Long-Term Persistence of blaCTX-M-15 in Soil and Lettuce after Introducing Extended-Spectrum β-Lactamase (ESBL)-Producing Escherichia coli via Manure or Water
Abstract
:1. Introduction
2. Materials and Methods
2.1. Resistance Testing and Preparation of Inoculation Cultures
2.2. Lettuce-in-Pot: Growth Conditions and Inoculation
2.2.1. Setup
2.2.2. Water-Inoculation Experiments
2.2.3. Manure-Inoculation Experiment
2.3. Sampling and Culture-Based Approach
2.3.1. Starting Materials and Sampling Scheme
2.3.2. Lettuce Leave Sampling
2.3.3. Soil Sampling
2.4. ARG Detection in Re-Isolated E. coli pEK499 and Secondary MDR Colonies
2.5. Molecular Detection of blaCTX-M-15 in Plant and Soil
2.6. Statistics
3. Results
3.1. Survival of E. coli pEK499 in Lettuce-in-Pot System
3.1.1. Pre-Inoculation
3.1.2. From Inoculation to Harvest
3.2. Detection of blaCTX-M-15 in the Lettuce-in-Pot System
3.2.1. Pre-Inoculation
3.2.2. From Inoculation to Harvest
3.3. ARG Detection in Re-Isolated E. coli and Secondary Colonies
4. Discussion
4.1. Effect of Low- and High-Level Contamination by Irrigation
4.1.1. Survival Dynamics of ESBL-E. coli pEK499
4.1.2. Persistence of blaCTX-M-15
4.2. Effect of Contaminated Manure Application
4.2.1. Survival Dynamics of ESBL-E. coli pEK499
4.2.2. Persistence of blaCTX-M-15
4.3. Good Agricultural Practice Recommendations
5. Conclusions
Author Contributions
Funding
Acknowledgments
Conflicts of Interest
Appendix A
Primer | Sequence (5’-3’) | Product Size (bp) | Origin |
---|---|---|---|
aadA5_F | CTG CGA CAC TGG ACA CAA TC | 502 | This study |
aadA5_R | CGA GCA AGA GCA AGA ACG AC | ||
CTX-M-15_F | CCT TAG GTT GAG GCT GGG TG | 760 | This study |
CTX-M-15_R | TTG TTA GGA AGT GTG CCG CT | ||
tetA_F | CAG GCA GAG CAA GTA GAG GG | 634 | This study |
tetA_R | CCT CAA TTT CCT GAC GGG CT | ||
27F | AGA GTT TGA TCM TGG CTC AG | 1505 | [65] |
1492R | CGG TTA CCT TGT TAC GAC TT |
98 °C | Initial Denaturation | 30 sec | |
---|---|---|---|
98 °C | Denaturation | 10 sec | 30 cycles |
65 °C | Annealing | 30 sec | |
72 °C | Extension | 30 sec | |
72 °C | Final Extension | 10 min |
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Gekenidis, M.-T.; Rigotti, S.; Hummerjohann, J.; Walsh, F.; Drissner, D. Long-Term Persistence of blaCTX-M-15 in Soil and Lettuce after Introducing Extended-Spectrum β-Lactamase (ESBL)-Producing Escherichia coli via Manure or Water. Microorganisms 2020, 8, 1646. https://doi.org/10.3390/microorganisms8111646
Gekenidis M-T, Rigotti S, Hummerjohann J, Walsh F, Drissner D. Long-Term Persistence of blaCTX-M-15 in Soil and Lettuce after Introducing Extended-Spectrum β-Lactamase (ESBL)-Producing Escherichia coli via Manure or Water. Microorganisms. 2020; 8(11):1646. https://doi.org/10.3390/microorganisms8111646
Chicago/Turabian StyleGekenidis, Maria-Theresia, Serena Rigotti, Jörg Hummerjohann, Fiona Walsh, and David Drissner. 2020. "Long-Term Persistence of blaCTX-M-15 in Soil and Lettuce after Introducing Extended-Spectrum β-Lactamase (ESBL)-Producing Escherichia coli via Manure or Water" Microorganisms 8, no. 11: 1646. https://doi.org/10.3390/microorganisms8111646