Genetic characterization of 2008 reassortant influenza A virus (H5N1), Thailand

In January and November 2008, outbreaks of avian influenza have been reported in 4 provinces of Thailand. Eight Influenza A H5N1 viruses were recovered from these 2008 AI outbreaks and comprehensively characterized and analyzed for nucleotide identity, genetic relatedness, virulence determinants, and possible sites of reassortment. The results show that the 2008 H5N1 viruses displayed genetic drift characteristics (less than 3% genetic differences), as commonly found in influenza A viruses. Based on phylogenetic analysis, clade 1 viruses in Thailand were divided into 3 distinct branches (subclades 1, 1.1 and 1.2). Six out of 8 H5N1 isolates have been identified as reassorted H5N1 viruses, while other isolates belong to an original H5N1 clade. These viruses have undergone inter-lineage reassortment between subclades 1.1 and 1.2 and thus represent new reassorted 2008 H5N1 viruses. The reassorted viruses have acquired gene segments from H5N1, subclade 1.1 (PA, HA, NP and M) and subclade 1.2 (PB2, PB1, NA and NS) in Thailand. Bootscan analysis of concatenated whole genome sequences of the 2008 H5N1 viruses supported the reassortment sites between subclade 1.1 and 1.2 viruses. Based on estimating of the time of the most recent common ancestors of the 2008 H5N1 viruses, the potential point of genetic reassortment of the viruses could be traced back to 2006. Genetic analysis of the 2008 H5N1 viruses has shown that most virulence determinants in all 8 genes of the viruses have remained unchanged. In summary, two predominant H5N1 lineages were circulating in 2008. The original CUK2-like lineage mainly circulated in central Thailand and the reassorted lineage (subclades 1.1 and 1.2) predominantly circulated in lower-north Thailand. To prevent new reassortment, emphasis should be put on prevention of H5N1 viruses circulating in high risk areas. In addition, surveillance and whole genome sequencing of H5N1 viruses should be routinely performed for monitoring the genetic drift of the virus and new reassorted strains, especially in light of potential reassortment between avian and mammalian H5N1 viruses

Performance of the QuickVue Influenza A+B Rapid Test for Pandemic H1N1 (2009) Virus Infection in Adults

To investigate the diagnostic accuracy of the QuickVue® Influenza A+B rapid test we conducted a prospective observational study in which this rapid test was compared with a real-time reverse transcription polymerase chain reaction (RT-PCR) for pandemic influenza A H1N1 (2009) infection in Austrian adults. The sensitivity, specificity, and positive and negative predictive values of the QuickVue test compared with the RT-PCR were 26% (95% CI 18–35), 98% (95% CI 92–100), 94% (95% CI 80–99) and 50% (95% CI 42–58), respectively. The prevalence of pandemic H1N1 (2009) virus infection among the 209 patients included in the study was 57%. Our data suggest that a positive QuickVue test provides considerable information for the diagnosis of pandemic influenza A H1N1 (2009) virus infection in young adults but that a negative QuickVue test result should, if relevant for patient management or public health measures, be verified using PCR

Evaluation of a new, rapid, simple test for the detection of influenza virus

BACKGROUND: Influenza virus infections are responsible for significant morbidity and mortality in both pediatric and adult populations worldwide. Rapid and accurate diagnosis of influenza is necessary for appropriate patient management during the influenza season and for optimal utilization of anti-influenza therapy. We prospectively tested the accuracy of a simple and rapid diagnostic method. METHODS: Ninety-eight samples (nasal and pharyngeal swabs) from patients with upper respiratory tract infection symptoms who presented to primary healthcare centres in Barcelona (Spain) were prospectively analyzed. The samples were collected as part of influenza surveillance program. Samples that had enough volume to make the new test after aliquoting the amount needed to perform routine tests were included. None of the samples were pre-selected as a result of their status in relation to influenza virus. Samples were analyzed by in-house real-time PCR and Alere™ i Influenza A & B (Alere™ i), which uses isothermal amplification of nucleic acids for the qualitative detection of influenza A and B in nasal swabs transported in viral transport media. The two techniques were compared by positive percent agreement (PPA) and negative percent agreement (NPA). Statistical analysis was performed with Stata. RESULTS: Of the 98 samples analysed 90 were concordant; 46 (46.9%) were positive and 44 (44.9%) were negative. Five samples showed invalid results with the Alere™ i test and could be not re-tested due to insufficient sample volume and were not included in the final statistical analysis. In the 93 remaining samples, the Alere™ i test showed 97% of accuracy having correctly classified 90 samples. We obtained discordant results in 3 samples (3%). The PPA was 93.8% for influenza A and 94.1% for influenza B, and NPA was 100% for influenza A and influenza B virus. In addition, the Alere™ i was very rapid (15 minutes or less) and extremely easy to use. CONCLUSIONS: The Alere™ i test provided a good correlation compared to the real-time PCR test for the diagnosis of influenza. Since this method can be performed in minutes, it allows immediate, accurate clinical decisions to prescribe appropriate antiviral treatment or isolation of patients

