The commensal/pathogenic Escherichia coli affects humans and animals, being present in diverse environmental niches, possibly surviving due to its adaptation to transient plant hosts like crops, increasing the risk of foodborne diseases. E. coli interaction with the plant host remains unknown, particularly the impacts on photosynthesis. We hypothesize that E. coli influences the tomato transient host’s photosynthetic capacity. To validate this hypothesis, we exposed 57-day-old tomato plants (Solanum lycopersicum) to different inoculation conditions, namely, non-inoculated plants (negative control, C−); plants directly injected with E. coli SL6.1 (107 CFU/mL) (positive control, C+); plants irrigated one time with E. coli SL6.1 (107 CFU/mL); and plants chronically irrigated with E. coli SL6.1 (104 CFU/mL). No significant changes were observed in chlorophyll fluorescence, pigments’ contents, morphological aspects, and fruiting in all conditions. However, irrigated plants (chronically and one-time contaminated) had decreased stomatal conductance (gs, 31.07 and 34.42 mol m−2 s−1, respectively, vs. 53.43 and 48.08 mol m−2 s−1 in C− and C+, respectively), transpiration rate (E, 0.32 and 0.35 mol m−2 s−1 in chronically and one-time contaminated conditions vs. 0.57 and 0.48 mol m−2 s−1 in C− and C+, respectively), and a trend of increased intrinsic carboxylation (Ci, 384 and 361 ppm in chronically and one-time irrigated plants vs. 321 and 313 ppm in C− and C+, respectively). The one-time inoculated plants presented more severe effects than the remaining conditions, with lower net photosynthetic rate (PN, 0.93 vs. 3.94–5.96 μmol (CO2) m−2 s−1 in the other conditions), intrinsic water use efficiency (iWUE, 33.1 vs. 74.51–184.40 μmol (CO2)/ mmol (H2O) in the chronically irrigated and the control plants), and intrinsic carboxylation efficiency (iCE, 0.003 vs. 0.012–0.022 μmol (CO2)/ppm in the remaining conditions). Our data support that some observed effects are similar to those associated with phytopathogenic bacteria. Lastly, we propose that the decrease in some parameters of gas exchange requires direct contact with the leaf/stomata, and is mainly observed for high concentrations of E. coli.