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Proceedings of the National Academy of... Oct 2018is a peculiar rapid-growing (RGM) capable of surviving within eukaryotic cells thanks to an arsenal of virulence genes also found in slow-growing mycobacteria (SGM),...
is a peculiar rapid-growing (RGM) capable of surviving within eukaryotic cells thanks to an arsenal of virulence genes also found in slow-growing mycobacteria (SGM), such as A screen based on the intracellular survival in amoebae and macrophages (MΦ) of an transposon mutant library revealed the important role of MAB_0855, a yet uncharacterized Mycobacterial membrane protein Large (MmpL). Large-scale comparisons with SGM and RGM genomes uncovered MmpL12 proteins as putative orthologs of MAB_0855 and a locus-scale synteny between the and loci. A KO mutant of the gene, designated herein as , had impaired adhesion to MΦ and displayed a decreased intracellular viability. Despite retaining the ability to block phagosomal acidification, like the WT strain, the mutant was delayed in damaging the phagosomal membrane and in making contact with the cytosol. Virulence attenuation of the mutant was confirmed in vivo by impaired zebrafish killing and a diminished propensity to induce granuloma formation. The previously shown role of MmpL in lipid transport prompted us to investigate the potential lipid substrates of MmpL8 Systematic lipid analysis revealed that MmpL8 was required for the proper expression of a glycolipid entity, a glycosyl diacylated nonadecyl diol (GDND) alcohol comprising different combinations of oleic and stearic acids. This study shows the importance of MmpL8 in modifying interactions between the bacteria and phagocytic cells and in the production of a previously unknown glycolipid family.
Topics: Amoeba; Animals; Bacterial Proteins; Biological Transport; Cell Line; Cytosol; Glycolipids; Humans; Lipids; Macrophages; Membrane Proteins; Mice; Mycobacterium abscessus; Phagosomes; Virulence; Virulence Factors; Zebrafish
PubMed: 30301802
DOI: 10.1073/pnas.1812984115 -
Antimicrobial Agents and Chemotherapy Dec 2017Lung disease caused by is increasing, and current clarithromycin-based treatment regimens are only moderately effective. Here, we determined the effect of...
Lung disease caused by is increasing, and current clarithromycin-based treatment regimens are only moderately effective. Here, we determined the effect of clarithromycin-vancomycin combination against complex isolates Synergy was found with a fractional inhibitory concentration index (FICI) score of ≤0.5 and a 4- to 10-fold decrease in MIC.
Topics: Antibiotics, Antitubercular; Clarithromycin; Drug Synergism; Drug Therapy, Combination; Humans; Lung Diseases; Microbial Sensitivity Tests; Mycobacterium Infections, Nontuberculous; Mycobacterium abscessus; Treatment Outcome; Vancomycin
PubMed: 28923867
DOI: 10.1128/AAC.01298-17 -
Antimicrobial Agents and Chemotherapy Jan 2022Mycobacterium abscessus is an opportunistic pathogen notorious for its resistance to most classes of antibiotics and low cure rates. M. abscessus carries an array of...
Mycobacterium abscessus is an opportunistic pathogen notorious for its resistance to most classes of antibiotics and low cure rates. M. abscessus carries an array of mostly unexplored defense mechanisms. A deeper understanding of antibiotic resistance and tolerance mechanisms is pivotal in development of targeted therapeutic regimens. We provide the first description of all major transcriptional mechanisms of tolerance to all antibiotics recommended in current guidelines, using RNA sequencing-guided experiments. M. abscessus ATCC 19977 bacteria were subjected to subinhibitory concentrations of clarithromycin (CLR), amikacin (AMK), tigecycline (TIG), cefoxitin (FOX), and clofazimine (CFZ) for 4 and 24 h, followed by RNA sequencing. To confirm key mechanisms of tolerance suggested by transcriptomic responses, we performed time-kill kinetic analysis using bacteria after preexposure to CLR, AMK, or TIG for 24 h and constructed isogenic knockout and knockdown strains. To assess strain specificity, pan-genome analysis of 35 strains from all three subspecies was performed. Mycobacterium abscessus shows both drug-specific and common transcriptomic responses to antibiotic exposure. Ribosome-targeting antibiotics CLR, AMK, and TIG elicit a common response characterized by upregulation of ribosome structural genes, the regulon and transferases, accompanied by downregulation of respiration through Exposure to any of these drugs decreases susceptibility to ribosome-targeting drugs from multiple classes. The cytochrome bd-type quinol oxidase contributes to CFZ tolerance in M. abscessus, and the sigma factor but not antisigma factor MAB_3542c is involved in TIG resistance. The observed transcriptomic responses are not strain-specific, as all genes involved in tolerance, except , are found in all included strains.
