This was done using the disc diffusion technique on freshly prepared Mueller Hinton Agar. The test antibiotics used in the study were Ciprofloxacin (30 μg), Streptomycin (30 μg), Septrin (15 μg), Gentamycin (15 μg), Amoxicillin (30 μg), Ceftriazone (10 μg), Ciprofloxacin (30 μg), Pefloxacin (10 μg), Gentamycin (30 μg) and Augmentin (10 μg) (Becton Dickinson, USA). The diameters of the zones of inhibition were interpreted as resistance and susceptible according to the NCCLS guideline [38 ].
Ciprofloxacin
Manufactured by BD
Sourced in United States, Germany
Ciprofloxacin is a broad-spectrum antibiotic belonging to the fluoroquinolone class. It is a synthetic antimicrobial agent that inhibits the bacterial DNA gyrase and topoisomerase IV enzymes, which are essential for bacterial cell division and DNA replication.
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Lab products found in correlation
Gentamicin, by BD (42 mentions)
Tetracycline, by BD (30 mentions)
Ampicillin, by BD (27 mentions)
Chloramphenicol, by BD (26 mentions)
Cefotaxime, by BD (23 mentions)
Trimethoprim sulfamethoxazole, by BD (22 mentions)
Ceftazidime, by BD (18 mentions)
Imipenem, by BD (17 mentions)
Nalidixic acid, by BD (17 mentions)
Streptomycin, by BD (16 mentions)
Cefepime, by BD (15 mentions)
Amikacin, by BD (15 mentions)
Meropenem, by BD (14 mentions)
Kanamycin, by BD (14 mentions)
Erythromycin, by BD (13 mentions)
Cefoxitin, by BD (13 mentions)
Tobramycin, by BD (10 mentions)
Clindamycin, by BD (10 mentions)
Fosfomycin, by BD (9 mentions)
Amoxicillin clavulanic acid, by BD (9 mentions)
66 protocols using ciprofloxacin
1
Antibiotic Susceptibility Profiling
2
Antimicrobial Resistance Profiling of Bacteria
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Antimicrobial resistance profiling was performed using the most representative antimicrobial agents from the different antibiotic families, which are of great clinical and epidemiological relevance. Kirby-Bauer disk diffusion or broth microdilution methods (in the case of colistin) were done according to Clinical and Laboratory Standards Institute (CLSI) guidelines (CLSI, 2020 ). Escherichia coli ATCC 25922 was used as a control strain. The antimicrobials tested by disk diffusion were: amoxicillin/clavulanate (20/10 μg), ampicillin (10 μg), aztreonam (30 μg), cefepime (30 μg), cefotaxime (30 μg), ceftazidime (30 μg), chloramphenicol (30 μg), ciprofloxacin (5 μg), fosfomycin (200 μg/50 μg of glucose-6-phosphate), gentamicin (10 μg), imipenem (10 μg), meropenem (10 μg), and trimethoprim-sulfamethoxazole (1.25/23.75 μg) (Becton Dickinson). Extended-spectrum beta-lactamases were screened by the ESBL test following the CLSI guidelines (CLSI, 2020 ). Isolates were classified as susceptible, resistant to 1 or 2 antimicrobial categories, multidrug-resistant (MDR) if resistant to at least one agent in ≥ 3 antimicrobial categories; or extensively drug-resistant (XDR), if resistant to at least one agent in all but two or fewer antimicrobial categories (Magiorakos et al., 2012 (link)).
3
Antibiotic Susceptibility Profiling of Bacterial Isolates
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For identification, colonies from different agar media were subjected to MALDI-TOF (VITEK MS – bioMerieux), and N196 automated biochemical testing was performed using the VITEK 2 system (bioMerieux).
Susceptibility testing was performed as recommended by the European Committee on Antimicrobial Susceptibility testing (EUCAST)16 . Interpretation of zone diameters was performed according to EUCAST 2017.
