PQS signaling under anaerobic conditions. (A) PQS production in the supernatant of aerobic, anaerobic, or oxygen-limited cultures. PQS was detected by TLC analysis. Lane 1, synthetic PQS; lane 2, aerobic culture; lane 3, anaerobic culture; lane 4, oxygen-limited culture with LBN medium; lane 5, synthetic PQS. (B) Comparison of PQS production between aerobic, anaerobic, and oxygen-limited cultures. Bar 1, aerobic culture; bar 2, anaerobic culture; bar 3, oxygen-limited culture with LBN medium. PQS production was compared by measuring the band intensity of the TLC plates with an image analyzer. ND, not detected. (C) pqsA expression in PAO1 and a ΔpqsR mutant under anaerobic conditions. pMEXpqsA or pMEX9 was introduced in each cells. C23O activity (nanomoles of product/minute/milligram of protein) was measured in cells cultured for 12 h (stationary phase). C23O activity of pMEXpqsA-introduced cells was measured and was normalized by the C23O activity of pMEX9-introduced cells. Data displayed are means ± standard deviations of three independent experiments.