Complete genome sequence of the fire blight pathogen Erwinia pyrifoliae DSM 12163T and comparative genomic insights into plant pathogenicity

Abstract

Background: Erwinia pyrifoliae is a newly described necrotrophic pathogen, which causes fire blight on Asian (Nashi) pear and is geographically restricted to Eastern Asia. Relatively little is known about its genetics compared to the closely related main fire blight pathogen E. amylovora.

Results: The genome of the type strain of E. pyrifoliae strain DSM 12163T, was sequenced using both 454 and Solexa pyrosequencing and annotated. The genome contains a circular chromosome of 4.026 Mb and four small plasmids. Based on their respective role in virulence in E. amylovora or related organisms, we identified several putative virulence factors, including type III and type VI secretion systems and their effectors, flagellar genes, sorbitol metabolism, iron uptake determinants, and quorum-sensing components. A deletion in the rpoS gene covering the most conserved region of the protein was identified which may contribute to the difference in virulence/host-range compared to E. amylovora. Comparative genomics with the pome fruit epiphyte Erwinia tasmaniensis Et1/99 showed that both species are overall highly similar, although specific differences were identified, for example the presence of some phage gene-containing regions and a high number of putative genomic islands containing transposases in the E. pyrifoliae DSM 12163T genome.

Conclusions: The E. pyrifoliae genome is an important addition to the published genome of E. tasmaniensis and the unfinished genome of E. amylovora providing a foundation for re-sequencing additional strains that may shed light on the evolution of the host-range and virulence/pathogenicity of this important group of plant-associated bacteria.

Figure 1

Circular representation of the chromosome of E. pyrifoliae DSM 12163^T. Circles (from outside to inside) First: scale bar in kb; second and third: predicted coding sequences of E. pyrifoliae DSM 12163^T chromosome on the leading and lagging strand, respectively (colors according to COGs); fourth: coding sequences of the hrp/hrc T3SS (red), the inv/spa T3SS (green), the T6SS clusters (blue), the flagellar genomic island (yellow), dispersed flagellar gene clusters (purple) and the EPS biosynthetic cluster (orange); fifth, G+C content; sixth, G+C skew.

Figure 2

Comparison of the annotated sequences of E. pyrifoliae DSM 12163^T plasmids pEP5, pEP3 and pEP2.6. Genes with high homology are indicated in the same color. Grey shading indicate regions with DNA sequence identity.

Figure 3

Electropherogram of amplified rpoS genes from different Erwinia species. Lane 1: E. amylovora CFBP 1232^T; lane 2: E. amylovora OR29; lane 3: E. pyrifoliae DSM 12163^T; lane 4: E. pyrifoliae CFBP 4174 and lane 5: E. billingae LMG 2613^T. A minus sign denotes the negative control (-); M denotes the marker (Fermentas GeneRuler DNA Ladder Mix). Relevant marker sizes (in bp) are indicated at the left side of the figure. The rpoS gene was amplified using primer set Ep-rpoS-F (5'-AGTACTGGCACGAGTTCTGTTAGA-3') and Ep-rpoS-R (5'-TGCAGTATTTCACGCAGACGACGC-3'). The expected amplicon size of an intact rpoS gene is 1109 bp; for the 140 bp deletion the amplicon is 969 bp.

Figure 4

Comparison of inv/spa-type Type III Secretion Systems in E. pyrifoliae DSM 12163^T and E. tasmaniensis Et1/99. For the comparison, the annotation of the E. tasmaniensis Et1/99 clusters was manually checked. Previously denoted pseudogenes were shown to have close orthologs in E. pyrifoliae DSM 12163^T, while "missing" genes in E. tasmaniensis Et1/99 [16] could be found. Blocks of related genes are shaded grey. Putative T3SS core genes are colored green, with low homology genes in light green. Effectors are colored red, regulatory genes black and chaperones blue. Genes with no homology are in white.

Figure 5

Gene organization of Type VI Secretion System (T6SS) gene clusters. (A) Large T6SS gene clusters. (B) Small T6SS gene clusters. Blocks of related genes are shaded grey. Putative core genes are colored green, putative effectors red, putative signal transducers black, conserved genes between clusters grey and genes without related or homologues in all other clusters white. The frameshift in EPYR_02110/EPYR_02111 is indicated by a red flash.

Figure 6

Comparison of the exopolysaccharide (EPS) biosynthetic clusters of E. pyrifoliae DSM 12163^T (middle), E. amylovora Ea7/74 (top) and E. tasmaniensis Et1/99 (bottom). Identical colours indicate identical predicted functions. White arrows indicate flanking genes probably not involved in EPS biosynthesis. The numbers between the gene clusters indicate the sequence identity of the translated gene products indicated.

Figure 7

Mauve progressive alignment of the genomes of E. pyrifoliae DSM 12163^T (top) and E. tasmaniensis Et1/99 (bottom). Some relevant features within regions that have large differences in the alignment are indicated with arrows, plasmids are indicated as horizontal bars. Abbreviations: GI: genomic islands; T1RS: type 1 restriction system; T2RS: type 2 restriction sytem; Tn: transposase, T3SS: type 3 secretion system, T4SS: type 4 secretion system.