Lipoprotein Processing Is Essential for Resistance of
            <i>Mycobacterium tuberculosis</i>
            to Malachite Green

Banaei, Niaz; Kincaid, Eleanor Z.; Lin, S.-Y. Grace; Desmond, Edward; Jacobs, William R.; Ernst, Joel D.

doi:10.1128/aac.00647-09

Cited by 30 publications

(37 citation statements)

References 20 publications

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“…Second, increased permeability of cells exposed to cell wall targeting antibiotics could facilitate the access of malachite green to its molecular target(s). Consistent with this interpretation, two recent reports demonstrated that mutant strains of mycobacteria with increased cell wall permeability are hypersensitive to malachite green (3,4). Third, antibiotic-mediated killing has been linked to the production of reactive oxygen species (ROS) in Gram-negative and Gram-positive bacteria (7), and malachite green has been shown to trigger ROS production via an unknown mechanism (18).…”

Section: Discussionsupporting

confidence: 50%

Malachite Green Interferes with Postantibiotic Recovery of Mycobacteria

Gelman

McKinney

Dhar

2012

Antimicrob Agents Chemother

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The genus Mycobacterium comprises slow-growing species with generation times ranging from hours to weeks. The protracted incubation time before colonies appear on solid culture medium can result in overgrowth by faster-growing microorganisms. To prevent contamination, the solid media used in laboratories and clinics for cultivation of mycobacteria contain the arylmethane compound malachite green, which has broad-spectrum antimicrobial activity. Malachite green has no impact on the plating efficiency of mycobacteria when cells are grown under normal conditions. However, we found that malachite green interfered with colony formation when bacteria were preexposed to antibiotics targeting cell wall biogenesis (isoniazid, ethionamide, ethambutol). This inhibitory effect of malachite green was not observed when bacteria were preexposed to antibiotics targeting cellular processes other than cell wall biogenesis (rifampin, moxifloxacin, streptomycin). Sputum specimens from tuberculosis patients are routinely evaluated on solid culture medium containing high concentrations of malachite green. This practice could lead to underestimation of bacterial loads and overestimation of chemotherapeutic efficacy. Mycobacterium tuberculosis grows very slowly, with a population doubling time of ϳ22 h. Consequently, detection of M. tuberculosis by outgrowth of CFU on solid culture medium requires weeks or months (17). Despite being labor-intensive and time-consuming, enumeration of CFU by plating sputum cultures on solid medium continues to be the "gold standard" for evaluating the effectiveness of antituberculosis chemotherapy (6,19). Contamination by faster-growing microorganisms is prevented by supplementing the medium with compounds that suppress the growth of common species in the human flora.In clinical practice, sputum samples from tuberculosis patients are routinely plated on Lowenstein-Jensen or Middlebrook 7H11 agar. These media contain high concentrations of malachite green, a diamino-triphenylmethane dye with broad-spectrum antimicrobial activity against Gram-positive and Gram-negative bacteria (1). The mechanistic basis of malachite green's antimicrobial activity is not understood, nor is it clear why mycobacteria can survive and replicate in the presence of high concentrations of this compound (1,12). This paper shows that malachite green interferes with the recovery of mycobacteria on solid culture medium following exposure to certain antibiotics. This inhibitory effect was specific to antibiotics that target cell wall biogenesis (isoniazid, ethionamide, ethambutol) and was not observed with antibiotics that target other cellular processes (rifampin, moxifloxacin, streptomycin). These observations may have implications for clinical practices because underestimation of bacterial loads in patients undergoing chemotherapy could result in overestimation of therapeutic efficacy. MATERIALS AND METHODSBacteria and culture conditions. Wild-type Mycobacterium smegmatis (strain mc 2 155) and M. tuberculosis (strain Erdman) were s...

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Section: Discussionsupporting

confidence: 50%

Malachite Green Interferes with Postantibiotic Recovery of Mycobacteria

Gelman

McKinney

Dhar

2012

Antimicrob Agents Chemother

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“…3B). Similar diminished ability of Mtb mutants to form colonies on solid medium has been previously attributed to disruption of cell wall integrity, resulting in an increased susceptibility to chaotropic or bacteriostatic agents (such as Malachite green, ordinarily incorporated in solid media for mycobacterial culture) (26). However, the persistence of the deficiency to form colony on Malachite green-free agar medium (Fig.…”

Section: Initiation and Elongation Of The Fasc Mannosyl Appendages Rementioning

confidence: 81%

Bacterial protein-O-mannosylating enzyme is crucial for virulence ofMycobacterium tuberculosis

Liu

Tonini

Malaga

et al. 2013

Proc. Natl. Acad. Sci. U.S.A.

107

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A posttranslational protein O-mannosylation process resembling that found in fungi and animals has been reported in the major human pathogen Mycobacterium tuberculosis (Mtb) and related actinobacteria. However, the role and incidence of this process, which is essential in eukaryotes, have never been explored in Mtb. We thus analyzed the impact of interrupting O-mannosylation in the nonpathogenic saprophyte Mycobacterium smegmatis and in the human pathogen Mtb by inactivating the respective putative protein mannosyl transferase genes Msmeg_5447 and Rv1002c. Loss of protein O-mannosylation in both mutant strains was unambiguously demonstrated by efficient mass spectrometry-based glycoproteomics analysis. Unexpectedly, although the M. smegmatis phenotype was unaffected by the lack of manno-proteins, the Mtb mutant had severely impacted growth in vitro and in cellulo associated with a strong attenuation of its pathogenicity in immunocompromised mice. These data are unique in providing evidence of the biological significance of protein O-mannosylation in mycobacteria and demonstrate the crucial contribution of this protein posttranslational modification to Mtb virulence in the host.protein glycosylation | O-glycosylation | proteomics

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“…The lspA mutant showed increased sensitivity to first-line antituberculosis drugs and was essential for resistance to malachite green (Banaei et al, 2009). Existing information for this gene is limited and no information is available regarding its regulation.…”

Section: Introductionmentioning

confidence: 98%