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      Diversity and Homogeneity among Small Plasmids of Aeromonas salmonicida subsp. salmonicida Linked with Geographical Origin

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          Abstract

          Furunculosis, which is caused by Aeromonas salmonicida subsp. salmonicida, is a major salmonid disease in fish farms worldwide. Several plasmids found in this bacterium confer phenotypes such drug resistance and virulence. Small plasmids (pAsa1, pAsa2, pAsa3, and pAsal1) related to ColE1- and ColE2-type replicons are usually present in its normal plasmidome. In the present study, with the objective to investigate if these plasmids display particularities related to the origin of the isolates bearing them, a total of 153 isolates, including 78 new and 75 previously described, were analyzed for the presence of small plasmids by PCR and DNA restriction fragment profiling. A geographical dichotomy between Canadian and European isolates for their propensity to do not have pAsa3 or pAsal1 was found. In addition, the genotyping analysis led to the identification of two European isolates harboring an unusual pAsal1. An investigation by next-generation sequencing (NGS) of these two isolates shed light on two pAsal1 variants (pAsal1C and pAsal1D). As with pAsal1B, another pAsal1 variant previously described, these two new variants bore a second insertion sequence (IS AS5) in addition to the usual IS AS11. The characterization of these variants suggested that they could predominate over the wild-type pAsal1 in stressful conditions such as growth at temperatures of 25°C and above. To obtain a comprehensive portrait of the mutational pressure on small plasmids, 26 isolates whose DNA had been sequenced by NGS were investigated. pAsa3 and pAsal1 were more prone to mutations than pAsa1 and pAsa2, especially in the mobA gene, which encodes a relaxase and a primase. Lastly, the average copy number of each plasmid per cell was assessed using raw sequencing data. A clear trend with respect to the relative proportion per cell of each plasmid was identified. Our large-scale study revealed a geographical dichotomy in small plasmid repertoire in addition to a clear trend for pAsa3 and pAsal1 to be more frequently altered. Moreover, we present the discovery of two new variants of pAsal1: pAsal1C and pAsal1D.

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          Most cited references 38

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          Improved tools for biological sequence comparison.

          We have developed three computer programs for comparisons of protein and DNA sequences. They can be used to search sequence data bases, evaluate similarity scores, and identify periodic structures based on local sequence similarity. The FASTA program is a more sensitive derivative of the FASTP program, which can be used to search protein or DNA sequence data bases and can compare a protein sequence to a DNA sequence data base by translating the DNA data base as it is searched. FASTA includes an additional step in the calculation of the initial pairwise similarity score that allows multiple regions of similarity to be joined to increase the score of related sequences. The RDF2 program can be used to evaluate the significance of similarity scores using a shuffling method that preserves local sequence composition. The LFASTA program can display all the regions of local similarity between two sequences with scores greater than a threshold, using the same scoring parameters and a similar alignment algorithm; these local similarities can be displayed as a "graphic matrix" plot or as individual alignments. In addition, these programs have been generalized to allow comparison of DNA or protein sequences based on a variety of alternative scoring matrices.
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            ISfinder: the reference centre for bacterial insertion sequences

            ISfinder () is a dedicated database for bacterial insertion sequences (ISs). It has superseded the Stanford reference center. One of its functions is to assign IS names and to provide a focal point for a coherent nomenclature. It is also the repository for ISs. Each new IS is indexed together with information such as its DNA sequence and open reading frames or potential coding sequences, the sequence of the ends of the element and target sites, its origin and distribution together with a bibliography where available. Another objective is to continuously monitor ISs to provide updated comprehensive groupings or families and to provide some insight into their phylogenies. The site also contains extensive background information on ISs and transposons in general. Online tools are gradually being added. At present an online Blast facility against the entire bank is available. But additional features will include alignment capability, PsiBLAST and HMM profiles. ISfinder also includes a section on bacterial genomes and is involved in annotating the IS content of these genomes. Finally, this database is currently recommended by several microbiology journals for registration of new IS elements before their publication.
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              A5-miseq: an updated pipeline to assemble microbial genomes from Illumina MiSeq data.

              Open-source bacterial genome assembly remains inaccessible to many biologists because of its complexity. Few software solutions exist that are capable of automating all steps in the process of de novo genome assembly from Illumina data.
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                Author and article information

                Contributors
                Journal
                Front Microbiol
                Front Microbiol
                Front. Microbiol.
                Frontiers in Microbiology
                Frontiers Media S.A.
                1664-302X
                23 November 2015
                2015
                : 6
                Affiliations
                1Département de Biochimie, de Microbiologie et de Bio-informatique, Faculté des Sciences et de Génie, Université Laval Quebec City, QC, Canada
                2Institut de Biologie Intégrative et des Systèmes, Université Laval Quebec City, QC, Canada
                3Centre de Recherche de l'Institut Universitaire de Cardiologie et de Pneumologie de Québec Quebec City, QC, Canada
                Author notes

                Edited by: Estelle Jumas-Bilak, Université de Montpellier, France

                Reviewed by: Antonio C. M. Correia, Universidade de Aveiro, Portugal; Carlos R. Osorio, University of Santiago de Compostela, Spain

                *Correspondence: Steve J. Charette steve.charette@ 123456bcm.ulaval.ca

                This article was submitted to Aquatic Microbiology, a section of the journal Frontiers in Microbiology

                †These authors have contributed equally to this work.

                Article
                10.3389/fmicb.2015.01274
                4655240
                Copyright © 2015 Attéré, Vincent, Trudel, Chanut and Charette.

                This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) or licensor are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.

                Page count
                Figures: 4, Tables: 1, Equations: 0, References: 46, Pages: 9, Words: 7234
                Funding
                Funded by: Natural Sciences and Engineering Research Council of Canada 10.13039/501100000038
                Funded by: Fonds de Recherche du Québec - Nature et Technologies 10.13039/501100003151
                Funded by: Ressources Aquatiques Québec
                Funded by: Ministère de l'Agriculture, des Pêcheries et de l'Alimentation 10.13039/100008777
                Categories
                Microbiology
                Original Research

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