A Systematic Analysis of miRNA Transcriptome in Marek’s Disease Virus-Induced Lymphoma Reveals Novel and Differentially Expressed miRNAs

Marek’s disease is a lymphoproliferative neoplastic disease of the chicken, which poses a serious threat to poultry health. Marek’s disease virus (MDV)-induced T-cell lymphoma is also an excellent biomedical model for neoplasia research. Recently, miRNAs have been demonstrated to play crucial roles in mediating neoplastic transformation. To investigate host miRNA expression profiles in the tumor transformation phase of MDV infection, we performed deep sequencing in two MDVinfected samples (tumorous spleen and MD lymphoma from liver), and two non-infected controls (non-infected spleen and lymphocytes). In total, 187 and 16 known miRNAs were identified in chicken and MDV, respectively, and 17 novel chicken miRNAs were further confirmed by qPCR. We identified 28 down-regulated miRNAs and 11 up-regulated miRNAs in MDVinfected samples by bioinformatic analysis. Of nine further tested by qPCR, seven were verified. The gga-miR-181a, gga-miR- 26a, gga-miR-221, gga-miR-222, gga-miR-199*, and gga-miR-140* were down-regulated, and gga-miR-146c was upregulated in MDV-infected tumorous spleens and MD lymphomas. In addition, 189 putative target genes for seven differentially expressed miRNAs were predicted. The luciferase reporter gene assay showed interactions of gga-miR-181a with MYBL1, gga-miR-181a with IGF2BP3, and gga-miR-26a with EIF3A. Differential expression of miRNAs and the predicted targets strongly suggest that they contribute to MDV-induced lymphomagenesis

Genome-wide identification of copy number variations between two chicken lines that differ in genetic resistance to Marek’s disease

Copy number variation (CNV) is a major source of genome polymorphism that directly contributes to phenotypic variation such as resistance to infectious diseases. Lines 63 and 72 are two highly inbred experimental chicken lines that differ greatly in susceptibility to Marek’s disease (MD), and have been used extensively in efforts to identify the genetic and molecular basis for genetic resistance to MD. Using next generation sequencing, we present a genome-wide assessment of CNVs that are potentially associated with genetic resistance to MD. Three chickens randomly selected from each line were sequenced to an average depth of 20×. Two popular software, CNVnator and Pindel, were used to call genomic CNVs separately. The results were combined to obtain a union set of genomic CNVs in the two chicken lines. A total of 5,680 CNV regions (CNVRs) were identified after merging the two datasets, of which 1,546 and 1,866 were specific to the MD resistant or susceptible line, respectively. Over half of the line-specific CNVRs were shared by 2 or more chickens, reflecting the reduced diversity in both inbred lines. The CNVRs fixed in the susceptible lines were significantly enriched in genes involved in MAPK signaling pathway. We also found 67 CNVRs overlapping with 62 genes previously shown to be strong candidates of the underlying genes responsible for the susceptibility to MD. Our findings provide new insights into the genetic architecture of the two chicken lines and additional evidence that MAPK signaling pathway may play an important role in host response to MD virus infection. The rich source of line-specific CNVs is valuable for future disease-related association studies in the two chicken lines.https://doi.org/10.1186/s12864-015-2080-

A genome-wide association study explores the genetic determinism of host resistance to Salmonella pullorum infection in chickens

International audienceAbstractBackgroundSalmonella infection is a serious concern in poultry farming because of its impact on both economic loss and human health. Chicks aged 20 days or less are extremely vulnerable to Salmonella pullorum (SP), which causes high mortality. Furthermore, an outbreak of SP infection can result in a considerable number of carriers that become potential transmitters, thus, threatening fellow chickens and offspring. In this study, we conducted a genome-wide association study (GWAS) to detect potential genomic loci and candidate genes associated with two disease-related traits: death and carrier state.MethodsIn total, 818 birds were phenotyped for death and carrier state traits through a SP challenge experiment, and genotyped by using a 600 K high-density single nucleotide polymorphism (SNP) array. A GWAS using a single-marker linear mixed model was performed with the GEMMA software. RNA-sequencing on spleen samples was carried out for further identification of candidate genes.ResultsWe detected a region that was located between 33.48 and 34.03 Mb on chicken chromosome 4 and was significantly associated with death, with the most significant SNP (rs314483802) accounting for 11.73% of the phenotypic variation. Two candidate genes, FBXW7 and LRBA, were identified as the most promising genes involved in resistance to SP. The expression levels of FBXW7 and LRBA were significantly downregulated after SP infection, which suggests that they may have a role in controlling SP infections. Two other significant loci and related genes (TRAF3 and gga-mir-489) were associated with carrier state, which indicates a different polygenic determinism compared with that of death. In addition, genomic inbreeding coefficients showed no correlation with resistance to SP within each breed in our study.ConclusionsThe results of this GWAS with a carefully organized Salmonella challenge experiment represent an important milestone in understanding the genetics of infectious disease resistance, offer a theoretical basis for breeding SP-resistant chicken lines using marker-assisted selection, and provide new information for salmonellosis research in humans and other animals

