Effects Of Benzoic Acid,Bacillus Coagulans And Oregano Oil On Growth Performance And Gut Health In Weaned Piglets

Benzoic acid,bacillus coagulans and oregano oil.as feed additives,have been widely researched and applied in weanling pig diets.Howexver,the experimental results of these feed additives were vary widely and the effect of a single additive was limited.In recent years,dietary combined supplementation of organic acids,probiotics and essential oils have received attention due to their potential ’synergistic’ and ’additive’ benefits on growth performance under normal physiological conditions.However.there are few reports about the effect of benzoic acid,bacillus coagulans and oregano oil combined supplementation on growth performance and intestinal health of weaned piglets,especially in the pathogen challenge conditions.Therefore,in this study,we conducted to investigate the protective effects of benzoic acid,bacillus coagulans and oregano oil combined supplementation on growth performance,immune function and intestinal health of weaned piglets under normal condition and enterotoxigenic Escherichia coli(ETEC)challenge condition and to examine the underlying mechanism.Effects of benzoic acid,bacillus coagulans and oregano oil on the growth performance,immune function and gut health in weaned pigletsThis experiment was conducted to investigate the effect of benzoic acid,bacillus coagulans and oregano oil combined supplementation on growth performance,immune function and intestinal health of weaned piglets.A total of 25 healthy crossbred piglets(Duroc×Landrace×Yorkshire,7.65±0.3± kg)with 24±1 days of old were randomly assigned to five treatments according to their initial body weight.Each treatment consisted of five replicates(one replicate per cage)with one piglets per replicate.Treatments included:basal diets group(CON);20 g/t colistin sulfate+40 g/t bacitracin zinc(AT,antibiotic group);3000 g/t benzoic acid+400 g/t bacillu.s coaguluans group(AB);3000 g/t benzoic acid +400 g/t oregano oil group(AO);3000 g/t benzoic acid+400 g/t bacillus coaguloans+400 g/t oregano oil group(ABO).The trial lasted for 26 days.The results showed that:(1)When compared with the control group,dietary AB supplementation enhanced ADG and reduced the F/G of weaned piglets(P<0.05);the other groups also improve the growth performance of piglets,but the difference was not significant.Meanwhile,no significant difference was observed on piglets diarrhea among the treatments(P>0.05),but the diarrhea incidence of piglets in dietary AB,AO and ABO groups were decreased by 33.37%.66.63%and 41.71%,respectively(P>0.05).(2)Dietary AB supplementation increased the mRNA expression level of SGLT1 in the jejunal mucosa of piglets(P<0.05).when compared with the control group.(3)When compared with the control group,dietary AO supplementation decreased serum TNF-a concentrations of weaned piglets(P<0.05),dietary ABO supplementation decreased serum TNF-a and IL-1β concentrations(P<0.05).In addition,dietary ABO also decreased IL-1β concentrations in the jejunal mucosa compared with the control group and antibiotic group(P<0.05).(4)When compared with the control group,dietary AB supplementation increased Btifidobacterium populations in jejunal digesta and Bifidobacterium and Bacillus populations in cecal digesta(P<0.05).dietary AO supplementation increased Bifidobacterium populations in jejunal digesta,dietary ABO supplementation increased Bifidobacterium populations in jejunal and cecal digesta and decreased Escherichia coli populations in cecal digesta(P<0.05).Moreover,dietary AB and ABO supplementation increased Bacillus populations in cecal digesta compared with the antibiotic group(P<0.05).(5)When compared with the control group and antibiotic group,dietary ABO supplementation decreased the LPS concentrations and DAO activity in serum of piglets(P<0.05).When compared with the control group,dietary AB supplementation increased the mRNA expression levels of Claudin-1K Occludin and Mucin2 in the jejunal mucosa(P<0.05).Meanwhile,higher Occludin expression level were observed in ABO group(P<0.05).