拟杆菌作为粪便污染指示微生物的初探

Preliminary study on bacteriodes as the potential fecal contamination indicator bacteria

目的 通过外环境中拟杆菌属菌量与大肠菌群存活菌量之间的相关性分析,探讨拟杆菌作为粪便污染指示微生物,并采用实时荧光定量PCR实现快速检测的可能性.方法 用大肠菌群最大或然数（most probable number,MPN）计数法和实时荧光定量PCR法分别定时检测模拟水样中大肠菌群的存活菌量及拟杆菌属菌量,探讨二者的污染检出时限和相关性.同时分别检测在3个时间点采集4个位点的12份河流水样,比较二者的检出情况.结果 MPN计数法检测大肠菌群需要72 h,而实时荧光定量PCR法检测拟杆菌不超过3 h;室内外模拟水样大肠菌群污染检出时限分别为＞40 d和9 d,而拟杆菌的污染检出时限分别为13 d和5 d;室外模拟水样拟杆菌属菌量（从第1天的8.3×106拷贝/ml下降到第5天的小于检出限104拷贝/ml）和大肠菌群存活菌量（从第1天的大于4.3×106MPN/100 ml下降到第5天的2.4×103 MPN/100 ml）之间呈正相关;12份河水水样两种方法检出均为阳性,符合率为100%,均说明二者对粪便污染的检测结果具有很好的一致性.结论 拟杆菌有可能作为粪便污染的指示微生物,并通过实时荧光定量PCR实现快速检测.

Objective To explore the possibility of Bacteriodes spp. as fecal contamination indicator bacteria with real-time quantitative PCR （RT-PCR） assay through analyzing the correlation between Bacteriodes spp. and coliform group in external environment. Methods Quantity of coliform group and Bacteriodes in water samples were detected by most-probable-number method （MPN） and RT-PCR,respectively, and their detection correlation was evaluated with linear correlation analysis. Both methods were also applied to detect the contaminated time limits and river water samples collected at four sampling sites in three different times. Results Seventy two hours were needed for the numeration of coliform group with MPN method,while RT-PCR could detect Bacteriodes within 3 hours. The contaminated time limit of indoor and outdoor water samples of coliform group was more than 40 days and 9 days, and Bacteriodes 13 days and 5 days,respectively. Also,the positive correlation between the quantity of Bacteroides and coliform group in outdoor water samples was obtained,the quantity of Bacteroides was from 8. 3 × 106 copise/ml to less than 104 copise/ml during the first day to the fifth day,while coliform group was 4. 3 × 106 MPN/100 ml to 2.4 × 103 MPN/100 ml. A 100% coincidence rate of the detection results with both methods was also observed. These results indicated that the detection results of both methods had perfect consistency. Conclusion Bacteriodes spp. can be potentially used as fecal contamination indicator bacteria with RT-PCR rapid detection.