Fig 1. Disruption of the sRNA B11 causes rough morphology with reduced GPL content in M. abscessus.
A. The indicated strains were grown in liquid media with Tween 80, diluted and plated on solid media with and without Tween 80. Colonies were imaged after 7 days of growth. See S1E Fig for exact mutations of ∆B11+B11mutated. B. Northern blotting of duplicate RNA samples from the indicated strains confirmed that B11 expression was reduced in the transposon mutant, absent from the deletion mutant, and restored in the complemented mutant. C. Thin layer chromatography analysis of surface lipids extracted from the indicated strains grown on 7H10-ADC agar plates. Lipids extracted from equivalent amounts of bacterial cells were analyzed on silica gel 60-precoated TLC plates F254 (Merck) developed in the solvent system chloroform:methanol:water (90:10:1, v/v/v). The plate was revealed by spraying with α-naphthol and heating. D. Quantitative PCR reveals that expression of two key GPL biosynthesis genes is modestly but significantly reduced in strains lacking B11. * P < 0.05; ** P < 0.01; *** P < 0.001; ANOVA with Tukey post-test. qPCR comparisons where P > 0.05 are not shown. qPCR data represent triplicate log-phase cultures. All experiments shown in this figure were performed at least twice and representative data are shown.