Prevalence and molecular analysis of macrolide-resistant Moraxella catarrhalis clinical isolates in Japan, following emergence of the highly macrolide-resistant strain NSH1 in 2011

Although Moraxella catarrhalis is known to be susceptible to macrolides, highly macrolide-resistant M. catarrhalis isolates have recently been reported in Japan and China. In this study, we investigated the prevalence of macrolide-resistant M. catarrhalis isolates in Tokyo and Chiba, Japan, and studied the mechanisms underlying their resistance. Specifically, we determined the susceptibility of 593 clinical isolates (collected between December 2011 and May 2014) to erythromycin, using the disk diffusion method. For isolates with erythromycin resistance, we identified the MICs of seven antimicrobial agents, including macrolides, and used PFGE to analyse the clonal spread. We also performed sequencing analysis to investigate macrolide-resistance targets. Thirteen isolates (2.2 %) were found to be resistant to erythromycin, showing a high MIC90 to erythromycin, clarithromycin, clindamycin and azithromycin. However, those isolates, in addition to 156 randomly selected erythromycin-susceptible strains, were susceptible to amoxicillin-clavulanate, cefixime and levofloxacin. The 13 highly macrolide-resistant isolates were classified into 10 clades and harboured three or four A2058T-mutated 23S rRNA alleles. Three highly macrolide-resistant isolates also exhibited mutations in ribosomal proteins L4 (V27A and R161C) and L22 (K68T). To the best of our knowledge, we have demonstrated for the first time that, whilst the prevalence of macrolide-resistant M. catarrhalis isolates is low in clinical settings in Japan, genetically diverse isolates with high-level macrolide resistance due to the acquisition of an A2058T mutation in the 23S rRNA have already spread. Our study therefore lays the basis for epidemiological studies of macrolide-resistant M. catarrhalis clinical isolates.