IAV is a critically important pathogen in terms of human health, food security and animal welfare. Despite coordinated international surveillance and control strategies, IAV regularly causes globally significant outbreaks of disease. Genome editing has the potential to generate livestock that are resistant to IAV, thereby reducing the burden on the farming economy in addition to limiting the source of future pandemic strains. In chickens, IAV relies on the host protein ANP32A to replicate. However, infection with high MOI can lead to breakthrough infections generating IAV escape mutants which have evolved to replicate independently of ANP32A. Therefore, additional high confidence host targets must be identified which may prevent the formation of ANP32A escape mutants. Here, we performed a genome-scale CRISPR/Cas9 KO survival screen in chicken lung epithelial cells (CLEC213) using the avian UDL 3:5 reassortant IAV strain. Several genes were enriched after consecutive challenges, with most guides found to target genes involved in sialic acid biosynthesis and N-linked glycosylation. These include SLC35A1, SLC35A2, MOGS and MGAT1. In addition, a novel host dependency factor was identified previously not associated with IAV infection. Subsequent validation experiments show this gene reduces IAV infection of CLEC213 cells for multiple IAV strains and can increase cell survival post infection. Trait stacking KO of this gene with ANP32A in CLEC213 cells is underway to determine its impact on IAV escape mutant generation.