Topics: Anti-Bacterial Agents; Clarithromycin; Humans; Kinetics; Microbial Sensitivity Tests; Mycobacterium Infections, Nontuberculous; Mycobacterium abscessus; RNA; Sequence Analysis, RNA
PubMed: 34633851
DOI: 10.1128/AAC.01509-21 -
Nature Microbiology Sep 2022The medical and scientific response to emerging and established pathogens is often severely hampered by ignorance of the genetic determinants of virulence, drug...
The medical and scientific response to emerging and established pathogens is often severely hampered by ignorance of the genetic determinants of virulence, drug resistance and clinical outcomes that could be used to identify therapeutic drug targets and forecast patient trajectories. Taking the newly emergent multidrug-resistant bacteria Mycobacterium abscessus as an example, we show that combining high-dimensional phenotyping with whole-genome sequencing in a phenogenomic analysis can rapidly reveal actionable systems-level insights into bacterial pathobiology. Through phenotyping of 331 clinical isolates, we discovered three distinct clusters of isolates, each with different virulence traits and associated with a different clinical outcome. We combined genome-wide association studies with proteome-wide computational structural modelling to define likely causal variants, and employed direct coupling analysis to identify co-evolving, and therefore potentially epistatic, gene networks. We then used in vivo CRISPR-based silencing to validate our findings and discover clinically relevant M. abscessus virulence factors including a secretion system, thus illustrating how phenogenomics can reveal critical pathways within emerging pathogenic bacteria.
Topics: Genome, Bacterial; Genome-Wide Association Study; Humans; Mycobacterium Infections, Nontuberculous; Mycobacterium abscessus; Virulence Factors
PubMed: 36008617
DOI: 10.1038/s41564-022-01204-x -
Journal of Clinical Microbiology Jun 2020Members of the complex (MABC) are multidrug-resistant nontuberculous mycobacteria and cause opportunistic pulmonary infections in individuals with cystic fibrosis (CF)....
Members of the complex (MABC) are multidrug-resistant nontuberculous mycobacteria and cause opportunistic pulmonary infections in individuals with cystic fibrosis (CF). In this study, genomic analysis of MABC isolates was performed to gain greater insights into the epidemiology of circulating strains in Ireland. Whole-genome sequencing (WGS) was performed on 70 MABC isolates that had been referred to the Irish Mycobacteria Reference Laboratory between 2006 and 2017 across nine Irish health care centers. The MABC isolates studied comprised 52 isolates from 27 CF patients and 18 isolates from 10 non-CF patients. WGS identified 57 (81.4%) as subsp, 10 (14.3%) as subsp. , and 3 (4.3%) as subsp. Forty-nine (94%) isolates from 25 CF patients were identified as subsp, whereas 3 (6%) isolates from 2 CF patients were identified as subsp. Among the isolates from non-CF patients, 44% (8/18) were identified as subsp, 39% (7/18) were identified as subsp. , and 17% (3/18) were identified as subsp. WGS detected two clusters of closely related subsp isolates that included isolates from different CF centers. There was a greater genomic diversity of MABC isolates among the isolates from non-CF patients than among the isolates from CF patients. Although WGS failed to show direct evidence of patient-to-patient transmission among CF patients, there was a predominance of two different strains of subsp Furthermore, some MABC isolates were closely related to global strains, suggesting their international spread. Future prospective real-time epidemiological and clinical data along with contemporary MABC sequence analysis may elucidate the sources and routes of transmission among patients infected with MABC.
Topics: Genomics; Humans; Ireland; Mycobacterium Infections, Nontuberculous; Mycobacterium abscessus; Nontuberculous Mycobacteria
PubMed: 32295892
DOI: 10.1128/JCM.00295-20 -
Antimicrobial Agents and Chemotherapy Jan 2020Pulmonary infection with the multidrug-resistant complex (MABSC) is difficult to treat in individuals with cystic fibrosis (CF). MABSC grows as biofilm aggregates in CF...
