畜產環境人畜禽葡萄球菌分離株腸毒素分析

Analysis of Staphylococcal Enterotoxins from Workers- and Livestock Animals-Isolates

陳仁信(Ren-Shinn Chen)

指導教授 ： 周晉澄

國立臺灣大學/生物資源暨農學院/獸醫學研究所/碩士(2005年)

https://doi.org/10.6342/NTU.2005.00750

摘要

本研究從不同畜殖環境(雞場、豬場、牛場及肉品市場)中分離出之疑似563株葡萄球菌中，調查產毒金黃色葡萄球菌之腸毒素型與抗Oxacillin分布及其親源性關係。葡萄球菌來源以雞源、牛源及人源為主。抗Oxacillin金黃色葡萄球菌（Oxacillin resistant Staphylococcus aureus, ORSA）以篩選平板試驗(screening plate test)共篩選出23株，其中以人源87﹪（20/23）比例最高，顯示人源為主要污染來源。以Staphylocooccal Enterotoxin-Reversed Passive Latex Agglutination,（SET-RPLA）免疫分析套組及五組具特異性聚合酶鏈鎖反應（polymerase chain reaction, PCR）引子，檢測A, B, C, D, E 五種不同型別葡萄球菌腸毒素，結果共檢出21株產毒葡萄球菌菌株，毒素類型所佔比例依序為腸毒素B型38﹪（8/21）、腸毒素A型24﹪（5/21）、腸毒素C型24﹪（5/21）及同時含腸毒素A與B型14﹪（3/21），其中包含15株ORSA產毒菌株(A與B型3株，A型2株，B型6株及C型4株)及6株非ORSA產腸毒素葡萄球菌菌株（A型3株，B型2株及C型1株），D與E型腸毒素則皆未檢出。SET-RPLA與PCR方法檢測結果並不一致，以PCR方法之敏感性較高。將產毒葡萄球菌菌株及產毒ORSA菌株以脈衝式膠體電泳（pulsed-field gel electrophoresis, PFGE）分析，結果發現雞場間人源產腸毒素葡萄球菌株相似度高，其有地域性分佈關係，其中3株來自同一雞場分離之菌株相似性最高，此可能與單一封閉型水濂式雞場環境有關，肉品拍賣市場由於人員與拍賣豬隻來自各地，菌株相似性較低。肉品市場人員工作年資愈短其分離出之ORSA菌株及產毒株有較高比例之情形，但經卡方統計分析（Chi-square test）發現，肉品市場工作人員檢出的ORSA菌株及產毒葡萄球菌菌株陽性率與工作人員的工作年資、工作地點都不相關（P > 0.05）。本研究建議宜持續監控相關畜禽場與肉品市場工作人員之ORSA菌株及產毒葡萄球菌污染情形，並進一步探討分析環境與人畜之關係。

關鍵字

葡萄球菌腸毒素 ； 聚合酶鏈鎖反應 ； 脈衝式膠體電泳

並列摘要

This study analyzed 563 suspected Staphylococci isolates from poultry farms, swine herds, bovine herds previously, and newly sampling isolates from meat market in order firstly to distinguish Staphylococcus aureus and then to characterize oxacillin resistant Staphylococcus aureus (ORSA), types of enterotoxin production and bacterial genetic similarity. Staphylococci were mainly collected from chicken, cattle and human origins. Twenty-three ORSAs were identified by use of screening plate test and 20 of these strains were from human source. A higher contamination potential from human origin was thus suspected. Five types of bacterial enterotoxin (A to E) were detected by using Staphylocooccal Enterotoxin Rreversed Passive Latex Agglutination (SET-RPLA) and also by five groups of polymerase chain reaction (PCR) primers. Twenty-one enterotoxin producing strains were identified. The type of enterotoxins were type B 38% (8/21), type A 24% (5/21), type C 24% (5/21) and type A&B 14% (3/21). Fifteen ORSA strains composed 3 strains of type A&B, 2 strains of type A, 6 strains of type B and 4 strains of type C, and 6 non-ORSA Staphylococci producing toxins composed 3 strains of type A, 2 strains of type B and 1 type C. No type D and E toxin producing strain were detected. The agreement of detection between RPLA and PCR were only 69% and PCR had a better sensitivity. Genetic similarity of staphylocooccal enterotoxin strains and ORSA strains by using pulse-field electrophoresis indicated that human source strains collected from poultry farm were high. Three enterotoxin strains from the same poultry farm had highest similarity that was due to close-raising environment. Genetic similarities of bacteria collected from meat market were low, and that might due to open environment and high diversity of animal and human from different origins. Workers from meat market with shorter working experience had higher isolation percentage of ORSA and staphylocooccal enterotoxin strains but with no statistical differences (P > 0.05). The isolation was also found to be no difference between workers’ working environment in meat market. The study suggests a necessary continuous monitoring of ORSA and staphylocooccal enterotoxin strains in worker at farm and meat market, and also need to analyze the relationship between environment, human and animals.

並列關鍵字

staphylococcal enterotoxin ； PCR ； PFGE

參考文獻

洪淑慎、張翠瑛、柯鍚津。1992。利用酵素免疫法檢測金黃色葡萄球菌腸毒素之評估。食品科學20：394-4-1。

柯鍚津、張翠瑛。1995。以逆被動乳膠凝集方法偵測食品中產金黃色葡萄球菌及腸毒素。藥物食品分析3：57-63。

曾浩洋、陳桐榮、虞積凱。1994b。應用聚合酶鏈反應於數種不同食品中腸毒素型金黃色葡萄球菌之檢測。藥物食品分析2：217-224。

洪淑慎、黃翠萍、柯鍚津。1993。以酵素色疫套組偵測米飯中之金黃色葡萄球菌腸毒素。藥物食品分析1：297-304。

高偉霞、施養志。1993。即食生菜中金黃色葡萄球菌之調查。藥物食品分析。1：107-115。