沙门氏菌选择性化学限定培养基的研制

Development of selective chemically defined medium for Salmonella

目的拟用氨基酸、维生素、无机盐替代牛肉膏和蛋白胨,开发质量可控的化学限定沙门氏菌选择性培养基。方法分别研究20种氨基酸、11种维生素或4种无机盐对大肠杆菌ATCC 25922生长特性的影响,并确定生长最适浓度,替换亚硫酸铋(BS)琼脂或Hektoen Enteric(HE)琼脂中的牛肉膏和蛋白胨,研制出化学限定BS琼脂或HE琼脂培养基。结果在36℃下培养48 h,基础培养基中添加天冬酰胺、天冬氨酸、缬氨酸、盐酸硫胺或核黄素有利于大肠杆菌ATCC 25922生长,OD_(595)值均高于0.1,而添加丝氨酸、赖氨酸盐酸盐、异亮氨酸、尼克酸、生物素或磷酸氢二钠对细菌增殖没有促进作用,OD_(595)值几乎为0。将35种营养物质按照最佳浓度替代牛肉膏、蛋白胨制成化学限定BS琼脂或HE琼脂,经过质控菌株验证,化学限定BS琼脂或HE琼脂在菌体生长指数、菌落形态和特殊显色反应等方面均与商业培养基相似。结论化学限定BS琼脂和HE琼脂均符合国标GB 4789.28—2013对参考培养基的质控要求。

Objective Amino acids, vitamins and inorganic salts were used to replace beef extract and peptone to develop a selective medium with controllable quality for Salmonella. Methods Effects of 20 kinds of amino acids, 11 kinds of vitamins or 4 kinds of inorganic salts on the growth characteristics of Escherichia coli ATCC 25922 were studied, and the optimal concentration was determined. The beef extract and peptone in bismuth sulfite(BS) agar or Hektoen Enteric(HE) agar were replaced, and the chemically defined BS agar or HE agar was developed. Results Adding asparagine, aspartic acid, valine, thiamine hydrochloride or riboflavin to the basic medium was the most favorable for the growth of Escherichia coli ATCC 25922, and the OD_(595)were all higher than 0.1. However, adding serine, lysine hydrochloride, isoleucine, nicotinic acid, biotin or disodium hydrogen phosphate to the basic medium did not promote the proliferation of Escherichia coli, and the OD_(595)was almost 0. The chemically defined BS agar or HE agar was prepared by replacing beef extract, peptone with 35 kinds of nutrients according to the optimal concentration. After verification of quality control strains, chemically defined BS agar or HE agar was similar to commercial medium in cell growth index, colony morphology and special color reaction. Conclusion Chemically defined BS agar or HE agar meet the quality requirements of national standard.