Genetically engineered bacterial strains constructed as a whole-cell biosensor for specific volatiles identification of infected potato tubers with a soft rot disease

Abstract

The potato (Solanum tuberosum) is the most globally popular non-cereal crop. During postharvest storage, the potato is susceptible to various diseases. Specifically, the symptoms of tuber soft rot can be identified visibly only after irreversible damage has already been caused. The existing technologies are not optimized for the identification of hidden diseases, thus, an early and sensitive diagnostic tool is needed. An attractive early-stage technology is based on monitoring the volatile organic compounds (VOCs) as indicators of pathogen presence. In this study, genetically engineered bacterial strains were constructed as a whole-cell biosensor for specific volatiles identification of infected potato tubers with soft rot disease. Identified volatiles were chosen (i.e., 1-Octanol, 2-Phenylethyl Alcohol, and 1-Octen-3-ol) as model toxicants and used for the identification of the responsive promoters in the host cell Escherichia coli through RNA sequencing and bioinformatic analysis. Furthermore, the promoter activity was verified by a fusion to luciferase-harboring plasmid. The bioluminescent bacterial cells monitor the changes in the VOC profile, and a signal change is then generated, reflecting the infection status of the potatoes. The efficacy of the recombinant bacteria to monitor potato soft rot disease was investigated in solution, surrounding air near pure volatiles, and around potato tubers contaminated by Pectobacterium. The results demonstrated that the new strains were sensitive to the presence of these VOCs, each to a different extent and in a different environment (solution or surrounding air). All of them except oxyR could detect differences between infected and uninfected potato tubers. The preferred bioreporter to detect Pectobacterium infection in whole potato tubers is dnaK. This strain responds to all tested volatiles in the liquid and gaseous states and has shown the highest response differences as a reaction to the infection presence in the whole potato tubers. Bacterial monitoring of the volatile profile results in identifying specific signatures that will allow quick and accurate detection of the presence of pathogens in stored crops, prior to visible disease symptoms.