Cefuroxime

Antimicrobial susceptibility testing of E. coli isolates were performed by the Kirby-Bauer disk diffusion assay on Mueller-Hinton agar medium (Oxoid, Basingstoke, England) as recommended by Clinical Laboratory Standard Institute (CLSI, 2011), against 15 antimicrobial agents from different categories including: amikacin (30 μg), amoxicillin (10 μg), amoxicillin-clavulanic acid (30 μg), ceftazidime (30 μg), ceftriaxone (30 μg) cefuroxime (30 μg), chloramphenicol (30 μg), ciprofloxacin (5 μg), gentamicin (10 μg), nalidixic acid (30 μg), nitrofurantoin (50 μg), ofloxacin (5 μg), tetracycline (30 μg), tobramicin (10 μg) and trimethoprim- sulfamethoxazole (25 μg) (Oxoid, Basingstoke, England). E. coli isolate was considered non-susceptible to an antimicrobial agent when it tested resistant, intermediate or non-susceptible when using clinical breakpoints as interpretive criteria, provided by the CLSI, (2011). MDR patterns of E. coli isolates were defined as non-susceptibility to at least one agent in three or more antimicrobial categories (Magiorakos et al., 2012 ).

Pure colonies from each sample with E. coli and P. aeruginosa contamination were randomly selected for antibiotic susceptibility testing. Pure isolates of E. coli, and P. aeruginosa were subjected to antibiotic susceptibility testing using the Kirby–Bauer Disc Diffusion method on Mueller–Hinton agar, as recommended by Clinical Laboratory Standards Institute (CLSI) guidelines [23 ]. Zones of inhibition were measured in millimeters and recorded for each antibiotic.
Antibiotics used for E. coli isolates included amoxicillin–clavulanate (20/10 µg), aztreonam (30 µg), ertapenem (10 µg), gentamicin (10 µg), chloramphenicol (20/10 µg), ciprofloxacin (5 µg), cefuroxime (30 µg), ceftriaxone (30 µg) and trimethoprim–sulphamethoxazole (1.25/23.75 µg) (Oxoid, Hampshire, UK). For P. aeruginosa, piperacillin–tazobactam (100/10µg), aztreonam (30 µg), gentamicin (10 µg) and ciprofloxacin (5 µg) (Oxoid, Hampshire, UK) were used.

Reagents for gold nanoparticle synthesis, including cetrimonium bromide (CTAB) and gold salts were ordered from Sigma-Aldrich (Saint Louis, USA). Colorimetric and fluorescent probes: 3-(4,5-Dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT), resazurin sodium salt, 2ʹ,7ʹ-dichlorofluorescin diacetate (DCFH-DA), N-phenyl-1-naphthylamine (NPN), 2-nitrophenyl β-d-galactopyranoside (ONPG), and propidium iodide (PI) were purchased from Sigma-Aldrich (Saint Louis, USA). Conventional antibiotics (amoxicillin/clavuronic acid, ampicillin, cefuroxime, ceftazidime, gentamicin, amikacin, tobramycin, ciprofloxacin, trimethoprim/sulfamethoxazole, nitrofurantoin, and doxycycline) were purchased from OXOID (Hampshire, United Kingdom) and Polfa Tarchomin (Warsaw, Poland).

Antimicrobial susceptibility tests of the three ESBL-producing isolates and seven non-ESBL-producing isolates were evaluated using the disk diffusion method to Piperacillin 100 μg (PIP), Moxalactam 30 μg (MOX), Ceftazidime 30 μg (CAZ), Cefixime 5 μg (CFM), Cefepime 30 μg (FEP), Cefotaxime 30 μg (CTX), Cephalexin 30 μg (CL), Caphazolin 30 μg (CZ), Ceftriaxone 30 μg (CRO), Cefoxitin 30 μg (FOX), Piperacillin/Tazobactam 100/10 μg (TZP), Cefuroxime 30 μg (CXM), Cefaclor 30 μg (CEC), Ampicillin/Sulbactam 10/10 μg (SAM), Cefoperazone 75 μg (CFP), Ceftizoxime 30 μg (ZOX), Aztreonam 30 μg (ATM), Meropenem 10 μg (MEM), Imipenem 10 μg (IPM), Kanamycin 30 μg (K), Streptomycin 10 μg (S), Ofloxacin 5 μg (OFX), Norfloxacin 10 μg (NOR), CiprOfloxacin 5 μg (CIP); Gatifloxacin 5 μg (GTX), Chloramphenicol 30 μg (C), Azithromycin 15 μg (AZM), Doxycycline 30 μg (TE), Minocycline 30 μg (MH), Compound Sulfamethoxazole 23.75/1.25 μg (SMZ), and Trimethoprim 5 μg (TMP) (Oxoid, Basingstoke, United Kingdom). The results were interpreted according to the Clinical and Laboratory Standards Institute (CLSI) breakpoints. Escherichia coli ATCC 25,922 was used as a control for antimicrobial susceptibility testing.

Susceptibility to antimicrobials was determined by Kirby–Bauer disk diffusion method on Mueller Hinton agar (MHA). The antibiotics used in the study included imipenem (IMP), meropenem (MEM), piperacillin (PRL), ampicillin/sulbactam (SAM), amoxicillin/clavulanic acid (AMC), cefepime (FEP), cefuroxime (CXM), cephalothin (KF), ceftriaxone (CRO), aztreonam (ATM), kanamycin (K), streptomycin (S), gentamicin (CN), nalidixic acid (NA), levofloxacin (LEV), norfloxacin (NOR), ciprofloxacin (CIP), trimethoprim/sulfamethoxazole (SXT), tetracycline (TE), furadantin (F), and chloramphenicol (C) (Oxoid, United Kingdom). The inoculated plates were incubated for 24 h aerobically at 37°C. The diameters of the zones of inhibition was interpreted according to the Clinical Laboratory Standards Institute (CLSI) guidelines (CLSI, 2016 ).
The minimum inhibitory concentration (MIC) of colistin was determined by broth microdilution method recommended by the joint CLSI-EUCAST Polymyxin Breakpoints Working Group¹.