scholarly journals Toward a Phage Cocktail for Tuberculosis: Susceptibility and Tuberculocidal Action of Mycobacteriophages against Diverse Mycobacterium tuberculosis Strains

mBio ◽  
2021 ◽  
Vol 12 (3) ◽  
Author(s):  
Carlos A. Guerrero-Bustamante ◽  
Rebekah M. Dedrick ◽  
Rebecca A. Garlena ◽  
Daniel A. Russell ◽  
Graham F. Hatfull
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Genome Engineering ◽  
Therapeutic Potential ◽  
Mycobacterium Abscessus ◽  
Cross Resistance ◽  
Multiple Drug ◽  
Drug Resistant ◽  
Content Type ◽  
Phage Cocktail ◽  
Mdr Tb

ABSTRACT The global health burden of human tuberculosis (TB) and the widespread antibiotic resistance of its causative agent Mycobacterium tuberculosis warrant new strategies for TB control. The successful use of a bacteriophage cocktail to treat a Mycobacterium abscessus infection suggests that phages could play a role in tuberculosis therapy. To assemble a phage cocktail with optimal therapeutic potential for tuberculosis, we have explored mycobacteriophage diversity to identify phages that demonstrate tuberculocidal activity and determined the phage infection profiles for a diverse set of strains spanning the major lineages of human-adapted strains of the Mycobacterium tuberculosis complex. Using a combination of genome engineering and bacteriophage genetics, we have assembled a five-phage cocktail that minimizes the emergence of phage resistance and cross-resistance to multiple phages, and which efficiently kills the M. tuberculosis strains tested. Furthermore, these phages function without antagonizing antibiotic effectiveness, and infect both isoniazid-resistant and -sensitive strains. IMPORTANCE Tuberculosis kills 1.5 million people each year, and resistance to commonly used antibiotics contributes to treatment failures. The therapeutic potential of bacteriophages against Mycobacterium tuberculosis offers prospects for shortening antibiotic regimens, provides new tools for treating multiple drug-resistant (MDR)-TB and extensively drug-resistant (XDR)-TB infections, and protects newly developed antibiotics against rapidly emerging resistance to them. Identifying a suitable suite of phages active against diverse M. tuberculosis isolates circumvents many of the barriers to initiating clinical evaluation of phages as part of the arsenal of antituberculosis therapeutics.

scholarly journals Comparison of In Vitro Activity and MIC Distributions between the Novel Oxazolidinone Delpazolid and Linezolid against Multidrug-Resistant and Extensively Drug-Resistant Mycobacterium tuberculosis in China

2018 ◽  
Vol 62 (8) ◽  
Author(s):  
Zhaojing Zong ◽  
Wei Jing ◽  
Jin Shi ◽  
Shu'an Wen ◽  
Tingting Zhang ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Novel Mutation ◽  
Multidrug Resistant ◽  
23S Rrna ◽  
Drug Resistant ◽  
Content Type ◽  
Extensively Drug Resistant ◽  
Significant Difference ◽  
Mdr Tb

