Article Figures & Data
Figures
- FIG 1
Differences in tailocin survival and proportion of surviving phenotypes between stationary- and log-phase cultures of Pph. (A) Cultures were treated with a lethal dose of tailocin (100 AU), and viable cells before and after 1 h of treatment were enumerated. (B) Tailocin death curve of the two cultures of Pph upon exposure to tailocin (100 AU). Viable populations were enumerated before and after 1, 4, 8, and 24 h of treatment (see Fig. S2). Three independent experiments were performed with 3 to 6 biological replicates per time point. The mean and standard error of the mean are graphed. A P value of < 0.05 or different letters indicate significant differences between stationary- and log-phase treatments for a given time point, as analyzed in SAS 9.4 with proc Glimmix. (C) Percentage of surviving colony phenotypes upon tailocin retreatment. Randomly selected surviving colonies (n = 12 to 44 for each growth phase and hours of treatment from three independent experiments) were subcultured and treated again with tailocin, and the percentage of the surviving phenotype was calculated.
- FIG 2
Dynamics of tailocin survival with concentrated tailocin treatment. (A) Cultures were treated with a high dose of tailocin (900 AU), and viable populations pre- and posttreatment were determined. Experiments were repeated at least three times with 3 to 6 biological replicates per time point. The mean and standard error of the mean are graphed. A P value of <0.05 indicates significant difference within grouped bars, as analyzed in SAS 9.4 with proc Glimmix. (B) Percentage of surviving colony phenotypes after treatment with concentrated (900 AU) tailocin for 1 hour. Although most of the surviving colonies were either incomplete resistant or resistant, persistent cells were maintained even at high tailocin concentration. Surviving colonies were tested during three independently repeated experiments for both cultures.
- FIG 3
Treatment response of tailocin persistent and resistant lines. (A) Reduction in the population of tailocin persistent and resistant mutant lines upon retreatment with tailocin. Log cultures of each line were treated with 900 AU of tailocin, and the change in the population was calculated after an hour of tailocin exposure. At least three separate colonies of each line were tested, and experiments were repeated a minimum of three times. Means of the difference in log-transformed viable population pre- and posttreatment are graphed. Error bars indicate the standard error of the mean. (B) Assessment of the response of mutant lines to tailocin in overlay conditions. Dilutions of tailocins (shown on the leftmost column) were spotted over the culture lawn of each of the lines. The yellow line indicates the dilution up to which visible killing was consistently observed. HPL, high persistent-like; IR, incomplete resistant; R, resistant.
- FIG 4
Dynamics of tailocin survival for the high persistent-like (HPL) mutant. Cultures were treated with various tailocin doses, and viable cells pre- and posttreatment were enumerated. Means and standard errors from three independently repeated experiments with at least three biological replicates per experiment are reported. More killing occurred at high tailocin concentrations, and no difference in stationary and log phases was observed. P values of >0.05 indicate no significant differences within grouped bars, as analyzed in SAS 9.4 with proc Glimmix.
- FIG 5
Genomic mapping of mutations and in planta fitness of selected mutants. (A) Map of predicted lipopolysaccharide O-antigen biogenesis region (PSPPH_0945 to PSPPH_0966) of Pph that showed pronounced effects in tailocin persistence and resistance. Refer to Table 1 for a complete list of genes at this and other genomic locations. (B) Strains and mutants were syringe infiltrated into green bean primary leaves. Experiments were repeated at least twice with 8 biological replications per strain. A representative experiment is presented. Error bars indicate standard error of the mean. Different letters indicate a significant difference (P < 0.05) for a given time point. Pph, wild-type strain; ΔhrpL::Pph, Pph ΔhrpL type III secretion system mutant; HPL, high persister-like; IR, incomplete resistant; R, complete resistant.
- FIG 6
A visual representation of the various tailocin surviving subpopulations and their phenotypes upon exposure to tailocin.
Tables
- TABLE 1
Tailocin sensitivity, predicted loci of mutation, and effect on target gene of tailocin high persistent and resistant mutants and selected allele-swapped strains
↵a WT, wild type; HPL, high persister-like; IR, incomplete resistant; R, resistant; *, indicates allele-swapped strains to confirm mutant phenotype.
Supplemental material
- Supplemental file 1 -
Tables S1 and S2 and Fig. S1 to S7
PDF, 1.3M
- Supplemental file 1 -
