Volatile Organic Compounds Inhibition of Legionella

This assay was used for environmental bacterial isolates which totally inhibit the growth of L. pneumophila on agar plate. A 6-well plate assay was thus designed to physically separate environmental isolates from L. pneumophila Lens in order to determine whether emitted volatiles organic compounds (VOCs) could inhibit or not the growth of L. pneumophila. Each well was filled with 5 mL of BCYE. Then, 10 μL of a suspension of GFP-expressing L. pneumophila Lens (Bigot et al., 2013 (link)) (OD_600nm adjusted to 0.1) were spotted onto both upper sides of the plate. Finally, 40 μL of a suspension of selected isolate (OD_600nm adjusted to 1) were spotted onto the upper center of the plate. Depending of the tested isolate, plates were incubated for 48 h at 22, 30, or 37°C. A second step of incubation was done for 48 h at 37°C. Anti-Legionella activity mediated by VOCs production was quantified by measuring the fluorescence of GFP with a TriStar² LB 942 Microplate Reader (Berthold Technologies, Bad Wildbad, Germany).