| Shiga toxin-producing Escherichia coli(STEC)can cause foodborne diseases.STEC is a normal flora in the intestines of ruminants,cattle and sheep,and can cause human infections with STEC through contaminated meat,vegetables and water.Because STEC pollution involves the entire food production chain and the environment,it brings great difficulties to the diagnosis,prevention and control of this disease.This article compares the different selective STEC culture media,the isolation and identification of STEC strains,the evaluation of cytotoxicity,and the isolation and preliminary application of STEC phage..1 Isolation and identification of Shiga toxin-producing Escherichia from ruminantsShiga toxin-producing Escherichia coli(STEC)can cause foodborne diseases.It often exists in the intestines of ruminants,such as cow and sheep,as normal flora.In this study,we isolated and identified Shiga toxin-producing Escherichia coli isolates from cow and sheep feces samples,and determinaed Shiga toxin subtypes.Totally11isolates were isolated from sheeps from Jiangsu,Yunnan,and Hebei,and 11 strains from cows from Xinjiang were collected.[Methods]We conducted the separation and identification through bacterial selective culture and detection of specific genes stx1and stx2.In this study,the separation and identification result revealed that 11isolates of sheep origin were isolated with an isolation rate of 17.5%(11/63);1 isolate from cow origin was isolated with an isolation rate of 0.7%(1/134).The results indicated the isolation rate of STEC strain from sheep The results showed that the prevalence of STEC in China was improved,and the proportion of STX1+STX2genotypes was increased,and the isolation rate of STX2A and STX2C subtypes was higher.Therefore,in addition to cattle,sheep as the host of STEC strains should also receive more attention.2.pathogenic potential analysis of Shiga toxin-producing Escherichia from ruminants.2 Pathogenic potential analysis of Shiga toxin-producing Escherichia from ruminantsShiga toxin-producing Escherichia coli(STEC)can cause foodborne diseases.It often exists in the intestines of ruminants,such as cow and sheep,as normal flora.In this study,we isolated and identified Shiga toxin-producing Escherichia coli isolates from cow and sheep feces samples,and determinaed Shiga toxin subtypes.Totally11isolates were isolated from sheeps from Jiangsu,Yunnan,and Hebei,and 11 strains from cows from Xinjiang were collected.The pathogenic potential of STEC isolates was analyzed by Vero cytotoxicity test and hemolytic activity test.5 out of 11sheep-derived isolates are highly toxic to Vero cells,and 3 have hemolytic activity;5out of 11 bovine-derived isolates are strongly toxic to Vero cells,and 3 have hemolytic activity.The results show that the pathogenic potential of STEC isolates from sheep is higher than that of bovine strains.Therefore,in addition to cattle,sheep should also receive more attention as the host of STEC strains.3 Analysis of Whole Gene Sequencing Results of Shiga Toxin Escherichia coliShiga toxin-producing Escherichia coli(STEC)can cause foodborne diseases.STEC often exists in the intestines of ruminants such as cattle and sheep in the form of normal flora.In this study,four strains with high pathogenic potential were selected for full gene sequencing based on the Vero cell model and hemolytic activity test.And analyze the serotypes,virulence genes and drug resistance genes of these STEC isolates to understand the threat of these STEC isolates to human health.[Method]Through the NCBI website and Center for Genomic Epidemiology website,the serotype,virulence gene and drug resistance gene were analyzed.EC276,EC278 are O157:H7 serotypes,and EC623 is a non-O157 STEC strain.The virulence genes of EC276 and EC278 are nle,ast A,kat P and tox B,hemolysis genes are eae and ehx A,and adhesion-related genes are iha and iss.EC200 virulence gene sub A,EC623non-virulence gene.EC276 and EC278 have great pathogenic potential and need to be paid attention to.4 Isolation and Purification of Shiga-producing Escherichia coli Phage and Biological Characteristics of Broad Spectral Phage PH444 applicationThe O157:H7 serotype related to severe disease outbreak has attracted wide attention.But,in recent years,non-O157 STEC has also played an important role in disease epidemic.The reason why it is difficult to control is that STEC is a normal flora in the intestine of ruminants but infects humans.Bacteriophage is an alternative way instead of antibiotic to control pathogenic bacteria.The present work extracted and analyzed a bacteriophage named PH444 from sewage at Nanjing sawage.PH444 can specifically infect and lyse six E.coli O157:H7 and six non-O157 STEC isolates.According to the one-step growth analysis,our extracted PH444 showed the great burst size of 94PFU/cell and the short latency of 10 min.The whole genome sequence and genomic analysis indicated that the PH444 is a T7-like coliphage which harbors a 115111-bp double-stranded DNA genome and as a member of family Podoviridae.In addition,we also found that PH444 was belong to lytic phage after screening by online tool PHACTS,indicating this phage suitable for clinic application.Bacterial load could be reduced from 3×10~3PFU/m L to 0 within 1 hour in milk after treating with PH444.On the basis of the above features,Phage PH444 has potential application value in phage therapy. |
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