Biological assessment of disease resistance in plants often relies on subjective visual assessment by experts, is costly and time consuming, and may be subject to high environmental and experimental variation resulting in low heritability. In vivo imaging technology is a non-invasive, rapid and unbiased tool that can be used to study the plant host-pathogen interaction and assess disease severity. We developed a strain of X. gardneri that expresses bioluminescence (lux operon) as a way to visualize and quantify infection. The objectives of this study were to determine the correlation between luminescence signals and bacterial populations and to make comparisons between resistant and susceptible tomato lines. Four week-old tomato seedlings were sprayed with a bacterial suspension (10⁸ CFU/ml) and kept under high humidity. Leaflets were excised and luminescence signals were measured at 3, 4, 5, 8, and 9 days post-inoculation (dpi). Bacterial counts were estimated using serial dilution plating after each time point. Regression analysis displayed a positive relationship between bacterial counts and luminescence in infected plant tissue (P= <.0001, R=.71). Tomato lines with different resistance loci were evaluated at 9 dpi in a completely random design. Quantification of bacterial populations using IVIS could distinguish between susceptible and resistant lines and the heritability was 0.24. This study illustrates the use of IVIS for phenotypic evaluation of disease resistance in tomato. Future studies will focus on evaluating larger collections of germplasm for resistance.