Antiviral activity of myricetin glycosylated compounds isolated from Marcetia taxifolia against chikungunya virus.

The chikungunya virus (CHIKV) has produced epidemic outbreaks of significant public health impact. The clinical symptoms of this disease are fever, polyarthralgia, and skin rash, generally self-limiting, although patients may develop a chronic disabling condition or suffer lethal complications. Unfortunately, there is no specific treatment or vaccine available. Thus, the search for effective therapies to control CHIKV infection is an urgent need. This study evaluated the antiviral activity of flavonoids isolated from Marcetia taxifolia by in vitro and in silico analysis. Cytotoxicity of compounds was determined by MTT assay and viral load was assessed in cell substrates supernatants by plaque-forming and RT-qPCR assays. Selected molecules were analyzed by molecular docking assays. Myricetin 3-rhamnoside (MR) and myricetin 3-(6-rhamnosylgalactoside) (MRG) were tested for antiviral assays and analyzed by the TCID50 method and RT-qPCR. MR exhibited dose-dependent antiviral activity, reducing viral titer at concentrations of 150-18.8 µg/mL by at least 1-log. Similarly, MRG showed a significant decrease in viral titer at concentrations of 37.5, 9.4, and 2.3 µg/mL. RT-qPCR analysis also displayed a substantial reduction of CHIKV RNA for both flavonoids. Furthermore, molecular docking of the selected flavonoids proposed the nsP3 macrodomain as a possible target of action. Our study reveals that MR and MRG could be considered promising anti-CHIKV therapeutic agents. Molecular modeling studies showed MR and MRG ligands with a high affinity for the N-terminal region of the nsP3 macrodomain, postulating them as a potential target of action for the CHIKV control.

Dissemination of the Omicron Variant and Its Sub-Lineages among Residents and Travelers in Its First Year of Emergence in Venezuela.

The emergence of the SARS-CoV-2 Variant of Concern (VOC), Omicron, has been characterized by an explosive number of cases in almost every part of the world. The dissemination of different sub-lineages and recombinant genomes also led to several posterior waves in many countries. The circulation of this VOC and its major sub-lineages (BA.1 to BA.5) was monitored in community cases and in international travelers returning to Venezuela by a rapid partial sequencing method. The specific sub-lineage assignment was performed by complete genome sequencing. Epidemic waves of SARS-CoV-2 cases were observed among international travelers during 2022, a situation not seen before December 2021. The succession of the Omicron VOC sub-lineages BA.1 to BA.5 occurred sequentially, except for BA.3, which was almost not detected. However, the sub-lineages generally circulated two months earlier in international travelers than in community cases. The diversity of Omicron sub-lineages found in international travelers was related to the one found in the USA, consistent with the most frequent destination of international travel from Venezuela this year. These differences are compatible with the delay observed sometimes in Latin American countries in the circulation of the different lineages of the Omicron VOC. Once the sub-lineages were introduced in the country, community transmission was responsible for generating a characteristic distribution of them, with a predominance of sub-lineages not necessarily similar to the one observed in travelers or neighboring countries.

Web-tools for the genomic analysis of the 2022 Monkeypox virus global outbreak / Herramientas web para el análisis genómico del virus de la viruela símica durante el brote mundial de 2022

Abstract The resources and platforms available on the internet for collecting and sharing information and performing genomic sequence analysis have made it possible to follow closely the evolution the evolution of SARS-CoV-2. However, the current monkeypox outbreak in the world brings us back to the need to use these resources to appraise the extent of this outbreak. The objective of this work was an analysis of the information presented so far in the genomic database GISAID EpiPox™, using various tools available on the web. The results indicate that the monkeypox outbreak is referred as MPXV clade II B.1 lineage and sub-lineages, isolated from male patients mainly from the European and American continents. In the current scenario, the access to genomic sequences, epidemiological information, and tools available to the scientific community is of great importance for global public health in order to follow the evolution of pathogens.

