-
Antimicrobial Agents and Chemotherapy Sep 2022Nontuberculous mycobacteria (NTM) infections are increasing worldwide. Mycobacterium avium complex (MAC) and the M. abscessus species are the most commonly cultured NTM...
Nontuberculous mycobacteria (NTM) infections are increasing worldwide. Mycobacterium avium complex (MAC) and the M. abscessus species are the most commonly cultured NTM and treatment options are limited, especially for the M. abscessus species. In this study, the activity of eravacycline, a new tetracycline derivative, was tested against 110 clinical isolates of NTM. MIC testing was performed as recommended by the Clinical and Laboratory Standards Institute against 60 isolates of rapidly growing mycobacteria (RGM), of which ~70% were tetracycline resistant. These included M. abscessus subsp. (8 isolates), M. abscessus subsp. (5), M. chelonae (10), (3), M. fortuitum group (20) including 12 doxycycline-resistant isolates, and M. mucogenicum group (10) including three doxycycline-resistant isolates. Due to trailing, eravacycline MICs were read at 80% and 100% inhibition. Eravacycline was active against all RGM species, with MIC ranges of ≤0.015 to 0.5 and ≤0.015 to 0.12 μg/mL for 100% and 80% inhibition, respectively. For M. abscessus subsp. , MIC values were 0.12 and 0.03 μg/mL with 100% and 80% inhibition, respectively. MICs for tigecycline were generally within 1 to 2 dilutions of the 100%-inhibition eravacycline MIC values. Fifty isolates of slowly growing mycobacteria (SGM) species, including 16 isolates of MAC, were also tested. While there was no trailing observed in most SGM, the eravacycline MICs were higher (MIC range of >8 μg/mL), except for M. kansasii and M. marinum which had MIC values of 1 μg/mL. This study supports further evaluation of eravacycline, including clinical trials for the development of RGM treatment regimens, especially for M. abscessus.
Topics: Anti-Bacterial Agents; Doxycycline; Humans; Microbial Sensitivity Tests; Mycobacterium; Mycobacterium Infections, Nontuberculous; Mycobacterium avium Complex; Nontuberculous Mycobacteria; Tetracycline; Tetracyclines; Tigecycline
PubMed: 35943269
DOI: 10.1128/aac.00689-22 -
Microbial Cell Factories Mar 20239α-hydroxyandrost-4-ene-3,17-dione (9-OHAD) is a significant intermediate for the synthesis of glucocorticoid drugs. However, in the process of phytosterol...
Improving the production of 9α-hydroxy-4-androstene-3,17-dione from phytosterols by 3-ketosteroid-Δ-dehydrogenase deletions and multiple genetic modifications in Mycobacterium fortuitum.
BACKGROUND
9α-hydroxyandrost-4-ene-3,17-dione (9-OHAD) is a significant intermediate for the synthesis of glucocorticoid drugs. However, in the process of phytosterol biotransformation to manufacture 9-OHAD, product degradation, and by-products restrict 9-OHAD output. In this study, to construct a stable and high-yield 9-OHAD producer, we investigated a combined strategy of blocking Δ‑dehydrogenation and regulating metabolic flux.
RESULTS
Five 3-Ketosteroid-Δ-dehydrogenases (KstD) were identified in Mycobacterium fortuitum ATCC 35855. KstD2 showed the highest catalytic activity on 3-ketosteroids, followed by KstD3, KstD1, KstD4, and KstD5, respectively. In particular, KstD2 had a much higher catalytic activity for C9 hydroxylated steroids than for C9 non-hydroxylated steroids, whereas KstD3 showed the opposite characteristics. The deletion of kstDs indicated that KstD2 and KstD3 were the main causes of 9-OHAD degradation. Compared with the wild type M. fortuitum ATCC 35855, MFΔkstD, the five kstDs deficient strain, realized stable accumulation of 9-OHAD, and its yield increased by 42.57%. The knockout of opccr or the overexpression of hsd4A alone could not reduce the metabolic flux of the C22 pathway, while the overexpression of hsd4A based on the knockout of opccr in MFΔkstD could remarkably reduce the contents of 9,21 ‑dihydroxy‑20‑methyl‑pregna‑4‑en‑3‑one (9-OHHP) by-products. The inactivation of FadE28-29 leads to a large accumulation of incomplete side-chain degradation products. Therefore, hsd4A and fadE28-29 were co-expressed in MFΔkstDΔopccr successfully eliminating the two by-products. Compared with MFΔkstD, the purity of 9-OHAD improved from 80.24 to 90.14%. Ultimately, 9‑OHAD production reached 12.21 g/L (83.74% molar yield) and the productivity of 9-OHAD was 0.0927 g/L/h from 20 g/L phytosterol.
