La hemolisina termoestable directa, tdhA, es parte de un transposón compuesto funcional en Vibrio parahaemolyuticus RIMD 2210633
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Fecha
2010
Profesor/a Guía
Facultad/escuela
Idioma
es
Título de la revista
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Título del volumen
Editor
Universidad Andrés Bello
Nombre de Curso
Licencia CC
Licencia CC
Resumen
RESUMEN: Vibrio parahaemolyticus es una bacteria de origen marino y agente causal de un síndrome
diarreico por consumo de alimentos marinos contaminados. En Chile, este cuadro se produce cada
verano con un número de casos que superan los 1000 afectados. La capacidad de generar la
gastroenteritis se relaciona con la presencia de una hemolisina codificada por los genes tdhA y tdhS.
Existe una fuerte correlación entre cepas clínicas y los genes tdh, el que además se encuentra
presente en V. cholerae, V. mimicus y V. hollisae. La presencia en otras especies se debería a la
capacidad de este gen de movilizarse lateralmente lo que se deduce de la existencia de secuencias de
inserción (IS) flanqueando el gen tdhA. Durante el desarrollo de este trabajo se ha demostrado que
flanqueando al gen tdhA se encuentra una IS 551 pb río abajo, la que también está presente repetida
e invertida 2.364 pb río arriba. Al considerar la disposición de ambas IS en función del gen tdhA se
observa una estructura similar a la de un transposón compuesto. Se analizó por Southern blot la
presencia del transposón, encontrándose en 12 de 20 de las cepas analizadas, tanto clínicas como
ambientales. Para evaluar si el transposón era funcional, éste fue clonado en un vector y
posteriormente transformado en S. Typhi. Al sobreexpresar la transposasa se generaron mutantes
auxótrofas como consecuencia del movimiento del transposón en el cromosoma de la bacteria
receptora. Esto nos permite concluir que el gen de virulencia tdhA es parte de un transposón
compuesto funcional.
ABSTRACT: Vibrio parahaemolyticus is a marine bacterium that causes human gastroenteritis. Most clinical isolates produce a major virulence factor known as the thermostable direct hemolysin (TDH). However have been isolated V. parahaemolyticus strains lacking tdh genes from patients with gastroenteritis. Besides this, exist evidence that support that TDH is not exclusive of Vibrio parahaemolytcus and is present in different Vibrio species, probably by lateral transfer. Recently, it has been demonstrated that TDH encoded by the tdh gene, in a clinical strain, was partially deleted by the transposition of the insertion sequence, ISVpa3. In order to add to understand the changes to the characteristics of V. parahaemolyticus occur during infection, we examined the surrounding tdhA gene, of the sequence strain RIMD2210633, for the presence of more insertion sequences. Was founded two 849‐bp IS flanking the tdhA gene forming a structure like a composite transposon. The presence of the transposase was detected by Westtern blott. The functionality of the transposón was demonstrated by the generation of auxotophic mutants as consequences of the interference of metabolic genes by the transposón. Therefore the evidences suggest the presence of a functional transposase that can be responsible of the transpose action of the composite transposon and the generation of strain lacking tdhA gene.
ABSTRACT: Vibrio parahaemolyticus is a marine bacterium that causes human gastroenteritis. Most clinical isolates produce a major virulence factor known as the thermostable direct hemolysin (TDH). However have been isolated V. parahaemolyticus strains lacking tdh genes from patients with gastroenteritis. Besides this, exist evidence that support that TDH is not exclusive of Vibrio parahaemolytcus and is present in different Vibrio species, probably by lateral transfer. Recently, it has been demonstrated that TDH encoded by the tdh gene, in a clinical strain, was partially deleted by the transposition of the insertion sequence, ISVpa3. In order to add to understand the changes to the characteristics of V. parahaemolyticus occur during infection, we examined the surrounding tdhA gene, of the sequence strain RIMD2210633, for the presence of more insertion sequences. Was founded two 849‐bp IS flanking the tdhA gene forming a structure like a composite transposon. The presence of the transposase was detected by Westtern blott. The functionality of the transposón was demonstrated by the generation of auxotophic mutants as consequences of the interference of metabolic genes by the transposón. Therefore the evidences suggest the presence of a functional transposase that can be responsible of the transpose action of the composite transposon and the generation of strain lacking tdhA gene.
Notas
Tesis (Doctor en Biotecnología)
Palabras clave
Contaminación de Alimentos Marinos., Bacterias.