The clinical relevance of structural modifications in the cell envelope glycans of Mycobacterium abscessus

Abstract

Mycobacterium abscessus is one of the leading causes of pulmonary disease caused by nontuberculous mycobacteria (NTMs) globally. As an opportunistic pathogen, individuals who have dysfunctional immunological and/or mechanical defenses are at the greatest risk of acquiring M. abscessus, but individuals with cystic fibrosis (CF) are the most disproportionately affected. Treatment of NTM pulmonary disease (NTM-PD) is complicated by the need to speciate the causative NTM to design an appropriate multidrug treatment regimen. Detecting NTMs is also challenging due to the low sensitivity associated with the reliance upon bacterial cultures. These hurdles in diagnosis and treatment are compounded by the propensity of M. abscessus pulmonary infections to become chronic, prolonging exposure of mycobacteria to the selective pressures of the human lung that can drive within-host adaptation. One of the genes identified to be under evolutionary pressure during chronic M. abscessus pulmonary infection was ubiA, a cell envelope biosynthetic gene. UbiA catalyzes the biosynthetic pathway of the arabinose donor required for the biosynthesis of two important molecules in the cell envelope, arabinogalactan (AG) and lipoarabinomannan (LAM). The mycobacterial cell envelope is a notorious barrier that protects mycobacteria and enables mycobacterial survival in harsh environments, and it is also the primary interface by which mycobacteria interact with its surrounding environment, including the human lung. As such, the patient-derived mutations in ubiA were unlikely to be functionally silent and were anticipated to modulate how M. abscessus interacted with the human lung environment to the benefit of M. abscessus. On the other hand, while modifications of the cell envelope presumably benefit mycobacteria, that does not preclude the use of cell envelope components as biomarkers of NTM infection for the benefit of clinics. The reliance of diagnostics and NTM monitoring/screening upon culturing for mycobacteria from patient samples presents challenges with detection sensitivity following standard sample decontamination procedures and long wait times for cultures to grow. Though molecular tests and protein-based mass spectrometric methods are used in diagnostic laboratories to accurately speciate NTMs (and subspeciate in the case of M. abscessus), these methods are culture-dependent. The discovery of a novel methyl modification of phosphatidyl-myo-inositol mannosides (PIMs) from M. abscessus positioned methylated PIMs (mPIMs) as a potential biomarker candidate. PIMs are a class of glycolipids that are abundant in the cell envelope of mycobacteria and closely related actinomycetes (e.g., Corynebacterium spp., Nocardia spp.) and essential for mycobacterial growth. PIMs are also released by mycobacteria and have been previously detected in exhaled breath condensates from patients with tuberculosis. Importantly, mPIMs were highly anticipated to differentiate M. abscessus from Mycobacterium avium complex infections, the leading cause of NTM-PD in the United States. For these reasons, we investigated the utility of using mPIMs as a culture-independent biomarker of M. abscessus. The research conducted in this dissertation will demonstrate the continued relevance of studying the mycobacterial cell envelope with respect to host-pathogen interactions and as a diagnostic tool. Patient-derived mutations in ubiA, a cell envelope biosynthetic gene, can have far-reaching consequences on the immunomodulatory properties of M. abscessus and its ability to evade cellular uptake and establish robust biofilms that enable mycobacterial persistence. Additionally, we have evaluated and found mPIMs to be largely specific to M. abscessus and constitutively expressed, making mPIMs a promising biomarker of M. abscessus infection.