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

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 (ISAS5) in addition to the usual ISAS11. 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.

SUBMITTER: Attere SA

PROVIDER: S-EPMC4655240 | biostudies-literature | 2015

REPOSITORIES: biostudies-literature

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

Attéré Sabrina A SA Vincent Antony T AT Trudel Mélanie V MV Chanut Romain R Charette Steve J SJ

Frontiers in microbiology 20151123

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 bea ...[more]

PMID: 26635745

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Project description:In Aeromonas salmonicida subsp. salmonicida, a bacterium that causes fish disease, there are two types of small plasmids (<15 kbp): plasmids without known function, called cryptic plasmids, and plasmids that bear beneficial genes for the bacterium. Four among them are frequently detected in strains of A. salmonicida subsp. salmonicida: pAsa1, pAsa2, pAsa3, and pAsal1. The latter harbors a gene which codes for an effector of the type three secretion system, while the three others are cryptic. It is currently unclear why these cryptic plasmids are so highly conserved throughout strains of A. salmonicida subsp. salmonicida. In this study, three small plasmids, named pAsa10, pAsaXI and pAsaXII, are described. Linked to tetracycline resistance, a partial Tn1721 occupies half of pAsa10. A whole Tn1721 is also present in pAsa8, another plasmid previously described in A. salmonicida subsp. salmonicida. The backbone of pAsa10 has no relation with other plasmids described in this bacterium. However, the pAsaXI and pAsaXII plasmids are derivatives of cryptic plasmids pAsa3 and pAsa2, respectively. pAsaXI is identical to pAsa3, but bears a transposon with a gene that encodes for a putative virulence factor. pAsaXII, also found in Aeromonas bivalvium, has a 95% nucleotide identity with the backbone of pAsa2. Like pAsa7, another pAsa2-like plasmid recently described, orf2 and orf3 are missing and are replaced in pAsaXII by genes that encode a formaldehyde detoxification system. These new observations suggest that transposons and particularly Tn1721 are frequent and diversified in A. salmonicida subsp. salmonicida. Moreover, the discovery of pAsaXI and pAsaXII expands the group of small plasmids that are derived from cryptic plasmids and have a function. Although their precise roles remain to be determined, the present study shows that cryptic plasmids could serve as moldable vectors to acquire mobile elements such as transposons. Consequently, they could act as key agents of the diversification of virulence and adaptive traits of Aeromonas salmonicida subsp. salmonicida.

Project description:Background Aeromonas salmonicida subsp. salmonicida, the etiologic agent of furunculosis, is a major pathogen of fisheries worldwide. Despite the identification of several virulence factors the pathogenesis is still poorly understood. We have used high-throughput proteomics to display the differences between in vitro secretome of A. salmonicida wild-type (wt, hypervirulent, JF5054) and T3SS-deficient (isogenic DeltaascV, extremely low-virulent, JF2747) strains in exponential (GP) and stationary (SP) phases of growth. Results Among the different experimental conditions we obtained semi-quantitative values for a total of 2136 A. salmonicida proteins. Proteins of specific A. salmonicida species were proportionally less detected than proteins common to the Aeromonas genus or those shared with other Aeromonas species, suggesting that in vitro growth did not induce the expression of these genes. Four detected proteins which are unidentified in the genome of reference strains of A. salmonicida were homologous to components of the conjugative T4SS of A. hydrophila pRA1 plasmid. Polypeptides of three proteins which are specific to the 01-B526 strain were also discovered. In supernatants (SNs), the number of detected proteins was higher in SP (326 for wt vs 329 for mutant) than in GP (275 for wt vs 263 for mutant). In pellets, the number of identified proteins (a total of 1536) was approximately the same between GP and SP. Numerous highly conserved cytoplasmic proteins were present in A. salmonicida SNs (mainly EF-Tu, EF-G, EF-P, EF-Ts, TypA, AlaS, ribosomal proteins, HtpG, DnaK, peptidyl-prolyl cis-trans isomerases, GAPDH, Enolase, FbaA, TpiA, Pgk, TktA, AckA, AcnB, Mdh, AhpC, Tpx, SodB and PNPase), and several evidences support the theory that their extracellular localization was not the result of cell lysis. According to the Cluster of Orthologous Groups classification, 29% of excreted proteins in A. salmonicida SNs were currently poorly characterized. Conclusions In this part of our work we elucidated the whole in vitro exoproteome of hypervirulent A. salmonicida subsp. salmonicida and showed the secretion of several highly conserved cytoplasmic proteins with putative moonlighting functions and roles in virulence. All together, our results offer new information about the pathogenesis of furunculosis and point out potential candidates for vaccine development.

Dataset Information

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

Publications

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

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