Draft Genome Sequence of Pseudomonas sp. Strain M47T1, Carried by Bursaphelenchus xylophilus Isolated from Pinus pinaster

Abstract

The draft genome sequence of Pseudomonas sp. strain M47T1, carried by the Bursaphelenchus xylophilus pinewood nematode, the causative agent of pine wilt disease, is presented. In Pseudomonas sp. strain M47T1, genes that make this a plant growth-promoting bacterium, as well as genes potentially involved in nematotoxicity, were identified.

GENOME ANNOUNCEMENT

Pine wilt disease (PWD) is caused by Bursaphelenchus xylophilus, the pinewood nematode (PWN), which is a serious pest and pathogen of forest trees of the Pinus species (5). Strains of the genus Pseudomonas were suggested to play a role in the disease (10), but their specific functions and contributions to the disease are still unclear. Isolation of Pseudomonas sp. strain M47T1 was achieved from trails made on R2A by B. xylophilus from a Pinus pinaster with PWD (7). Sequencing of the 16S rRNA gene was performed, and the strain was tested for the ability to kill nematodes (7). Culture supernatants of Gram-negative Pseudomonas sp. strain M47T1 (classified as Pseudomonas sp. M47T1 [7]) killed B. xylophilus nematodes in vitro within 24 h of incubation (D. N. Proença, unpublished results).

Reads were generated by 454 GS FLX sequencing (4), and raw data were assembled by using GS De Novo Assembler (Newbler), version 2.5.3. The assembled contigs were submitted to the RAST annotation server for subsystem classification and functional annotation (1). Coding sequences (CDSs) were assigned using BLASTp with KEGG Orthology (KO). The G+C content was calculated using an in-house Perl script. The NCBI Prokaryotic Genomes Automatic Annotation Pipeline (PGAAP) was employed for gene annotation in preparation for submission to GenBank (http://www.ncbi.nlm.nih.gov/genomes/static/Pipeline.html).

The draft genome sequence of Pseudomonas sp. strain M47T1 comprises 6,311,977 bases, representing a 49-fold coverage of the genome. The assembled genome consists of 72 large contigs with more than 500 bp (N₅₀ contig size was approximately 87,580 bp). The G+C content was 61.80%. The genome encodes 5,678 putative coding sequences (CDSs). The draft genome sequence contains six ribosomal RNAs and 69 tRNA loci. For the CDSs, 82.4%, corresponding to 4,679 proteins, could be assigned to Cluster of Orthologous Groups (COG) families (8). Comparison with genome sequences available at RAST showed that 30 of the closest strains belong to eight genera (Azoarcus, Azotobacter, Cellvibrio, Chromohalobacter, Hahella, Marinobacter, Marinomonas, and Shewanella) outside the pseudomonads.

The genome of Pseudomonas sp. strain M47T1 carries multiple genes potentially involved in nematotoxic activity such as bacteriocin and colicin V biosynthesis, the YdhE/NorM gene encoding a homolog of a multidrug and toxin extrusion (MATE) family efflux pump, three genes coding for Rhs family proteins that may be involved in adhesion or virulence (3, 6), and a gene encoding a calcium-binding protein from the bacterial RTX toxin (repeats in toxins) family (9).

Moreover, strain M47T1 also contains genes that are potentially involved in promoting plant growth, such as genes encoding ferric siderophores and for phenazine biosynthesis (2). This strain also has a set of genes typical for plant niche adaptation, including the genes for NodD, which regulates the nodulation genes, and nitrogen regulatory protein P-II (ammonia assimilation) and acetoin (diacetyl) reductase for plant protection against fungal and bacterial infections. In short, the genomic information suggests a positive interaction with the plant but a negative interaction with the PWN worm. Thus, Pseudomonas sp. strain M47T1 may be a good candidate to be developed as a bionematocide against an important pest of coniferous plants.

Nucleotide sequence accession numbers.

This Whole Genome Shotgun project has been deposited at DDBJ/EMBL/GenBank under accession no. AJWX00000000. The version described in this paper is the first version, AJWX01000000. The genome project data are also available at GenBank under the genome Bioproject ID PRJNA162465.