Effects Of Yeast On Immune Responses And Intestinal Barrier Function In Broilers And The Related Mechanisms

Three experiments were conducted to investigate the effects of yeast addition on immune responses and intestinal barrier function in broiler chickens and the related mechanism(s).Exp.1 was conducted to probe the effects of Saccharomyces cerevisiae supplementation on inflammatory responses and intestinal morphological structure in broiler chickens challenged with lipopolysaccharide(LPS)through a 3×2 factorial arrangement.A total of 480 one-day-old male broiler chicks were divided into 6 groups.Birds were fed a basal diet without or with 0.5,5 g/kg S.cerevisiae(1.0×1010CFU/g)and intraperitoneally injected with LPS or saline at an interval of 24 h between 21 and 27 d of age.Results showed that LPS challenge impaired the growth performance,as well as induced the systemic inflammation and intestinal structure damage.Supplemental S.cerevisiae at 0.5 g/kg elevated serum anti-NDV titers at 3 h post the first injection,increased serum a-acidoglycoprotein level and splenic TLR4 expression in unchallenged birds,decreased IL-1β expression in spleen at 8 h post the first and the last injection and increased ileal villus width and villus surface area.Comparatively,supplemental S.cerevisiae at 5 g/kg alleviated LPS-induced increases in serum lysozyme and diamine oxidase activities and splenic TLR4 expression at 3 h post the first injection.It also induced an increase in splenic IL-10 expression at 3 h post the first and the last injection coupled with a reduction of TLR4 expression in spleen at 8 h post the first injection.Besides,the ratio of villus height to crypt depth of jejunum and ileum was increased in birds treated with 5 g/kg S.cerevisiae.In conclusion,supplemental S.cerevisiae failed to modify the performance of broilers,but exerted positive effects on immune responses and intestinal morphological structure of birds.Overall,supplemental S.cerevisiae at 0.5 g/kg improved the innate immunity and intestinal morphological structure of broiler chickens as well as attenuated LPS-induced inflammation to a degree,while the addition at 5 g/kg could be more favorable to mitigate LPS-induced inflammation and intestinal structure damage.Exp.2 was conducted to investigate the effects of S.cerevisiae supplementation on intestinal immunity and barrier function in broilers challenged with Escherichia coli.In a 4×2 factorial arrangement,720 one-day-old male broiler chicks were allocated into 8 groups.Birds were fed a basal diet without or with 0.5 g/kg mannan oligosaccharide(MOS),S.cerevisiae strains 1 and 2(1.0×101 CFU/g).Birds were orally gavaged with E.coli culture or saline from 8 to 11 d of age.Results showed that E.coli challenge reduced the growth performance,induced intestinal inflammation and barrier dysfunction of broilers.Dietary treatment tended to reduce feed conversion ratio during the grower period(1-21 d)and increase ileal villus height and the ratio of villus height to crypt depth on d 14.Dietary treatment also decreased ileal myeloperoxidase activity on d 14,and alleviated the increases in serum diamine oxidase activitie along with ileal TLR4,NF-κB and IL-1β expression of challenged birds on d 14.Furthermore,ileal TNF-a expression was lower in both S.cerevisiae 1 and 2-treated birds,while ileal occludin expression was higher in S.cerevisiae 2 group as compared to control.MOS addition ’attenuated E.coli-induced reduction of IL-10 and occluding expression in ileum on d 14.The 16S rDNA sequencing revealed that E.coli challenge exerted some effect on microbial community structure of ileum.Addition of MOS conduced to maintain the stability of richness of some bacteria species(Reuteri lactobacillus and E.coli),whereas S.cerevisiae addition could increase the alpha diversity of ileal microbial community.Collectively,supplemental 0.5 g/kg MOS and these two kinds of yeast products all attenuated intestinal inflammation and barrier dysfunction induced by E.coli in broiler chickens.Exp.3 was aimed to investigate the effects of Kluyveromyces marxianus supplementation on immune response,intestinal structure and