Ocorrência e caracterização molecular de Bartonella spp. e Mycoplasma spp. em quirópteros amostrados no Brasil
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Data
2017-02-15
Autores
Ikeda, Priscila [UNESP]
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Editor
Universidade Estadual Paulista (Unesp)
Resumo
Doenças transmitidas por vetores artrópodes tem se tornado cada vez mais importantes para a saúde humana e animal. Neste sentido, o papel dos animais selvagens como reservatórios na transmissão destas enfermidades vem sendo investigado. A ordem Chiroptera é considerada o segundo maior grupo de mamíferos no mundo, hospedando importantes patógenos zoonóticos, tais como vírus e bactérias. Bartonella spp. e Mycoplasma spp. são bactérias que parasitam eritrócitos de diferentes espécies de mamíferos, incluindo humanos, podendo causar manifestações clínicas diversas. O presente estudo objetivou pesquisar a ocorrência e analisar filogeneticamente Bartonella spp. e Mycoplasma spp. em quirópteros amostrados no Brasil. Para tal, foram colhidas 325 amostras de sangue e/ou tecidos (fígado, coração e baço) de 162 quirópteros pertencentes a 19 espécies distribuídas em quatro famílias distintas de cinco estados: Mato Grosso, Pará, Paraná, São Paulo e Tocantins. Destas, três amostras mostraram-se negativas para o gene endógeno de referência (GAPDH), sendo excluídas das análises. Portanto, do total de 322 amostras, 17 (5,28%) mostraram-se positivas para Bartonella spp. por meio da PCR em tempo real baseada no gene nuoG. A quantificação de um fragmento do gene nuoG de Bartonella spp. por microlitro variou de 4,4 x 10⁰ a 6,95 x10³ cópias/μL nas amostras de sangue e teciduais dos quirópteros. Ainda, 45 amostras (13,97%) mostraram-se positivas para hemoplasmas por meio da PCR convencional baseada no gene 16S rRNA. Destas amostras foram obtidas sete sequências para Bartonella (nuoG [n=3], gltA [n=2], rpoB [n=1], ftsZ [n=1]), e cinco sequências de um fragmento do gene 16S rRNA de Mycoplasma spp. Nas análises filogenéticas, as sequências de Bartonella spp. posicionaram-se proximamente a genótipos de Bartonella spp. detectados em quirópteros amostrados em países da América Latina. Todas as cinco sequências de hemoplasmas formaram um grupo monofilético, tanto pela análise de Máxima Verossimilhança quanto pela Inferência Bayesiana, mostrando-se filogeneticamente relacionadas a Mycoplasma coccoides. As amostras positivas para Bartonella spp. foram provenientes das espécies Phyllostomus hastatus, Carollia perspicillata, Sturnira lilium, Glossophaga soricina e Natalus espiritosantensis. Trata-se da primeira descrição de Bartonella spp. na última espécie de quiróptero elencada. Os quirópteros positivos para hemoplasmas pertenciam às espécies Artibeus planirostris, Eptesicus sp., Eumops auripendulus, Glossophaga soricina, Molossus molossus, Molossus rufus, Myotis nigricans e Sturnira lilium, sendo a primeira evidência da circulação deste patógeno nestas espécies. Os genótipos de Bartonella spp. obtidos mostraram-se filogeneticamente próximos a genótipos detectados em quirópteros amostrados em outras regiões do mundo. Ainda, evidenciou-se a presença de genótipos filogeneticamente díspares entre si em diferentes regiões do Brasil. Já os genótipos de Mycoplasma spp. obtidos mostraram-se próximos entre si como um grupo monofilético, mas distantes dos previamente detectados em quirópteros dos Estados Unidos da América e Espanha. Portanto, o presente trabalho evidenciou, pela primeira vez, a circulação de Bartonella spp. e hemoplasmas entre quirópteros amostrados no Brasil e identificou novas espécies de quirópteros como hospedeiros das bactérias estudadas.
