Selection of Resistance to Clarithromycin in Mycobacterium abscessus Subspecies

Abstract

Mycobacterium abscessus is an emerging pathogen against which clarithromycin is the main drug used. Clinical failures are commonly observed and were first attributed to acquired mutations in rrl encoding 23S rRNA but were then attributed to the intrinsic production of the erm(41) 23S RNA methylase. Since strains of M. abscessus were recently distributed into subspecies and erm(41) sequevars, we investigated acquired clarithromycin resistance mechanisms in mutants selected in vitro from four representative strains. Mutants were sequenced for rrl, erm(41), whiB, rpIV, and rplD and studied for seven antibiotic MICs. For mutants obtained from strain M. abscessus subsp. abscessus erm(41) T28 sequevar and strain M. abscessus subsp. bolletii, which are both known to produce effective methylase, rrl was mutated in only 19% (4/21) and 32.5% (13/40) of mutants, respectively, at position 2058 (A2058C, A2058G) or position 2059 (A2059C, A2059G). No mutations were observed in any of the other genes studied, and resistance to other antibiotics (amikacin, cefoxitin, imipenem, tigecycline, linezolid, and ciprofloxacin) was mainly unchanged. For M. abscessus subsp. abscessus erm(41) C28 sequevar and M. abscessus subsp. massiliense, not producing effective methylase, 100% (26/26) and 97.5% (39/40) of mutants had rrl mutations at position 2058 (A2058C, A2058G, A2058T) or position 2059 (A2059C, A2059G). The remaining M. abscessus subsp. massiliense mutant showed an 18-bp repeat insertion in rpIV, encoding the L22 protein. Our results showed that acquisition of clarithromycin resistance is 100% mediated by structural 50S ribosomal subunit mutations for M. abscessus subsp. abscessus erm(41) C28 and M. abscessus subsp. massiliense, whereas it is less common for M. abscessus subsp. abscessus erm(41) T28 sequevar and M. abscessus subsp. bolletii, where other mechanisms may be responsible for failure.

Clinical Characteristics and Treatment Outcomes of Patients with Acquired Macrolide-Resistant Mycobacterium abscessus Lung Disease

“…The frequencies of mutations at position 2058 or 2059 differ between studies. Some studies showed that the majority of mutational changes involved A2059 (17,29), whereas other studies found that most mutations involved A2058 (18,30). In our study, 23% involved A2059, with the remaining 77% involving A2058, and there was no difference in clarithromycin MICs based on mutation position, although only small numbers of isolates were studied.…”

“…In our study, of the 13 isolates with rrl mutations, 85% had the T28 sequevar and 15% had the C28 sequevar of erm(41). A recent study of mutants selected in vitro for acquired macrolide resistance found that only 19% of mutants derived from the T28 sequevar had rrl mutations, whereas 100% of mutants derived from the C28 sequevar had rrl mutations (29). In clinical isolates, rrl mutations were observed more frequently in isolates of the T28 sequevar than in isolates of the C28 sequevar (18,30).…”

Whole-Genome Sequencing for Predicting Clarithromycin Resistance in Mycobacterium abscessus

“…Mougari and colleagues found that in 39/40 M. abscessus subsp. massiliense isolates selected for clarithromycin resistance, this could be explained by an rrl mutation at positions 2270 and 2271, with a further sample containing an rplV insertion (20). All of our isolates contained this insertion (which was also present in the reference sequence with GenBank accession number NC_010397.1), which was associated with susceptibility to clarithromycin, except in the presence of a relevant rrl mutation.…”

PCR amplification of the erm(41) gene can be used to predict the sensitivity of Mycobacterium�abscessus complex strains to clarithromycin

“…Clarithromycin (41) were detected in this study; however, numerous studies have revealed that both resistance mechanisms can occur concurrently (36,38), and that a functional erm(41) gene does not exclude selection for rrl mutations (44). Transcriptome analysis of M. abscessus complex revealed the presence of several novel open reading frames, which could be activated by stressful conditions, such as hypoxia, to persist (45).…”

“…Conversely, several studies have reported on acquired resistance due to rrl mutations in the absence of any macrolide exposure (16,37). However, an in vitro study reported that rrl mutations at position 2058 or 2059 were observed during incubation with clarithromycin (38). In East Asia, low dose macrolides are often administered for prolonged periods to treat chronic respiratory disorders, such as diffuse panbronchiolitis, to stimulate an immunomodulatory effect (39).…”