Resumen Los recursos y plataformas disponibles en Internet para recopilar, compartir información y realizar análisis de secuencias genómicas han permitido seguir de cerca la evolución del SARS-CoV-2. El actual brote global de viruela del mono en el mundo, requiere de nuevo utilizar estos recursos para conocer el alcance de este brote. El objetivo de este trabajo fue un análisis de la información presentada hasta el momento en la base de datos genómica EpiPox™ de GISAID, utilizando diversas herramientas disponibles en la web. Los resultados indican que el brote de la viruela del mono o símica está referido al linaje y sub-linajes B.1 del clado II de MPXV, aislado principalmente de pacientes hombres de Europa y América. En el escenario actual, el acceso a las secuencias genómicas, la información epidemiológica, y las herramientas disponibles para la comunidad científica son de gran importancia para la salud pública mundial con el fin de seguir la evolución de los patógenos.

Sub-lineages of the Omicron variant of SARS-CoV-2: characteristic mutations and their relation to epidemiological behavior / Sub-linajes de la variante Ómicron del SARS-CoV-2: mutaciones características y su relación con el comportamiento epidemiológico

Abstract By the end of 2021, the Omicron variant of SARS-CoV-2, the coronavirus responsible for COVID-19, emerges, causing immediate concern, due to the explosive increase in cases in South Africa and a large number of mutations. This study describes the characteristic mutations of the Omicron variant in the Spike protein, and the behavior of the successive epidemic waves associated to the sub-lineages throughout the world. The mutations in the Spike protein described are related to the virus ability to evade the protection elicited by current vaccines, as well as with possible reduced susceptibility to host proteases for priming of the fusion process, and how this might be related to changes in tropism, a replication enhanced in nasal epithelial cells, and reduced in pulmonary tissue; traits probably associated with the apparent reduced severity of Omicron compared to other variants.

Resumen A finales de 2021 surge la variante Omicron del SARS-CoV-2, el coronavirus responsable de la COVID-19, causando preocupación inmediata, debido al aumento explosivo de casos en Suráfrica, y a su gran cantidad de mutaciones. Este estudio describe las mutaciones características de la variante Ómicron en la proteína de la Espiga (S) y el comportamiento de las sucesivas olas epidémicas asociadas a la circulación de sus sub-linajes en todo el mundo. Las mutaciones en la proteína S descritas están relacionadas con su capacidad para evadir la protección provocada por las vacunas actuales, así como su posible susceptibilidad reducida a las proteasas del hospedero para la preparación del proceso de fusión. Se infiere cómo esto podría estar relacionado con su cambio en el tropismo, con una replicación mayor en las células epiteliales nasales y menor en el tejido pulmonar, rasgos probablemente asociados a su aparente menor gravedad en comparación con otras variantes.

Description of a One-Year Succession of Variants of Interest and Concern of SARS-CoV-2 in Venezuela.

Some of the lineages of SARS-CoV-2, the new coronavirus responsible for COVID-19, exhibit higher transmissibility or partial resistance to antibody-mediated neutralization and were designated by WHO as Variants of Interests (VOIs) or Concern (VOCs). The aim of this study was to monitor the dissemination of VOIs and VOCs in Venezuela from March 2021 to February 2022. A 614 nt genomic fragment was sequenced for the detection of some relevant mutations of these variants. Their presence was confirmed by complete genome sequencing, with a correlation higher than 99% between both methodologies. After the introduction of the Gamma VOC since the beginning of the year 2021, the variants Alpha VOC and Lambda VOI were detected as early as March 2021, at a very low frequency. In contrast, the Mu VOI, detected in May 2021, was able to circulate throughout the country. After the detection of the Delta VOC in June 2021, it became the predominant circulating variant. With the arrival of the Omicron VOC in December, this variant was able to displace the Delta one in less than one month.

Detection of the Omicron variant of SARS-CoV-2 in international travelers returning to Venezuela.

