Effects Of The Synbiotic Containing Lactulose And Bacillus Coagulans On Growth,Intestinal Barrier And Immune Function Of Weaned Piglets

The pig industry is critical for feeding the people,and the weaning stage is one of the key periods in pig raising.Weaning of piglets can easily lead to weaning stress and causes many problems such as reduced growth performance,intestinal damage and weakened immune system in piglets.With the prohibition of use of antibiotic growth promoters,it is urgent to find a safe and efficient alternative to improve the the resistance to stress of weaned piglets.Therefore,this study was conducted to investigate the effects of the synbiotic containing lactulose and Bacillus coagulans on growth performance,nutrient apparent digestibilities and blood indexes of weaned piglets.Meanwhile,an immune stress model was established by intraperitoneal injection of lipopolysaccharide(LPS)to further evaluate the protective effects of the synbiotic on intestinal barrier and immune function of weaned piglets.1 Effects of the synbiotic containing lactulose and Bacillus coagulans on growth performance,nutrient apparent digestibilities and blood indexes of weaned pigletsIn the study,twenty-four healthy 27 or 28-day-old crossbred(Duroc×Landrace×Yorkshire)weaned piglets with initial body weight of 9.09±0.12 kg were randomly divided into 3 groups:the control group(CTR,n=12),the chlortetracycline group(ANT,n=6)and the synbiotics group(SYN,n=6).Piglets in the CTR group were fed a basal diet(antibiotic-free),piglets in the ANT group were fed the basal diet plus 75 mg/kg chlortetracycline,and those in the SYN group were fed the basal diet plus the synbiotic containing 10 g/kg lactulose+2×109 CFU/kg Bacillus coagulans.The experiment lasted 29 days.The results are described as follows.On days 1,15 and 29 of the experiment,there was no significant difference in the body weight(BW)among 3 groups of piglets(P>0.05).During week 1 to 2,the synbiotic containing lactulose and Bacillus coagulans reduced the feed to gain ratio of weaned piglets(P<0.05),and there was no difference between the ANT group and the other 2 groups(P>0.05).During day 1 to 29,there was no significant difference in fecal score among the 3 groups(P>0.05).Compared with the CTR group,the apparent digestibilities of gross energy and ether extract of the ANT group and SYN group were increased(P<0.05),and there were no differences between the ANT and SYN groups(P>0.05).In addition,chlortetracycline increased crude protein apparent digestibility(P<0.05),but there was no difference between the SYN group and the other 2 groups(P>0.05).There were no differences in blood routine index among the 3 groups(P>0.05).Compared with the CTR group,chlortetracycline and the synbiotic containing lactulose and Bacillus coagulans each decreased serum total bilirubin(TBIL)content of weaned piglets(P<0.05),and there was no difference between these two groups(P>0.05).In summary,adding the synbiotic containing lactulose and Bacillus coagulans to antibiotic-free diet increased the apparent digestibilities of gross energy and ether extract,and decreased the serum TBIL content and the feed to gain ratio in the weaned piglets.2 Effects of the synbiotic containing lactulose and Bacillus coagulans on the intestinal barrier function of weaned piglets challenged with lipopolysaccharideThis experiment was conducted to investigate the effects of the synbiotic containing lactulose and Bacillus coagulans on intestinal morphology and barrier function of weaned piglets challenged with LPS.The experimental animals and feeding management were the same as described in Chapter 1.After the 29-day feeding experiment,the 12 piglets in the CTR group were randomly divided into the control group(CTR-saline,n=6)and the lipopolysaccharide group(CTR-LPS,n=6).On 33 d and 34 d,the weaned piglets of the CTR.LPS group,ANT-LPS group(piglets of the ANT group in Chapter 1)and SYN-LPS group(piglets of the SYN group in Chapter 1)were intraperitoneally injected with Lipopolysaccharide at a dose of 100 μg/kg BW,whereas the CTR-saline group was injected with the same volume of 0.9%NaCl solution.A 10%D-xylose was orally administrated to all piglets of 1 mL/kg BW at the same time.Piglets were killed 4 hours after the injection.Serum was collected and used for biochemical analysis,and intestinal tissue and mucosa samples were collected for morphological examination,relative gene and protein expression analyses.The results are described as follows:LPS induced lower