Doré J, Sghir A, Hannequart-Gramet G, Corthier G, Pochart P
Laboratoire d'Ecologie et Physiologie du Système digestif, Institut National de la Recherche Agronomique, Jouy-en-Josas, France.
Syst Appl Microbiol. 1998 Mar;21(1):65-71. doi: 10.1016/S0723-2020(98)80009-X.
Colonic Bacteroides include several species which, by their population level and activities, are significant contributers to the metabolic activity and health of man and animals. Yet, the understanding of their ecology has been hampered by the lack of highly specific and reliable enumeration techniques. Based on 16S rRNA sequence comparisons within the available database, we have designed an 18-mer oligonucleotide that targets a region common to-and specific for the Bacteroides-Porphyromonas-Prevotella group. We have tested the specificity of the probe and its usefulness for studies of human faecal samples. Under experimentally optimized hybridization conditions, the probe was shown to similarly recognize the rDNA obtained from 40 strains representing 8 species of the Bacteroides-Porphyromonas-Prevotella group. Importantly, it did not recognize 31 strains of microorganisms representing 8 genera of the dominant human faecal microbiota. Among selected colonies of dominant microorganisms of the faecal flora of two human individuals, strains identified as B. vulgatus by immunoblots using a species-specific monoclonal antibody were all detected by the probe. Colony hybridization was used to enumerate total Bacteroides-group microorganisms in faecal specimen from children and adults. The probe described therein was further used in quantitative RNA blots to monitor fluctuations of the Bacteroides-group versus Bifidobacterium genus in frozen faecal samples from a child between 85 and 125 days of age. It will be applicable to similar investigations of other anaerobic environments.
结肠拟杆菌包括几个菌种，就其数量水平和活性而言，它们对人和动物的代谢活动及健康有重要贡献。然而，由于缺乏高度特异性和可靠的计数技术，对其生态学的理解受到了阻碍。基于现有数据库内的16S rRNA序列比较，我们设计了一种18聚体寡核苷酸，它靶向拟杆菌 - 卟啉单胞菌 - 普雷沃菌属共有的且特异的一个区域。我们测试了该探针的特异性及其在人类粪便样本研究中的实用性。在实验优化的杂交条件下，该探针能同样识别从代表拟杆菌 - 卟啉单胞菌 - 普雷沃菌属8个菌种的40株菌株中获得的rDNA。重要的是，它不能识别代表人类粪便中主要微生物群8个属的31株微生物。在两个人类个体粪便菌群的优势微生物的选定菌落中，用种特异性单克隆抗体通过免疫印迹鉴定为普通拟杆菌的菌株均被该探针检测到。菌落杂交用于计数儿童和成人粪便标本中拟杆菌属微生物的总数。其中所述的探针进一步用于定量RNA印迹，以监测一名85至125日龄儿童的冷冻粪便样本中拟杆菌属与双歧杆菌属的波动情况。它将适用于其他厌氧环境的类似研究。
