Expression data from broiler chickens in response to the combined stressors of catching, transport and slaughter
ABSTRACT: The process of commercial catching, transport and slaughter (CTS) is known to be an acute stressful event in broiler chickens. Corticosteroid concentrations increase, impacting measures of IGF-1, growth hormone and metabolites of the immune system from blood plasma samples. We used ARK-Genomics chicken 20K oligo array, a two channel DNA microarray, to investigate the significantly differentially expressed genes in the livers of chickens during CTS. Overall design: We investigate the differences of gene expression profiles in hepatic tissues between control birds (n=10) and birds experiencing CTS (n=10) using an ARK-Genomics chicken 20K oligo array, a two channel DNA array (http://www.ark-genomics.orgmicroarray) with full dye swap.
INSTRUMENT(S): ARK-Genomics G. gallus 20K v2 (single gene version)
Project description:The process of commercial catching, transport and slaughter (CTS) is known to be an acute stressful event in broiler chickens. Corticosteroid concentrations increase, impacting measures of IGF-1, growth hormone and metabolites of the immune system from blood plasma samples. We used ARK-Genomics chicken 20K oligo array, a two channel DNA microarray, to investigate the significantly differentially expressed genes in the livers of chickens during CTS. We investigate the differences of gene expression profiles in hepatic tissues between control birds (n=10) and birds experiencing CTS (n=10) using an ARK-Genomics chicken 20K oligo array, a two channel DNA array (http://www.ark-genomics.orgmicroarray) with full dye swap.
Project description:BACKGROUND: High throughput gene expression studies using oligonucleotide microarrays depend on the specificity of each oligonucleotide (oligo or probe) for its target gene. However, target specific probes can only be designed when a reference genome of the species at hand were completely sequenced, when this genome were completely annotated and when the genetic variation of the sampled individuals were completely known. Unfortunately there is not a single species for which such a complete data set is available. Therefore, it is important that probe annotation can be updated frequently for optimal interpretation of microarray experiments. RESULTS: In this paper we present OligoRAP, a pipeline to automatically update the annotation of oligo libraries and estimate oligo target specificity. OligoRAP uses a reference genome assembly with Ensembl and Entrez Gene annotation supplemented with a set of unmapped transcripts derived from RefSeq and UniGene to handle assembly gaps. OligoRAP produces alignments of each oligo with the reference assembly as well as with unmapped transcripts. These alignments are re-mapped to the annotation sources, which results in a concise, as complete as possible and up-to-date annotation of the oligo library. The building blocks of this pipeline are BioMoby web services creating a highly modular and distributed system with a robust, remote programmatic interface.OligoRAP was used to update the annotation for a subset of 791 oligos from the ARK-Genomics 20 K chicken array, which were selected as starting material for the oligo annotation session of the EADGENE/SABRE Post-analysis workshop. Based on the updated annotation about one third of these oligos is problematic with regard to target specificity. In addition, the accession numbers or ids the oligos were originally designed for no longer exist in the updated annotation for almost half of the oligos. CONCLUSION: As microarrays are designed on incomplete data, it is important to update probe annotation and check target specificity regularly. OligoRAP provides both and due to its design based on BioMoby web services it can easily be embedded as an oligo annotation engine in customised applications for microarray data analysis. The dramatic difference in updated annotation and target specificity for the ARK-Genomics 20 K chicken array as compared to the original data emphasises the need for regular updates.
Project description:The family concerns 46 offspring of the sire BC1 n°1940 (produced from an F1 male obtained by-intercrossing two fat (FL) and lean (LL) meat-type chicken lines that divergently selected on fatness [Leclercq et al. 1980]) mated with 8 LL females. Hepatic transcriptomic profiles were obtained using the chicken 20K oligochip (ARK-genomics). The 46 chickens were fed ad libitum using a conventional starter diet (0-3 weeks: 12.8 MJ of metabolizable energy) and then a growing broiler diet (4-9 weeks: 13.0 Mj of metabolizable energy). Light/dark periods were 24h light for the first 2 days, then 14h light/10h night up to slaughtering. At 4 weeks of age, blood samples were collected for DNA extraction and genotyping. At 9 weeks of age, the birds were fed ad libitum for 4 hours minimum after overnight fasting and then weighed and sacrificed by electrical stunning in the experimental processing plant (Station de Recherches Avicoles, Inra, Nouzilly, France). Following sacrifice, livers were collected, quickly frozen in liquid nitrogen and stored at −80°C until RNA extraction for transcriptome analyses. Overall design: Five micrograms of total RNA from each sample were labeled with Cy5 and then hybridized to the microarray with a same Cy3-labeled reference probe. The reference RNA pool was made from an equal amount of RNA derived from all liver samples.
