Quantification of differential toxin expressions and their relation to distinct lifespans of bacterial subpopulations associated with diverse host immune mechanisms

Abstract

AbstractAn assortment of robust intracellular defence mechanisms are critical for restricting proliferation of pathogens and maintaining sanctity of the cytosol. Defect in these mechanisms could be exploited by the pathogens for creation of a safe sanctuary which can act as a transient reservoir for periodic dissemination into the host. While residing inside the host cell, pore forming toxins secreted by the pathogens compromises the integrity of the vacuole and exposes the microbe to diverse intracellular defence mechanisms. However, the correlation between toxin expression levels and consequent pore dynamics, fostering pathogen’s intracellular life, remains largely unexplored. In this study, usingStreptococcus pneumoniae(SPN) and its secreted pore forming toxin pneumolysin (Ply), as model systems, we explored various facets of host-pathogen interactions in host cytosol, governed by the toxin expression and the resultant pore formation. The extent of damage on the endosomal membrane was found to dictate subsequent interaction with different host endosomal damage sensors. This in turn governed the routes of SPN clearance, revealing multiple layers of defence mechanisms at host’s disposal for counteracting invaded pathogens. A subset of SPN population producing extremely low amount of Ply inflicted minimal damage to the endomembrane, precluding decoration by endomembrane damage sensors and significantly prolonging its intracellular persistence. Such long persisting bacterial population could be key for pathogenic transmission or ensuing invasive disease. Using time-lapse fluorescence imaging, we monitored lifespans of different pneumococcal population subsets inside host cells. After quantitative analysis of various timescales such as pore formation time, vacuolar or cytosolic residence time and total degradation time, we developed a mathematical model that could correlate these to intravacuolar accumulation of Ply monomers. By proposing events like pore formation and vacuolar degradation of SPN as first passage processes, our theoretical modelling yields estimates of Ply production rate, burst size, and threshold Ply quantities which triggers these outcomes. Collectively, we present a general method by which intracellular lifespans of pathogens could be correlated to differential levels of toxins that they produce.