern hemisphere from 2010 through 2013 influenza seasons and strains isolated in South America revealed several amino acid substitutions. Mapping of these substitutions revealed that most of them are located at the surface of the protein and do not interfere with the active site. 3.4% of the strains enrolled in these studies carried the H275Y substitution that confers resistance to oseltamivir. Strains isolated in South America differ from vaccine in two predicted B-cell epitope regions present at positions 102-103 and 351-352 of the NA protein. Moreover, vaccine and strains isolated in Paraguay differ also in an epitope present at position 229. These differences among strains isolated in South America and vaccine strain suggests that these epitopes may not be present in strains isolated in this region. A potential new N-linked glycosylation site was observed in the NA protein of an H1N1pdm IAV strain isolated in Brazil. The results of these studies revealed several genetic and antigenic differences in the NA of H1N1pdm IAV among vaccine and strains circulating in South America. All these findings contribute to our understanding of the course of genetic and antigenic evolution of H1N1pdm IAV populations circulating in the South American region and, consequently, contribute to the study and selection of future and more appropriate vaccines and anti-viral drugs.
[Show abstract][Hide abstract] ABSTRACT: The human genome contains a large number of endogenous retroviruses (HERVs). Their reactivation has frequently been observed in patients with cancer. Considering their role in the carcinogenesis process, we aimed to study the possible relationship between HERVs gene expression and Chronic Lymphocytic Leukemia (CLL). We focused on two viral genes: gag and np9, the latter presumably an oncogene. We found that the transcriptional activity of HERV-K np9 gene was greater in CLL patients than in healthy donors. However, gag expression was not significantly increased.
These findings suggest a noteworthy relationship between CLL disease and HERV-K np9 expression.
Full-text · Article · Jul 2014 · Leukemia Research Reports
[Show abstract][Hide abstract] ABSTRACT: It is widely accepted that the majority of cancers result from multiple cellular events leading to malignancy after a prolonged period of clinical latency, and that the immune system plays a critical role in the control of cancer progression. Bovine leukemia virus (BLV) is an oncogenic member of the Retroviridae family. Complete genomic sequences of BLV strains isolated from peripheral blood mononuclear cells (PBMC) from cattle have been previously reported. However, a detailed characterization of the complete genome of BLV strains directly isolated from bovine tumors is much needed in order to contribute to the understanding of the mechanisms of leukemogenesis induced by BLV in cattle. In this study, we performed a molecular characterization of BLV complete genomes from bovine B-cell lymphosarcoma isolates. A nucleotide substitution was found in the glucocorticoid response element (GRE) site of the 5[prime] long terminal repeat (5[prime]LTR) of the BLV isolates. All amino acid substitutions in Tax previously found to be related to stimulate high transcriptional activity of 5[prime]LTR were not found in these studies. Amino acid substitutions were found in the nucleocapsid, gp51 and G4 proteins. Premature stop-codons in R3 were observed. Few mutations or amino acid substitutions may be needed to allow BLV provirus to achieve silencing. Substitutions that favor suppression of viral expression in malignant B cells might be a strategy to circumvent effective immune attack.
Full-text · Article · Mar 2013 · Veterinary Research
[Show abstract][Hide abstract] ABSTRACT: West Nile virus (WNV) is a member of the family Flaviviridae and its genome consists of an 11-kb single-stranded, positive-sense RNA. WNV is maintained in an enzootic cycle between mosquitoes and birds, but can also infect and cause disease in horses and humans, which serve as incidental dead-end hosts. Understanding the extent and causes of biases in codon usage is essential to the comprehension of viral evolution. In this study, we performed a comprehensive analysis of 449 WNV strains, for which complete genome sequences are available. Effective number of codons (ENC) indicates that the overall codon usage among WNV strains is only slightly biased. Codon adaptation index (CAI) values found for WNV genes are different from the CAI values found for human genes. The relative synonymous codon usage among WNV strains isolated from birds, equines, humans and mosquitoes are roughly similar and are influenced by the relative dinucleotide frequencies. Taking together, the results of this work suggest that WNV genomic biases are the result of the evolution of genome composition, the need to escape the antiviral cell responses and a dynamic process of mutation and selection to re-adapt its codon usage to different environments.
