Análisis genómico-transcripcional de la respuesta del patógeno de peces Piscirickettsia salmonis frente a antibióticos utilizados en la industria acuícola
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Fecha
2016
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Profesor/a Guía
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Idioma
es
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Universidad Andrés Bello
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Licencia CC
Licencia CC
Resumen
La bacteria intracelular Piscirickettsia salmonis (P. salmonis) es el agente causal de
la Septicemia Rickettsial del Salmón (SRS), la cual afecta cultivos principalmente en
etapas de mar. Cada año aumenta la cantidad de antibióticos utilizados por la industria
acuícola como método de tratamiento, generando condiciones en el ambiente que permitan
la selección de cepas resistentes a estos. Estudios sugieren que incluso la exposición a
bajas concentraciones de antibióticos podría seleccionar bacterias resistentes. En el caso de
P. salmonis, a la fecha es limitada la información de los mecanismos moleculares que
determinan la resistencia a diversos antibióticos. El objetivo general de este trabajo es
identificar genes de P. salmonis potencialmente involucrados en procesos de resistencia a
antibióticos y analizar sus niveles de expresión. Los objetivos específicos son: i) Identificar
y analizar proteínas predichas potencialmente involucrados en la expulsión y
metabolización de antibióticos en los genomas de diferentes cepas de P. salmonis, ii)
Identificar la Concentración Mínima Inhibitoria (CIM) para diferentes antibióticos en las
cepas de P. salmonis bajo estudio, iii) Evaluar la expresión de genes potencialmente
involucrados en procesos de resistencia antibiótica frente a concentraciones subinhibitorias
del antibiótico florfenicol y iv) Integrar información con datos disponibles en
bases de datos públicas y generar un modelo preliminar de resistencia antibiótica en P
salmonis. Los resultados de determinación de la concentración mínima inhibitoria
utilizando antibióticos regularmente usados en la industria indican que existen distintos
grados de inhibición frente a un gradiente de concentración de antibióticos, diferencias que
se hacen más marcadas entre cepas ambientales y de referencia. Con el objetivo de explicar
estas diferencias se clasificaron y compararon, mediante herramientas bioinformáticas,
genes potencialmente involucrados en resistencia entre los genomas secuenciados de las
cepas de P. salmonis bajo estudio. Nuestros resultados indican que no existen grandes
diferencias entre los genes identificados, ya que todas las cepas poseen una amplia
maquinaria genética que le permitiría sobrevivir frente a diferentes clases de antibióticos.
Como las cepas cuentan con un paquete genético similar, se evaluó diferencias en la
expresión de genes seleccionados como potencialmente determinantes en resistencia frente
a concentraciones sub-inhibitorias. Se observaron diferencias significativas en la expresión
de algunos de estos genes, lo que se podría asociar a posibles determinantes en las cepas
ambientales, expuestas constantemente a las condiciones operacionales descritas
(concentraciones sub-inhibitorias del antibiótico florfenicol). Finalmente se concluye que
las diferencias observadas en la respuesta de cepas ambientales y de referencia podrían dar
pie a estudios más representativos para la generación de nuevos enfoques en el estudio de
mecanismos de resistencia a antibióticos en ambientes marinos de producción intensiva.
The intracellular bacterium Piscirickettsia salmonis (P. salmonis) is the causal agent of the Salmonid Rickettsial Septicaemia (SRS), which mainly affects cultures in sea stages. As a method of treatment, the amount of antibiotics increases every year, generating environmental conditions which allow the proliferation of resistant strains. Studies suggest that even low concentrations of antibiotics would allow this proliferation. In the case of P. salmonis, the information about the molecular mechanisms behind the antibiotic resistance is restricted. The main goal of this work is to identify P. salmonis genes potentially involved in the antibiotic-resistance processes and analyze their expression levels. The specific objectives are: i). Identify and analyze predicted proteins potentially involved in the expulsion and metabolization of antibiotics in the genomes of different strains of P. salmonis. ii). Determine the Minimum Inhibitory Concentration (MIC) for different antibiotics in the strains of P. salmonis studied. iii) Evaluate the expression of genes potentially involved in antibiotic resistance processes in sub-inhibitory antibiotics of antibiotics and iv) Integrate information with the available data in public databases and generate a preliminar model of antibiotic resistance in P. salmonis. The results of the determination of the MIC using antibiotics regularly used in the industry, indicate that exist different grades of inhibition in a gradient of antibiotic concentration, differences which accentuate between environmental and reference strains. With the objective of explaining these differences, genes potentially involved in antibiotic resistance were classified and compared between the sequenced genomes of the P. salmonis strains under study through the use of bioinformatic tools. Our results show that no significant differences exist between the identified genes, given that all the strains possess a wide genetic machinery that would allow them to survive different classes of antibiotics. Since the strains have a similar genetic repertoire, differences between the expressions of the selected genes potentially involved in antibiotic resistance in sub-inhibitory concentration of antibiotics were evaluated. Significant differences were observed in the expression of some of these genes, which could be associated to possible determinants in the environmental strains, constantly exposed to the described operational conditions. Finally, we conclude that the differences observed in the response of environmental and reference strains could give cause for more representative studies for the generation of new approaches in the study of the mechanisms of antibiotic resistance in sea environments of mass production.
The intracellular bacterium Piscirickettsia salmonis (P. salmonis) is the causal agent of the Salmonid Rickettsial Septicaemia (SRS), which mainly affects cultures in sea stages. As a method of treatment, the amount of antibiotics increases every year, generating environmental conditions which allow the proliferation of resistant strains. Studies suggest that even low concentrations of antibiotics would allow this proliferation. In the case of P. salmonis, the information about the molecular mechanisms behind the antibiotic resistance is restricted. The main goal of this work is to identify P. salmonis genes potentially involved in the antibiotic-resistance processes and analyze their expression levels. The specific objectives are: i). Identify and analyze predicted proteins potentially involved in the expulsion and metabolization of antibiotics in the genomes of different strains of P. salmonis. ii). Determine the Minimum Inhibitory Concentration (MIC) for different antibiotics in the strains of P. salmonis studied. iii) Evaluate the expression of genes potentially involved in antibiotic resistance processes in sub-inhibitory antibiotics of antibiotics and iv) Integrate information with the available data in public databases and generate a preliminar model of antibiotic resistance in P. salmonis. The results of the determination of the MIC using antibiotics regularly used in the industry, indicate that exist different grades of inhibition in a gradient of antibiotic concentration, differences which accentuate between environmental and reference strains. With the objective of explaining these differences, genes potentially involved in antibiotic resistance were classified and compared between the sequenced genomes of the P. salmonis strains under study through the use of bioinformatic tools. Our results show that no significant differences exist between the identified genes, given that all the strains possess a wide genetic machinery that would allow them to survive different classes of antibiotics. Since the strains have a similar genetic repertoire, differences between the expressions of the selected genes potentially involved in antibiotic resistance in sub-inhibitory concentration of antibiotics were evaluated. Significant differences were observed in the expression of some of these genes, which could be associated to possible determinants in the environmental strains, constantly exposed to the described operational conditions. Finally, we conclude that the differences observed in the response of environmental and reference strains could give cause for more representative studies for the generation of new approaches in the study of the mechanisms of antibiotic resistance in sea environments of mass production.
Notas
Tesis (Magíster en Biotecnología)
Palabras clave
Rickettsiosis, Salmón, Enfermedades, Antibióticos en Veterinaria