Tween 80 induces a carbon flux rerouting in Mycobacterium tuberculosis

2020 ◽  
Vol 170 ◽  
pp. 105795
Author(s):  
Ray-Dean Pietersen ◽  
Ilse du Preez ◽  
Du Toit Loots ◽  
Mari van Reenen ◽  
Derylize Beukes ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Carbon Flux ◽  
Tween 80

scholarly journals Glucan Is a Component of the Mycobacterium tuberculosis Surface That Is Expressed In Vitro and In Vivo

2002 ◽  
Vol 70 (5) ◽  
pp. 2566-2575 ◽  
Author(s):  
J. Reid Schwebach ◽  
Aharona Glatman-Freedman ◽  
Leslie Gunther-Cummins ◽  
Zhongdong Dai ◽  
John B. Robbins ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Cell Envelope ◽  
Tween 80 ◽  
Enzyme Linked Immunosorbent Assay ◽  
In Vitro Growth ◽  
Comprehensive Picture ◽  
The Media ◽  
Surface Polysaccharide

ABSTRACT The outermost layer of Mycobacterium tuberculosis is composed primarily of two polysaccharides, glucan (GC) and arabinomannan. To analyze the surface polysaccharide composition of M. tuberculosis, we generated a monoclonal antibody (MAb) that binds M. tuberculosis GC and is known as MAb 24c5. Immunofluorescence and whole-mount immunoelectron microscopy indicated that GC is on the outermost portion of the bacteria. M. tuberculosis strains Erdman and CDC 1551 were analyzed for their ability to bind MAb 24c5 after in vitro growth in media with and without the detergent Tween 80. MAb 24c5 bound to Erdman and CDC 1551 at all culture times with only slightly greater apparent affinity after extended culture in the absence of Tween 80, indicating that a stable amount of GC polysaccharide antigen is associated with the cell surface of M. tuberculosis. An enzyme-linked immunosorbent assay indicated that GC is antigenically similar to glycogen, and the amount of GC antigen increased in the media of M. tuberculosis cultures grown either with or without the detergent Tween 80. Other nontuberculosis mycobacteria have antigenically similar GCs on their surfaces after in vitro growth. Inoculation of mice with live bacilli but not inoculation with dead bacilli elicited a strong antibody response to GC consistent with production of this antigen in vivo. Our results provide a more comprehensive picture of the M. tuberculosis cell envelope and the conditions that allow expression of M. tuberculosis GC.

scholarly journals Expression of a Mycobacterium tuberculosis Arabinomannan Antigen In Vitro and In Vivo

2001 ◽  
Vol 69 (9) ◽  
pp. 5671-5678 ◽  
Author(s):  
J. Reid Schwebach ◽  
Arturo Casadevall ◽  
Rachel Schneerson ◽  
Zhongdong Dai ◽  
Xiaojuan Wang ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Culture Media ◽  
Extended Period ◽  
Tween 80 ◽  
Surface Expression ◽  
Dependent Manner ◽  
Bacteria Growth ◽  
Growth Changes

ABSTRACT The outermost layer of Mycobacterium tuberculosiscontains two major polysaccharides, arabinomannan (AM) and glucan (GC). We studied the in vitro and in vivo expression of anM. tuberculosis AM antigen using monoclonal antibody (MAb) 9d8 (2a), an isotype-switched variant of the immunoglobulin G3 (IgG3) MAb 9d8. MAb 9d8 had been previously shown to bind M. tuberculosis AM and the M. tuberculosis surface. Our in vitro experiments showed that MAb 9d8(2a) bound strongly to whole-cell M. tuberculosis Erdman but not to the CDC 1551 strain grown in medium for an extended period. However, AM antigen was detected in the culture supernatant of both strains, and its concentration increased in a time-dependent manner. The detection of AM antigen from both strains was decreased in the presence of Tween 80. In mice infected with M. tuberculosis Erdman, AM antigen accumulated in organ homogenates concomitant to an increase in bacterial organ burden and an increase in IgG and IgM titer to AM. These results (i) indicate that the surface expression of AM during in vitro growth changes with culture age, is strain dependent, and is affected by the presence of Tween 80 in the culture media; (ii) show that AM is produced by bacteria growth in vivo; and (iii) demonstrate that the amount of in vivo-detected AM can be dependent on the number of bacteria in the infected organ.