CONCLUSIONS
KstD2 and KstD3 are the main dehydrogenases that lead to 9-OHAD degradation. Hsd4A and Opccr are key enzymes regulating the metabolic flux of the C19- and C22-pathways. Overexpression of fadE28-29 can reduce the accumulation of incomplete degradation products of the side chains. According to the above findings, the MF-FA5020 transformant was successfully constructed to rapidly and stably accumulate 9-OHAD from phytosterols. These results contribute to the understanding of the diversity and complexity of steroid catabolism regulation in actinobacteria and provide a theoretical basis for further optimizing industrial microbial catalysts.
Topics: Phytosterols; Mycobacterium fortuitum; Androstenedione; Oxidoreductases; Steroids
PubMed: 36922830
DOI: 10.1186/s12934-023-02052-y -
Microbial Drug Resistance (Larchmont,... Jul 2023There is a scarcity of data regarding the antimicrobial susceptibility testing profiles of nontuberculous mycobacterial (NTM) in Israel and other Middle Eastern...
There is a scarcity of data regarding the antimicrobial susceptibility testing profiles of nontuberculous mycobacterial (NTM) in Israel and other Middle Eastern countries. We aimed to describe the antimicrobial susceptibility profiles of NTM in Israel. A total of 410 clinical isolates of NTM, identified to the species level using matrix-assisted laser desorption ionization-time of flight mass spectrometry or gene sequencing, were included. Minimum inhibitory concentrations for slowly growing mycobacteria (SGM) and rapidly growing mycobacteria (RGM) for 12 and 11 drugs were determined using the Sensititre SLOMYCOI and RAPMYCOI broth microdilution plates, respectively. complex (MAC) was the most frequently isolated species ( = 148; 36%), followed by ( = 93; 23%), group ( = 62; 15%), ( = 27; 7%), and ( = 22; 5%) accounting together for 86% of isolates. The most active agents against SGM were amikacin (98%/85%/100%) and clarithromycin (97%/99%/100%), followed by moxifloxacin (25%/10%/100%) and linezolid (3%/6%/100%) for MAC, , and , respectively. For RGM, the most active agents were amikacin (98%/100%/88%) followed by linezolid (48%/80%/100%) and clarithromycin (39%/28%/94%) for group, , and , respectively. These findings can assist in guiding the treatment of NTM infections.
