rRNA-targeted fluorescent in situ hybridization analysis of bacterial community structure in river water

• Published in: Microbiology (Society for General Microbiology) Vol. 144; no. 8; pp. 2085 - 2093
• Main Authors: Kenzaka, Takehiko, Yamaguchi, Nobuyasu, Tani, Katsuji, Nasu, Masao
• Format: Journal Article
• Language: English
• Published: Reading Soc General Microbiol 01.08.1998; Society for General Microbiology
• Subjects: Acinetobacter; Aeromonas; Alcaligenes; Bacteria - genetics; Bacteria - isolation & purification; Bacteriological methods and techniques used in bacteriology; Bacteriology; Biological and medical sciences; Burkholderia; Cytophaga; Diazonium Compounds; Flavobacterium; Fluorescein-5-isothiocyanate; Fluorescent Dyes; Fresh Water; Fundamental and applied biological sciences. Psychology; In Situ Hybridization, Fluorescence; Microbiology; Naphthalenes; Oligonucleotide Probes - chemical synthesis; Pseudomonas; RNA Probes - chemical synthesis; RNA, Ribosomal, 16S - analysis; RNA, Ribosomal, 23S - analysis; Species Specificity; Vibrio; Water Microbiology; Freshwater environment; Molecular hybridization; Investigation method; Fluorescent labelling; Ribosomal RNA; Bacteria; Population structure; River water; Microbial community
• ISSN: 1350-0872; 1465-2080
• DOI: 10.1099/00221287-144-8-2085

Faculty of Pharmaceutical Sciences, Osaka University, 1-6 Yamada-oka, Suita, Osaka 565-0871, Japan ABSTRACT An improved in situ hybridization technique, HNPP-FISH, using 2-hydroxy-3-naphthoic acid 2'-phenylanilide phosphate (HNPP) and Fast Red TR was applied to analyse the community structure of planktonic bacteria in river water. Oligonucleotide probes specific for the domain Bacteria (EUB338) and five bacterial groups [ Flavobacterium-Cytophaga; Burkholderia-Pseudomonas (rRNA III)-authentic Alcaligenes; Vibrio-Aeromonas; Pseudomonas (rRNA I); the genus Acinetobacter ] were used to investigate the bacterial community structure at two sites differing in organic carbon pollution level. At the eutrophic site, 54-68% of all cells visualized by staining with DAPI (4',6-diamidino-2-phenylindole) could be detected with probe EUB338. In samples from the oligotrophic site, 39-45% of the total cells hybridized with EUB338. At the eutrophic site, approximately 50% of the total cells were identified with the five group-specific probes; the bacterial community structure was dominated by the Flavobacterium-Cytophaga group and Burkholderia-Pseudomonas (rRNA III)-authentic Alcaligenes group. At the oligotrophic site, only 26-38% of the total cells were identified with the five group-specific probes. The community structure at the oligotrophic site was similar to that at the eutrophic site, although the percentage of EUB338-detectable cells differed. No appreciable change was found in the community structure during the sampling period at either site. The improved HNPP-FISH technique should be a useful tool for the analysis of microbial community composition. * Author for correspondence: Masao Nasu. Tel: +81 6 879 8170. Fax: +81 6 879 8174. e-mail: nasu@phs.osaka-u.ac.jp Keywords: rRNA-targeted oligonucleotide probes, fluorescent in situ hybridization, river, bacterial community structure