, 2006; Park et al., 2007; Tamang et al., 2008; Carattoli, 2009; Strahilevitz et al., 2009). The aim of this study was to demonstrate the expression of an inducible acquired pACBL in S. marcescens and Escherichia coli isolates from the same patient. Moreover, as the E. coli isolate

showed reduced susceptibility to quinolones, plasmid-encoded quinolone resistance (PMQR) were also screened. Bacterial isolates were recovered from a urine specimen collected during nephrostomy in a 68-year-old patient who had initially undergone BCG instillation therapy and was later treated surgically by radical EPZ5676 mouse cyst-prostatectomy for a vesicle and ureteral transitional cell carcinoma. This patient carried an ileal conduit. Conjugation assays were performed using the broth mating method at 37 °C. Entinostat price Escherichia coli and S. marcescens isolates suspected to harbour pACBL were used as donor strains. As a recipient strain, we used the E. coli HB101 (UA6190), which expresses a green fluorescent protein marker and is resistant to rifampin, gentamicin

and kanamycin. Briefly, donor and recipient cells from exponentially growing cultures [3 h at 37 °C with agitation in Luria–Bertani (LB) media] were mixed with a donor/recipient ratio of 1 : 1 and incubated overnight at 37 °C. Transconjugants were selected on LB agar supplemented with ceftazidime (10 μg mL−1) and rifampin (100 μg mL−1) and were exposed to UV illumination. Isolates were identified using the API System 20E (bioMérieux, Marcy l’Étoile, France). The disc diffusion susceptibility test was performed on both donor and transconjugant strains, according to Clinical

Laboratory Standards Institute guidelines, using commercially available discs (Neo-Sensitabs, Rosco Diagnostica S/A, Taastrup, Denmark). The antimicrobial agents included were ampicillin, piperacillin, cephalotin, cefuroxime, cefotaxime, ceftazidime, cefepime, aztreonam, imipenem, cefoxitin, amoxicillin–clavulanic acid, piperacillin–tazobactam, nalidixic acid, ciprofloxin, sulphonamides, trimethoprim, trimethoprim–sulphamethoxazole, chloramphenicol, rifampin, tetracycline, gentamicin, kanamycin, tobramycin, amikacin and streptomycin. The inducible AmpC β-lactamase was suspected when antagonism between oxyimino-β-lactams and imipenem or cefoxitin was observed from on primary antibiogram plates. The presence of scattered colonies in the inhibition halo of cefoxitin, cefotaxime, ceftazidime and aztreonam was also examined (Mirelis et al., 2006). Antimicrobial resistance genes present in donor and transconjugant strains were studied. ampC genes were characterized using a previously described multiplex PCR (Pérez-Pérez & Hanson, 2002). Specific primers used to obtain the complete blaDHA-1 gene sequence were: DHA-1A 5′-CTG ATG AAA AAA TCG TTA TC-3′ and DHA-1B 5′-ATT CCA GTG CAC TCC AAA ATA-3′. PCR conditions were one cycle of denaturation at 95 °C for 5 min, followed by 30 cycles at 95 °C for 1 min, annealing at 55 °C for 1 min and elongation at 72 °C for 1 min.