Infection and Drug Resistance (Nov 2018)
Novel mechanism responsible for high-level macrolide resistance in Moraxella catarrhalis
Abstract
Ayako Kasai,1 Ayaka Ohta,1 Yuina Maeda,1 Kageto Yamada,2 Kazuyuki Tao,3 Ryoichi Saito1 1Department of Molecular Microbiology, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University, Bunkyo-ku, Tokyo 113-8510, Japan; 2Department of Clinical Laboratory, Toho University Medical Center Omori Hospital, Ota-ku, Tokyo 143-8541, Japan; 3Radioisotope Center, University of Tokyo, Bunkyo-ku, Tokyo 113-0032, Japan Background: High-level macrolide-resistant Moraxella catarrhalis strains have been isolated; however, the underlying mechanism has not been well elucidated. We investigated the role of mutations in the 23S rRNA gene and the L4 and L22 ribosomal proteins using spontaneous erythromycin-resistant mutants and transformants. Materials and methods: The erythromycin-susceptible M. catarrhalis ATCC25238 and clinical isolate Mc19 were used as parental strains. To obtain spontaneous erythromycin-resistant mutants, in vitro stepwise selection was performed using brain–heart infusion agar plates containing various concentrations of erythromycin. The role of the mutations identified in the spontaneous mutants was validated using transformation experiments. Results: We obtained two spontaneous mutants with high-level resistance to erythromycin, S25-32-af10 and S19-256-af10, from ATCC25238 and Mc19, respectively. S25-32-af10 exhibited mutations of Q61R in L4 and Insertion98SRADRIS in L22. S19-256-af10 exhibited three C2611T-mutated alleles in the 23S rRNA gene and G65A in L4. Transformants with single mutations identified in S25-32-af10 or S19-256-af10 showed higher erythromycin and azithromycin minimum inhibitory concentrations (MICs) than those of each parental strain. However, transformants with multiple mutations identified in S25-32-af10 or S19-256-af10 showed macrolide MICs similar to those of each parental strain. Conclusion: Our results provide the first evidence suggesting that Q61R in L4 and Insertion98SRADRIS in L22 are involved in the synergistic acquisition of high-level resistance to both 14- and 15-member macrolides, and that C2611T in the 23S rRNA gene and G65A in L4 also synergistically contribute toward conferring high-level 14-member macrolide resistance to M. catarrhalis. Keywords: macrolide, Moraxella catarrhalis, ribosomal protein, 23S rRNA gene