The following antibiotics were used: ampicillin (10 μg), amoxicillin/clavulanic acid (20 μg/10 μg), piperacillin/tazobactam (100 μg/10 μg), cefalexin (30 μg), cefuroxime (30 μg), cefoxitin (30 μg), cefotaxime (5 μg), ceftazidime (10 μg), cefepime (30 μg), imipenem (10 μg), meropenem (10 μg), amikacine (30 μg), gentamicin (10 μg), trimethoprim/sulfamethoxazole (1.25 μg/23.75 μg), ciprofloxacin (5 μg), and moxifloxacin (5 μg) (Becton Dickinson and Company, Sparks, MD, USA, BD BBL™). Sensi-DiscTM paper discs (BD) were used.
Susceptibility testing was performed as recommended by the European Committee on Antimicrobial Susceptibility testing (EUCAST)16 . Interpretation of zone diameters was performed according to EUCAST 2017.
The following antibiotics were used: ampicillin (10 μg), amoxicillin/clavulanic acid (20 μg/10 μg), piperacillin/tazobactam (100 μg/10 μg), cefalexin (30 μg), cefuroxime (30 μg), cefoxitin (30 μg), cefotaxime (5 μg), ceftazidime (10 μg), cefepime (30 μg), imipenem (10 μg), meropenem (10 μg), amikacine (30 μg), gentamicin (10 μg), trimethoprim/sulfamethoxazole (1.25 μg/23.75 μg), ciprofloxacin (5 μg), and moxifloxacin (5 μg) (Becton Dickinson and Company, Sparks, MD, USA, BD BBL™). Sensi-DiscTM paper discs (BD) were used.
4
Antimicrobial Susceptibility Testing
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Bacterial cell suspensions in saline were normalized at 0.5 McFarland standard and swabbed onto MH agar plates. Disks containing ciprofloxacin (5 μg), imipenem (10 μg), gentamicin (10 μg), novobiocin (30 μg), erythromycin (15 μg), rifampicin (5 μg), or tobramycin (10 μg) (Becton Dickinson) were placed on the surface of the inoculated plates, and growth inhibition halo diameters were measured after 24-h incubation at 37°C.
5
Antimicrobial Susceptibility Testing Protocol
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Antimicrobial susceptibility testing was performed using the disk-diffusion method and the antibiotics ampicillin (AM), amoxicillin-clavulanic acid (AMC), cefotaxime (CTX), nalidixic acid (Na), ciprofloxacin (CIP), gentamicin (GM), kanamycin (K), streptomycin (S), sulfamethoxazole (Su), trimethoprim (TMP), tetracycline (T), and chloramphenicol (C) (Becton Dickinson, Heidelberg, Germany). Results were interpreted according to Clinical and Laboratory Standards Institute (CLSI) performance standards (CLSI, 2016 ). For sulfamethoxazole, for which breakpoints are not listed separately from trimethoprim, an inhibition zone of ≤10 mm was interpreted as resistant. Isolates displaying resistance to three or more classes of antimicrobials (counting β-lactams as one class) were defined as multidrug-resistant (MDR). Synergistic effects between AMC and CTX were regarded as an indication of the presence of an ESBL producer (Kaur et al., 2013 (link)).
6
Antibiotic Sensitivity Profiling of Bacterial Strains
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Sensitivity to many different antibiotics was assessed by the Kirby– Bauer disc diffusion test. Bacterial cell suspensions in saline were normalized at 0.5 McFarland Standard and swabbed onto MH agar plates, using disks containing ciprofloxacin (5 µg), novobiocin (30 µg), rifampicin (5 µg), erythromycin (15 µg), streptomycin (10 µg), tobramycin (10 µg), imipenem (10 µg), and colistin (10 µg) (Becton Dickinson). Growth inhibition halos were measured after 24 h of growth at 22, 30, or 37 °C.
colistin sensitivity was also assessed with minimum inhibitory concentration (MIC) assay using the broth microdilution method. Briefly, strains were cultured in MH at 37 °C for 8 h, and then refreshed at 5 × 105 cells/ml in the same medium in the presence of increasing concentrations of colistin (up to 16 μg/mL). MIC was defined as the lowest concentration of antibiotics for which no visible growth was observed after 24 h at 37 °C. Each strain was tested in at least three independent experiments.
colistin sensitivity was also assessed with minimum inhibitory concentration (MIC) assay using the broth microdilution method. Briefly, strains were cultured in MH at 37 °C for 8 h, and then refreshed at 5 × 105 cells/ml in the same medium in the presence of increasing concentrations of colistin (up to 16 μg/mL). MIC was defined as the lowest concentration of antibiotics for which no visible growth was observed after 24 h at 37 °C. Each strain was tested in at least three independent experiments.
