Draft Genome Sequence of Erwinia billingiae OSU19-1, Isolated from a Pear Tree Canker.

Plant-associated Erwinia include pathogenic and nonpathogenic species. We report the 5.6-Mb genome sequence of Erwinia billingiae OSU19-1, isolated from a canker on a pear tree inoculated with Erwinia amylovora. OSU19-1 and a closely related European isolate, E. billingiae Eb661(T), share many similarities including 40 kb of plasmid sequence.

GENOME ANNOUNCEMENT

Plant-associated Gram-negative Erwinia spp. belong to the Enterobacteriaceae family. Species include plant epiphyte Erwinia billingiae and pathogenic Erwinia amylovora, which causes fire blight (1, 2). We isolated E. billingiae OSU19-1 from a canker on the mid-trunk cambium of a Concorde pear tree inoculated with E. amylovora 1 year prior; others also found E. billingiae in necrotic tissue of trees with fire blight (1–3). We surface sterilized tissue in 10% bleach, rinsed with sterile water, pulverized tissue in sterile water, and isolated bacteria on LB agar at 25°C. We used a MoBio PowerSoil kit to purify genomic DNA from bacteria cultured in LB broth (4).

To compare E. billingiae OSU19-1 with Eb661T, the type strain isolated in England in 1959, we sequenced the genome of OSU19-1 using an Illumina MiSeq to generate 250-bp paired-end reads. Low-quality read pairs flagged by the MiSeq were removed prior to quality trimming with Sickle (quality score = 30; minimum length = 50), yielding 2,172,379 read pairs; single reads were discarded. To obtain optimum coverage (~65×), we used 663,000 read pairs to complete four de novo assemblies with ABySS, Celera Assembler, IDBA, and Velvet (5–9). Optimum k-mer lengths of 65, 120, and 87 were used for the ABySS, IDBA, and Velvet assemblies, respectively. We used Minimus2 to merge the Celera and Velvet assemblies; ABySS and IDBA assemblies were merged similarly (10). These combined assemblies were merged into a consensus alignment, which was validated using REAPR (11). The final assembly contained 5,602,087 bp (55% G/C) in 32 contigs (N50, 409,442 bp; maximum, 668,909).

Annotation using RAST (12) predicted 4,931 protein coding sequences and 76 RNAs. BLASTn analysis of recA, gyrA, gyrB, and gpd genes revealed 98 to 99% identity between OSU19-1 and its closest known relative, E. billingiae Eb661T (3); small-subunit rRNA genes were identical. OSU19-1 encodes several quorum-sensing systems, including LuxS, which produces autoinducer-2, and two homoserine lactone synthases. Similarly, Eb661T produces an acyl-homoserine lactone (13) and autoinducer-2 (14). OSU19-1 and Eb661 lack a type III secretion system, which is important for pathogenicity of E. amylovora (3).

The 5.37-Mb genome of Eb661T has a chromosome of 5,100,168 bp and two plasmids, pEB102 (102 kb) and pEB170 (170 kb) (3). OSU19-1 lacks significant similarity to pEB170, but contigs 2 (98,580 bp) and 10 (40,687 bp) share 26,170 bp and 14,490 bp (94 to 96% identity) with different regions of pEB102. The portion of contig 2 homologous to pEB102 includes four genes resembling those in the integrative conjugative element Genomic Island-1 of Pseudomonas fluorescens Pf-5 (15). Outside this shared region, contig 2 encodes RepA and ParAB proteins; it also encodes DNA repair proteins UmuC and RadC, which may help E. billingiae survive exposure to UV radiation. Study of genetic diversity in E. billingiae increases understanding of this microbe’s lifestyle and its potential for biocontrol of fire blight (3, 13).

Nucleotide sequence accession numbers.

This whole-genome shotgun project has been deposited at DDBJ/EMBL/GenBank under the accession no. LHXI00000000. The version described in this paper is version LHXI01000000.