Cell wall synthesizing complexes in Mycobacteriales.

Members of the order Mycobacteriales are distinguished by a characteristic diderm cell envelope, setting them apart from other Actinobacteria species. In addition to the conventional peptidoglycan cell wall, these organisms feature an extra polysaccharide polymer composed of arabinose and galactose, termed arabinogalactan. The nonreducing ends of arabinose are covalently linked to mycolic acids (MAs), forming the immobile inner leaflet of the highly hydrophobic MA membrane. The contiguous outer leaflet of the MA membrane comprises trehalose mycolates and various lipid species. Similar to all actinobacteria, Mycobacteriales exhibit apical growth, facilitated by a polar localized elongasome complex. A septal cell envelope synthesis machinery, the divisome, builds instead of the cell wall structures during cytokinesis. In recent years, a growing body of knowledge has emerged regarding the cell wall synthesizing complexes of Mycobacteriales., focusing particularly on three model species: Corynebacterium glutamicum, Mycobacterium smegmatis, and Mycobacterium tuberculosis.

Effects of benzothiazinone and ethambutol on the integrity of the corynebacterial cell envelope.

The mycomembrane (MM) is a mycolic acid layer covering the surface of Mycobacteria and related species. This group includes important pathogens such as Mycobacterium tuberculosis, Corynebacterium diphtheriae, but also the biotechnologically important strain Corynebacterium glutamicum. Biosynthesis of the MM is an attractive target for antibiotic intervention. The first line anti-tuberculosis drug ethambutol (EMB) and the new drug candidate, benzothiazinone 043 (BTZ) interfere with the synthesis of the arabinogalactan (AG), which is a structural scaffold for covalently attached mycolic acids that form the inner leaflet of the MM. We previously showed that C. glutamicum cells treated with a sublethal concentration of EMB lose the integrity of the MM. In this study we examined the effects of BTZ on the cell envelope. Our work shows that BTZ efficiently blocks the apical growth machinery, however effects in combinatorial treatment with ß-lactam antibiotics are only additive, not synergistic. Transmission electron microscopy (TEM) analysis revealed a distinct middle layer in the septum of control cells considered to be the inner leaflet of the MM covalently attached to the AG. This layer was not detectable in the septa of BTZ or EMB treated cells. In addition, we observed that EMB treated cells have a thicker and less electron dense peptidoglycan (PG). While EMB and BTZ both effectively block elongation growth, BTZ also strongly reduces septal cell wall synthesis, slowing down growth effectively. This renders BTZ treated cells likely more tolerant to antibiotics that act on growing bacteria.