Topics: Humans; Nontuberculous Mycobacteria; Mycobacterium Infections, Nontuberculous; Linezolid; Anti-Bacterial Agents; Amikacin; Clarithromycin; Israel; Microbial Sensitivity Tests
PubMed: 37219996
DOI: 10.1089/mdr.2023.0024 -
Applied Microbiology and Biotechnology Oct 2023Biofilm formation by Mycobacterium fortuitum causes serious threats to human health due to its increased contribution to nosocomial infections. In this study, the first... (Comparative Study)
Comparative Study
Biofilm formation by Mycobacterium fortuitum causes serious threats to human health due to its increased contribution to nosocomial infections. In this study, the first comprehensive global proteome analysis of M. fortuitum was reported under planktonic and biofilm growth states. A label-free Q Exactive Quadrupole-Orbitrap tandem mass spectrometry analysis was performed on the protein lysates. The differentially abundant proteins were functionally characterized and re-annotated using Blast2GO and CELLO2GO. Comparative analysis of the proteins among two growth states provided insights into the phenotypic switch, and fundamental pathways associated with pathobiology of M. fortuitum biofilm, such as lipid biosynthesis and quorum-sensing. Interaction network generated by the STRING database revealed associations between proteins that endure M. fortuitum during biofilm growth state. Hypothetical proteins were also studied to determine their functional alliance with the biofilm phenotype. CARD, VFDB, and PATRIC analysis further showed that the proteins upregulated in M. fortuitum biofilm exhibited antibiotic resistance, pathogenesis, and virulence. Heatmap and correlation analysis provided the biomarkers associated with the planktonic and biofilm growth of M. fortuitum. Proteome data was validated by qPCR analysis. Overall, the study provides insights into previously unexplored biochemical pathways that can be targeted by novel inhibitors, either for shortened treatment duration or for eliminating biofilm of M. fortuitum and related nontuberculous mycobacterial pathogens. KEY POINTS: • Proteomic analyses of M. fortuitum reveals novel biofilm markers. • Acetyl-CoA acetyltransferase acts as the phenotype transition switch. • The study offers drug targets to combat M. fortuitum biofilm infections.
Topics: Mycobacterium fortuitum; Biofilms; Microscopy, Electron, Scanning; Proteome; Metabolic Networks and Pathways; Acetyl-CoA C-Acetyltransferase; Quorum Sensing
PubMed: 37542577
DOI: 10.1007/s00253-023-12705-y -
Ocular Immunology and Inflammation Jan 2022To report clinical features and outcomes of nontuberculous mycobacteria (NTM) presenting as uveitis in HIV positive patients.
PURPOSE
To report clinical features and outcomes of nontuberculous mycobacteria (NTM) presenting as uveitis in HIV positive patients.
MATERIALS AND METHODS
Retrospective study of HIV positive patients who were diagnosed as uveitis due to NTM.
RESULTS
Six eyes of four HIV positive patients with NTM were studied. Average age at presentation was 35.5 years (range 30-38). With specific PCR primers, was detected in three patients (75%) and in one patient (25%). Culture was positive in two cases. Two eyes (33.33%) each had endophthalmitis and necrotizing retinitis like picture, one eye (16.66%) each had chorioretinitis and frosted branch angitis like. Visual acuity improved in two eyes (33.33%), worsened in three eyes (50%), and remained unchanged in one eye (16.6%).
CONCLUSIONS
NTM infection is a unique entity in immunosuppressed with poor visual outcome. PCR forms a useful tool for rapid diagnosis and timely initiation of specific anti-tuberculosis therapy.
Topics: Adult; HIV Seropositivity; Humans; Mycobacterium; Mycobacterium Infections, Nontuberculous; Nontuberculous Mycobacteria; Retrospective Studies; Uveitis
PubMed: 32813606
DOI: 10.1080/09273948.2020.1788610 -
Acta Microbiologica Et Immunologica... Jun 2023Mycobacterium fortuitum is a clinically important species among nontuberculous mycobacteria (NTM). Treatment of diseases caused by NTM is challenging. The aim of this...
Frequency of mutations in erm(39) related to clarithromycin resistance and in rrl related to linezolid resistance in clinical isolates of Mycobacterium fortuitum in Iran.
Mycobacterium fortuitum is a clinically important species among nontuberculous mycobacteria (NTM). Treatment of diseases caused by NTM is challenging. The aim of this study was identification of drug susceptibility and detection of mutations in erm(39) related to clarithromycin resistance and in rrl related to linezolid resistance in clinical isolates of M. fortuitum in Iran. In the study, 328 clinical NTM isolates were subjected to identification based on rpoB and 15% of isolates were assigned to M. fortuitum. Minimum inhibitory concentration for clarithromycin and linezolid was determined by E-test. Altogether 64% of M. fortuitum isolates showed resistanc to clarithromycin and 18% of M. fortuitum isolates showed resistance to linezolid. PCR and DNA sequencing were performed in erm(39) and in rrl genes for detection of mutations related to clarithromycin and linezolid resistance, respectively. Sequencing analysis revealed (84.37%) single nucleotide polymorphisms in the erm(39). A total 55.55% of M. fortuitum isolates harbored an A→G, 14.81% harbored an C→A, 29.62% harbored an G→T mutation in erm(39) at position 124, 135, 275. Seven strains harbored point mutation in the rrl gene either at T2131C or at A2358G. Our findings showed M. fortuitum isolates have become a serious problem with high-level antibiotic resistance. The existence of drug resistance to clarithromycin and linezolid indicates more attention to the study of drug resistance in M. fortuitum.
