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首页> 外文期刊>Journal of Clinical Microbiology >Rapid and Sensitive Assay for Detection of Enterotoxigenic Bacteroides fragilis
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Rapid and Sensitive Assay for Detection of Enterotoxigenic Bacteroides fragilis

机译：快速灵敏的检测肠毒素的脆弱拟杆菌

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Bacteroides fragilis is an obligatory anaerobic, gram-negative bacterium found among the normal intestinal flora of humans. Enterotoxigenic strains of B. fragilis (ETBF) have been associated with diarrheal diseases in humans and animals. The enterotoxin of ETBF induces fluid changes in ligated intestinal segments and cytotoxic response in HT29/C1 cells. By using a pair of monoclonal antibodies (MAbs; MAb C3 and MAb 4H8) specific for the lipopolysaccharide of B. fragilis, an assay based on immunomagnetic separation (IMS) in combination with PCR (IMS-PCR) was developed. After DNA extraction, a 294-bp fragment was amplified. The specificity of the IMS-PCR assay was evaluated by adding previously isolated and confirmed ETBF strains to normal fecal samples. All fecal samples to which ETBF strains were added were positive, showing a 100% specificity. The spiked fecal samples were also used for evaluation of the sensitivity of the assay. The detection limit was found to be ～50 CFU/g of feces. By this method 10 clinical fecal samples (5 from patients with diarrhea and 5 from healthy controls) were examined. The results of PCR were in accordance with the results of the HT29/C1 cell assay for all samples. The minimum time to retrieval of the final result by the IMS-PCR method is 36 h. The proposed IMS-PCR assay is rapid and sensitive for the direct detection of ETBF in stool samples.
机译：脆弱的拟杆菌（Bacteroides fragilis）是在人类正常肠道菌群中发现的一种必不可少的厌氧革兰氏阴性细菌。 B型肠毒素菌株。脆弱类杆菌（ETBF）与人类和动物的腹泻病有关。 ETBF的肠毒素在HT29 / C1细胞中诱导结扎的肠段液体变化和细胞毒性反应。通过使用一对特异于B脂多糖的单克隆抗体（MAbs; MAb C3和MAb 4H8）。脆弱类，建立了一种基于免疫磁分离（IMS）结合PCR（IMS-PCR）的检测方法。 DNA提取后，扩增出294bp的片段。通过向正常粪便样品中添加先前分离和确认的ETBF菌株，评估IMS-PCR分析的特异性。所有添加了ETBF菌株的粪便样品均为阳性，显示出100％的特异性。加标的粪便样品也用于评估测定的灵敏度。粪便的检出限为〜50 CFU / g。通过这种方法，检查了10个临床粪便样本（5个来自腹泻患者，5个来自健康对照）。 PCR结果与所有样品的HT29 / C1细胞测定结果一致。 IMS-PCR方法检索最终结果的最短时间为36小时。所提出的IMS-PCR分析方法快速，灵敏，可直接检测粪便中的ETBF。

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