Brief Summary of Minutes of Annual Meeting

Objective 1: Identify and characterize genes and their relationships to disease resistance in poultry with an emphasis on the major histocompatibility complex as well as other genes encoding alloantigens, communication molecules and their receptors and other candidate systems.

3.1.1 Genomics of response to avian pathogenic E. coli (APEC)
In this project, we are analyzing host transcriptional response to vaccination and/or infection with avian pathogenic E. coli (APEC). Our overall objective is to identify genes, signaling pathways and biological networks associated with infection and resistance to APEC in chickens. Broiler chicks were experimentally vaccinated at 2 weeks of age and challenged at 4 weeks of age with APEC. At 1 and 5 days post-infection, whole blood was collected and peripheral blood leukocytes isolated. Gross lesions in the air sacs, pericardium and liver at necropsy were used to assign pathology category, as either mild or severe, to the non-vaccinated, challenged chicks. Therefore, ten treatment groups were classified by a priori factors of vaccination status, challenge status, day post-infection, and the a posteriori factor of pathology severity within the non-vaccinated, challenged groups. In recent years, in cooperation with H. Zhou (UC-Davis), we have conducted and published microarray analyses of the spleen and peripheral WBC transcriptome evaluated utilizing a chicken 44K Agilent microarray (Sandford et al. 2011; 2012; 2013) and in cooperation with Qinghua Nie (South China Agricultural University) analysis of Solexa sequencing of spleen (Nie et al. 2012).

Our current phase of the APEC study is deep-sequence based transcriptome analyses of the primary immune tissues, conducted as part of the dissertation studies of H. Sun. This focus on the primary lymphoid tissues will give novel insights into the earliest transcriptional changes induced in immune-system cells in response to bacterial infections.

From four individuals of each of the six treatment groups that were not vaccinated, the mRNA has been isolated from bursa, thymus and bone marrow. From these, individual libraries were constructed, and subsequent deep sequencing was done. All the individuals have good read quality. as the quality score is more than 32 in FASTQC. Alignment was then done by using TopHat2 and SAMtool. Thousands of genes in total were detected as differentially expressed for the six treatments in each tissue, using edgeR. Generally, APEC infection resulted in more up-regulation than down-regulation in all three tissues, which is consistent with the results of the first phase of APEC study on spleen and WBC. In bone marrow, thymus, and bursa, hundreds to thousands of differentially expressed genes were found over time in challenged severe lesion groups (CS5 vs. CS1), and contrasting challenged birds with severe lesions with other groups, within day (CS1 vs. NC1, CS5 vs. CM5 and CS5 vs. NC5). Also, many genes in three contrasts examining pathology and infection status (CS5 vs. NC5, CS5 vs. and CM5, CS5 vs. CS1) were significantly differentially expressed in all three tissues. Little difference was seen between infected birds with mild lesions and the non-infected groups at either time.

In bone marrow and thymus, hundreds of differentially expressed genes existed in challenged birds with severe lesions relative to control, and severe relative to mild, over time; which suggests interaction between treatment and time. However, almost no differentially expressed genes were found in bursa in mild relative to control, in severe relative to control, and severe relative to mild over time, suggesting that no interaction exists between treatment and time in bursa. Analysis of expressed genes by GO term and pathway analysis is commencing.

3.2 Objective 2: Identify and characterize environmental, dietary and physiologic factors that modulate immune system development, optimal immune function and immune system related disease resistance and welfare in poultry genetic stocks.

3.2.1 Interaction of heat stress and inflammatory response in poultry Within a USDA-AFRI funded Climate Change project (PD: C. Schmidt, UDEL), we are investigating the interaction of two putative stressors: heat stress and exposure to an inflammation-inducing PAMP (LPS). Birds of two genetic lines (broiler, Fayoumi) that were either exposed to daily cyclic heat episodes or kept at control temperatures were injected with either LPS or saline. Measures of body temperature and blood gas parameters were taken on the live birds, and tissues were collected for future analyses of transcriptomes. This will be part of the dissertation studies of A. Bjorkquist. In the past year, only the experimental design and planning was accomplished for the use of the already collected tissues.

3.3 Objective 3: Develop, evaluate and characterize methodologies, reagents and genotypes to assess immune function and disease resistance to enhance production efficiency through genetic selection in poultry.

3.3.1 Genetic population development, maintenance, and characterization Iowa State University maintains 13 chicken genetic lines. In the past year, they were reproduced in one generation. The ISU genetic lines are of two basic genetic structures:

(a) highly inbred lines or (b) advanced intercross lines (AIL). Highly inbred lines (60 – 100 generations of sib-mating) of defined MHC type were maintained, with the inbreeding of the earliest line starting in 1925. Lines are primarily of egg-type origin, but also include the non-commercial Fayoumi and Spanish lines, and one broiler line. Birds of the MHC-defined inbred lines were serologically typed each generation with line-specific anti-erythrocyte antisera to verify line purity (about 800 birds each generation); all birds are typed as chicks and the potential breeders are typed a second time before mating takes place. A non-inbred broiler breeder line is also maintained, as well as two advanced intercross lines (now at generation F21) initiated by crossing outbred broiler males with females of two distinct, highly inbred lines (Leghorn and Fayoumi).

Cheng, H.H., Kaiser, P., and Lamont, S.J. 2013. Integrated genomic approaches to enhance genetic resistance in chickens. Annu. Rev. Anim. Biosci. 2013. 1:239–260

Coble, D.J., Sandford, E. E., Ji, T., Abernathy, J., Fleming, D., Zhou, H., and Lamont, S.J. 2013. Impacts of Salmonella enteritidis infection on liver transcriptome in broilers. Genesis 51:357–364

BOOKS AND CHAPTERS IN BOOKS:

Cheng, H.H. and Lamont, S.J. 2013 Genetics of disease resistance. Pp. 70-86. In: Diseases of Poultry. 13th ed. D. E. Swayne, J.R. Glisson, L.R. McDougald, V. Nair, L. Nolan, and D.L. Suarez, Eds. Wiley-Blackwell, Ames

Lamont, S.J., Dekkers, J.C.M., and Zhou, H. 2014. Immunogenetics and mapping immunological functions. Pp. 205-221. In: Avian Immunology. F. Davison, B. Kaspars, P. Kaiser, K.A. Schat, Eds., Elsevier, London, San Diego

PUBLISHED ABSTRACTS:

Wang, Y., Li, J., Li, Q., Hu, X., Li, N. Hu, S., Brahmakshatriy, V., Lupiani, B., Reddy, S., Lamont, S.J., and Zhou, H. 2013. Effects of avian influenza virus infection on the transcriptome and the DNA methylome in two genetically distinct chicken lines using next generation sequencing. Plant & Animal Genome XXI, January 2013, San Diego, CA

Wang, Y., Lupiani, B., Reddy, S., Wang, H., Chen, R., Lamont, S.J., and Zhou, H. 2013. Lung transcriptome following avian influenza virus infection in two genetically distinct chicken inbred lines using RNA-seq. Epigenetics Conference, June 2013, Japan

Zhou, H.J., Wang, Y., Lamont, S.J., and Ross, P. 2013. Re-annotation of chicken genome using RNA-seq data. Proc. Poultry Sci. Ann. Mtg. San Diego, CA

Zavelo. A.E., Schmidt, C.J., Rothschild, M.F., Persia, M.E., Lamont, S.J., and Ashwell, C.M. 2013. Major histocompatibility complex