Abstract
Dieback, caused by Erwinia psidii is currently one of the most severe emerging diseases of Eucalyptus spp. in Brazil. Because of its recent report, little is known about the mechanisms underlying bacterial infection. We studied the colonization and movement of E. psidii in host tissue using a strain labeled with green fluorescent protein. We were able to transform E. psidii with pGreen-TIR and to demonstrate plasmid stability in the absence of antibiotic selection both in vitro and in vivo. We demonstrate that tissue colonization by E. psidii is not restricted to the inoculation point (leaf axil). E. psidii colonizes the xylem vessels, sclerenchyma and parenchyma of leaves and stems of eucalypt. At 35 days after inoculation, the bacterium was found at 5 cm above the inoculation point, indicating that it colonizes the plant acropetally. Confocal microscopy analysis revealed that when root-inoculated the bacterium penetrates the primary and secondary roots and reaches the xylem, but it was never found in the root crown or stem, irrespective of the evaluation time. Our results provide insights into the biology of the E. psidii-eucalypt interaction, which requires a better understanding in order to design efficient strategies for pathogen control and disease management.
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Acknowledgements
The authors are grateful to CMPC Celulose Riograndense, Fibria, FAPEMIG and CNPq for financial support. The authors thank Doctor Alan Collmer, Plant Pathology and Plant-Microbe Biology Section at Cornell University, for providing the pGreen-TIR plasmid; Nucleus of Microscopy and Microanalysis at the Universidade Federal de Viçosa and Clonar Resistência a Doenças Florestais for the propagation of the plant material used in the inoculation experiments; the Organization of American States (OAS) and Coimbra Group of Brazilian Universities (GCUB) for Doctoral Scholarships to C. N. Montoya-Estrada.
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Figure S1.
Fluorescence of Erwinia psidii LPF534R expressing the pGreen-TIR plasmid. Bacterial cells were observed by fluorescence microscopy (100x) with filter of 470 to 490 nm. (PNG 3199 kb)
Figure S2.
Stability of the pGreen-TIR plasmid in Erwinia psidii LPF534R. (a) Stability of the pGreen-TIR plasmid in Erwinia psidii LPF534R determined as the percentage of the number of colony forming units (CFU) on culture medium containing gentamycin compared to the number of CFU on culture medium without the antibiotic. The experiment was carried out with three biological and two technical repetitions. (b) Fluorescence of Erwinia. psidii LPF534R transformed with pGreen-TIR on solid LB medium amended with gentamicin, visualized with a bench UV-lamp. (JPG 1004 kb)
Figure S3.
Roots of eucalypt clone CLR360 inoculated with Erwinia psidii LPF534T by root immersion. (PNG 1736 kb)
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Montoya-Estrada, C.N., Costa, C.R., Badel, J.L. et al. Root infection and aerial colonization of eucalypt host plants by Erwinia psidii. Trop. plant pathol. 44, 251–257 (2019). https://doi.org/10.1007/s40858-018-0264-7
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DOI: https://doi.org/10.1007/s40858-018-0264-7