Molecular Epidemiology and Evolution of Human Enterovirus Serotype 68 in Thailand, 2006–2011

BACKGROUND: Publications worldwide have reported on the re-occurrence of human enterovirus 68 (EV68), a rarely detected pathogen usually causing respiratory illness. However, epidemiological data regarding this virus in particular on the Asian continent has so far been limited. METHODOLOGY/FINDINGS: We investigated the epidemiology and genetic variability of EV68 infection among Thai children with respiratory illnesses from 2006-2011 (n = 1810). Semi-nested PCR using primer sets for amplification of the 5'-untranslated region through VP2 was performed for rhino-enterovirus detection. Altogether, 25 cases were confirmed as EV68 infection indicating a prevalence of 1.4% in the entire study population. Interestingly, the majority of samples were children aged >5 years (64%). Also, co-infection with other viruses was found in 28%, while pandemic H1N1 influenza/2009 virus was the most common co-infection. Of EV68-positive patients, 36% required hospitalizations with the common clinical presentations of fever, cough, dyspnea, and wheezing. The present study has shown that EV68 was extremely rare until 2009 (0.9%). An increasing annual prevalence was found in 2010 (1.6%) with the highest detection frequency in 2011 (4.3%). Based on analysis of the VP1 gene, the evolutionary rate of EV68 was estimated at 4.93 × 10(-3) substitutions/site/year. Major bifurcation of the currently circulating EV68 strains occurred 66 years ago (1945.31 with (1925.95-1960.46)95% HPD). Among the current lineages, 3 clusters of EV68 were categorized based on the different molecular signatures in the BC and DE loops of VP1 combined with high posterior probability values. Each cluster has branched off from their common ancestor at least 36 years ago (1975.78 with (1946.13-1984.97)95% HPD). CONCLUSION: Differences in epidemiological characteristic and seasonal profile of EV68 have been found in this study. Results from Bayesian phylogenetic investigations also revealed that EV68 should be recognized as a genetically diverse virus with a substitution rate identical to that of enterovirus 71 genotype B (4.2 × 10(-3 )s/s/y)

Genetic Structure of Human A/H1N1 and A/H3N2 Influenza Virus on Corsica Island: Phylogenetic Analysis and Vaccine Strain Match, 2006–2010

Background: The aim of this study was to analyse the genetic patterns of Hemagglutinin (HA) genes of influenza A strains circulating on Corsica Island during the 2006-2009 epidemic seasons and the 2009-2010 pandemic season. [br/] Methods: Nasopharyngeal samples from 371 patients with influenza-like illness (ILI) were collected by General Practitioners (GPs) of the Sentinelles Network through a randomised selection routine. [br/] Results: Phylogenetic analysis of HA revealed that A/H3N2 strains circulating on Corsica were closely related to the WHO recommended vaccine strains in each analyzed season (2006-2007 to 2008-2009). Seasonal Corsican influenza A/H1N1 isolated during the 2007-2008 season had drifted towards the A/Brisbane/59/2007 lineage, the A/H1N1 vaccine strain for the 2008-2009 season. The A/H1N1 2009 (A/H1N1pdm) strains isolated on Corsica Island were characterized by the S220T mutation specific to clade 7 isolates. It should be noted that Corsican isolates formed a separate sub-clade of clade 7 as a consequence of the presence of the fixed substitution D222E. The percentages of the perfect match vaccine efficacy, estimated by using the p(epitope) model, against influenza viruses circulating on Corsica Island varied substantially across the four seasons analyzed, and tend to be highest for A/H1N1 compared with A/H3N2 vaccines, suggesting that cross-immunity seems to be stronger for the H1 HA gene. [br/] Conclusion: The molecular analysis of the HA gene of influenza viruses that circulated on Corsica Island between 2006-2010 showed for each season the presence of a dominant lineage characterized by at least one fixed mutation. The A/H3N2 and A/H1N1pdm isolates were characterized by multiples fixation at antigenic sites. The fixation of specific mutations at each outbreak could be explained by the combination of a neutral phenomenon and a founder effect, favoring the presence of a dominant lineage in a closed environment such as Corsica Island

