RESUMO
The sequence of the Leishmania tarentolae SSU rRNA (small subunit or 18S rRNA) gene was completely determined from 2 different strains and used to determine phylogenetic relationships between this organism and other trypanosomatids. Extensive structural similarities were observed between L. tarentolae and mammalian leishmanias the SSU rRNA. Phylogenetic reconstructions, using distance matrix or parsimony methods, showed large evolutionary distances between trypanosomes, either African and American, and L. tarentolae. Further analysis using intergenic rDNA spacer (IGS) sequences as probes in dot blot experiments confirmed the results obtained with the SSU rDNA comparisons. The data presented here clearly indicate that L. tarentolae is closely related to the mammalian parasite Leishmania donovani and highly divergent from trypanosomes.
Assuntos
Leishmania/classificação , Leishmania/genética , RNA de Protozoário/genética , RNA Ribossômico 18S/genética , Animais , Sequência de Bases , DNA Ribossômico/genética , Leishmania donovani/classificação , Leishmania donovani/genética , Dados de Sequência Molecular , Filogenia , Homologia de Sequência do Ácido Nucleico , Especificidade da EspécieRESUMO
The genus Acanthamoeba comprises free-living amebae identified as opportunistic pathogens of humans and other animal species. Morphological, biochemical and molecular approaches have shown wide genetic diversity within the genus. In an attempt to determine the genetic relatedness among isolates of Acanthamoeba we analyzed randomly amplified polymorphic DNA (RAPD) profiles of 11 Brazilian isolates from cases of human keratitis and 8 American type culture collection (ATCC) reference strains. We found that ATCC strains belonging to the same species present polymorphic RAPD profiles whereas strains of different species show very similar profiles. Although most Brazilian isolates could not be assigned with certainty to any of the reference species, they could be clustered according to pattern similarities. The results show that RAPD analysis is a useful tool for the rapid characterization of new isolates and the assessment of genetic relatedness of Acanthamoeba spp. A comparison between RAPD analyses and morphological characteristics of cyst stages is also discussed.
Assuntos
Ceratite por Acanthamoeba/parasitologia , Acanthamoeba/genética , DNA de Protozoário/análise , Técnica de Amplificação ao Acaso de DNA Polimórfico , Acanthamoeba/classificação , Acanthamoeba/crescimento & desenvolvimento , Animais , Primers do DNA/análise , Variação Genética , Humanos , Estágios do Ciclo de Vida/genéticaRESUMO
Fifty-four species or isolates of insect trypanosomatids were examined for the presence of selected restriction enzyme sites in the small (SSU) and large (LSU) rRNA coding units of ribosomal genes. In the SSU, sites for Eco RI, Bgl II, Pst I, and Hind III were found to occur at the same location for all species examined, thus displaying a universal distribution among trypanosomatids. In the LSU, a site for Bgl II in the 24S-alpha sequence and sites for Hind III and Pst I in the 24S-beta sequence were found in all species examined. In contrast, a site for Pvu II in the SSU exhibited a genus-related distribution, being present in Crithidia and Herpetomonas but absent in Phytomonas. A site for Hind III in the 24S-alpha sequence of the LSU also exhibited genus-restricted distribution. The site was present in Crithidia but absent in Phytomonas and Herpetomonas. These findings were confirmed by dot hybridization with a synthetic oligonucleotide complementary to the 18S rRNA sequence containing the Pvu II site. Results point to the usefulness of restriction markers as diagnostic tools for distinguishing the lower trypanosomatid genera Crithidia, Herpetomonas, and Phytomonas at the same time revealing a marked complexity within the genus Leptomonas.
Assuntos
DNA Ribossômico/genética , RNA Ribossômico 18S/genética , Trypanosomatina/genética , Amidoidrolases/genética , Animais , Anticorpos Monoclonais , Arginase/genética , Sequência de Bases , Southern Blotting , Enzimas de Restrição do DNA , DNA de Protozoário/genética , Dados de Sequência Molecular , Sondas de Oligonucleotídeos , RNA Ribossômico/genética , Trypanosomatina/classificaçãoRESUMO
Hybridization using kDNA and rDNA sequences as probes was performed to study phylogenetic relatedness of different species of trypanosomatids. Using this approach, we identified five organisms which had been classified as Phytomonas and Herpetomonas that were more closely correlated to each other phylogenetically than to any other species or isolates from either genera. These findings raise doubts about the validity of the current classification of Trypanosomatidae. Finally, we demonstrated the usefulness of kDNA sequences as an alternative to genomic sequences in obtaining phylogenetic information on trypanosomatids.
Assuntos
DNA de Cinetoplasto/genética , DNA Ribossômico/genética , Trypanosomatina/classificação , Trypanosomatina/genética , Animais , Sequência de Bases , Clonagem Molecular , Dados de Sequência Molecular , Hibridização de Ácido Nucleico , Filogenia , Mapeamento por Restrição , Especificidade da EspécieRESUMO
The genus Acanthamoeba comprises free-living amebae identified as opportunistic pathogens of humans and other animal species. Morphological, biochemical and molecular approaches have shown wide genetic diversity within the genus. In an attempt to determine the genetic relatedness among isolates of Acanthamoeba we analyzed randomly amplified polymorphic DNA (RAPD) profiles of 11 Brazilian isolates from cases of human keratitis and 8 American type culture collection (ATCC) reference strains. We found that ATCC strains belonging to the same species present polymorphic RAPD profiles whereas strains of different species show very similar profiles. Although most Brazilian isolates could not be assigned with certainty to any of the reference species, they could be clustered according to pattern similarities. The results show that RAPD analysis is a useful tool for the rapid characterization of new isolates and the assessment of genetic relatedness of Acanthamoeba spp. A comparison between RAPD analyses and morphological characteristics of cyst stages is also discussed (au)