Complete sequence and detailed analysis of the first indigenous plasmid from Xanthomonas oryzae pv. oryzicola

Abstract

BackgroundBacterial plasmids have a major impact on metabolic function and adaptation of their hosts. An indigenous plasmid was identified in a Chinese isolate (GX01) of the invasive phytopathogen Xanthomonas oryzae pv. oryzicola (Xoc), the causal agent of rice bacterial leaf streak (BLS). To elucidate the biological functions of the plasmid, we have sequenced and comprehensively annotated the plasmid.MethodsThe plasmid DNA was extracted from Xoc strain GX01 by alkaline lysis and digested with restriction enzymes. The cloned and subcloned DNA fragments in pUC19 were sequenced by Sanger sequencing. Sequences were assembled by using Sequencher software. Gaps were closed by primer walking and sequencing, and multi-PCRs were conducted through the whole plasmid sequence for verification. BLAST, phylogenetic analysis and dinucleotide calculation were performed for gene annotation and DNA structure analysis. Transformation, transconjugation and stress tolerance tests were carried out for plasmid function assays.ResultsThe indigenous plasmid from Xoc strain GX01, designated pXOCgx01, is 53,206-bp long and has been annotated to possess 64 open reading frames (ORFs), including genes encoding type IV secretion system, heavy metal exporter, plasmid stability factors, and DNA mobile factors, i.e., the Tn3-like transposon. Bioinformatics analysis showed that pXOCgx01 has a mosaic structure containing different genome contexts with distinct genomic heterogeneities. Phylogenetic analysis indicated that the closest relative of pXOCgx01 is pXAC64 from Xanthomonas axonopodis pv. citri str. 306. It was estimated that there are four copies of pXOCgx01 per cell of Xoc GX01 by PCR assay and the calculation of whole genome shotgun sequencing data. We demonstrate that pXOCgx01 is a self-transmissible plasmid and can replicate in some Xanthomonas spp. strains, but not in Escherichia coli DH5α. It could significantly enhance the tolerance of Xanthomonas oryzae pv. oryzae PXO99A to the stresses of heavy metal ions. The plasmid survey indicated that nine out of 257 Xoc Chinese isolates contain plasmids.ConclusionspXOCgx01 is the first report of indigenous plasmid from Xanthomonas oryzae pv. oryzicola, and the first completely sequenced plasmid from Xanthomonas oryzae species. It is a self-transmissible plasmid and has a mosaic structure, containing genes for macromolecule secretion, heavy metal exportation, and DNA mobile factors, especially the Tn3-like transposon which may provide transposition function for mobile insertion cassette and play a major role in the spread of pathogenicity determinants. The results will be helpful to elucidate the biological significance of this cryptic plasmid and the adaptive evolution of Xoc.Electronic supplementary materialThe online version of this article (doi:10.1186/s12866-015-0562-x) contains supplementary material, which is available to authorized users.

Adaptation of genetically monomorphic bacteria: evolution of copper resistance through multiple horizontal gene transfers of complex and versatile mobile genetic elements

“…It contained genes that are syntenic and similar to genes from the plasmid‐encoded Xanthomonas Tn Xo19 , a Tn 3 ‐like transposon (Niu et al . ). This includes a transposon‐related cointegrate protein tnpT , a cointegrate protein tnpS , a transposase tnpA , a DNA recombination protein and a DNA helicase, that all display nucleotide identity (NI) between 70% and 90% with their Tn Xo19 homologues, on 94%, 70%, 26%, 100% and 100% of TnPLH201.1's gene length, respectively.…”

“…oryzae , (Niu et al . )], a heavy metal efflux protein, a copper‐sensing transcriptional repressor (which binds to a gene promoter and to copper with a higher affinity to copper) and a metal binding exoribonuclease (which might help degrade the transcriptional repressor in the presence of copper).…”

Identification of new protein-coding genes with a potential role in the virulence of the plant pathogen Xanthomonas euvesicatoria

“…Interestingly, XCV_PG01 , located between XCV0062 and XCV0063, encodes a putative serine/threonine phosphatase of the 2C family, which was previously overlooked. Furthermore, we identified a putative YecR-like lipoprotein, XCV _PG06, which was recently annotated in X. oryzicola [ 27 ] . A special case is XCV_PG30 , which is encoded antisense to gcvP , predicted to encode a metal-dependent RNase.…”

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