Protein identification before and after glyphosate exposure in Lolium multiflorum genotypes.

BACKGROUND: Weeds reduce crop yields, and among the methods used to control these plants, the use of chemicals is preferred. However, the repeated application of herbicides with the same mechanism of action selects for resistant populations. The aim of this study was to evaluate glyphosate resistance in Lolium multiflorum (Lam.) and relate the resistance to protein expression in the absence and presence of the herbicide using a metabolic-proteomic approach. RESULTS: Glyphosate resistance was confirmed, with a sevenfold difference in resistance between susceptible and resistant genotypes. Among the possible mechanisms affecting resistance, mutations in the enzyme 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS), herbicide differential translocation and overexpression of EPSPS are suggested. Susceptible plants had higher growth than did resistant plants in the absence of the herbicide, in addition to greater expression of protein groups related to photosynthesis and to tolerance to biotic and abiotic stresses. With application of glyphosate, resistant plants maintained their metabolism and began to express EPSPS and other candidate proteins related to herbicide resistance. CONCLUSIONS: In the absence of glyphosate, the susceptible plants would replace the resistant plants over time, and abiotic or biotic stresses would accelerate this process. Resistance in plants resulted from a combination of target-site and non-target-site resistance mechanisms. We identified several candidate proteins that could be investigated in future studies on glyphosate resistance. © 2017 Society of Chemical Industry.

Emergence of Hyper-Resistant Escherichia coli MG1655 Derivative Strains after Applying Sub-Inhibitory Doses of Individual Constituents of Essential Oils.

The improvement of food preservation by using essential oils (EOs) and their individual constituents (ICs) is attracting enormous interest worldwide. Until now, researchers considered that treatments with such antimicrobial compounds did not induce bacterial resistance via a phenotypic (i.e., transient) response. Nevertheless, the emergence of genotypic (i.e., stable) resistance after treatment with these compounds had not been previously tested. Our results confirm that growth of Escherichia coli MG1655 in presence of sub-inhibitory concentrations of the ICs carvacrol, citral, and (+)-limonene oxide do not increase resistance to further treatments with either the same IC (direct resistance) or with other preservation treatments (cross-resistance) such as heat or pulsed electric fields (PEF). Bacterial mutation frequency was likewise lower when those IC's were applied; however, after 10 days of re-culturing cells in presence of sub-inhibitory concentrations of the ICs, we were able to isolate several derivative strains (i.e., mutants) displaying an increased minimum inhibitory concentration to those ICs. Furthermore, when compared to the wild type (WT) strain, they also displayed direct resistance and cross-resistance. Derivative strains selected with carvacrol and citral also displayed morphological changes involving filamentation along with cell counts at late-stationary growth phase that were lower than the WT strain. In addition, co-cultures of each derivative strain with the WT strain resulted in a predominance of the original strain in absence of ICs, indicating that mutants would not out-compete WT cells under optimal growth conditions. Nevertheless, growth in the presence of ICs facilitated the selection of these resistant mutants. Thus, as a result, subsequent food preservation treatments of these bacterial cultures might be less effective than expected for WT cultures. In conclusion, this study recommends that treatment with ICs at sub-inhibitory concentrations should be generally avoided, since it could favor the emergence of hyper-resistant strains. To ascertain the true value of EOs and their ICs in the field of food preservation, further research thus needs to be conducted on the induction of increased transient and stable bacterial resistance via such antimicrobial compounds, as revealed in this study.

Inhibition of Staphylococcus aureus cocktail using the synergies of oregano and rosemary essential oils or carvacrol and 1,8-cineole.

This study assessed the inhibitory effects of the essential oils (EOs) from Origanum vulgare L. (OVEO) and Rosmarinus officinalis L. (ROEO), as well as of the its majority individual constituents (ICs) carvacrol (CAR) and 1,8-cineole (CIN), respectively, combined at subinhibitory concentrations against a cocktail of Staphylococcus aureus. The Minimum inhibitory Concentration (MIC) of both OVEO and CAR against S. aureus cocktail was 1.25 µL/mL, while for ROEO and CIN the MIC value was 10 µL/mL. The Fractional Inhibitory Concentration Index of the combined EOs or ICs was ≤0.5 indicating a synergic interaction. The incorporation of OVEO and ROEO or CAR and CIN at different combinations in cheese and meat broths caused a decrease (p ≤ 0.05) of initial counts of S. aureus. Combined application of 1/8 MIC OVEO and 1/4 MIC ROEO or 1/4 MIC CAR and 1/4 MIC CIN in meat and cheese samples reduced (p ≤ 0.05) the viable cells counts and caused morphological changes in S. aureus cells, such as cell shrinkage and appearance of blebbing-like structures on cell surfaces. However, in cheese and meat samples the decrease in viable cell counts was smaller (p ≤ 0.05) than that observed in cheese and meat broths. These findings reinforce the potential of the use of OVEO and ROEO or CAR and CIN in combination to control S. aureus in cheese and meat matrices.