Isolation,Identification And Preliminary Application Of Lytic Phage To Cow Escherichia And Streptococcus

Cow endometritis is a common disease in dairy farming and is one of the main causes of infertility in dairy cows.It also causes a sharp drop in milk production in dairy cows,and increases feeding costs and reduces breeding efficiency,which seriously restricts the development of dairy industry.With the unreasonable use of antibiotics,many drug-resistant strains have emerged,making the treatment of cow endometritis caused by bacterial infections encounter unprecedented difficulties.As a natural killer of bacteria,bacteriophage has renewed people's attention.In this study,Escherichia coli and Streptococcus from dairy cow endometritis saved in the laboratory were used as host bacteria,and one Escherichia coli phage EP-CE and one Streptococcus bacteriophage SP48 were isolated and purified from the sewage by double-layer plate method.The nucleic acid types of the two phages were identified,and biological characteristics and preliminary application studies were carried out.The results showed that the phage EP-CE and phage SP48 isolated in this study had strong specificity and were not sensitive to ether and chloroform.Both are double-stranded DNA viruses,the genome size of phage EP-CE is about 50 kb,and the genome size of phage SP48 is about 61 kb.Transmission electron microscopy showed that the phage EP-CE belonged to the family of myoviruses.The head is icosahedral symmetrical structure with a retractable tail that the tail and tail plates can be seen.The head is about 70.9±0.35 nm and the width is about 68.4±0.48 nm.The Tail is about 90.1±5.39 nm;phage SP48 belongs to the family of long-tailed virions.The head is icosahedral symmetrical structure with a long tail and no tail wire and tail plate.The head is about 79.0±0.33 nm,the width is about 73±0.38 nm,and the tail is about 354±0.25 nm.Phage EP-CE has good stability at 40~55 °C or pH 6~9 acid and alkali environment,the optimal multiplicity of infection is 0.01,the incubation period is about 20 min,the burst period is about 100 min,and the average burst amount is about 26 PFU/cell;phage SP48 has good stability at 40~45 °C or pH 5~10 acid and alkali environment,the optimal multiplicity of infection is 10,the incubation period is about 30 min,and the burst period is about 80 min,and the average burst amount is about 45 PFU/cell.EP-CE and SP48 have strong in vitro lytic activity against Escherichia coli and Streptococcus,respectively.Safety tests show that the two phages have no toxic side effects on mice.Uterine perfusion phage mixed preparation can reduce the uterine tissue lesions of endometritis in rabbits caused by host bacteria.The number of colonies in group A antibiotic treatment group,group B phage treatment group and E group prevention group is lower than that of group C infection group.The difference between group A,group B and group E was significantly higher than that of group C(P<0.01).The effect of group A antibiotic treatment group is higher than that of group B phage treatment group,but the difference is not significant.It can be seen that EP-CE and SP4 have certain preventive and therapeutic effects on endometritis in rabbits caused by host bacteria.