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| 題 名 | 含10% NaCl及1% Sodium Pyruvate的HiMedia TSB用於食品中金黃色葡萄球菌最確數檢測方法的適合性=Suitablity of HiMedia TSB w/10% NaCl and 1% Sodium Pyruvate in the Surveillance of Staphylococcus aureus in Foods Using the Most Probable Method |
|---|---|
| 作 者 | 秦蓓; 沈嘉恬; 葉國裕; 蔡宗佑; 蔡文城; | 書刊名 | 檢驗及品保雜誌 |
| 卷 期 | 4:2 2015.05[民104.05] |
| 頁 次 | 頁58-66 |
| 分類號 | 412.37 |
| 關鍵詞 | 金黃色葡萄球菌; MPN計數法; HiMedia TSB培養基; 世代期; HiMedia TSB; Detection of staphylococcus aureus; MPN method; Generation time; |
| 語 文 | 中文(Chinese) |
| 中文摘要 | 食品中金黃色葡萄球菌(Staphylococcus aureus)的檢測需要定量,當預期檢體中只含低菌量時,美國食品藥物檢驗局(Food and Drug Administration, FDA)出版的Bacteriological Analytical Manual 及台灣衛福部食品藥物管理署(TFDA)所公告的食品中金黃色葡萄球菌檢驗方法建議利用tryptic soy broth (TSB) w/10% NaCl & 1% sodium pyruvate 培養基進行三階三支的最確數(most probable number, MPN)計數法,本研究利用添加10% NaCl & 1% sodium pyruvate 的兩種不同廠牌(Difco, B-D, USA 及HiMedia, India)TSB 培養基評估S. aureus ATCC 25923 的生長世代期以了解其應用於食品中金黃色葡萄球菌定量檢測的效能。生長世代期的評估係將測試菌分別種入兩種廠牌的100 mL TSB w/10% NaCl & 1% sodium pyruvate 培養基,然後在不同時間(0, 2, 4, 6, 8, 11, 24 與30 小時)取出定量(1 mL)的培養液進行系列稀釋,然後將不同稀釋倍數接種液以tryptic soy agar (TSA)進行傾注平板法的生菌計數,操作雙重複。TSA 於35℃培養24~48小時後,取30~300 CFU 的平板進行計數,加以平均後,根據稀釋倍數換算生菌數,分別得到不同時間的生長菌量,再以Excel 製作semi-log 圖表,並計算世代期。結果指出B-D Difco 廠牌TSB 的遲緩期(lag phase)為8 小時,但能在24 小時達到靜止期(菌數最高峰)而符合檢驗時程;至於HiMedia 廠牌的遲緩期長達51 小時,需在73 小時以後,才能達到靜止期,因此不符合檢驗時程。另外以TSB 為滋養性(非選擇性)增菌培養基,以上述方式同樣地進行Escherichia coli, Pseudomonas aeruginosa, Salmonella enterica, S. aureus 生長世代期的測試,結果發現此四種菌在B-D Difco/HiMedia 兩種廠牌TSB 的世代期分別為21.2/20.6,42.1/40.7,19.8/17.9 及34.2/30.1 分鐘,兩種廠牌均能在24 小時達到靜止期。根據上述的發現,HiMedia 的TSB 培養基僅適用於食品中病原菌的一般滋養性增菌,而不能用於食品中金黃色葡萄球菌的最確數測試,此發現可做為食品廠品管室的參考,以避免偽陰性的產生。 |
| 英文摘要 | Generally, quantitation is required to determine the presence of Staphylococcus aureus in foods. The most probable number (MPN) method is recommended for routine surveillance of products that are expected to contain small numbers of S. aureus. The MPN method is also used for analyzing food samples that are expected to contain large populations of competing species. The “Bacteriological Analytical Manual” published by the US Food and Drug Administration (FDA), and the “Detection methods for S. aureus in Foods” issued by the Taiwan FDA (TFDA) recommend that three tubes of tryptic soy broth (TSB) containing 10% NaCl and 1% sodium pyruvate must be inoculated with 1 mL portions of decimal dilution of each sample. In order to determine the generation time of S. aureus ATCC 25923, this study used 10% NaCl and 1% sodium pyruvate to supplement each TSB sample obtained from two different providers (Difco, B-D, USA and HiMedia, India). The generation time for a bacterial species in a specific medium can be used to determine the efficacy of that medium in detecting pathogens in foods. The generation time of S. aureus ATCC 25923 was determined by separately inoculating the organisms in each of the two flasks containing each provider’s TSB supplemented with 10% NaCl and 1% sodium pyruvate (100 mL). Two flasks were incubated at 35℃ and then 1 mL portions from each flask were individually removed to perform a series of decimal dilutions at different periods of time (0, 2, 4, 6, 8, 11, 24 and 30 hrs.). Thereafter, a total viable count in tryptic soy agar (TSA) was determined by the pour plate count method. TSA were inoculated in duplicates and then incubated at 35℃ for 24-48 hrs. TSA plates containing 30-300 CFU were selected to count the number of existing colonies. The average number was obtained from the sum of two plates; this number was then multiplied by the sample dilution factor to obtain the total viable count (TVC) at different incubation time periods. TVC, in turn, was used for making semi-log graphs via Excel calculation, and then the generation time of the organism was determined. Results indicate that the lag phase of S. aureus ATCC 25923 grown in the B-D Difco TSB w/10% NaCl and 1% sodium pyruvate is 8 hours, but it can reach its stationary phase within 24 hrs. Thus, it complies with the method promulgated by TFDA; while the lag phase for S. aureus ATCC 25923 in HiMedia takes as long as 51 hours and reaches the stationary phase after 73 hours of incubation. Thus, it does not comply with the method. In addition, according to the aforementioned procedures, we used TSB as an enriched (non-selective) medium to determine the generation time for Escherichia coli, Pseudomonas aeruginosa, Salmonella enterica, and S. aureus. Results indicate that the generation time for these four organisms in B-D Difco/HiMedia TSB were 21.2/20.6, 42.1/40.7,19.8/17.9, and 34.2/30 minutes respectively. In addition, all organisms in the TSB of both brands reached their stationary phases with in 24 hours. Based on the above findings, we concluded that HiMedia TSB can only be suitable for use as an enriched medium; it was not suitable for detecting S. aureus in foods using the MPN method. Laboratory workers should use precautionary measures while using HiMedia TSB so as to avoid false negatives during the detection of S. aureus in foods. |
本系統中英文摘要資訊取自各篇刊載內容。