以改良增殖方法和免疫抗體法快速檢測截切蔬菜中Escherichia coli O157菌株   全文下載

Development of A Modified Enrichment Method for the Rapid Immunoassay of Escherichia coli O157 Strains in Fresh Cut Vegetables

Escherichia coli O157:H7 為病原菌，能附著於人體大腸之微細絨毛，造成腹瀉、糞便帶血、出血性結腸炎和溶血性尿毒症候群。Escherichia coli O157:H7 分離和鑑定非常費時，且判斷困難。本研究中截切蔬菜接種Escherichia coli O157:H7，先以含novobiocin之mEC 42℃增殖培養，6小時後，再以sorbitol MacConkey broth 添加tellurite 和cefixime 兩種抗生素，於35 ℃增殖18 小時。以此二段式增殖之細菌培養液以免疫層析法（immunostrip）或自動化酵素鍵結螢光免疫法（mini-VIDAS）篩選含E. coli O157之樣品，可以降低免疫層析法和mini-VIDAS 檢測造成之偽陽性機率。即使食物樣品中只含約10 CFU/g 之E. coli O157 菌體，經此二段式增殖，皆可達到mini-VIDAS最低偵測菌數為105 CFU/mL和immunostrip最低偵測菌數為107CFU/mL之要求。增殖液最後以複式聚合?連鎖反應（multiplex polymerase chain reaction）來確認食品中是否含有E. coli O157:H7 。因此以此方式可以快速篩選食品中含E. coli O157之樣品。

severe syndromes, including diarrhea, hemorrhagic colitis, hemolytic uremic syndrome, and thrombotic thrombocytopenic purpura. It is difficult to isolate and identify this pathogen. The purpose of this research was to establish a two-stage enrichment method for the detection of E. coli O157:H7 in fresh cut vegetables. Fresh cut vegetable samples were first enriched in modified EC broth with novobiocin at 42°C for 6 hr, and then transferred to sorbitol MacConkey broth containing cefixime and tellurite at 35°C for 18 hr. Finally, the enriched broth was verified for the existence of E. coli O157 by using an automated enzyme-linked fluorescence immunoassay (mini-VIDAS) or an immunostrip test. This two-stage enrichment method would decrease the false-positive E. coli O157 tested by immunostrip or mini-VIDAS. The pathogen in food sample could be as low as 10 CFU/g and selectively enriched to 105 or 107 CFU/mL required for the detection by mini-VIDAS or immunostrip. The presence of E. coli O157:H7 in enriched broth was confirmed by multiplex polymerase chain reaction (PCR). Thus, the two-stage enrichment method can be used to rapidly screen food samples for E. coli O157.