Vitek 2 densichek

A sterile microloop was used to take a few colonies of a pure culture cultivated on Luria-Bertani Broth (Biocorp, Warszawa, Poland) for 18 to 24 h. A bacterial suspension was adjusted to McFarland Turbidity Standard of 0.5–0.63 in 3 mL of a 0.45% sodium chloride solution with a VITEK^® 2 DensiChek (bioMérieux, Warszawa, Poland). The GN card was put on the cassette and placed in the instrument. The time interval between suspension preparation and card filling was less than 30 min to avoid changes in turbidity. Cards were incubated at 35.5 ± 1 °C. Each card was removed from the incubator every 15 min and automatically subjected for colorimetric readings. Results were read after 10 h – 18 h of incubation.

Six antibiotic-resistant bacterial strains (Gram-positive and Gram-negative) were used in this investigation to determine the antibacterial activities of the honey: Staphylococcus aureus (ATCC 25923), Streptococcus mutans (1815T), Proteus vulgaris (ATCC 13315), Pseudomonas aeruginosa (ATCC 27853), Klebsiella pneumoniae (ATCC 27736), and Escherichia coli (ATCC 35218). The identification and susceptibility patterns of all the clinical isolates were performed using the VITEK 2 compact system of the Department of Zoonotic Diseases, National Research Centre (Cairo, Egypt), which kindly provided and maintained these bacterial strains. Muller Hinton Agar (MHA) was used to subculture the bacterial strains. Then, the incubation was carried out at 37 °C overnight. A homogeneous suspension was obtained from a single colony of the tested microorganism using a sterile loop and inoculating it in 3 mL of Muller Hinton Broth (Sigma-Aldrich, St. Louis, MO, USA). To provide 0.5 McFarland  =  1 – 2 × CFU/mL, the suspension was standardized using a calibrated VITEK 2 DENSICHEK (BioMérieux, Inc., Marcy l′Etoile, France) [25 (link),26 (link)].