Analysis of species and intra-species associations between the Mycobacterium abscessus complex strains using pulsed-field gel electrophoresis (PFGE) and multi-locus sequence typing (MLST)

Jeon, Se Mi; Lim, Na Ra; Kwon, Seung Jik; Shim, Tae Sun; Park, Mi Sun; Kim, Bum Joon; Kim, Seong Han

doi:10.1016/j.mimet.2014.05.024

Cited by 11 publications

(15 citation statements)

References 29 publications

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“…massiliense), and CIP108541 (M. abscessus subsp. bolletii) served as reference controls (11,14,34,35). H37Rv (M. tuberculosis) was used as a separate species reference control.…”

Section: Methodsmentioning

confidence: 99%

“…bolletii, on the other hand, exhibit inducible resistance (9, 15). As a result, lung diseases caused by different M. abscessus subspecies exhibit contrasting responses to macrolide-based antibiotic therapy (1,8,11,(16)(17)(18)(19). As such, identification of isolates at the subspecies level, combined with antimicrobial susceptibility testing, is very important for clinicians to make proper therapeutic decisions.…”

mentioning

confidence: 99%

“…The most meaningful genomic difference between the three subspecies is that M. abscessus subsp. massiliense possesses an incomplete and inactive erm gene, which is 276 bp in length and harbors deletion mutations at two positions (10)(11)(12)(13). M. abscessus subsp.…”

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confidence: 99%

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Evaluation of Matrix-Assisted Laser Desorption Ionization−Time of Flight Mass Spectrometry for Identification of Mycobacterium abscessus Subspecies According to Whole-Genome Sequencing

Luo

Liu

et al. 2016

J Clin Microbiol

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cThis study was undertaken to evaluate the utility of matrix-assisted laser desorption ionization-time of flight mass spectrometry with the Vitek MS Plus system for identifying Mycobacterium abscessus subspecies in order to facilitate more rapid and appropriate therapy. A total of 175 clinical M. abscessus strains were identified by whole-genome sequencing analysis: 139 Mycobacterium abscessus subsp. abscessus and 36 Mycobacterium abscessus subsp. massiliense. The research-use-only (RUO) Saramis Knowledge Base database v.4.12 was modified accordingly by adding 40 M. abscessus subsp. abscessus and 19 M. abscessus subsp. massiliense reference spectra to construct subspecies SuperSpectra. A blind test, used to validate the remaining 116 isolates, yielded 99.1% (n ‫؍‬ 115) reliability and only 0.9% (n ‫؍‬ 1) error for subspecies identification. Among the two subspecies SuperSpectra, two specific peaks were found for M. abscessus subsp. abscessus and four specific peaks were found for M. abscessus subsp. massiliense. Our study is the first to report differential peaks 3,354.4 m/z and 6,711.1 m/z, which were specific for M. abscessus subsp. massiliense. Our research demonstrates the capacity of the Vitek MS RUO Saramis Knowledge Base database to identify M. abscessus at the subspecies level. Moreover, it validates the potential ease and accuracy with which it can be incorporated into the IVD system for the identification of M. abscessus subspecies. Mycobacterium abscessus is one of the most common pathogens isolated from patients with cystic fibrosis and the second most prevalent, rapidly growing nontuberculous mycobacteria (NTM) species causing NTM pulmonary diseases (1, 2). Although it might not be the most virulent NTM pathogen, inherent antibiotic resistance makes it notoriously difficult to treat (3-7). Among multiple resistance mechanisms, resistance to macrolides is noteworthy (8). In this regard, acquired resistance to clarithromycin is always associated with a mutation in the 23S rRNA gene; intrinsic resistance is conferred mainly by the erythromycin methylase (erm) gene (3). The erm gene plays a significant role in the drug susceptibility and antibiotic resistance of M. abscessus. Mycobacterium abscessus is divided into three subspecies: Mycobacterium abscessus subsp. massiliense, Mycobacterium abscessus subsp. bolletii, and Mycobacterium. abscessus subsp. abscessus (9). The most meaningful genomic difference between the three subspecies is that M. abscessus subsp. massiliense possesses an incomplete and inactive erm gene, which is 276 bp in length and harbors deletion mutations at two positions (10-13). M. abscessus subsp. massiliense can acquire clarithromycin resistance as a consequence of a 23S rRNA mutation at position A2058 (12,14). Due to an inactive erm gene, M. abscessus subsp. massiliense does not exhibit inducible resistance after exposure to macrolides. M. abscessus subsp. abscessus and M. abscessus subsp. bolletii, on the other hand, exhibit inducible resistance (9, 15). As a result, lung diseas...