BACKGROUND: By the end of 2021, the SARS-CoV-2 Variant of Concern (VOC) Delta was predominant in most of the world. At the end of November, the Omicron variant was first detected in South Africa. This variant was immediately classified as VOC, due to the explosive increase of cases in South Africa, and the great number of mutations exhibited by this new lineage. Since then, Omicron VOC displaced Delta one in almost every country. Venezuela implemented in May 2021 molecular testing of all the passengers arriving at Venezuelan airports. METHODS: In this study, we analyzed the presence of variants of SARS-CoV-2 in those positive samples, by sequencing a small fragment of the Spike genomic region. RESULTS: The Omicron variant was found in passengers arriving to Venezuela from the beginning of December. Complete genome analysis confirmed the presence of the Omicron VOC. The detection of this VOC coincided with an unprecedented increase in the frequency of passengers with positive nucleic acid testing. CONCLUSIONS: Genomic surveillance of samples for international travelers returning to Venezuela allowed us to rapidly detect the introduction of the Omicron variant in the country.

Detection of the Omicron variant of SARSCoV-2 by restriction analysis targeting the mutations K417N and N440K of the spike protein / Detección de la Variante Ómicron del SARS-CoV-2 por análisis de restricción de dos mutaciones de la proteína de la espiga

Abstract By the end of 2021, the Omicron variant of concern (VOC) emerges in South Africa. This variant caused immediate concern, due to the explosive increase in cases associated with it and the large number of mutations it exhibits. In this study, the restriction sites that allow detecting the mutations K417N and N440K in the Spike gene are described. This analysis allows us to propose a rapid method for the identification of cases infected with the Omicron variant. We show that the proposed methodology can contribute to provide more information on the prevalence and rapid detection of cases of this new VOC.

Resumen Para finales de 2021 surge la variante de preocupación (VOC por sus siglas en inglés) Ómicron en Sudáfrica. Esta variante causó de forma inmediata preocupación, debido al aumento explosivo de casos asociados a ella y al gran número de mutaciones que exhibe. En este estudio, se describen los sitios de restricción que permiten detectar dos de estas mutaciones en el gen de la espiga, las mutaciones K417N y N440K. Este análisis permite proponer un método rápido para la identificación de casos infectados con la variante Ómicron. Mostramos que la metodología propuesta puede contribuir a proporcionar más información sobre la prevalencia y a detectar rápidamente los casos de esta nueva VOC.

First Case of Monkeypox in Venezuela: Partial Complete Genome Sequence Allowed Its Grouping into the West African Clade II.

The ongoing epidemic of monkeypox virus (MPXV) infection has already reached more than 50,000 persons worldwide until the end of August 2022. We report the first case detected in Venezuela. The patient reported traveling from Spain and contact with friends tested positive for MPXV after his return. Partial complete genome phylogenetic analysis allowed to group the isolate within the clade II of MPXV, the major one circulating worldwide. No other case of MPXV has been detected until the end of August 2022 in the country, although the presence of undiagnosed cases due to the fear of stigmatization cannot be ruled out.

Introduction and rapid dissemination of SARS-CoV-2 Gamma Variant of Concern in Venezuela.

In less than two years since SARS-CoV-2 emerged, the new coronavirus responsible for COVID-19, has accumulated a great number of mutations. Many of these mutations are located in the Spike protein and some of them confer to the virus higher transmissibility or partial resistance to antibody mediated neutralization. Viral variants with such confirmed abilities are designated by WHO as Variants of Concern (VOCs). The aim of this study was to monitor the introduction of variants and VOCs in Venezuela. A small fragment of the viral genome was sequenced for the detection of the most relevant mutations found in VOCs. This approach allowed the detection of Gamma VOC. Its presence was confirmed by complete genome sequencing. The Gamma VOC was detected in Venezuela since January 2021, and in March 2021 was predominant in the East and Central side of the country, representing more than 95% of cases sequenced in all the country in April-May 2021. In addition to the Gamma VOC, other isolates carrying the mutation E484K were also detected. The frequency of this mutation has been increasing worldwide, as shown in a survey of sequences carrying E484K mutation in GISAID, and was detected in Venezuela in many probable cases of reinfection. Complete genome sequencing of these cases allowed us to identify E484K mutation in association with Gamma VOC and other lineages. In conclusion, the strategy adopted in this study is suitable for genomic surveillance of variants for countries lacking robust genome sequencing capacities. In the period studied, Gamma VOC seems to have rapidly become the dominant variant throughout the country.

