Bisson J W, Cabelli V J
Appl Environ Microbiol. 1979 Jan;37(1):55-66. doi: 10.1128/aem.37.1.55-66.1979.
A membrane filter procedure has been developed for the rapid quantitation of C. perfringens in the aquatic environment. Background growth is inhibited by the use of D-cycloserine, polymyxin B sulfate, and incubation at 45 degrees C. Differential characteristics include the fermentation of sucrose, production of acid phosphatase, and the absence of beta-D-glucosidase activity. The medium is prepared as follows (in grams per 100 ml of distilled water): tryptose, 3.0; yeast extract, 2.0; sucrose, 0.5; L-cysteine, 0.1; MgSO4. 7H2O, 0.01; bromocresol purple, 0.004; and agar, 1.5. The ingredients are dissolved, and the pH is adjusted to 7.6. After autoclaving at 121 degrees C for 15 min, the medium is allowed to cool at 50 degrees C, and the following are added per 100 ml: D-cycloserine, 40 mg; polymyxin B sulfate, 2.5 mg; indoxyl-beta-D-glucoside, 60 mg; 2.0 ml of a filter-sterilized 0.5% phenolpthalein diphosphate solution; and 0.2 ml of a filter-sterilized 4.5% FeCl3.6H2O solution. Enumeration of C. perfringens in a water sample is completed within 18 to 24 h. The verification of typical colonies was 93%. The average recovery from peptone-water spore suspensions of five strains was 79%, and that from filter-sterilized seawater suspensions was 90%. The precision of the method was approximately equal to that expected from random error alone. Confirmed recoveries of C. perfringens from water and sewage samples generally were greater than those by the Bonde pour tube method.
已开发出一种膜过滤程序，用于快速定量水生环境中的产气荚膜梭菌。通过使用D - 环丝氨酸、硫酸多粘菌素B以及在45℃下培养来抑制背景生长。鉴别特征包括蔗糖发酵、酸性磷酸酶的产生以及β - D - 葡萄糖苷酶活性的缺失。培养基按如下方法制备（每100毫升蒸馏水的克数）：胰蛋白胨，3.0；酵母提取物，2.0；蔗糖，0.5；L - 半胱氨酸，0.1；MgSO₄·7H₂O，0.01；溴甲酚紫，0.004；琼脂，1.5。将成分溶解，调节pH至7.6。在121℃高压灭菌15分钟后，让培养基在50℃冷却，每100毫升加入以下成分：D - 环丝氨酸，40毫克；硫酸多粘菌素B，2.5毫克；吲哚酚 - β - D - 葡萄糖苷，60毫克；2.0毫升经滤膜除菌的0.5%酚酞二磷酸溶液；以及0.2毫升经滤膜除菌的4.5% FeCl₃·6H₂O溶液。水样中产气荚膜梭菌的计数在18至24小时内完成。典型菌落的验证率为93%。五株菌株在蛋白胨水芽孢悬液中的平均回收率为79%，在经滤膜除菌的海水悬液中的平均回收率为90%。该方法的精密度大致等同于仅由随机误差所预期的精密度。从水和污水样品中经确认的产气荚膜梭菌回收率通常高于邦德倾注管法。
