HiMedia – M1354 M-CP Agar Base

Principle and Interpretation
Clostridial species are one of the major causes of food poisoning/gastro-intestinal illnesses. They are gram-positive, spore-forming rods that occur naturally in the soil (2). Among the family are: Clostridium botulinum which produces one of the most potent toxins in existence ; Clostridium tetani, causative agent of tetanus; and Clostridium perfringens commonly found in wound infections and diarrhoea cases. The use of toxins to damage the host is a method deployed by many bacterial pathogens. The major virulence factor of C. perfringens is the CPE enterotoxin, which is secreted upon invasion of the host gut, and contributes to food poisoning and other gastrointestinal illnesses (2). Several solid media have been devised for quantitation of C. perfringens. The selectivity of the media is achieved by incorporation of one or more antibiotics that inhibit certain anaerobes or facultative anaerobes. M-CP Agar Base is prepared as per the formula of Armon and Payment (1). It is also recommended by the Directive of the Council of the European Union 98/83/EC (3) for isolation and enumeration of Clostridium perfringens from water sample using membrane filtration technique. Tryptose, yeast extract provide nitrogenous and carbonaceous compounds, long chain amino acids, vitamin B complex and other essential growth nutrients ompounds while sucrose is the fermentable carbohydrate. Bromocresol purple serves as a pH indicator. Indoxylb-D-glucoside is a chromogenic substrate for b-D-glucosidase or cellobiose and phenolphthalein diphosphate for the detection of acid phosphatase. The addition of D-cycloserine and polymyxin B (FD153) makes the medium inhibitory to accompanying non-clostridial microflora and thus allows analysis of both clostridial vegetative cells and spores. Further selectivity is provided by incubation under anaerobic conditions. Yellow (cellobiose-negative) colonies becoming old rose to pink-red upon exposure to ammonia fumes for 30 seconds are considered to be presumptive C. perfringens. Colour differentiation on M-CP Agar Base is sometimes difficult, so typical colonies (yellow turning into pink) as well as a typical colonies (green or those that remain yellow upon exposure to ammonia fumes) are picked for confirmation. Presumptive C. perfringens can be confirmed by sulphite reduction, gram-positive, sporulating rods, non-motile, reduction of nitrate, gelatine liquefaction, lactose fermentation and other biochemical tests (4).

原理與解釋
梭菌屬物種是食物中毒/胃腸疾病的主要原因之一。它們是革蘭氏陽性，孢子形成的桿，天然存在於土壤中（2）。其中包括：肉毒桿菌（Clostridium botulinum），它可以產生一種最有效的毒素;破傷風梭菌（Clostridium tetani），破傷風的致病因子;產氣莢膜梭菌（Clostridium perfringens）常見於傷口感染和腹瀉病例。 使用毒素來破壞宿主是許多細菌病原體所採用的方法。產氣莢膜梭菌的主要毒力因子是CPE腸毒素，其在宿主腸道入侵時分泌，並導致食物中毒和其他胃腸道疾病（2）。已經設計了幾種固體培養基用於產氣莢膜梭菌的定量。通過摻入一種或多種抑制某些厭氧菌或兼性厭氧菌的抗生素來實現培養基的選擇性。 M-CP瓊脂基質按照Armon和Payment（1）的配方製備。歐盟理事會指令98/83 / EC（3）也建議使用膜過濾技術從水樣中分離和計數產氣莢膜梭菌。 胰蛋白腖，酵母提取物提供含氮和含碳化合物，長鏈氨基酸，維生素B複合物和其他必需的生長營養素，而蔗糖是可發酵的碳水化合物。溴甲酚紫可作為pH指示劑。 Indoxylb-D-葡萄糖苷是b-D-葡糖苷酶或纖維二糖和酚酞二磷酸的顯色底物，用於檢測酸性磷酸酶。添加D-環絲氨酸和多粘菌素B（FD153）使培養基抑制伴隨的非梭菌微生物群落，從而允許分析梭菌營養細胞和孢子。通過在厭氧條件下孵育提供進一步的選擇性。黃色（纖維二糖陰性）菌落在暴露於氨氣30秒後變為粉紅色，被認為是假定的產氣莢膜梭菌。 M-CP瓊脂基質上的顏色分化有時是困難的，因此挑選典型的菌落（黃色變成粉紅色）以及典型的菌落（綠色或暴露於氨煙霧時保持黃色的菌落）用於確認。推定的產氣莢膜梭菌可以通過亞硫酸鹽還原，革蘭氏陽性，孢子形成棒，非運動，硝酸鹽還原，明膠液化，乳糖發酵和其他生化測試來證實（4）。