Study On Pituitary Gland And Intestinal Of Zhedong White Goose During Different Seasons And Reproductive Periods Based On Omics Technology

The goose(Answer cygnoides)is herbivorous waterfowl,and well known for its strong adaptability,rapid growth,rich nutrients and low input requirement.However,the seasonal reproduction and strong broodiness resulted in the low laying performance,which directly affect the number of offspring breeding,and hindered the goose industry development.In this study,Zhedong white geese,a typical seasonal breeding poultry,were taken as the research object,and two experimental studies were carried out by using high-throughput sequencing techniques such as transcriptome sequencing(RNA-seq),whole genome bisulfite sequencing(WGBS)and 16S rDNA sequencing.This research includes two aspects:first,geese were studied in different seasons of pituitary gland mRNA expression level and DNA methylation level changes at whole genome level,preliminarily explored the related genes and mechanisms involved in regulating the seasonal reproduction of Zhedong white goose.In addition,the differences of pituitary gland transcriptome expression,as well as microbiota characteristics of intestinal of geese at different reproductive periods during breeding season were studied,aiming to explore the key genes and mechanisms that regulate the broodiness,and the synergistic relationship between the intestinal microbiota and reproductive status.The results are as follow:Through the transcriptome sequencing analysis of pituitary samples at the autumn equinox(AE),winter solstice(WS),spring equinox(SE)and summer solstice(SS),we finally obtained more than 7.4G(gigabases)of available data from each sample.Pairwise comparisons WS-vs-AE,SE-vs-WS,SS-vs-SE and AE-vs-SS resulted in 1139,33,704 and 3503 differently expressed genes(DEGs),respectively(P_adjusted<0.05),including reproduction genes(GnRHR?PRLR?ESR2 and VIPR),circadian clock genes(Clock,Per2,Npas2,Bmal2 and NR1D2),and transcription factors(POU1F1,Pitx2,NR5A1,NR4A2 and SREBF2).GO and KEGG enrichment analysis of DEGs revealed that they were mainly involved in biological processes such as 'response to oxidative stress','oxidation-reduction process','response to steroid hormone' and 'hormone-mediated signaling pathway',and interact with GnRH signaling pathway,and steroid biosynthesis.The pituitary glands methylation maps of 12 Zhedong white geese from four seasons were conducted through WGBS analysis.The raw data exceeding the genome coverage of 17× was obtained,in which the C site coverage exceeded 8×.There were 10476,83 3 6,21693,23989 differentially methylated regions(DMRs)between WS-vs-AE,SE-vs-WS,SS-vs-SE and AE-vs-SS,respectively,which were mainly distributed in gene intron region when mapped to genome.The differentially methylated genes were significantly enriched in molecular function such as 'organic cyclic compound binding','nucleic acid binding',and biological process like‘metabolic process',‘macromolecule biosynthesis process' and‘gene expression'.Pathway enrichment analysis revealed that MAPK signaling pathway,FOXO signaling pathway and calcium signaling pathway may played an important role in regulating seasonal reproduction.We delineated the transcriptome of Zhedong white goose pituitary glands at laying(ZDL),broody(ZDB)and recovery(ZDR)periods using RNA-seq,and the 9 samples produced a total of 535,239,542 clean reads,which were mapped to 16,151 genes.Differences in gene expression between reproduction periods were examined,and 582,979,33 differentially expressed genes(DEGs)were screened by pairwise comparisons of ZDB-vs-ZDL,ZDR-vs-ZDB and ZDR-vs-ZDL,respectively(fold change>1.5,P<0.05).GO analysis of the DEGs revealed significant enrichment in 'calcium ion binding','secretory granule'and‘G-protein receptor activity'.Pathways the neuroactive ligand-receptor interaction,protein processing in endoplasmic reticulum and calcium signaling pathway were crucial for the control of broodiness of geese.Further analysis revealed that genes PRL,TSH?,THR? and GnRHR2 may act as central modulators of hormonal.and behavioral regulation mechanism associated with broodiness.16S rDNA sequencing was performed to characterize the duodenal microbial communities in Zhedong white geese at different reproductive periods(laying,broody and recovery).A total of 4,748 OTUs were obtained.At the phylum level,in laying geese,Firmicutes accounted for the highest proportion(52.29%),followed by Actinobacteria(15.99%)and Proteobacteria(12.11%);Cyanobacteria(45.69%),Firmicutes(27.29%)and Proteobacteria(18.74%)were the dominant bacteria in broody geese;the microbial communities of the recovery geese were mainly composed of Spirochaetes(42.01%),Firmicutes(22.52%)and Cyanobacteria(18.61%).Additionally,the microbial richness and diversity of laying group were significantly higher than other groups(P<0.05).Principal coordinate analysis and similarity analysis showed that there were significant differences in the structure of the microbial community among three groups(P<0.05).