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Doctoral Thesis
DOI
https://doi.org/10.11606/T.11.2016.tde-07012016-175356
Document
Author
Full name
Thays Benites Camargo Pereira
E-mail
Institute/School/College
Knowledge Area
Date of Defense
Published
Piracicaba, 2015
Supervisor
Committee
Bedendo, Ivan Paulo (President)
Flores, Daniela
Gasparoto, Maria Cândida de Godoy
Mello, Ana Paula de Oliveira Amaral
Rezende, Dalilla Carvalho
Title in Portuguese
Identificação de diversidade genética em fitoplasmas com base na análise molecular dos gene 16S rRNA, SecY e Proteína Ribossomal
Keywords in Portuguese
Caracterização Multilocus
Ferramentas moleculares
Mollicutes
Procariotos sem parede celular
Abstract in Portuguese
Os fitoplasmas são organismos procariotos sem parede celular, que habitam as células do floema das plantas e são transmitidos, principalmente, por cigarrinhas sugadoras de floema. Este patógeno é responsável por causar doenças em diversas espécies vegetais, provocando a expressão de diversos sintomas, entre os quais podem ser destacados a redução do tamanho foliar, amarelecimento das folhas, superbrotamento de ramos e enfezamento da planta hospedeira. Entre os anos de 2013 e 2014, plantas de três espécies vegetais com sintomas característicos de infecção causada por fitoplasmas foram observadas no estado de São Paulo. O DNA total foi extraído a partir de plantas de couve-flor com sintomas de nanismo, malformação da inflorescência e necrose dos vasos do floema; de plantas da ornamental conhecida por árvore-da-felicidade com sintomas amarelecimento e redução do limbo foliar; e de plantas de Alho-poró com sintomas de enfezamento. Por meio do teste de PCR conduzido em sua maioria com os primers P1A/16S-SR foi possível detectar fitoplasmas nas três espécies vegetais testadas. Os fragmentos genômicos correspondentes ao 16S rRNA dos fitoplasmas foram sequenciados e identificados. Nas plantas de couve-flor e da árvore-da-felicidade foram identificados fitoplasmas afilados ao grupo 16SrVII-B, através da análise virtual de RFLP, cálculo do coeficientes de similaridade (F) e análise filogenética. Para ambas as espécies, este é o primeiro relato da ocorrência de representantes deste grupo, nas condições brasileiras. Nas plantas de Alho-poró foram identificados fitoplasmas afiliados ao subgrupo 16SrIII-J e ao grupo 16SrXIII, com base na análise dos genes 16S rRNA, SecY e proteína ribossomal, conduzidas através de análise virtual de RFLP, valores de coeficientes de similaridade e análise filogenética. Para o fitoplasma membro do grupo 16SrXIII foram identificadas quatro estirpes, uma delas afiliada ao subgrupo 16SrXIII-E e as demais como prováveis representantes de novos subgrupos. O presente estudo se constitui em uma contribuição ao conhecimento de novos patossistemas envolvendo fitoplasmas, bem como à caracterização molecular de fitoplasmas baseadas em diferentes genes marcadores, atualmente usados para a classificação de representantes deste grupo emergente de agentes causais de doenças de plantas.
Title in English
Identification of genetic diversity in phytoplasma based on molecular analysis of the 16S rRNA, SecY and ribosomal protein genes
Keywords in English
Cell wall-less prokaryotes
Molecular tools
Multilocus characterization
Abstract in English
The phytoplasmas are prokaryotic organisms without cell walls, which inhabit the phloem cells of plants and are transmitted mainly by leafhoppers sucking the phloem. This pathogen is responsible for causing diseases in various plant species, leading to expression of many symptoms, among which can be highlighted the reduction of leaf size, leaf yellowing, shoots proliferation and stunting of the plant host. Between the years 2013 and 2014, three plant species with characteristic symptoms of infection caused by phytoplasma were observed in São Paulo. Total DNA was extracted from cauliflower plants with symptoms of stunting, malformation of the inflorescence and necrosis of the phloem; ornamental plants known for Ming Aralia with symptoms yellowing and little leaves; and leek plants with symptoms of stunting. Through the PCR test conducted mostly with the primers P1A/16S-SR was possible to detect phytoplasmas in the three species of plants. The genomic fragments corresponding of 16S rRNA gene of the phytoplasma were sequenced and identified. In plants of cauliflower and Ming Aralia were identified phytoplasmas belonging to 16SrVII-B group through virtual RFLP analysis, calculation of similarity coefficients (F) and phylogenetic analysis. For both species, this is the first report of the occurrence of representatives of this group, in Brazilian conditions. In leek plants were identified phytoplasmas affiliated with 16SrIII-J subgroup, and the 16SrXIII group, based on the analysis of the 16S rRNA, SecY and ribosomal protein genes, conducted through virtual analysis of RFLP, similarity coefficient values and phylogenetic analysis. For the phytoplasma member of group 16SrXIII were identified four strains, one affiliated with 16SrXIII-E subgroup and the others as probable representatives of new subgroups. This study constitutes a contribution to knowledge of new pathosystems involving phytoplasmas and the molecular characterization of phytoplasma based on different marker genes, currently used for the classification of representatives of this emerging group of plant pathogens.
 
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Publishing Date
2016-01-14
 
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