Metagenomics And Transcriptomics Reveal The Genetic Mechanism Of Abdominal Fat Deposition In Duck

China is one of big countries yielding and consumering duck in the world.The production of duck meat has steadily occupied the first place in the world.As the growth rate of meat ducks increased significantly,the deposition of abdominal fat is also increasing,which reduces the feed conversion rate and growth performance of ducks and affects ducks' health status.Therefore,from the perspective of production performance and health,it is a key objective for meat duck breeding to reduce duck abdominal fat deposition;meanwhile,revealing the genetic mechanism of abdominal fat deposition of meat ducks has become a hot spot for domestic and international research.In this study,the F2 generation resource population of the cross between Cherry Valley Peking Duck Progenitor and Runzhou crested white duck was used as the research object.The correlation between the phenotypes was determined by collecting and analyzing the data of body weight,abdominal fat weight and abdominal fat rate of 304 42-day F2 generation ducks.16S rRNA sequencing was performed on the intestinal contents of five segments-duodenum(SC),jejunum(KC),ileum(AC),caecum(MC)and rectum(ZC)of F2 generation ducks with high abdominal fat rate(HF)and low abdominal fat rate(LF),to identify the intestinal flora that affects abdominal fat deposition.The metagenomic analysis of cecal contents of duck with high and low abdominal fat rates was conducted to find the differential genes affecting abdominal fat deposition,and the functional enrichment analysis of the differential genes was conducted to explore the response mechanism of microorganisms to abdominal fat deposition.In addition,the transcriptome analysis of the abdominal fat of F2 generation ducks with high and low abdominal fat rate was performed to screen the differentially expressed mRNAs and lncRNAs affecting abdominal fat deposition.Finally,the results of 16S rRNA,metagenome and transcriptome analysis were combined to screen the key flora and genes that affect fat deposition and perform preliminary functional validation.The main results are as follows:(1)By analyzing the phenotypic data of body weight,abdominal fat weight and abdominal fat percentage of 304 ducks,the results showed that the abdominal fat percentage of ducks in the F2 generation resource population had individual differences and conformed to a normal distribution,with the lowest value of 0.217%and the highest value of 2.255%,and the mean value of 1.17%.Analysis of the effect of gender on abdominal fat percentage showed that there were more female ducks in the high abdominal lipid rate range(1.5%,2.25%)with a male to female ratio of 0.5313,and more male ducks in the low abdominal lipid rate range(0,0.75%),which indicated that the distribution of abdominal lipid rate in F2 generation ducks was related to sex.Body weight was significantly positively correlated with abdominal fat weight(P<0.05),and abdominal fat weight was significantly positively correlated with abdominal fat rate(P<0.05).Abdominal fat weight and body weight will affect the abdominal fat rate of ducks.(2)OTU clustering results of 16S rRNA sequencing from five intestinal sites showed that showed that MC-L had the highest number of OTUs,followed by ZC-L and ZC-H,and SC-H had the lowest number of OTUs.Alpha diversity analysis showed that several groups of MC-L,MC-H,SC-L,ZC-L and KC-L had higher flora diversity.Beta diversity results showed that intra-group concordance was better for each sample from MC site,followed by HC,ZC and SC,and the worst concordance was found in KC.Enrichment results of KEGG pathway showed that microorganisms from MC site were more enriched in the pathway of carbohydrate metabolism compared to other groups.(3)Metagenomic sequencing of cecum contents revealed 18 microbial genes enriched in the pathway related to lipid metabolism,including PKS9/KSA1?fab I?pqsD?bgsB?fabD?relA?AvrBs2?gigX3?fabGl and glpD.LDA Effect Size(LEfSe)analysis showed that the bacteria with differences among different groups were related to fat metabolism:Bacteroidetes,Firmicutes,Ruminococcus,Lachnospiraceae,Desulfovibrionales,Deltaproteobacteria,Actinobacteria and other 12 groups of bacteria.(4)Transcriptome sequencing of duck abdominal lipid tissues with high and low abdominal lipid rates identified 633 differentially expressed mRNAs and 214 differentially expressed lnCRNAs.Key differential genes such as FOXO1,FASN,ELOVL6,EHHADH,PPARA,ACSL1,PLA2G4B,SCD1 and key differential lncRNAs such as XR₀03493025.1,XR₀03499737.1,TCONS₀0005095,TCONS₀0013863,XR₀02405656.2,TCONS₀0050591 and TCONS₀0059689.were screened by GO and KEGG enrichment analysis.Bacteroidetes in duck's cecum are negatively correlated with down-regulated genes TMF1,STRBP,TRRAP and LAMA3.Firmicutes are positively correlated with the above 4 down-regulated genes,and also have positive correlations with MANBAL and Fam234a.Therefore,it can be speculated that the above bacterial community interacts with the above differential genes to jointly regulate the deposition of abdominal fat.(5)A preliminary functional validation of the FOXO1 gene in ducks showed that the relative mRNA expression of FOXO1 gene in the abdominal fat tissue was significantly different between the high and low abdominal fat rate groups(P<0.01),and the relative expression level of FOXO1 gene in the high abdominal fat rate group was lower than that of the low abdominal fat rate group.FOXO1 gene may play a negative role in regulating fat deposition in ducks.In conclusion,there are individual differences in the abdominal fat rate of the F2 generation ducks of the cherry valley Peking duck ancestor and Runzhou Crested White Duck.Both abdominal fat weight and body weight will affect the abdominal fat rate of ducks.There were significant differences in microbial structure and function in different parts of intestinal tract,and the cecum might be the key part affecting abdominal fat deposition in ducks.The differences of Bacteroidetes,Firmicutes,Actinobacteria and other bacteria affected the expression of FOXOl,FASN,EHHADH,ACSBG2,AASDH and PPARA pathway genes involved in fat metabolism,which further affects the synthesis and breakdown of fat.In addition,FOXO1 gene plays a negative regulatory role in duck abdominal fat deposition,and the identified SNP locus(g.59359C>T)can be used as a molecular marker affecting duck abdominal fat deposition in ducks.The above research results provide a theoretical basis for improving duck abdominal fat deposition performance.