Coupled DNA-labeling and sequencing approach enables the detection of viable-but-non-culturable Vibrio spp. in irrigation water sources in the Chesapeake Bay watershed

Environmental Microbiome

Emmanuel F. Mongodin¹  Leena Malayil²  Amy R. Sapkota²  Suhana Chattopadhyay² 

[1] Institute for Genome Sciences, University of Maryland School of Medicine, Baltimore, MD, USA;Maryland Institute for Applied Environmental Health, University of Maryland School of Public Health, College Park, MD, USA;

关键词: Viable-but-nonculturable;    Vibrio;    DNA-labeling;    Sequencing;    Reclaimed water;    Brackish water;    BrdU;   

DOI  :  10.1186/s40793-021-00382-1

来源: Springer

【 摘 要 】

Nontraditional irrigation water sources (e.g., recycled water, brackish water) may harbor human pathogens, including Vibrio spp., that could be present in a viable-but-nonculturable (VBNC) state, stymieing current culture-based detection methods. To overcome this challenge, we coupled 5-bromo-2′-deoxyuridine (BrdU) labeling, enrichment techniques, and 16S rRNA sequencing to identify metabolically-active Vibrio spp. in nontraditional irrigation water (recycled water, pond water, non-tidal freshwater, and tidal brackish water). Our coupled BrdU-labeling and sequencing approach revealed the presence of metabolically-active Vibrio spp. at all sampling sites. Whereas, the culture-based method only detected vibrios at three of the four sites. We observed the presence of V. cholerae, V. vulnificus, and V. parahaemolyticus using both methods, while V. aesturianus and V. shilonii were detected only through our labeling/sequencing approach. Multiple other pathogens of concern to human health were also identified through our labeling/sequencing approach including P. shigelloides, B. cereus and E. cloacae. Most importantly, 16S rRNA sequencing of BrdU-labeled samples resulted in Vibrio spp. detection even when our culture-based methods resulted in negative detection. This suggests that our novel approach can effectively detect metabolically-active Vibrio spp. that may have been present in a VBNC state, refining our understanding of the prevalence of vibrios in nontraditional irrigation waters.

【 授权许可】

CC BY