ABSTRACT Phagocytosis of Mycobacterium tuberculosis (Mtb) aggregates, rather than similar numbers of single bacilli, induces host macrophage death and favors bacterial growth. Here, we examined whether aggregation contributes to enhanced Mtb pathogenicity in vivo in rabbit lungs. Rabbits were exposed to infectious aerosols containing mainly Mtb-aggregates (Mtb-AG) or Mtb-single cells (Mtb-SC). The lung bacterial load, histology, and immune cell composition were investigated over time. Genome-wide transcriptome analysis, cellular and tissue-level assays, and immunofluorescent imaging were performed on lung tissue to define and compare differential immune activation and pathogenesis between Mtb-AG and Mtb-SC infection. Lung bacillary loads, disease scores, lesion size, and structure were significantly higher in Mtb-AG than in Mtb-SC infected animals. A differential immune cell distribution and activation were noted in the lungs and spleen of the two groups of infected animals. Mtb-AG infected animals also showed early induction of inflammatory network genes associated with necrosis and reduced host cell viability. Consistently, larger lung granulomas with clumped Mtb, extensive necrotic foci, and elevated matrix metalloproteases expression were observed in Mtb-AG infected rabbits. Our findings suggest that bacillary aggregation increases Mtb fitness for improved growth and accelerated lung inflammation and cell death, thereby exacerbating disease pathology in the lungs.

Aggregation state of Mycobacterium tuberculosis impacts host immunity and augments pulmonary disease pathology