Isolated colonies of CV026 (LB agar – 48 hours at 28 °C) were used to make a bacterial suspension in LB medium adjusted to 2.0 McFarland scale. The bacterial suspension was used to inoculate the test medium in a proportion of 10% v/v. The test medium was then supplemented with the cognate autoinducer (N-hexanoyl-l -homoserine 96% lactone, Sigma-Aldrich), at a final concentration of 10 µM. A 1.95 mL aliquot of this culture was transferred to a conical falcon tube (v = 15 mL) and supplemented with 50 µL of the tested molecules in an acetone solution of 25%, in different final concentrations (2.5 mM, 1.25 mM, 625 µM and 312 µM). The assays were incubated under agitation at 200 rpm at 30 °C for 24 hours and then were put in a kiln to completely evaporate the liquid medium. Violacein was solubilized from dried bacterial pellets with 1 mL of DMSO. After 24 h of agitation, the absorbance of soluble violacein was measured at 630 nm using a microplate reader (Biotek – ELx808™).
The same experimental conditions were repeated, in parallel, in the absence of the cognate auto-inducer (“blank”) for evaluating the bacterial growth by measuring turbidity (630 nm) after 24 hours under agitation at 200 rpm and 30 °C in a microplate reader (Biotek – ELx808™).
The same experimental conditions were repeated, in parallel, in the absence of the cognate auto-inducer (“blank”) for evaluating the bacterial growth by measuring turbidity (630 nm) after 24 hours under agitation at 200 rpm and 30 °C in a microplate reader (Biotek – ELx808™).