A Genome-Wide SNP Scan Reveals Novel Loci for Egg Production and Quality Traits in White Leghorn and Brown-Egg Dwarf Layers

Availability of the complete genome sequence as well as high-density SNP genotyping platforms allows genome-wide association studies (GWAS) in chickens. A high-density SNP array containing 57,636 markers was employed herein to identify associated variants underlying egg production and quality traits within two lines of chickens, i.e., White Leghorn and brown-egg dwarf layers. For each individual, age at first egg (AFE), first egg weight (FEW), and number of eggs (EN) from 21 to 56 weeks of age were recorded, and egg quality traits including egg weight (EW), eggshell weight (ESW), yolk weight (YW), eggshell thickness (EST), eggshell strength (ESS), albumen height(AH) and Haugh unit(HU) were measured at 40 and 60 weeks of age. A total of 385 White Leghorn females and 361 brown-egg dwarf dams were selected to be genotyped. The genome-wide scan revealed 8 SNPs showing genome-wise significant (P<1.51E-06, Bonferroni correction) association with egg production and quality traits under the Fisher's combined probability method. Some significant SNPs are located in known genes including GRB14 and GALNT1 that can impact development and function of ovary, but more are located in genes with unclear functions in layers, and need to be studied further. Many chromosome-wise significant SNPs were also detected in this study and some of them are located in previously reported QTL regions. Most of loci detected in this study are novel and the follow-up replication studies may be needed to further confirm the functional significance for these newly identified SNPs

Feather colour affects the aggressive behaviour of chickens with the same genotype on the dominant white (I) locus.

Aggression in chickens is a serious economic and animal welfare issue in poultry farming. Pigmentation traits have been documented to be associated with animal behaviour. Chicken pecking behaviour has been found to be related to feather colour, with premelanosome protein 17 (PMEL17) being one of the candidate genes. In the present study, we performed a genotypic and phenotypic association analysis between chicken plumage colour (red and white) and aggressive behaviour in an F1 hybrid group generated by crossing the autosomal dominant white-feathered breed White Leghorn (WL) and the red-feathered breed Rhode Island Red (RIR). In genetic theory, all the progeny should have white feathers because WL is homozygous autosomal dominant for white feathers. However, we found a few red-feathered female chickens. We compared the aggressiveness between the red and white females to determine whether the feather colour alone affected the behaviour, given that the genetic background should be the same except for feather colour. The aggressiveness was recorded 5 days after sexual maturity at 26 weeks. Generally, white plumage hens showed significantly higher aggressiveness compared to the red ones in chasing, attacking, pecking, and threatening behaviour traits. We assessed three candidate feather colour genes-PMEL17, solute carrier family 45 member 2 (SLC45A2), and SRY-box 10 (SOX10)-to determine the genetic basis for the red and white feather colour in our hybrid population; there was no association between the three loci and feather colour. The distinct behavioural findings observed herein provide clues to the mechanisms underlying the association between phenotype and behaviour in chickens. We suggest that mixing red and white chickens together might reduce the occurrence of aggressive behaviours

Active Yeast but Not Henhouse Environment Affects Dropping Moisture Levels in Egg-Laying Hens

