Flavobacterium columnare: caracterização molecular, patogenicidade e perfil de susceptibilidade a antimicrobianos
Carregando...
Data
2021-07-29
Autores
Título da Revista
ISSN da Revista
Título de Volume
Editor
Universidade Estadual Paulista (Unesp)
Resumo
Flavobacterium columnare é uma bactéria oportunista que causa grande impacto na aquicultura mundial. Neste estudo, investigamos as características moleculares, a patogenicidade da bactéria ao pacu, Piaractus mesopotamicus e susceptibilidade antimicrobiana de F. columnare isolada de tilápia-do-Nilo, Oreochromis niloticus. A patogenicidade ao pacu foi avaliada através da infecção experimental por imersão. As características moleculares da bactéria foram determinadas através da técnica PCR-RFLP e sequenciamento de nucleotídeos com análise filogenética do gene 16S rRNA. A susceptibilidade à enrofloxacina (ENRO), florfenicol (FFC), oxitetraciclina (OTC), tianfenicol (TAP) e suas combinações foram avaliadas pelos métodos da microdiluição em caldo e teste de sinergismo, respectivamente. Como resultado, após a infecção experimental, colônias rizóides foram isoladas do rim cranial do pacu, após a realização da técnica da Reação em Cadeia da Polimerase (PCR) para o gene 16S rDNA, confirmando tratar-se de F. columnare. Após análise por PCR-RFLP com a endonuclease Hae III, confirmou-se que a cepa pertence ao genomovar II. A análise filogenética do gene 16S rRNA demonstrou que a cepa de F. columnare está intimamente relacionada ao grupo genético tailandês (GG4), sendo esta a primeira ocorrência desta linhagem no Brasil. Para o teste de susceptibilidade aos antimicrobianos, a cepa foi classificada como tipo selvagem para todos os antimicrobianos testados. Em relação ao sinergismo entre os antimicrobianos, encontramos sinergismo entre tianfenicol e a enrofloxacina (ΣFIC = 0,5), atividade aditiva com a combinação de florfenicol com tianfenicol (ΣFIC = 0,75) e antagonismo na combinação de oxitetraciclina com enrofloxacina. Os resultados deste estudo são importantes para o controle da columnariose na piscicultura brasileira, pois demonstram que apesar da cepa ser altamente virulenta, os antimicrobianos mais utilizados na piscicultura brasileira são eficientes contra ela.
Flavobacterium columnare is an important pathogen of freshwater fish that has severe impacts on the commercial aquaculture sector. In this study, we investigated the molecular characteristics, pathogenicity against pacu, Piaractus mesopotamicus, and antimicrobial susceptibility of a virulent F. columnare isolated from Nile tilapia, Oreochromis niloticus, cultured in Brazil. The pathogenicity against pacu was analyzed by experimental columnaris infection by immersion challenge. Bacteria molecular characteristics were determined through PCR-RFLP, multiplex PCR, nucleotide sequencing and phylogenetic analysis of 16S rRNA gene. Susceptibility to enrofloxacin (ENRO), florfenicol (FFC), oxytetracycline (OTC), thiamphenicol (TAP), and their combinations were assessed by broth microdilution and checkerboard methods, respectively. As a result, after experimental infection, rhizoid-type colonies were isolated from the cranial kidney of pacu, which after performing PCR for 16S rDNA gene, were identified as F. columnare, and then submitted to PCR-RFLP using restriction enzyme Hae III demonstrated to belong to genomovar II, phylogenetic analyses reveals the strain is closely related to Thai isolates (GG4), this linage was also identified in an outbreak in Brazilian fish farming for the first time. For the AST test the strain was classified as wild-type for all tested antimicrobials. Regarding the synergism between antimicrobials, we found synergism between thiamphenicol and enrofloxacin (ΣFIC = 0.5), additive activity with the combination of florfenicol with thiamphenicol (ΣFIC = 0.75), and antagonism in the combination of oxytetracycline with enrofloxacin. These results are very important because making it possible to assess the susceptibility of F. columnare to the main antimicrobials used in Brazilian fish farming, as well as to obtain a synergistic effect between thiamphenicol and enrofloxacin.
Flavobacterium columnare is an important pathogen of freshwater fish that has severe impacts on the commercial aquaculture sector. In this study, we investigated the molecular characteristics, pathogenicity against pacu, Piaractus mesopotamicus, and antimicrobial susceptibility of a virulent F. columnare isolated from Nile tilapia, Oreochromis niloticus, cultured in Brazil. The pathogenicity against pacu was analyzed by experimental columnaris infection by immersion challenge. Bacteria molecular characteristics were determined through PCR-RFLP, multiplex PCR, nucleotide sequencing and phylogenetic analysis of 16S rRNA gene. Susceptibility to enrofloxacin (ENRO), florfenicol (FFC), oxytetracycline (OTC), thiamphenicol (TAP), and their combinations were assessed by broth microdilution and checkerboard methods, respectively. As a result, after experimental infection, rhizoid-type colonies were isolated from the cranial kidney of pacu, which after performing PCR for 16S rDNA gene, were identified as F. columnare, and then submitted to PCR-RFLP using restriction enzyme Hae III demonstrated to belong to genomovar II, phylogenetic analyses reveals the strain is closely related to Thai isolates (GG4), this linage was also identified in an outbreak in Brazilian fish farming for the first time. For the AST test the strain was classified as wild-type for all tested antimicrobials. Regarding the synergism between antimicrobials, we found synergism between thiamphenicol and enrofloxacin (ΣFIC = 0.5), additive activity with the combination of florfenicol with thiamphenicol (ΣFIC = 0.75), and antagonism in the combination of oxytetracycline with enrofloxacin. These results are very important because making it possible to assess the susceptibility of F. columnare to the main antimicrobials used in Brazilian fish farming, as well as to obtain a synergistic effect between thiamphenicol and enrofloxacin.
Descrição
Palavras-chave
Genética microbiana, Microbiologia, Reação em cadeia da polimerase, Filogenia, Peixes de água doce