Fig. 4. HS upon exposure to kanamycin is not a consequence of slow growth or generally decreased metabolic activity.

ns, not significant; *P < 0.05; ***P < 0.001; ****P <0.0001. Error bars: ±SD. (A) ArgD-E105X with an atc-inducible copy of WT argD (E105X::argD) was grown in 0, 0.8, or 25 ng/ml of atc. Left: Growth curves. Symbols, mean OD₆₀₀ of two to three replicates. Right: Growth rate relative to WT mean [bars, means of four to five replicate cultures (symbols) pooled from two independent experiments] and kanamycin survival (filters above bars). Generation times compared to WT by one-way analysis of variance (ANOVA). Growth curves and filters representative of two independent experiments. (B) Generation times of WT versus ArgD-E105X in Sauton’s (unpaired two-tailed t test). Bars, means of N independent experiments (symbols; N = 5 for WT and 3 for E105X) with three to four replicates each. (C) Kanamycin survival of WT and ArgD-E105X in Sauton’s. Representative of four to five independent experiments (E105X in Sauton’s, 4; WT in Sauton’s versus 7H9, 5). (D and E) Metabolites from ΔargA, ΔargA::argA, ΔargD, and ΔargD::argD provisionally identified by accurate mass and retention time. (D) Colors reflect mean log₂ FC relative to WT from three independent experiments (three replicates each). N-A-l-Glu-5-SA, N-acetyl-l-glutamate-5-semialdehyde; deoxy-MTA, deoxy-methylthioadenosine; SAM, S-adenosylmethionine. (E) Mean log₂ FC compared to a theoretical mean of 0 (two-tailed one-sample t test). Dotted lines, cutoffs (significance, P = 0.05; FC, ±2). (F) Strains from (D) were incubated with resazurin. Fluorescence (excitation 544 nm, emission 590 nm) was normalized to OD₆₀₀ before addition of resazurin and then to WT mean. Bars, means of eight replicate cultures (symbols) pooled from two independent experiments (exp.). Means compared to a theoretical mean of 1 (two-tailed one-sample t test). HK, heat-killed. See also data S1.