Full-text · Article · Jan 2013 · Infection, genetics and evolution: journal of molecular epidemiology and evolutionary genetics in infectious diseases
[Show abstract][Hide abstract] ABSTRACT: Background
Influenza A virus (IAV) is a member of the family Orthomyxoviridae and contains eight segments of a single-stranded RNA genome with negative polarity. The first influenza pandemic of this century was declared in April of 2009, with the emergence of a novel H1N1 IAV strain (H1N1pdm) in Mexico and USA. Understanding the extent and causes of biases in codon usage is essential to the understanding of viral evolution. A comprehensive study to investigate the effect of selection pressure imposed by the human host on the codon usage of an emerging, pandemic IAV strain and the trends in viral codon usage involved over the pandemic time period is much needed.
We performed a comprehensive codon usage analysis of 310 IAV strains from the pandemic of 2009. Highly biased codon usage for Ala, Arg, Pro, Thr and Ser were found. Codon usage is strongly influenced by underlying biases in base composition. When correspondence analysis (COA) on relative synonymous codon usage (RSCU) is applied, the distribution of IAV ORFs in the plane defined by the first two major dimensional factors showed that different strains are located at different places, suggesting that IAV codon usage also reflects an evolutionary process.
A general association between codon usage bias, base composition and poor adaptation of the virus to the respective host tRNA pool, suggests that mutational pressure is the main force shaping H1N1 pdm IAV codon usage. A dynamic process is observed in the variation of codon usage of the strains enrolled in these studies. These results suggest a balance of mutational bias and natural selection, which allow the virus to explore and re-adapt its codon usage to different environments. Recoding of IAV taking into account codon bias, base composition and adaptation to host tRNA may provide important clues to develop new and appropriate vaccines.
[Show abstract][Hide abstract] ABSTRACT: The first influenza pandemic of this century was declared in April of 2009, with the emergence of a novel H1N1 influenza A virus strain (H1N1pdm). Understanding the evolution of H1N1pdm populations within the South American region is essential for studying global diversification, emergence, resistance and vaccine efficacy. In order to gain insight into these matters, we have performed a Bayesian coalescent Markov Chain Monte Carlo analysis of hemagglutinin (HA) and neuraminidase (NA) gene sequences of all available and comparable HA and NA sequences obtained from H1N1pdm IAV circulating in the South American region. High evolutionary rates and fast population growths characterize the population dynamics of H1N1pdm strains in this region of the world. A significant contribution of first codon position to the mean evolutionary rate was found for both genes studied, revealing a high contribution of non-synonymous substitutions to the mean substitution rate. In the 178days period covered by these studies, substitutions in all HA epitope regions can be observed. HA substitutions D239G/N and Q310H have been observed only in Brazilian patients. While substitution D239G/N is not particularly associated to a specific genetic lineage, all strains bearing substitution Q310H were assigned to clade 6, suggesting a founder effect. None of the substitutions found in the NA proteins of H1N1pdm strains isolated in South America appears sufficiently close to affect the drug binding pocket for the three NA inhibitor antivirals tested. A more detailed analysis of NA proteins revealed epitope differences among 2010 vaccine and H1N1pdm IAV strains circulating in the South American region.
