Accumulation of Norfloxacin by
            <i>Bacteroides fragilis</i>

Ricci, Vito; Piddock, Laura J. V.

doi:10.1128/aac.44.9.2361-2366.2000

Cited by 19 publications

(16 citation statements)

References 28 publications

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“…1B). Similar data were obtained for some fluoroquinolones in Pseudomonas aeruginosa (42) and Bacteroides fragilis (48). Overall, these results suggest that fluoroquinolone uptake is by simple passive diffusion in M. hominis.…”

Section: Resultssupporting

confidence: 76%

“…Thus the absolute amount of drug that can be retained seems larger at 37°C than at 4°C in the absence of an energizing agent, suggesting that the rise in temperature could allow the unmasking of binding sites and that the fluoroquinolone uptake could be influenced by the membrane fluidity. These data have been previously described for several other bacteria (15,42,48). To determine the saturability of accumulation into M. hominis, the effect of exogenous concentrations of CIP and PEF was studied over a concentration range of 0 to 250 g/ml.…”

Section: Resultsmentioning

confidence: 99%

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Evidence of Active Efflux in Resistance to Ciprofloxacin and to Ethidium Bromide by Mycoplasma hominis

Raherison

Gonzalez

Renaudin

et al. 2002

Antimicrob Agents Chemother

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The uptake of fluoroquinolones was characterized for the fluoroquinolone-susceptible strain PG21 of Mycoplasma hominis. Accumulation of fluoroquinolones appeared to occur by passive diffusion. Addition of arginine as the energizer significantly reduced the uptake of fluoroquinolones, suggesting the presence of an energy-dependent efflux process. Reserpine and orthovanadate, two multidrug pump inhibitors, increased significantly the ciprofloxacin (CIP) uptake. In contrast, such a strong effect was not observed for moxifloxacin and pefloxacin uptakes. Two ethidium bromide (EtBr)-resistant strains, selected in vitro, showed a resistance profile compatible with a multidrug-resistant phenotype, with increased MICs for the hydrophilic fluoroquinolones, CIP and norfloxacin, EtBr, and acriflavine. Taking the EtBr-resistant strain RB1La as a model, a significant decrease of the CIP and EtBr uptakes was observed compared to the reference strain PG21. In the presence of reserpine and orthovanadate, both inhibitors of ATP-dependent efflux pumps, the CIP uptake increased significantly, reaching approximately the same level as that of the susceptible strain. Similar results were obtained with EtBr uptake and efflux experiments. Our data suggest the presence of an active efflux system, possibly an ABC-type efflux pump, implicated in the resistance to CIP and unrelated compounds like EtBr in the human mycoplasma M. hominis.Mycoplasma hominis is a cause of urogenital tract infections and has been implicated in extragenital infections as well, especially in immunocompromised patients. This wall-less organism belongs to the class Mollicutes, characterized by a low GϩC content and having arisen from a common ancestor with the gram-positive bacteria, like some clostridia (53). M. hominis is capable of metabolizing arginine as an energy source but does not ferment glucose (44). We recently reported in vitro and in vivo fluoroquinolone-resistant mutants of M. hominis associated with alterations in the topoisomerase II target genes of these antibiotics (9-10).To date, no other mechanism of fluoroquinolone resistance has been identified in M. hominis. However, a second mechanism of resistance to fluoroquinolones, namely active efflux of the drug, might be expected, as already described for various bacteria. Indeed, energy-dependent efflux as a mechanism of quinolone resistance has been found in an increasing number of gram-negative and gram-positive bacteria (for reviews, see references 45 and 46), although the major form of quinolone resistance found in bacteria is altered DNA gyrase and topoisomerase IV (22). Furthermore, the role of an efflux mechanism in fluoroquinolone-resistant clinical isolates has been described for several bacterial species (6,12,16,34,43). This active efflux involves multidrug resistance (MDR) pumps, which transport structurally unrelated compounds, including different classes of antibiotics, antiseptics, and cationic dyes, such as ethidium bromide (EtBr) and acriflavine. These pumps are classified in two grou...

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Section: Resultssupporting

confidence: 76%

Section: Resultsmentioning

confidence: 99%

Evidence of Active Efflux in Resistance to Ciprofloxacin and to Ethidium Bromide by Mycoplasma hominis

Raherison

Gonzalez

Renaudin

et al. 2002

Antimicrob Agents Chemother

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“…Varon et al (15) also suggested that parC and gyrA of S. pneumoniae might be interchangeable initial targets of certain fluoroquinolones, such as ciprofloxacin, sparfloxacin, and moxifloxacin. Due to the overwhelming amount of data supporting the idea of a role of topoisomerase IV (ParC/ParE) in (7,13). More recently Miyamae et al described bexA from B. thetaiotaomicron (8).…”

mentioning

confidence: 99%

Role of Topoisomerase Mutations and Efflux in Fluoroquinolone Resistance of Bacteroides fragilis Clinical Isolates and Laboratory Mutants

