BT3560-3563 is required for bile-dependent biofilm formation. (A and C) A 96-well plate crystal violet biofilm assay after 48-h growth in BHIS or BHIS + 0.5% bile extract. (B) Schema of the BT3560-3563 genetic locus. Red arrows indicate transposon insertion points. N/A, not annotated; EEP, exonuclease/endonuclease/phosphatase family. (D) A 96-well plate crystal violet biofilm assay after 48-h growth in BHIS + 0.5% bile extract. WT in BHIS is not represented. (A, C, and D) Mean of WT in BHIS is adjusted to 100%. Min-max boxplot of 6 to 18 biological replicates for each strain, each replicate is the mean of two technical replicates. *P value <0.05, **P value <0.005, ***P value <0.0005, Mann–Whitney U test.