Impact of veterinary antibiotics on plasmid-encoded antibiotic resistance transfer

Abstract

Objectives: Resistance genes can be genetically transmitted and exchanged between commensal and pathogenic bacterial species, and in different compartments including the environment, or human and animal guts (One Health concept). The aim of our study was to evaluate whether subdosages of antibiotics administered in veterinary medicine could enhance plasmid transfer and, consequently, resistance gene exchange in gut microbiota.

Methods: Conjugation frequencies were determined with Escherichia coli strains carrying IncL- (blaOXA-48) or IncI1-type (blaCTX-M-1) plasmids subjected to a series of subinhibitory concentrations of antibiotics used in veterinary medicine, namely amoxicillin, ceftiofur, apramycin, neomycin, enrofloxacin, colistin, erythromycin, florfenicol, lincomycin, oxytetracycline, sulfamethazine, tiamulin and the ionophore narasin. Treatments with subinhibitory dosages were performed with and without supplementation with the antioxidant edaravone, known as a mitigator of the inducibility effect of several antibiotics on plasmid conjugation frequency (PCF). Expression of SOS-response associated genes and fluorescence-based reactive oxygen species (ROS) detection assays were performed to evaluate the stress oxidative response.

Results: Increased PCFs were observed for both strains when treating with florfenicol and oxytetracycline. Increased expression of the SOS-associated recA gene also occurred concomitantly, as well as increased ROS production. Addition of edaravone to the treatments reduced their PCF and also showed a decreasing effect on SOS and ROS responses for both plasmid scaffolds.

Conclusions: We showed here that some antibiotics used in veterinary medicine may induce transfer of plasmid-encoded resistance and therefore may contribute to the worldwide spread of antibiotic resistance genes.

Figure 1.

ROS experiment by fluorimetry. (a) ROS production for E. coli (IncL) after treatment with antibiotics; (b) ROS production for E. coli (IncL) after incubation with florfenicol, with and without edaravone; (c) ROS production for E. coli (IncL) after incubation with oxytetracycline, with and without edaravone; (d) ROS production for E. coli (IncI1) after treatment with antibiotics; (e) ROS production for E. coli (IncI1) after incubation with florfenicol, with and without edaravone; (f) ROS production for E. coli (IncI1) after incubation within oxytetracycline, with and without edaravone. IF/OD, intensity of fluorescence/OD; CTRL, control (no treatment); FLO, florfenicol (4 mg/L for both isolates); OTC, oxytetracycline (IncL, 0.5 mg/L; IncI1, 256 mg/L), NEO, neomycin (1 mg/L for both isolates); EDA, edaravone (0.1 mM). Data are presented as mean and SD. */#, P < 0.05 (*, against the control; #, against the treated only with antibiotics); **/##, P ≤ 0.01; ***/###, P ≤ 0.001; ****/####, P ≤ 0.0001.