Topics: Clarithromycin; Linezolid; Mycobacterium fortuitum; Iran; Anti-Bacterial Agents; Nontuberculous Mycobacteria; Mutation; Microbial Sensitivity Tests; Drug Resistance, Bacterial
PubMed: 37224008
DOI: 10.1556/030.2023.02020 -
HIV Medicine Mar 2022In this study, the distribution of nontuberculous mycobacteria (NTM) strains in patients with and without HIV/AIDS in Chongqing, China was evaluated.
INTRODUCTION
In this study, the distribution of nontuberculous mycobacteria (NTM) strains in patients with and without HIV/AIDS in Chongqing, China was evaluated.
METHODS
A retrospective study was performed in January-December 2020 at Chongqing Public Health Medical Center. NTM strains were assessed by a multi locus phylogenetic analysis. The distribution of NTM strains in HIV/AIDS and non-HIV/AIDS groups was compared. CD4+ cell counts, imaging changes, and characteristics of mycobacterial species were determined.
RESULTS
In total, 324 patients with NTM infection (50 patients with HIV/AIDS and 274 patients without HIV/AIDS) were included. The most common etiological agent was M. abscessus (29%), followed by M. paraintracellulare (12%) and M. colombiense (11%). Predominant NTM species were M. avium (26%), M. colombiense (24%), and M. kansasii (18%) in patients with HIV/AIDS and were M. abscessus (32%), M. paraintracellulare (13%), M. fortuitum (10%), and M. intracellulare (10%) in patients without HIV/AIDS. For a CD4+ cell count of <200/μl, the predominant species were M. aviumin the HIV/AIDS group and M. abscessus in the non-HIV/AIDS group. With respect to radiologic characteristics, different NTM strains were associated with distinct imaging manifestations; for example, M. marseillense, M. kansasii, and M. parasenchytosis were more likely to induce cavities. Imaging cavities, bronchiectasis, and acinar-like changes were more common in the non-HIV/AIDS groups.
CONCLUSIONS
The infection rates of HIV and NTM in Chongqing are high, while M. abscessus, M. paraintracellulare, and M. colombiense are the main pathogens causing NTM diseases in Chongqing, and NTM strains differed significantly between patients with and without HIV/AIDS. Monitoring these indicators can help develop prevention strategies.
Topics: Acquired Immunodeficiency Syndrome; HIV Infections; Humans; Mycobacterium Infections, Nontuberculous; Nontuberculous Mycobacteria; Phylogeny; Retrospective Studies
PubMed: 35293104
DOI: 10.1111/hiv.13249 -
Infection and Drug Resistance 2022Nontuberculous mycobacteria (NTM) can cause pulmonary and extrapulmonary diseases. Tedizolid (TZD) is a new oxazolidinone with in vitro activity against NTM such as...
OBJECTIVE
Nontuberculous mycobacteria (NTM) can cause pulmonary and extrapulmonary diseases. Tedizolid (TZD) is a new oxazolidinone with in vitro activity against NTM such as complex (MAC), , and complex. The aim of this study was to evaluate the TZD susceptibility profiles of clinical isolates of NTM.
METHODS
The microdilution method was used to identify the minimum inhibitory concentration (MIC) of TZD and linezolid (LZD) for 133 clinical NTM isolates. Broth microdilution chequerboard assays were used to investigate the synergistic effects of TZD and three antibiotics on two reference isolates and eleven clinical isolates of NTM.
