Modulation of Streptomycin Killing Rate against Mature Escherichia Coli Biofilms in the Presence of Medicinal Plant Extracts

Background. Medicinal plant extracts exhibiting pro- and antioxidant properties may affect antibiotic-induced killing of biofilm-producing bacteria in both synergistic and antagonistic modes. Better understanding of these alternations is required to adjust antibiotic therapy and herbal medicine in order to exclude unwanted losses of antibiotic efficiency.Aim: to study modulation modes of streptomycin killing rate against mature biofilms of Escherichia coli in the presence of different doses of commonly used medicinal plant extracts.Materials and methods. Pharmacodynamic parameter killing rate and mass biofilm formation were determined in the presence of streptomycin and medicinal plant extracts.Results. Synergism was found between 100 mg/ml streptomycin and low doses (0.83 mg of dry herb/ml) of green, black tea, Arctostaphylos uva-ursi, Betula pendula and Laminaria japonica against killing mature biofilms. Alternatively, high doses (6.64 mg of dry herb/ml) of green, black tea and Vaccinium vitis-ideae demonstrated antagonism, decreasing killing rate and enhancing biofilm formation. Presumably, high doses of the extracts were sufficient to enhance biofilm formation blocking penetration of streptomycin through enlarged biofilm matrix and diminishing the killing rate.Conclusions. Widely consumed as soft beverages or for prophylactic purposes green, black tea and V. vitis-ideae could promote strong antagonistic effects with streptomycin. These extracts can stimulate biofilm production, making benefit for commensal microbiota, but have clinical relevance due to a significant reduction in the lethal efficiency of streptomycin in biofilms of pathogenic strains. This highlights the need of careful antibiotic prescription scheme adjustment when choosing appropriate combinations of plant extracts and antibiotics to achieve a synergistic effect

Study of the effect of protein synthesis inhibitors on growing <i>Escherichia coli</i> bacteria using electrochemical sensors

Background. The study of the mechanisms of action of antibiotics requires the integrated use of traditional microbiological and physicochemical methods.   The aim. To study the response of Escherichia coli bacteria to the action of four antibiotics, inhibitors of protein synthesis, using combined approach.   Methods. Bacteria were grown under aerobic conditions on minimal M9 medium with glucose. Tetracycline, kanamycin, streptomycin and chloramphenicol have been tested. The effect of antibiotics on survival (CFU) and growth rate was determined. Respiratory activity, sulfide production, extracellular potassium, as well as pH and Eh of the medium were measured using electrochemical sensors directly in a growing culture in the “real time”, membrane potential was measured using a DiBAC dye and a Leica DM2000 fluorescent microscope.   Results. The tested antibiotics were divided into two groups according to their properties. Tetracycline and chloramphenicol showed a pronounced bacteriostatic effect, growth inhibition began immediately after the addition of antibiotics and occurred at a high rate. Both antibiotics inhibited respiration, stimulated sulfide production and an Eh jump. Bacteria treated with tetracycline and chloramphenicol retained the ability to maintain membrane potential and intracellular potassium better. Inhibition of respiration led to a decrease in glucose catabolism, as evidenced by a lower rate of acidification of the medium compared to the control. Growth inhibition with streptomycin and kanamycin was initiated with a 30-minute delay. Both antibiotics showed a bactericidal effect, did not stimulate sulfide production and Eh jump, did not inhibit respiration, but caused a drop in membrane potential and intracellular potassium. High respiratory activity promoted glucose catabolism, as evidenced by the rapid acidification of the medium. Of interest is the detection of kanamycin-induced sulfide production during E. coli growth on MOPS medium.   Conclusion. An analysis of the data obtained indicates that the use of electrochemical sensors in combination with traditional methods is a promising approach to studying the mechanisms of action of antibiotics

MICROBIAL TEST-SYSTEMS FOR EVALUATION OF E. COLI SUSCEPTIBILTY TO ANTIBIOTICS

Susceptibility of Escherichia coli to antibiotics was estimated by determination of the minimum inhibitory concentrations (MICs). To evalu-ate gene expression we measured в-galactosidase activity in strains carrying gene fusions using plate reader. It has been shown that treatment with polyphe-nols and some plant extracts significantly decreased bacteriostatic effects of antibiotics, to a larger extent, ciprofloxacin. A positive correlation was found among protective effects of polyphenols and extracts and their antioxidant properties.Работа поддержана грантами Президента РФ МК1763.2012.4, РФФИ-Урал № 10-04-96017, а также грантом Президиума УрО РАН для молодых ученых 11-4-ИП-181

IN GLUTATHIONE-DEFICIENT STRAIN ESCHERICHIA COLI UNDER ANTIBIOTICS TREATMENT

The results of the present study indicate that cell treatment with antibiotics leads to Eh shift and change of oxygen pressure level in glutathione-deficient strain.Работа выполнена при поддержке грантом Президиума УрО РАН для молодых ученых 14-4-НП-126, а также грантом РФФИ-Урал №14-04-96031

INVESTIGATING THE ROLE OF STRESS-REGULONS IN MODULATING EFFECTS OF PLANT EXTRACTS ON BACTERIAL BIOFILMS

Among 18 plant extracts studied black tea, A. uva ursi, V. vitis-idaea, T. сordata, B. pendula and Z. mays stimulated biofilm formation, while the extracts of A. millefolium, U. dioica and L. japonica demonstrated inhibiting effects. Some of the extracts also inhibited rpoS expression both in biofilms and planktonic cultures. To elucidate the role of RpoS sigma-S factor in the observed effects of the plant extracts, we studied influence of different doses of pure plant polyphenol quercetin on biofilm formation in strains with different levels of RpoS. It was found that negative influence on biofilm formation was due to the RpoS excess. Collectively, our data indicate the possibility of RpoS-independent pathway of biofilm formation regulation in the pres-ence of plant extracts containing polyphenols.Исследование выполнено при поддержке грантом РФФИ-Урал № 14-04-96031, грантом №12-И-4-2072 по Программе интеграционных проектов Президиума УрО РАН, а также грантом Президиума УрО РАН для молодых ученых 14-4-НП-126