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PubMed1993

Pharmacology of the antimycobacterial drugs.

The management of MDR-TB requires that the clinician become familiar with the "second-line" antimycobacterial agents. These drugs are generally less potent and frequently more toxic than isoniazid and rifampin. Because they are less active, innovative dosing schedules may allow us to take advantage of the few strengths that they possess. This approach will require further research into the dose-response relationships for each agent. Based on our current knowledge of these drugs, practical guidelines for their use have been described. These guidelines include the gradual escalation of the oral doses of PAS, cycloserine, and ethionamide over several days, and the intravenous administration of streptomycin and capreomycin. Both ciprofloxacin and ofloxacin may be used for the treatment of MDR-TB, but data from clinical trials are currently lacking. Finally, because patients with AIDS appear to develop antimycobacterial drug malabsorption over the course of their HIV infection, therapeutic drug monitoring can be used to verify drug absorption in the individual patient. This approach may improve therapy for that patient and prevent the selection of additional drug resistance.

Antitubercular Agents↗
PubMed1987

Comparison of ribosomes and ribosomal proteins of sensitive and resistant mycobacteria.

Ribosomal proteins from Mycobacterium tuberculosis H37 Rv were analyzed by polyacrylamide gel electrophoresis and 73 proteins were separated. The 30S subunit consists of 26 proteins, the 50S subunit of 41 proteins, 6 proteins of the ribosomes could not be shown in the subunits. The molecular weights for the proteins of the 30S subunit are 9100-43500, and for the proteins of the 50S subunit 8000-46000. The sedimentation coefficient have values of 28.8S and 47.7S. It was demonstrated that streptomycin-resistant cells in comparison to sensitive cells have two additional 30S proteins. Moreover, the protein L34 has changed its position. Kanamycin- and capreomycin-resistant cells have also two additional proteins, but viomycin shows no changes.

Cross Reactions↗
PubMed1992

Spectrum of drugs against atypical mycobacteria: how valid is the current practice of drug susceptibility testing and the choice of drugs?

The in vitro activity of 13 drugs against 552 clinical isolates of atypical mycobacteria representing 12 species was performed in 7H11 agar medium at the National Reference Laboratory for Mycobacteria, using the 1% proportion method. All the species tested were resistant to isoniazid and pyrazinamide. In general, clofazimine and D-cycloserine showed the widest spectrum of activity except in the case of Mycobactrium fortuitum and M. chelonei which were resistant to both drugs, and the M. szulgai and M. terrae complex which was resistant to D-cycloserine. The next broad-spectrum drug was ethionamide, followed by ansamycin, rifampin, capreomycin, kanamycin, streptomycin and ethambutol. Among the fluoroquinolones, both ciprofloxacin and ofloxacin were active against M. xenopi, M. gordonae and M. fortuitum whereas M. kansasii and M. gastri were sensitive to ofloxacin only. When the species were listed in respect of the number of drugs to which they were susceptible (less than 10% of resistant strains), they were classified as follows; 7/13 drugs for M. kansasii, M. gastri and M. xenopi; 6/13 for M. gordonae; 5/13 for M. marinum; 3/13 for M. szulgai; 2/13 for M. fortuitum; 1/13 for the M. avium, M. scrofulaceum, M. simiae, and M. terrae complex, and none of the 13 in the case of M. chelonei. These results are discussed in relation to the multiple drug resistance of atypical mycobacteria. We conclude that the critical concentrations of drugs established for M. tuberculosis are not appropriate for atypical mycobacteria.

Antitubercular Agents↗
PubMed2024

Evaluating culture-free targeted next-generation sequencing for diagnosing drug-resistant tuberculosis: a multicentre clinical study of two end-to-end commercial workflows.

