Differential MicroRNA Expression And T-cell Receptor Repertoires Of The Chicken And Duck Infected By H5N1-subtype Highly Pathogenic Avian Inlfuenza

Chicken and duck show large divergence in susceptibility to H5N1highly pathogenicavian influenza virus (HPAIV). Various studies have been focusing on impact of thecontinuing evolution of avian influenza virus and host adaption to it. Few studiesexplored the mechanism underlining the divergent susceptibility of the two birdspecies. This study aimed to explore the difference in miRNA repertoires andexpression patterns in spleen, thymus and bursa of Fabricius of chicken and duckinfected by H5N1HPAIV to explain the immunity divergence of the two species.We analyzed microRNA (miRNA) expression of spleen, thymus and bursa of theFabricius of chicken and duck with and without H5N1HPAIV infection byhigh-throughput sequencing. Using these unbiased small RNA data sets of chickenand duck, firstly, we identified486and262mature miRNAs respectively for chickenand duck. The262duck miRNAs belonged to83miRNA families (128chickenmiRNA families). Secondly, the expression of conserved miRNA in immune organs ofSPF chickens showed much divergence. And the diverged miRNA expression showedtissue-specificity. Thirdly, the relative expression of miRNAs in the immune organs ofchickens and ducks with and without H5N1HPAIV-infection were clustered across alllibraries of chickens and ducks. The data indicated that the miRNA expression inimmune organs of chicken showed more divergence after H5N1HPAIV infection thanthose in ducks. MiRNA expression in the spleen and bursa of Fabricius showed highcorrelation with those in SPF ducks. However, in the thymus of infected chickens, themiRNA expression was distinct with those in SPF chickens and ducks. This might berelated to the dysfunction of the thymus of the infected chickens.Through miRNA expression analysis and targets prediction, we constructed themiRNA regulatory networks of chickens and ducks. Through digital gene expressionin duck spleen, we investgated the changes in the pathways related to immuneresponses. The up-regulated genes were highly enriched in JAK-STAT, WNT,RIG-like and Toll-like pathways.We analysed the T-cell receptor genes of chickens and ducks. First, we cloned the cDNA and the genomic sequences of TCR β chain in chickens and ducks by5’RACEand Genome Walking methods. Three Vβ genes were found for ducks, of which Vβ3was a homologue of Vβ1. Phylogenetic analysis of the Vβ genes of birds andmammals revealed that Vβ1and Vβ2of bird species showed different features inevolution. Second, using high-throuthput sequencing, we investigated the features ofTCRβ CDR3of chickens and ducks with and without H5N1HPAIV infection. Thedata revealed the features of VDJ rearangement and CDR3sequences of chicken andduck TCR β chain. Third, we investgated the clonotypes of TCRβ CDR3s. Thediversity of TCRβ CDR3clonotypes were estimated using the preseqR package. Theresult indicated that there were more CDR3clonotypes in duck thymus than that ofchickens. And there are more CDR3clonotypes in chickens and ducks infected byH5N1HPAIV.We provide annotation of duck miRNAs using high-throughput sequencing andbioinformatics tools. The comparison of miRNA expression patterns in immuneorgans of chicken and duck provides evidence of their difference in susceptibility toH5N1avian influenza. We constructed the regulatory networks related to immuneresponses of chickens and ducks. We revealed the features of TCRβ genes, VDJrecombination and CDR3clonotypes.