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

Abstract

A worldwide increase in the Mycobacterium abscessus (M. abscessus) complex has been observed. Therefore, the aim of the present study was to investigate the diversity of the rrl and erm(41) genes, both of which are associated with macrolide sensitivity in the M. abscessus complex. The current study also examined the efficacy of mass spectrometry as an alternative to molecular testing to classify subspecies of the M. abscessus complex. A total of 14 strains of the M. abscessus complex were obtained, and based on conventional analyses using housekeeping genes, 57% were determined to be M. abscessus subsp. abscessus, 43% were M. abscessus subsp. massiliense, and none were identified as M. abscessus subsp. bolletii. However, depending on the strain, it was not always possible to distinguish between the subspecies by mass spectrometry. Consequently, PCR products for the rrl and erm(41) genes were directly sequenced. Overall, 7.1% of the strains were identified to have a rrl mutation, and 92.9% carried a T at position 28 of erm (41). Results presented here suggest that the principal cause of treatment failure for M. abscessus complex infections is inducible macrolide resistance encoded by the erm(41) gene. From a strictly pragmatic standpoint, the phenotypic function of a putative erm(41) gene is the most important piece of information required by clinicians in order to prescribe an effective treatment. Although PCR amplification of erm(41) is not sufficient to differentiate between the M. abscessus complex subspecies, PCR can be easily and efficiently used to predict the sensitivity of members of the M. abscessus complex to clarithromycin. PCR amplification of the erm(41) gene can be used to predict the sensitivity of Mycobacterium abscessus complex strains to clarithromycin

The Use of Comparative Genomic Analysis for the Development of Subspecies-Specific PCR Assays for Mycobacterium abscessus

“…Due to the presence of an inducible functional erythromycin resistance methylase ( erm 41) gene that confers macrolide resistance within the subspecies, this gene has also been used as a target in identifying MABC. Nevertheless, this target has also showed discrepancies as only two of the three subspecies ( M. abscessus and M. massiliense ) have been differentiated ( Mase et al., 2019 ). In addition, some M. massiliense strains have been identified as having an intact erm (41) gene instead of the expected deleted gene ( Shallom et al., 2013 ).…”

“…PCR-based tests offer a rapid and relatively simple alternative, which have been able to differentiate between M. abscessus and M. massiliense , but not M. bolletii ( Nakanaga et al., 2014 ; Mase et al., 2019 ). However, there is a limited number of PCR-based methods that have been developed which are able to differentiate between all the three subspecies ( Minias et al., 2020 ; Yoshida et al., 2021 ).…”

A novel DNA chromatography method to discriminate Mycobacterium abscessus subspecies and macrolide susceptibility

“…MABC is the most frequent clinical isolate of rapidly growing mycobacteria and an increased emergence has recently been observed in Japan and other developed countries [42] , [43] , [44] . Commercially available DNA-DNA hybridization kits or MALDI-TOF MS are available in clinical laboratories in Japan and Taiwan to identify mycobacterium isolates [ 45 , 46 ], but they cannot discriminate between either subspecies or the macrolide-susceptible erm (41) truncation and T28C polymorphism [47] . Since macrolide susceptibility is crucial for effective treatment of MABC infection, here we developed a novel multiplex PCR and DNA chromatography method to identify subspecies and macrolide susceptibility.…”

Mycobacterioses Induced by Mycobacterium abscessus: Case Studies Indicating the Importance of Molecular Analysis for the Identification of Antibiotic Resistance

“…The amplification of the erm(41) gene with an incomplete product (product size 397 bp) identified M. a. massiliense. The full length erm(41) gene products (size 673 bp) indicated M. a. abscessus or M. a. bolletii, and further typing was based on the variability of nucleotides in select positions, as previously reported and shown in Table 2 [16,17]. Subspecies identification of M. a. was further confirmed by PCR method based on variable-number, tandem-repeat analysis, as previously described by Choi et al, 2011 [39].…”

“…Constitutive resistance in M. a. occurs rarely and develops on the basis of spontaneous point mutation in the region of the rrl gene with subsequent selection of the strain during therapy with macrolides. Inducible resistance is connected with the presence of the functional erm(41) gene, which codes for the methylase that alters the target point for the macrolides on the ribosome [16,17]. The inducible resistance typically does not manifest by culture method during the standard incubation period in the in vitro tests for the determination of sensitivity to macrolides, but is observed later on after an extension of the incubation period to 14 days [18,19].…”