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Evaluation of method utilizing 16S rRNA amplicon sequencing for analysis of live bacterial community


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A combination of selective membrane-permeable dye, propidium monoazide (PMA) with PCR (PMA-PCR) and RNA targeted reverse transcription with PCR (RT-PCR) have been developed to distinguish viable from dead bacteria in various water environments. The objective of this study was to investigate the effective method to detect viable bacterial community using 16S rRNA amplicon sequencing combined with PMA-PCR and RT-PCR. Microbial community analysis of heat-treated influent spiked with two viable enteric bacteria showed that 16S rRNA sequencing combined with PMA-PCR and RT-PCR methods were applicable to the detection of physiologically active bacterial community. RT-PCR method was able to detect active bacteria more selectively than PMA-PCR method. In contrast, nested-PCR which composed of two-step PCR after PMA treatment showed no difference in the microbial community from PMA-PCR method. Four environmental samples were assessed the viability of bacterial community and the relationship between the abundance and activity using PCR, PMA-PCR and RT-PCR method.


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  • Article Type
    departmental bulletin paper
  • Data Source
    • IRDB
    • CiNii Articles

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