Virus research
-
A wide range of sensitivity has been reported for rapid influenza antigen tests (RIAT). In this study, we analyzed the viral loads of 778 pandemic H1N1- and 227 seasonal H3N2-virus positive clinical specimens collected during the same period and found that viral loads in pandemic H1N1 viruses was characterized by lower copy numbers than seasonal H3N2 viruses. Among various factors including the timing of specimen collection, patient age, patient gender and subtype of influenza, we found that the subtype of influenza was the most important determinant of viral load. ⋯ Based on three strategies, including cut-off values, performance on a subset of clinical specimens and evaluated performance curve of the Espline influenza A&B-N RIAT, the clinical sensitivities were 48.7-55.9% for pandemic H1N1 and 64.0-70.5% for seasonal H3N2 viruses in this study. These results indicate that the distributions of viral loads of different influenza A subtypes substantially influence the sensitivity of RIAT for clinical specimens. The lower sensitivity of RIAT for pandemic H1N1 than seasonal H3N2 virus is mainly due to differences in viral load in clinical samples rather than a diminished capacity of RIAT itself to detect these two subtypes of influenza A viruses.
-
HIV-1 Vpr, an accessory protein with multiple functions, is involved in the induction of apoptosis, cell cycle G2 arrest, and modulation of gene expression. Many functions of this protein have been documented for the wild-type subtype B Vpr, however the functionality of other subtypes has not sufficiently been addressed. In this study, the functionality of Subtype B Vpr, 6 subtype C mutant Vpr proteins and the consensus sequence of subtype C Vpr were compared with each other. ⋯ Most natural mutations in Vpr not only do not disturb the functions of the protein but also potentiate the protein for an increased functionality. The natural mutations of Vpr may thus not always be regarded as defective mutations. The study suggests the adaptive role of the natural mutations commonly found in subtype C Vpr.
-
SARS coronavirus (SARS-CoV) emerged in 2002 as an important cause of severe lower respiratory tract infection in humans and in vitro models of the lung are needed to elucidate cellular targets and the consequences of viral infection. The severe and sudden onset of symptoms, resulting in an atypical pneumonia with dry cough and persistent high fever in cases of severe acute respiratory virus brought to light the importance of coronaviruses as potentially lethal human pathogens and the identification of several zoonotic reservoirs has made the reemergence of new strains and future epidemics all the more possible. In this chapter, we describe the pathology of SARS-CoV infection in humans and explore the use of two models of the human conducting airway to develop a better understanding of the replication and pathogenesis of SARS-CoV in relevant in vitro systems. ⋯ SARS-CoV GFP replicated to similar titers as wild type viruses in Vero E6, MA104, and CaCo2 cells. In addition, SARS-CoV replication in airway epithelial cultures generated from Golden Syrian hamster tracheas reached similar titers to the human cultures by 72 h post-infection. Efficient SARS-CoV infection of ciliated cell-types in HAE provides a useful in vitro model of human lung origin to study characteristics of SARS-CoV replication and pathogenesis.
-
Comparative Study
Evaluating the 3C-like protease activity of SARS-Coronavirus: recommendations for standardized assays for drug discovery.
Although the initial outbreaks of the deadly coronavirus that causes severe acute respiratory syndrome (SARS-CoV) were controlled by public health measures, the development of vaccines and antiviral agents for SARS-CoV is essential for improving control and treatment of future outbreaks. One potential target for SARS-CoV antiviral drug development is the 3C-like protease (3CLpro). This enzyme is an attractive target since it is essential for viral replication, and since there are now a number of high resolution X-ray structures of SARS-CoV 3CLpro available making structure-based drug-design possible. ⋯ Moreover, we provide experimental evidence showing that the activity of 3CLpro enzymatic is significantly reduced when non-native sequences or affinity-tags are added to the N- or C-termini of the enzyme, or when the enzyme used in assays is at concentrations below the equilibrium dissociation constant of the 3CLpro dimer. We demonstrate for the first time the utility of a highly sensitive and novel Alexa488-QSY7 FRET-based peptide substrate designed for routine analysis and high-throughput screening, and show that kinetic constants determined from FRET-based assays that are uncorrected for inner-filter effects can lead to artifacts. Finally, we evaluated the effects of common assay components including DTT, NaCl, EDTA and DMSO on enzymatic activity, and we recommend standardized assay conditions and constructs for routine SARS-CoV 3CLpro assays to facilitate direct comparisons between SARS-CoV 3CLpro inhibitors under development worldwide.
-
Hepatitis C virus (HCV) has been the subject of intense research and clinical investigations due to its worldwide prevalence and major role in chronic liver disease. Like most RNA viruses, HCV circulates in vivo as a complex population of different but closely related viral variants, commonly referred to as a quasispecies. Recent studies suggest that ribavirin might exert an antiviral effect against HCV through both mutagenic effect and an impairment of RNA replication. ⋯ In children with chronic HCV infection, the viral population is initially highly homogeneous, but diversifies during prolonged infection which seems to be a common event during chronic hepatitis C in childhood. Coinfection of human immunodeficiency virus 1 (HIV-1) patients by HCV can complicate the treatment of these patients with highly active antiretroviral therapy (HAART). HIV coinfection is associated with a decrease of HCV quasispecies variability, which appears to be reversed by effective HAART.