Fluoroquinolone resistance in clinical and environmental isolates of Escherichia coli in Mexico City.

AIMS: To assess the different phenotypes and mechanisms of fluoroquinolone (FQ) resistance in clinical and environmental isolates of Escherichia coli. METHODS AND RESULTS: We compared FQ-resistant E. coli isolates, measuring minimal inhibitory concentrations (MIC) of ciprofloxacin, along with susceptibility to other antibiotics. We also searched for the presence of efflux pumps, using efflux inhibitors, and for plasmid-borne FQ-resistance by PCR. We found that, aside from the higher FQ-resistance prevalence among clinical strains, environmental ones resist much lower concentrations of ciprofloxacin. Efflux pumps mediate fluoroquinolone resistance as frequently among environmental isolates than in clinical strains. Plasmid-borne qnrA genes were not detected in any resistant strain. CONCLUSIONS: Environmental FQ-resistant strains may have a nonclinical origin and/or a selective pressure different from the clinical use of FQs. SIGNIFICANCE AND IMPACT OF THE STUDY: The identification of the source of low-level FQ-resistant strains (ciprofloxacin MIC c. 8 microg ml(-1)) in the environment could be important to curb the rapid emergence and spread of FQ-resistance in clinical settings, as these strains can easily become fully resistant to FQ concentrations achievable in fluids and tissues during therapy.

Efficacy and safety of sequential intravenous/oral moxifloxacin vs intravenous/oral amoxicillin/clavulanate for complicated skin and skin structure infections.

BACKGROUND: Complicated skin and skin structure infections (cSSSIs) are an important healthcare concern worldwide, as they can be life-threatening and challenging to treat. cSSSIs are normally managed using a combination of surgical intervention and prompt antibiotic use. New therapeutic options, including novel antibiotics, are required to improve outcomes in terms of duration of illness and to reduce the consumption of healthcare resources. METHODS: This was a prospective, randomized, open-label, parallel-group, multinational clinical study comparing sequential intravenous/oral (iv/po) moxifloxacin, 400 mg once daily, and iv amoxicillin/clavulanate, 1,000 mg/ 200 mg three times daily followed by po amoxicillin/ clavulanate, 500 mg/125 mg three times daily, for 7-21 days in hospitalized patients. RESULTS: A total of 804 patients were enrolled (mean age 51.8 years). The most common clinical diagnosis was complicated erysipelas (32.1% moxifloxacin; 30.0% amoxicillin/ clavulanate) and major abscess (31.1% moxifloxacin; 29.3% amoxicillin/clavulanate). Overall clinical success rates at the test-of-cure (TOC) visit (14-28 days post-treatment) for the per-protocol population (primary efficacy variable) were 80.6% (254/315) for patients in the moxifloxacin group and 84.5% (268/317) for those receiving amoxicillin/clavulanate (95% confidence interval [CI] -9.41, 2.18). Similar results were obtained for the intention-to-treat population (95% CI -7.56, 4.31). In both treatment groups, the highest clinical success rates were recorded for patients with complicated erysipelas, major abscess, surgical wound infection, and cellulitis. The lowest clinical cure rates were reported for diabetic foot infection and necrotizing fasciitis. In the microbiologically evaluable population, the bacteriological success rate (eradication and presumed eradication) was 76.0% (127/ 167) in the moxifloxacin group and 81.4% (140/172) in the amoxicillin/clavulanate group (95% CI -12.96, 4.41). Staphylococcus aureus (137 isolates) and Escherichia coli (50 isolates) were the most frequently isolated skin pathogens. Adverse event rates were comparable between treatment groups. CONCLUSIONS: Treatment with sequential iv/po moxifloxacin monotherapy once daily is clinically comparable to that with iv/po amoxicillin/clavulanate three times daily in the management of cSSSIs. Moxifloxacin's simple dose regimen offers an advantage over amoxicillin/clavulanate and represents a valuable addition to current antibiotic regimens used in the treatment of cSSSIs.

Differential activities of the SoxR protein of Escherichia coli: SoxS is not required for gene activation under iron deprivation.

