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

Leena Malayil,Emmanuel F Mongodin,Amy R Sapkota,Suhana Chattopadhyay

doi:10.1186/s40793-021-00382-1

Leena Malayil, Emmanuel F Mongodin + Show 2 more

Open Access

https://doi.org/10.1186/s40793-021-00382-1

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Abstract

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.

Highlights

As global freshwater resources are rapidly being depleted—due to population growth, climate change, over pumping of aquifers and other factors—states and nations are relying more heavily on nontraditional irrigation water sources to ensure agricultural water security and prevent foodMalayil et al Environmental Microbiome (2021) 16:13 sorghum, soybeans and tomatoes [29, 32, 57, 58, 64]
V. parahaemolyticus isolates were predominantly obtained from tidal brackish water (9 isolates), and two isolates were obtained from reclaimed water
The nontraditional irrigation water sources tested in our study harbored diverse bacterial communities, and these water sources would likely require treatment prior to their use for food crop irrigation

Summary

Introduction

As global freshwater resources are rapidly being depleted—due to population growth, climate change, over pumping of aquifers and other factors—states and nations are relying more heavily on nontraditional irrigation water sources (e.g., recycled water, brackish water) to ensure agricultural water security and prevent foodMalayil et al Environmental Microbiome (2021) 16:13 sorghum, soybeans and tomatoes [29, 32, 57, 58, 64]. Brackish water sources are known to harbor important human pathogens. Vibrio spp. have been recovered from surface waters, such as rivers, creeks, and irrigation canals [37, 52, 65], as well as reclaimed water [25, 52, 53]. Human Vibrio infections can occur among people consuming raw or undercooked shellfish and among those working or recreating in contaminated waters [14, 15, 17, 56]. If Vibrio-contaminated water is used to irrigate food crops that are eaten raw, this practice could represent an additional exposure pathway for human Vibrio infections [28, 76]

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