BACKGROUND: Drug-resistant tuberculosis remains a major obstacle in ending the global tuberculosis epidemic. Deployment of molecular tools for comprehensive drug resistance profiling is imperative for successful detection and characterisation of tuberculosis drug resistance. We aimed to assess the diagnostic accuracy of a new class of molecular diagnostics for drug-resistant tuberculosis. METHODS: We conducted a prospective, cross-sectional, multicentre clinical evaluation of the performance of two targeted next-generation sequencing (tNGS) assays for drug-resistant tuberculosis at reference laboratories in three countries (Georgia, India, and South Africa) to assess diagnostic accuracy and index test failure rates. Eligible participants were aged 18 years or older, with molecularly confirmed pulmonary tuberculosis, and at risk for rifampicin-resistant tuberculosis. Sensitivity and specificity for both tNGS index tests (GenoScreen Deeplex Myc-TB and Oxford Nanopore Technologies [ONT] Tuberculosis Drug Resistance Test) were calculated for rifampicin, isoniazid, fluoroquinolones (moxifloxacin, levofloxacin), second line-injectables (amikacin, kanamycin, capreomycin), pyrazinamide, bedaquiline, linezolid, clofazimine, ethambutol, and streptomycin against a composite reference standard of phenotypic drug susceptibility testing and whole-genome sequencing. FINDINGS: Between April 1, 2021, and June 30, 2022, 832 individuals were invited to participate in the study, of whom 720 were included in the final analysis (212, 376, and 132 participants in Georgia, India, and South Africa, respectively). Of 720 clinical sediment samples evaluated, 658 (91%) and 684 (95%) produced complete or partial results on the GenoScreen and ONT tNGS workflows, respectively, with 593 (96%) and 603 (98%) of 616 smear-positive samples producing tNGS sequence data. Both workflows had sensitivities and specificities of more than 95% for rifampicin and isoniazid, and high accuracy for fluoroquinolones (sensitivity approximately ≥94%) and second line-injectables (sensitivity 80%) compared with the composite reference standard. Importantly, these assays also detected mutations associated with resistance to critical new and repurposed drugs (bedaquiline, linezolid) not currently detectable by any other WHO-recommended rapid diagnostics on the market. We note that the current format of assays have low sensitivity (≤50%) for linezolid and more work on mutations associated with drug resistance is needed. INTERPRETATION: This multicentre evaluation demonstrates that culture-free tNGS can provide accurate sequencing results for detection and characterisation of drug resistance from Mycobacterium tuberculosis clinical sediment samples for timely, comprehensive profiling of drug-resistant tuberculosis. FUNDING: Unitaid.

Humans↗
PubMed2002

Comparative study on the use of solid media: Löwenstein-Jensen and Ogawa in the determination of anti-tuberculosis drug susceptibility.

A total of 53 Mycobacterium tuberculosis strains were examined to evaluate the consistency of susceptibility tests using two solid egg-based culture media, Ogawa and Löwenstein-Jensen (L-J), in order to assess the inter-media differences in susceptibility tests for anti-tuberculosis drugs. Fifty-three M. tuberculosis strains were tested for resistance against isoniazid, rifampicin, ethambutol, streptomycin and other alternative drugs on L-J and Ogawa media. Data from isoniazid (INH) revealed 31 strains were susceptible and 11 strains were resistant in L-J medium. On the other hand, one strain showed resistance against INH only on Ogawa medium. This gave a consistency ratio of 97.7% for INH between these two media. Similarly, the methods had 100% compatibility with rifampicin, 86.0% with ethambutol and 88.4% with streptomycin. When 41 of these strains were tested against ethionamide, enviomycin, para -aminosalicylate, sparfloxacin, and levofloxacin, the same numbers of strains were shown to be resistant or susceptible to L-J and Ogawa media. Similarly, the consistency ratio was 75.6% for kanamycin and 61.0% for capreomycin. There are inconsistencies between results obtained with L-J and Ogawa media, Ogawa medium had a tendency to produce results that indicated more resistant strains. However, the differences were statistically negligible.

Antitubercular Agents↗
PubMed1990

Immunosuppression by gramicidin S of experimental autoimmune uveoretinitis, pinealitis and autoimmune encephalomyelitis.

Using an in vitro lymphocyte proliferation assay we screened several cyclic peptide antibiotics (bacitracin, oleandomycin, capreomycin, colistin, virginiamycin, and gramicidin S) for their immunosuppressive activity. Gramicidin S (GrS) was found to inhibit [3H]-thymidine incorporation into concanavalin A-stimulated and E coli lipopolysaccharide-stimulated lymphocytes. In vivo studies, experimental autoimmune uveoretinitis (EAU) and pinealitis were induced in female Lewis rats by immunization with bovine S-antigen and experimental autoimmune encephalomyelitis (EAE) were induced by immunization of rats with rat brain homogenates. GrS suppressed the onset of these inflammatory diseases at nontoxic concentrations. Evidence was obtained that GrS inhibits [3H]-thymidine incorporation into lymphocytes by preventing transport of the compound across the membrane. Since GrS binds to various cell membranes, GrS would suppress the proliferation of not only lymphocytes but also of other immune cells by modifying cell membrane properties. The present study indicates that a search for compounds which cause proper cell membrane modification should be a worthwhile strategy for development of immunosuppressive drugs.