ABSTRACT Oxazolidinones are efficacious in treating mycobacterial infections, including tuberculosis (TB) caused by drug-resistant Mycobacterium tuberculosis. In this study, we compared the in vitro activities and MIC distributions of delpazolid, a novel oxazolidinone, and linezolid against multidrug-resistant TB (MDR-TB) and extensively drug-resistant TB (XDR-TB) in China. Additionally, genetic mutations in 23S rRNA, rplC, and rplD genes were analyzed to reveal potential mechanisms underlying the observed oxazolidinone resistance. A total of 240 M. tuberculosis isolates were included in this study, including 120 MDR-TB isolates and 120 XDR-TB isolates. Overall, linezolid and delpazolid MIC90 values for M. tuberculosis isolates were 0.25 mg/liter and 0.5 mg/liter, respectively. Based on visual inspection, we tentatively set epidemiological cutoff (ECOFF) values for MIC determinations for linezolid and delpazolid at 1.0 mg/liter and 2.0 mg/liter, respectively. Although no significant difference in resistance rates was observed between linezolid and delpazolid among XDR-TB isolates (P > 0.05), statistical analysis revealed a significantly greater proportion of linezolid-resistant isolates than delpazolid-resistant isolates within the MDR-TB group (P = 0.036). Seven (53.85%) of 13 linezolid-resistant isolates were found to harbor mutations within the three target genes. Additionally, 1 isolate exhibited an amino acid substitution (Arg126His) within the protein encoded by rplD that contributed to high-level resistance to linezolid (MIC of >16 mg/liter), compared to a delpazolid MIC of 0.25. In conclusion, in vitro susceptibility testing revealed that delpazolid antibacterial activity was comparable to that of linezolid. A novel mutation within rplD that endowed M. tuberculosis with linezolid, but not delpazolid, resistance was identified.

scholarly journals Genotypic Analysis of Genes Associated with Independent Resistance and Cross-Resistance to Isoniazid and Ethionamide in Mycobacterium tuberculosis Clinical Isolates

2015 ◽  
Vol 59 (12) ◽  
pp. 7805-7810 ◽  
Author(s):  
Johana Rueda ◽  
Teresa Realpe ◽  
Gloria Isabel Mejia ◽  
Elsa Zapata ◽  
Juan Carlos Rozo ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Multidrug Resistant ◽  
Cross Resistance ◽  
Resistance Mutations ◽  
Content Type ◽  
Genotypic Analysis ◽  
Mdr Tb ◽  
Proportion Method ◽  
Emergence Of Resistance ◽  
Phenotypic Susceptibility

ABSTRACTEthionamide (ETH) is an antibiotic used for the treatment of multidrug-resistant (MDR) tuberculosis (TB) (MDR-TB), and its use may be limited with the emergence of resistance in theMycobacterium tuberculosispopulation. ETH resistance inM. tuberculosisis phenomenon independent or cross related when accompanied with isoniazid (INH) resistance. In most cases, resistance to INH and ETH is explained by mutations in theinhApromoter and in the following genes:katG,ethA,ethR,mshA,ndh, andinhA. We sequenced the above genes in 64M. tuberculosisisolates (n= 57 ETH-resistant MDR-TB isolates;n= 3 ETH-susceptible MDR-TB isolates; andn= 4 fully susceptible isolates). Each isolate was tested for susceptibility to first- and second-line drugs using the agar proportion method. Mutations were observed in ETH-resistant MDR-TB isolates at the following rates: 100% inkatG, 72% inethA, 45.6% inmshA, 8.7% inndh, and 33.3% ininhAor its promoter. Of the three ETH-susceptible MDR-TB isolates, all showed mutations inkatG; one had a mutation inethA, and another, inmshAandinhA. Finally, of the four fully susceptible isolates, two showed no detectable mutation in the studied genes, and two had mutations inmshAgene unrelated to the resistance. Mutations not previously reported were found in theethA,mshA,katG, andndhgenes. The concordance between the phenotypic susceptibility testing to INH and ETH and the sequencing was 1 and 0.45, respectively. Among isolates exhibiting INH resistance, the high frequency of independent resistance and cross-resistance with ETH in theM. tuberculosisisolates suggests the need to confirm the susceptibility to ETH before considering it in the treatment of patients with MDR-TB.

scholarly journals Aminoglycoside Cross-Resistance in Mycobacterium tuberculosis Due to Mutations in the 5′ Untranslated Region ofwhiB7

2013 ◽  
Vol 57 (4) ◽  
pp. 1857-1865 ◽  
Author(s):  
Analise Z. Reeves ◽  
Patricia J. Campbell ◽  
Razvan Sultana ◽  
Seidu Malik ◽  
Megan Murray ◽  
...  
Keyword(s):  
Antibiotic Resistance ◽  
Mycobacterium Tuberculosis ◽  
Untranslated Region ◽  
Regulatory Region ◽  
Cross Resistance ◽  
Drug Resistant ◽  
Second Line ◽  
Content Type ◽  
Low Level ◽  
Level Resistance