microbiota as well as serum metabolome in broiler chickens.A total of 960 female broiler chicks were divided into 8 goups with 8 replicates of each.Birds were fed basal diet without or with 0.25,0.5,1.0,1.5,2.0,2.5 g/kg K.marxianus(2.0×1010 CFU/g)and 0.5 g/kg S.cerevisiae(1.0×1010 CFU/g).Results showed that increasing K.marxianus addition linearly reduced feed conversion ratio of broilers during the grower period(1-21 d)and overall period(1-35 d).Dietary treatment showed a trend to increase relative thymus weight,serum IgG and lysozyme levels on d 21.Thereinto,increasing K.marxianus addition linearly increased relative thymus weight and quadratically increased serum IgG and lysozyme levels,with the medium dose(1.0 g/kg)being the most effective.S.cerevisiae addition elevated jejunal sucrase activity on 21,while increasing K.marxianus addition linearly increased ileal sucrase activity on d 21 along with ileal maltase activity on d 35,linearly increased the ratio of villus height to crypt depth of jejunum and ileum coupled with ileal villus height on d 21,as well as linearly increased ileal mucin-2,claudin-1,SGLT-1 expression on d 21.Supplemental K.marxianus at 0.5,1.5,2.5 g/kg could improve ileal microbial community structure,as it elevated Firmicutes abundance and lowered Cyanobacteria abundance as well as increased Lactobacillus abundance.The improvement of ileal microbiota was greatest in broilers fed with 2.5 g/kg,which also decreased the abundance of harmful germs(Rickettsia,Pseudomonas and Acinetobacter)in ileum.Collectively,K.marxianus supplementation was beneficial for feed efficiency and innate immunity,as well as intestinal structure and function in broilers.However,there was no obvious difference between the effects of K.marxianus and S.cerevisiae in broiler chickens.The immune function seem to be optimized at medium dose(1.0 g/kg)of K.marxianus addition,while supplemental K.marxianus at high level(2.5 g/kg)was more effective for feed efficiency,intestinal structure and function of broilers.And this could be associated with the improvement of intestinal microbial community structure after K.marxianus addition.Moreover,the LC-MS was used to analyze the serum metabolic profiling of birds from control and high-dose groups.Results showed that high dose of K.marxianus addition affected serum metabolic profiling.The concentrations of serum Gly-Gln,Arg-Glu,ethyl-benzeneacetic acid,glutamate,ornithine,2-amino-butanoic acid,phenylacetyl-L-glutamine,etc,,were up-regulated,while the concentrations of sphingomyelin,bilirubin,nicotinuric acid,1,2-Di-(9Z-octadecenoyl)-sn-glycero-3-phosphocholine,levulinic acid,etc.,were down-regulated in supplemented group as compared with control group.KEGG annotation revealed that these differences metabolites was mainly involved in the metabolic process of amino acids,nucleotides,bile acid,lipids and carbohydrate.The metabolism pathway analysis using MetaboAnalyst indicated two critical metabolic pathways:glutamine and glutamate metabolism as well as alanine,aspartate and glutamate metabolism.High dose of K.marxianus(2.5 g/kg)addition activated the metabolic process of glutamine and glutamate in these two metabolic pathways,and may primarily promoted the citric acid cycle and the biosynthesis of amino acids and nucleotides,which may conduce to the improvement of intestinal health and feed efficiency of broiler chickens.In conclusion,dietary S.cerevisiae supplementation at 0.5 g/kg improved the humoral immunity and intestinal morphological structure,as well as attenuated E.coli-induced intestinal inflammation and barrier dysfunction in broiler chickens,which could be attributed to the suppression of TLR4 signal and the increase of intestinal microbial diversity.Supplemental K.marxianus improved the feed efficiency,innate immunity and intestinal structure of broiler chickens.The immune function was optimized at 1.0 g/kg of K.marxianus addition,while the addition at 2.5 g/kg was more effective for feed efficiency and intestinal structure of broilers,which might be associated with the improvement of gut microbiota and activation of the metabolic pathways of glutamine and glutamate.