Study On Selective Differential Medium Of Three Pathogenic Vibrios

Aquatic products are the survival foundation material and important protein source of human. It is loved by the consumers with a continuous increase of consumption. Pathogenic Vibrio in the aquatic products was the main pathogenic bacteria which causes human diseases. It can cause the intestinal diseases which spread seasonally and epidemically, such as severe diarrhea, wound infection and even septicemia. Research and setting up new techniques and methods of rapid detection of Pathogenic Vibrio were taken seriously by research institutions of various countries. The selective differential medium can not only meet the need of the rate of inspection, but also reflect the contamination status of the aquatic products. Moreover, it can overcome the weak points of the standard method, which was time-consuming, cockamamie and high-cost.The researches carried out in this paper would be described as follows:1. Study on Vibrio hollisae Differential Medium(VhDM)To save enrichment culture time and enhance the detection efficiency, the enrichment mediums of Vibrio hollisae(Vh) were optimized based on nutrient requirements and living environment demands. The results showed as follow:Vh enrichment medium compositions for mixture peptone2.11%(tryptone:fish peptone=l:l), yeast extract0.210%, NaCl2.13%, pH7.5; temperature of culturing was36±1℃. After8hours culturing, the bacteria liquid concentration of Vh enrichment medium was1.39times than that of Vh cultured on APW (GB/T4789.7-2008) under the same condition, it was obvious.Based on the specific enzyme systems and the corresponding metabolisms of Vh, according to the resistibility to different antibacterial ingredients, VhDM was designed for the rapid separation, identification or detection of Vh. The tests of color effects, plating efficiency, sensitivity and specificity were done to evaluate the efficiency of VhDM. The results showed that:VhDM has good selectivity and specificity for Vh. After incubation for16-24h at36±1℃, The Vh formed yellow colonies on VhDM. The other strains did not form colonies or formed red colonies.VhDM showed a mean plating efficiency of Vh cells of (90.65±1.72)%and its detection limit was101cfu/mL. Accordingly, VhDM is suitable for kinds of inspection agencies to detect Vh, which is simple operation, lower cost, convenient for observation and so on.2. Study on Vibrio flurialis Differential Medium(VfDM)To save enrichment culture time and enhance the detection efficiency, the enrichment mediums of Vibrio flurialis(Vf) were optimized based on nutrient requirements and living environment demands. The results showed as follow:Vf enrichment medium compositions for mixture peptone2.24%(tryptone:fish peptone=l:3), yeast extract0.204%, NaCl1.81%, pH8.0; temperature of culturing was36±1℃. After8hours culturing, the bacteria liquid concentration of Vf enrichment medium was1.59times than that of Vfcultured on APW (GB/T4789.7-2008) under the same condition, it was obvious.Based on the specific enzyme systems and the corresponding metabolisms of Vf according to the resistibility to different antibacterial ingredients, VfDM was designed for the rapid separation, identification or detection of Vf. The tests of color effects, plating efficiency, sensitivity and specificity were done to evaluate the efficiency of VfDM. The results showed that:VhDM has good selectivity and specificity for Vf. After incubation for18-24h at36±1℃, The Vf formed yellow colonies on VfDM. The other strains did not form colonies or formed red colonies.VfDM showed a mean plating efficiency of Vf cells of (92.80±2.88)%and its detection limit was101cfu/mL. Accordingly, VfDM is suitable for kinds of inspection agencies to detect Vf, which is simple operation, lower cost, convenient for observation and so on.3. Study on Vibrio furnissii Differential Medium(VfuDM)To save enrichment culture time and enhance the detection efficiency, the enrichment mediums of Vibrio furnissii(Vfu) were optimized based on nutrient requirements and living environment demands. The results showed as follow:Vfu enrichment medium compositions for mixture peptone2.08%(tryptone:fish peptone=l:3), yeast extract0.211%, NaCl1.94%, pH8.5; temperature of culturing was36±1℃. After8hours culturing, the bacteria liquid concentration of Vfu enrichment medium was1.48times than that of Vh cultured on APW (GB/T4789.7-2008) under the same condition, it was obvious.Based on the specific enzyme systems and the corresponding metabolisms of Vfu, according to the resistibility to different antibacterial ingredients, VfuDM was designed for the rapid separation, identification or detection of Vfu. The tests of color effects, plating efficiency, sensitivity and specificity were done to evaluate the efficiency of VfuDM. The results showed that:VfuDM has good selectivity and specificity for Vfu. After incubation for16-24h at36±1℃, The Vfu formed yellow colonies on VfuDM. The other strains did not form colonies or formed red colonies.VfuDM showed a mean plating efficiency of Vfu cells of (90.65±1.72)%and its detection limit was101cfu/mL. Accordingly, VhDM is suitable for kinds of inspection agencies to detect Vfu, which is simple operation, lower cost, convenient for observation and so on.We have successfully established an isolation and identification method for Vibrio hollisae, Vibrio flurialis, and Vibrio furnissii. As compared with the conventional methods (TCBS combined with the API20E system or biochemistry assessor of Vibrionaceae), vibrio enrichment mediums combined with three Selective Differential medias respectively, used for isolation and detection of Pathogenic Vibrio, possess the advantages of simple operation, economic, convenient for observation, high sensitivity and specificity, which could lay the foundation for the further study.