Blossom End Rot Tomato Fruit Diagnosis for <i>In Situ</i> Cell Analyses with Real Time Pico-Pressure Probe Ionization Mass Spectrometry

Abstract

By combining a cell pressure probe and an Orbitrap mass spectrometer, quantitative snapshot profiles of metabolites of in situ plant single cells during development of blossom end rot in tomato (Solanum lycopersicum L.) fruit were analyzed while tomato plants were grown hydroponically in a greenhouse. By using the pressure probe, cell turgor, cell volume, cell wall elastic modulus, and hydraulic conductivity of plasma membrane were measured, followed by managed cytoplasm sampling, and osmotic and water potentials determination. It was found that the cell bursting results from alterations in water relations and cell wall properties accompanied with changes in metabolites of cells located at blossom end rot area in tomato fruit. From the water relations point of view, the loss of the elasticity and cell wall weakening, with decreased water potential and excess turgor resulted in mechanical rupture of membrane and cell wall. Abscisic acid was detected in damaged cells as a possible evidence of triggered precocious maturity. Simultaneously, a sharp rise in the concentration of phenols (coumarinate-glucoside and chlorogenic acid) and salicylic acid and decline in ascorbic acid reflected the activation of cell death process that would facilitate the deterioration of cell wall and plasma membrane.

Advances in mass spectrometry based single-cell metabolomics

“…Pressure driven microsampling using silicone oil has been used for sampling the cytosol of single cells from tomato plants, enabling the detection of sugars, amino acids etc. 49,50 Since the sampled volume was known, the concentration of some metabolites in the cytosol could be determined by external calibration. 49,50 Nanopipettes have more recently been refined for pressure driven microsampling using double-barrel nanopipettes (theta pipettes) for repeated single-cell analysis.…”

“…49,50 Since the sampled volume was known, the concentration of some metabolites in the cytosol could be determined by external calibration. 49,50 Nanopipettes have more recently been refined for pressure driven microsampling using double-barrel nanopipettes (theta pipettes) for repeated single-cell analysis. 51 Both barrels were filled with perfluorodecalin (PFD), a liquid that is immiscible in both aqueous and organic solvents.…”