Intracellular growth of Mycobacterium tuberculosis after macrophage cell death leads to serial killing of host cells.

Deeqa Mahamed,Lance Oom,Gila Lustig,Alexander S Pym,Jessica Hunter,Chanelle Mc Arthur,Yashica Ganga,Colisile Mathonsi,Gopalkrishna Sreejit,Myshnee Naicker,Steven Skroch,Emily B Wong,Mikael Boulle,Kelly Pillay,Alex Sigal,Moosa Suleman,Sanisha Rampersad,Oana Catinas

doi:10.7554/elife.22028

Abstract

A hallmark of pulmonary tuberculosis is the formation of macrophage-rich granulomas. These may restrict Mycobacterium tuberculosis (Mtb) growth, or progress to central necrosis and cavitation, facilitating pathogen growth. To determine factors leading to Mtb proliferation and host cell death, we used live cell imaging to track Mtb infection outcomes in individual primary human macrophages. Internalization of Mtb aggregates caused macrophage death, and phagocytosis of large aggregates was more cytotoxic than multiple small aggregates containing similar numbers of bacilli. Macrophage death did not result in clearance of Mtb. Rather, it led to accelerated intracellular Mtb growth regardless of prior activation or macrophage type. In contrast, bacillary replication was controlled in live phagocytes. Mtb grew as a clump in dead cells, and macrophages which internalized dead infected cells were very likely to die themselves, leading to a cell death cascade. This demonstrates how pathogen virulence can be achieved through numbers and aggregation states.

Highlights

Tuberculosis is characterized by the formation of granulomas, cellular structures which attempt to ‘wall off’ infection by surrounding it with cells of the immune system (Ramakrishnan, 2012; Russell, 2007; Russell et al, 2010)
We infected human monocyte derived macrophages (MDM) with the virulent M. tuberculosis H37Rv strain labelled with red fluorescent protein (RFP) or mCherry expressed under the control of a constitutively active promoter
To examine whether Mycobacterium tuberculosis (Mtb) fluorescence is a valid measure of Mtb number, we tracked the increase in Mtb by fluorescence versus colony forming units (CFU) over 3 days of growth

Summary

Introduction

Tuberculosis is characterized by the formation of granulomas, cellular structures which attempt to ‘wall off’ infection by surrounding it with cells of the immune system (Ramakrishnan, 2012; Russell, 2007; Russell et al, 2010). Granulomas do not always succeed in containing Mtb infection, and different granulomas in the same lung can control the infection or support the growth of the bacilli (Barry et al, 2009; Lenaerts et al, 2015; Lin et al, 2014; Kaplan et al, 2003). While Mtb is considered an intracellular pathogen infecting live cells, its proliferation during active pulmonary tuberculosis occurs in an environment containing many dead cells and cell remnants (Kaplan et al, 2003; Hunter, 2011; Hunter et al, 2007; Welsh et al, 2011; Irwin et al, 2015)

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Conclusion