Abstract
Alternation of bacterial antioxidant defense pathways might affect susceptibility to antibiotics in dual ways. Using a relatively simple model based on wild-type and oxyR Escherichia coli mature biofilms, their counterpart planktonic cultures and exponentially growing planktonic cultures, we explored the role of OxyR-mediated metabolism alternations in modulation of susceptibility to antibiotics ciprofloxacin and cefotaxime. All three types of cultures were placed in fresh medium,1 h after antibiotics were added and incubation continued further for 2 h. Killing rates of antibiotics were determined, biofilm eradication using crystal violet assay was estimated, expression of rpoS, katG, sulA genes as well as HPI and HPII catalase activity were measured. Biofilms of both strains were more recalcitrant to ciprofloxacin and cefotaxime at all tested concentrations compared to exponentially growing planktonic cultures. In oxyR biofilms killing rate of ciprofloxacin was lower, and killing rate of cefotaxime was higher compared to the parental strain. Compared to biofilms, wild-type biofilm counterpart planktonic cultures showed higher tolerance to low doses of ciprofloxacin, while oxyR plankton demonstrated higher tolerance to cefotaxime. Higher recalcitrance of oxyR biofilms to ciprofloxacin may be caused by an increase in persister cells under conditions of enhanced oxidative stress and activated SOS response.
Highlights
Ciprofloxacin- and cefotaxime-treated urinary tract infections are often associated with biofilmproducing bacteria such as Escherichia coli which demonstrated resistance to both antibiotics in 49.9 and 28% of described cases, respectively [1]
OxyR factor stimulates transcription of genes involved in bacterial defense against peroxide stress, decreasing the levels of H2O2 and unincorporated iron, thereby reducing DNA damage caused by Fenton chemistry and preventing cell death [8]
Determination of β-galactosidase and catalase activity We explored the role of OxyR, RpoS and SOS regulons in susceptibility to antibiotics measuring βgalactosidase activity in reporter strains carrying fusions of the genes katG, rpoS and sulA with the gene lacZ [17] using a SmartSpec Plus Spectrophotometer (Bio-Rad, USA)
Summary
Ciprofloxacin- and cefotaxime-treated urinary tract infections are often associated with biofilmproducing bacteria such as Escherichia coli which demonstrated resistance to both antibiotics in 49.9 and 28% of described cases, respectively [1]. Determination of β-galactosidase and catalase activity We explored the role of OxyR, RpoS and SOS regulons in susceptibility to antibiotics measuring βgalactosidase activity in reporter strains carrying fusions of the genes katG, rpoS and sulA with the gene lacZ [17] using a SmartSpec Plus Spectrophotometer (Bio-Rad, USA). To previously published data [9], oxyR mutant demonstrated an increase in biofilm formation after 22 h of static incubation compared to the wild-type strain.
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