Investigating the effect of a Mycobacterium tuberculosis inhA promoter mutation in cell envelope composition and morphology
type: CreativeWork
description: Isoniazid (INH) resistance in Mycobacterium tuberculosis (M.tb) compromises the effectiveness of global tuberculosis treatment programs, with the inhA promotor mutation (IPM) serving as a driver of low-level drug resistance. IPMs are seen more frequently in multi-drug-resistant isolates and are considered a gateway to the development of extensive drug resistance. IPMs contribute resistance to INH and ethionamide which are first and second-line drugs respectively that target mycolic acid (MA) synthesis. MA are characteristic components of mycobacterial species’ cell wall and contribute to the impermeability of cell wall by virtue of its hydrophobicity. IPMs result in a transcriptional upregulation of inhA, and mabA, which are genes involved in MA synthesis, and hemZ which is involved in heme synthesis through the incorporation of iron. It is unknown how the upregulation of genes involved in MA synthesis may affect the cell wall, therefore, this project aimed to investigate molecular and microscopic differences in the cell wall between IPM and wild-type (WT) strain. To investigate the influence of IPM on overall cell wall morphology multiple complementary techniques were employed. Scanning electron microscopy (SEM) was implemented to visualise extracellular surface features and determine cell length, while transmission electron microscopy (TEM) was used to examine ultrastructural cell wall layer distances. To further understand the architecture and organisation of M.tb cell wall we assessed membrane fluidity using DPH, a membrane fluorescent probe. In parallel, a preliminary investigation of cell wall integrity using propidium iodide was performed. Finally, to determine the influence of IPM on MA, two extraction protocols were compared and validated. The resulting extracts were firstly evaluated by thin-layer chromatography, followed by mass spectrometry and nuclear magnetic resonance spectrometry to confirm MA extraction. We found that even though two genetically ide…
abstract: Isoniazid (INH) resistance in Mycobacterium tuberculosis (M.tb) compromises the effectiveness of global tuberculosis treatment programs, with the inhA promotor mutation (IPM) serving as a driver of low-level drug resistance. IPMs are seen more frequently in multi-drug-resistant isolates and are considered a gateway to the development of extensive drug resistance. IPMs contribute resistance to INH and ethionamide which are first and second-line drugs respectively that target mycolic acid (MA) synthesis. MA are characteristic components of mycobacterial species’ cell wall and contribute to the impermeability of cell wall by virtue of its hydrophobicity. IPMs result in a transcriptional upregulation of inhA, and mabA, which are genes involved in MA synthesis, and hemZ which is involved in heme synthesis through the incorporation of iron. It is unknown how the upregulation of genes involved in MA synthesis may affect the cell wall, therefore, this project aimed to investigate molecular and microscopic differences in the cell wall between IPM and wild-type (WT) strain. To investigate the influence of IPM on overall cell wall morphology multiple complementary techniques were employed. Scanning electron microscopy (SEM) was implemented to visualise extracellular surface features and determine cell length, while transmission electron microscopy (TEM) was used to examine ultrastructural cell wall layer distances. To further understand the architecture and organisation of M.tb cell wall we assessed membrane fluidity using DPH, a membrane fluorescent probe. In parallel, a preliminary investigation of cell wall integrity using propidium iodide was performed. Finally, to determine the influence of IPM on MA, two extraction protocols were compared and validated. The resulting extracts were firstly evaluated by thin-layer chromatography, followed by mass spectrometry and nuclear magnetic resonance spectrometry to confirm MA extraction. We found that even though two genetically ide…
author: Harris, Labeeqah
language: en
date published: 2026-03
date modified: 2026-07-24T04:40:06Z
provider: Stellenbosch University
publisher: Networked Digital Library of Theses and Dissertations
type: WebPage
source organization: Stellenbosch : Stellenbosch University