Background Recent studies have revealed the existence of genetic diversity in swine influenza viruses (SIVs) in the world. In Thailand, there has been a little information on the molecular characteristics of the SIVs since the first isolation of viruses of H1N1 and H3N2 subtypes in the late 1970s. Our previous study demonstrated that Thai H1N1 SIVs possessed the classical swine H1 and avian‐like swine N1 genes (Takemae et al., Proceedings of the Options for the Control of Influenza VI.2007;350–353). Objectives In the present study, we genetically characterized 12 SIVs including those of H1N1, H1N2 and H3N2 subtypes isolated between 2000 and 2005. Methods We determined the entire nucleotide sequences of the eight gene segments of those isolates. Results Phylogenetic analysis revealed the existence of nine distinct genotypes amongst the Thai SIVs. These genotypes arose from multiple introductions of classical swine, avian‐like swine and human viruses. The existence of two distinct sublineages within classical swine H1 and NS, avian‐like swine PA and M and human H3 and N2 genes of the Thai SIVs suggested that introduction of viruses of classical swine, avian‐like swine and human origins occurred twice respectively into the Thai pig population. The predominance of avian‐like swine genes amongst the Thai SIVs was evident. In particular, three polymerase (PB1, PB2 and PA) and matrix genes of avian‐like swine origin were retained in all the Thai SIVs examined. Conclusions These observations may suggest that genes of avian‐like swine lineages have some advantages to be maintained in pigs as seen in the SIVs established through multiple introductions in other regions.
Alterations of the receptor-binding properties of swine influenza A viruses (SIVs) during their isolation in embryonated chicken eggs have not been well studied. In this study, the receptorbinding properties of classical H1 SIVs isolated solely in eggs or Madin-Darby canine kidney (MDCK) cells were examined. Sequencing analysis revealed substitutions of D190V/N or D225G in the haemagglutinin (HA) proteins in egg isolates, whereas MDCK isolates retained HA genes identical to those of the original viruses present in the clinical samples. Egg isolates with substitution of either D190V/N or D225G had increased haemagglutinating activity for mouse and sheep erythrocytes, but reduced activity for rabbit erythrocytes. Additionally, egg isolates with D225G had increased haemagglutination activity for chicken erythrocytes. A direct binding assay using a sialyl glycopolymer that possessed either a 5-N-acetylneuraminic acid (Neu5Ac) a2,6galactose (Gal) or a Neu5Aca2,3Gal linkage revealed that the egg isolates used in this study showed higher binding activity to the Neu5Aca2,3Gal receptor than MDCK isolates. Increased binding activity of the egg isolates to the Neu5Aca2,3Gal receptor was also confirmed by haemagglutination assay with resialylated chicken erythrocytes by Galb1,3/4GlcNAca2,3-sialyltransferase. These observations were reinforced by flow-cytometric and N-glycan analyses of the erythrocytes. The a2,3-linked sialic acids were expressed predominantly on the surface of mouse and sheep erythrocytes. Chicken erythrocytes expressed Neu5Aca2,3Gal more abundantly than Neu5Aca2,6Gal, and rabbit erythrocytes expressed both 5-N-glycolylneuraminic acid (Neu5Gc) a2,6Gal and Neu5Aca2,6Gal. Our results demonstrate clearly that classical H1 SIVs undergo alterations in receptor-binding activity associated with an amino acid substitution in the HA protein during isolation and propagation in embryonated chicken eggs.
BackgroundUnderstanding swine influenza virus (SIV) ecology has become more and more important from both the pig industry and public health points of views. However, the mechanism whereby SIV occurs in pig farms is not well understood. The purpose of this study was to develop a proper strategy for SIV surveillance.FindingsWe conducted longitudinal monitoring in 6 farrow-to-finish farms in the central region of Thailand from 2008 to 2009. Nasal swabs and serum samples were collected periodically from clinically healthy pigs consisting of sows, fattening pigs, weaned piglets and pigs transferred from other farms. A total of 731 nasal swabs were subjected to virus isolation and 641 serum samples were subjected to detection of SIV antibodies against H1 and H3 subtypes using the hemagglutination inhibition test and ELISA. Twelve SIVs were isolated in this study and eleven were from piglets aged 4 and 8 weeks. Phylogenetical analysis revealed that SIVs isolated from different farms shared a common ancestor. Antibodies against SIVs were detected in fattening pigs on farms with no SIV isolation in the respective periods studied. These observations suggested that piglets aged 8 weeks or younger could be a main target for SIV isolation. Farm-to-farm transmission was suggested for farms where pigs from other farms are introduced periodically. In addition, antibodies against SIVs detected in fattening pigs could be a marker for SIV infection in a farm.ConclusionsThe present study provided important information on SIV surveillance that will enable better understanding of SIV ecology in farrow-to-finish farms.
ABSTRACT. The adjuvant effect of chicken interferon-γ (ChIFN-γ) was examined for protecting chickens against intestinal colonization of Salmonella Enteritidis (SE) following oral exposure. Ten 7-week-old chickens per group were immunized with inactivated SE twice with or without co-administration of ChIFN-γ intramuscularly, and all chickens were challenged with SE. Sera collected from immunized groups with or without ChIFN-γ, and from unimmunized group were measured for SE antibody by agglutination test. The levels of antibodies were raised by 1 week post-immunization and did not show any difference between groups with and without ChIFN-γ. No antibodies were detected in unimmunized group before challenge. Fecal samples from each group were cultured at 1, 4, 7, and 13 days postchallenge to determine the incidence of intestinal colonization and the numbers of SE shed into the environment. Co-administration of ChIFN-γ, significantly reduced the incidence of intestinal colonization (P<0.05). At 13 days post-challenge, the bacterial counts of SE in organs were also reduced in ChIFN-γ administered group. These data suggest co-administration of ChIFN-γ with SE antigen enhances protection against SE challenge without acceleration of antibody production.
ABSTRACT. Bioactive recombinant chicken interferon-α (ChIFN-α) was expressed in a baculovirus system. For easy purification, it was expressed as ChIFN-α bearing histidine hexamer (His-tag) at C-terminal, designated ChIFN-αHis. The expressed proteins were detected by SDS-PAGE analysis with Coomassie brilliant blue staining as around 23 and 19 kDa bands thought to be immature and matured ChIFN-αHis respectively. The purified ChIFN-αHis with a nickel chelated column showed anti-viral activity in vitro.
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