When Escherichia coli cells are under superoxide stress, proteins SoxR and SoxS, acting sequentially, control the expression of a set of repair and defense genes. One of these genes, fumC, encoding fumarase C, was reported to be also activated by iron deprivation in a soxRS-dependent manner. However, the same condition failed to induce the expression of a soxS'::lacZ fusion. The expression of acnA (aconitase A) is also activated by SoxR alone when under iron deprivation, but not of sodA (Mn-superoxide-dismutase). SoxR completely inhibited the migration of a DNA fragment containing the promoter region of fumC, in gel-shift experiments. SoxR might bind to a different region than SoxS within the fumC promoter, or an unknown intermediate other than SoxS might be acting. It is possible that the regulatory role of SoxR is more complex than previously considered.

Comparative in vitro activity of moxifloxacin by E-test against Streptococcus pyogenes.

Macrolides are currently used to treat Streptococcus pyogenes infections where allergy or resistance prevents the use of penicillin. However, growing macrolide resistance is now seen worldwide, with rates of 5%-40% being reported. In this context it is therefore important to have other therapeutic options. The aim of this study was to ascertain the potential role of moxifloxacin, a third-generation fluoroquinolone, in the treatment of infections caused by group A S. pyogenes. The antimicrobial susceptibilities of S. pyogenes isolated from 197 adult patients with pharyngotonsillitis were analyzed by the E-test. Twelve percent of the isolates were resistant to macrolides, and 5% showed diminished susceptibility toward penicillin; none of the strains were resistant to cefotaxime or to moxifloxacin (90% minimum inhibitory concentration, 0.25 microg/mL). Therefore, moxifloxacin may be a therapeutic option in the management of S. pyogenes infections when penicillin cannot be used or when macrolide resistance may be a local issue. Clinical studies of moxifloxacin in pharyngotonsillitis are warranted.

Regulatory locus soxRS partially protects Escherichia coli against ozone.

Ozone is one of the major city air pollutants. Since it is known to induce the overexpression of superoxide-dismutase in various models, and is also a powerful oxidant, we tested if ozone can induce the expression of the soxRS regulon of Escherichia coli, which is activated by superoxide and nitric oxide. A sub-lethal exposure to ozone was unable to activate the expression of soxS'::lacZ transcriptional fusions. However, cells lacking the soxRS locus were more susceptible than wild-type to ozone-mediated killing. Constitutive expression of the soxRS regulon did not increase the resistance to ozone. Ozone might be exerting a selective pressure upon oxidative-stress defense mechanisms in airborne bacteria.

Mercury induces multiple antibiotic resistance in Escherichia coli through activation of SoxR, a redox-sensing regulatory protein.

Sub-inhibitory mercury concentrations are capable of partially activating SoxR, as shown by the augmented expression of a soxS'::lacZ fusion, and a diminished sensitivity to antibiotics caused by mercury treatment. Mercury may elevate the intracellular concentration of superoxide or perhaps act as a putative metal ligand for SoxR.

A possible role for plasmids in mediating the cell-cell proximity required for gene flux.

One of the major requirements for successful gene flux is a close proximity between participating organisms. In previous articles, we have proposed that plasmids act as powerful vehicles transporting genes collected by integration and transposition, mainly via the process of conjugation. However, in addition to conjugation, there are other processes, also mediated by plasmids, in which different cells come into very close contact with each other, such as symbiosis and the formation of multi-specific cellular communities. There is evidence that suggests that such intimate associations between cells may facilitate gene transfer events, even between distantly related organisms. Examples of symbiotic endosymbiotic, and parasitic associations provide evidence in support of the role of plasmids in bridging the genetic gap between species. In this purely theoretical article we attempt to conceptualize existing data on this subject, provide new insights and present testable predictions on how plasmids may facilitate gene flux by bringing cells together.

Two-stage control of an oxidative stress regulon: the Escherichia coli SoxR protein triggers redox-inducible expression of the soxS regulatory gene.

