洁净环境样本低浓度微生物高效富集检测及应用

Study on the convenient and efficient enrichment effect of new modified magnetic beads on low level microorganisms in the environment.

为了研究对洁净环境样本的低浓度微生物监测方法,制备了低浓度的革兰氏阳性菌和阴性菌各3种,使用修饰后的新型磁珠对模拟样本中的微生物进行富集评价实验.革兰氏染色和菌落计数结果表明,磁珠对3种革兰氏阳性菌和3种阴性菌均具有显著富集浓缩作用,与大肠杆菌相比,磁珠对金黄色葡萄菌的富集浓缩比更高.使用磁珠富集液对自来水、桶装水和瓶装矿泉水中的微生物进行非特异性吸附后,比较上清和磁珠-微生物复合物的细菌16S rDNA基因拷贝数和菌落总数指标,结果表明磁珠对自来水和桶装水中的微生物均具有显著的吸附效果(P<0.01,P<0.001).通过比较,抽滤法和磁珠法对自来水中的微生物富集效果有差异(P=0.002),2种方法对桶装水和矿泉水的富集效果无显著差异(P=0.362).采集实际空气样本进行磁珠富集微生物的菌落计数及细菌16S rDNA基因拷贝数进行分析,磁珠-微生物复合物中的绝对基因拷贝数和菌落总数均显著高于磁分离后上清液中基因拷贝数和菌落计数(P=0.0001).此方法对微生物具有一定的高效富集作用.

In order to study the monitoring method of low-concentration microorganisms in clean environment,three kinds of low concentrated Gram-positive and negative bacteria,were prepared to evaluate the enriched and concentrated efficiency of microorganisms.The experiment showed the bacteria concentrated by the modified magnetic beads had a higher enrichment and concentration ratio compared with E.coli and S.aureus.After non-specific enrichment of microorganisms in tap water,drinking water and bottled water,there were significant difference among these three types of water for the 16S rDNA gene copy numbers(P<0.01),and the significant difference was also observed in the total number of colonies between the supernatant and the enriched microbial complex(P<0.001).Significant difference between filtering method and magnetic-bead enrichment(P=0.002)for tap water,but no significant differences for drinking and bottled water(P=0.362).The colony counting and 16S rDNA gene copy number of bacteria enriched by magnetic beads were analyzed.The absolute gene copy numbers and the total number of colonies in the magnetic-bead microbial complex were significantly higher than those in the supernatant after magnetic separation(P=0.0001).This method has some high efficiency enrichment effect on microorganisms.