Fig. 1: M. tuberculosis modulates its retrograde transport to the perinuclear region to avert delivery to the lysosomes.

a Calculation of perinuclear index of Mtb distribution inside macrophages. Green- Mtb cells, red- 3 μm concentric ring, and blue- 6 μm concentric ring. b Fractional distance of Mtb distribution inside macrophages. Green—bacterial cells. a, b were created using BioRender.com. c THP-1 macrophages were infected with GFP-MtbH37Rv (live and heat-killed) for 3 h. The figure shows the representative images of THP-1 macrophages infected with GFP-MtbH37Rv (live and heat-killed). Violin plots depicting perinuclear index (d, n = 70–75) and fractional distance (e, n > 90) of GFP-MtbH37Rv in cells were quantified from experiments described in (c) (One-tailed, Mann–Whitney U test, Confidence Interval-95%). Values are represented as arbitrary units (A.U.) in violin plots. f THP-1 macrophages were infected with GFP-MtbH37Rv to analyze whether Mtb’s spatial localization affects its delivery to lysosomes. Inset shows an enlarged region of interest containing Mtb. g Percent colocalization of GFP-MtbH37Rv with lysosomes (stained with anti-LAMP1) at different distances from the nucleus (stained with Hoechst dye) (n = 110). Data were quantified from experiments described in f (One-tailed, unpaired Student’s t test, CI-95%). h C3HeB/FeJ mice were infected with MtbH37Rv for two weeks. The representative image shows lung cells infected with MtbH37Rv (green) and its distance from the nucleus (blue, stained with DAPI). i Lung sections described in h were utilized for immunostaining against LAMP-1 protein. j Percent colocalization of MtbH37Rv with lysosomes at different distances from the nucleus (n = 45) was quantified from experiments described in (i) (One-tailed, unpaired Student’s t test, CI-95%). Data in d, e represent Mtb distribution in four quarters with median (central line), first quartile (upper line), and third quartile (lower line) from three independent biological experiments. Data in g, j represent mean ± SEM from three independent biological experiments. Scale Bar: 10 μm for c, f and 2 μm for h, i.