Biofilms Formed by Mycobacterium tuberculosis on Cement, Ceramic, and Stainless Steel Surfaces and Their Controls

2014 ◽  
Vol 77 (4) ◽  
pp. 599-604 ◽  
Author(s):  
VICTORIA ADETUNJI ◽  
ADEREMI KEHINDE ◽  
OLAYEMI BOLATITO ◽  
JINRU CHEN
Keyword(s):  
Stainless Steel ◽  
Mycobacterium Tuberculosis ◽  
Incubation Time ◽  
Liver Extract ◽  
Tween 80 ◽  
Hot Water ◽  
The Third ◽  
Steel Surfaces ◽  
Biofilm Development

This study assessed the biofilms formed by selected strains of Mycobacterium tuberculosis and investigated the efficacy of three different treatments to control the biofilms. Two M. tuberculosis strains were inoculated separately in 150 ml of Middlebrook 7H9–Tween 80 (0.1%) broth with 5% liver extract and 10% oleic albumin dextrose catalase (OADC) supplement, 5% liver extract alone, or 10% OADC alone in sterile jars, each containing a 2-cm2 coupon of cement, ceramic, or stainless steel for biofilm development at 37°C, with agitation for 2, 3, or 4 weeks. Biofilms on the coupons were exposed to 10 ml of 2% sanitizer A or 0.5% sanitizer B at 28 and 45°C and to hot water at 85°C for 5 min. Residual biofilms on treated and untreated coupons were assessed. Both strains of M. tuberculosis formed biofilms on the three surfaces; however, one strain formed more biofilms. More biofilms were formed when media containing 5% liver extract was used. Biofilm mass increased as incubation time increased until the third week. More biofilms were formed on cement than on ceramic and stainless steel coupons. Sanitizing treatments at 45°C removed more biofilms than those at 28°C. However, neither treatment completely eliminated the biofilms.

THE INDUCTION BY LYSOZYME OF AN L-TYPE GROWTH IN MYCOBACTERIUM TUBERCULOSIS

1966 ◽  
Vol 12 (1) ◽  
pp. 11-16 ◽  
Author(s):  
Hilda Pope Willett ◽  
Harshad Thacore
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Tween 80 ◽  
Tetraacetic Acid ◽  
Ethylenediamine Tetraacetic Acid ◽  
Solid Media

Lysozyme in combination with ethylenediamine tetraacetic acid (EDTA) gives rise to spheroplasts when Mycobacterium tuberculosis is cultured in a nutritionally complete Tween 80 – albumin medium. These spheroplasts do not multiply in this medium, but when such forms, stabilized by the presence of sucrose and Mg++, are inoculated into PPLO agar or broth an L-form of growth is initiated. A CO2requirement was demonstrated by L-forms grown on solid media.

scholarly journals Comparison of the Construction of Unmarked Deletion Mutations in Mycobacterium smegmatis, Mycobacterium bovis Bacillus Calmette-Guérin, and Mycobacterium tuberculosis H37Rv by Allelic Exchange

1999 ◽  
Vol 181 (16) ◽  
pp. 4780-4789 ◽  
Author(s):  
Martin S. Pavelka ◽  
William R. Jacobs
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Mycobacterium Bovis ◽  
Mycobacterium Smegmatis ◽  
Tween 80 ◽  
Defined Medium ◽  
Lysine Biosynthesis ◽  
Rich Medium ◽  
Deletion Mutations ◽  
Allelic Exchange ◽  
Slow Growing

ABSTRACT Until recently, genetic analysis of Mycobacterium tuberculosis, the causative agent of tuberculosis, was hindered by a lack of methods for gene disruptions and allelic exchange. Several groups have described different methods for disrupting genes marked with antibiotic resistance determinants in the slow-growing organismsMycobacterium bovis bacillus Calmette-Guérin (BCG) and M. tuberculosis. In this study, we described the first report of using a mycobacterial suicidal plasmid bearing the counterselectable marker sacB for the allelic exchange of unmarked deletion mutations in the chromosomes of two substrains ofM. bovis BCG and M. tuberculosis H37Rv. In addition, our comparison of the recombination frequencies in these two slow-growing species and that of the fast-growing organismMycobacterium smegmatis suggests that the homologous recombination machinery of the three species is equally efficient. The mutants constructed here have deletions in the lysA gene, encoding meso-diaminopimelate decarboxylase, an enzyme catalyzing the last step in lysine biosynthesis. We observed striking differences in the lysine auxotrophic phenotypes of these three species of mycobacteria. The M. smegmatis mutant can grow on lysine-supplemented defined medium or complex rich medium, while the BCG mutants grow only on lysine-supplemented defined medium and are unable to form colonies on complex rich medium. The M. tuberculosis lysine auxotroph requires 25-fold more lysine on defined medium than do the other mutants and is dependent upon the detergent Tween 80. The mutants described in this work are potential vaccine candidates and can also be used for studies of cell wall biosynthesis and amino acid metabolism.