Mutations in the SARS-CoV-2 spike protein modulate the virus affinity to the human ACE2 receptor, an in silico analysis.

The increasing number of SARS-CoV-2 variants associated with highly transmissible phenotypes is a health-public concern in the current pandemic scenario. Herein, we developed a comprehensive in silico analysis of the changes occurring upon mutations in the viral spike. We focused on mutants located in the receptor-binding domain of the viral spike protein and analyzed whether these mutants modulate the interaction with the human host receptor angiotensin-converting enzyme II (ACE2). Thirty-two highly prevalent mutants were retrieved from the GISAID database, and their structural models were built using the SWISS-Model server. The stabilization effect for each mutation was assessed by the DUET and DeepDGG software. By applying molecular docking using both Z-Dock and Haddock software we found that multiple mutations, including A475V, V455E, V445L, and V445I, resulted in the higher binding free energy as compared to the wild type (WT) spike protein, thus had a destabilizing effect on the binding to ACE2. On the other hand, several mutants, including the most prevalent N501Y and B.1.1.7 variants, as well as the K444R, L455F, Q493R, and Y505W variants exhibited lower binding free energy as compared to the WT spike. These mutants showed an increased number of electrostatic interactions with ACE2 than the WT spike protein, and they changed the interaction pattern of the neighboring residues. Together, the results presented in this study contribute to a better understanding of the changes in the interaction between SARS-CoV-2 and the human host ACE2 receptor associated with point mutations in the viral spike protein.

SARS-CoV-2 genetic diversity in Venezuela: Predominance of D614G variants and analysis of one outbreak.

SARS-CoV-2 is the new coronavirus responsible for COVID-19 disease. The first two cases of COVID-19 were detected in Venezuela on March 13, 2020. The aim of this study was the genetic characterization of Venezuelan SARS-CoV-2 isolates. A total of 7 full SARS-CoV-2 genome sequences were obtained by Sanger sequencing, from patients of different regions of Venezuela, mainly from the beginning of the epidemic. Ten out of 11 isolates (6 complete genomes and 4 partial spike genomic regions) belonged to lineage B, bearing the D614G mutation in the Spike protein. Isolates from the first outbreak that occurred in the Margarita Island harbored an in-frame deletion in its sequence, without amino acids 83-85 of the NSP1 of the ORF1. The search for deletions in 48,635 sequences showed that the NSP1 gene exhibit the highest frequency of deletions along the whole genome. Structural analysis suggests a change in the N-terminal domain with the presence of this deletion. In contrast, isolates circulating later in this island lacked the deletion, suggesting new introductions to the island after this first outbreak. In conclusion, a high diversity of SARS-CoV-2 isolates were found circulating in Venezuela, with predominance of the D614G mutation. The first small outbreak in Margarita Island seemed to be associated with a strain carrying a small deletion in the NSP1 protein, but these isolates do not seem to be responsible for the larger outbreak which started in July.

Understanding Severe Acute Respiratory Syndrome Coronavirus 2 Replication to Design Efficient Drug Combination Therapies.

BACKGROUND: The emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and its disease CO-VID-19 has strongly encouraged the search for antiviral compounds. Most of the evaluated drugs against SARS-CoV-2 derive from drug repurposing of Food and Drug Administration-approved molecules. These drugs have as target three major processes: (1) early stages of virus-cell interaction, (2) viral proteases, and (3) the viral RNA-dependent RNA polymerase. SUMMARY: This review focused on the basic principles of virology and pharmacology to understand the importance of early stages of virus-cell interaction as therapeutic targets and other main processes vital for SARS-CoV-2 replication. Furthermore, we focused on describing the main targets associated with SARS-CoV-2 antiviral therapy and the rationale of drug combinations for efficiently suppressing viral replication. Key Messages: We hypothesized that blocking of both entry mechanisms could allow a more effective antiviral effect compared to the partial results obtained with chloroquine or its derivatives alone. This approach, already used to achieve an antiviral effect higher than that offered by every single drug administered separately, has been successfully applied in several viral infections such as HIV and HCV. This review will contribute to expanding the perception of the possible therapeutic targets in SARS-CoV-2 infection and highlight the benefits of using combination therapies.

