| It has been proved that montmorillonite(MMT)can improve the gut health of weaned piglets and broilers due to its special spatial structure and physicochemical property.However,whether MMT can improve the intestinal mucosal barrier functions of laying hens remain unknown.Bacillus subtilis(BS)is one of the most widely used probiotics for poultry production.It can enhance gut function and production performance in poultry,but the probiotic mechanism requires further research.In addition,it has been reported that there are synergistic effects between MMT and BS in inhibition of pathogenic bacteria and protection of the intestinal barrier.The combination of MMT and probiotic was more effective than MMT or the probiotic in improving the intestinal microbiota,gut barrier,and growth performance.Thus,we hypothesized that the combination of BS and MMT may also have synergistic effects in improving intestinal barrier functions and production performance of laying hens.Therefore,this study was conducted to investigate the effects of normal dietary(mycotoxin concentration meets the national hygienical standard for feeds)supplementation with MMT on the performance and intestinal barrier functions of laying hens.Furthermore,we evaluated the combination of BS and MMT on the production performance of laying hens and its mechanism to provide a theoretical reference for the scientific use of MMT and BS on laying hens.Chapter 2 to 4 Effect of MMT on performance,egg quality,antioxidant ability,and intestinal functions of laying hensFour hundred and eighty 75-wk-old laying hens(Lohmann Brown)were randomly assigned to 5 treatments with 8 replicates per treatment and 12 hens in each replicate.Hens in the control group were fed a basal diet,and hens in experimental groups(group MMT)were fed the basal diet supplemented with 0.3,0.6,0.9,or 1.2g/kg of MMT,respectively.The adjustment period lasted for 7 d,and the experimental period lasted for 70 d.The results showed as follows:1.As dietary supplementation of MMT increased,egg weight at d 35,shell thickness at d 70,and yolk index at d 35 and d 70 increased in a linear manner(P<0.05);the activities of T-SOD at d 35 and GSH-Px at d 70 in the serum,and GSH-Px activity in the liver increased in a linear and quadratic manner(P<0.05);the activities of ALP(d 35)and ALT(d 70)in the serum decreased in a linear or quadratic manner(P<0.05).The laying rate and daily egg weight during wks 1–5 of the experiment,and shell strength at d 35 tended to increase linearly(0.05?P<0.10),and feed:egg during 1 to 5 wks of the experiment tended to decrease linearly as dietary MMT increased(P=0.058).Dietary inclusion of 0.9 g MMT/kg tended to improve hens'performance during wks 1–10 of the experiment.Compared with the control,the egg mass of the group fed 0.9 g MMT/kg increased significantly(P<0.05)during wks 1–5 of the experiment,the TC concentration of the group fed 0.9 g/kg in the serum increased significantly(P<0.05)at d 70,and the T-AOC in the serum of the groups fed 0.3 and 0.9 g MMT/kg increased significantly at d 35(P<0.05).2.Dietary supplementation with MMT tended to improve the Shannon index(P=0.096),and decrease the Simpson index(P=0.095).At the genus level,compared with the control,the Ruminococcus relative content of the group MMT was significantly increased(P<0.05),the relative contents of Dorea and Blautia in groups fed 0.6 and 1.2 g/kg of MMT were significantly increased(P<0.05),and the relative content of Paraprevotella in the group fed 0.6 g MMT/kg increased significantly(P<0.05).The relative contents of Escherichia coli and Salmonella in the duodenal digesta decreased linearly and quadratically(P<0.05)as MMT increased in the diets.The relative content of Escherichia coli in the duodenum in groups fed 0.6,0.9,or 1.2g MMT/kg was significantly decreased(P<0.05),and the Salmonella relative content of groups fed 0.9 and 1.2 g MMT/kg was significantly decreased(P<0.05)compared with the control.3.As dietary MMT concentration increased,the villus height:crypt depth in the ileum and villus width in the duodenum changed in a linear and quadratic manner(P<0.05),the mRNA expression of Claudin-1 and Claudin-5 in the jejunum increased in a linear manner(P<0.05),the mRNA expression of NF-?B P65 and TNF-?of jejunum and the concentration of endotoxin in the serum decreased in a linear and quadratic manner(P<0.05).Compared with the control,the level of sIgA in the duodenum of groups fed 0.3,0.6,or 1.2 g/kg of MMT was significantly increased(P<0.05),the sIgA concentration in the jejunum and endotoxin level of ileum in groups fed 0.6 and 0.9 g MMT/kg increased significantly(P<0.05),and the MDA content of ileum in groups fed 0.6 and 0.9 g/kg of MMT was significantly decreased(P<0.05).In conclusion,supplemental MMT can partially improve the performance and egg quality,such as yolk index,shell thickness,and shell strength in laying hens through increase the antioxidant capability,protection the liver function,and improvement of the intestinal microflora and gut barrier functions.Chapter 5 to 6 Effects of the combination of