[Show abstract][Hide abstract] ABSTRACT: Coxsackie B viruses (CVB) are associated with serious illnesses in humans. In this study, the patterns of synonymous codon usage in CVB have been studied through multivariate statistical methods. Effective number of codons (ENC) indicates that the overall extent of codon usage bias in CVB is not significant. The relative dinucleotide abundances suggest that codon usage bias in CVB genomes is influenced by underlying biases of dinucleotide frequencies. The distribution of CVB ORFs along the plane defined by the first two axes of correspondence analysis (COA) showed that different genotypes, as well as strains known to infect different cell types, are located at different places in the plane suggesting that CVB codon usage is reflecting an evolutionary process. The results of these studies suggest that CVB genomic biases are the result of co-evolution of translation adaptation to different cell environments and probably the need to escape anti-viral cell defenses.
[Show abstract][Hide abstract] ABSTRACT: The Rotavirus genus belongs to the family Reoviridae and its genome consist of 11 segments of double-stranded RNA. Group A rotaviruses (RV-A) are the main etiological agent of acute viral gastroenteritis in infants and young children worldwide. Understanding the extent and causes of biases in codon usage is essential to the understanding of viral evolution. However, the factors shaping synonymous codon usage bias and nucleotide composition in human RV-A are currently unknown. In order to gain insight into these matters, we analyzed the codon usage and base composition constraints on the two genes that codify the two outer capsid proteins (VP4 [VP8*] and VP7) of 58 PG2 RV-A strains isolated in Brazil and investigated the possible key evolutionary determinants of codon usage bias. The results of these studies revealed that the frequencies of codon usage in both RV-A proteins studied are significantly different than the ones used by human cells. In order to observe if similar trends of codon usage are found when RV-A complete genomes are considered, we compare these results with results found using a dataset of 10 reference strains for whom the complete codes of the 11 segments are known. Similar results were obtained using capsid proteins or complete genomes. The general correlations found between the position of each sequence on the first axis generated by correspondence analysis and the relative dinucleotide abundances indicate that codon usage in RV-A can also be strongly influenced by underlying biases in dinucleotide frequencies. CpG and GpC containing codons are markedly suppressed. Thus, the results of this study suggest that RV-A genomic biases are the result of the evolution of genome composition in relation to host adaptation and the ability to escape antiviral cell responses.
Full-text · Article · Apr 2011 · Infection, genetics and evolution: journal of molecular epidemiology and evolutionary genetics in infectious diseases
[Show abstract][Hide abstract] ABSTRACT: The subtype diversity of the hepatitis C virus (HCV) genotypes is unknown in Venezuela.
Partial sequencing of the NS5B region was performed in 310 isolates circulating in patients from 1995 to 2007. In the samples collected between 2005 and 2007, HCV genotype 1 (G1) was the most common genotype (63%), composed as expected of mainly G1a and G1b. G2 was the second most common genotype (33%), being G2a almost absent and G2j the most frequent subtype. Sequence analysis of the core region confirmed the subtype assignment performed within the NS5b region in 63 isolates. The complete genome sequence of G2j was obtained. G2j has been described in France, Canada and Burkina Fasso, but it was not found in Martinique, where several subtypes of G2 circulate in the general population. Bayesian coalescence analysis indicated a most recent common ancestor (MRCA) of G2j around 1785, before the introduction of G1b (1869) and G1a (1922). While HCV G1a and G1b experienced a growth reduction since 1990, coincident with the time when blood testing was implemented in Venezuela, HCV G2j did not seem to reach growth equilibrium during this period.
Assuming the introduction of G2j from Africa during the slave trade, the high frequency of G2j found in Venezuela could suggest: 1- the introduction of African ethnic groups different from the ones introduced to Martinique or 2- the occurrence of a founder effect. This study represents an in-depth analysis of the subtype diversity of HCV in Venezuela, which is still unexplored in the Americas and deserves further studies.
