Genome Sequences of Pseudomonas fragi Strains A22 and B25

Abstract

Pseudomonas fragi A22 is a novel isolate that produces bead-like particles (A22B) in its cell wall. To explore the genetic basis for the formation of A22B, P. fragi A22 and the type strain of the species, P. fragi B25, were subjected to genome sequence analysis. Here, we report the draft genome sequences and automatic annotation of both strains. These data offer a solid base for related studies of P. fragi, including comparative genomics, proteomics, and gene mining.

GENOME ANNOUNCEMENT

A Gram-negative bacterial strain, A22, was isolated while screening lipid-accumulating microorganisms by using Sudan black staining. The strain was identified as Pseudomonas fragi with a unique property to produce bead-like particles (A22B) in its cell wall. The A22B component extracted from the bead-like particles of strain A22 possesses the following properties: Sudan black staining positive but water soluble; retained in dialysis tubing with a molecular mass cutoff of 8,000 Da; and no detectable amino or amino acid residues in its hydrolysates (data not shown). Both morphological and chemical analyses indicate that A22B is different from the poly(β-hydroxybutyrate) (PHB) produced by Pseudomonas fluorescens (5), capsule of Azotobacter vinelandii (4, 9), lipopolysaccharide from Escherichia coli (7), and protease liquid beads from P. fragi B25 (3). Comparative genomics and proteomics are efficient methods to explore the genetic basis for the formation of A22B, but to date, no genome sequence has been reported for the P. fragi species. Therefore, both the type strain of this species, P. fragi B25 (ATCC 4973), and the novel isolate (A22) were subjected to genome sequence analysis.

The genomic DNA was isolated and purified from strains A22 and B25 by a previously described method (11), and DNA fragments were prepared by using Illumina paired-end sample preparation kits (Illumina, San Diego, CA). Libraries were constructed and sequenced at the Beijing Genomics Institute (Shenzhen, China) by using the Illumina Solexa GA instrument. For both strains (A22 and B25), libraries containing ∼500-bp inserts were sequenced with the paired-end strategy of 90-bp reads to produce 600 Mb and 660 Mb of filtered sequences, representing 118-fold and 131-fold coverage of the genome, respectively; further, libraries with fragments of about 2,000 bp were constructed and sequenced by using the same methods to produce 300 Mb of filtered sequences. Genome annotation was performed at the NCBI Prokaryotic Genomes Automatic Annotation Pipeline (PGAAP), and the open reading frames (ORFs) and tRNA and rRNA genes were identified by Glimmer 3.02, GeneMarkS, tRNAscan-SE, RNAmmer, KEGG, and COG databases (1, 2, 6, 8, 10, 12).

The strain A22 assembly has a total of 5,074,987 nucleotides (nt) spread across 114 contigs, with an average G+C content of 58.64%, 4,631 ORFs, 61 tRNA genes, and 48 rRNA genes. The type strain B25 assembly has a total of 5,021,250 nt spread across 227 contigs, with an average G+C content of 59.37%, 4,645 ORFs, 65 tRNA genes, and 15 rRNA genes.

Comparing the contigs of strains A22 and B25 with the genome in the database, the maximum coverage was 61% and 74% with Pseudomonas fluorescens SBW25, respectively. The preliminary results indicate some notable sequence differences and specific arrangements, biosynthesis pathway construction by genome collinearity analysis, and COG function classification between strains A22 and B25. Complete genome sequences will be obtained in the future, which will also include the organization of some typical polyester biosynthesis pathways and functional genes.

Nucleotide sequence accession numbers.

The Whole Genome Shotgun project for P. fragi A22 has been deposited at DDBJ/EMBL/GenBank under accession numbers AHZY01000001 to AHZY01000114. The Whole Genome Shotgun project for P. fragi B25 has been deposited at DDBJ/EMBL/GenBank under accession numbers AHZX01000001 to AHZX01000227.