FIGURE 1: Construction and characterization of L. pneumophila strain encoding the LuxR operon. (a) Construction of the Lp icmRp–LuxR strain via single homologous recombination event with the suicide plasmid pSR47::icmRp–LuxR, which encodes the LuxR operon cloned downstream of a 863 nt region of homology containing the icmR promoter region from the Lp01 strain. (b) Luminescence output of the LuxR-encoding Lp as compared to the parental strain. Simultaneous luminescence and optical density measurements are shown from serial dilutions prepared from two-day heavy patches grown on CYA plates. (c) Luminescence output of the Lp icmRp–LuxR measured every ten min in axenically grown AYE liquid cultures in the presence/absence of chloramphenicol over the indicated time period. Each time point represents an average of a technical triplicate. (d) Bacterial growth of Lp icmRp–LuxR measured from bioluminescence output when bacteria were cultured in the absence or presence of BMDMs. (e) Growth comparison between in the indicated Lp strains in BMDMs infections measured by a CFU growth assay. (f) Growth kinetics of Lp icmRp–LuxR is shown for various MOIs. A vertical line indicates the peak of the growth curve for each infection. (g) The peak time and amplitude of the growth curve for each MOI from (f) are shown. (d-g) Each data point represents an average from technical triplicates ± SD. (b-g) Representative data from one of three biological replicates are shown.