Vector-borne diseases have become increasingly important to human and animal health. The role of wild animals as reservoirs in the transmission of these diseases has been investigated. The Chiroptera Order is considered the second largest group of mammals in the world, hosting important zoonotic virus and bacteria. Bartonella spp. and Mycoplasma spp. are bacteria that parasites different mammals species' erythrocytes, including humans, causing different clinic manifestations. The present work aimed at investigating the occurrence and assessing the phylogenetic positioning of Bartonella spp. and Mycoplasma spp. in bats sampled in Brazil. A total of 325 blood and/or tissue (liver, spleen and heart) samples were collected from 162 bats belonging to 19 species distributed in four different families from five states: Mato Grosso, Pará, Paraná, São Paulo and Tocantins. Three samples showed negative results in the conventional PCR based on GAPDH gene and excluded from analysis. Seventeen (5,28%) out of 322 bats' samples were positive to qPCR for Bartonella spp. based on nuoG gene. The quantification of a Bartonella spp. nuoG gene fragment per microlitro in bats´ blood and tissues ranged from 4,4 x 10⁰ a 6,95 x10³ copies/μL. On the other hand, 45 (13,97%) samples were positive to cPCR assays for hemoplasmas based on 16S rRNA gene. Seven sequences were obtained for Bartonella spp. (nuoG [n=3], gltA [n=2], rpoB [n=1], ftsZ [n=1]), and five 16S rRNA sequences were obtained for Mycoplasma spp. In the phylogenetic analysis, the found Bartonella spp. sequences clustered with Bartonella spp. genotypes previously detected in bats sampled in Latin America countries. All five hemoplasmas sequences clustered together as a monophyletic group and closely related to Mycoplasma coccoides, by Maximum Likelihood and Bayesian Inference analyses. The biological samples positive for Bartonella spp. were collected from Phyllostomus hastatus, Carollia perspicillata, Sturnira lilium, Glossophaga soricina and Natalus espiritosantensis bats species. This is the first report of occurrence of Bartonella spp. in Natalus espiritosantensis. The positive biological samples for hemoplasmas were collected from animals belonging to the species Artibeus planirostris, Eptesicus sp., Eumops auripendulus, Glossophaga soricina, Molossus molossus, Molossus rufus, Myotis nigricans and Sturnira lilium, which represented the first evidence of the circulation of this pathogen in these species. The Bartonella spp. genotypes showed a phylogenetic relation with genotypes detected in bats sampled in other regions in the world; additionally, genotypes detected in bats from different regions of Brazil were phylogenetic distant from each other. However, Mycoplasma spp. genotypes obtained from bats’ biological samples from Brazil showed a closely relation among each other, comprising a monophyletic group, but distant from genotypes obtained in bats from the United States of America and Spain. Therefore, this is the first record of circulation of Bartonella spp. and Mycoplasma spp. among bats from Brazil, describing new species of bats as hosts of the studied bacteria.
Vector-borne diseases have become increasingly important to human and animal health. The role of wild animals as reservoirs in the transmission of these diseases has been investigated. The Chiroptera Order is considered the second largest group of mammals in the world, hosting important zoonotic virus and bacteria. Bartonella spp. and Mycoplasma spp. are bacteria that parasites different mammals species' erythrocytes, including humans, causing different clinic manifestations. The present work aimed at investigating the occurrence and assessing the phylogenetic positioning of Bartonella spp. and Mycoplasma spp. in bats sampled in Brazil. A total of 325 blood and/or tissue (liver, spleen and heart) samples were collected from 162 bats belonging to 19 species distributed in four different families from five states: Mato Grosso, Pará, Paraná, São Paulo and Tocantins. Three samples showed negative results in the conventional PCR based on GAPDH gene and excluded from analysis. Seventeen (5,28%) out of 322 bats' samples were positive to qPCR for Bartonella spp. based on nuoG gene. The quantification of a Bartonella spp. nuoG gene fragment per microlitro in bats´ blood and tissues ranged from 4,4 x 10⁰ a 6,95 x10³ copies/μL. On the other hand, 45 (13,97%) samples were positive to cPCR assays for hemoplasmas based on 16S rRNA gene. Seven sequences were obtained for Bartonella spp. (nuoG [n=3], gltA [n=2], rpoB [n=1], ftsZ [n=1]), and five 16S rRNA sequences were obtained for Mycoplasma spp. In the phylogenetic analysis, the found Bartonella spp. sequences clustered with Bartonella spp. genotypes previously detected in bats sampled in Latin America countries. All five hemoplasmas sequences clustered together as a monophyletic group and closely related to Mycoplasma coccoides, by Maximum Likelihood and Bayesian Inference analyses. The biological samples positive for Bartonella spp. were collected from Phyllostomus hastatus, Carollia perspicillata, Sturnira lilium, Glossophaga soricina and Natalus espiritosantensis bats species. This is the first report of occurrence of Bartonella spp. in Natalus espiritosantensis. The positive biological samples for hemoplasmas were collected from animals belonging to the species Artibeus planirostris, Eptesicus sp., Eumops auripendulus, Glossophaga soricina, Molossus molossus, Molossus rufus, Myotis nigricans and Sturnira lilium, which represented the first evidence of the circulation of this pathogen in these species. The Bartonella spp. genotypes showed a phylogenetic relation with genotypes detected in bats sampled in other regions in the world; additionally, genotypes detected in bats from different regions of Brazil were phylogenetic distant from each other. However, Mycoplasma spp. genotypes obtained from bats’ biological samples from Brazil showed a closely relation among each other, comprising a monophyletic group, but distant from genotypes obtained in bats from the United States of America and Spain. Therefore, this is the first record of circulation of Bartonella spp. and Mycoplasma spp. among bats from Brazil, describing new species of bats as hosts of the studied bacteria.
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Palavras-chave
Análise filogenética, Bartonella, Mycoplasma, Quirópteros, Phylogenetic analysis, Bartonella, Hemotropic micoplasmas, Chiroptera