Application of Pseudomurein Endoisopeptidase to Fluorescence In Situ Hybridization of Methanogens within the Family Methanobacteriaceae

• DOI: 10.1128/aem.01499-06
• Publication Date: 2006-11-06T18:53:46Z
• Authors: Kohei Nakamura; Takeshi Terada; Yuji Sekiguchi; Naoya Shinzato; Xian-Ying Meng; Miho Enoki; Yoichi Kamagata
• Keywords: Methanobacterium

ABSTRACT In situ detection of methanogens within the family Methanobacteriaceae is sometimes known to be unsuccessful due difficulty in permeability oligonucleotide probes. Pseudomurein endoisopeptidase (Pei), a lytic enzyme that specifically acts on their cell walls, was applied prior 16S rRNA-targeting fluorescence hybridization (FISH). For this purpose, pure cultured family, Methanobacterium bryantii , Methanobrevibacter ruminantium Methanosphaera stadtmanae and Methanothermobacter thermautotrophicus together with -containing syntrophic acetate-oxidizing coculture, endosymbiotic an anaerobic ciliate, upflow sludge blanket (UASB) granule were examined. Even without Pei treatment, cells are relatively well hybridized However, almost none cocultured UASB hybridized. treatment able increase probe ratio every specimen, particularly specimen had shown little hybridization. Interestingly, hybridizing signal intensity coculture H 2 -producing syntroph significantly improved by pretreatment, whereas culture same strain. We found difference attributed differences wall thicknesses between two conditions. These results indicate effective for FISH analysis show impermeability probe.