Aggregated Mycobacterium tuberculosis enhances the inflammatory response

Abstract

Mycobacterium tuberculosis (Mtb) readily aggregates in culture and Mtb aggregates in the lung were observed in experimental Mtb infection. However, the physiological consequences of Mtb aggregation are incompletely understood. Here we examined the human macrophage transcriptional response to aggregated Mtb relative to infection with non-aggregated single or multiple bacilli per host cell. Infection with aggregated Mtb led to an early upregulation of pro-inflammatory associated genes and enhanced TNFα signaling via the NFκB pathway. Both these pathways were significantly upregulated relative to infection with single bacilli, and TNFα signaling was also significantly elevated relative to infection with multiple non-aggregated Mtb. Secretion of TNFα and downstream cytokines were also enhanced. On a longer timescale, aggregate infection led to overall increased acidification per macrophage and a high proportion of death in these cells after aggregate phagocytosis. Host cell death did not occur when Mtb aggregates were heat killed despite such clumps being readily picked up. To validate that Mtb aggregates do occur in the human lung, we document Mtb aggregates surrounding a cavity in a human TB lesion. Aggregates may therefore be present in some lesions and elicit a stronger inflammatory response resulting in recruitment of additional phagocytes and their subsequent death, potentially leading to necrosis and transmission.

Single cell preparations ofMycobacterium tuberculosisdamage the mycobacterial envelope and disrupt macrophage interactions

“…However, there was a wide distribution in cell envelope thickness, which may contribute to heterogeneity in macrophage responses to bacilli on a cell-to-cell level. Finally, we considered that differences in the degree of aggregation in the different bacterial preparations may account for differences in inflammatory potential or intracellular survival (Rodel et al, 2021), but heterogeneity in this aspect of the bacterial population was unlikely to explain the differences in outcomes. There may be other aspects of underlying heterogeneity in our samples that contribute to their distinct behavior or confound bulk measurements.…”

“…For example, the observation that Mtb forms serpentine cords in vivo dates back to the earliest descriptions of the bacteria. Aggregated Mtb are found at the periphery of human necrotic granulomas, in alveolar macrophages of Mtb-infected patients, and are exhaled by infected individuals (Dinkele et al, 2021;Rodel et al, 2021;Ufimtseva et al, 2018). The literature describing the impact of aggregation on host interactions is difficult to interpret in light of our findings, as many of these studies used agitation with glass beads, filtration, sonication, or some combination of these procedures to generate the dispersed samples (Kolloli et al, 2021;Mahamed et al, 2017;Rodel et al, 2021).…”

“…Aggregated Mtb are found at the periphery of human necrotic granulomas, in alveolar macrophages of Mtb-infected patients, and are exhaled by infected individuals (Dinkele et al, 2021;Rodel et al, 2021;Ufimtseva et al, 2018). The literature describing the impact of aggregation on host interactions is difficult to interpret in light of our findings, as many of these studies used agitation with glass beads, filtration, sonication, or some combination of these procedures to generate the dispersed samples (Kolloli et al, 2021;Mahamed et al, 2017;Rodel et al, 2021). Thus, bacterial aggregation is likely an important virulence property of Mtb, which investigators overlook in the effort to generate single cell suspensions; at the same time, in generating single cell suspensions, investigators introduce the potential for experimental artifact.…”