Research On Metabonomics And Transcriptomics Of Ascites Syndrome In Broiler Chickens

Ascites syndrome (AS) is a nutritional metabolic disease of poultry The previous researches on AS mainly focused on histopathologic, blood and biochemical parameters, oxidative stress and the expression of single gene or protein. There is no report of systemic researches on ascites pathogenesis using high-throughput omics technology. Through establishing AS model and AS-resistant model, the metabolic biomarkers and the differentially expressed genes (DEGs) of AS broilers and AS-resistant broilers were decteted using ultra-performance liquid chromatography coupled with quadruple time-of-flight high-sensitivity mass spectrometry (UPLC-QTOF/HSMS) metabolomic and RNA-seq transcriptomicExperiment1was designed to investigate the DEGs and metabolism biomarkers of AS broilers. A total of2160-day-old male Ross308broilers were randomLy assigned to two groups (control group and low-temperature treatment) with six replicates of eighteen broilers each. The serum of6healthy chickens from the control group and6AS chickens from the AS group were selected for the metabolomic profile analysis, while the liver tissues of3healthy chickens from the control group and3AS chickens from the AS group were selected for the transcriptomic profile analysis at21and35days. Results:(1) Compared with the normal group, the average F/G and AS morbility were significantly increased, while the ADG were significantly decreased in the AS group. Meanwhile, the hematocrit, heart index, ascites heart index and liver index were significantly increased in21and35days in the AS group. The impact of AS on pulmonary vascular remodeling was shown up in the significantly increased thickness of pulmonary vascular in external diameter range from20to50mm,50to100mm and100to200mm in both21and35days.(2) Metabolomic results revealed that in the AS group the concentrations of PC(20:1/18:3),PC(14:1/22:1),PC(22:4/14:0),PC(20:4/18:0)and LysoPC(18:0) were up-regulated, while dihydroxyacetone, ursodeoxycholic acid, cycloserine, Indoleacrylic acid, Methyl amyl armillaria, hypoxanthine, inosine, LysoPC(20:4)、LysoPC(16:0)、LysoPC(18:2) were down-regulated in21days. In35days, concentrations of dihydroxyacetone, ursodeoxycholic acid, cycloserine, Indoleacrylic acid, valine and tryptophan were down-regulated, while LysoPC(20:4)、 LysoPC(16:0)、LysoPC(18:0)、LysoPC(18:1)、LysoPC(18:2)、PC(14:1/20:1、PC(20:4/18:0)、 PC(14:1/22:1) were up-regulated.(3) Upon comparison with the control group, a total of287and390genes were identified as DEGs in21and35days respectively, among which90genes were up-regulated and197genes were down-regulated in21days, while212genes were up-regulated and178genes were down-regulated in35days in the AS group. DEGs in21days mainly significantly down-regulated lipid metabolism, and up-regulated the production of precursors metabolites and energy, the signal path of adipose cytokines. DEGs in35days mainly significantly down-regulated lipid metabolism, cell differentiation and enzyme linked receptor protein signaling pathway, up-regulated protein polymerization, among which the GO:0006629-lipid metabolic process had the maximum numbers of enrichment genes (n=9),GO:0008203-cholesterol metabolic process had the lowest p value of fold enrichment (0.000966) while GO:0006695-cholesterol biosynthetic process had the highest fold enrichment (46.67).Those resultes indicated that there would be some metabolism disorders of lipid, protein and glucose, especially glycerophospholipid, while genes of lipid metabolism were changed, especially genes of steroid.Experiment2was carried out to investigate the resistant DEGs and metabolism biomarkers of resistant AS broilers. A total of1080-day-old male Ross308broilers were randomLy assigned six replicates of eighteen broilers each. The serum of6ascites chickens from the susceptible group and6ascites resistant chickens from the resistant group were selected for the metabolomic profile analysis, while the liver tissues of3healthy chickens from the resistant group and3AS chickens from the susceptible group were selected for the transcriptomic profile analysis at21and35days. Results:(1) Compared with the susceptible group, the hematocrit of resistant group were significantly decreased in21and35days (P<0.01), the AHI significantly increased in35days (P<0.01). Meanwhile, the thickness of pulmonary vascular significantly decreased in35days (P<0.01).(2) Metabolomic results reveal that in the resistant group the concentrations of dihydroxyacetone, ursodeoxycholic acid, cicloserine, hypoxanthine, tryptophan,4-O-Methylmelleolide were up-regulated, while LysoPC(18:0), PE(18:3/P-16:0), PC(20:1/18:3)、DG(24:1/22:6/0:0), PS(18:2/18:0), PI(16:0/16:0), PS(18:0/18:1), PS(14:1/14:0) were down-regulated in21days. In35days, concentrations of dihydroxyacetone,4-O-Methylmelleolide, LysoPC(18:2)、PC(14:1/22:1) were down-regulated, while LysoPC(18:0)、 LysoPE(20:1/0:0),LysoPC(16:0),LysoPE(16:0/0:0),hypoxanthine were up-regulated.(3) Upon comparison with the susceptible group, a total of413and214genes were identified as DEGs in21and35days respectively, among which244genes were up-regulated and169genes were down-regulated in21days, while99genes were up-regulated and115genes were down-regulated in35days in the AS group. DEGs in21days mainly participated in the down-regulation of oxygen transport, defensive reaction and protein modification and the up-regulation of cell morphological formation, neurodevelopment, cell adhesion function and signal pathway of TGF-beta, among which among which the GO:0050896-response to stimulus had the maximum numbers of enrichment genes (n=15), GO:0015669-gas transport process had the lowest p value of fold enrichment (0.000966) the highest fold enrichment (59.64). DEGs in35days mainly participated in the down-regulating oxygen transport.Those resultes indicated that up-regulated phospholipids(phosphatidyl serine, phosphatidyl ethanolamine, phosphatidyl inositol, diphosphatidyl glycerol, lysoPhosphatidyl ethanolamine) and down-regulated DEGs of oxygen transport (HBZ, HBAA, HBAD, RHAG)would be the key characters of AS-resistant chickens compared to AS-susceptible chickens.In conclusions, both metabonomics and transcriptomics data revealed some metabolism disorders of Iipids in AS broilers. Dihydroxyacetone may be the biomarker of low temperature induced-AS. Glycerol phospholipid compounds oxidized to lyso-glycerol phospholipid compounds would be the key mechanism of AS chickens in early stage.The combined strategy of metabonomics and transcriptomics provides comprehensive information of metabolic characters of AS and corresponding mechanisms to AS-related metabolic disorders from a systematic view.