Abstract
Herbicides are frequently released into both rural and urban environments. Commercial herbicide formulations induce adaptive changes in the way bacteria respond to antibiotics. Salmonella enterica sv. Typhimurium and Escherichia coli were exposed to common co-formulants of formulations, and S. enterica sv. Typhimurium was exposed to active ingredients dicamba, 2,4-D and glyphosate to determine what ingredients of the commercial formulations caused this effect. Co-formulants Tween80 and carboxymethyl cellulose induced changes in response, but the pattern of the responses differed from the active ingredients, and effect sizes were smaller. A commercial wetting agent did not affect antibiotic responses. Active ingredients induced changes in antibiotic responses similar to those caused by complete formulations. This occurred at or below recommended application concentrations. Targeted deletion of efflux pump genes largely neutralized the adaptive response in the cases of increased survival in antibiotics, indicating that the biochemistry of induced resistance was the same for formulations and specific ingredients. We found that glyphosate, dicamba, and 2,4-D, as well as co-formulants in commercial herbicides, induced a change in susceptibility of the potentially pathogenic bacteria E. coli and S. enterica to multiple antibiotics. This was measured using the efficiency of plating (EOP), the relative survival of the bacteria when exposed to herbicide and antibiotic, or just antibiotic, compared to survival on permissive media. This work will help to inform the use of non-medicinal chemical agents that induce changes in antibiotic responses.
Highlights
The widespread use of antibiotics has led to antibiotic-resistant human pathogens
Bacteria were cultured at 37 C using LB Lennox (Invitrogen, Auckland, NZ) medium supplemented as appropriate with ampicillin (Amp; AppliChem, Dunedin, NZ), chloramphenicol (Cam; Sigma, Auckland, NZ), ciprofloxacin (Cip; Pentex, Auckland, NZ), kanamycin (Kan; Gibco, Auckland, NZ) or tetracycline (Tet; Sigma, Auckland, NZ)
Bacteria were spread onto solid media with increasing concentrations of antibiotic, with either no or a single defined concentration of active ingredient
Summary
The widespread use of antibiotics has led to antibiotic-resistant human pathogens. Multi-drug-resistant pathogens are a serious but common occurrence complicating treatment, increasing morbidity and mortality, and resulting in increased costs to health systems [1,2,3]. While mutations and gene acquisition through horizontal gene transfer that lead to changes in the minimum inhibitory concentration (MIC) of antibiotics in clinical settings have received the most attention, microbes in many environments are exposed to antibiotics at sub-inhibitory levels. This can have a wide range of physiological consequences [4]. Small increases in the MIC, either through mutations that incrementally increase resistance or through an adaptive response, extend the range of sub-lethal concentrations and the probability of treatment failure [5, 6]
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