Project description:Zoonotic influenza A viruses of avian origin can cause severe disease in individuals, or even global pandemics, and thus pose a threat to human populations. Waterfowl and shorebirds are believed to be the reservoir for all influenza A viruses, but this has recently been challenged by the identification of novel influenza A viruses in bats. The major bat influenza A virus envelope glycoprotein, haemagglutinin, does not bind the canonical influenza A virus receptor, sialic acid or any other glycan, despite its high sequence and structural homology with conventional haemagglutinins. This functionally uncharacterized plasticity of the bat influenza A virus haemagglutinin means the tropism and zoonotic potential of these viruses has not been fully determined. Here we show, using transcriptomic profiling of susceptible versus non-susceptible cells in combination with genome-wide CRISPR-Cas9 screening, that the major histocompatibility complex class II (MHC-II) human leukocyte antigen DR isotype (HLA-DR) is an essential entry determinant for bat influenza A viruses. Genetic ablation of the HLA-DR α-chain rendered cells resistant to infection by bat influenza A virus, whereas ectopic expression of the HLA-DR complex in non-susceptible cells conferred susceptibility. Expression of MHC-II from different bat species, pigs, mice or chickens also conferred susceptibility to infection. Notably, the infection of mice with bat influenza A virus resulted in robust virus replication in the upper respiratory tract, whereas mice deficient for MHC-II were resistant. Collectively, our data identify MHC-II as a crucial entry mediator for bat influenza A viruses in multiple species, which permits a broad vertebrate tropism.
Project description:Purpose: Next-generation sequencing (NGS) on targeted locus in Sindbis genome to determine frequency changes of artificial microRNAs expressed by viruses after passaging in cancer and normal cells Methods: RNA was harvested in Trizol 488 (Thermo Fisher). RNA was extracted using the manufacturer’s protocol and quantified by nanodrop. Sequencing was done by SeqMatic on a MiSeq v3 platform generating 75bp reads. Adapters were trimmed using Trimmomatic and adapter-free reads represent artificial microRNAs encoded by Sindbis virus in a sample. Results: We have identified changes in artificial microRNA frequency after passaging virus pool in cancer and normal cells and have identified microRNAs increasing viral fitness in cancer cells. Conclusions: Our study represents artificial microRNAs which target pathways that can aid oncolytic viral replication in cancer cells.
Project description:We have developed a generally adaptable, novel high-throughput chromosome conformation capture assay for use in trans (V3C-seq) that allows genome-wide identification of the direct associations of a lytic virus genome with discreet regions of the cellular chromosome. Upon infection, the parvovirus Minute Virus of Mice genome associated directly with sites of cellular DNA damage. These sites also exhibited damage in uninfected cells when cycling through S-phase. As infection proceeded, new sites of DNA damage were induced, and virus subsequently also associated with these.
Project description:We performed whole genome single nucleotide polymorphism (SNP) based analysis of all available Venezuelan equine encephalitis (VEE) virus antigenic complex genomes and developed a high resolution genome-wide SNP microarray. We used the SNP microarray to analyze a broad panel of VEEV isolates, found excellent concordance between array and sequence based genotypes for previously sequenced strains, and genotyped unsequenced strains.