ABSTRACT
Mutation in any of five key amino acid residues (at positions 26, 27, 30, 31 and 34) within the M2 protein of influenza A viruses leads to resistance against the amantodine class of anti-influenza drugs. In this study, a pyrosequencing method was described to rapidly detect established five molecular markers of resistance to M2 blockers, amantadine. The residues L26, V27, A30, S31 and G34 in the M2 protein were targeted for pyrosequencing, and 94 avian influenza viruses were used to perform the amantadine resistance analysis. Our results showed that most of avian influenza viruses were amantadine resistant, Mutations V27I and S31N were founded in these isolates.
Subject(s)
Animals , Amantadine , Therapeutic Uses , Antiviral Agents , Therapeutic Uses , Chickens , Drug Resistance, Viral , Genetics , Influenza A virus , Genetics , Influenza in Birds , Drug Therapy , Virology , Reverse Transcriptase Polymerase Chain ReactionABSTRACT
Thirteen isolates of Class I Newcastle disease virus obtained from healthy poultry in China during 2008 were characterized genotypically in this study. All the isolates were proved to be lentogenic strains based on the deduced amino acid sequence of the Fusion protein gene. Molecular epidemiological analysis showed that 13 isolates could be subdivided into 2 distinct genotypes, 11 isolates belonged to genotype 2, and other 2 isolates belonged to genotype 3. Results indicated two genotypes of Class I Newcastle disease virus might widely exist in domestic poultry in China.
Subject(s)
Animals , Humans , Birds , China , Epidemiology , Genotype , Molecular Epidemiology , Methods , Newcastle Disease , Epidemiology , Virology , Newcastle disease virus , Classification , Genetics , Virulence , Phylogeny , Reverse Transcriptase Polymerase Chain Reaction , Viral Fusion Proteins , GeneticsABSTRACT
<p><b>OBJECTIVE</b>To design and rapidly evaluate a TaqMan assay for detecting influenza A viruses.</p><p><b>METHODS</b>The probe and the primers of the assay were designed with the software packages of DNA Star and Primer Premier 5.0. Their specificity and conservation were verified through Blast in GenBank and electronic hybridization. The assay's sensitivity was compared with the standard RT-PCR.</p><p><b>RESULTS</b>The designed primers and probe were confirmed to be very specific and conserved. The assay was 3-27 folds more sensitive than the standard RT-PCR. The RT and PCR steps could be simplified into one step.</p><p><b>CONCLUSION</b>The TaqMan Real-time PCR assay is specific, sensitive and easy to perform.</p>