[Isolation, identification and genetic analysis of an H1N1 subtype isolate of swine influenza virus]

In 2006, a swine influenza virus (SIV) isolate was isolated from 30 nasal swabs samples collected from pigs with clinical syndromes of swine influenza in a pig farm of Liaoning Province. The virus isolate was studied and identified by the growth in 9-11 days old chicken embryo, hemagglutination (HA) assay, hemagglutination inhibition (HI) assay, reverse transcription-polymerase chain reaction assays (RT-PCR) for its genetic subtype, whole gene sequence analysis and animal trial for its virulence. The virus isolate demonstrated the hemagglutination activity. Result of HI test against H1 subtype of SIV was positive, however, the results were negative when the HI studies were conducted using SIV H3 subtype virus and Newcastle Disease Virus (NDV). Eight gene segments of the virus isolate were amplified by RT-PCR. Phylogenetic analysis of the gene sequence of the virus isolate by using DNAstar software program revealed that the isolate have the H1 HA gene, by comparing to the sequences of H1-H16 in the GenBank. Furthermore, sequencing results also demonstrated that the virus isolate's NA gene belongs to N1 subtype. Therefore, the subtype of the SIV isolate is H1N1. The results of sequence analysis indicated that the genome of the SIV-H1N1 LN strain includes 8 fragments, among which only M protein gene is not swine originated. All other 7 fragments have close relationship with the domestic standard swine H1N1 strains. Results suggested that the SIV isolate LN strain might be created by a possible triple reassortants among the classic swine influenza virus, human influenza-like virus, and avian influenza-like virus. Piglets were inoculated with the SIV LN strain virus preparations and the virus caused the typical clinical symptoms of swine influenza in the inoculated piglets. This study, the isolation, identification and genetic analysis of the SIV LN strain provided useful information and basic data for the further investigation of epidemic principles and patterns of swine influenza virus in China.