The Study Of Antagonistic Activities Against Campylobacter Jejuni By Lactic Acid Bacteria

Campylobacter jejuni,as a zoonotic pathogen,is one of the main pathogens for infectious diarrhea.At present,antibiotics are most widely used as medicines for C.jejuni infection.Long-term use of antibiotics will bring the drug residue and resistance in body.It has been wildly reported that probiotics were used as the substitution for antibiotics to prevent C.jejuni infection.In this study,firstly,the mouse-infected model and the poultry-carried model were used to investigate the antagonistic activities against C.jejuni by different species of lactic acid bacteria.The difference of the antagonism by lactic acid bacteria in these two models was analyzed.Secondly,the changes of gut microbiota caused by the intervention or treatment of lactic acid bacteria in different models were analyzed and the relationship between the host's antagonistic effects and gut microbiota were investigates.Furthermore,a Caenorhabditis elegans model was constructed to investigate the mechanism of lactic acid bacteria on alleviateion about C.jejuni infection.Finally,the correlation between the growth characteristics of lactic acid bacteria and its antagonistic effect on C.jejuni infection was analyzed.Combined with genome comparison analysis,it was determined that the capabilities of film formation and cell adhesion of lactic acid bacteria directly affected its antagonistic effect on C.jejuni.The main research results of this article are as follows:1.A mouse model of Toxoplasma gondii and C.jejuni co-infection was constructed to eliminate the colonization resistance of normal mice to pathogenic bacteria.Through the intervention of lactic acid bacteria on mice,the colonization amount of C.jejuni in feces,fecal occult blood,intestines Pathology,cytokine levels in serum,and short-chain fatty acids in feces were analyzed to find strains that could significantly antagonize C.jejuni and improve immunity in mice.At the same time,C.jejuni colonization,growth characteristics(weight,thymus weight and index,growth plate width coefficient)in the cecum content of broilers,the transcription level of C.jejuni virulence genes in the cecum content,short-chain fatty acids in the cecum content were also analyzed in chicken-C.jejuni model.The lactic acid bacteria that significantly reduced the carrying of C.jejuni and improved its metabolic system and growth characteristics in broilers were screened.Through correlation analysis,it was found that in two different animal models,the antagonistic ability of lactic acid bacteria to C.jejuni showed strong consistency.2.The changes on the gut microbiota between two models were measured.In mice,lactic acid bacteria would reduce the abundance of Proteobacteria which represented a wide range of pathogens at the phylum level.At the genus level,the regulation of Lactobacillus and Bifidobacterium improved the body's immune system The reduction of Unclassified Enterobacteriaceae indicated the intestinal immune function.The increase of Unclassified Clostridiales and Allobaculum lead to the production of butyric acid and repaired the intestinal barrier.The experimental results show that the intervention of lactic acid bacteria would regulate the short-chain fatty acids of the host as a carbon source and energy by affecting the abundance of intestinal flora.The level of the intestinal epithelial mucosal immune system was repaired and the mouse immune system was regulated.In broiler caecum,the improvement of Lactobacillus,Peptostreptococcus and Bacteroides herald the regulation of the immune system and metabolic system and improved the growth and development of the host.Lachnospira and Clostridium was involved in the growth and maturation of T cells.It was related to the regulation of the immune system.The abundance changes affected the development of immune organs.The intake of lactic acid bacteria could further regulate the synthesis of SCFAs by regulating the intestinal flora and assisting in regulating the host metabolic system.The development and maturation of the immune system inhibited the proliferation of C.jejuni in the host and improved the production level of poultry.The study analyzed the difference of intestinal flora in the two models.After the intervention of lactic acid bacteria,the changes in gut microbiota of the two animal models were common and specific,which indicated the gut microbiota to participate in the lactic acid bacteria against pathogenic bacteria.But it was not the determinant of lactic acid bacteria.3.Building C.jejuni's C.elegans infection model,we investigated whether this model was suitable for large-scale and maximum simulation of mammalian lactic acid bacteria antagonist C.jejuni screening model.Through the longevity,lactic acid bacteria and C.jejuni colonization,morphological size,pharyngeal aspiration,defense gene expression and other series of indicators,we found that the nematode infection model can better simulate the infection of C.jejuni in mice and is suitable for large-scale useing.It was found that increasing the expression of defense genes is a mechanism for lactic acid bacteria to relieve C.jejuni infection in C.elegans.Considered the high homology between the nematode gene and mammals,it was found that the activation of defense genes is the mechanism for lactic acid bacteria to relieve C.jejuni infection in the host.The chronic and acute oxidative damage models of C.elegans were constructed using hydrogen peroxide and walnut quinone.Under lactic acid bacteria,the nematode lifespan,antioxidant enzyme levels,ROS content and transcription levels of some immune genes were measured.The nematode model was linearly fitted with the mouse and poultry models,and it was found that they had a strong correlation and a higher degree of fit with the mouse model.The results reflected that the nematode could simulate the antagonistic situation in vivo.It confirmed the mechanism of lactic acid bacteria alleviating C.jejuni infection by activating defense genes in the body.4.Correlation analysis was measured between the biological characteristics of different species of lactic acid bacteria(growth capacity,acid production capacity,acid and bile salt resistance,biofilm formation ability,adhesion to HT-29 cells)and the growth of C.jejuni and the inhibition of the transcription level of C.jejuni virulence factors.It was found that the self-forming biofilm of lactic acid bacteria and the ability to adhere to cells were most related to its ability to antagonize C.jejuni.It was investigated whether lactic acid bacteria would interfere with C.jejuni quorum sensing.It was found that when the concentration of C.jejuni reached 10~6 CFU/m L,the aggregation of C.jejuni would form an obvious quorum sensing.At this time,the LuxS gene related to QS system would be affected.It is completely activated and regulated by lactic acid bacteria.Not only that,the metabolites of lactic acid bacteria also affect C.jejuni's quorum sensing system.The cell model has also been introduced to verify the regulation of lactic acid bacteria C.jejuni AI-2/LuxS QS system.Taking Lactobacillus plantarum as an example,the core potential genes that antagonize C.jejuni are analyzed,it is found that the functional gene components related to cell membrane tissue synthesis and carbohydrate transport and metabolism are the largest.The presence of these core functional genes gives lactic acid bacteria a strong Membrane capacity,adhesion to the host and colonization capacity in the host,which also confirmed the ability of the lactic acid bacteria to form a biofilm and adhesion to HT-29 cells and its antagonistic ability to C.jejuni Related to this conclusion.