平板显色法同时计数生乳中的大肠菌群和菌落总数

Simultaneous Coliform and Aerobic Plate Counts in Raw Milk using the Chromogenic Plate Method

摘要:

为提高大肠菌群和菌落总数的计数效率，选用2-硝基苯基-β-D-吡喃半乳糖苷作为平板培养基中的显色底物，利用大肠菌群lacZ基因编码的β-半乳糖苷酶将其分解产生黄色化合物，使得大肠菌群菌落区分于其他菌株，实现大肠菌群和菌落总数的同时计数，优化该培养基配方，评估培养基的特异性、准确性以及在生乳中的应用效果。最终确定该培养基配方：胰蛋白胨12.5 g/L、酵母浸粉2.5 g/L、磷酸氢二钾2.75 g/L、磷酸二氢钾1.75 g/L、氯化钠5.0 g/L、琼脂15.0 g/L，2-硝基苯基-β-D-吡喃半乳糖苷0.1 g/L，pH值为7.0。在此基础上，添加0.05 g/L异丙基-β-D-硫代半乳糖苷或2.0 g/L乳糖均能够增强菌落显色效果，而葡萄糖则会抑制黄色化合物生成。特异性实验中，仅大肠菌群菌落颜色呈黄橙色，其他菌株则呈乳白色。在生乳样本中，该培养基与国标方法对于大肠菌群和菌落总数的计数结果基本一致。因此，该复合显色培养基能够大大提高大肠菌群和菌落总数的计数效率，降低检测成本，为生乳的卫生指标检测提供一种可行方案。

Abstract:

In order to improve the counting efficiency of coliforms and the aerobic plate count, 2-Nitrophenyl-β-D-Galactopyranoside was selected as the chromogenic substrate in the plate culture medium. The β-Galactosidase encoded by the lacZ gene in coliforms decomposes it to produce yellow compounds, making the coliforms different from other strains. This was used for simultaneous enumeration of coliforms and the aerobic plate count. The medium formula was optimized to evaluate the specificity, accuracy, and application in raw milk. The medium formula was finally determined as follows: casein tryptone 12.5 g/L, yeast extract fermentation 2.5 g/L, potassium dihydrogen phosphate 2.75 g/L, potassium dihydrogen phosphate 1.75 g/L, sodium chloride 5.0 g/L, agar 15.0 g/L, 2-Nitrophenyl-β-D-Galactopyranoside 0.1 g/L; pH value 7.0. Based on this, adding 0.05 g/L Isopropyl-β-D-thiogalactoside or 2.0 g/L lactose enhanced the colony color effect, while glucose inhibited the formation of yellow compounds. In the specificity test, the color of coliforms was yellow to orange, whereas other strains were milky white. In raw milk samples, the coliform and aerobic plate counts in this medium were basically consistent with the national, standard method. In conclusion, the complex chromogenic medium can greatly improve the efficiency of coliform and aerobic plate counts, reduce the detection cost, and provide a feasible scheme for the detection of health indicators in raw milk.