Dropping moisture (DM) refers to the water content in feces. High DM negatively affects poultry production, environment, production costs, and animal health. Heredity, nutrition, environment, and disease may affect DM level. DM has medium inheritability and is related to cage height in henhouses. We examined the relationship among DM level, production performance, and environmental factors at different locations at the same henhouse height and effects of three types of additives. We measured the correlation between environmental factors including temperature, humidity, CO2 concentration, absolute pressure, and DM levels and laying performance of 934 Rhode Island Red hens. DM level was not significantly associated with environmental factors or production performance. We divided 64 persistently high DM hens into control and treatment groups supplied with different additives (probiotics, anisodamine, and antibiotics). DM levels, laying performance, egg quality, and serum biochemical indices were determined. Compared with the control and antibiotics, probiotics significantly reduced DM levels and eggshell strength while improving yolk color but did not significantly affect production performance. The additives reduced the b value of eggshell color; compared with probiotics, anisodamine decreased serum globulin levels. Exogenous active yeast supplementation can significantly reduce DM levels

Analysis of miRNA Expression Profiling of RIP2 Knockdown in Chicken HD11 Cells When Infected with Avian Pathogenic E. coli (APEC)

Colibacillosis is an acute and chronic avian disease caused by avian pathogenic E. coli (APEC). Previous studies have demonstrated that RIP2 plays a significant role in APEC infection. Moreover, increasing evidence indicates that microRNAs (miRNAs) are involved in host&ndash;pathogen interactions and the immune response. However, the role of miRNAs in the host against APEC infection remains unclear. Herein, we attempted to reveal new miRNAs potentially involved in the regulation of the immune and inflammatory response against APEC infection, with a particular focus on those possibly correlated with RIP2 expression, via miRNA-seq, RT-qPCR, Western blotting, dual-luciferase reporter assay, and CCK-8. The results showed that a total of 93 and 148 differentially expressed (DE) miRNAs were identified in the knockdown of RIP2 cells following APEC infection (shRIP2+APEC) vs. knockdown of RIP2 cells (shRIP2) and shRIP2 vs. wild-type cells (WT), respectively. Among those identified DE miRNAs, the biological function of gga-miR-455-5p was investigated. It was found that gga-miR-455-5p regulated by RIP2 was involved in the immune and inflammatory response against APEC infection via targeting of IRF2 to modulate the expression of type I interferons. Additionally, RIP2 could directly regulate the production of the type I interferons. Altogether, these findings highlighted the crucial role of miRNAs, especially gga-miR-455-5p, in host defense against APEC infection

Gene expression profiling of RIP2-knockdown in HD11 macrophages — elucidation of potential pathways (gene network) when challenged with avian pathogenic E.coli (APEC)

Background: Receptor interacting serine/threonine kinase 2 (RIP2), ubiquitous in many tissue/cell types, is the key regulator of immune and inflammatory responses for many diseases, including avian pathogenic E. coli (APEC), which causes a wide variety of localized or systemic infections. However, the molecular mechanisms by which RIP2 drives its transcriptional program to affect immune and inflammatory response upon APEC infection remains poorly understood. Results: In this study, RNA-seq and bioinformatics analyses were used to detect gene expression and new direct/indirect RIP2 targets in the treatments of wild type HD11 cells (WT), RIP2 knockdown cells (shRIP2), APEC stimulation cells (APEC), and RIP2 knockdown cells combined with APEC infection (shRIP2 + APEC). The results revealed that a total of 4691 and 2605 differentially expressed genes (DEGs) were screened in shRIP2 + APEC vs. APEC and shRIP2 vs. WT, respectively. Functional annotation analysis showed that apoptosis, MAPK, p53, Toll-like receptor, and Nod-like receptor signaling pathways were involved in APEC-induced RIP2 knockdown HD11 cells. By analyzing the enriched pathway and gene networks, we identified that several DEGs, including HSP90AB1, BID, and CASP9 were targeted by RIP2 upon APEC infection. Conclusion: As a whole, this study can not only provide data support for constructing gene networks of RIP2 knockdown with APEC challenge but also provide new ideas for improving the immune and inflammatory response.This article is published as Sun, H., Yang, Y., Cao, Y. et al. Gene expression profiling of RIP2-knockdown in HD11 macrophages — elucidation of potential pathways (gene network) when challenged with avian pathogenic E.coli (APEC). BMC Genomics 23, 341 (2022). https://doi.org/10.1186/s12864-022-08595-5. Posted with permission. This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/