Microbial Tracking During Plant Growth Using Fluorescence Reporter Strains

This work implemented the microbial seed coating technique to apply a plant beneficial microbe, Burkholderia contaminans (B. contaminans), to a Kennedy Space Center (KSC) tomato variety (Solanum lycopersici) of space crop to assess an ability to mitigate a Fusarium wilt infection (by Fusarium oxysporum f. sp. lycopersici (FOL)) of the tomato plant. Fusarium wilt is a common tomato plant pathogenic infection. We pursued this study due that fact that tomatoes are commonly grown in closed environment agriculture settings on Earth and have recently become the subject of space crop science investigations on the ISS. Furthermore, Fusarium oxysporum is a fungal plant pathogen that presented itself on the ISS upon the first tech demo growth experiment of Zinnia flowers suggesting there may be a future potential risk for tomato crops to be similarly affected. For this current study, we strove to evaluate a countermeasure for FOL infection specific to tomato plants, and to understand where the microbes travel throughout the plant so that methods for the proper containment of the introduced microbes could be understood. Here, a fluorescently labeled B. contaminans, Pseudomonas fulva (P. fulva), and Panteoa agglomerans (P. agglomerans) strains were employed to allow for microbial enumeration and visual documentation of the retention for our introduced strain on the host plant using fluorescence-based microscopy. The B. contaminans strain was evaluated for the ability to inhibit FOL and was found effective in a plate-based assay. We observed that B. contaminans applied to the roots could subsequently be found on the shoots of plants at 28 days although in much lower abundance. In addition, we assessed commercially available food grade probiotics known for their benefit to the human gut for their added plant beneficial traits, but we were unable to successfully transform the fluorescence reporter into these gram-positive strains, and none showed inhibition of fungal growth in the plate-based assay. Here we deliver a summary of the effective dosage of B. contaminans that was applied to the tomato seed using the seed film technique and the controlled environment testing of tomato plants over three grow outs that were conducted with a combination of applied seed film, B. contaminans, and FOL to test the effectiveness of the seed coat to counter FOL infection. It was found that the plate-based assay using B. contaminans demonstrated efficacy as an anti-fungal agent; however, the practical application using tomato may have been limited within our sealed vessel growth set up and did not confer any discernable mitigation of FOL infection. Future work could look to optimize growth setting parameters that are more like actual tomato growth conditions in closed environment agriculture to further test for fungal-protective effects, but any effort would still need to account for the proper containment and handling of the FOL organism in KSC labs.