Protozoan predation of Escherichia coli in hydroponic media of leafy vegetables

Abstract

ABSTRACT Multiple outbreaks of food poisoning associated with fresh vegetable consumptions have occurred in many countries. Numerous reports have described human pathogenic bacteria, such as Escherichia coli O157:H7 and Salmonella spp., that can internalize into fresh vegetables via root or leaf surfaces. While attempting to obtain the threshold concentration of internalization of E. coli inoculated into hydroponic medium during vegetable cultivation, we observed a rapid decrease in E. coli numbers. In the present study, we determined that the rapid decline in E. coli was not due to a physiological change into a viable but non-culturable (VNC) state. The population crash was instead caused by true bacterial death, as the rapid descent was also confirmed by micro-colony fluorescence in situ hybridization, a culture-independent method that can detect VNC cells. We next monitored the number of E. coli inoculated into intact or filter-sterilized hydroponic medium after cultivation of various types of plants. We found that the number of E. coli in intact hydroponic medium decreased markedly, whereas the level in filter-sterilized hydroponic medium was completely unchanged. This result suggests that biotic factors were present that could be eliminated by filtering. Robust predation of E. coli by protozoa (ciliates and flagellates) was observed using fluorescently labeled bacteria incorporated into the hydroponic medium. Finally, morphological identification of flagellates by scanning electron microscopy revealed the presence of a species of Stramenopiles. These findings suggest the importance of protozoa as bacterial feeders in hydroponic systems and hence the use of these organisms as potential control agents of human pathogenic bacteria.