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      Nationwide surveillance of antimicrobial susceptibility of 509 rapidly growing mycobacteria strains isolated from clinical specimens in Japan

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          Abstract

          This study aimed to identify effective treatments against rapidly growing mycobacteria (RGM) infections by investigating the minimum inhibitory concentrations (MIC) of 24 antimicrobial agents and their molecular mechanisms of resistance. In total, 509 clinical RGM isolates were identified by analyzing the sequences of three housekeeping genes ( hsp65, rpoB, and sodA), and their susceptibilities to 24 antimicrobial agents were tested. We also performed sequencing analysis of antimicrobial resistance genes ( rrl, rrs, gyrA, and gyrB). To identify Mycobacteroides abscessus group subspecies, we performed PCR-based typing and determined the sequevar of erm(41). We identified 15 RGM species, most of which were susceptible to amikacin and linezolid. Among these species, arbekacin and sitafloxacin had the lowest MIC among the same class of antimicrobials. The MIC of rifabutin for M. abscessus subsp. abscessus (MAB) was lower than that for M. abscessus subsp. massiliense (MMA). The proportion of MAB isolates with MIC ≤ 2 mg/L for rifabutin was significantly higher than that of MMA [MAB: 50/178 (28.1%) vs. MMA: 23/130 (17.7%); p = 0.041]. In summary, our study revealed the antimicrobial susceptibility profile of 15 RGM species isolated in Japan and indicated that arbekacin, sitafloxacin, and rifabutin may be possible therapeutic options for RGM infections.

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          Most cited references31

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          Rapid identification of mycobacteria to the species level by polymerase chain reaction and restriction enzyme analysis.

          A method for the rapid identification of mycobacteria to the species level was developed on the basis of evaluation by the polymerase chain reaction (PCR) of the gene encoding for the 65-kDa protein. The method involves restriction enzyme analysis of PCR products obtained with primers common to all mycobacteria. Using two restriction enzymes, BstEII and HaeIII, medically relevant and other frequent laboratory isolates were differentiated to the species or subspecies level by PCR-restriction enzyme pattern analysis. PCR-restriction enzyme pattern analysis was performed on isolates (n = 330) from solid and fluid culture media, including BACTEC, or from frozen and lyophilized stocks. The procedure does not involve hybridization steps or the use of radioactivity and can be completed within 1 working day.
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            A novel gene, erm(41), confers inducible macrolide resistance to clinical isolates of Mycobacterium abscessus but is absent from Mycobacterium chelonae.

            Mycobacterium abscessus infections tend to respond poorly to macrolide-based chemotherapy, even though the organisms appear to be susceptible to clarithromycin. Circumstantial evidence suggested that at least some M. abscessus isolates might be inducibly resistant to macrolides. Thus, the purpose of this study was to investigate the macrolide phenotype of M. abscessus clinical isolates. Inducible resistance to clarithromycin (MIC > 32 microg/ml) was found for 7 of 10 clinical isolates of M. abscessus previously considered susceptible; the remaining 3 isolates were deemed to be susceptible (MIC
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              rpoB-based identification of nonpigmented and late-pigmenting rapidly growing mycobacteria.

              Nonpigmented and late-pigmenting rapidly growing mycobacteria (RGM) are increasingly isolated in clinical microbiology laboratories. Their accurate identification remains problematic because classification is labor intensive work and because new taxa are not often incorporated into classification databases. Also, 16S rRNA gene sequence analysis underestimates RGM diversity and does not distinguish between all taxa. We determined the complete nucleotide sequence of the rpoB gene, which encodes the bacterial beta subunit of the RNA polymerase, for 20 RGM type strains. After using in-house software which analyzes and graphically represents variability stretches of 60 bp along the nucleotide sequence, our analysis focused on a 723-bp variable region exhibiting 83.9 to 97% interspecies similarity and 0 to 1.7% intraspecific divergence. Primer pair Myco-F-Myco-R was designed as a tool for both PCR amplification and sequencing of this region for molecular identification of RGM. This tool was used for identification of 63 RGM clinical isolates previously identified at the species level on the basis of phenotypic characteristics and by 16S rRNA gene sequence analysis. Of 63 clinical isolates, 59 (94%) exhibited 3% partial rpoB gene sequence divergence from the corresponding type strain; they belonged to three taxa related to M. mucogenicum, Mycobacterium smegmatis, and Mycobacterium porcinum. For M. abscessus and M. mucogenicum, this partial sequence yielded a high genetic heterogeneity within the clinical isolates. We conclude that molecular identification by analysis of the 723-bp rpoB sequence is a rapid and accurate tool for identification of RGM.
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                Author and article information

                Contributors
                kikuchi.ken@twmu.ac.jp
                Journal
                Sci Rep
                Sci Rep
                Scientific Reports
                Nature Publishing Group UK (London )
                2045-2322
                9 June 2021
                9 June 2021
                2021
                : 11
                : 12208
                Affiliations
                [1 ]GRID grid.410818.4, ISNI 0000 0001 0720 6587, Department of Infectious Diseases, , Tokyo Women’s Medical University, ; 8-1 Kawada-Cho, Shinjuku-ku, Tokyo, 162-8666 Japan
                [2 ]GRID grid.39158.36, ISNI 0000 0001 2173 7691, Department of Respiratory Medicine, Faculty of Medicine and Graduate School of Medicine, , Hokkaido University, ; Kita 14-jo Nishi 5-chome, Kita-ku, Sapporo-shi, Hokkaido, 001-0014 Japan
                [3 ]GRID grid.419151.9, ISNI 0000 0001 1545 6914, Department of Mycobacterium Reference and Research, The Research Institute of Tuberculosis, , Japan Anti-Tuberculosis Association, ; Tokyo, 204-0022 Japan
                [4 ]GRID grid.410848.1, BML, Inc, ; Matoba 1361-1, Kawagoe-shi, Saitama, 350-1101 Japan
                Article
                91757
                10.1038/s41598-021-91757-4
                8190260
                34108590
                b2bfd7b1-0a71-4db1-952a-4f00307468d5
                © The Author(s) 2021

                Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/.

                History
                : 23 February 2021
                : 31 May 2021
                Funding
                Funded by: FundRef http://dx.doi.org/10.13039/501100001691, Japan Society for the Promotion of Science;
                Award ID: JP20K22755
                Award ID: JP19K08938
                Award Recipient :
                Categories
                Article
                Custom metadata
                © The Author(s) 2021

                Uncategorized
                antimicrobial resistance,clinical microbiology
                Uncategorized
                antimicrobial resistance, clinical microbiology

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