Amino Acid Sequence↗
PubMed1999

Activity of 16 antimicrobial agents against drug-resistant strains of Mycobacterium tuberculosis.

The in vitro activity of 16 antimicrobial agents against 46 drug-resistant strains of Mycobacterium tuberculosis recently isolated from Italian patients was determined. As for first-line antituberculosis drugs, while isoniazid was ineffective against all the strains tested, resistance to streptomycin, rifampicin, pyrazinamide, and ethambutol was 80.4%, 71.7%, 39.1%, and 8.7%, respectively. Among second-line antituberculous drugs, resistance to ciprofloxacin, ofloxacin, and sparfloxacin and to amikacin and kanamycin was around 20%. About 10% of the strains were resistant to capreomycin and cycloserine and 4.3% were resistant to ethionamide; no strain was found to be resistant to thiacetazone, para-aminosalicylic acid, and viomycin. Although all strains displayed a rather continuous distribution of minimal inhibitory concentrations (MICs), a bimodal distribution was observed for rifampicin, amikacin, and kanamicin, with very high MIC values for resistant strains; relatively low MICs were found for fluoroquinolone-resistant strains. Among the small number of strains resistant to second-line agents, low resistant levels were observed. Restriction fragment length polymorphism analysis showed few strain clusters with resistance to first-line antituberculous drugs and aminoglycosides, fluoroquinolones, or both. Altogether, these results showed that second-line agents were still active against the isoniazid-resistant and multiply first-line resistant strains tested, with none or low resistance levels; these observations can be of importance for the treatment of multidrug-resistant tuberculosis in Italy.

Antitubercular Agents↗
PubMed1996

Prevention and treatment of drug-resistant tuberculosis.

Therapy recommended for preventing and treating drug-resistant tuberculosis is discussed. Drug-resistant strains of Mycobacterium tuberculosis can be transmitted by an infected individual, or resistance can be acquired during therapy for drug-susceptible disease. At least until susceptibility test data are available, the recommended initial treatment for tuberculosis consists of isoniazid, rifampin, pyrazinamide, and either ethambutol or streptomycin. Resistance has led to variations on this regimen that sometimes include more toxic alternative drugs, including ethionamide, aminosalicylic acid, cycloserine, and capreomycin, as well as ciprofloxacin and ofloxacin. Drug regimens used for retreatment usually include the alternative drugs. Success in treating drug-resistant tuberculosis varies. Therapy to prevent resistant tuberculosis is recommended for any individual with a positive skin-test result and any of the following: infection with the human immunodeficiency virus (HIV), close contact with a newly diagnosed patient, recent conversion to a positive skin-test result, or a predisposing medical condition. Although isoniazid is the only drug with FDA-approved labeling for use as prophylaxis in patients with latent tuberculosis, the Centers for Disease Control and Prevention suggests several two-drug combinations. Fluoroquinolones are recommended for both treatment and prevention. Patient compliance and HIV infection are special issues in managing resistant tuberculosis. Drug-resistant tuberculosis is a growing problem that must be addressed through appropriate prophylactic and treatment measures.

AIDS-Related Opportunistic Infections↗
PubMed1998

A survey of drug-resistant Mycobacterium tuberculosis and its relationship to HIV infection.

OBJECTIVE: To compare the prevalence of drug-resistant tuberculosis disease among patients with and without HIV infection. DESIGN: An historical prospective evaluation of patients with culture-proven tuberculosis reported between January 1988 and December 1995. SETTING: A major metropolitan county public health department. PATIENTS: A total of 802 consecutive culture-positive tuberculosis patients were eligible for inclusion in the study. HIV serologic testing and drug susceptibilities were completed on 741 (92%) eligible patients. Of these patients, 646 tested HIV-negative and 95 (12.8%) tested HIV-positive. Patients not tested for HIV (n = 59) and without drug susceptibilities (n = 2) were excluded from the analyses. Outpatient management was based on a policy of universal directly observed therapy. MAIN OUTCOME MEASURES: Patient HIV status, initial drug resistance and acquired drug resistance. Isolates were characterized for resistance to isoniazid, rifampin, rifabutin, ethambutol, streptomycin, capreomycin, kanamycin and ethionamide. Determination of initial resistance was based on the first available susceptibility study and acquired resistance on subsequent susceptibility studies. RESULTS: Initial drug resistance was found in 55 (8.5%) HIV-negative patients and four (4.2%) HIV-positive patients. Acquired drug resistance occurred in five (0.8%) HIV-negative patients and one (1.1%) HIV-positive patient. These differences were not statistically significant. CONCLUSIONS: HIV infection is not a risk factor for drug-resistant tuberculosis. Increased drug resistance in HIV infected tuberculosis patients reflects a failure of tuberculosis control in the underlying population.