MMT and BS on performance,plasma parameters,nutrient utilization,and intestinal function of laying hensThe trial used a 2×2 factorial random design.A total of 360 healthy 29-wk-old Brown Lohmann hens were randomly arranged into 4 groups with 6 replicates per group and 15 hens in each replicate,and the hens in those 4 groups were fed the basal diet(control group),basal diet+0.5 g/kg of MMT(group MMT),basal diet+5×10~8cfu BS/kg(group BS),and basal diet+0.5 g/kg of MMT+5×10~8 cfu BS/kg(group MMT+BS),respectively.The adjustment period lasted for 7 d,and the experimental period lasted for 70 d.The results showed as follows:1.Dietary supplementation of MMT increased the laying rate,daily egg weight,egg weight(d 70),shell thickness,albumen height,Haugh unit(d 35),the activity of GSH-Px and IL-2 concentration in the plasma,and the mRNA expression of CAT1 in the jejunum(P<0.05),and decreased the feed:egg,and the activity of ALT and the concentration of ACTH in the plasma(P<0.05).Dietary inclusion of BS increased the laying rate,daily egg weight,the activities of T-SOD and GSH-Px and the concentrations of IgG and FSH in the plasma,and the apparent utilization of dry mater,ether extract,crude protein,and energy(P<0.05).There were an interaction between MMT and BS supplementation on the yolk index(d 35),yolk weight(d 70),the concentrations of GLU,T-AOC,and IL-4 in the plasma,and the mRNA expression of CAT1 and GLUT2 in the jejunum(P<0.05).Compared with the control,the laying rate,daily egg weight,and the level of IL-2 in the plasma of group MMT+BS increased significantly(P<0.05).The feed:egg in the group MMT+BS decreased significantly compared with the control and group BS(P<0.05),and the T-SOD activity of plasma and the apparent utilization of energy in the group MMT+BS were significantly increased compared to results from the control and group MMT(P<0.05).Compared with groups MMT and BS,the yolk color of group MMT+BS was significantly increased(P<0.05).The activity of GSH-Px in the plasma and egg weight at d 70 in the group MMT+BS increased significantly compared with other groups.2.The MMT supplementation decreased the relative contents of Clostridium perfringens and Salmonella in the intestine and the mRNA expression of MyD88 in the jejunum(P<0.05),and increased the concentration of sIgA in the duodenum,and Occludin and Mcuin-5ac mRNA expression in the jejunum(P<0.05).Dietary supplementation with BS decreased the relative contents of Clostridium perfringens and Salmonella in the intestine,the activity of DAO and endotoxin level in the plasma,the mRNA expression of TLR-4,MyD88,NF-?B P65,TNF-?,and IL-1?in the intestine,and the protein expression of NF-?B P65 in the jejunum(P<0.05),and increased the villus height and villus height:crypt depth,and the protein expression of I?B-?in the jejunum(P<0.05).At the genus level,compared with the control,the relative contents of Megamonas and Lactobacillus in cecum of the group BS were significantly increased(P<0.05);the Megamonas relative content of the group MMT+BS increased significantly compared with the control and group MMT(P<0.05),the relative content of Clostridium in the group MMT+BS was significantly decreased compared to results from the control(P<0.05),and the Butyricimonas relative content of group MMT+BS increased significantly compared with the group BS(P<0.05).The relative contents of Clostridium perfringens,Escherichia coli,and Salmonella,and the mRNA expression of MyD88 and IL-1?in the intestine of the group MMT+BS decreased significantly compared with the control(P<0.05),and the mRNA expression of Occludin of jejunum in the group MMT+BS was significantly increased compared with the group BS(P<0.05).In addition,the mRNA expression of TLR-4 and the protein expression of NF-?B P65 in jejunum of the group MMT+BS decreased significantly and the I?B-?protein expression in jejunum increased significantly compared with the control and the group MMT(P<0.05).In conclusion,dietary supplementation with MMT can improve the production performance in laying hens through improvement of antioxidant capability and immune function,increase the expression of Occludin and Mcuin-5ac in the intestine,and regulation the intestinal microflora and reduction stress response.Supplemental BS can improve hens'performance through promotion the antioxidant capability and immune function,improvement of intestinal morphology and the apparent utilization of nutrients and intestinal micro-ecology environment,reduction the intestinal permeability,and regulation the signal pathway of TLR and the secretion of FSH.The combination of MMT and BS can improve the production performance of laying hens through increase the antioxidant and immune functions,improvement of gut morphology and intestinal microflora and the apparent utilization of energy,and regulation the signaling pathway of TLR.Taken together,the combination of MMT and BS have synergistic effects on improvement of the performance,antioxidant capability,and gut functions in laying hens to an extent. |
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