Study On Rapid Identification Of Vibrio Vulnificus In Aquatic Products

Vibrio vulnificus is a pathogen widely distributed in lakes and oceans, which can cause primary septicemia and soft tissue infections. Therefore, detection technology of Vibrio vulnificus is gradually aroused great concern by many researchers. The objective of this study was to establish and validate the selective medium for vibrio vulnificus, including optimization of the culture medium, screening of growth-promoting factors and antibacterial drugs, and research of mixed indicator. The experimental results were as follows:1The effects of incubation temperature, initial pH, sodium chloride, peptone, yeast extract were studied on the production of vibrio vulnificus. The software of Design Expert7.1.6was used to optimize the results of response surface methodology (RSM), and the optimized medium was comprised of peptone6g/L, beef extract1.0g/L, yeast extract4g/L, NaCl10g/L, FePO4·4H2O0.01g/L, pH7.5.2According to the single factor experiment,0.01%potassium chloride and0.03%magnesium sulfate were the most conducive to the growth of Vibrio vulnificus. Moreover, polymyxin E and polymyxin B were found not sensitive to Vibrio vulnificus through the experiment of drug sensitivity. According to the difference between minimum inhibitory concentration of the antimicrobial agents to Vibrio vulnificus and antibacterial spectrum, the final concentration were defined as100μg/ml and40μg/ml for polymyxin E and polymyxin B, respectively.3Vibrio vulnificus can decompose the cellobiose in the medium to small-molecular organic acids then the pH of the medium was changed. Single indicator was deficited in colour change or insensitive, therefore, it is necessary to optimize the mixed indicator. Bromothymol blue and bromocresol purple, Cresol red, bromocresol green, methyl orange and bromophenol bluewere used in our study Single factor experimentshowed that the mixed indicator of bromothymol blue and bromocresol green was better than the other combinations. The most sensitive effect was found when bromothymol blue and bromocresol green were mixed according to the proportion of1:2(v/v), and the colour was changed from yellow to blue in the pH range from4.31to5.47. The best concentration of this mixed indicator used in medium was1.0%(v/v).4The composition of selective medium and culture condition for vibrio vulnificus were defined based on above results. This medium can be used separate vibrio vulnificus specifically. When vibrio vulnificus grow in this medium for24hours, it would present yellow colonies. Our result of rapid detection of vibrio vulnificus using VVSM is feasible, and has higher detection efficiency than traditional methods. It is more convenient operation, more easy to observe the results, particularly easy to generalize in the instrument and equipment conditions at low level, which has a high practical value.