Decreasing prevalence of Hepatitis B and absence of Hepatitis C Virus infection in the Warao indigenous population of Venezuela.

Prevalence and molecular epidemiology studies for hepatitis B (HBV) and C (HCV) virus are scarce in Warao Amerindians from Venezuela, where an epidemic of human immunodeficiency virus type 1 (HIV-1) has recently been documented. To carry out a molecular epidemiology analysis of hepatitis B (HBV) and C (HCV) virus in Warao individuals from the Delta Amacuro State of Venezuela. A total of 548 sera were tested for serological and molecular markers for HBV and HCV. The prevalence of active infection (presence of HBV surface antigen, HBsAg), exposure to HBV (presence of Antibody to HBV core antigen, anti-HBc) and anti-HCV, was 1.8%, 13% and 0% respectively. HBV exposure was significantly lower in men below 18 years old and also lower than rates previously reported in other Amerindian communities from Venezuela. Thirty one percent (31%, 25/80) of individuals without evidence of HBV infection exhibited anti-HBs titer ≥ 10U.I / ml, being significantly more frequent in individuals younger than 20 years. A higher HBV exposure was observed among HIV-1 positive individuals (33% vs 11%, p <0.005). A high prevalence of occult HBV infection was also observed (5.6%, 11/195). Phylogenetic analysis of S gene and complete HBV genomes showed that F3 is the only circulating subgenotype, different from the F2 subgenotype found in 1991 in this population. These results suggest a recent introduction of subgenotype F3, with a low divergence among the isolates. These results highlight the importance of molecular epidemiology studies for viral control, and support the effectiveness of vaccination in reducing transmission of HBV.

The role of the glycosyl moiety of myricetin derivatives in anti-HIV-1 activity in vitro.

BACKGROUND: Plant extracts are sources of valuable compounds with biological activity, especially for the anti-proliferative activity against pathogens or tumor cells. Myricetin is a flavonoid found in several plants that has been described as an inhibitor of Human immunodeficiency virus type 1 (HIV-1) through its action against the HIV reverse transcriptase, but myricetin derivatives have not been fully studied. The aim of this study was to evaluate the anti-HIV-1 activity of glycosylated metabolites obtained from Marcetia taxifolia and derived from myricetin: myricetin rhamnoside and myricetin 3-(6-rhamnosylgalactoside). METHODS: Compounds were obtained from organic extracts by maceration of aerial parts of M. taxifolia. All biological assays were performed in the MT4 cell line. Antiviral activity was measured as inhibition of p24 and reverse transcriptase with a fluorescent assay. RESULTS: Both flavonoids have antiviral activity in vitro, with an EC50 of 120 µM for myricetin 3-rhamnoside (MR) and 45 µM for myricetin 3-(6-rhamnosylgalactoside) (MRG), both significantly lower than the EC50 of myricetin (230 µM). Although both compounds inhibited the reverse transcriptase activity, with an IC50 of 10.6 µM for MR and 13.8 µM for MRG, myricetin was the most potent, with an IC50 of 7.6 µM, and an inhibition greater than 80%. Molecular docking approach showed correlation between the free energy of binding with the assays of enzyme inhibition. CONCLUSIONS: The results suggest that glycosylated moiety might enhance the anti-HIV-1 activity of myricetin, probably by favoring the internalization of the flavonoid into the cell. The inhibition of the HIV-1 reverse transcriptase is likely responsible for the antiviral activity.

The Evolving HIV-1 Epidemic in Warao Amerindians Is Dominated by an Extremely High Frequency of CXCR4-Utilizing Strains.