Project description:The family concerns 44 offspring of the sire BC1 n°1998 (produced from a F1 male obtained by-intercrossing two fat (FL) and lean (LL) meat-type chicken lines that divergently selected on fatness [Leclercq et al 1980] mated with 8 LL females. Hepatic transcriptomic profiles were obtained using a chicken 20K oligochips (ARK-genomics). The 44 chickens were fed ad libitum using a conventional starter diet (0-3 weeks: 12.8 MJ of metabolizable energy) and then a growing broiler diet (4-9 weeks: 13.0 Mj of metabolizable energy). Light/dark periods were 24h light for the first 2 days, then 14h light/10h night up to slaughtering. At 4 weeks of age, blood samples were collected for DNA extraction and genotyping. At 9 weeks of age, the birds were fed ad libitum for 4 hours minimum after overnight fasting and then weighed and sacrificed by electrical stunning in the experimental processing plant (Station de Recherches Avicoles, Inra, Nouzilly, France). Following sacrifice, livers were collected, quickly frozen in liquid nitrogen and stored at −80°C until RNA extraction for transcriptome analyses Overall design: Five micrograms of total RNA from each sample were labelled with Cy5 and then hybridized to the microarray with a same Cy3-labeled reference probe. The reference RNA pool was made from an equal amount of RNA derived from all liver samples.
Project description:The avian coronavirus infectious bronchitis virus (IBV) S1 subunit of the spike (S) glycoprotein mediates viral attachment to host cells and the S2 subunit is responsible for membrane fusion. Using IBV Arkansas-type (Ark) S protein histochemistry, we show that extension of S1 with the S2 ectodomain improves binding to chicken tissues. Although the S1 subunit is the major inducer of neutralizing antibodies, vaccination with S1 protein has been shown to confer inadequate protection against challenge. The demonstrated contribution of S2 ectodomain to binding to chicken tissues suggests that vaccination with the ectodomain might improve protection compared to vaccination with S1 alone. Therefore, we immunized chickens with recombinant trimeric soluble IBV Ark-type S1 or S-ectodomain protein produced from codon-optimized constructs in mammalian cells. Chickens were primed at 12days of age with water-in-oil emulsified S1 or S-ectodomain proteins, and then boosted 21days later. Challenge was performed with virulent Ark IBV 21days after boost. Chickens immunized with recombinant S-ectodomain protein showed statistically significantly (P<0.05) reduced viral loads 5days post-challenge in both tears and tracheas compared to chickens immunized with recombinant S1 protein. Consistent with viral loads, significantly reduced (P<0.05) tracheal mucosal thickness and tracheal lesion scores revealed that recombinant S-ectodomain protein provided improved protection of tracheal integrity compared to S1 protein. These results indicate that the S2 domain has an important role in inducing protective immunity. Thus, including the S2 domain with S1 might be promising for better viral vectored and/or subunit vaccine strategies.
Project description:BACKGROUND: The development of microarray resources for the chicken is an important step in being able to profile gene expression changes occurring in birds in response to different challenges and stimuli. The creation of an immune-related array is highly valuable in determining the host immune response in relation to infection with a wide variety of bacterial and viral diseases. RESULTS: Here we report the development of chicken immune-related cDNA libraries and the subsequent construction of a microarray containing 5190 elements (in duplicate). Clones on the array originate from tissues known to contain high levels of cells related to the immune system, namely Bursa, Peyers patch, thymus and spleen. Represented on the array are genes that are known to cluster with existing chicken ESTs as well as genes that are unique to our libraries. Some of these genes have no known homologies and represent novel genes in the chicken collection. A series of reference genes (ie. genes of known immune function) are also present on the array. Functional annotation data is also provided for as many of the genes on the array as is possible. CONCLUSION: Six new chicken immune cDNA libraries have been created and nearly 10,000 sequences submitted to GenBank [GenBank: AM063043-AM071350; AM071520-AM072286; AM075249-AM075607]. A 5 K immune-related array has been developed from these libraries. Individual clones and arrays are available from the ARK-Genomics resource centre.
Project description:Despite reducing the prevalent foodborne pathogen Campylobacter jejuni in chickens decreases campylobacteriosis, few effective approaches are available. The aim of this study was to use microbial metabolic product bile acids to reduce C. jejuni chicken colonization. Broiler chicks were fed with deoxycholic acid (DCA), lithocholic acid (LCA), or ursodeoxycholic acid (UDCA). The birds were also transplanted with DCA modulated anaerobes (DCA-Anaero) or aerobes (DCA-Aero). The birds were infected with human clinical isolate C. jejuni 81-176 or chicken isolate C. jejuni AR101. Notably, C. jejuni 81-176 was readily colonized intestinal tract at d16 and reached an almost plateau at d21. Remarkably, DCA excluded C. jejuni cecal colonization below the limit of detection at 16 and 28 days of age. Neither chicken ages of infection nor LCA or UDCA altered C. jejuni AR101 chicken colonization level, while DCA reduced 91% of the bacterium in chickens at d28. Notably, DCA diet reduced phylum Firmicutes but increased Bacteroidetes compared to infected control birds. Importantly, DCA-Anaero attenuated 93% of C. jejuni colonization at d28 compared to control infected birds. In conclusion, DCA shapes microbiota composition against C. jejuni colonization in chickens, suggesting a bidirectional interaction between microbiota and microbial metabolites.