ABSTRACTSince the discovery of streptomycin's bactericidal activity againstMycobacterium tuberculosis, aminoglycosides have been utilized to treat tuberculosis (TB). Today, the aminoglycosides kanamycin and amikacin are used to treat multidrug-resistant (MDR) TB, and resistance to any of the second-line injectable antibiotics, including kanamycin, amikacin, or capreomycin, is a defining characteristic of extensively drug-resistant (XDR) TB. Resistance to kanamycin and streptomycin is thought to be due to the acquisition of unlinked chromosomal mutations. However, we identified eight independent mutations in the 5′ untranslated region of the transcriptional activatorwhiB7that confer low-level resistance to both aminoglycosides. The mutations lead to 23- to 145-fold increases inwhiB7transcripts and subsequent increased expression of botheis(Rv2416c) andtap(Rv1258c). Increased expression ofeisconfers kanamycin resistance in these mutants, while increased expression oftap, which encodes an efflux pump, is a previously uncharacterized mechanism of low-level streptomycin resistance. Additionally, high-level resistance to streptomycin arose at a much higher frequency inwhiB7mutants than in a wild-type (WT) strain. AlthoughwhiB7is typically associated with intrinsic antibiotic resistance inM. tuberculosis, these data suggest that mutations in an uncharacterized regulatory region ofwhiB7contribute to cross-resistance against clinically used second-line antibiotics. As drug resistance continues to develop and spread, understanding the mechanisms and molecular basis of antibiotic resistance is critical for the development of rapid molecular tests to diagnose drug-resistant TB strains and ultimately for designing regimens to treat drug-resistant cases of TB.

scholarly journals Mycobacterium tuberculosis Subculture Results in Loss of Potentially Clinically Relevant Heteroresistance

2017 ◽  
Vol 61 (11) ◽  
Author(s):  
John Z. Metcalfe ◽  
Elizabeth Streicher ◽  
Grant Theron ◽  
Rebecca E. Colman ◽  
Renee Penaloza ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Multidrug Resistant ◽  
Multiple Drug ◽  
Drug Resistant ◽  
Genotypic Resistance ◽  
Content Type ◽  
Multiple Drug Resistant ◽  
Dynamic Loss ◽  
Tb Diagnostics ◽  
The Impact

ABSTRACT Multidrug-resistant tuberculosis (TB) presents a major public health dilemma. Heteroresistance, the coexistence of drug-resistant and drug-susceptible strains or of multiple drug-resistant strains with discrete haplotypes, may affect accurate diagnosis and the institution of effective treatment. Subculture, or passage of cells onto fresh growth medium, is utilized to preserve Mycobacterium tuberculosis cell lines and is universally employed in TB diagnostics. The impact of such passages, typically performed in the absence of drug, on drug-resistant subpopulations is hypothesized to vary according to the competitive costs of genotypic resistance-associated variants. We applied ultradeep next-generation sequencing to 61 phenotypically rifampin-monoresistant (n = 17) and preextensively (n = 41) and extensively (n = 3) drug-resistant isolates with presumptive heteroresistance at two time points in serial subculture. We found significant dynamic loss of minor-variant resistant subpopulations across all analyzed resistance-determining regions, including eight isolates (13%) whose antibiogram data would have transitioned from resistant to susceptible for at least one drug through subculture. Surprisingly, some resistance-associated variants appeared to be selected for in subculture.

scholarly journals Primary Clofazimine and Bedaquiline Resistance among Isolates from Patients with Multidrug-Resistant Tuberculosis

