Amoxicillin

The disk diffusion method on Muller-Hinton agar medium (MH) (Oxoid, UK) was performed to assess the sensitivity of Staphylococcus aureus, Escherichia coli, and Proteus spp. isolates against standard antibiotic disks of 8 mm, including Cefotaxime, Gentamicin, Ciprofloxacin, and Amoxicillin (Himedia, India). The isolates were suspended in sterile water and diluted according to MacFarland microbial suspension, which approximately contained 1×10⁸ CFU/ml. The cultures were incubated at 37°C for 18 hr, according to Kirby-Baur. The zones of inhibition were determined by the National Committee for Clinical Laboratories Standard rules [25 ]. All measurements were executed three times to obtain an average result. Activity Index (AI) and Fold Increase were calculated based on the inhibition zone diameters using Equation 2 and Equation 3 to compare the performance of Co nanoparticles compared to standard antibiotics [19 (link)–24 (link)].
Where Ic is the inhibition zone diameter of Co NPs and Ia is the inhibition zone diameter of antibiotics.

The antimicrobial susceptibility of bacterial isolates was performed by using Kirby Bauer Disk diffusion method on Mueller Hinton Agar (Hi-Media, India) according to the CLSI guidelines [15 ]. The antibiotic susceptibility pattern was examined by using commercial antibiotic discs including Amoxicillin (10μg), Ceftazidime (30μg), Gentamicin (10μg), Tetracycline (30μg), Nitrofurantoin (300μg), Cotrimoxazole (1.25μg), Nalidixic acid (30μg), Cefoxitin (30μg), Azithromycin (15μg), and Ciprofloxacin (5μg) (Hi-Media, India). The Escherichia coli isolate ATCC 25922 and Staphylococcus aureus isolate ATCC 25923 were used as reference organisms for quality control to antimicrobial susceptibility testing.

All of P. multocida and M. haemolytica isolates were tested for their susceptibility to amoxicillin (30 µg), cefotaxime (30 µg), amikacin (30 µg), gentamicin (10 µg), ciprofloxacin (5 µg), enrofloxacin (5 µg), tetracycline (30 µg), and chlortetracycline (30 µg) disks (HiMedia, Mumbai) using disk diffusion method [10 ]. Escherichia coli ATCC 25922 was used as a quality control strain. The interpretation of the results was based on the Clinical and Laboratory Standards Institute breakpoints [11 ].

All Listeria isolates were subjected to antibiotic sensitivity test against 12 different antimicrobial agents by agar diffusion method in Mueller-Hinton AGAR [15 ]. Listeria isolates were tested against amoxicillin (10 mg), cefixime (10 mg), chloramphenicol (25 mg), furazolidone (100 mg), co-trimoxazole (25 mg), oxytetracycline (30 mg), erythromycin (15 mg), chlortetracycline (30 mg), streptomycin (10 mg), and nitrofurazone (100 mg), neomycin (10 mg), doxycycline hydrochloride (30 mg), and antibiotic discs (HiMedia, Mumbai). The clinical breakpoints for Listeria susceptibility testing were defined according to the Clinical and Laboratory Standard Institute [16 ] and the isolates were grouped as sensitive, intermediary sensitive, and resistant against each antibiotic.

Isolates of Campylobacter spp. were tested via the disk diffusion method [37 (link)] using eight (8) commercially available antimicrobials in Bangladesh, viz., amoxicillin (30 μg), ciprofloxacin (5 μg), azithromycin (30 μg), erythromycin (30 μg), tetracycline (30 μg), streptomycin (10 μg), gentamicin (10 μg), and ceftriaxone (30 μg) (HiMedia, Mumbai, India). The zones of inhibition growth were evaluated as the diameter zone as per parameters specified by the Clinical and Laboratory Standard Institute (CLSI) [38 ], thus established as resistant (R), intermediate resistant (I) or susceptible (S) against the antimicrobial agents. In this testing, the E. coli ATCC 25922 strain was used as a quality control organism. All evaluations were validated by conducting at least two replications of the disk diffusion test. Multidrug resistance (MDR) was decided as the resistance to a minimum of three different classes of antimicrobial agents [39 (link)].