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“…massiliense), and CIP108541 (M. abscessus subsp. bolletii) served as reference controls (11,14,34,35). H37Rv (M. tuberculosis) was used as a separate species reference control.…”

Section: Methodsmentioning

confidence: 99%

mentioning

confidence: 99%

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Evaluation of Matrix-Assisted Laser Desorption Ionization−Time of Flight Mass Spectrometry for Identification of Mycobacterium abscessus Subspecies According to Whole-Genome Sequencing

Luo

Liu

et al. 2016

J Clin Microbiol

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“…Because M. massiliense differs from M. abscessus in its susceptibility to clarithromycin, distinguishing between the two organisms has clinical importance (6, 7). Furthermore, phylogenetic separation between three types of M. massiliense has also been reported (4,8), highlighting the epidemiological importance of their separation (7,9,10).Peptide nucleic acids (PNAs) are artificially synthesized DNA analogues with an uncharged peptide backbone (11,12). A PNA probe-based real-time PCR assay has been developed for the diagnosis of mycobacterial infections, particularly for distinguishing between Mycobacterium tuberculosis and nontuberculous mycobacteria (NTM) in clinical specimens (13,14 T , and M. xenopi ATCC 19250 T ), were used in this study.…”

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confidence: 90%

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confidence: 92%

Development of a Peptide Nucleic Acid-Based Multiprobe Real-Time PCR Method Targeting the hsp65 Gene for Differentiation among Mycobacterium abscessus Strains

Kim

Shim

et al. 2015

J Clin Microbiol

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c Recently, the need to distinguish between members of the Mycobacterium abscessus group has gained increasing attention. Here, we introduced a novel peptide nucleic acid (PNA) real-time PCR method targeting the hsp65 gene in order to distinguish between four subspecies within the M. abscessus group (M. abscessus and 3 types of M. massiliense).T he taxonomic status of the Mycobacterium abscessus group remains undetermined. The recent development of genetic investigation tools has revealed that the M. abscessus group can be further divided into three closely related taxa, i.e., M. abscessus, M. massiliense and M. bolletii (1-3). Recently, it was reported that M. massiliense can be further subdivided into three genotypes (here referred to as types I, II-1, and II-2) based on analyses of the hsp65 sequence (4, 5). Because M. massiliense differs from M. abscessus in its susceptibility to clarithromycin, distinguishing between the two organisms has clinical importance (6, 7). Furthermore, phylogenetic separation between three types of M. massiliense has also been reported (4,8), highlighting the epidemiological importance of their separation (7,9,10).Peptide nucleic acids (PNAs) are artificially synthesized DNA analogues with an uncharged peptide backbone (11,12). A PNA probe-based real-time PCR assay has been developed for the diagnosis of mycobacterial infections, particularly for distinguishing between Mycobacterium tuberculosis and nontuberculous mycobacteria (NTM) in clinical specimens (13,14 T , and M. xenopi ATCC 19250 T ), were used in this study. All clinical isolates were collected from the Asan Medical Center (Seoul, Republic of Korea) during the period from January 2004 to June 2011. This work was approved by the institutional review board of Seoul National University Hospital (C-1202-057-398) and the Asan Medical Center (2012-0170). The mycobacteria were maintained as a subculture of a frozen stock before use in molecular assays. Bacterial chromosomal DNA was extracted from the clinical isolates by the bead-beater-phenol extraction method, as described previously (15).The primers were designed using the Oligo software (version 6.5; Molecular Biology Insights). These primers produced 377-bp hsp65 amplicons (from nucleotides 380 to 756 in the M. abscessus hsp65 gene). We designed a set of three PNA probes for specific simultaneous detection in a single reaction of the four subspecies of the M. abscessus group (M. abscessus and specific types I, II-1, and II-2) using the LC-PDS (version 2.0) software. The probes were further developed according to the design guidelines of the PNA manufacturer, using three different single nucleotide polymorphisms (SNPs) within 604-bp hsp65 sequences (see Fig. S1 in the supplemental material). We used these reporter dyes to detect different sequences: 6-carboxyfluorescein (FAM) for the detection of M. abscessus and M. massiliense at the subspecies level, Hex for discriminating M. massiliense type I and type II, and Texas Red for discriminating M. massiliense types II-1 and I...

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A novel cluster of Mycobacterium abscessus complex revealed by matrix-assisted laser desorption ionization–time-of-flight mass spectrometry (MALDI-TOF MS)

Suzuki

Yoshida

et al. 2015

Diagnostic Microbiology and Infectious Disease

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Analysis of species and intra-species associations between the Mycobacterium abscessus complex strains using pulsed-field gel electrophoresis (PFGE) and multi-locus sequence typing (MLST)

Cited by 11 publications

References 29 publications

Evaluation of Matrix-Assisted Laser Desorption Ionization−Time of Flight Mass Spectrometry for Identification of Mycobacterium abscessus Subspecies According to Whole-Genome Sequencing

Evaluation of Matrix-Assisted Laser Desorption Ionization−Time of Flight Mass Spectrometry for Identification of Mycobacterium abscessus Subspecies According to Whole-Genome Sequencing

Development of a Peptide Nucleic Acid-Based Multiprobe Real-Time PCR Method Targeting the hsp65 Gene for Differentiation among Mycobacterium abscessus Strains

A novel cluster of Mycobacterium abscessus complex revealed by matrix-assisted laser desorption ionization–time-of-flight mass spectrometry (MALDI-TOF MS)

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