2017 ◽  
Vol 61 (6) ◽  
Author(s):  
Jian Xu ◽  
Bin Wang ◽  
Minghao Hu ◽  
Fengmin Huo ◽  
Shaochen Guo ◽  
...  
Keyword(s):  
Multidrug Resistant ◽  
Cross Resistance ◽  
Drug Resistant ◽  
Content Type ◽  
Tb Treatment ◽  
Resistant Tuberculosis ◽  
Multidrug Resistant Tuberculosis ◽  
Mdr Tb ◽  
Previously Treated ◽  
Alamarblue Assay

ABSTRACT Clofazimine has been repurposed for the treatment of tuberculosis, especially for multidrug-resistant tuberculosis (MDR-TB). To test the susceptibility to clofazimine of Mycobacterium tuberculosis clinical isolates, MICs of clofazimine were determined using the microplate alamarBlue assay (MABA) method for 80 drug-resistant isolates and 10 drug-susceptible isolates for comparison. For five clofazimine-resistant strains isolated from previously treated pre-extensively drug-resistant TB (pre-XDR-TB) and XDR-TB patients without prior exposure to clofazimine or bedaquiline, clofazimine MICs were ≥1.2 μg/ml. Four isolates with cross-resistance to bedaquiline had Rv0678 mutations. The other isolate with no resistance to bedaquiline had an Rv1979c mutation. This study adds to a recent study showing that 6.3% of MDR-TB patients without prior clofazimine or bedaquiline exposure harbored isolates with Rv0678 mutations, which raises concern that preexisting resistance to these drugs may be associated with prior TB treatment. Furthermore, we propose a tentative breakpoint of 1.2 μg/ml for clofazimine resistance using the MABA method. More-widespread surveillance and individualized testing for clofazimine and bedaquiline resistance, together with assessment of their clinical usage, especially among previously treated and MDR-TB patients, are warranted.

scholarly journals Carbapenems and Rifampin Exhibit Synergy against Mycobacterium tuberculosis and Mycobacterium abscessus

2015 ◽  
Vol 59 (10) ◽  
pp. 6561-6567 ◽  
Author(s):  
Amit Kaushik ◽  
Nayani Makkar ◽  
Pooja Pandey ◽  
Nicole Parrish ◽  
Urvashi Singh ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Clinical Utility ◽  
Mycobacterium Abscessus ◽  
Synergistic Activity ◽  
Drug Resistant ◽  
Content Type ◽  
Tb Treatment ◽  
Resistant Mutants ◽  
Multiple Drugs ◽  
Spontaneous Mutants

ABSTRACTAn effective regimen for treatment of tuberculosis (TB) is comprised of multiple drugs that inhibit a range of essential cellular activities inMycobacterium tuberculosis. The effectiveness of a regimen is further enhanced if constituent drugs act with synergy. Here, we report that faropenem (a penem) or biapenem, doripenem, or meropenem (carbapenems), which belong to the β-lactam class of antibiotics, and rifampin, one of the drugs that forms the backbone of TB treatment, act with synergy when combined. One of the reasons (carba)penems are seldom used for treatment of TB is the high dosage levels required, often at the therapeutic limits. The synergistic combination of rifampin and these (carba)penems indicates that (carba)penems can be administered at dosages that are therapeutically relevant. The combination of faropenem and rifampin also limits the frequency of resistant mutants, as we were unable to obtain spontaneous mutants in the presence of these two drugs. The combinations of rifampin and (carba)penems were effective not only against drug-sensitiveMycobacterium tuberculosisbut also against drug-resistant clinical isolates that are otherwise resistant to rifampin. A combination of doripenem or biapenem and rifampin also exhibited synergistic activity againstMycobacterium abscessus. Although the MICs of these three drugs alone againstM. abscessusare too high to be of clinical relevance, their concentrations in combinations are therapeutically relevant; therefore, they warrant further evaluation for clinical utility to treatMycobacterium abscessusinfection, especially in cystic fibrosis patients.

scholarly journals The Cyclic Peptide Ecumicin Targeting ClpC1 Is Active against Mycobacterium tuberculosis In Vivo

