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  • Title: Quantification of viable but nonculturable bacterial pathogens in anaerobic digested sludge.
    Author: Jiang Q, Fu B, Chen Y, Wang Y, Liu H.
    Journal: Appl Microbiol Biotechnol; 2013 Jul; 97(13):6043-50. PubMed ID: 22996281.
    Abstract:
    Enteric bacterial pathogens in sewage sludge easily become viable but nonculturable (VBNC) during anaerobic digestion, which escape detection by standard culture methods and pose a potential health risk. In this study, a method that is combining the standard culture method with the reverse transcription quantitative PCR (RT-qPCR) assay was developed for the quantification of bacterial pathogens in the VBNC state. The cycle threshold (CT) values from RT-qPCR assays were linear to the bacterial number in the range from 10⁹ to 10² most probable number (MPN) per reaction for Escherichia coli (R²=0.9964) and Salmonella typhimurium (R²=0.9938) and from 10⁹ to 10⁴ MPN per reaction for Shigella flexneri (R²=0.997), respectively. Mesophilic anaerobic digestion (MAD) caused the bacterial pathogens in sewage sludge entering into VBNC state with the incidence indexes of 0.01-1.12 for E. coli, 2.48-436.52 for S. typhimurium, and 4.17-6.61 for S. flexneri, respectively. Given different VBNC incidence indexes of bacterial pathogens in sewage sludge by MAD, the quantification results of VBNC pathogens using RT-qPCR could provide an improved evaluation of pathogen inactivation efficiency and biological safety in sludge anaerobic digestion.
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