RESULTS
The TZD MIC and MIC for complex were 2 and 4 μg/mL, 16 and >32 μg/mL for MAC, respectively. TZD exhibited lower MICs than that of LZD for most NTM, which were positively correlated. Due to the high MIC values of TZD against MAC, it is necessary to conduct drug sensitivity tests before TZD administration. TZD-clarithromycin combination had synergistic response on complex in 3 of the 8 isolates, which lasted only 3-5 days. TZD-cefoxitin had synergistic effect against all five isolates.
CONCLUSION
Our study demonstrates that TZD had greater in vitro potency than LZD, and synergy studies suggested that TZD may be an important component of multi-drug treatment regimen.
PubMed: 36045871
DOI: 10.2147/IDR.S362583 -
Cureus Jun 2023Introduction complex (MTBC), the primary cause of tuberculosis (TB), must be accurately identified to implement effective patient management and control strategies....
Introduction complex (MTBC), the primary cause of tuberculosis (TB), must be accurately identified to implement effective patient management and control strategies. Non-tuberculous mycobacteria (NTM) in suspected TB cases can result in erroneous diagnoses and needless treatment. Objective The study aimed to identify NTM in patients suspected of TB at a tertiary care hospital in central India using molecular methods. Methods This prospective study enrolled 400 suspected pulmonary and extra-pulmonary TB patients. Patients between the age of two to 90 years, of either gender, new and previously treated cases, Culture positive, patients with immune-compromised status, patients not responding to ATT, HIV positive and negative, and willing to give consent were included in the study. Liquid culture via the Mycobacterial growth indicator tube (MGIT) system was used to culture mycobacteria from clinical samples. The SD Bioline Ag MPT64 Test (Standard Diagnostics, South Korea) and in-house multiplex-PCR (mPCR) were used to differentiate between Mycobacterium tuberculosis complex and NTM species for the molecular identification of NTM GenoType® Mycobacterium Common Mycobacteria (CM) assay kit (HAIN Life Science, Nehren, Germany) was used following the manufacturer's protocol. Results Only 59/400 (14.7%) of the samples produced a positive result in MGIT culture, indicating the presence of mycobacteria, and 85.25% of the remaining 341 samples were negative for mycobacterial growth. Further investigation of these 59 cultures with mPCR and SD Bioline Ag MPT64 test showed that 12 (20.33%) cultures were determined to be NTM, while the remaining 47 (79.67%) were identified as MTBC. Genotype characterization with GenoType® mycobacterium CM assay kit revealed that five of the 12 NTM isolates (41.67%) showed patterns that were consistent with (, three (25%) showed patterns that were consistent with , and four (33.33%) showed patterns that were consistent with . Conclusion These results emphasize the value of molecular methods for precisely identifying mycobacterial species, particularly in suspected TB cases. The high prevalence of NTM in positive cultures emphasizes the significance of differentiating between MTBC and NTM to prevent misdiagnosis and ensure proper care. Understanding the epidemiology and clinical significance of these organisms in central India is made possible by the identification of particular NTM species.
PubMed: 37416024
DOI: 10.7759/cureus.39992 -
Microorganisms Nov 2021group (MFG) members are able to cause clinical mycobacteriosis in fish and other animals including humans. , , , , , , , , and were isolated from fish with...
Clinical Relevance and Environmental Prevalence of Group Members. Comment on Mugetti et al. Gene Sequencing and Phylogenetic Analysis: Powerful Tools for an Improved Diagnosis of Fish Mycobacteriosis Caused by Group Members. 2021, , 797.
group (MFG) members are able to cause clinical mycobacteriosis in fish and other animals including humans. , , , , , , , , and were isolated from fish with mycobacteriosis. In other animals only three MFG species have been isolated: from camels' milk, from cutaneous infections often described as "farcy", and from different domestic and wild mammals' species. Out of 17, only 3 MFG species (, and ) have never been reported in humans. A total of eight MFG members (, , , subsp. , , , , and ) have been isolated from both pulmonary and extrathoracic locations. In extrathoracic tissues five MFG species (, , , , and ) have been diagnosed and only one MFG member ( subsp. ) has been isolated from pulmonary infection.
PubMed: 34835470
DOI: 10.3390/microorganisms9112345