Antimicrobial Efflux Pumps and Mycobacterium tuberculosis Drug Tolerance: Evolutionary Considerations

Szumowski, John D.; Adams, Kristin N.; Edelstein, Paul H.; Ramakrishnan, Lalita

doi:10.1007/82_2012_300

Cited by 56 publications

(68 citation statements)

References 176 publications

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“…Importantly, we find that these same pumps also mediate tolerance to antitubercular drugs, presumably by efflux mechanisms (Fig. 5) (Adams et al 2011;Szumowski et al 2013). As predicted by this finding, the most actively growing bacteria within macrophages are also the most drug tolerant.…”

Section: What Does Not Kill Them Makes Them Stronger: Macrophages Maksupporting

confidence: 75%

“…In work that began with the zebrafish but moved quickly into a human macrophage cell line, we realized the importance of prior findings that M. tuberculosis expresses efflux pumps that are induced upon macrophage entry, several of which enable bacterial growth in macrophages and as such comprise host-induced virulence factors (Schnappinger et al 2003;Rengarajan et al 2005;Adams et al 2011;Szumowski et al 2013). Importantly, we find that these same pumps also mediate tolerance to antitubercular drugs, presumably by efflux mechanisms (Fig.…”

Section: What Does Not Kill Them Makes Them Stronger: Macrophages Makmentioning

confidence: 68%

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The Zebrafish Guide to Tuberculosis Immunity and Treatment

Ramakrishnan

2013

Cold Spring Harbor Symposia on Quantitative Biology

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During the past 12 years, we have developed the zebrafish as a model for the study of tuberculosis pathogenesis and immunology. We have taken advantage of the optical transparency and the genetic and pharmacological tractability of the developing zebrafish to monitor infection in real time. Detailed information about the sequential interactions among the host and the pathogen, the cell types, and the molecules involved has yielded surprising insights into this ancient disease. We have identified a number of host evasion strategies deployed by pathogenic mycobacteria as well as host responses that provide broad insights into host immunity. Many of these discoveries have relevance to human tuberculosis and suggest new therapeutic avenues for tuberculosis as well as other inflammatory diseases.

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Section: What Does Not Kill Them Makes Them Stronger: Macrophages Maksupporting

confidence: 75%

Section: What Does Not Kill Them Makes Them Stronger: Macrophages Makmentioning

confidence: 68%

The Zebrafish Guide to Tuberculosis Immunity and Treatment

Ramakrishnan

2013

Cold Spring Harbor Symposia on Quantitative Biology

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“…Estos resultados difieren de trabajos previos publicados en los cuales solo se reporta la presencia de perfiles de mutación en la posición S315T en aislados resistentes 7,19,20,22,31 , el 25,5% de aislados fenotipicamente resistentes que no presentaron la mutación estudiada, pudo deberse a la presencia de polimorfismos en regiones diferente a la posición S315T [15][16][17][18][19][20] en el gen katG u otros genes que aporten resistencia a Isoniacida 9-14 indiciendo a la sobreexpresión que superen el poder inhibitorio de INH 26,31 ; otra causa puede ser la presencia de mecanismos de resistencia diferente a las mutaciones tales como mecanismos de barrera (reducción de la permeabilidad 32-33 y bomba de eflujo [34][35][36][37] ). En cuanto a la presencia de un aislado sensible con la mutación S315T, posiblemente se deba a que el límite de la detección por PCR-RFLP empleada se encuentre cercano al 1% de resistentes que basado en el método de las proporciones pasa imperceptible y se considera como sensible, tambien podría deberse a una inadecuada identificación del perfil de resistencia mediante la prueba de las proporciones en el aislado sensible, y/o contaminación del aislado después de su conservación.…”

Section: Nuestros Resultados Mostraron Que Al Emplear Las Enzimasunclassified

Determinación de la mutación S315T del gen katG en aislados resistentes a Isoniacida de Mycobacterium tuberculosis mediante PCR-RFLP

Sánchez-Domínguez

Nicola-Salas

Morey-León

2018

Infect

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Cómo citar este artículo: J. Sánchez-Domínguez, et al. Determinación de la mutación S315T del gen katG en aislados resistentes a Isoniacida de Mycobacterium tuberculosis mediante PCR-RFLP. Infectio 2018; 22(4): 178-184 ResumenObjetivo: determinar la mutación S315T del gen katG en aislados de Mycobacterium tuberculosis resistentes a isoniacida mediante la técnica PCR-RFLP. Materiales y métodos: A partir de 68 aislados de Mycobacterium tuberculosis se realizó el análisis de polimorfismo en productos de amplificación de 1054 y 630 pb que contenían la mutación S315T del gen katG mediante PCR-RFLP empleando las enzimas de restricción MspI y SatI. Mediante SPSS se determinó sensibilidad, especificidad, valores predictivo positivo y negativo, coeficientes de probabilidad positivo y negativo. Resultados: El 74,46% de aislados fenotípicamente resistentes y 4,76% fenotípicamente sensibles presentaron la mutación del gen katG S315T. La PCR-RFLP para S315T del gen katG presentó 85,4% de sensibilidad y 95,2% de especificidad con MspI y 85,4% de sensibilidad y 94,4% de especificidad con SatI. Discusión: La PCR-RFLP tiene una alta capacidad resolutiva que depende de la enzima que se emplee como se observó en estudios previos. La presencia de la mutación S315T en pacientes vírgenes al tratamiento sugiere la circulación de aislados resistentes a Isoniacida. Conclusión: La PCR-RFLP resultó una alternativa válida y rápida para el diagnóstico de la resistencia a isoniacida, mediante la detección de la mutación S315T del gen katG en comparación con el método convencional de las proporciones.Palabras Clave: Mutaciones, Polimorfismo, Isoniacida, PCR-RFLP, Mycobacterium tuberculosis, katG. Determination of S315T mutation within the katG gene in isoniazid-resistant Mycobacterium tuberculosis isolate by PCR-RFLP AbstractObjetive: To determine the S315T mutation of the katG gene in Mycobacterium tuberculosis isoniazid-resistant isolates by PCR-RFLP. Materials and Methods: Polymorphism analysis of 1054 and 630 bp products containing the S315T mutation of the katG gene was performed by PCR-RFLP using the MspI and SatI restriction enzymes from 68 Mycobacterium tuberculosis isolates. Sensitivity, specificity, positive and negative predictive values, positive and negative likelihood ratio were determined using SPSS. Results: 74.46% of isoniazid-resistant and 4.76% of isoniazid-sensitive isolates showed the S315T mutation in katG gene. The PCR-RFLP for S315T of the katG gene had 85.4% sensitivity and 95.2% specificity with MspI and 85.4% sensitivity and 94.4% specificity with SatI. Discussion: The PCR-RFLP has a high resolutive capacity that depends on the enzyme that is used as it was observed in previous studies. The presence of the S315T mutation in treatment-naive patients suggests the circulation of isolates resistant to isoniazid. Conclusion: PCR-RFLP is a valid and rapid alternative for the diagnosis of isoniazid resistance, by detection of S315T mutation in the katG gene compared to the conventional method of proportions.