(6)When compared with the control group,dietary AB and ABO supplementation decreased the mRNA expression level of NOD1 in the jejunal mucosa of weaned piglets(P<0.05).In addition,the mRNA expression levels of IL-1β and TNF-α in dietary AB and ABO group were decreased by 40.0%and 26.0%,and 38.0%and 41.0%,respectively(P>0.05).Benzoic acid,bacillus coagulans and oregano oil alleviate intestinal injury caused by enterotoxigenic Escherichia coli in weaned piglets associated with modulation of Toll-like receptor 4 and nucleotide-binding oligomerisation domain protein signaling pathwaysThe present study was mainly conducted to determine whether dietary benzoic acid(A),bacillus coagulans(B)and oregano oil(O)combined supplementation could alleviate the damage of growth performance,immune function and intestinal health in weaned piglets challenged with enterotoxigenic Escherichia coli(ETEC),and to examine their underlying mechanism.A total of 30 healthy crossbred piglets(Duroc×Landrace×Yorkshire,7.64 ± 0.46 kg)with 24±1 days of old were randomly assigned to six treatments according to their initial body weight.Each treatment consisted of five replicates(one replicate per cage)with one piglets per replicate.Treatments included:basal diets group(CON);basal diets+ETEC(ETEC);20g/t colistin sulfate+40 g/t bacitracin zinc+ETEC(AT,antibiotic group);3000 g/t benzoic acid+400 g/t bacillus coagulans+ETEC(AB);3000 g/t benzoic acid+400 g/t oregano oil+ETEC(AO);3000 g/t benzoic acid+400 g/t bacillus coagulans+400 g/t oregano oil+ETEC(ABO).On day 22,pigs in CON were orally challenged with the sterile essential medium and pigs in other groups were orally infused with 3x1011cfu ETEC.The trial lasted for 26 days.The results were listed as follows:(1)During days 1 to 21(pre-challenge),AB and ABO diets increased the ADG and decreased the F/G significantly compared with that of the ETEC group(P<0.05,basal diet,without ETEC challenge).During days 22 to 26(post-challenge),ETEC challenge resulted in a 23.28%reduction of the ADG(P=0.181)and a 21.54%increasing of the F/G(P=0.23 7)compared with the CON group.However,pigs fed the AO and ABO diets significantly increased the ADG and decreased the F/G compared with ETEC group(P<0.05).During the overall period(d 1 to 26),compared with ETEC group,the AB,AO and ABO diets increased the ADG and decreased the F/G(P<0.05),no difference was observed between ETEC group and AT group(P>0.05).After ETEC challenge,the diarrhea incidence.average diarrhea index and diarrhea percentage were increased in the ETEC group compared with the CON group(P<0.05).However,compared with ETEC group,the AB and ABO diets all decreased the diarrhea incidencee,average diarrhea index and diarrhea percentage(P<0.05).Furthermore，the AO diets also decreased the diarrhea incidence and average diarrhea index compared with the ETEC group(P<0.05).(2)Compared with the CON group.ETEC challenge decreased the mRNA expression level of SGLT1(P<0.05).However,the increased mRNA abundance of SGLT1 GLUT2 and PepTl in the jejunal mucosa was observed in ABO group compared with the ETEC group(P<0.05).Additionally,higher PepTl mRNA expression levels were observed in AB groups compared with the ETEC group(P<0.05).(3)Compared with the CON group,ETEC challenge increased serum TNF-a and IL-1β concentrations(P<0.05).However，dietary AO and ABO decreased serum TNF-a and IL-1β concentrations compared with that of ETEC group(P<0.05).Meanwhile,lower serum TNF-a concentrations were observed in AB groups when compared with ETEC group(P<0.05).(4)Compared with the CON group,ETEC challenge increased TNF-a and IL-1βconcentrations and and decreased sIgA concentration in the jejunal mucosa(P<0.05).In contrast,dietary ABO decreased TNF-a and IL-1 concentrations and increased sIgA concentration in the jejunal mucosa compared with the ETEC group(P<0.05).Meanwhile,the IL-1β concentration in the jejunal mucosa was lower in AO group than that of ETEC group(P<0.05).