Intein prp8 em fungos dermatófitos: identificação molecular e aspectos evolutivos.
Carregando...
Data
2018-02-08
Autores
Garces, Hans Garcia
Título da Revista
ISSN da Revista
Título de Volume
Editor
Universidade Estadual Paulista (Unesp)
Resumo
Os dermatófitos são um grupo de fungos constituídos pelos gêneros Trichophyton, Epidermophyton e Microsporum que têm a habilidade de degradar a queratina. É por essa razão que podem colonizar a pele do homem e dos animais, embora também possam crescer no ambiente, geralmente em solos com restos de queratina. As características morfológicas permitem a identificação e classificação taxonômica das espécies, mas realizar um diagnóstico baseado nestas características pode levar a erros. As técnicas moleculares ajudam a realizar diagnósticos mais rápidos baseados em uma identificação molecular e também têm possibilitado importantes avanços quanto aos estudos filogenéticos, sendo as sequências ITS1-5.8S-ITS2 as mais utilizadas. No entanto, estes marcadores não são suficientes para identificar e elucidar todas as relações filogenéticas e taxonômicas dos dermatófitos devido à ampla variedade de espécies existentes, -sendo necessário novos marcadores genéticos, como o intein PRP8. Os inteins são elementos genéticos parasitas, de natureza proteica, presentes em alguns fungos e estão associados a importantes genes altamente conservados. Ointein PRP8 está localizado nogenePRP8 que codifica para a proteína PRP8 associada ao complexo do spliciosoma. O presente estudo visa caracterizar o intein PRP8 em fungos dermatófitos, de forma a empregar estes elementos genéticos como marcadores moleculares para uma correta identificação destas espécies e elucidações das relações filogenéticas do grupo. Para realizar este objetivo, foram caracterizadasmolecularmente 45 cepas usando as regiões nucleares ribossomais ITS1-5.8S-ITS2 e D1/D2 e posteriormente a região do intein PRP8 de 40 cepas foi sequenciada para determinar as características do intein. A presença do intein foi comprovada em todas as espécies avaliadas, apresentando um full-intein com uma Homing Endonuclease presumivelmente ativa. As construções filogenéticas obtidas usando as sequências do intein são correspondentes as construídas a partir de genes nucleares ribossomais, demonstrando a validade do intein para ser incluído em futuros estudos filogenético associados a taxonomia e determinação de espécies. O polimorfismo na região da Homing Endonuclease permitiu a identificação de algumas espécies de Microsporum mediante um ensaio de PCR-Eletroforese.
Dermatophytes are a fungal group composed by the genera Trichophyton, Epidermophyton and Microsporum with the ability to degrade keratin. That is why they can colonize the skin of man and animals, although they can also grow in the environment, usually in soils with remains of keratin. The morphological characteristics allow the identification and taxonomic classification of the species, but a diagnosis based on these characteristics is difficult. Molecular techniques made diagnoses based on molecular identification faster and made possible important advances in phylogenetic studies, with ITS1-5.8s-ITS2 sequences being the most used. However, these markers are not enough to elucidate all the phylogenetic relationships and taxonomic of dermatophytes due to the wide variety of existing species, and new genetic markers may be needed for correct identification and phylogenetic studies, such as the PRP8 intein. Inteins are parasitic genetic elements of a protein nature present in some fungi and are associated with important highly conserved genes. The PRP8 intein is located within the PRP8 gene coding for the PRP8 protein associated with the spliciosome complex. The present study aims to characterize the PRP8 intein in dermatophyte fungi, in order to use these genetic elements as molecular markers for a correct identification of these species and elucidations of the phylogenetic relationships of the group. To accomplish this goal, 45 strains were molecularly characterized using the ITS1-5.8S-ITS2 and D1 / D2 ribosomal nuclear regions and subsequently the PRP8 intein region of 40 strains was sequenced to determine the intein characteristics. The presence of intein was confirmed in all evaluated species, presenting a full-intein with a presumably active Homing Endonuclease. The phylogenetic construction obtained using the intein sequences correspond to those constructed from ribosomal nuclear genes, demonstrating the intein validity to be included in future phylogenetic studies associated with taxonomy and species determination. Polymorphism in the Homing Endonuclease region allowed the identification of some Microsporum species by a PCR-Electrophoresis assay.
Dermatophytes are a fungal group composed by the genera Trichophyton, Epidermophyton and Microsporum with the ability to degrade keratin. That is why they can colonize the skin of man and animals, although they can also grow in the environment, usually in soils with remains of keratin. The morphological characteristics allow the identification and taxonomic classification of the species, but a diagnosis based on these characteristics is difficult. Molecular techniques made diagnoses based on molecular identification faster and made possible important advances in phylogenetic studies, with ITS1-5.8s-ITS2 sequences being the most used. However, these markers are not enough to elucidate all the phylogenetic relationships and taxonomic of dermatophytes due to the wide variety of existing species, and new genetic markers may be needed for correct identification and phylogenetic studies, such as the PRP8 intein. Inteins are parasitic genetic elements of a protein nature present in some fungi and are associated with important highly conserved genes. The PRP8 intein is located within the PRP8 gene coding for the PRP8 protein associated with the spliciosome complex. The present study aims to characterize the PRP8 intein in dermatophyte fungi, in order to use these genetic elements as molecular markers for a correct identification of these species and elucidations of the phylogenetic relationships of the group. To accomplish this goal, 45 strains were molecularly characterized using the ITS1-5.8S-ITS2 and D1 / D2 ribosomal nuclear regions and subsequently the PRP8 intein region of 40 strains was sequenced to determine the intein characteristics. The presence of intein was confirmed in all evaluated species, presenting a full-intein with a presumably active Homing Endonuclease. The phylogenetic construction obtained using the intein sequences correspond to those constructed from ribosomal nuclear genes, demonstrating the intein validity to be included in future phylogenetic studies associated with taxonomy and species determination. Polymorphism in the Homing Endonuclease region allowed the identification of some Microsporum species by a PCR-Electrophoresis assay.
Descrição
Palavras-chave
Dermatófitos, Intein PRP8, Identificação molecular, filogenia, Dermatophytes, PRP8 intein, molecular identification, phylogeny