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“…Based on the role of MarR in regulating efflux pumps in E. coli, we tested the efflux pump inhibitors verapamil, chlorpromazine, and carbonyl cyanide 3-chlorophenyl hydrazone (CCCP) (30)(31)(32)(33). Verapamil and chlorpromazine compounds had modest potentiating effects on the MIC of MP-III-71 to wild-type H37Rv and complemented mutant 1; however, they did not alter the MIC of the resistant strain, mutant 1 (see Table S4 in the supplemental material).…”

Section: Resultsmentioning

confidence: 99%

Mutation of Rv2887 , a marR -Like Gene, Confers Mycobacterium tuberculosis Resistance to an Imidazopyridine-Based Agent

Winglee

Lun

Pieroni

et al. 2015

Antimicrob Agents Chemother

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Drug resistance is a major problem in Mycobacterium tuberculosis control, and it is critical to identify novel drug targets and new antimycobacterial compounds. We have previously identified an imidazo[1,2-a]pyridine-4-carbonitrile-based agent, MP-III-71, with strong activity against M. tuberculosis. In this study, we evaluated mechanisms of resistance to MP-III-71. We derived three independent M. tuberculosis mutants resistant to MP-III-71 and conducted whole-genome sequencing of these mutants. Loss-of-function mutations in Rv2887 were common to all three MP-III-71-resistant mutants, and we confirmed the role of Rv2887 as a gene required for MP-III-71 susceptibility using complementation. The Rv2887 protein was previously unannotated, but domain and homology analyses suggested it to be a transcriptional regulator in the MarR (multiple antibiotic resistance repressor) family, a group of proteins first identified in Escherichia coli to negatively regulate efflux pumps and other mechanisms of multidrug resistance. We found that two efflux pump inhibitors, verapamil and chlorpromazine, potentiate the action of MP-III-71 and that mutation of Rv2887 abrogates their activity. We also used transcriptome sequencing (RNA-seq) to identify genes which are differentially expressed in the presence and absence of a functional Rv2887 protein. We found that genes involved in benzoquinone and menaquinone biosynthesis were repressed by functional Rv2887. Thus, inactivating mutations of Rv2887, encoding a putative MarR-like transcriptional regulator, confer resistance to MP-III-71, an effective antimycobacterial compound that shows no cross-resistance to existing antituberculosis drugs. The mechanism of resistance of M. tuberculosis Rv2887 mutants may involve efflux pump upregulation and also drug methylation.T uberculosis (TB) is a devastating disease that infects one-third of the world's population and killed 1.5 million people in 2013 (1). TB is caused by Mycobacterium tuberculosis and is challenging and time-consuming to treat. Standard TB treatment is currently 6 months long and involves a 2-month intensive phase consisting of treatment with four antibiotics (isoniazid, rifampin, ethambutol, and pyrazinamide) followed by a 4-month continuation phase (treatment with isoniazid and rifampin only). However, despite this combination therapy, drug resistance is on the rise, and in 2013, there were an estimated 480,000 cases of multidrug-resistant (MDR) TB, which is defined as TB caused by bacteria that are resistant to at least isoniazid and rifampin. These cases require up to 2 years of treatment, but drug resistance is developing even under these conditions, and in 2014, 9% of MDR TB cases were extensively drug resistant (XDR), meaning that they were also resistant to isoniazid, rifampin, a fluoroquinolone, and an injectable anti-TB drug (typically an aminoglycoside) (1). Thus, there is a great need both for new antibiotics to treat M. tuberculosis and for new drug targets that avoid cross-resistance to currently used therapies.O...

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Antimicrobial Efflux Pumps and Mycobacterium tuberculosis Drug Tolerance: Evolutionary Considerations

Cited by 56 publications

References 176 publications

The Zebrafish Guide to Tuberculosis Immunity and Treatment

The Zebrafish Guide to Tuberculosis Immunity and Treatment

Determinación de la mutación S315T del gen katG en aislados resistentes a Isoniacida de Mycobacterium tuberculosis mediante PCR-RFLP

Mutation of Rv2887 , a marR -Like Gene, Confers Mycobacterium tuberculosis Resistance to an Imidazopyridine-Based Agent

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