Estudo metabolômico de fungos do gênero Colletotrichum e de organismos marinhos produtores de tambjaminas

Página do item simplificado
dc.contributorUniversidade de São Paulo
dc.contributor.author1
dc.date.issued2020-09-01
dc.description.abstract<span style=\"font-weight: 400;\">O presente trabalho descreve o estudo metabol&ocirc;mico de fungos do g&ecirc;nero <span style=\"font-weight: 400;\">Colletotrichum<span style=\"font-weight: 400;\"> e de organismos marinhos produtores de tambjaminas <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;\"> e <span style=\"font-weight: 400;\">Roboastra ernsti<span style=\"font-weight: 400;\">. Fungos do g&ecirc;nero <span style=\"font-weight: 400;\">Colletotrichum<span style=\"font-weight: 400;\"> causam doen&ccedil;as em uma grande variedade de plantas de diversas fam&iacute;lias. Metab&oacute;litos secund&aacute;rios produzidos por estes micro-organismos podem constituir parte do processo de virul&ecirc;ncia associada &agrave; sua fitopatogenicidade. Apesar de muitos metab&oacute;litos de linhagens de <span style=\"font-weight: 400;\">Colletotrichum<span style=\"font-weight: 400;\"> j&aacute; terem sido isolados e identificados, a fitopatogenicidade associada a fungos deste g&ecirc;nero ainda n&atilde;o est&aacute; bem estabelecida. Com o intuito de se investigar a diferen&ccedil;a no metabolismo de linhagens endof&iacute;ticas e fitopatog&ecirc;nica de <span style=\"font-weight: 400;\">Colletotrichum<span style=\"font-weight: 400;\"> spp., 25 isolados de <span style=\"font-weight: 400;\">Colletotrichum<span style=\"font-weight: 400;\"> foram investigados, dentro os quais 16 eram endof&iacute;ticos e 9 fitopatog&ecirc;nicos. Os extratos dos meios de cultivo foram fracionados e analisados por UPLC-IMS-QToF-HRMS. Os dados brutos foram processados empregando-se ferramentas <span style=\"font-weight: 400;\">in-house<span style=\"font-weight: 400;\"> e <span style=\"font-weight: 400;\">molecular networking<span style=\"font-weight: 400;\">. A avalia&ccedil;&atilde;o dos resultados permitiu concluir que linhagens endof&iacute;ticas e fitopatog&ecirc;nicas de <span style=\"font-weight: 400;\">Colletotrichum<span style=\"font-weight: 400;\"> spp. possuem metabolismo consideravelmente diferente. Linhagens endof&iacute;ticas produzem compostos que s&atilde;o comuns &agrave; varias linhagens do grupo; j&aacute; linhagens fitopatog&ecirc;nicas produzem compostos mais espec&iacute;ficos a cada um dos isolados. O processo de desreplica&ccedil;&atilde;o dos resultados de<span style=\"font-weight: 400;\"> molecular networking <span style=\"font-weight: 400;\">utilizando-se uma biblioteca <span style=\"font-weight: 400;\">in-house<span style=\"font-weight: 400;\"> de compostos isolados de <span style=\"font-weight: 400;\">Colletotrichum<span style=\"font-weight: 400;\"> spp. permitiu a identifica&ccedil;&atilde;o de 5 compostos que apresentam atividade fitot&oacute;xica nas amostras relacionadas &agrave;s linhagens fitopatog&ecirc;nicas. Tais resultados s&atilde;o forte indicativo de que a fitopatogenicidade de linhagens de <span style=\"font-weight: 400;\">Colletotrichum<span style=\"font-weight: 400;\"> podem estar associada &agrave; produ&ccedil;&atilde;o de compostos fitot&oacute;xicos. Tambjaminas s&atilde;o um grupo de