Ricci

Peterson

Rotschafer

et al. 2004

Antimicrob Agents Chemother

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Twelve laboratory mutants and 32 ciprofloxacin-resistant isolates of Bacteroides fragilis were examined for the mechanism(s) of fluoroquinolone resistance. Five mutants had mutations in gyrA. One mutant and two clinical isolates contained a mutation in gyrB. Eight mutants and five clinical isolates accumulated significantly less ciprofloxacin than did wild-type isolates; the mutants and clinical isolates were restored to wild-type characteristics when carbonyl cyanide m-chlorophenylhydrazone was used.DNA gyrase is the primary target of fluoroquinolones for most gram-negative bacteria (3,4,6). DNA topoisomerase IV is also a primary target for specific fluoroquinolones in certain gram-positive bacteria (15,16). Studies to explore mechanisms of fluoroquinolone resistance in Bacteroides fragilis have identified mutations in gyrA as well as active efflux of norfloxacin (2, 7, 9, 10). However, analysis of the literature suggests that gyrA is a secondary (not primary) target of fluoroquinolones in B. fragilis (2, 10). The aim of this study was to identify the mechanism(s) of resistance in fluoroquinolone-resistant B. fragilis selected in vitro from agar and a pharmacodynamic (PD) model compared with those of 32 ciprofloxacin-resistant clinical isolates from the United States.The MIC of each antibiotic was determined for each strain with the agar doubling-dilution method described by the British Society for Antimicrobial Chemotherapy (1). Mutants were selected as described previously (5) from B. fragilis NCTC9343/ ATCC25285 (Z14) on Wilkins and Chalgren agar in a single step at 2ϫ MIC of trovafloxacin, moxifloxacin, and ciprofloxacin at mutation frequencies of 2.6 ϫ 10 Ϫ10 , 6.6 ϫ 10 Ϫ9 , and 4.5 ϫ 10 Ϫ9 , respectively. In vitro mutants were also selected in a PD model (11, 12) from Z14, a clinical isolate (Z54), and ATCC 23745 (Z55). From Z14, three mutants (Z50, Z72, and Z73) were selected with trovafloxacin and two mutants (Z69 and Z70) were selected with moxifloxacin. From Z54, one mutant (Z51) was selected with levofloxacin and one mutant (Z52) was selected with trovafloxacin. From Z55, two mutants (Z48 and Z49) were selected with sparfloxacin.Except for garenoxacin and isolate Z72, the MICs of all agents were 2-to 16-fold higher for all mutants than those seen with their parent strain (Table 1). Mutants selected with trovafloxacin on agar (isolate Z56) had a smaller decrease in susceptibility to garenoxacin than mutants selected in the PD model (Z50 and Z72). To study any role of efflux in fluoroquinolone resistance, the efflux inhibitor carbonyl cyanide m-chlorophenylhydrazone (CCCP) was used. For all parent strains and mutants (except Z69 and Z70), CCCP lowered the MIC of ciprofloxacin two-to fourfold. A total of 32 ciprofloxacin-resistant strains isolated in 1999 from a tertiary hospital in the United States were also investigated. Two isolates were inhibited by Ն8 g of ciprofloxacin/ml, 15 were inhibited by Ն16 g of ciprofloxacin/ml, 10 were inhibited by Ն32 g of ciprofloxacin/ml, and 5 were inhibited by Ն64 g...

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“…Transport of material from the cytoplasm across bacterial membranes requires energy in the form of a proton gradient, and disruptors of the proton motive force are often used to examine the role of efflux in resistance to antibiotics (50,55). A proton motive force inhibitor, CCCP, significantly increases T. denticola's susceptibility to killing by h␤D-3, but it had no effect on T. denticola's sensitivity to h␤D-2; similar results were observed for incubation times ranging from 0.5 to 4 h (data not shown).…”

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confidence: 99%

Mechanisms of Decreased Susceptibility to β-Defensins byTreponema denticola

Brissette¹,

Lukehart

2007

Infect Immun

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Treponema denticola, a periodontal pathogen, is relatively resistant to human beta-defensins, which are small cationic antimicrobial peptides produced by a number of cells, including the gingival epithelium. Using two independent methods, we previously demonstrated that T. denticola proteases are not responsible for decreased vulnerability to defensins. In this study, we confirmed that the major outer membrane protease, dentilisin, is not responsible for T. denticola insensitivity to defensins and examined several other possible mechanisms, including reduced binding to the bacterial surface and efflux pump activity. It has been suggested that some bacteria mask their surfaces with serum proteins. T. denticola grown in a serum-free medium did not exhibit increased susceptibility to human beta-defensin 2 and 3 (h␤D-2 and h␤D-3, respectively), suggesting that cloaking of the outer surface with host proteins is not involved in defensin resistance. Nonetheless, we demonstrated that T. denticola binds significantly less h␤D-2 and -3 than susceptible organisms bind, suggesting that the unusual outer membrane composition of T. denticola may discourage cationic peptide binding. Efflux pumps have been shown to mediate resistance to antibiotics and cationic peptides in other bacteria, and their role in T. denticola's relative resistance to ␤-defensins was investigated. Three inhibitors of bacterial ATP-binding cassette (ABC) efflux pumps had no effect on T. denticola's susceptibility to h␤D-2 or -3. In contrast, a proton motive force inhibitor, carbonyl cyanide 3-chlorophenylhydrazone, increased the susceptibility of T. denticola to killing by h␤D-3, demonstrating a potential role for efflux pumps (other than ABC pumps) in resistance to this peptide. Our data suggest that the combination of decreased defensin binding and efflux of any peptide which enters the cytoplasm may explain T. denticola's relative resistance to human beta-defensins.

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Accumulation of Norfloxacin by Bacteroides fragilis

Cited by 19 publications

References 28 publications

Evidence of Active Efflux in Resistance to Ciprofloxacin and to Ethidium Bromide by Mycoplasma hominis

Evidence of Active Efflux in Resistance to Ciprofloxacin and to Ethidium Bromide by Mycoplasma hominis

Role of Topoisomerase Mutations and Efflux in Fluoroquinolone Resistance of Bacteroides fragilis Clinical Isolates and Laboratory Mutants

Mechanisms of Decreased Susceptibility to β-Defensins byTreponema denticola

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