Phage-Bacteria Interaction Dynamics

Infection experiments were carried out in glass vials containing 6 ml of M9 media with 0.2% glucose supplemented with 100 µM Baicalein (Cayman Chemical) dissolved in DMSO, and 10⁶ cfu/ml P. aeruginosa WT. The same experiment was carried out concurrently with the QS mutant in the presence or absence of 2 µM 3OC12-HSL and 10 µM C4-HSL (Sigma) dissolved in DMSO. Control treatments received the same amount of DMSO. Total amounts of 10⁴ plaque forming units (pfu) (low phage), 10⁷ pfu (mid phage) or 10⁹ pfu (high phage) were added at the same time as the bacteria. Microcosms were incubated at 37 °C while shaking at 180 rpm. Every 24 h microcosms were sampled and transferred to fresh media at a 1:100 dilution. Samples from the infected cultures were taken prior to each transfer and chloroform treated to kill bacteria as described previously [38 (link)]. 5 µl of a serial dilution of chloroform treated sample was spotted onto a lawn of sensitive bacteria (CRISPR-KO strain). Plaques were counted and densities were calculated as pfu/ml. Bacteria were sampled at 3 days post infection to analyse the resistance mechanisms that evolved in response to phage infection.