alcal&oacute;ides quimicamente relacionados &agrave; prodigiosina e prodigionina, isoladas de bact&eacute;rias e organismos marinhos como asc&iacute;dias, briozo&aacute;rios e nudibr&acirc;nquios. Al&eacute;m de apresentarem atividades biol&oacute;gicas diversificadas, as tambjaminas est&atilde;o possivelmente relacionadas &agrave; mecanismos de defesa dos organismos a partir dos quais s&atilde;o isoladas. Diversos estudos de organismos marinhos produtores de tambjaminas envolvidos em rela&ccedil;&otilde;es presa-predador demonstraram que estes alcal&oacute;ides foram detectados nos extratos tanto das presas quanto dos predadores. Nesse sentido, foram obtidos os extratos de <span style=\"font-weight: 400;\">V. dentata, T. stegosauriformis, T. brasiliensis e R. ernsti<span style=\"font-weight: 400;\">. A partir da an&aacute;lise dos extratos org&acirc;nicos e aquosos dos organismos marinhos por HPLC-UV-ELSD-MS e UPLC-QToF-MS/MS, foi poss&iacute;vel confirmar a presen&ccedil;a de 6 tambjaminas conhecidas: tambjaminas A, C, D e K e os alde&iacute;dos das tambjaminas A e B. A quantifica&ccedil;&atilde;o do conte&uacute;do de tambjaminas nas amostras de diferentes invertebrados por HPLC com detector de fluoresc&ecirc;ncia demonstrou que <span style=\"font-weight: 400;\">T. stegosauriformis<span style=\"font-weight: 400;\"> e <span style=\"font-weight: 400;\">T. brasiliensis<span style=\"font-weight: 400;\"> possuem conte&uacute;do de tambjaminas de 8 e 14 vezes superiores &agrave; sua presa <span style=\"font-weight: 400;\">V. dentata<span style=\"font-weight: 400;\">, enquanto que <span style=\"font-weight: 400;\">R. ernsti<span style=\"font-weight: 400;\"> possui concentra&ccedil;&atilde;o de tambjaminas entre 3 e 5 vezes superiores &agrave; suas presas <span style=\"font-weight: 400;\">T. brasiliensis<span style=\"font-weight: 400;\"> e <span style=\"font-weight: 400;\">T. stegosauriformis<span style=\"font-weight: 400;\">. Adicionalmente, an&aacute;lises metabol&ocirc;micas n&atilde;o-direcionadas do extrato do manto de <span style=\"font-weight: 400;\">R. ernsti <span style=\"font-weight: 400;\">permitiram verificar a presen&ccedil;a de 6 novas tambjaminas. A s&iacute;ntese, purifica&ccedil;&atilde;o e caracteriza&ccedil;&atilde;o das novas tambjaminas permitiu confirmar a estrutura de 1 das tambjaminas in&eacute;ditas.
dc.description.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.
dc.formatapplication/pdf
dc.identifier.doi10.11606/T.75.2020.tde-28082020-152400
dc.identifier.urihttps://www.teses.usp.br/teses/disponiveis/75/75133/tde-28082020-152400/
dc.languagept
dc.rights.holder1
dc.subjectmetabolômica
dc.subjectColletotrichum
dc.subjecttambjaminas
dc.subjectfungos endofíticos
dc.subjectfungos fitopatogênicos
dc.subjectorganismos marinhos
dc.subjectphytopathogenic fungi
dc.subjectmetabolomics
dc.subjectColletotrichum
dc.subjectmarine organisms
dc.subjectendophytic fungi
dc.subjecttambjamines
dc.titleEstudo metabolômico de fungos do gênero Colletotrichum e de organismos marinhos produtores de tambjaminas
dc.title.alternativeMetabolomic study of fungi of the genus Colletotrichum and tambjamines- producing marine organisms
dc.typeTese de Doutorado
usp.advisorBerlinck, Roberto Gomes de Souza
usp.date.defense2020-04-16
Arquivos
Coleções
Teses e Dissertações (BDTD USP)