Determination of fluoroquinolones concentration among biofilm-forming and non-forming uropathogenic Acinetobacter Calcoaceticus-Baumannii complex and their correlation to fluoroquinolone-resistant genes
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Abstract
Acinetobacter spp., the predominant cause of urinary tract infection (UTI), is increasingly resistant to fluoroquinolones. Resistance to fluoroquinolones poses a serious health problem in the treatment of urinary tract infections, which are mostly associated with chromosomal mutations encoded by topoisomerase IV. The objective of the study was to detect the blaOXA-23, qnrB, and acc(6l)-lb-cr genes in fluoroquinolone-resistant biofilm-producing ACB complex isolated from UTI-suspected patients.
A hospital-based cross-sectional study was carried out at the Nepal Police Hospital, Kathmandu, from April 2019 to January 2021. A total of 598 urine specimens were processed, and 78 ACB complex were isolated from a total of 366 positive cultures, which were identified based on colony morphology, Gram’s staining reaction, and various biochemical tests. The antimicrobial susceptibility pattern was determined by the modified Kirby-Bauer disk diffusion method, and the result was interpreted according to CLSI guidelines. The minimum inhibitory concentration (MIC) values of levofloxacin and ciprofloxacin for fluoroquinolone-resistant ACB were determined by the broth dilution method. The fluoroquinolone-resistant ACB was selected for blaOXA-23, qnrB, and acc(6l)-lb-cr gene detection by polymerase chain reaction.
ACB complex was isolated more in females 26 (55.31%); outpatients 42 (53.84); and in the age group of 21–40 years 27 (34.61%). The majority of ACB complex isolates were sensitive to Ciprofloxacin 5 (6.41%) and Levofloxacin 10 (12.82%). However, most of them were resistant to Ciprofloxacin 64 (82.05%) and Levofloxacin 60 (76.92%). A total of 78 (21.31%) ACB complex isolates were found to be multidrug resistant. The MICs of Ciprofloxacin and Levofloxacin ranged from 4 to 256 μl/ml. Among 78 (82.05%) fluoroquinolone-resistant ACB isolates, the blaOXA-23 gene was found on 32 (41.02%) isolates, the acc(6l)-Ib-cr gene on 3 (3.84%) isolates, and the qnrB gene on 1 (1.28%) isolates.
The presence of blaOXA-23, acc(6l)-Ib-cr, and qnrB genes and biofilm formation lead to the emergence of fluoroquinolone-resistant strains of the ACB complex. So, the identification of responsible genes is required for AMR surveillance and guidelines for empirical therapy, which aid in preventing the emergence of antibiotic-resistant isolates.
README
Determination of fluoroquinolones concentration among biofilm-forming and non-forming uropathogenic Acinetobacter Calcoaceticus-Baumannii complex and their correlation to fluoroquinolone-resistant genes
Dataset DOI link: https://doi.org/10.5061/dryad.2z34tmptj
Resistance to antimicrobials is rapidly emerging among uropathogens, chiefly attributed to the extensive use and misuse of antimicrobial agents. For example, fluoroquinolone, which was once considered a highly effective broad-spectrum antibiotic to treat infectious diseases, is currently of poor functionality because of the emergence of fluoroquinolone-resistant bacteria, limiting the selection for treating infections. Of the several resistance mechanisms to fluoroquinolones, plasmid-mediated quinolone resistance (PMQR) (blaoxa-23, aac(6')-Ib-cr, qnrB) and biofilm formation—the polymicrobial community of microbial cells enveloped by extracellular polymeric substances—have been identified as the major resistance mechanisms to fluoroquinolones in the Acinetobacter Calcoaceticus-Baumannii complex (ACB complex).
Herein, the dataset is of hospital-visiting patients with urinary tract infections caused by non-lactose-fermenting bacteria, the ACB complex. The dataset comprises a single sheet. The sheet details microbiological findings comprising antimicrobial (drug) resistance patterns, biofilm formers or non-formers, inhibitory concentrations of ciprofloxacin and levofloxacin, and the presence or absence of plasmid-mediated quinolone resistance genes. Both ciprofloxacin and levofloxacin are fluoroquinolone antibiotics, and their inhibitory concentrations refer to the lowest concentration of an antibiotic that will inhibit the visible growth of a ACB complex after overnight incubation at 37°C for 24 hours.
Data were anonymized with the code. Urine samples from which ACB complex strains were isolated were coded with serial alphanumeric numbers (S1, S23, S847, etc.).
The units of the study variables were standard and as follows:
(a) Inhibitory concentrations μg/mL
(b) Optical density (ODtest) AU/cm
(c) Cutoff optical density (ODc) AU/cm
(d) Antimicrobial susceptibility testing mm
Biofilm was quantified by measuring the absorbance (optical density) at 630nm following the solubilization of the attached biofilm in 95% ethanol. The grading of biofilm formation was done as per the chart
Biofilm formation Rule
a) None -
b) Weak ODtest < ODc
c) Moderate ODc < ODtest < 2×ODc
d) Strong 2×ODc < ODtest < 4×ODc
ODtest [(OD 1 + OD 2 + and OD 3) / 3] was determined as the average of ODs. The cut–off optical density (ODc) was set as three standard deviations above the mean OD of the negative control. Negative control was the sterile broth.
Disc diffusion or antimicrobial susceptibility testing was done with the Kirby Bauer Disc Diffusion method. The inhibitory concentrations were determined by the gold standard broth microdilution technique. Biofilm formers were detected by the gold standard microtiter plate method. Molecular characterization of plasmid-mediated quinolone resistance genes was done by conventional polymerase chain reaction.
Except for values of inhibitory concentrations, all other data were qualitative and were calculated as frequency (percentage) i.e., n (%) in SPSS version 17.0. Quantitative variables (i.e., Minimum Inhibitory concentrations 50/ Minimum Inhibitory concentrations 90) were calculated as median (interquartile range). MIC50 and MIC 90 were defined as the minimum concentration at which 50% and 90% of the ACB complex strains were inhibited, respectively. While quantitative variables were analyzed using a t-test for statistical correlations, qualitative variables were tested using a chi-square test at a 95% confidence interval.
ACB complex = Acinetobacter calcoaceticus-baumannii complex, AST = antimicrobial susceptibility testing, MIC = minimum inhibitory concentration, n/a in file = not available, OD 1= first optical density, OD 2= second optical density, OD 3= third optical density, ODc = cut-off optical density, CIP = ciprofloxacin, LF = levofloxacin, AMP = ampicillin, CTX = cefotaxime, GEN = gentamicin, COT = cotrimoxazole, C = chloramphenicol, AMC = amoxicillin clavulanate, CPM = cefepime, IMP = imipenem, CL = colistin
Readers may access the data from the Dryad repository or with a request email to the corresponding author, Ajaya Basnet (xlcprk@gmail.com), of the article.