创伤弧菌产铁载体菌株的筛选及其诱导条件的响应面优化

Screening of siderophore-producing Vibrio vulnificus and optimization of induction conditions using response surface methodology

创伤弧菌是一种重要的人鱼共患致病菌,根据目前已报道的致病因子,产铁载体是其主要的致病因子之一。以海水中筛选分离出的创伤弧菌为实验菌株,通过改良金氏培养基(Modified King B,MKB)筛选产铁载体菌株,通过平板覆盖法筛选高产铁载体菌株及紫外可见分光光度计法测定铁载体活性单位(siderphore units,SU)。基于MKB培养基,选取产铁载体能力最高的V.vulnificus Y8菌株,利用响应面法对其铁载体活性单位的显著影响因素包括初始p H、装液量及碳氮比(C/N)进行优化。响应面优化结果表明:在初始p H7.84、装液量81.53 m L及碳氮比(C/N)3.06的条件下,铁载体活性单位预测值为90.7404%,并通过三次平行验证实验证明,实际平均SU值和预测SU值接近,较优化前提高了23.73%。响应面优化了MKB诱导产铁载体条件,实现铁载体的高效诱导,为日后的进一步研究奠定基础。

Vibrio vulnificus is an important pathogen.When eating infected fish, human will suffer from serious disease.To date, many potential virulence factors of Vibrio vulnificus have been identified.Among the reported factors, Siderophore is a well- known crucial virulence factor.In this study, the siderophore-producing strains were screened by using MKB （ Modified King B） medium which isolated from the seawater.MKB-CAS （Chrome azuorl S）overlay plate method and UV-VIS Spectrophotometer were used for the screening of high yield siderophore- producing strain and detection of SU （ siderphore units） respectively. V.vulnificusY8 strain was optimized which had the highest siderophore- producing capacity. The optimization of significant factors on SU including the initial pH, medium capacity and nitrogen ratio （C/N）was determined by response surface methodology. These results indicated that the value of SU was expected to 90.7404% under the optimum condition of initial pH7.84, medium capacity of 81.53 mL and C/N of 3.06.Three parallel verification results indicated that the maximum SU value was consistent with the mean SU value and the optimized SU value was increased by 23.73% compared to the previously measured value.The best optimization conditions was obtained by using response surface methodology which would lay the foundation of the further study in the future.