villus height(P<0.05)and lower ratio of villus height to crypt depth(P<0.05)in small intestine,and caused severe villi damage and clustered microvilli.However,chlortetracycline and the synbiotic containing lactulose and Bacillus coagulans attenuated these changes in intestinal morphology(P<0.05),and reduced the damage of villi and the shedding of microvilli.LPS induced higher serum diamine oxidase activities(P<0.05),while adding the synbiotic containing lactulose and Bacillus coagulans attenuated intestinal leaking(P<0.05).There was no difference in intestinal morphology between the ANT-LPS group and other 2 groups(P>0.05).Compared with the CTR-saline group,LPS induced lower serum D-xylose level(P<0.05),but there were no differences in serum D-xylose level among the CTR-LPS group,ANT-LPS group and SYN-LPS group(P>0.05).LPS induced high mRNA expression related to tight junction in the j ejunum(ZO-1,ZO-2,CLDN3)(P<0.05)and ileum(ZO-1,ZO-2,CLDN-2,CLDN-3)(P<0.05).In addition,significant decreases of ZO-2,OCLN and CLDN-3 protein levels in jejunum were also observed(P<0.05).However,chlortetracycline and the synbiotic containing lactulose and Bacillus coagulans reduced the mRNA expression related to tight junction in the jejunum and ileum(P<0.05),and up-regulated ZO-2,OCLN and CLDN-3 protein levels in the jejunum(P<0.05).In summary,adding the synbiotic containing lactulose and Bacillus coagulans to antibioticfree diet partially alleviated the destruction of intestinal morphology and barrier function induced by LPS,and improved the intestinal resistance to inflammation of weaned piglets.3 Effects of the synbiotic containing lactulose and Bacillus coagulans on the intestinal immune function of weaned piglets challenged with lipopolysaccharideThis experiment was conducted to investigate the effects of the synbiotic containing lactulose and Bacillus coagulans on body temperature,blood indexes and cytokines of weaned piglets challenged with LPS.The experiment design and sample collection were the same as Chapter 2.Compared with the CTR-saline group,the body temperature of weaned piglets in the CTR-LPS group significantly increased from 1 to 4 hours after LPS injection(P<0.05).At 3 hours after LPS injection,the body temperature of weaned piglets in the SYNLPS group was significantly decreased(P<0.05),but there was no significant difference between the ANT-LPS group and the other 2 groups(P>0.05).Compared with the CTR-saline group,the white blood cell count,neutrophil count and lymphocyte count of weaned piglets in the CTR-LPS group were significantly reduced(P<0.05).There were no significant differences in blood routine index among the CTR-LPS group,ANT-LPS group and SYN-LPS group(P>0.05).After LPS injection,the concentrations of TBIL,triglyceride(TG),cholesterol(CHOL),low-density lipoprotein cholesterol(LDL-C)and the activity of aspartate aminotransferase(AST)in serum of weaned piglets were significantly increased(P<0.05),and the concentration of serum glucose(GLU)was significantly decreased(P<0.05).Compared with the CTR-LPS group and SYN-LPS group,Chlortetracycline significantly decreased serum CHOL and LDL-C contents of weaned piglets challenged with LPS(P<0.05),but the synbiotic containing lactulose and Bacillus coagulans had no significant effect on serum biochemical indexes(P>0.05).LPS injection and different treatments after LPS injection had no significant effect on serum immunoglobulin(Ig)G and IgA(P>0.05).Compared with the CTR-saline group,CTR-LPS significantly up-regulated the mRNA expression of Toll-like receptor(TLR)4,myeloid differentiation factor(MyD)88,nuclear transcription factor(NF)-κB,tumor necrosis factor(TNF)-α,interleukin(IL)-1β and IL-10 in the jejunum of weaned piglets(P<0.05),down-regulated the mRNA expression of TLR4 and TNF-α in the ileum(P<0.05),increased the levels of IL-1β,IL-6,IL-8,IL-12,TNF-α,interferon(IFN)-γ,IL-10 in serum(P<0.05)and the levels of IL-4,IL-6,IL-12,IFNy and IL-10 in the j ejunum(P<0.05).However,chlortetracycline and the synbiotic containing lactulose and Bacillus coagulans significantly alleviated the LPS-induced inflammation of weaned piglets(P<0.05).In summary,adding the synbiotic containing lactulose and Bacillus coagulans to antibiotic-free diet partially inhibited the jejunum TLR4/MyD88/NF-κB signaling pathway and reduced the mRNA expression of TNF-α and IL-1β,regulated the levels of cytokines of j ejunum and serum,alleviated the stress response induced by LPS,and improved the resistance to inflammation of weaned piglets.