Optimization Of Co-culture Medium For Food Borne Escherichia Coli And Salmonella And Its Application In ?PADs Test Paper Preparing

WHO lists Escherichia coli,Salmonella,Listeria Monocytogenes and Staphylococcus aureus as the most important of four foodborne pathogens.Food-borne diseases play an important role in food safety,which can lead to intestinal infections,food poisoning and zoonosis.The existing foodborne pathogenic microorganism detection techniques are either cumbersome,laborious effort,long reporting time(such as traditional culture identification methods),strong dependence on equipment and professional technicians,high cost,and is not suitable for the rapid and convenient detection of animal foods in the production line and in the market.The low cost multi linked convenient test paper technology is one of the ways to realize the rapid inspection technology for the first line screening and quality control of the production line.However,one of the key problems that need to be solved is to complete the co culture of multiple(two and/ or three)foodborne pathogens.In this study,the co-culture conditions of Escherichia coli,Salmonella and/ or Listeria monocytogenes were optimized,and the optimum co-culture conditions of the two and / or three strains were screened.The microfluidic paper based analytical devices(?PADs)was prepared,which laid a foundation for the research and development of a new technology for the convenient detection of edible bacteria.The main contents of this study are as follows:1.Optimization and improvement of Co-culture medium for Salmonella,Escherichia coli and Listeria monocytogenes and its ?PADs Assembly: The "SEL" co culture medium developed by Hyochin and Arun was used to verify the compound culture medium(SEL)of Salmonella,Escherichia coli O157:H7 and Listeria monocytogens.The optimum number of co-cultured bacteria,the best culture time,the improved process and the assembly of duplex and / or triplex ?PAD were selected.Results: SEL can co culture two species of Salmonella and Escherichia coli,and the growth curves of the two bacteria are similar when they are co-cultured for about 8 h.With the exception of Escherichia coli,the target bacteria cultured in unmodified SEL medium showed unstable coloration or very little coloration,whether in single culture or co-culture.It was found that acridine yellow was the reason that affected the coloration of Salmonella.After removing acridine yellow,the Escherichia coli and Salmonella in modified SEL medium could be co-cultured and cocolored.2.The optimum preparation conditions for two fast test strips of Escherichia coli and Salmonella enteritidis(E.coli+S.en-?PADs): screening test: The best co culture conditions of Escherichia coli and Salmonella enteritidis were screened out,E.coli+S.en-?PADs is prepared.The optimum preparation conditions were screened by response surface method.The optimum color conditions for screening E.coli+S.en-?PADs are as follows: The co culture time of Escherichia coli and Salmonella enteritis was 8 h,the dosage of substrate was 5 ?L,the dosage of bacterial liquid was 90 ?L,the reaction temperature was 37?,the reaction time was 1.5 h,and the p H value of increasing bacteria was 7.5.3.Detection performance measurement of E.coli+S.en-?PADs: The prepared E.coli+S.en-?PADs was used for the detection of animal food samples,and compared with the national standard method to verify the detection effect.Compared with the national standard method,the detection effect of E.coli+S.en-?PADs isas follows: The coincidence rate of Escherichia coli detected by market meat was 100 %,the coincidence rate of Salmonella was 90 %,and the total coincidence rate was 95 %.Conclusion:In this study,the co culture and coloration of Escherichia coli and Salmonella were solved,and two test paper(E.coli+S.en-?PADs)of Escherichia coli and Salmonella were prepared by enzyme substrate.It takes only 10 hours to achieve the results of culture from the culture to the report.The accuracy of the national standard method is achieved,and the time is saved and the cost of convenience and shortcut is lower than that of the national standard method.This research provides technical support for the development and application of the convenient detection technology of Escherichia coli and Salmonella.