[Show abstract][Hide abstract] ABSTRACT: Dengue virus (DENV) is a member of the genus Flavivirus of the family Flaviviridae. DENV are comprised of four distinct serotypes (DENV-1 through DENV-4) and each serotype can be divided in different genotypes. Currently, there is a dramatic emergence of DENV-3 genotype III in Latin America. Nevertheless, we still have an incomplete understanding of the evolutionary forces underlying the evolution of this genotype in this region of the world. In order to gain insight into the degree of genetic variability, rates and patterns of evolution of this genotype in Venezuela and the South American region, phylogenetic analysis, based on a large number (n = 119) of envelope gene sequences from DENV-3 genotype III strains isolated in Venezuela from 2001 to 2008, were performed.
Phylogenetic analysis revealed an in situ evolution of DENV-3 genotype III following its introduction in the Latin American region, where three different genetic clusters (A to C) can be observed among the DENV-3 genotype III strains circulating in this region. Bayesian coalescent inference analyses revealed an evolutionary rate of 8.48 x 10⁻⁴ substitutions/site/year (s/s/y) for strains of cluster A, composed entirely of strains isolated in Venezuela. Amino acid substitution at position 329 of domain III of the E protein (A→V) was found in almost all E proteins from Cluster A strains.
A significant evolutionary change between DENV-3 genotype III strains that circulated in the initial years of the introduction in the continent and strains isolated in the Latin American region in recent years was observed. The presence of DENV-3 genotype III strains belonging to different clusters was observed in Venezuela, revealing several introduction events into this country. The evolutionary rate found for Cluster A strains circulating in Venezuela is similar to the others previously established for this genotype in other regions of the world. This suggests a lack of correlation among DENV genotype III substitution rate and ecological pattern of virus spread.
[Show abstract][Hide abstract] ABSTRACT: The first influenza pandemic of this century was declared in April of 2009, with the emergence of a novel H1N1 influenza A virus strain (H1N1pdm). Understanding the evolution of H1N1pdm strains within the South American region is essential for studying global diversification, emergence and resistance, as well as determining vaccine efficacy. In order to gain insight into these matters, phylogenetic analysis was performed using 29 hemagglutinin (HA) gene sequences from H1N1pdm strains isolated in South America. The results of these studies revealed that clade 7 was the dominant H1N1pdm lineage in South America. None of the strains isolated in South America clustered together with the 2010 H1 vaccine strain. Amino acid substitutions P100S, S220T and I338V were found in almost all HAs of South American H1N1pdm strains.
Full-text · Article · Oct 2010 · Archives of Virology
[Show abstract][Hide abstract] ABSTRACT: Group A rotavirus (RV-A) genotype PG9 has emerged as one of the leading causes of gastroenteritis in children worldwide and currently is recognized as one of the five most common genotypes detected in humans. High intragenotype diversity in G9 RV-A has been observed, and nowadays, based on the genetic variability of the VP7 gene, six different phylogenetic lineages and eleven sublineages were described.
To study the degree of genetic variation and evolution of Brazilian PG9 RV-A strains.
Phylogenetic analysis of 19 PG9 RV-A strains isolated from 2004 to 2007 in five different Brazilian states was conducted using the NSP1, NSP3, NSP5, VP4 and VP7 genes. For the VP4 and VP7 genes, 3D protein structure predictions were generated to analyze the spatial distribution of amino acid substitutions observed in Brazilian strains.
Based on the phylogenetic analyses, all Brazilian strains clustered within lineage G9-III and P-3 for VP7 and VP4, respectively, and were classified as genotype A1, T1 and H1 for the NSP1, NSP3 and NSP5 genes, respectively. Interestingly, all the strains isolated in Acre State (Northern Brazil) formed a closely related cluster clearly separated from the other Brazilian and prototype strains with regard to the five genes studied. Unique amino acid substitutions were observed in Acre strains in comparison with the prototype and Brazilian strains.
Inclusion of Acre strains in the phylogenetic analysis revealed the presence of a novel genetic variant and demonstrated a diversification of PG9 rotaviruses in Brazil.
Full-text · Article · Apr 2010 · Journal of clinical virology: the official publication of the Pan American Society for Clinical Virology