AIDS Serodiagnosis↗
PubMed1980

Studies on the cross-resistance of Mycobacterium tuberculosis, strain H37Rv, to aminoglycoside- and peptide-antibiotics.

Various phenotypes of the resistance to aminoglycoside- and peptide-antibodies of Mycobacterium tuberculosis strain H37Rv were produced by single- and/or two-step selection of the parent strain. Mutants obtained by single-step selection with antibiotics were classified into ten phenotypes; one of single resistance, two of triple resistance, three of quadruple resistance, and four of sextuple resistance. There were two kinds of sextuple resistance (high resistance to enviomycin, viomycin, capreomycin, kanamycin, lividomycin). One was isolated from the parent strain by single-step selection and could be eliminated by mutation to isoniazid resistance, the other was obtained by two-step selections and was not eliminated by mutation to isoniazid resistance. Interaction between mutation to streptomycin resistance and mutation to quadruple resistance (4R phenotype) was observed. Streptomycin resistance interfered with the formation of the 4R phenotype and produced a different phenotype, KR instead of the 4R phenotype. The existence of mutation of the 4R phenotype did not usually interfere with mutation to streptomycin resistance, but a small portion of the mutants with the 4R phenotype were altered in their phenotype from 4R to KR after addition of the mutation to streptomycin resistance. This effect of the mutation to streptomycin resistance was not observed in mutants which already had a mutation to klR phenotype (mutation to low concentrations of kanamycin only).

Aminoglycosides↗
PubMed1984

In vitro antimicrobial susceptibility of a Mycobacterium sp. isolated from patients with Crohn's disease.

The in vitro susceptibility of three strains of an unclassified Mycobacterium sp., isolated from three patients with Crohn's disease, to 23 antimicrobial agents was determined by a modified broth dilution method with 7H9 broth containing oleic acid-albumin-dextrose-catalase, Tween 80, and mycobactin J. All three strains were susceptible to streptomycin, viomycin, rifampin, clofazimine, cefazolin, amikacin, and kanamycin and resistant to p-aminosalicylic acid, cycloserine, 2-thiophenecarboxylic acid hydrazide, trimethoprim, diaminodiphenylsulfone, sulfamethoxazole, sulfadimethoxine, polymyxin B, metronidazole, neomycin, and carbenicillin. Variable results between strains were encountered with ethambutol, ethionamide, capreomycin, amoxicillin, and cephalothin.

Adolescent↗
PubMed1993

Therapy of multidrug-resistant tuberculosis: lessons from studies with mice.

The activities of antituberculosis agents were evaluated in a murine tuberculosis model using a drug-resistant isolate. A multidrug-resistant clinical isolate from a recent outbreak of tuberculosis in the New York State correctional system was used for infection. Approximately 10(7) viable Mycobacterium tuberculosis ATCC 49967 (strain CNL) organisms were given intravenously to 4-week-old female outbred mice. Treatment was started 1 day after infection and given for 4 weeks. Spleens and lungs were homogenized, and viable cell counts were determined. Statistical analysis indicated that ethionamide, sparfloxacin, ofloxacin, capreomycin, clarithromycin, and clofazimine are active in the murine test system with this multidrug-resistant tuberculosis isolate. Sparfloxacin is the most active quinolone. Despite in vitro resistance, isoniazid has moderate activity. In vitro susceptibility data coupled with evaluation of agents against drug-resistant isolates in the murine system should provide information necessary to design clinical trials for treatment of infections with these organisms.

Animals↗
PubMed1973

Antimicrobial susceptibility of Nocardia species.

Variations of inoculum, pH, and composition of the agar assay media each influenced the in vitro susceptibility of Nocardia organisms (30 strains of N. asteroides and 6 strains of N. brasiliensis) to one or more antimicrobial agents incorporated in solid agar and incubated at 37 C for 48 h. Among newer antimicrobial agents, doxycycline, minocycline, and 7-halogenated lincomycin analogues exhibited the most consistent in vitro inhibitory activity; among older agents, viomycin, erythromycin, and streptomycin exhibited more selective in vitro inhibitory activity, but against a significant percentage of strains tested. N. brasiliensis was conspicuously more susceptible to erythromycin and gentamicin and more resistant to viomycin and capreomycin than was N. asteroides. Cycloserine demonstrated no significant in vitro inhibitory activity when assayed in appropriately augmented alanine-free medium. The outstanding feature of this in vitro survey was the marked variation in susceptibility among individual strains of N. asteroides, which variation deserves more careful consideration in future clinical or experimental therapeutic trials.

