Combined microscopy and molecular analyses show phloem occlusions and cell wall modifications in tomato leaves in response to ‘Candidatus Phytoplasma solani’

• Published in: Journal of microscopy (Oxford) Vol. 263; no. 2; pp. 212 - 225
• Main Authors: MARCO, F. DE, PAGLIARI, L., DEGOLA, F., BUXA, S. V., LOSCHI, A., DINANT, S., HIR, R. LE, MORIN, H., SANTI, S., MUSETTI, R.
• Format: Journal Article
• Language: English
• Published: England Wiley Subscription Services, Inc 01.08.2016; Wiley
• Subjects: Callose; Cell Wall - metabolism; Cell Wall - microbiology; Flowers & plants; Glucans - metabolism; Glucosyltransferases - genetics; Glucosyltransferases - metabolism; Infections; Integrated approach; Leaves; Life Sciences; Lycopersicon esculentum - cytology; Lycopersicon esculentum - enzymology; Lycopersicon esculentum - metabolism; Lycopersicon esculentum - microbiology; Microscopy - methods; phloem; Phloem - cytology; Phloem - metabolism; Phloem - microbiology; phytoplasma; Phytoplasma - pathogenicity; Phytoplasma - physiology; Plant Leaves - cytology; Plant Leaves - enzymology; Plant Leaves - metabolism; Plant Leaves - microbiology; Proteins; P‐proteins; tomato; phloem; phytoplasma; tomato; P-proteins; Callose
• ISSN: 0022-2720; 1365-2818
• DOI: 10.1111/jmi.12426

Summary Callose deposition, phloem‐protein conformational changes and cell wall thickening are calcium‐mediated occlusions occurring in the plant sieve elements in response to different biotic and abiotic stresses. However, the significance of these structures in plant–phytoplasma interactions requires in‐depth investigations. We adopted a novel integrated approach, based on the combined use of microscopic and molecular analyses, to investigate the structural modifications induced in tomato leaf tissues in presence of phytoplasmas, focusing on vascular bundles and on the occlusion structures. Phloem hyperplasia and string‐like arrangement of xylem vessels were found in infected vascular tissue. The diverse occlusion structures were differentially modulated in the phloem in response to phytoplasma infection. Callose amount was higher in midribs from infected plants than in healthy ones. Callose was observed at sieve plates but not at pore‐plasmodesma units. A putative callose synthase gene encoding a protein with high similarity to Arabidopsis CalS7, responsible for callose deposition at sieve plates, was upregulated in symptomatic leaves, indicating a modulation in the response to stolbur infection. P‐proteins showed configuration changes in infected sieve elements, exhibiting condensation of the filaments. The transcripts for a putative P‐protein 2 and a sieve element occlusion‐related protein were localized in the phloem but only the first one was modulated in the infected tissues. Lay description Phytoplasmas are plant‐pathogenic prokaryotes belonging to the class Mollicutes, a group of wallless microorganisms phylogenetically related to low G+C Gram‐positive bacteria. They have been associated with several hundred diseases affecting economically important crops, such as ornamentals, vegetables, fruit trees and grapevine. In host plants, phytoplasmas are restricted to the sieve tubes of the phloem and are transmitted among plants by phloem sap feeding leafhoppers or psyllids in a persistent manner. Phytoplasmas are obligate intracellular parasites, inducing characteristic symptoms in host plants, such as low growth rate, stunting, yellowing or reddening of the leaves, reduced leaf size, shortening of internodes and loss of apical dominance. These lead to reduced yields, proliferation of shoots or roots, witches’ brooming, general decline and, sometimes, death of the plant. Several symptoms affect flowers, including virescence, phyllody and sterility. Despite their economic incidence, phytoplasmas remain as the most poorly characterized plant pathogens, primarily because efforts at in vitro culture, gene delivery and mutagenesis have been unsuccessful. Alteration in phloem function is one of the most dramatic effects of phytoplasma infection in plants because it leads to the impairment of assimilate translocation. Histological studies on several plant species infected by phytoplasmas showed that the first detectable anatomical aberration is an abnormal deposition of callose in the phloem regions which might be followed by a collapse of sieve elements and companion cells. Phloem occlusion has been described as a defence mechanism leading to the formation of physical barriers aimed to achieve pathogen containment. Nevertheless, almost nothing is understood about the intimate events occurring between phytoplasmas and phloem cells. The presented data give, for the first time, a precise, punctual description about the different sites, in which occlusions occur in tomato leaf tissue. The results described here are obtained by using an integrated approach, based on the combined use of microscopic and molecular analyses.