(5)Compared with the CON group,ETEC challenge increased MDA concentrations in the serum and jejunal mucosa(P<0.05),and decreased serum T-AOC and T-SOD activity(P<0.05),and trended to decrease T-AOC and T-SOD activity in the jejunal mucosa of piglets(P<0.10).However,dietary AB,AO and ABO increased T-AOC and T-SOD activity in the serum and jejunal mucosa compared with that of ETEC group(P<0.05).Meanwhile,the MDA concentration in the serum and jejunal mucosa was lower in ABO group than that of ETEC group(P<0.05).(6)Compared with the CON group,ETEC challenge increased Escherichia coli population(P<0.05)and trended to decrease Lactobacillus and Bacillus populations in cecal digesta(P<0.10).However,dietary AT increased Lactobacillus and Bacillus populations in cecal digesta compared with the ETEC group(P<0.05).Dietary AB decreased Escherichia coli population in jejunal digesta and increased Bacillus population in cecal digesta compared with the ETEC groups(P<0.05).Dietary AO increased Lactobacillus population and decreased Escherichia coli population in the jejunal digesta and increased Lactobacillus,Bacillus and Total bacteria populations in cecal digesta compared with ETEC group(P<0.05).Dietary ABO increased Lactobacillus and Bifidobacterium populations in jejunal digesta and increased Lactobacillus,Bacillus and Total bacteria populations in cecal digesta,and decreased Escherichiacoli populations in jejunal and cecal digesta compared with ETEC group(P<0.05).(7)Compared with the CON group,ETEC challenge increased LPS concentration in serum(P<0.05).Lower LPS concentrations were observed in AT,AB,AO and ABO groups when compared with the ETEC group(P<0.05).Compared with the CON group,ETEC challenge trended to decreased the mRNA expression of Claudin-1(P<0.10).However,pigs fed with AT diets had higher Occludin and ZO-1 mRNA expression levels than the ETEC groups(P<0.05).The Claudin-1 mRNA expression in the jejunal mucosa were higher in AO group than that of ETEC group(P<0.05).Meanwhile,higher Claudin-1 and Occludin mRNA expression levels were observed in ABO group compared with the ETEC group(P<0.05).(8)ETEC challenge decreased mucin2 mRNA abundance in jejunal mucosa compared with the CON group(P<0.05).In contrast.dietary AT,AB,AO and ABO resulted in increased mucin2 expression compared with the ETEC group(P<0.05).(9)Compared with the CON group,ETEC challenge increased TLR4,MYD88,P38MAPK,NF-KBp65 and RIPK2 mRNA expression(P<0.05)and trended to increase NOD2 mRNA expression(P<0.10)in the jejunal mucosa of weaned piglets.The increased P38MAPK and RIPK2 mRNA abundance in the jejunal mucosa of weaned piglets challenged with ETEC was reduced by AB treatment(P<0.05).The NF-KBp65 and RIPK2 mRNA expression in the jejunal mucosa was lower in AO group than that of ETEC group(P<0.05).Additionally,the mRNA expression of TLR4.MYD88,IRAK1,P38MAPK,NF-Bp65.TNF-α，NOD2 and RIPK2 in the jejunal mucosa was decreased in ABO group when compared with the ETEC group(P<0.05).In conclusion,under normal feeding conditions,benzoic acid,bacillus coagulans and oregano oil combined supplementation could improve the growth performance and reduce the diarrhea of weaned pigs via regulating the immune function,improving the intestinal microecological balance and intestinal barrier integrity,and regulating the nutrient transporter related genes expressions;dietary benzoic acid+bacillus coagulans supplementation had the best effect on improving the growth performance and intestinal health,dietary benzoic acid+bacillus cougulans+oregano oil supplementation had the best effect on improving immune function of the weaned piglets.ETEC challenge decreased the growth performance and increased the diarrhea of weaned piglets via decreasing immune function and antioxidant capacity,and impairing the intestinal barrier integrity and function.However,dietary benzoic acid+bacillus coaguans+oregano oil supplementation alleviated the ETEC-induced injury on growth performance and diarrhea of weaned pigs via improving the utilization of nutrients,immune function,antioxidant capacity and intestinal barrier function,which was possibly associated with the decreasing the expression of intestinal pro-inflammatory cytokines via inhibition of TLR4 and NOD2 signaling pathways.