Time-Lapse Atomic Force Microscopy Reveals New End Take Off (Neto) Dynamics in Mycobacteria

Abstract

Tuberculosis is a lung infection caused by Mycobacterium tuberculosis. One third of the world population is thought to be infected. Antibiotic treatment with antibiotics lasts 6 month, which is difficult to achieve in low-income countries. The uncommon ability of tuberculosis bacteria to persist during long periods is linked to various factors, one of which potentially being cell growth asymmetry. However, growth symmetry or asymmetry of mycobacteria remains a controversial topic as opposing descriptions have been reported in the literature. Observing bacterial growth at the single cell level is inherently challenging due to the diffraction limit of optical microscopes and the small size of the cells. We built a new dedicated platform for long-term, correlated Atomic Force Microscopy (AFM) and fluorescence microscopy and obtained time-lapses of bacterial growth at an unprecedented resolution. Surprisingly, the AFM high-resolution time-lapses did not match any of the two previously reported models of pure symmetry and pure asymmetry. Mycobacterial pole growth dynamics was rather an intermediate between the two, reminiscent of the well-known “new end take off” mechanism for fission yeast. Using AFM nanomanipulation we show that growth asymmetry is not a physical occlusion phenomenon, but is inherent to mycobacterial growth. Finally, using fluorescent photoconversion microscopy we identify biomolecular mechanisms linked to growth asymmetry, which in the future could be used to develop novel treatments for tuberculosis infection.