Susceptibility of Xanthomonas maltophilia to six quinolones and study of outer membrane proteins in resistant mutants selected in vitro

Abstract

The in vitro susceptibilities of 75 clinical isolates of Xanthomonas maltophilia to nalidixic acid, five fluoroquinolones, latamoxef, and doxycycline were determined. Spontaneous mutants were selected, at a frequency of about 10'-to 10-from four strains by culturing the strains in the presence of each quinolone.Selection in the presence of nalidixic acid provided mutants that were either resistant only to that compound or that exhibited cross-resistance to all the fluoroquinolones tested. Cross-resistance was always observed for mutants selected on any of the five fluoroquinolones. It was always associated with chloramphenicol resistance and, frequently, with doxycycline resistance. The electrophoretic alterations of the outer membrane proteins of the mutants suggest that different mechanisms may be involved in quinolone resistance in X. maltophilia.Xanthomonas maltophilia has recently been isolated with increasing frequency and is mainly responsible for nosocomial infections in compromised hosts (17). Its natural resistance restricts the choice of antibiotics for use in the treatment of infected patients (1, 2, 11, 16), and quinolones have been considered as a possible option for therapy (17). The agar diffusion method has revealed a particular phenotype of quinolone susceptibility for this species (12). In the current study, we determined the MICs of nalidixic acid, pefloxacin, ciprofloxacin, ofloxacin, temafloxacin, sparfloxacin, latamoxef, and doxycycline for 75 clinical isolates ofX. maltophilia. Since the emergence of mutants resistant to quinolones and unrelated drugs has been described in several species of gram-negative bacilli (5-7, 14, 18, 20), especially Pseudomonas aeruginosa (8,15), from 4 of the 75 strains of X. maltophilia, we selected mutants that were resistant to six quinolones. The resistance patterns of the mutants were studied, and the electrophoretic profiles of the outer membrane proteins (OMPs) were analyzed for each type of mutant.MICs were determined by a twofold serial agar dilution method on Mueller-Hinton agar by using a replicating spot device, with an inoculum of about 104 CFU per spot.Spontaneous mutants were selected from four susceptible strains by spreading approximately 2 x 108 CFU onto plates of Mueller-Hinton agar containing each of the six quinolones at concentrations of 4, 8, and 16 times the respective MICs for the strains. After 48 h at 37°C, the frequencies of mutation were evaluated and the MICs of the six quinolones, doxycycline, latamoxef, and chloramphenicol were determined. The mutants were classified according to their patterns of resistance to these nine antibiotics. From each type of mutant, OMP fractions were prepared as described previously (6) and were submitted to sodium dodecyl sulfatepolyacrylamide gel electrophoresis for separation (13.5% acrylamide, 0.36% bisacrylamide).

A changing pattern of susceptibility of Xanthomonas maltophilia to antimicrobial agents: implications for therapy

“…The impressive activity exhibited by minocycline against X maltophilia has rarely been described in prior reports (9,14). In contrast, borderline activity of doxycycline (9,12,21) and poor activity of tetracycline (12,24) have been documented previously.…”

“…The increasing resistance of X. maltophilia to the commercially available quinolones is worrisome and requires further investigation (3, 4, 10, 12, 16, 17, 21-23, 30, 33). Although the investigational quinolones show good in vitro activity compared with that of ciprofloxacin and other quinolones (3,13,17,19,21,22,29,30,32,33), their clinical usefulness needs to VOL. 38,1994 on be determined after further in vitro investigation of, for example, their stability to selection of resistant strains.…”

Pulsed-field gel electrophoresis of restriction-digested genomic DNA and antimicrobial susceptibility of Xanthomonas maltophilia strains from Brazil, Switzerland and the USA

“…The incidence of clinical isolation of X. maltophilia is increasing, possibly in part because of the selective pressure from some new antimicrobial agents (Marshall et al, 1989). Strains of X. maltophilia are usually resistant to aminoglycosides and most /Mactams, show variable susceptibility to fluoroquinolones and are generally susceptible to trimethoprim/sulphamethoxazole (Neu, Saha & Chin, 1989;Lecso-Bornet et al, 1992). Low outer membrane permeability and constitutive overproduction of /J-lactamase appears to be responsible for the generally high degree of antibiotic resistance (Mett et al, 1988).…”

Activities of Ciprofloxacin and Moxifloxacin against Stenotrophomonas maltophilia and Emergence of Resistant Mutants in an In Vitro Pharmacokinetic-Pharmacodynamic Model

“…Moreover, acquired resistances to these antibiotics are frequent (2 to 10%, 10 to 29%, and 21 to 53%, respectively) (18). Indeed, mutants exhibiting pleiotropic resistance, including to quinolones, are readily selected in vitro (1,19,22,39,40) as in vivo (19,36,38). In contrast to other gram-negative organisms, the primary determinant of quinolone resistance in S. maltophilia has been identified as the overproduction of efflux pumps, SmeDEF being the most clearly involved (1)(2)(3), rather than mutations in the type II topoisomerases targets (30,35).…”