Virologic and clinical characteristics of HBV genotypes/subgenotypes in 487 Chinese pediatric patients with CHB

<p>Abstract</p> <p>Background</p> <p>The association of hepatitis B virus (HBV) genotypes/subgenotypes with clinical characteristics is increasingly recognized. However, the virologic and clinical features of HBV genotypes/subgenotypes in pediatric patients remain largely unknown.</p> <p>Methods</p> <p>Four hundred and eighty-seven pediatric inpatients with CHB were investigated, including 217 nucleos(t)ide analog-experienced patients. HBV genotypes/subgenotypes and reverse transcriptase (RT) mutations were determined by direct sequencing. The stage of fibrosis and degree of inflammatory activity were evaluated by the Metavir score system.</p> <p>Results</p> <p>Among 487 enrolled pediatric patients, HBV genotype C2 and B2 were the most two prevalent (73.7% and 21.1%). Comparing with HBV/B2 infected patients, no significant difference was observed in the incidence rate and mutant patterns of lamivudine- or adefovir-resistant mutations in HBV/C2 infected patients (<it>P </it>> 0.05). Importantly, we found that the degree of hepatic inflammation degree, fibrosis stage and ALT level were significantly higher in HBV/C2-infected HBeAg positive patients than it was in HBV/B2-infected ones.</p> <p>Conclusions</p> <p>The pediatric patients with HBV/C2 infection might be more susceptible to develop severe liver pathogenesis.</p

Direct association between pharyngeal viral secretion and host cytokine response in severe pandemic influenza

<p>Abstract</p> <p>Background</p> <p>Severe disease caused by 2009 pandemic influenza A/H1N1virus is characterized by the presence of hypercytokinemia. The origin of the exacerbated cytokine response is unclear. As observed previously, uncontrolled influenza virus replication could strongly influence cytokine production. The objective of the present study was to evaluate the relationship between host cytokine responses and viral levels in pandemic influenza critically ill patients.</p> <p>Methods</p> <p>Twenty three patients admitted to the ICU with primary viral pneumonia were included in this study. A quantitative PCR based method targeting the M1 influenza gene was developed to quantify pharyngeal viral load. In addition, by using a multiplex based assay, we systematically evaluated host cytokine responses to the viral infection at admission to the ICU. Correlation studies between cytokine levels and viral load were done by calculating the Spearman correlation coefficient.</p> <p>Results</p> <p>Fifteen patients needed of intubation and ventilation, while eight did not need of mechanical ventilation during ICU hospitalization. Viral load in pharyngeal swabs was 300 fold higher in the group of patients with the worst respiratory condition at admission to the ICU. Pharyngeal viral load directly correlated with plasma levels of the pro-inflammatory cytokines IL-6, IL-12p70, IFN-γ, the chemotactic factors MIP-1β, GM-CSF, the angiogenic mediator VEGF and also of the immuno-modulatory cytokine IL-1ra (p < 0.05). Correlation studies demonstrated also the existence of a significant positive association between the levels of these mediators, evidencing that they are simultaneously regulated in response to the virus.</p> <p>Conclusions</p> <p>Severe respiratory disease caused by the 2009 pandemic influenza virus is characterized by the existence of a direct association between viral replication and host cytokine response, revealing a potential pathogenic link with the severe disease caused by other influenza subtypes such as H5N1.</p

Catalysing sustainable fuel and chemical synthesis

Concerns over the economics of proven fossil fuel reserves, in concert with government and public acceptance of the anthropogenic origin of rising CO2 emissions and associated climate change from such combustible carbon, are driving academic and commercial research into new sustainable routes to fuel and chemicals. The quest for such sustainable resources to meet the demands of a rapidly rising global population represents one of this century’s grand challenges. Here, we discuss catalytic solutions to the clean synthesis of biodiesel, the most readily implemented and low cost, alternative source of transportation fuels, and oxygenated organic molecules for the manufacture of fine and speciality chemicals to meet future societal demands