[Resistance to quinolones and beta-lactams in Salmonella enterica due to mutations in topoisomerase-encoding genes, altered cell permeability and expression of an active efflux system].
BACKGROUND
The mechanisms of resistance to fluoroquinolones and beta-lactams were studied in isolates of Salmonella enterica resistant to both antimicrobial groups, isolated over time from two patients treated with fluoroquinolones.


METHODS
The clonal relationships among the various strains was established by serotyping and pulsed-field gel electrophoresis. MICs for beta-lactams, quinolones, chloramphenicol and tetracycline were determined. Presence of beta-lactamases was ruled out by a colorimetric assay. Quinolone resistance-determining regions of the gyrA, gyrB, parC, and parE genes were sequenced, and the relevance of the mutations in these regions was evaluated by complementation assays. Outer membrane protein profiles, the effect of phenylalanyl-arginyl-naphthylamide (PAN, 20 mg/l) on the MICs of several quinolones, and norfloxacin accumulation in the absence and in the presence of a metabolic inhibitor were also determined.


RESULTS
The following mutations were found: gyrA (Asp87 --> Gly; Ser83 --> Phe; Asp87 --> Lys), gyrB (Ser463 --> Phe) and parC (Glu84 --> Gly). Altered outer membrane protein profiles, including decreased expression of a porin equivalent to OmpF from Escherichia coli was observed. Active efflux of norfloxacin was proved in both a clinical isolate and a mutant obtained in vitro. In the presence of PAN, nalidixic acid MICs decreased 4-32 times (except in one strain), pefloxacin MICs decreased 4-16 times for 5 out of 9 evaluated strains, and MICs of both norfloxacin and ciprofloxacin did not change or changed within a single dilution step.


CONCLUSIONS
Quinolone-resistance is the consequence of a combination of mutations in topoisomerase-encoding genes, altered permeability and active efflux. Altered permeability and active efflux would also contribute to decreased susceptibility to beta-lactams.
Miró
Elisenda
E
Servicio de Microbiología, Hospital de la Santa Creu i Sant Pau, Barcelona, Spain. emiro@hsp.santpau.es
Vergés
Clara
C
García
Isabel
I
Mirelis
Beatriz
B
Navarro
Ferrán
F
Coll
Pere
P
Prats
Guillermo
G
Martínez-Martínez
Luis
L
D000328:Adult; D000368:Aged; D019943:Amino Acid Substitution; D000890:Anti-Infective Agents / Q000493:pharmacokinetics / Q000494:pharmacology* / Q000627:therapeutic use; D001425:Bacterial Outer Membrane Proteins / Q000235:genetics* / Q000502:physiology; D001426:Bacterial Proteins / Q000235:genetics* / Q000502:physiology; D001693:Biological Transport, Active / Q000235:genetics; D002463:Cell Membrane Permeability; D027081:DNA Gyrase / Q000235:genetics* / Q000502:physiology; D004252:DNA Mutational Analysis; D027101:DNA Topoisomerase IV / Q000235:genetics* / Q000502:physiology; D004269:DNA, Bacterial / Q000235:genetics; D004151:Dipeptides / Q000494:pharmacology; D024901:Drug Resistance, Multiple, Bacterial / Q000235:genetics*; D005759:Gastroenteritis / Q000188:drug therapy / Q000382:microbiology*; D006801:Humans; D008297:Male; D008826:Microbial Sensitivity Tests; D008875:Middle Aged; D009154:Mutation; D020125:Mutation, Missense; D018272:Porins / Q000235:genetics / Q000502:physiology; D015363:Quinolones / Q000493:pharmacokinetics / Q000494:pharmacology* / Q000627:therapeutic use; D012480:Salmonella Infections / Q000188:drug therapy / Q000382:microbiology*; D019779:Salmonella enterica / Q000187:drug effects / Q000235:genetics* / Q000302:isolation & purification; D013997:Time Factors; D018440:beta-Lactam Resistance / Q000235:genetics
D000890:Anti-Infective Agents; D001425:Bacterial Outer Membrane Proteins; D001426:Bacterial Proteins; D004269:DNA, Bacterial; D004151:Dipeptides; D018272:Porins; D015363:Quinolones; C474925:phenylalanylarginine-naphthylamide; D027101:DNA Topoisomerase IV; D027081:DNA Gyrase