Fig. 1. Medium-dependent growth defects of ∆mptA.

a Proposed pathway for PIM, LM, and LAM biosynthesis. Bold text, enzymes. Ara-T, arabinosyltransferase; Man-T, mannosyltransferase. Cartoon made with BioRender. b Anti-MptA immunoblot of cell lysates prepared from WT, ∆mptA, ∆mptA L5::mptA (WT), and ∆mptA L5::mptA (D129A) cells. Blot is representative of two independent experiments with similar results. c Lipoglycans extracted from WT, ∆mptA, ∆mptA L5::mptA (WT), and ∆mptA L5::mptA (D129A) cells, visualized by glycan staining. Gel is representative of two independent experiments with similar results. d Representative example images comparing colony size between WT, ∆mptA, ∆mptA L5::mptA (WT), and ∆mptA L5::mptA (D129A) on Middlebrook 7H10 and LB solid agar media grown at 30 °C or 37 °C. e Quantification of colony area for each condition described in Fig. 1d. n = 8, 22, 14, 53 (7H10 30 °C); 15, 15, 21, 9 (7H10 37 °C); 7, 16, 23, 26 (LB 30 °C); 9, 34, 22, 32 (LB 37 °C) colonies for WT, ∆mptA, ∆mptA L5::mptA (WT), and ∆mptA L5::mptA (D129A), respectively. A box spanning the interquartile range (IQR) is drawn from the first quartile to the third quartile with a horizontal line indicating the median. The whiskers extend from the box to the farthest data point within 1.5x IQR from the box. Dots beyond the whiskers indicate potential outliers. Statistical significance was determined by one-way ANOVA and Tukey post-hoc test. f Cross-sectional imaging of microcolonies from WT and ∆mptA strains stained with SYTO9 (live, green) and propidium iodide (dead, red) grown as micro-aggregates in LB medium. Images are representative of results obtained from three independent experiments with similar results. Source data for panels b, c, and e are provided in the Source Data file. Colony image data for panels d and e are available at https://github.com/IanLairdSparks/Sparks_2023. Raw micro-aggregate data images for panel f can be accessed via 10.5061/dryad.1vhhmgr1w⁷².