Study On The Antibacterial Effect And Mechanism Of Plant Polysaccharide Comined With LP-2 During The Infection Of Escherichia Coli And Salmonella

The study provides research on the issue of diarrhea in weaned piglets,which has been perplexing the breeding industry.Bacterial diarrhea is the main cause of diarrhea in weaned piglets,leading to high morbidity and mortality,antibiotic resistance,and wide distribution.Enterotoxigenic E.coli（ETEC）and Salmonella are two significant pathogens causing diarrhea in weaned piglets.ETEC disrupts the balance of intestinal flora,damages the normal structure of the intestines,and induces intestinal inflammatory reactions,which are harmful to the production of weaned piglets.Salmonella infection has a significant impact on food safety and causes high morbidity and mortality in humans due to diarrhea.The clinical isolation rate of pathogenic E.coli and Salmonella,as well as their antibiotic resistance,is increasing steadily.Probiotics can inhibit pathogenic bacteria,while plant polysaccharides can regulate the host’s immune function by promoting the secretion of cytokines by intestinal immune cells.Lactobacillus plantarum LP-2 has been found to effectively inhibit pathogenic Escherichia coli and Salmonella.However,the inhibitory effect and mechanism of Lactobacillus plantarum LP-2 on E.coli and Salmonella infection are not yet clear.Therefore,in this study,mice were infected with E.coli ATCC10667 and Salmonella typhimurium SL1344 to establish diarrhea models.The study aimed to investigate the effects and differences of eight different plant polysaccharides,such as Angelica sinensis polysaccharide extract and Poria cocos polysaccharide extract,on the inhibitory effects of LP-2 against Salmonella and Escherichia coli.Additionally,the study explored the combined use of plant polysaccharide extracts and Lactobacillus plantarum LP-2.The specific research results are as follows:（1）Biological characteristics of LP-2 and its effect on porcine intestinal epithelial cell IPEC-J2Isolation and identification of LP-2 demonstrated that its fermentation supernatant could inhibit the growth of ETEC.Acid resistance,bile salt resistance,high-temperature resistance,and drug sensitivity tests showed that LP-2 had good adaptability and potential for clinical application.Metabolomics analysis of the LP-2 fermentation supernatant revealed higher levels of acetic acid,lactic acid,and other organic acids.Co-fermentation with Angelica extract or Poria cocos extract significantly enhanced the inhibitory effect of LP-2 fermentation supernatant.（2）The Effect of polysaccharide extract and LP-2 fermentation supernatant on bacteriostasis of Escherichia coli and Salmonella in vitroUsing the Oxford Cup method,it was observed that Lactobacillus plantarum LP-2,Pediococcus lactis PA-1,Lactobacillus plantarum LP-3,and Enterococcus faecium W had certain bacteriostatic effects on ETEC,with LP-2 showing the best results.Fermentation with traditional Chinese medicine extracts,except for non-aqueous extracts of Salvia miltiorrhiza,promoted the growth and propagation of LP-2.Significant increases in the number of LP-2 were observed after fermentation.Moreover,Angelica sinensis polysaccharides（ASP）,Poria cocos polysaccharide extract（PCP）,and Portulaca oleracea polysaccharide extract（POP）significantly enhanced the antibacterial effect of LP-2.The above results show that the metabolites produced in the supernatant of LP-2can inhibit the growth of ETEC,E.coli O157 and Salmonella enterica,Salmonella enteritidis and Salmonella pullorum,ASP,PCP and other traditional Chinese medicine polysaccharide extract can promote the growth of LP-2,and promote its production of up-regulation of metabolites,enhance the inhibitory effect of metabolites on E.coli and Salmonella.（3）Establishment of a mouse model of immune protection by plant polysaccharide combined with LP-2 and analysis of intestinal micro floraC₅₇BL/6,KM and ICR mice were infected with different gradients of bacteria,and the appropriate challenge doses were obtained respectively.ATCC10667 and SL1344 were9.6×10⁷CFU/m L and 7.3×10⁷CFU/m L,respectively.The ICR mice immunized with LP-2 and LP-2 combined with PCP and ASP at the dose of 1.0×10⁹CFU/m L were used as the infection prevention animal model.ICR mice were then infected with SL1344 at1.0×10⁷CFU/m L and ATCC10667 at 3.0×10⁸CFU/m L.The results showed that LP-2could significantly reduce the intestinal damage caused by SL1344 and ATCC10067,and the addition of PCP and ASP could significantly enhance the inhibition of LP-2 on pathogenic bacteria and reduce the degree of intestinal damage.After 21 days of normal intragastric treatment,KM mice feces were collected for 16S r DNA in the LP-2 treatment group,ASP combined with the LP-2 treatment group and control group.At the same time,KM mice were infected with SL1344 1.0×10⁷CFU/m L,and mice feces were collected for16S r DNA.Compared with the control group,SL1344 increased the abundance of Proteobacteria and decreased the abundances of Bacteroidetes（Including Bacteroidetes Proteus）and Firmicutes（Including Lactobacillus）.LP-2 treatment alone and ASP combined treatment up-regulated the flora abundance of Firmicutes and Verrucomicrobia flora,and down-regulated the flora abundance of Bacteroidetes.In summary,the mice models infected by SL1344 and ATCC10667 were constructed in this study,and it was found that LP-2 produced bacteriostatic effects on Escherichia coli and Salmonella through metabolites including lactic acid and acetic acid.In this study,LP-2,which can replace the antibacterial effect of antibiotics,was found as probiotics,PCP,and ASP,which can promote the growth and colonization of probiotics and enhance the antibacterial effect,as prebiotics,and the combined microecological preparation is particularly important to control the diarrhea infection of weaned piglets caused by bacteria such as Salmonella and ETEC.At the same time,it is necessary to develop products that do not effect or promote healthy breeding while focusing on prevention and treatment,and clarify their modes and mechanisms to lay the foundation for the development and application of new drugs.