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The abundant extrachromosomal DNA content of the Spiroplasma citri GII3-3X genome

Université Victor Ségalen Bordeaux 2, UMR 1090 Génomique Diversité Pouvoir Pathogène, BP 81, F-33883 Villenave d'Ornon, France.
BMC Genomics (Impact Factor: 4.04). 02/2008; 9:195. DOI: 10.1186/1471-2164-9-195
Source: PubMed

ABSTRACT Spiroplama citri, the causal agent of citrus stubborn disease, is a bacterium of the class Mollicutes and is transmitted by phloem-feeding leafhopper vectors. In order to characterize candidate genes potentially involved in spiroplasma transmission and pathogenicity, the genome of S. citri strain GII3-3X is currently being deciphered.
Assembling 20,000 sequencing reads generated seven circular contigs, none of which fit the 1.8 Mb chromosome map or carried chromosomal markers. These contigs correspond to seven plasmids: pSci1 to pSci6, with sizes ranging from 12.9 to 35.3 kbp and pSciA of 7.8 kbp. Plasmids pSci were detected as multiple copies in strain GII3-3X. Plasmid copy numbers of pSci1-6, as deduced from sequencing coverage, were estimated at 10 to 14 copies per spiroplasma cell, representing 1.6 Mb of extrachromosomal DNA. Genes encoding proteins of the TrsE-TraE, Mob, TraD-TraG, and Soj-ParA protein families were predicted in most of the pSci sequences, in addition to members of 14 protein families of unknown function. Plasmid pSci6 encodes protein P32, a marker of insect transmissibility. Plasmids pSci1-5 code for eight different S. citri adhesion-related proteins (ScARPs) that are homologous to the previously described protein P89 and the S. kunkelii SkARP1. Conserved signal peptides and C-terminal transmembrane alpha helices were predicted in all ScARPs. The predicted surface-exposed N-terminal region possesses the following elements: (i) 6 to 8 repeats of 39 to 42 amino acids each (sarpin repeats), (ii) a central conserved region of 330 amino acids followed by (iii) a more variable domain of about 110 amino acids. The C-terminus, predicted to be cytoplasmic, consists of a 27 amino acid stretch enriched in arginine and lysine (KR) and an optional 23 amino acid stretch enriched in lysine, aspartate and glutamate (KDE). Plasmids pSci mainly present a linear increase of cumulative GC skew except in regions presenting conserved hairpin structures.
The genome of S. citri GII3-3X is characterized by abundant extrachromosomal elements. The pSci plasmids could not only be vertically inherited but also horizontally transmitted, as they encode proteins usually involved in DNA element partitioning and cell to cell DNA transfer. Because plasmids pSci1-5 encode surface proteins of the ScARP family and pSci6 was recently shown to confer insect transmissibility, diversity and abundance of S. citri plasmids may essentially aid the rapid adaptation of S. citri to more efficient transmission by different insect vectors and to various plant hosts.

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    • "We designed the primer pair Scif⁄Scir based on the pE gene of pSci plasmids (Breton et al. 2008). Seven circular multicopied plasmids have been reported in S. citri (Saillard et al. 2008), and CDS pE region is common to all six of the pSci plasmids (Breton et al. 2008). From 43 symptomatic plants examined in this research, 79% were positive by primer pair pScif⁄pScir, while nested PCR of the spiralin gene based on primer pairs D⁄D¢ (Foissac et al. 1996) followed by F1⁄R1 (Khanchezar et al. 2010) in the second round resulted in detection of only 36.6% infection in symptomatic samples. "
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    • "cutes and other bacteria by leafhoppers. For some S. citri strains, although tools such as random mutagenesis through transposition, site-directed mutagenesis via homologous recombination , plasmid vectors, and inducible gene expression systems have been reported (Renaudin and Bove 1994; Ye et al. 1994; Foissac et al. 1997; Melcher et al. 1999; Duret et al. 2005; Berho et al. 2006; Breton et al. 2008; Saillard et al. 2008), genetic tools for the study of S. citri remain limited. The work reported here demonstrates that transposomes have the potential of being useful tools. "
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