Table 1.
Bacterial strains and primers used in this study.
Fig 1.
Production of non-attached biofilm and eDNA by 30-84ZN (no phenazine) 30-84PCA, 30–84 wild type, and 30-84O*.
(A) Bacteria were grown in AB minimal media + 2% casamino acid in static plates for up to 72 h. Biofilm matrix production by P. chlororaphis 30–84 and derivatives at 48 and 72 h was quantified by weight. (B) Visualization of biofilm formed after culturing in static plates for 72 h and being collected by centrifugation. (C) Production of eDNA by different derivatives was quantified using the double stranded DNA quantifying fluorescent dye assay from Invitrogen. The data are the average of three biological replicates and error bars indicate the standard deviation.
Fig 2.
Production of biofilm matrix and eDNA in the presence or absence of DNase I.
(A) Biofilm production after bacterial strains were grown in static plates in AB minimal media + 2% casamino acid either with or without DNase I for 72 h and biofilms were collected by centrifugation. Data are the weight of the biofilm matrix. (B) Production of eDNA in the presence or absence of DNase I in the same experiment. The data represent the average of at least three biological replicates and error bars indicate the standard deviation.
Fig 3.
The number of differentially expressed genes in 30-84PCA, wild type and 30-84O* compared with the 30-84ZN.
Differential expressed genes are those exhibiting over twofold change and a P value < 0.05. (A) Phenazine Induced and Suppressed genes are those that are expressed in at a higher or lower level, respectively, by the phenazine producing strains compared to 30-84ZN. (B) Venn diagram showing the number of genes differentially expressed in 30-84PCA, wild type and 30-84O* compared with 30-84ZN.
Table 2.
Selected genes that are differentially expressed by all phenazine producers: e.g. by 30-84PCA, wild type, and 30-84O*compared with 30-84ZN.
Table 3.
Genes annotated as R2 type pyocin genes belonging to a 13 Kb open reading frame spanning a bacteriophage-derived gene cluster from Pchl3084_1194–1233.
Fig 4.
The effect of holin mutation on eDNA and biofilm matrix production.
(A) Production of eDNA by P. chlororaphis 30–84 wild type and the holin mutant after growing 48 and 72 hr in static culture. (B) Biofilm matrix production by 30–84 wild type and the holin mutant at 72 h. The data are the average of three biological replicates and error bars indicate the standard deviation.
Table 4.
Selected genes that were changed in expression (relative to 30-84ZN) only in wild type and 30-84O* biofilm matrices, e.g. are specifically affected by 2-OH-PCA production.
Fig 5.
Possible metabolic fates for chorismic acid in P. chlororaphis 30–84.
Chorismic acid is produced via the shikimic acid pathway and is a metabolic precursor for a number of different end products in pseudomonads, including phenazines (indicated by solid lines). The negative symbols (-) indicate competing pathways in which transcript levels for key genes are reduced in 2-OH-PCA producing strains compared to 30-84ZN (Table 4).