2014 ◽  
Vol 59 (2) ◽  
pp. 880-889 ◽  
Author(s):  
Wei Gao ◽  
Jin-Yong Kim ◽  
Jeffrey R. Anderson ◽  
Tatos Akopian ◽  
Seungpyo Hong ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
High Throughput Screening ◽  
Cyclic Peptide ◽  
Genome Mining ◽  
Subcutaneous Administration ◽  
Multiple Drug ◽  
Drug Resistant ◽  
Content Type

ABSTRACTDrug-resistant tuberculosis (TB) has lent urgency to finding new drug leads with novel modes of action. A high-throughput screening campaign of >65,000 actinomycete extracts for inhibition ofMycobacterium tuberculosisviability identified ecumicin, a macrocyclic tridecapeptide that exerts potent, selective bactericidal activity againstM. tuberculosisin vitro, including nonreplicating cells. Ecumicin retains activity against isolated multiple-drug-resistant (MDR) and extensively drug-resistant (XDR) strains ofM. tuberculosis. The subcutaneous administration to mice of ecumicin in a micellar formulation at 20 mg/kg body weight resulted in plasma and lung exposures exceeding the MIC. Complete inhibition ofM. tuberculosisgrowth in the lungs of mice was achieved following 12 doses at 20 or 32 mg/kg. Genome mining of lab-generated, spontaneous ecumicin-resistantM. tuberculosisstrains identified the ClpC1 ATPase complex as the putative target, and this was confirmed by a drug affinity response test. ClpC1 functions in protein breakdown with the ClpP1P2 protease complex. Ecumicin markedly enhanced the ATPase activity of wild-type (WT) ClpC1 but prevented activation of proteolysis by ClpC1. Less stimulation was observed with ClpC1 from ecumicin-resistant mutants. Thus, ClpC1 is a valid drug target againstM. tuberculosis, and ecumicin may serve as a lead compound for anti-TB drug development.

scholarly journals 1H-Benzo[d]Imidazole Derivatives Affect MmpL3 in Mycobacterium tuberculosis

2019 ◽  
Vol 63 (10) ◽  
Author(s):  
Małgorzata Korycka-Machała ◽  
Albertus Viljoen ◽  
Jakub Pawełczyk ◽  
Paulina Borówka ◽  
Bożena Dziadek ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Mycolic Acid ◽  
Cross Resistance ◽  
Metabolic Labeling ◽  
Drug Resistant ◽  
Content Type ◽  
Antitubercular Drugs ◽  
Imidazole Derivatives ◽  
Trehalose Dimycolate

ABSTRACT 1H-benzo[d]imidazole derivatives exhibit antitubercular activity in vitro at a nanomolar range of concentrations and are not toxic to human cells, but their mode of action remains unknown. Here, we showed that these compounds are active against intracellular Mycobacterium tuberculosis. To identify their target, we selected drug-resistant M. tuberculosis mutants and then used whole-genome sequencing to unravel mutations in the essential mmpL3 gene, which encodes the integral membrane protein that catalyzes the export of trehalose monomycolate, a precursor of the mycobacterial outer membrane component trehalose dimycolate (TDM), as well as mycolic acids bound to arabinogalactan. The drug-resistant phenotype was also observed in the parental strain overexpressing the mmpL3 alleles carrying the mutations identified in the resistors. However, no cross-resistance was observed between 1H-benzo[d]imidazole derivatives and SQ109, another MmpL3 inhibitor, or other first-line antitubercular drugs. Metabolic labeling and quantitative thin-layer chromatography (TLC) analysis of radiolabeled lipids from M. tuberculosis cultures treated with the benzoimidazoles indicated an inhibition of trehalose dimycolate (TDM) synthesis, as well as reduced levels of mycolylated arabinogalactan, in agreement with the inhibition of MmpL3 activity. Overall, this study emphasizes the pronounced activity of 1H-benzo[d]imidazole derivatives in interfering with mycolic acid metabolism and their potential for therapeutic application in the fight against tuberculosis.

