Phytotoxic Tryptoquialanines Produced In Vivo by Penicillium digitatum Are Exported in Extracellular Vesicles

Author(s):Show less -

Costa, Jonas Henrique ^[1] ; Bazioli, Jaqueline Moraes ^{[2, 1]} ; Barbosa, Luidy Darllan ^[1] ; Theodoro dos Santos Junior, Pedro Luis ^[1] ; Reis, Flavia C. G. ^{[3, 4]} ; Klimeck, Tabata ^[3] ; Crnkovic, Camila Manoel ^[5] ; Berlinck, Roberto G. S. ^[6] ; Sussulini, Alessandra ^[1] ; Rodrigues, Marcio L. ^{[3, 7]} ; Fill, Taicia Pacheco ^[1]

Affiliation

^[1] Univ Estadual Campinas, Inst Chem, CP 6154, Campinas, SP - Brazil

^[2] Univ Estadual Campinas, Fac Pharmaceut Sci, Campinas, SP - Brazil

^[3] Fundacao Oswaldo Cruz FIOCRUZ, Inst Carlos Chagas, Curitiba, Parana - Brazil

^[4] Fiocruz MS, Ctr Desenvolvimento Tecnol Saude CDTS, Rio De Janeiro - Brazil

^[5] Univ Sao Paulo, Sch Pharmaceut Sci, Dept Biochem & Pharmaceut Technol, Sao Paulo, SP - Brazil

^[6] Univ Sao Paulo, Inst Quim Sao Carlos, CP 780, Sao Carlos, SP - Brazil

^[7] Univ Fed Rio de Janeiro, Inst Microbiol Paulo Goes IMPG, Rio De Janeiro, RJ - Brazil

• Document type: Journal article
• Source: MBIO; v. 12, n. 1 JAN-FEB 2021.
• Web of Science Citations: 4

Abstract

Penicillium digitatum is the most aggressive pathogen of citrus fruits. Tryptoquialanines are major indole alkaloids produced by P. digitatum. It is unknown if tryptoquialanines are involved in the damage of citrus fruits caused by P. digitatum. To investigate the pathogenic roles of tryptoquialanines, we initially asked if tryptoquialanines could affect the germination of Citrus sinensis seeds. Exposure of the citrus seeds to tryptoquialanine A resulted in a complete inhibition of germination and an altered metabolic response. Since this phytotoxic effect requires the extracellular export of tryptoquialanine A, we investigated the mechanisms of extracellular delivery of this alkaloid in P. digitatum. We detected extracellular vesicles (EVs) released by P. digitatum both in culture and during infection of citrus fruits. Compositional analysis of EVs produced during infection revealed the presence of a complex cargo, which included tryptoquialanines and the mycotoxin fungisporin. The EVs also presented phytotoxicity activity in vitro and caused damage to the tissues of citrus seeds. Through molecular networking, it was observed that the metabolites present in the P. digitatum EVs are produced in all of its possible hosts. Our results reveal a novel phytopathogenic role of P. digitatum EVs and tryptoquialanine A, implying that this alkaloid is exported in EVs during plant infection. IMPORTANCE During the postharvest period, citrus fruits can be affected by phytopathogens such as Penicillium digitatum, which causes green mold disease and is responsible for up to 90% of total citrus losses. Chemical fungicides are widely used to prevent green mold disease, leading to concerns about environmental and health risks. To develop safer alternatives to control phytopathogens, it is necessary to understand the molecular basis of infection during the host-pathogen interaction. In the P. digitatum model, the virulence strategies are poorly known. Here, we describe the production of phytotoxic extracellular vesicles (EVs) by P. digitatum during the infection of citrus fruits. We also characterized the secondary metabolites in the cargo of EVs and found in this set of molecules an inhibitor of seed germination. Since EVs and secondary metabolites have been related to virulence mechanisms in other host-pathogen interactions, our data are important for the comprehension of how P. digitatum causes damage to its primary hosts. (AU)

• FAPESP's process: 17/24462-4 - Understanding the virulence strategies of Penicillium digitatum against the host citrus
• Grantee: Taicia Pacheco Fill
• Support Opportunities: Regular Research Grants
• FAPESP's process: 13/50228-8 - Biodiversity components, and their metabolic characters, of Brazilian Islands: an integrated approach
• Grantee: Roberto Gomes de Souza Berlinck
• Support Opportunities: BIOTA-FAPESP Program - Thematic Grants
• FAPESP's process: 19/06359-7 - Understanding the chemical factors involved in the endophytism x pathogenicity of different Penicillium brasilianum isolates
• Grantee: Taicia Pacheco Fill
• Support Opportunities: Regular Research Grants
• FAPESP's process: 15/01017-0 - Multi-use instrument approved and awarded in the project 2013/50228-8, name of instrument: ultra-performance liquid chromatograph coupled with a high resolution mass spectrometer
• Grantee: Roberto Gomes de Souza Berlinck
• Support Opportunities: Multi-user Equipment Program
• FAPESP's process: 19/11563-2 - Prospecting for anticancer secondary metabolites produced by phytopathogenic fungi
• Grantee: Jaqueline Moraes Bazioli
• Support Opportunities: Scholarships in Brazil - Doctorate (Direct)