Surveillance activities and molecular analysis of H5N1 highly pathogenic avian influenza viruses from Thailand, 2004-2005.

Avian influenza (AI) outbreaks were first reported in Thailand in January 2004. In the past 2 yr, AI viruses have caused three epidemic waves. Disease prevention and control in all aspects have been actively carried out. Active and passive surveillance based on clinical observation and laboratory analysis were intensively conducted, as well as monitoring of genetic variation of the viruses. H5N1 viruses isolated from different avian species from different cases and locations were selected. We have sequenced specific genes (HA, NA, M, Ns, and part of PB2 genes) of 58 H5N1 isolates, as well as whole genome sequencing of 21 Thai influenza A (H5N1) viruses isolated during the 2004-2005 outbreak. Cluster analysis study showed that AI isolates were identified as highly pathogenic avian influenza (HPAI) and belonged to genotype Z. The virus had a multiple basic amino acid motif at the cleavage site of HA, deletions in the NA stalk region, a five amino acid deletion in the NS1 gene, and genetic markers for amantadine resistance in the M2 gene. All 58 H5N1 isolates were closely related and grouped into the same cluster, together with isolates from wild birds, cats, tigers, and humans. Phylogenetic analysis also revealed that Thai isolates were in the same cluster as Vietnamese isolates but aligned in a different cluster from Indonesian, Hong Kong, and Chinese viruses. In addition, genetic analysis showed that most avian influenza virus (AIV) isolates from Thailand had no major genetic changes in each gene such as HA (HA cleavage site, receptor binding site, N-link glycosylation site), NA (NA stalk region, oseltamivir resistance marker), M (the amantadine resistance marker, host specificity site), NS (five amino acid deletion site), and PB2 (host specificity site). All Thai poultry isolates contained the amantadine resistance marker while none of them had the oseltamivir resistance marker. To this end, the molecular characterization of H5N1 viruses from Thailand showed that there were no significant point mutations in the critical regions, and there was no evidence of changes in the viruses that indicate they are capable of sustained human-to-human transmission.