Abstract:

【Objective】With the development of intensive and large-scale broiler breeding, the impact of environmental conditions on the growth of livestock and poultry has become increasingly prominent. Under high-density feeding conditions, a large amount of toxic and harmful gases is generated constantly, among which ammonia is the most harmful gas which reduces the performance of livestock and poultry and threatens animal health. The purpose of this experiment was to study the effects of chronic ammonia stress on serum metabolites in broiler chickens, and to explore the changes of metabolism caused by ammonia stress in terms of small molecules and metabolic pathways, so as to provide data support for reasonable regulation of ammonia in the poultry house. 【Method】Ninety-six 21-day-old healthy AA broilers were randomly divided into control group and experimental group by a single factor completely random design, with 4 replicates in each group and 12 chickens per replicate. The control group ammonia concentration was 0 mg·kg ^-1, the test group ammonia concentration was 45 mg·kg ^-1. The experiment was carried out in an artificial simulated respiratory chamber, which was fully automated to control the temperature and humidity and the other environmental conditions. Broilers were raised in the nets and the feed and water were provided ad libitum during the entire experimental period. The trial lasted 21 days with 24 h lighting program. They were routinely managed according to the AA broiler feeding manual and immunized according to the routine procedures. Two broiler chickens pre replicate from each treatment were chosen randomly and blood samples were collected from the wing vein at 42 day-old. After pretreatment of serum samples, the metabolites were detected by GC-MS technology, and identified by mass spectral database. Principal component analysis (PCA), partial least-squares discriminant analysis (PLSDA), orthogonal partial least squares discriminant analysis (OPLSDA) and t-test were used to find the different metabolites, and then the bioinformatics methods was used to analyze the differential metabolic pathways. 【Result】Results showed that: (1) A total of 204 metabolites were detected in serum of broiler chickens by GC-MS combining with rapid identification of the detected metabolites performed with a mass spectrometry database. Metabolomics data were imported into SIMCA-P software for multivariate statistical analysis, and then t-test. Twenty-three different metabolites were identified, of which 19 were up-regulated and 4 were down-regulated. (2) Differential metabolites mainly involved energy metabolism (lactic acid, α-ketoglutaric acid), amino acid metabolism (L-Allothreonine acid, L-Homoserine, Nicotinylglycine), fatty acid metabolism (stearic acid, linolenic acid, linoleic acid, cholesterol) and nucleotide metabolism (hypoxanthine, uracil, thymine) and so on. (3) Metabolic pathway enrichment analysis showed that ammonia stress mainly affected the fat metabolism pathway in serum of broiler chickens, including linoleic acid metabolism, arachidonic acid metabolism and α-linolenic acid metabolism. 【Conclusion】 It can be seen that GC-MS can detect serum metabolites comprehensively, and screen differential metabolites accurately. Chronic ammonia stress significantly affected serum metabolites content of broilers, mainly affected fat metabolism pathway.

Key words: broiler, metabonomics, serum, metabolites, GC-MS