Echeverría Ancín, Myriam

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https://academica-e.unavarra.es/handle/2454/48926

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Echeverría Ancín

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Myriam

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Producción Agraria

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751875

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810079

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2014 - 20193
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    PublicationOpen Access
    The toxic guardians: multiple toxin-antitoxin systems provide stability, avoid deletions and maintain virulence genes of Pseudomonas syringae virulence plasmids
    (BMC, 2019) Bardají Goikoetxea, Leire; Añorga García, Maite; Echeverría Ancín, Myriam; Ramos, Cayo; Murillo Martínez, Jesús; Institute for Multidisciplinary Research in Applied Biology - IMAB
    Background: Pseudomonas syringae is a y-proteobacterium causing economically relevant diseases in practically all cultivated plants. Most isolates of this pathogen contain native plasmids collectively carrying many pathogenicity and virulence genes. However, P. syringae is generally an opportunistic pathogen primarily inhabiting environmental reservoirs, which could exert a low selective pressure for virulence plasmids. Additionally, these plasmids usually contain a large proportion of repeated sequences, which could compromise plasmid integrity. Therefore, the identification of plasmid stability determinants and mechanisms to preserve virulence genes is essential to understand the evolution of this pathogen and its adaptability to agroecosystems. Results: The three virulence plasmids of P. syringae pv. savastanoi NCPPB 3335 contain from one to seven functional stability determinants, including three highly active toxin-antitoxin systems (TA) in both pPsv48A and pPsv48C. The TA systems reduced loss frequency of pPsv48A by two orders of magnitude, whereas one of the two replicons of pPsv48C likely confers stable inheritance by itself. Notably, inactivation of the TA systems from pPsv48C exposed the plasmid to high-frequency deletions promoted by mobile genetic elements. Thus, recombination between two copies of MITEPsy2 caused the deletion of an 8.3 kb fragment, with a frequency of 3.8 ± 0.3 x 10-3. Likewise, one-ended transposition of IS801 generated plasmids containing deletions of variable size, with a frequency of 5.5 ± 2.1 x 1 0- 4, of which 80% had lost virulence gene idi. These deletion derivatives were stably maintained in the population by replication mediated by repJ, which is adjacent to IS801. IS801 also promoted deletions in plasmid pPsv48A, either by recombination or one-ended transposition. In all cases, functional TA systems contributed significantly to reduce the occurrence of plasmid deletions in vivo. Conclusions: Virulence plasmids from P. syringae harbour a diverse array of stability determinants with a variable contribution to plasmid persistence. Importantly, we showed that multiple plasmid-borne TA systems have a prominent role in preserving plasmid integrity and ensuring the maintenance of virulence genes in free-living conditions. This strategy is likely widespread amongst native plasmids of P. syringae and other bacteria.
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    PublicationOpen Access
    Papel de ginA y ginB en la movilidad de la isla genómica GInt
    (2014) Echeverría Ancín, Myriam; Bardají Goikoetxea, Leire; Escuela Técnica Superior de Ingenieros Agrónomos; Nekazaritza Ingeniarien Goi Mailako Eskola Teknikoa
    La transferencia horizontal de genes, mediada por elementos genéticos móviles tales como las islas genómicas, es uno de los principales mecanismos de generación de variabilidad genética en bacterias, permitiendo su adaptación a nuevos nichos ecológicos. En el grupo de Patología Vegetal de la UPNA se ha descrito, en especies de Pseudomonas, una nueva familia de islas genómicas, denominadas GInt. Éstas se caracterizan por contener en su extremo 5’ el operón gin, constituido por cuatro genes, ginA, ginB, ginC y ginD, tres de los cuales codifican recombinasas de tirosina. Este tipo de proteínas participan en la escisión, integración y otros reordenamientos de ADN mediante recombinación específica de sitio. Los GInts se escinden formando moléculas circulares extracromosómicas, o episomas. En islas genómicas ya descritas, éste es un paso fundamental previo a la transferencia horizontal del elemento. Como parte de la caracterización funcional de los GInts se evaluó la implicación de los genes ginA y ginB en la formación de episomas. Mediante mutagénesis dirigida, utilizando la técnica de intercambio alélico, se generaron mutantes no polares de ginA y de ginB. Estos fueron incapaces de generar episomas, a tenor de los resultados de PCR anidada. Sin embargo, los mutantes complementados en trans, con una copia silvestre del gen en cuestión, restauraron completamente el fenotipo. Los resultados obtenidos en este trabajo demuestran que ginA y ginB son esenciales en la movilidad de los GInts ya que participan en el proceso de formación de estos intermediarios circulares.
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    PublicationOpen Access
    Four genes essential for recombination define GInts, a new type of mobile genomic island widespread in bacteria
    (Nature Publishing Group, 2017) Bardají Goikoetxea, Leire; Echeverría Ancín, Myriam; Rodríguez Palenzuela, Pablo; Martínez García, Pedro M.; Murillo Martínez, Jesús; Producción Agraria; Nekazaritza Ekoizpena
    Integrases are a family of tyrosine recombinases that are highly abundant in bacterial genomes, actively disseminating adaptive characters such as pathogenicity determinants and antibiotics resistance. Using comparative genomics and functional assays, we identified a novel type of mobile genetic element, the GInt, in many diverse bacterial groups but not in archaea. Integrated as genomic islands, GInts show a tripartite structure consisting of the ginABCD operon, a cargo DNA region from 2.5 to at least 70 kb, and a short AT-rich 3′ end. The gin operon is characteristic of GInts and codes for three putative integrases and a small putative helix-loop-helix protein, all of which are essential for integration and excision of the element. Genes in the cargo DNA are acquired mostly from phylogenetically related bacteria and often code for traits that might increase fitness, such as resistance to antimicrobials or virulence. GInts also tend to capture clusters of genes involved in complex processes, such as the biosynthesis of phaseolotoxin by Pseudomonas syringae. GInts integrate site-specifically, generating two flanking direct imperfect repeats, and excise forming circular molecules. The excision process generates sequence variants at the element attachment site, which can increase frequency of integration and drive target specificity.