Isolation And Identification Of Broad-Spectrum Phage Of Multiple Drug Resistant Salmonella And Its Iherapeutic Study In Mice

The Salmonella hosts is very extensive.Salmonella not only can cause livestock and poultry and other animal disease,but also make people sick.Animals often show typhoid,enteritis,sepsis and other clinical symptoms,when the animals infected with Salmonella.At present,antibiotics are the first choice for the prevention and treatment of Salmonella infection.However,due to the large number of antibiotics,Salmonella resistance more and more serious,multi-resistant strains are also increasing.Therefore,how to effectively control Salmonella,and prevent the growing drug resistance is today's urgent need to solve.Phage is a natural enemy of bacteria.The ratio of phage to bacteria in the environment is about 10:1.Its structure is simple,mainly by the protein shell and nucleic acid composition,and widely present in nature,its type is a wide variety,the number is large.It is generally associated with its sensitive host bacteria and is easy to separate.The phage has become a research hotspot scientists,due to efficient specific,non-toxic side effects,and widely distributed,easy to prepare and other advantages in the cleavage of host bacteria.The purpose of this study is to isolate and screen a broad spectrum of bacteriophage of Salmonella,identify its biological characteristics and explore the potential of this phage in the treatment of mice infected with Salmonella.1.Isolation,Purification and Biological Characteristics of PhageIn this study,Salmonella phage was isolated from the fecal samples of the farm by double-layer plate culture using the multidrug-resistant Salmonella CS20130523-001-1,and the phage cleavage spectrum was determined.The morphological,thermostability,pH stability,optimal infection number(MOI)and one-step growth curve were studied For one of the strong spectrum of Salmonella phage.Results:Four phages(SaFB4,SaFB13,SaFB14 and SaFB15)of Salmonella were isolated.Their cleavage rates were 35.61%,54.15%,53.17%and 57.07%for the 16 serotypes of 205 strains of Salmonella,respectively,The cleavage spectrum of SaFB4 was relatively narrow,which could cleave 73 strains of Salmonella in 9 serotypes,and the other three could cleave 109-117 Salmonella from 10-12 serotypes.And their cleavage rate of Salmonella enteritidis(39)and Salmonella typhimurium(56)was 90%and 70%,respectively,while the four phages can also crack Salmonella pullorum,chicken Salmonella typhi.The results showed that the phage SaFB14 is amember of the Siphoviridae family in the order of Caudovirales.The optimum growth temperature was 37 ?.the optimum pH was 7.0,and its pH tolerance was larger,because it could grow in the range of pH 3-10.The optimal infection number(MOI)was 0.1,and the titer was as high as 2.6 × 1010 PFU/mL.Its latency time was 10 min,the outbreak time was 70 min,the burst was 23.In conclusion,the four phages isolated from this experiment belonged to broad-spectrum lytic phage,and had strong ability to decompose Salmonella common in clinical practice.Phage SaFB14 is a common long tail phage,and had strong tolerance to acid and alkali.2.Study on the therapeutic effect of phage SaFB14 on infected miceThe safety evaluation of phage SaFB14 was carried out by using animal challenge test,the minimum lethal dose of attack bacteria was determined,and then explore the therapeutic conditions and effects of intraperitoneal injection of SaFB14 on infected Salmonella mice.The results showed that phage SaFB14 had no obvious toxicity to mice.The minimum lethal dose of multi-resistant Salmonella CS20130523-001-1 was 2 × 109 CFU/only.When the single dose of phage was given,the optimal treatment time was 0-lh after infection,and the optimal dose was 2×1010 CFU/mouse.And the survival rate of the mice was 40%(6/15).The survival rate of the mice was significantly increased(60%)for 3 days of treatment with the best mode of administration.It can be seen that SaFB 14 has a therapeutic effect on mice infected with Salmonella CS20130523-001-1 and 2 × 1010 CFU/only 1 h in the infection of bacteria for 3 consecutive days of injection treatment better.