Traditional approaches to assess the immune response of chickens to infection are through animal trials, which are expensive, require enhanced biosecurity, compromise welfare, and are frequently influenced by confounding variables. Since the chicken embryo becomes immunocompetent prior to hatch, we here characterized the transcriptional response of selected innate immune genes to Newcastle disease virus (NDV) infection in chicken embryos at days 10, 14, and 18 of embryonic development. The results suggest that the innate immune response 72 h after challenge of 18-day chicken embryo is both consistent and robust. The expression of CCL5, Mx1, and TLR3 in lung tissues of NDV challenged chicken embryos from the outbred Kuroiler and Tanzanian local ecotype lines showed that their expression was several orders of magnitude higher in the Kuroiler than in the local ecotypes. Next, the expression patterns of three additional innate-immunity related genes, IL-8, IRF-1, and STAT1, were examined in the highly congenic Fayoumi (M5.1 and M15.2) and Leghorn (Ghs6 and Ghs13) sublines that differ only at the microchromosome bearing the major histocompatibility locus. The results show that the Ghs13 Leghorn subline had a consistently higher expression of all genes except IL-8 and expression seemed to be subline-dependent rather than breed-dependent, suggesting that the innate immune response of chicken embryos to NDV infection may be genetically controlled by the MHC-locus. Taken together, the results suggest that the chicken embryo may represent a promising model to studying the patterns and sources of variation of the avian innate immune response to infection with NDV and related pathogens.

Newcastle disease virus (NDV) is a threat to the global poultry industry, but particularly for smallholder farmers in low- and middle-income countries. Previous reports suggest that some breeds of chickens are less susceptible to NDV infection, however, the mechanisms contributing to this are unknown. We here examined the comparative transcriptional responses of innate immune genes to NDV infection in inbred sublines of the Fayoumi and Leghorn breeds known to differ in their relative susceptibility to infection as well as at the microchromosome bearing the major histocompatability complex (MHC) locus. The analysis identified a set of five core genes, Mx1 , IRF1 , IRF7 , STAT1 , and SOCS1 , that are up-regulated regardless of subline. Several genes were differentially expressed in a breed- or subline-dependent manner. The breed-dependent response involved TLR3 , NOS2 , LITAF , and IFIH1 in the Fayoumi versus IL8 , CAMP , and CCL4 in the Leghorn. Further analysis identified subline-dependent differences in the pro-inflammatory response within the Fayoumi breed that are likely influenced by the MHC. These results have identified conserved, breed-dependent, and subline-dependent innate immune responses to NDV infection in chickens, and provide a strong framework for the future characterization of the specific roles of genes and pathways that influence the susceptibility of chickens to NDV infection.

Newcastle disease virus (NDV) causes substantial economic losses to smallholder farmers in low- and middle-income countries with high levels of morbidity and mortality in poultry flocks. Previous investigations have suggested differing levels of susceptibility to NDV between specific inbred lines and amongst breeds of chickens, however, the mechanisms contributing to this remain poorly understood. Studies have shown that some of these differences in levels of susceptibility to NDV infection may be accounted for by variability in the innate immune response amongst various breeds of poultry to NDV infection. Recent studies, in inbred Fayoumi and Leghorn lines, uncovered conserved, breed-dependent, and subline-dependent responses. To better understand the role of innate immune genes in engendering a protective immune response, we assessed the transcriptional responses to NDV of three highly outbred Tanzanian local chicken ecotypes, the Kuchi, the Morogoro Medium, and the Ching’wekwe. Hierarchical clustering and principal coordinate analysis of the gene expression profiles of 21-day old chick embryos infected with NDV clustered in an ecotype-dependent manner and was consistent with the relative viral loads for each of the three ecotypes. The Kuchi and Morogoro Medium exhibit significantly higher viral loads than the Ching’wekwe. The results show that the outbred ecotypes with increased levels of expression of CCL4, NOS2, and SOCS1 also had higher viral loads. The higher expression of SOCS1 is inconsistent with the expression in inbred lines. These differences may uncover new mechanisms or pathways in these populations that may have otherwise been overlooked when examining the response in highly inbred lines. Taken together, our findings provide insights on the specific conserved and differentially expressed innate immune-related genes involved the response of highly outbred chicken lines to NDV. This also suggests that several of the specific innate immunity related genes identified in the current investigation may serve as markers for the selection of chickens with reduced susceptibility to NDV.