We previously reported a high prevalence of HIV-1 infection in Warao Amerindians from Venezuela due to the rapid spread of a single B subtype strain. In this study we evaluated the coreceptor use of the HIV-1 strains infecting this Amerindian community. Sequences of the HIV-1 V3 loop from 56 plasma samples were genotyped for coreceptor use. An extremely high frequency of CXCR4 strains was found among HIV-1-infecting Waraos (47/49, 96%), compared to HIV-1 strains infecting the non-Amerindian Venezuelan population (35/79, 44%, p < 0.00001). Evolutionary analysis showed that a significant number of infections occurred between 1 and 12 months before collection and that a great proportion (50-70%) of HIV-1 transmissions occurred within the very early phase of infection (≤12 months). This is consistent with an initial infection dominated by an X4 strain or a very rapid selection of X4 variants after infection. This Amerindian population also exhibits the highest prevalence of tuberculosis in Venezuela, being synergistically bad prognostic factors for the evolution of morbidity and mortality in this vulnerable population.

Genetic diversity of hepatitis B virus and hepatitis C virus in human immunodeficiency virus type 1-co-infected patients from Venezuela.

The aim of this study was to evaluate the prevalence and genetic diversity of hepatitis B virus (HBV) and hepatitis C virus (HCV) in human immunodeficiency virus type 1 (HIV-1)-co-infected Venezuelan patients. The prevalence of HBV and HCV markers of infection in HIV-1 patients was 14% for anti-hepatitis B core antigen, 3% for hepatitis B surface antigen and 0.7% for anti-HCV, respectively. HBV prevalence was higher than HCV, as expected for a country where sexual intercourse, not intravenous drug use, is the main mode of HIV-1 transmission. The HCV genotype distribution in HIV-1-co-infected patients was similar to that obtained in HCV-mono-infected patients, but genotype 1a was more frequent in HIV-1-infected patients. The HBV genotype distribution exhibited differences between mono-infected and HIV-1-co-infected individuals. HBV F3 was the most common subgenotype in both groups, followed by F1b in HIV-1 co-infection and F2 in HBV mono-infection. In addition, genotype G (single infection) was found in an HIV-1-co-infected individual. A high prevalence of occult HBV infection was detected in HIV-1-co-infected naïve patients (18%), with F2 being the most common genotype (75%). To the best of our knowledge, these results correspond to the first description of frequency and molecular characterization of HBV and HCV in HIV-1 Venezuelan patients.

HIV-1 epidemic in Warao Amerindians from Venezuela: spatial phylodynamics and epidemiological patterns.

OBJECTIVES: We previously reported HIV-1 infection in Warao Amerindians from Venezuela. The aim of this study was to evaluate the extent and the dynamic of HIV-1 dissemination in eight Warao communities. DESIGN AND SETTING: HIV-1 infection was evaluated in 576 Warao Amerindians from the Orinoco Delta. Partial HIV-1 pol sequences were analyzed to reconstruct the spatiotemporal and demographic dynamics of the epidemic. RESULTS: HIV-1 antibodies were present in 9.55% of Warao Amerindians, ranging from 0 to 22%. A significantly higher prevalence was found in men (15.6%) compared with women (2.6%), reaching up to 35% in men from one community. All but one isolates were classified as subtype B. Warao's HIV-1 subtype-B epidemic resulted from a single viral introduction at around the early 2000s. After an initial phase of slow growth, the subtype B started to spread at a fast rate (0.8/year) following two major routes of migration within the communities. CONCLUSION: A dramatic high prevalence was documented in almost all the communities of Warao Amerindians from the Orinoco Delta tested for HIV-1 infection. This epidemic resulted from the dissemination of a single HIV-1 subtype B founder strain introduced about 10 years ago and its size is probably doubling every year, creating a situation that can be devastating for this vulnerable Amerindian group.

Prevalence of amino acid mutations in hepatitis C virus core and NS5B regions among Venezuelan viral isolates and comparison with worldwide isolates.

