Visualizing pathogen internalization pathways in fresh tomatoes using MicroCT and confocal laser scanning microscopy

Authors: Zhou, Bin; Luo, Yaguang - Sunny; Bauchan, Gary; FENG, H - University Of Illinois; Stommel, John

Submitted to: Food Control
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 9/26/2017
Publication Date: 9/27/2017
Citation: Zhou, B., Luo, Y., Bauchan, G.R., Feng, H., Stommel, J.R. 2017. Visualizing pathogen internalization pathways in fresh tomatoes using MicroCT and confocal laser scanning microscopy. Food Control. 85:276-282.

Interpretive Summary: In packing houses, tomatoes are typically washed in dump tanks to remove debris and soil, and maintain quality and shelf life. However, pathogens may enter the tomato during the washing process due to infiltration through the tomato stem scar. Once internalized in the fruit, it is impossible to remove or inactivate the pathogens without compromising the quality. In this study, the authors used novel a technology to visualize the pathway of pathogen internalization in the tomato fruit. The results showed that pathogens were confined in xylem vessels and concentrated immediately beneath the stem-scar, and that pathogen cell populations declined with vertical distance from the stem-scar. The findings are anticipated to facilitate the produce industry in developing science-based food safety practices and interventions for controlling pathogen internalization and improving food safety.

Technical Abstract: Pathogen contamination of fresh produce significantly impacts public health and the produce industry's economic well-being. In tomato fruits, studies have shown that the stem-scar plays an important role in pathogen infiltration. However, the exact mechanisms and pathways for pathogen movement inside tomatoes are not well documented. This study examined in detail the pathway of pathogen internalization in tomato fruit. Tomato fruit were infused with potassium iodine and examined with Xradia Bio MicroCT at 0.5× ~ 4×. Micro CT images revealed that infiltrated potassium iodine moved along the vascular bundles inside the fruit. This result led us to investigate whether vascular tissues could be a similar means for pathogenic Escherichia coli infiltration. Tomato fruits were infiltrated with red fluorescent microspheres and E. coli O157:H7 -pGFP. Stem scars and core tissues were subsequently excised after fruit surface disinfection and observed using a Zeiss 710 laser confocal microscope, or stomached, cultured, and enumerated for infiltrated E. coli O157:H7. Populations of internalized E. coli O157:H7 were confined in xylem vessels and concentrated immediately beneath the stem-scar, with a sharp decline in population with vertical distance from the stem-scar. Our observations suggest that 1) vascular bundles, especially the xylem vessels in tomato fruit, play a critical role in pathogen internalization, and 2) pathogen internalization is a passive, rather than active process, as the infiltration and movement of both microspheres (non-living) and E. coli O157:H7 cells (living) behaved similarly inside tomatoes. This study presents the first visual evidence of the critical role of vascular bundles in pathogen internalization in tomato fruit. The finding is important for developing science-based food safety practices, interventions in controlling pathogen internalization, and insight for developing pathogen internalization resistant tomato varieties.