Identification and quantification of microbial contaminants of fecal origin are major priority in the control of drinking and recreational water qualities. In this study, we proposed a new PCR-based approach using 16S rRNA gene markers of enteric anaerobes, Bacteroides-Prevotella spp. for discriminating human, cow, and pig fecal contamination in environmental waters without culturing indicator organisms. Human-, cow-, and pig-specific Bacteroides-Prevotella 16S rRNA genetic markers were identified. We also investigated the relationships between level of host-specific Bacteroides markers and the presence of enteric bacterial pathogens. Additionally, the persistence of these genetic markers in non-host water environments was investigated.
16S rRNA-based molecular methods such as polymerase chain reaction (PCR), real-time and multiplex PCR, and terminal restriction fragment length polymorphism (T-RPLP) techniques were used to identify and quantify the host-specific Bacteroides-Prevotella. Additionally, we developed a novel fluorescence-based live/dead staining technique combined with Fluorescent in situ hybridization (FISH) to study the persistence of genetic markers in natural water.
Current research topics
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Publications（2007-）
- Estimation of concentration ratio of indicator to pathogen-related gene in environmental water based on left-censored data
Kato, T., Kobayashi, A., Ito, T., Miura, T., Ishii, S., Okabe, S. and Sano, D.
Journal of Water and Health, 2015, in press. - Microfluidic quantitative PCR for simultaneous quantification of multiple viruses in environmental water samples
Ishii, S., Kitamura, G., Segawa, T., Kobayashi, A., Miura, T., Sano, D. and Okabe, S.
Applied and Environmental Microbiology, 2014, 80(24), 7505-7511. - Water quality monitoring and risk assessment by simultaneous multipathogen quantification
Ishii, S., Nakamura, T., Ozawa, S., Kobayashi, A., Sano, D. and Okabe, S.
Environmental Science and Technology, 2014, 48(9), 4744-4749. - Use of a genetically-engineered Escherichia coli strain as a sample process control for quantification of the host-specific bacterial genetic markers
Kobayashi, A., Sano, D., Taniuchi, A., Ishii, S. and Okabe, S.
Applied Microbiology and Biotechnology, 2013, 97(20), 9165-9173. - Chicken- and duck-associated Bacteroides-Prevotella genetic markers for detecting fecal contamination in environmental water
Kobayashi, A., Sano, D., Hatori, J., Ishii, S. and Okabe, S.
Applied Microbiology and Biotechnology, 2013, 97(16), 7427-7437. - Simultaneous quantification of multiple food and waterborne pathogens by use of microfluidic quantitative PCR
Ishii, S., Segawa, T. and Okabe, S.
Applied and Environmental Microbiology, 2013, 79(9), 2891-2898. - Effects of temperature and predator to the persistence of host-specific Bacteroides-Prevotella genetic markers in water
Kobayashi, A., Sano, D. and Okabe, S.
Water Science and Technology, 2013, 67(4), 838-845. - Qualitative and quantitative estimation of host-specific fecal pollution using Bacteroides-Prevotella 16S rRNA genetic markers by T-RFLP and real-time PCR analyses
Savichtcheva, O. and Okabe, S.
Water Science and Technology, 2009, 59(9), 1831-1840. - Persistence of host-specific Bacteroides-Prevotella 16S rRNA genetic markers in environmental waters: Effects of temperature and salinity
Okabe, S., and Shimazu, Y.
Applied Microbiology and Biotechnology, 2007, 76(4), 935-944. - Quantification of host-specific Bacteroides-Prevotella 16S rRNA genetic markers for assessment of fecal pollution in freshwater
Okabe, S., Okayama, N., Savichtcheva, O., and Ito, T.
Applied Microbiology and Biotechnology, 2007, 73(4), 890-901. - Relationships between Bacteroides 16S rRNA genetic markers and presence of bacterial enteric pathogens and conventional fecal indicators
Savichtcheva, O., Okayama, N., and Okabe, S.
Water Research, 2007, 41(16), 3615-3628.