7
Antibacterial Activity of Plant Extracts
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Standard strains of bacteria — Escherichia coli (ATCC 25922), Pseudomonas aeruginosa (ATCC 27853), Staphylococcus aureus (ATCC 25923), Shigella flexneri (ATCC 12022), and Salmonella enterica serovar Typhimurium (S. Typhimurium; ATCC 14028) — were obtained from the Aklilu Lemma Institute of Pathobiology (ALIPB), Addis Ababa University and the Ethiopian Public Health Institute. The bacteria were selected based on availability and by considering the traditional use of the experimental plants. Standard antibacterial disks used for the study were amoxicillin (30 μg/disk, Oxoid) and ciprofloxacin (5 μg/disk, Becton Dickinson).
8
Antibiotic Susceptibility Testing Protocol
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Susceptibility testing was performed by Kirby Bauer disk diffusion method using 22 antibiotics: amikacin, ampicillin, cefpodoxime, ceftazidime, ceftriaxone, cefuroxime, ciprofloxacin, ertapenem, imipenem, nalidixic acid, nitrofurantoin, tetracycline, trimethoprim/sulfamethoxazole (Becton Dickinson, NJ), ampicillin-sulbactam, aztreonam, cefazoline, cefepime, cefotaxime, ceftiofur, chloramphenicol, kanamycin, and streptomycin (Oxoid), according to the Clinical and Laboratory Standards Institute, CLSI.18 ESCs and ciprofloxacin-resistant isolates were selected and cryopreserved for further analysis. Multiresistance was defined as the resistance to three or more structural classes of antibiotics19 (link) and M100 from CLSI (30th edition, 2020) was used for interpretation and antimicrobial class classification (Glossary I).20 Isolates showing an intermediate resistant pattern were included in the resistant group for the analysis of antibiotic susceptibility but excluded from the minimal inhibitory concentration (MIC) assays. MICs were performed manually using the microdilution method on isolates that showed resistance to cefotaxime, ciprofloxacin, and nalidixic acid as recommended by CLSI.21 Concentrations from 0.125 (μg/mL) to 256 (μg/mL) were evaluated for each antibiotic.
9
Antibiotic Susceptibility of A. baumannii
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Random strains of A. baumannii (11 from TVGH, 5 from HBTZH, and 2 from ATCC) were selected to check their antibiotic susceptibility using an agar disk diffusion test against several antibiotics, including ciprofloxacin (5 µg), levofloxacin (5 µg), imipenem (10 µg), piperacillin-tazobactam (110 µg), and ampicillin (10 µg) (Becton-Dickinson, Sparks, MD, USA). The diameter of the inhibition zone was measured and classified according to the Clinical and Laboratory Standard Institute [21 ].
10
Antibiotic Susceptibility Profiling
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Antibiotic susceptibility testing of ampicillin, ciprofloxacin and high-level gentamicin (Becton Dickinson, Franklin Lakes, NJ, USA) was performed by a disc diffusion method. Minimum inhibitory concentrations (MICs) of vancomycin, teicoplanin, linezolid, daptomycin and tigecycline were evaluated with E-test strips (AB Biodisk, Solna, Sweden). The results were interpreted according to the recommendations by the Clinical and Laboratory Standards Institute [20] . The susceptibility for tigecycline was defined based on the European Committee on Antimicrobial Susceptibility Testing criteria (MIC≤0.25 µg/mL) [21] . Staphylococcus aureus ATCC29213 and Enterococcus faecalis ATCC29212 were used as control strains.
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