Escherichia coli responds to the redox stress imposed by superoxide-generating agents such as paraquat by activating the synthesis of as many as 80 polypeptides. Expression of a key group of these inducible proteins is controlled at the transcriptional level by the soxRS locus (the soxRS regulon). A two-stage control system was hypothesized for soxRS, in which an intracellular redox signal would trigger the SoxR protein as a transcriptional activator of the soxS gene and the resulting increased levels of SoxS protein would activate transcription of the various soxRS regulon genes (B. Demple and C.F. Amábile Cuevas, Cell 67:837-839, 1990). We have constructed operon fusions of the E. coli lac genes to the soxS promoter to monitor soxS transcription. Expression from the soxS promoter is strongly inducible by paraquat in a manner strictly dependent on a functional soxR gene. Several other superoxide-generating agents also trigger soxR(+)-dependent soxS expression, and the inductions by paraquat and phenazine methosulfate were dependent on the presence of oxygen. Numerous other oxidative stress agents (H2O2, gamma rays, heat shock, etc.) failed to induce soxS, while aerobic growth of superoxide dismutase-deficient bacteria triggered soxR-dependent soxS expression. These results indicate a specific redox signal for soxS induction. A direct role for SoxR protein in the activation of the soxS gene is indicated by band-shift and DNase I footprinting experiments that demonstrate specific binding of the SoxR protein in cell extracts to the soxS promoter. The mode of SoxR binding to DNA appears to be similar to that of its homolog MerR in that the SoxR footprint spans the -10 to -35 region of the soxS promoter.

Decreased resistance to antibiotics and plasmid loss in plasmid-carrying strains of Staphylococcus aureus treated with ascorbic acid.

The effect of ascorbic acid on plasmid-coded antibiotic resistance in Staphylococcus aureus was investigated. Several strains of S. aureus were cultured in the presence of 1 mM ascorbate for 6 h. This treatment induced an increased loss of resistance markers in 4 of 6 strains tested, and agarose gel electrophoresis showed this disappearance of plasmid DNA in ascorbate-induced susceptible colonies. The presence of ascorbate induced a 50-75% decrease in minimal inhibitory concentrations of different antibiotics for resistant strains. When ascorbate is added, formerly subinhibitory concentrations of penicillin or tetracycline have an increased inhibitory effect on resistant strains and even induced the death of 25-93% of the initial population. These results suggest that ascorbate can induce the loss of several plasmids of S. aureus, and that the levels of antibiotic resistance are also affected by the presence of this compound.

Molecular characterization of the soxRS genes of Escherichia coli: two genes control a superoxide stress regulon.

The soxR locus of Escherichia coli K12 mediates transcriptional activation of a complex oxidative stress regulon in response to superoxide-generating (redox-cycling) agents. We have cloned the soxR locus, which is positioned near the uvrA gene at 92.2 min on the genetic map, by monitoring complementation of a delta soxR mutation. Subclones from the soxR region in the delta soxR strain simultaneously restored cellular resistance to the redox-cycling agent phenazine methosulfate and inducibility of at least two of the regulon proteins, glucose-6-phosphate dehydrogenase and endonuclease IV, by paraquat, another redox-cycling agent. DNA sequence analysis revealed the presence of two genes involved in activating the soxR regulon. These genes, named soxR and soxS, are arranged divergently with their 5' ends separated by only 85 bp. The predicted 12.9-kDa SoxS protein is related to the AraC family of one-component gene regulators, but corresponds only to the putative DNA-binding regions of these proteins. The 17.1-kDa SoxR protein bears significant homology only to the MerR family of proteins including a predicted DNA-binding helix-turn-helix and a cluster of cysteine residues positioned similarly to those that regulate the activity of MerR in response to Hg2+. This suggests that SoxR could be a metal-binding gene regulator that acts as the intracellular sensor for superoxide. SoxS is evidently the proximal activator of the regulon genes: antibiotic resistance and high-level expression of at least three of the regulon proteins was effected in vivo by the individual expression of SoxS, but not of SoxR, whether or not the cells were exposed to paraquat. These data, together with the recently reported paraquat-inducibility of the soxS gene (Wu, I., and Weiss, B. (1990) J. Bacteriol. 173, 2864-2871), indicate that SoxR and SoxS may constitute a novel type of two-component regulatory system in which the two proteins act sequentially to activate transcription of the various regulon genes in response to superoxide stress.

Loss of penicillinase plasmids of Staphylococcus aureus after treatment with L-ascorbic acid.

When 2 clinical strains of plasmid-mediated penicillin-resistant Staphylococcus aureus were treated with 1 mM sodium ascorbate for 6 h, 12-35% colony-forming units (CFU) irreversibly lost their ability to produce beta-lactamase. Agarose gel electrophoresis showed the disappearance of the plasmid bands from the lysates of colonies in which susceptibility to penicillin was induced by ascorbate treatment. Since ascorbic acid is a compound that is completely non-toxic, the possibility of its therapeutic use in the treatment of multiresistant bacterial diseases is proposed.