BACKGROUND: Recent reports show that R70Q and L/C91M amino acid substitutions in the core from different hepatitis C virus (HCV) genotypes have been associated with variable responses to interferon (IFN) and ribavirin (RBV) therapy, as well to an increase of hepatocellular carcinoma (HCC) risk, liver steatosis and insulin resistance (IR). Mutations in NS5B have also been associated to IFN, RBV, nucleoside and non-nucleoside inhibitors drug resistance. The prevalence of these mutations was studied in HCV RNA samples from chronically HCV-infected drug-naïve patients. METHODS: After amplification of core and NS5B region by nested-PCR, 12 substitutions were analyzed in 266 Venezuelan HCV isolates subtype 1a, 1b, 2a, 2c, 2b, 2j (a subtype frequently found in Venezuela) and 3a (n = 127 and n = 228 for core and NS5B respectively), and compared to isolates from other countries (n = 355 and n = 646 for core and NS5B respectively). RESULTS: R70Q and L/C91M core substitutions were present exclusively in HCV G1b. Both substitutions were more frequent in American isolates compared to Asian ones (69% versus 26%, p < 0.001 and 75% versus 45%, p < 0.001 respectively). In Venezuelan isolates NS5B D310N substitution was detected mainly in G3a (100%) and G1a (13%), this later with a significantly higher prevalence than in Brazilian isolates (p = 0.03). The NS5B mutations related to IFN/RBV treatment D244N was mainly found in G3a, and Q309R was present in all genotypes, except G2. Resistance to new NS5B inhibitors (C316N) was only detected in 18% of G1b, with a significantly lower prevalence than in Asian isolates, where this polymorphism was surprisingly frequent (p < 0.001). CONCLUSIONS: Genotypical, geographical and regional differences were found in the prevalence of substitutions in HCV core and NS5B proteins. The substitutions found in the Venezuelan G2j type were similar to that found in G2a and G2c isolates. Our results suggest a high prevalence of the R70Q and L/C91M mutations of core protein for G1b and D310N substitution of NS5B protein for the G3a. C316N polymorphism related with resistance to new NS5B inhibitors was only found in G1b. Some of these mutations could be associated with a worse prognosis of the disease in HCV infected patients.

Evidence of at least two introductions of HIV-1 in the Amerindian Warao population from Venezuela.

BACKGROUND: The Venezuelan Amerindians were, until recently, free of human immunodeficiency virus (HIV) infection. However, in 2007, HIV-1 infection was detected for the first time in the Warao Amerindian population living in the Eastern part of Venezuela, in the delta of the Orinoco river. The aim of this study was to analyze the genetic diversity of the HIV-1 circulating in this population. METHODOLOGY/PRINCIPAL FINDINGS: The pol genomic region was sequenced for 16 HIV-1 isolates and for some of them, sequences from env, vif and nef genomic regions were obtained. All HIV-1 isolates were classified as subtype B, with exception of one that was classified as subtype C. The 15 subtype B isolates exhibited a high degree of genetic similarity and formed a highly supported monophyletic cluster in each genomic region analyzed. Evolutionary analyses of the pol genomic region indicated that the date of the most recent common ancestor of the Waraos subtype B clade dates back to the late 1990s. CONCLUSIONS/SIGNIFICANCE: At least two independent introductions of HIV-1 have occurred in the Warao Amerindians from Venezuela. The HIV-1 subtype B was successfully established and got disseminated in the community, while no evidence of local dissemination of the HIV-1 subtype C was detected in this study. These results warrant further surveys to evaluate the burden of this disease, which can be particularly devastating in this Amerindian population, with a high prevalence of tuberculosis, hepatitis B, among other infectious diseases, and with limited access to primary health care.

Genetic history of hepatitis C virus in Venezuela: high diversity and long time of evolution of HCV genotype 2.

BACKGROUND: The subtype diversity of the hepatitis C virus (HCV) genotypes is unknown in Venezuela. METHODOLOGY/PRINCIPAL FINDINGS: 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. CONCLUSIONS/SIGNIFICANCE: 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.