Comparative analysis of mobilizable genomic islands - PubMed (original) (raw)

Comparative Study

. 2013 Feb;195(3):606-14.

doi: 10.1128/JB.01985-12. Epub 2012 Nov 30.

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Comparative Study

Comparative analysis of mobilizable genomic islands

Aurélie Daccord et al. J Bacteriol. 2013 Feb.

Abstract

Mobilizable genomic islands (MGIs) are small genomic islands of less than 35 kbp containing an integrase gene and a sequence that resembles the origin of transfer (oriT) of an integrating conjugative element (ICE). MGIs have been shown to site-specifically integrate and excise from the chromosome of bacterial hosts and hijack the conjugative machinery of a coresident ICE to disseminate. To date, MGIs have been described in three strains belonging to three different Vibrio species. In this study, we report the discovery of 11 additional putative MGIs found in various species of Vibrio, Alteromonas, Pseudoalteromonas, and Methylophaga. We designed an MGI capture system that allowed us to relocate chromosomal MGIs onto a low-copy-number plasmid and facilitate their isolation and sequencing. Comparative genomics and phylogenetic analyses of these mobile genetic elements revealed their mosaic structure and their evolution through recombination and acquisition of exogenous DNA. MGIs were found to belong to a larger family of genomic islands (GIs) sharing a similar integrase gene and often integrated into the same integration site yet exhibiting a different mechanism of regulation of excision and mobilization. We found that MGIs can excise only when an ICE of the SXT/R391 family is coresident in the same cell, while GIs still excise regardless.

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Figures

Fig 1

Fig 1

MGI capture system. Conjugation between a donor strain bearing an ICE and an MGI with an intermediate strain (AD175) bearing R997 (1) but lacking attB and dapA yields exconjugants that contain the transferred MGI integrated into pVB201. These exconjugants cannot be directly selected because of the lack of a selective marker on the MGI; yet the resulting plasmid becomes mobilizable by R997 and transfer to a Δ_attB_ recipient strain (2), yielding exconjugants harboring pVB201::MGI (3) that can be selected using the chloramphenicol marker present on pVB201 and the ability of the recipient strain (AD157) to grow in the presence of tetracycline and in the absence of diaminopimelic acid (dap).

Fig 2

Fig 2

General structures of MGIs and phylogenetic analyses of the core genes. (A) Genetic structure of 12 sequenced MGIs; (B) atypical structure of GI_Vch_Hai3, from Vibrio cholerae H-09; (C) genetic content of GI_Spu_PO1 and GI_Vmi_USA1 from Shewanella sp. W3-18-1 and Vibrio mimicus VM573, respectively. Colors of the predicted open reading frames indicate the following putative functions: pink and red, DNA replication, recombination, and repair; purple, diguanylate cyclase/phosphodiesterase; blue, transcription; green, restriction modification systems; orange, putative HipA-like toxin; gray, unknown function. See Table S2 in the supplemental material for predictions of putative function of genes identified in variable regions.

Fig 3

Fig 3

Phylogenetic analysis of the MGI backbone. Phylogenetic trees based on the nucleotide sequences of the indicated core genes or region. Bootstrap values are indicated when over 80%. The individual scale bars represent genetic distances.

Fig 4

Fig 4

Phylogenetic analysis of MGIs and related GIs. (A) Tree based on protein sequences of Int from SXT, MGIs, and 31 related GIs; (B) phylogenetic tree based on the nucleotide sequences of the rpoB gene from the corresponding strains. Bootstrap values are indicated when over 80%. The individual scale bars represent genetic distances. Refer to Table S3 in the supplemental material for GI and strain details.

Fig 5

Fig 5

Excision of MGIs and related GIs. (A) Ethidium bromide-stained 1.5% agarose gels of PCR products amplified by nested PCR assays for detection of the excision of the MGIs and related GIs. Lanes: M, 2-log DNA ladder; 1, MGI_Vpa_Moz1; 2, MGI_Vch_Moz2; 3, MGI_Vch_Moz3; 4, MGI_Vch_Moz4; 5, MGI_Vch_Moz6; 6 and 7, MGI_Ama_Med1; 8, GI_Vmi_USA1 from Vibrio mimicus VM573; 9, GI_Spu_PO1 from Shewanella sp. W3-18-1. Lane 7, coresident R391 was present in the cells. (B) Model of excision/integration of MGI_Ama_Med1 by site-specific recombination (solid crossover) and by homologous recombination in the absence of R391 (dotted crossover). Refer to Fig. S3 in the supplemental material for details about the alignment of MGI_Ama_Med1 attachment sites.

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