Anti-Bacterial Agents↗
PubMed1998

Green fluorescent protein reporter microplate assay for high-throughput screening of compounds against Mycobacterium tuberculosis.

An optimal assay for high-throughput screening for new antituberculosis agents would combine the microplate format and low cost of firefly luciferase reporter assays and redox dyes with the ease of kinetic monitoring inherent in the BACTEC system. The green fluorescent protein (GFP) of the jellyfish Aequorea victoria is a useful reporter molecule which requires neither substrates nor cofactors due to the intrinsically fluorescent nature of the protein. The gene encoding a red-shifted, higher-intensity GFP variant was introduced by electroporation into Mycobacterium tuberculosis H37Ra and M. tuberculosis H37Rv on expression vector pFPV2. A microplate-based fluorescence assay (GFP microplate assay [GFPMA]) was developed and evaluated by determining the MICs of existing antimycobacterial agents. The MICs of isoniazid, rifampin, ethambutol, streptomycin, amikacin, ofloxacin, ethionamide, thiacetazone, and capreomycin, but not cycloserine, determined by GFPMA were within 1 log2 dilution of those determined with the BACTEC 460 system and were available in 7 days. Equivalent MICs of antituberculosis agents in the BACTEC 460 system for both the reporter and parent strains suggested that introduction of pFPV2 did not influence drug susceptibility, in general. GFPMA provides a unique tool with which the dynamic response of M. tuberculosis to the existing and potential antituberculosis agents can easily, rapidly, and inexpensively be monitored.

Anti-Infective Agents↗
PubMed2003

Activities of moxifloxacin alone and in combination with other antimicrobial agents against multidrug-resistant Mycobacterium tuberculosis infection in BALB/c mice.

The activity of moxifloxacin was enhanced by the addition of ethionamide but not by that of cycloserine, thiacetazone, capreomycin, para-aminosalicylic acid, or linezolid in BALB/c mice infected with a strain of Mycobacterium tuberculosis resistant to isoniazid, rifampin, and six other drugs. These observations are important for the therapy of multidrug-resistant tuberculosis.

Animals↗
PubMed2003

Resazurin microtiter assay plate testing of Mycobacterium tuberculosis susceptibilities to second-line drugs: rapid, simple, and inexpensive method.

The emergence of multidrug-resistant tuberculosis calls for new, rapid drug susceptibility tests. We have tested 150 Mycobacterium tuberculosis isolates against the second-line drugs ethionamide, kanamycin, capreomycin, ofloxacin, and para-aminosalicylic acid by the colorimetric resazurin microtiter assay and the proportion method. By visual reading, MICs were obtained after 8 days. A very good correlation between results by the colorimetric resazurin microtiter assay and the proportion method was obtained. The colorimetric resazurin microtiter assay is inexpensive, rapid, and simple to perform, and implementation of the assay is feasible for low-resource countries.

Antitubercular Agents↗
PubMed1976

Altered ribosomes in antibiotic-resistant mutants of Mycobacterium smegmatis.

Two alleles for viomycin-capreomycin resistance (vic) in Mycobacterium smegmatis affect ribosome structures. One (vicA) affects a component of 50S subunits and the other (vicB) affects a component of 30S subunits. The locus for neomycin-kanamycin resistance (nek), which is linked to vicA and vicB, affects a component of 30S subunits. Although the erythromycin resistance locus (ery) is linked to vic and nek, no ribosomal alterations could be detected. Mutations at the streptomycin locus (str) not linked to vic and nek caused alterations of 30S subunits.

Anti-Bacterial Agents↗
PubMed2005

A mutation in the decoding center of Thermus thermophilus 16S rRNA suggests a novel mechanism of streptomycin resistance.

A spontaneous kanamycin resistance and capreomycin resistance mutation, A1408G, in the decoding center of 16S rRNA, was identified in the extreme thermophile Thermus thermophilus. Unexpectedly, this mutation also confers resistance to streptomycin. We propose a novel mechanism of streptomycin resistance by which A1408G influences conformational changes in 16S rRNA during tRNA selection.

Anti-Bacterial Agents↗