RESUMO
Giardia duodenalis (syn. Giardia intestinalis and Giardia lamblia) is a widespread intestinal parasite in mammals, including humans and pets worldwide. It should be considered a species complex and comprises eight assemblage (A-H). This works aimed to determine the genotypic variability among G. duodenalis isolates from dogs from Minas Gerais state, Brazil. Fecal samples of 97 dogs, from 1-to-10 months old from 15 commercial kennels, were collected and analyzed by the zinc sulfate centrifugal flotation technique, to determine their positivity for G. duodenalis cysts. Cysts pellets were stored and submitted to PCR and nested-PCR reactions with gdh and tpi primers, and then sequencing. Among positive samples (n = 19), fragment amplifications of gdh and tpi genes was observed in 16 (84,2%) and 14 (73,6%), respectively. In total, 30 sequences were obtained. Sequencing analysis showed that for gdh, all isolates were identified as host-specific genotype D, and for tpi, besides host-specific genotype C, were also observed zoonotic genotypes A and B. This study provides, for the first time, current information about genetic characterization of G. duodenalis isolates found in dogs in Minas Gerais state.
Assuntos
DNA de Protozoário/isolamento & purificação , Doenças do Cão/parasitologia , Giardia lamblia/genética , Giardíase/veterinária , Distribuição por Idade , Animais , Brasil/epidemiologia , DNA de Protozoário/química , Doenças do Cão/epidemiologia , Cães , Fezes/parasitologia , Feminino , Giardia lamblia/classificação , Giardíase/epidemiologia , Giardíase/parasitologia , Glutamato Desidrogenase/genética , Masculino , Prevalência , Alinhamento de Sequência/veterinária , Análise de Sequência de DNA/veterinária , Triose-Fosfato Isomerase/genéticaRESUMO
BACKGROUND: Giardia duodenalis is a parasite of several mammalian species, including humans, distributed worldwide. This research aimed to identify the molecular assemblages/sub-assemblages of G. duodenalis and to determine the intra-assemblage genetic variation of the different genes of assemblages A and B in pre-school children in the cities of Araguari and Uberlândia, Minas Gerais, Brazil. METHODS: The molecular characterization followed ß-giardin (bg), glutamate dehydrogenase (gdh) and triose phosphate isomerase (tpi) protocols. RESULTS: Of 226 stool samples, G. duodenalis cysts were found in 45 (19.9%). The tpi gene was amplified in 34 samples: 16 assemblage A, 14 B and four mixed samples A/B. The gdh gene was amplified in 32 samples, including 14 A, 16 B and two A/B. For the bg gene, 19 samples were sequenced: nine assemblage A, five B, three E, and two mixed, A/E and B/E. Animal-specific assemblage E were identified by bg, but were not confirmed for other genes. Twelve samples were characterized by full agreement of the three genes. Two new multilocus genotyping (MLGs) for assemblage A and two new MLGs for assemblage B were also described. CONCLUSIONS: These findings substantiate the importance of using more than one gene protocol since the sensitivity and genetic variability changes with the locus used.Access numbers: The GenBank access numbers for the nucleotide sequences reported in this article are: JQ794877-JQ794890, JX033113-JX033118.
Assuntos
Proteínas do Citoesqueleto/genética , Genes de Protozoários , Genótipo , Giardia lamblia/genética , Giardíase/parasitologia , Glutamato Desidrogenase/genética , Proteínas de Protozoários/genética , Triose-Fosfato Isomerase/genética , Sequência de Bases , Brasil , Criança , Pré-Escolar , Cidades , Fezes , Feminino , Amplificação de Genes , Variação Genética , Técnicas de Genotipagem , Humanos , Lactente , Recém-Nascido , Masculino , OocistosRESUMO
Giardia duodenalis is a small intestinal protozoan parasite of several terrestrial vertebrates. This work aims to assess the genotypic variability of Giardia duodenalis isolates from cattle, sheep and pigs in the Southeast of Brazil, by comparing the standard characterization between glutamate dehydrogenase (gdh) and triose phosphate isomerase (tpi) primers. Fecal samples from the three groups of animals were analyzed using the zinc sulphate centrifugal flotation technique. Out of 59 positive samples, 30 were from cattle, 26 from sheep and 3 from pigs. Cyst pellets were stored and submitted to PCR and nested-PCR reactions with gdh and tpi primers. Fragment amplification of gdh and tpi genes was observed in 25 (42.4%) and 36 (61.0%) samples, respectively. Regarding the sequencing, 24 sequences were obtained with gdh and 20 with tpi. For both genes, there was a prevalence of E specific species assemblage, although some isolates have been identified as A and B, by the tpi sequencing. This has also shown a larger number of heterogeneous sequences, which have been attribute to mixed infections between assemblages B and E. The largest variability of inter-assemblage associated to the frequency of heterogeneity provided by tpi sequencing reinforces the polymorphic nature of this gene and makes it an excellent target for studies on molecular epidemiology.