The Research Of Oral Microcapsulated Phages Prophylaxis Against Mixed Escherichia Coli And Salmonella Infections In Weaned Piglets

Weaning is a severe stress for piglets,which can lead to a decrease in their immunity and make them susceptible to infection by pathogens,resulting in diarrhea and causing huge financial losses to pig farms.Escherichia coli and Salmonella are the two major pathogenic bacteria causing diarrhea in weaned piglets.However,the ban on antibiotics has presented a significant challenge to preventing and controlling diarrhea caused by Escherichia coli and Salmonella in weaned piglets on pig farms.To develop a new antibacterial agent,the phages are isolated from sewage,and their biological characteristics and whole genomes are analyzed.This will provide a new approach for pig farms to prevent diarrhea in weaned piglets caused by Escherichia coli and Salmonella.In this study,Escherichia coli H4 and Salmonella Typhimurium SM022are used as host strains to isolate phage ZH4 and NJ12,respectively.The biological characteristics and whole genome analysis of the two phages show that they could survive at 4℃-50℃and p H 4-10 without significant decrease.The nucleic acid types of both phages are ds DNA,with genome lengths of87,366 bp and 39,496 bp,GC contents of 38.84%and 49.89%,and a total of132 and 49 encoded CDSs without t RNA.The phage genomes do not contain antibiotic resistance and toxin-related genes.According to BLASTp comparison and evolutionary analysis,phage NJ12 and phage ZH4 belong to the subfamily Felixounalikevirus and the subfamily Kayfunavirus,respectively.In vitro antibacterial activity tests show that phage NJ12 and EP01 inhibit effectively the growth of SM022 and Escherichia coli O157:H7 GXEC-N07within 16h at an MOI of 1.The study also investigate the preventive effect of orally microencapsulated phage on diarrhea caused by a mixed infection of E.coli and Salmonella in piglets.A mixture of phage NJ12 and the previously reported EP01 are microencapsulated with sodium alginate.Eighteen piglets are randomly divided into the phage group(6 piglets),the phage-free group(6piglets),and the control group(6 piglets).The phage group is orally 6 g microencapsulated phage,both the phage-free group and the control group are orally 6 g microencapsulated PBS.Two hours later,the piglets in the phage and phage-free groups are fed feed contaminated with 6.0 m L(10~8 CFU/m L)of GXEC-N07 and 6.0 m L(10~8 CFU/m L)of SM022 every day.Samples are collected starting when all piglets in the phage-free group had diarrhea.The results show that compared with the phage group,the piglets in the phage-free group have more severe diarrhea,greater weight loss,higher neutrophil levels,higher SM022 and GXEC-N07 loads in the ileum(P<0.05),and more severe intestinal damage.In addition,compared with the phage-free group,oral phage significantly reduce the relative abundance of Enterobacteriaceae in the intestinal flora of piglets within the phage group(P<0.05).In conclusion,the phage NJ12 and EP01 show good antibacterial effects in vitro.Microencapsulated phage NJ12 and EP01 administer orally to weaned piglets effectively relieved diarrhea caused by a mixed infection of Escherichia coli O157:H7 GN07 and Salmonella Typhimurium SM022 and significantly reduced the relative abundance of Enterobacteriaceae in the ileum.This study will provide a reference and scientific basis for the application of phage as feed additives to prevent bacterial diarrhea in weaned piglets.