Hidden Resistome: Enrichment Reveals the Presence of Clinically Relevant Antibiotic Resistance Determinants in Treated Wastewater-Irrigated Soils

Roberto B.M. Marano, Chhedi Lal Gupta, Tamar Cozer, Edouard Jurkevitch, Eddie Cytryn*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

26 Scopus citations


Treated-wastewater (TW) irrigation transfers antibiotic-resistant bacteria (ARB) to soil, but persistence of these bacteria is generally low due to resilience of the soil microbiome. Nonetheless, wastewater-derived bacteria and associated antibiotic resistance genes (ARGs) may persist below detection levels and potentially proliferate under copiotrophic conditions. To test this hypothesis, we exposed soils from microcosm, lysimeter, and field experiments to short-term enrichment in copiotroph-stimulating media. In microcosms, enrichment stimulated growth of multidrug-resistant Escherichia coli up to 2 weeks after falling below detection limits. Lysimeter and orchard soils irrigated in-tandem with either freshwater or TW were subjected to culture-based, qPCR and shotgun metagenomic analyses prior, and subsequent, to enrichment. Although native TW- and freshwater-irrigated soil microbiomes and resistomes were similar to each other, enrichment resulted in higher abundances of cephalosporin- and carbapenem-resistant Enterobacteriaceae and in substantial differences in the composition of microbial communities and ARGs. Enrichment stimulated ARG-harboring Bacillaceae in the freshwater-irrigated soils, whereas in TWW-irrigated soils, ARG-harboring γ-proteobacterial families Enterobacteriaceae and Moraxellaceae were more profuse. We demonstrate that TW-derived ARB and associated ARGs can persist at below detection levels in irrigated soils and believe that similar short-term enrichment strategies can be applied for environmental antimicrobial risk assessment in the future.

Original languageAmerican English
Pages (from-to)6814-6827
Number of pages14
JournalEnvironmental Science & Technology
Issue number10
StatePublished - 18 May 2021

Bibliographical note

Funding Information:
This study was funded in part by the Chief Scientist of the Israeli Ministry of Agriculture (grant agreement 20-03-0032) and by the PRIMA programme (DSWAP, grant agreement 1822) within the EU Horizon 2020 Framework Programme for Research and Innovation. RM was funded by the European Union’s Horizon 2020 research and innovation programme under the Marie Skłodowska-Curie grant agreement 675530. The authors would like to thank Dr. Daniel Kurtzman, Dr. Asher Bar Tal, Prof. Shmuel Assouline, and Ido Nitzan for logistic and technical support with the orchard and lysimeter experiments, Dr. Shlomo Blum for technical MALDI-TOF support, and Dr. Max Kolton for critical review of the paper.

Publisher Copyright:
© 2021 American Chemical Society.


  • antibiotic resistance
  • contaminants of emerging concern
  • microbiome
  • resistome
  • soil
  • treated-wastewater irrigation


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