ABSTRACT
Phytophthora parasitica is an oomycete pathogen that infects a broad range of crops of worldwide economic interest; among them are citrus species. In general, some Citrus and the rootstocks of related genera offer considerable resistance against P. parasitica; therefore, understanding the mechanisms involved in the virulence of this pathogen is crucial. In this work, P. parasitica secondary metabolite production was studied using matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI) and ultrahigh-performance liquid chromatography coupled with electrospray ionization quadrupole time-of-flight tandem mass spectrometry (UHPLC/ESI-Q-TOF-MS) combined with chemometric tools, and its metabolic profile was evaluated under the influence of Citrus sunki (a highly susceptible host) and Poncirus trifoliata (a resistant genotype) extracts. The root extracts of Citrus sunki had an influence on the growth and hyphae morphology, and the root extracts of P. trifoliata had an influence on the zoospore behavior. In parallel, the spatial distribution of several metabolites was revealed in P. parasitica colonies using MALDI-MSI, and the metabolite ion of m/z 246 was identified as the protonated molecule of Arg-Ala. The MALDI-MSI showed variations in the surface metabolite profile of P. parasitica under the influence of the P. trifoliata extract. The P. parasitica metabolome analysis using UHPLC-ESI-Q-TOF-MS resulted in the detection of Arg-Gln (m/z 303.1775), as well as L-arginine (m/z 175.1191) and other unidentified metabolites. Significant variations in this metabolome were detected under the influence of the plant extracts when evaluated using UHPLC-ESI-Q-TOF-MS. Both techniques proved to be complementary, offering valuable insights at the molecular level when used to assess the impact of the plant extracts on microbial physiology in vitro. The metabolites identified in this study may play significant roles in the interaction or virulence of P. parasitica, but their functional characterization remains to be analyzed. Overall, these data confirm our initial hypotheses, demonstrating that P. parasitica has the capabilities of (i) recognizing host signals and altering its reproductive programing and (ii) distinguishing between hosts with varying responses in terms of reproduction and the production of secondary metabolites.
ABSTRACT
The bacteria of the genus Curtobacterium are usually seen as plant pathogen, but some species have been identified as endophytes of different crops and could as such present a potential for disease control and plant growth promotion. We have therefore applied the desorption electrospray ionization mass spectrometry imaging (DESI-MSI) in the direct analysis of living Curtobacterium sp. strain ER1/6 colonies to map the surface metabolites, and electrospray ionization tandem mass spectrometry (ESI-MS/MS) for characterization of these compounds. Several colony-associated metabolites were detected. The ESI-MS/MS showed characteristic fragmentations for phospholipids including the classes of glycerophosphocholine, glycerophosphoglycerol, and glycerophosphoinositol as well as several fatty acids. Although a secure identification was not obtained, many other metabolites were also detected for this bacteria species. Principal component analysis showed that fatty acids were discriminatory for Curtobacterium sp. ER1/6 during inoculation on periwinkle wilt (PW) medium, whereas phospholipids characterize the bacterium when grown on the tryptic soy agar (TSA) medium.