scholarly journals Phosphoenolpyruvate depletion mediates both growth arrest and drug tolerance of Mycobacterium tuberculosis in hypoxia

2021 ◽  
Vol 118 (35) ◽  
pp. e2105800118
Author(s):  
Juhyeon Lim ◽  
Jae Jin Lee ◽  
Sun-Kyung Lee ◽  
Seoyong Kim ◽  
Seok-Yong Eum ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Carbon Flux ◽  
Drug Tolerance ◽  
Drug Susceptibility ◽  
Tb Treatment ◽  
Metabolic Modulation ◽  
Repetitive Exposure ◽  
Metabolic Remodeling ◽  
Novel Strategy

Mycobacterium tuberculosis (Mtb) infection is difficult to treat because Mtb spends the majority of its life cycle in a nonreplicating (NR) state. Since NR Mtb is highly tolerant to antibiotic effects and can mutate to become drug resistant (DR), our conventional tuberculosis (TB) treatment is not effective. Thus, a novel strategy to kill NR Mtb is required. Accumulating evidence has shown that repetitive exposure to sublethal doses of antibiotics enhances the level of drug tolerance, implying that NR Mtb is formed by adaptive metabolic remodeling. As such, metabolic modulation strategies to block the metabolic remodeling needed to form NR Mtb have emerged as new therapeutic options. Here, we modeled in vitro NR Mtb using hypoxia, applied isotope metabolomics, and revealed that phosphoenolpyruvate (PEP) is nearly completely depleted in NR Mtb. This near loss of PEP reduces PEP-carbon flux toward multiple pathways essential for replication and drug sensitivity. Inversely, supplementing with PEP restored the carbon flux and the activities of the foregoing pathways, resulting in growth and heightened drug susceptibility of NR Mtb, which ultimately prevented the development of DR. Taken together, PEP depletion in NR Mtb is associated with the acquisition of drug tolerance and subsequent emergence of DR, demonstrating that PEP treatment is a possible metabolic modulation strategy to resensitize NR Mtb to conventional TB treatment and prevent the emergence of DR.

Carbon flux rerouting during Mycobacterium tuberculosis growth arrest

2016 ◽  
Vol 99 (6) ◽  
pp. 1179-1179 ◽  
Author(s):  
Lanbo Shi ◽  
Charles D. Sohaskey ◽  
Carmen Pheiffer ◽  
Pratik Datta ◽  
Michael Parks ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Carbon Flux ◽  
Growth Arrest

scholarly journals Acetyl-CoA-mediated activation of Mycobacterium tuberculosis isocitrate lyase 2

2019 ◽  
Vol 10 (1) ◽  
Author(s):  
Ram Prasad Bhusal ◽  
Wanting Jiao ◽  
Brooke X. C. Kwai ◽  
Jóhannes Reynisson ◽  
Annabelle J. Collins ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Carbon Flux ◽  
Isocitrate Lyase ◽  
Tca Cycle ◽  
Structural Rearrangement ◽  
Tricarboxylic Acid ◽  
Glyoxylate Shunt ◽  
Acetyl Coa ◽  
High Lipid ◽  
Lipid Metabolites

Abstract Isocitrate lyase is important for lipid utilisation by Mycobacterium tuberculosis but its ICL2 isoform is poorly understood. Here we report that binding of the lipid metabolites acetyl-CoA or propionyl-CoA to ICL2 induces a striking structural rearrangement, substantially increasing isocitrate lyase and methylisocitrate lyase activities. Thus, ICL2 plays a pivotal role regulating carbon flux between the tricarboxylic acid (TCA) cycle, glyoxylate shunt and methylcitrate cycle at high lipid concentrations, a mechanism essential for bacterial growth and virulence.

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