Molecular Characterization of Fluoroquinolone-Resistant Moraxella catarrhalis Variants Generated In Vitro by Stepwise Selection

Abstract

Moraxella catarrhalis causes respiratory infections. In this study, fluoroquinolone-resistant strains were selected in vitro to evaluate the mechanism of fluoroquinolone resistance. Strains with reduced fluoroquinolone susceptibility were obtained by stepwise selection in levofloxacin, and fluoroquinolone targets gyr and par were sequenced. Six novel mutations in GyrA (D84Y, T594dup, and A722dup), GyrB (E479K and D439N), and ParE (Q395R) involved in M. catarrhalis resistance to fluoroquinolones were revealed.KEYWORDS Moraxella catarrhalis, fluoroquinolone, gyr, par, quinolone resistancedetermining region M oraxella catarrhalis is a Gram-negative aerobic diplococcus that causes upper and lower respiratory tract infections (1). M. catarrhalis is known to be resistant to penicillin and first-generation cephalosporins, owing to its ability to produce BRO ␤-lactamase; however, it typically remains susceptible to other antibiotics, including fluoroquinolones (2-4). Fluoroquinolones act primarily by inhibiting DNA gyrase and topoisomerase IV. These two enzymes work together during DNA replication, transcription, recombination, and repair, which are essential for bacterial growth. Both DNA gyrase and topoisomerase IV are large, complex enzymes composed of two pairs of subunits. DNA gyrase comprises the GyrA and GyrB subunits, encoded by the gyrA and gyrB genes, respectively, while topoisomerase IV is composed of ParC and ParE, encoded by the parC and parE genes, respectively (5, 6).In Gram-negative cocci, such as Neisseria gonorrhoeae and N. meningitidis, the mechanism of fluoroquinolone resistance involves amino acid substitutions in the quinolone resistance-determining region (QRDR) of GyrA and ParC and reduced fluoroquinolone accumulation in the cells (7-9). In a recent study, we showed that T80I substitution in GyrA is involved in low-level fluoroquinolone resistance in M. catarrhalis clinical isolates (10). However, the molecular mechanism of high-level fluoroquinolone resistance in M. catarrhalis is unknown. Therefore, we selected fluoroquinolone-resistant M. catarrhalis strains in vitro and identified the mechanism of fluoroquinolone resistance.To obtain fluoroquinolone-resistant M. catarrhalis strains, we performed stepwise selection in brain heart infusion (BHI; BD, Tokyo, Japan) agar plates, as previously described, with modifications (11). The ATCC 49143 reference strain was used as the parental strain (P1) and cultured on BHI agar plates containing levofloxacin at half of the previously determined MIC. The agar plates were then incubated at 35°C for 10 h

Screening and Matching Amphiphilic Cationic Polymers for Efficient Antibiosis

“…As shown in Figure S15, mPB 35 /T 57 did not cause the resistance of E. coli or S. aureus (MIC 5 /MIC 0 < 4). 65 This was consistent with the description in the published literature that cationic amphiphilic polymers are not likely to cause bacterial resistance. 26 Eradication of E. coli Biofilm.…”

“…coli or S. aureus (MIC 5 /MIC 0 < 4) . This was consistent with the description in the published literature that cationic amphiphilic polymers are not likely to cause bacterial resistance …”

The resistance mechanisms of bacteria against ciprofloxacin and new approaches for enhancing the efficacy of this antibiotic

“…FQ targets gyr and par were also sequenced in another work on isolates with decreased FQ resistance that were produced by stepwise selection in levofloxacin. GyrA (D84Y, T594dup, and A722dup), GyrB (E479K and D439N), and ParE (Q395R) were shown to have six new mutations that contribute to M. catarrhalis resistance to FQs ( 131 ). According to a Polish study, M. catarrhalis FQ resistance is linked to amino acid changes in the gyrA and gyrB genes.…”

Alternations in DNA gyrase genes in low-level fluoroquinolone-resistant <em>Moraxella catarrhalis</em> strains isolated in Poland

“…Molecular basis of Gram-negative bacterial resistance to fluoroquinolones has already been described both for cocci and for bacilli such as, among others, Neisseria gonorrhoeae , N. meningitidis , Escherichia coli , Salmonella enterica , Shigella spp., Campylobacter jejuni , Haemophilus influenzae . 20 – 27 The present work describes 4 strains resistant to fluoroquinolones which possessed mutations in gyrA and gyrB genes within their genome. In these strains, the occurrence of mutations was connected to low-level resistance to levofloxacin and moxifloxacin as well as with resistance to nalidixic acid.…”

“…These mutations occurred in gyrA (Asp 84> Tyr, Thr594dup, and Ala722dup), gyrB (Glu 479> Lys and Asp 439> Asn), and parE (Gln 395> Arg), all of which are involved in M. catarrhalis resistance to fluoroquinolones. 27 Iwata et al, 13 in a work also conducted in Japan, analyzed a single clinical M. catarrhalis strain resistant to macrolides and fluoroquinolones (levofloxacin MIC of 4 mg/L). The researchers discovered the presence of mutation C239T (Thr 80> Ile) in gyrA gene and A1481G (Asp 494> Gly) in gyrB gene.…”