Metabolomic study of fungi of the genus Colletotrichum and tambjamines- producing marine organisms

Author(s)

Mirelle Takaki

• Document type: Doctoral Thesis
• Press: São Carlos.
• Institution: Universidade de São Paulo (USP). Instituto de Química de São Carlos (IQSC/BT)
• Defense date: 2020-04-16

Examining board members

Roberto Gomes de Souza Berlinck; Moacir Rossi Forim; Mario Sergio Palma; Edson Rodrigues Filho

• Advisor: Roberto Gomes de Souza Berlinck

Abstract

<span style=\"font-weight: 400;\">The present study describes the metabolomic investigation of fungi of the genus <span style=\"font-weight: 400;\">Colletotrichum<span style=\"font-weight: 400;\"> and tambjamine-producing marine organisms <span style=\"font-weight: 400;\">Virididentula dentata<span style=\"font-weight: 400;\">, <span style=\"font-weight: 400;\">Tambja stegosauriformis<span style=\"font-weight: 400;\">, <span style=\"font-weight: 400;\">Tambja brasiliensis<span style=\"font-weight: 400;\"> and <span style=\"font-weight: 400;\">Roboastra ernsti<span style=\"font-weight: 400;\">. Fungi belonging to the genus <span style=\"font-weight: 400;\">Colletotrichum<span style=\"font-weight: 400;\"> cause disease in a wide variety of plants from different families. Secondary metabolites produced by these microorganisms can be part of the virulence process associated with their phytopathogenicity. Although many metabolites of <span style=\"font-weight: 400;\">Colletotrichum<span style=\"font-weight: 400;\"> spp. have already been isolated and identified, the phytopathogenicity associated with fungi of this genus is not yet well established. In order to investigate the metabolic differences between endophytic and phytopathogenic strains of <span style=\"font-weight: 400;\">Colletotrichum<span style=\"font-weight: 400;\"> spp., culture media produced by twenty-five isolates of <span style=\"font-weight: 400;\">Colletotrichum<span style=\"font-weight: 400;\"> spp. were investigated, 16 of which were endophytic and 9 were phytopathogenic. The culture media were fractionated and analyzed by UPLC-IMS-QToF-HRMS. The raw data were processed using in-house tools and Molecular Networking. The evaluation of the results allowed us to conclude that endophytic and phytopathogenic strains of <span style=\"font-weight: 400;\">Colletotrichum<span style=\"font-weight: 400;\"> spp. have considerably different metabolism. Endophytic strains produce compounds that are common to several strains in the group; phytopathogenic strains produce compounds more specific to each isolate. Dereplication of the GNPS exported results using an in-house library of compounds isolated from <span style=\"font-weight: 400;\">Colletotrichum<span style=\"font-weight: 400;\"> fungi combined with <span style=\"font-weight: 400;\">in-silico<span style=\"font-weight: 400;\"> fragmentation tools allowed the identification of 5 compounds in samples related to phytopathogenic strains. Those compounds display phytotoxic activity. Such results are a strong indication that the phytopathogenicity of <span style=\"font-weight: 400;\">Colletotrichum<span style=\"font-weight: 400;\"> fungi may be associated with the production of phytotoxic compounds. Tambjamines are a group of alkaloids chemically related to prodigiosin and prodiginine, isolated from bacteria and marine organisms such as ascidians, bryozoans and nudibranchs. In addition to presenting diversified biological activities, tambjaminas are possibly related to the defense mechanisms of the organisms from which they are isolated. Several studies of tambjamine-producing marine organisms involved in prey-predator relationships have shown that these alkaloids were detected in extracts from both prey and predators. In this sense, extracts of <span style=\"font-weight: 400;\">V. dentata<span style=\"font-weight: 400;\">, <span style=\"font-weight: 400;\">T. stegosauriformis<span style=\"font-weight: 400;\">, <span style=\"font-weight: 400;\">T<span style=\"font-weight: 400;\">. <span style=\"font-weight: 400;\">brasiliensis<span style=\"font-weight: 400;\"> and <span style=\"font-weight: 400;\">R. ernsti <span style=\"font-weight: 400;\">were obtained. The analysis of organic and aqueous extracts from marine organisms by HPLC-UV-ELSD-MS and UPLC-QTOF-MS/MS allowed us to confirm the presence of 6 known tambjamines: tambjamines A, C, D and K, and the aldehydes of the tambjamines A and B. The quantification of tambjamine content in samples from different invertebrates performed by HPLC with a fluorescence detector demonstrated that <span style=\"font-weight: 400;\">T. stegosauriformis<span style=\"font-weight: 400;\"> and <span style=\"font-weight: 400;\">T. brasiliensis<span style=\"font-weight: 400;\"> have tambjamines content 8 and 14 times higher than their prey <span style=\"font-weight: 400;\">V. dentata<span style=\"font-weight: 400;\">, while <span style=\"font-weight: 400;\">R. ernsti<span style=\"font-weight: 400;\"> has tambjamines concentration between 3 and 5 times higher than their prey <span style=\"font-weight: 400;\">T. brasiliensis<span style=\"font-weight: 400;\"> and <span style=\"font-weight: 400;\">T. stegosauriformis<span style=\"font-weight: 400;\">. Additionally, non-targeted metabolomic analyzes of the extract from the mantle of <span style=\"font-weight: 400;\">R. ernsti<span style=\"font-weight: 400;\"> allowed to verify the presence of 6 new tambjamines. The synthesis, purification and characterization of the new tambjamines allowed to confirm the structure of 1 of the unprecedented tambjamines. (AU)

• FAPESP's process: 15/14114-3 - Which metabolomics components influence the phytopathogenicity of Colletotrichum spp.?
• Grantee: Mirelle Takaki
• Support Opportunities: Scholarships in Brazil - Doctorate