Pulmonary infection with the multidrug-resistant complex (MABSC) is difficult to treat in individuals with cystic fibrosis (CF). MABSC grows as biofilm aggregates in CF patient lungs, which are known to have anaerobic niches. How aggregation and anoxic conditions affect antibiotic tolerance is not well understood. We sought to determine whether disaggregation and oxygen availability sensitize MABSC isolates to recommended antibiotics. We tested the susceptibilities of 33 isolates from 22 CF patients with MABSC infection and a reference strain to the following antibiotics: amikacin, azithromycin, cefoxitin, ciprofloxacin, clarithromycin, imipenem, kanamycin, linezolid, moxifloxacin, rifampin, tigecycline, and sulfamethoxazole-trimethoprim. Isolates were grown in Mueller-Hinton broth with and without the disaggregating detergent Tween 80 (5%). Time-kill curves at days 1 and 3 were generated for oxic and anoxic amikacin treatment in 4-fold dilutions ranging from 2 to 512 mg liter Scanning electron microscopy was used to visualize the aggregation patterns, while confocal laser scanning microscopy and microrespirometry were used to visualize biofilm growth patterns. Disruption of MABSC aggregates increased susceptibility to amikacin, tigecycline, kanamycin, azithromycin, imipenem, cefoxitin, and clarithromycin ( < 0.05, = 29 to 31). Oxygenation enhanced the killing of disaggregated MABSC isolates by amikacin ( < 0.05) by 1 to 6 log units when 2 to 512 mg liter of amikacin was used. This study explains why current drug susceptibility testing results correlate poorly with treatment outcomes. The conditions achieved by oxic culturing of planktonic isolates do not resemble the hypoxic conditions in CF patient lungs. Biofilm disruption and increased O availability during antibiotic therapy may be new therapeutic strategies for chronic MABSC infection.
Topics: Adolescent; Aerobiosis; Anti-Bacterial Agents; Biofilms; Child; Cystic Fibrosis; Drug Resistance, Multiple, Bacterial; Female; Humans; Lung; Male; Microbial Sensitivity Tests; Mycobacterium Infections, Nontuberculous; Mycobacterium abscessus; Oxygen; Polysorbates; Surface-Active Agents; Young Adult
PubMed: 31740557
DOI: 10.1128/AAC.01212-19 -
PLoS Pathogens Mar 2023Mycobacterium abscessus is the most pathogenic species among the predominantly saprophytic fast-growing mycobacteria. This opportunistic human pathogen causes severe...
Mycobacterium abscessus is the most pathogenic species among the predominantly saprophytic fast-growing mycobacteria. This opportunistic human pathogen causes severe infections that are difficult to eradicate. Its ability to survive within the host was described mainly with the rough (R) form of M. abscessus, which is lethal in several animal models. This R form is not present at the very beginning of the disease but appears during the progression and the exacerbation of the mycobacterial infection, by transition from a smooth (S) form. However, we do not know how the S form of M. abscessus colonizes and infects the host to then multiply and cause the disease. In this work, we were able to show the hypersensitivity of fruit flies, Drosophila melanogaster, to intrathoracic infections by the S and R forms of M. abscessus. This allowed us to unravel how the S form resists the innate immune response developed by the fly, both the antimicrobial peptides- and cellular-dependent immune responses. We demonstrate that intracellular M. abscessus was not killed within the infected phagocytic cells, by resisting lysis and caspase-dependent apoptotic cell death of Drosophila infected phagocytes. In mice, in a similar manner, intra-macrophage M. abscessus was not killed when M. abscessus-infected macrophages were lysed by autologous natural killer cells. These results demonstrate the propensity of the S form of M. abscessus to resist the host's innate responses to colonize and multiply within the host.
Topics: Animals; Humans; Mice; Mycobacterium abscessus; Drosophila melanogaster; Phagocytes; Mycobacterium Infections; Mycobacterium; Drosophila; Mycobacterium Infections, Nontuberculous
PubMed: 36972320
DOI: 10.1371/journal.ppat.1011257 -
ChemistryOpen Mar 2020Non-tuberculous mycobacterium (NTM) infections, such as those caused by , are increasing globally. Due to their intrinsic drug resistance, pulmonary infections are...
Non-tuberculous mycobacterium (NTM) infections, such as those caused by , are increasing globally. Due to their intrinsic drug resistance, pulmonary infections are often difficult to cure using standard chemotherapy. We previously demonstrated that a piperidinol derivative, named PIPD1, is an efficient molecule both against and , the agent of tuberculosis, by targeting the mycolic acid transporter MmpL3. These results prompted us to design and synthesize a series of piperidinol derivatives and to determine the biological activity against . Structure-activity relationship (SAR) studies pointed toward specific sites on the scaffold that can tolerate slight modifications. Overall, these results identified FMD-88 as a new promising active analogue against . Also, we determined the pharmacokinetics properties of PIPD1 and showed that intraperitoneal administration of this compound resulted in promising serum concentration and an elimination half-life of 3.2 hours.