Project description:BACKGROUND: Infectious laryngotracheitis virus (ILTV; gallid herpesvirus 1) infection causes high mortality and huge economic losses in the poultry industry. To protect chickens against ILTV infection, chicken-embryo origin (CEO) and tissue-culture origin (TCO) vaccines have been used. However, the transmission of vaccine ILTV from vaccinated- to unvaccinated chickens can cause severe respiratory disease. Previously, host cell responses against virulent ILTV infections were determined by microarray analysis. In this study, a microarray analysis was performed to understand host-vaccine ILTV interactions at the host gene transcription level. RESULTS: The 44 K chicken oligo microarrays were used, and the results were compared to those found in virulent ILTV infection. Total RNAs extracted from vaccine ILTV infected chicken embryo lung cells at 1, 2, 3 and 4 days post infection (dpi), compared to 0 dpi, were subjected to microarray assay using the two color hybridization method. Data analysis using JMP Genomics 5.0 and the Ingenuity Pathway Analysis (IPA) program showed that 213 differentially expressed genes could be grouped into a number of functional categories including tissue development, cellular growth and proliferation, cellular movement, and inflammatory responses. Moreover, 10 possible gene networks were created by the IPA program to show intermolecular connections. Interestingly, of 213 differentially expressed genes, BMP2, C8orf79, F10, and NPY were expressed distinctly in vaccine ILTV infection when compared to virulent ILTV infection. CONCLUSIONS: Comprehensive knowledge of gene expression and biological functionalities of host factors during vaccine ILTV infection can provide insight into host cellular defense mechanisms compared to those of virulent ILTV.
Project description:From infection studies with cultured chicken cells and experimental mammalian hosts, it is well known that influenza viruses use the nonstructural protein 1 (NS1) to suppress the synthesis of interferon (IFN). However, our current knowledge regarding the in vivo role of virus-encoded NS1 in chickens is much more limited. Here, we report that highly pathogenic avian influenza viruses of subtypes H5N1 and H7N7 lacking fully functional NS1 genes were attenuated in 5-week-old chickens. Surprisingly, in diseased birds infected with NS1 mutants, the IFN levels were not higher than in diseased birds infected with wild-type virus, suggesting that NS1 cannot suppress IFN gene expression in at least one cell population of infected chickens that produces large amounts of the cytokine in vivo. To address the question of why influenza viruses are highly pathogenic in chickens although they strongly activate the innate immune system, we determined whether recombinant chicken alpha interferon (IFN-?) can inhibit the growth of highly pathogenic avian influenza viruses in cultured chicken cells and whether it can ameliorate virus-induced disease in 5-week-old birds. We found that IFN treatment failed to confer substantial protection against challenge with highly pathogenic viruses, although it was effective against viruses with low pathogenic potential. Taken together, our data demonstrate that preventing the synthesis of IFN is not the primary role of the viral NS1 protein during infection of chickens. Our results further suggest that virus-induced IFN does not contribute substantially to resistance of chickens against highly pathogenic influenza viruses.
Project description:Previous studies revealed that Igf2 and Mpr/Igf2r are imprinted in eutherian mammals and marsupials but not in monotremes or birds. Igf2 lies in a large imprinted cluster in eutherians, and its imprinting is regulated by long-range mechanisms. As a step to understand how the imprinted cluster evolved, we have determined a 490-kb chicken sequence containing the orthologs of mammalian Ascl2/Mash2, Ins2 and Igf2. We found that most of the genes in this region are conserved between chickens and mammals, maintaining the same transcriptional polarities and exon-intron structures. However, H19, an imprinted noncoding transcript, was absent from the chicken sequence. Chicken ASCL2/CASH4 and INS, the orthologs of the imprinted mammalian genes, showed biallelic expression, further supporting the notion that imprinting evolved after the divergence of mammals and birds. The H19 imprinting center and many of the local regulatory elements identified in mammals were not found in chickens. Also, a large segment of tandem repeats and retroelements identified between the two imprinted subdomains in mice was not found in chickens. Our findings show that the imprinted genes were clustered before the emergence of imprinting and that the elements associated with imprinting probably evolved after the divergence of mammals and birds.