Tetracycline

The susceptibility of the confirmed E. coli isolates to thirteen antibiotics was examined according to the Kirby–Bauer disk diffusion method [27 ]. The antibiotics were obtained from Bioanalyse (Ankara, Turkey) and included ampicillin (10 μg), amoxicillin (25 μg), amoxicillin-clavulanic acid (30 μg), cephalexin (30 μg), ceftriaxone (30 μg), gentamicin (10 μg), kanamycin (30 μg), tetracycline (30 μg), chloramphenicol (30 μg), ciprofloxacin (5 μg), levofloxacin (15 μg), trimethoprim-sulphamethoxazole (25 μg), and nitrofurantoin (300 μg). For the test, fresh culture suspensions were standardised to 0.5 McFarland (equivalent to 8 log cfu/mL). Using sterile cotton swabs, sterile prepoured plate count agar (PCA) (Laboratorios CONDA, Madrid, Spain) plates were swabbed with standardised culture suspensions and incubated at 37°C for 1 h. Antibiotic discs were then placed on the agar surface and incubated at 37°C for 24 h. The diameter of the inhibition zone was measured in mm (Figure 1). Isolates were categorized as resistant, intermediate, or susceptible according to the guidelines of the Clinical and Laboratory Standard Institute [27 ]. The multiple antibiotic resistance (MAR) index was computed as a/b, where a is the number of antibiotics the isolate was resistant to and b is the total number of antibiotics to which the isolate was exposed [28 (link)].

β-Lap was purchased from Solarbio Life Science (Beijing, China) and dissolved in dimethyl sulfoxide (DMSO). Gram’s crystal violet, a bacteriological stain, was obtained from TITAN BIOTECH. Phosphate buffered saline (PBS) tablets were obtained from Sigma Aldrich^®, Dorset, UK. Methanol (≥99.8%) was purchased from Sigma-Aldrich^® GmbH Chemie, Steinheim, France. Acetic acid (≥99.8%) was purchased from Sigma-Aldrich^® GmbH Chemie, Steinheim, Germany. Six antibiotics belonging to different classes and potencies were purchased from different pharmaceutical companies. Gentamicin (10 µg) from Becton Dickinson, NJ, USA, vancomycin (30 µg), amikacin (30 µg), tetracycline (30 µg), ciprofloxacin (5 µg) from Bioanalyse^® Ankara, Turkey, and amoxicillin (30 µg) from Oxoid™ Basingstoke, Hampshire, UK.

To determine antibiotic resistance profiles, the disk diffusion method was applied following the methodology described by European Committee on Antimicrobial Susceptibility Testing (EUCAST). Four antibiotics, representing distinct classes and modes of action, were selected (Gentamicin 10 µg, Amoxicillin 25 µg, Chloramphenicol 30 µg, Tetracycline 30 µg (Bioanalyse, Ankara, Turkey). According to EUCAST breakpoints [36 ] zones of inhibition (in mm) were determined.