A Rapid, Specific Membrane Filtration Procedure for Enumeration of Enterococci in Recreational Water

A two-step membrane filter (MF) method with mE medium, upon which the membrane must be incubated for 48 h and then transferred to a substrate medium to differentiate enterococci, is recommended by the U.S. Environmental Protection Agency to measure enterococci in fresh and marine recreational waters. The original mE medium was modified by reducing the triphenyltetrazolium chloride from 0.15 to 0.02 g/liter and adding 0.75 g of indoxyl β-d-glucoside per liter. The new MF medium, mEI medium, detected levels of enterococci in 24 h comparable to those detected by the original mE medium in 48 h, with the same level of statistical confidence. In addition, the use of mEI medium eliminated the need to transfer the membrane to a substrate medium to differentiate enterococci from other genera of the fecal streptococcal group. Colonies from mEI medium were examined to determine the rates of false-positive and false-negative occurrences. mEI medium had a false-positive rate of 6.0% and a false-negative rate of 6.5%. Interlaboratory testing of the MF method with mEI medium demonstrated that the relative reproducibility standard deviations among laboratories ranged from 2.2% for marine water to 18.9% for freshwater. The comparative recovery studies, specificity determinations, and multilaboratory evaluation indicated that mEI medium has analytical performance characteristics equivalent to those of mE medium. The simplicity of use and decreased incubation time with mEI medium will facilitate the detection and quantification of enterococci in fresh and marine recreational waters.

美国环境保护署推荐采用基于mE培养基（mE medium）的两步膜过滤（MF）法，用于检测淡水与海洋娱乐水域中的肠球菌：该方法需将滤膜置于培养基中孵育48小时，随后转移至底物培养基以区分肠球菌。原始mE培养基经改良，将氯化三苯基四氮唑的含量从0.15 g/L降至0.02 g/L，并添加每升0.75 g的吲哚基β-D-葡萄糖苷（indoxyl β-D-glucoside）。改良后的膜过滤培养基即mEI培养基（mEI medium），可在24小时内检出肠球菌，其检出水平与原始mE培养基48小时的检出结果相当，且统计置信水平一致。此外，使用mEI培养基无需再将滤膜转移至底物培养基，即可区分肠球菌与粪链球菌群的其他菌属。研究人员对mEI培养基上生长的菌落进行检测，以确定假阳性与假阴性发生率：mEI培养基的假阳性率为6.0%，假阴性率为6.5%。采用mEI培养基的膜过滤法的实验室间测试结果显示，实验室间的相对重复性标准差范围为海水的2.2%至淡水的18.9%。对比回收率研究、特异性测定及多实验室评估结果表明，mEI培养基的分析性能特征与原始mE培养基相当。mEI培养基操作简便且孵育时间缩短，将有助于淡水与海洋娱乐水域中肠球菌的检测与定量。