scholarly journals Genetic Diversity of Multi- and Extensively Drug-Resistant Mycobacterium tuberculosis Isolates in the Capital of Iran, Revealed by Whole-Genome Sequencing

2018 ◽  
Vol 57 (1) ◽  
Author(s):  
Farzam Vaziri ◽  
Thomas A. Kohl ◽  
Hasan Ghajavand ◽  
Mansour Kargarpour Kamakoli ◽  
Matthias Merker ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Whole Genome Sequencing ◽  
Genome Sequencing ◽  
Whole Genome ◽  
Drug Resistant ◽  
Content Type ◽  
Extensively Drug Resistant ◽  
Recent Transmission ◽  
Mdr Tb ◽  
Lineage 2

ABSTRACT The emergence and spread of multidrug resistant (MDR) Mycobacterium tuberculosis complex (MTBC) strains is a critical global health problem. Between 2014 and 2018, 606 MTBC strains were isolated from 13,892 suspected pulmonary tuberculosis (TB) patients in Tehran, Iran, including 16 (2.6%) MDR-TB cases. A combination of phenotypic and genotypic methods (whole-genome sequencing) was employed for the identification of additional drug resistances and strain-to-strain genetic distances as a marker for recent transmission events. MDR and extensively drug-resistant (XDR) TB cases were almost exclusively infected by lineage 2/Beijing strains (14/16, P < 0.001). We further showed that recent transmission and/or recent introduction of lineage 2/Beijing strains contribute to high XDR-TB rates among all MDR-TB cases and should be considered an emerging threat for TB control in Tehran. In addition, the extensive pre-existing drug resistance profiles of MDR/XDR strains will further challenge TB diagnostics in the region.

scholarly journals Analysis of drug resistance and mutation profiles in Mycobacterium tuberculosis isolates in a surveillance site in Beijing, China

2021 ◽  
Vol 49 (1) ◽  
pp. 030006052098493
Author(s):  
Jie Zhang ◽  
Yixuan Ren ◽  
Liping Pan ◽  
Junli Yi ◽  
Tong Guan ◽  
...  
Keyword(s):  
Drug Resistance ◽  
Mycobacterium Tuberculosis ◽  
Drug Testing ◽  
Multidrug Resistant ◽  
High Rate ◽  
Drug Resistant ◽  
Surveillance Site ◽  
Mdr Tb ◽  
Previously Treated Patients ◽  
Beijing China

Objective This study analyzed drug resistance and mutations profiles in Mycobacterium tuberculosis isolates in a surveillance site in Huairou District, Beijing, China. Methods The proportion method was used to assess drug resistance profiles for four first-line and seven second-line anti-tuberculosis (TB) drugs. Molecular line probe assays were used for the rapid detection of resistance to rifampicin (RIF) and isoniazid (INH). Results Among 235 strains of M. tuberculosis, 79 (33.6%) isolates were resistant to one or more drugs. The isolates included 18 monoresistant (7.7%), 19 polyresistant (8.1%), 28 RIF-resistant (11.9%), 24 multidrug-resistant (MDR) (10.2%), 7 pre-extensively drug-resistant (XDR, 3.0%), and 2 XDR strains (0.9%). A higher rate of MDR-TB was detected among previously treated patients than among patients with newly diagnosed TB (34.5% vs. 6.8%). The majority (62.5%) of RIF-resistant isolates exhibited a mutation at S531L in the DNA-dependent RNA polymerase gene. Meanwhile, 62.9% of INH-resistant isolates carried a mutation at S315T1 in the katG gene. Conclusion Our results confirmed the high rate of drug-resistant TB, especially MDR-TB, in Huairou District, Beijing, China. Therefore, detailed drug testing is crucial in the evaluation of MDR-TB treatment.

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