Topics: Antitubercular Agents; Biological Transport; Humans; Membrane Transport Proteins; Microbial Sensitivity Tests; Models, Molecular; Mycobacterium abscessus; Mycolic Acids; Structure-Activity Relationship; Tuberculosis
PubMed: 32211280
DOI: 10.1002/open.202000042 -
Antimicrobial Agents and Chemotherapy Feb 2020is intrinsically resistant to most antimicrobial agents. The emerging infections caused by and the lack of effective treatment call for rapid attention. Here, we...
is intrinsically resistant to most antimicrobial agents. The emerging infections caused by and the lack of effective treatment call for rapid attention. Here, we intended to construct a selectable marker-free autoluminescent strain (designated UAlMab) as a real-time reporter strain to facilitate the discovery of effective drugs and regimens for treating The UAlMab strain was constructed using the /Xer recombinase system. and activities of several drugs, including clofazimine and TB47, a recently reported cytochrome inhibitor, were assessed using UAlMab. Furthermore, the efficacy of multiple drug combinations, including the clofazimine and TB47 combination, were tested against 20 clinical isolates. The UAlMab strain enabled us to evaluate drug efficacy both and in live BALB/c mice in a real-time, noninvasive fashion. Importantly, although TB47 showed marginal activity either alone or in combination with clarithromycin, amikacin, or roxithromycin, the drug markedly potentiated the activity of clofazimine, both and This study demonstrates that the use of the UAlMab strain can significantly facilitate rapid evaluation of new drugs and regimens. The clofazimine and TB47 combination is effective against , and dual/triple electron transport chain (ETC) targeting can be an effective therapeutic approach for treating mycobacterial infections.
Topics: Amikacin; Animals; Anti-Bacterial Agents; Bacterial Proteins; Clarithromycin; Clofazimine; Drug Combinations; Drug Resistance, Multiple, Bacterial; Drug Synergism; Electron Transport; Electron Transport Complex III; Enzyme Inhibitors; Female; Genetic Engineering; Luminescence; Mice; Mice, Inbred BALB C; Microbial Sensitivity Tests; Mycobacterium Infections, Nontuberculous; Mycobacterium abscessus; Optical Imaging; Recombinases; Roxithromycin
PubMed: 31843996
DOI: 10.1128/AAC.01881-19 -
Annals of Clinical Microbiology and... Sep 2023This study investigated the differences in epidemiological and clinical data, and antimicrobial susceptibilities among different subspecies of Mycobacterium abscessus...
OBJECTIVES
This study investigated the differences in epidemiological and clinical data, and antimicrobial susceptibilities among different subspecies of Mycobacterium abscessus complex (MABSC) clinical isolates at a medical school in Thailand.
METHODS
A total of 143 MABSC clinical isolates recovered from 74 patients were genotypically analyzed for erm(41), rrl, and rrs mutations, and antimicrobial susceptibilities were determined using a broth microdilution method. Patient characteristics and clinical outcomes were reviewed from the medical records.
RESULTS
Seventy-four patients were infected with 28/74 (37.8%) M. abscessus subspecies abscessus (MAB), 43/74 (58.1%) M. abscessus subsp. massiliense (MMA), and 3/74 (4.1%) M. abscessus subsp. bolletii (MBO). The clinical findings and outcomes were generally indistinguishable between the three subspecies. All three subspecies of MABSC clinical isolates exhibited high resistance rates to ciprofloxacin, doxycycline, moxifloxacin, TMP/SMX, and tobramycin. MAB had the highest resistance rates to clarithromycin (27.8%, 20/72) and amikacin (6.9%, 5/72) compared to MBO and MMA, with p < 0.001 and p = 0.004, respectively. In addition, the rough morphotype was significantly associated with resistance to amikacin (8.9%, 5/56), clarithromycin (26.8%, 15/56), and imipenem (76.8%, 43/56) (p < 0.001), whereas the smooth morphotype was resistant to linezolid (57.1%, 48/84) (p = 0.002). In addition, T28 of erm(41), rrl (A2058C/G and A2059C/G), and rrs (A1408G) mutations were detected in 87.4% (125/143), 16.1% (23/143), and 9.1% (13/143) of MABSC isolates, respectively.
CONCLUSIONS
Three MABSC subspecies caused a variety of infections in patients with different underlying comorbidities. The drug susceptibility patterns of the recent circulating MABSC strains in Thailand were different among the three MABSC subspecies and two morphotypes.
Topics: Humans; Clarithromycin; Schools, Medical; Thailand; Mycobacterium abscessus; Amikacin; Mycobacterium Infections, Nontuberculous; Anti-Infective Agents
PubMed: 37735687
DOI: 10.1186/s12941-023-00637-4