Complete Genome Sequence of Listeria seeligeri, a Nonpathogenic Member of the Genus Listeria

Abstract

We report the complete and annotated genome sequence of the nonpathogenic Listeria seeligeri SLCC3954 serovar 1/2b type strain harboring the smallest completely sequenced genome of the genus Listeria.

Listeria seeligeri is one of seven species of the genus Listeria, a group of Gram-positive, motile, facultative anaerobic, low-GC-content, nonsporulating rods (3, 10, 12). To obtain a better understanding of the evolution of this nonpathogenic Listeria species, the type strain, SLCC 3954 (serovar 1/2b), a soil isolate from Germany (11), was sequenced using Sanger technology. Two small (1.5- to 2.5-kb) insert plasmid libraries were constructed with the TOPO Shotgun Subcloning Kit (Invitrogen) as described previously (9). Additionally, a small insert plasmid library (∼1.0 to 1.5 kb) and a medium insert plasmid library (∼5 kb) were constructed in the pUC19 cloning vector (New England Biolabs) by Qiagen (Hilden, Germany). A fosmid library harboring fragments of around 40 kb was created using the CopyControl Fosmid Library Production Kit (Epicentre) as published before (9).

Sequencing was performed by Agowa (Berlin, Germany), Qiagen (Hilden, Germany), and the Max-Planck-Institut (Köln, Germany) by using ABI Big Dye Terminator technology. A total of 30,961 and 1,305 high-quality reads from the shotgun, and fosmid libraries were used to generate a draft assembly with an overall coverage of ∼7-fold by using the Phred/Phrap/Consed assembly package (4, 5, 7). Contigs were linked by primer walking on shotgun clones and fosmids as well as by PCR gap closure followed by sequencing of the PCR product. Genome annotation was performed as described previously (9).

The genome of L. seeligeri consists of a circular chromosome of 2,797,636 bp and hence is slightly smaller than those of previously sequenced listerial strains (6, 9). Functional classification of genes obtained by mapping against clusters of orthologous groups (14) was predicted by Augur (1). Comparative analysis of these clusters indicates gene loss in categories such as amino acid/carbohydrate transport and metabolism as well as in transcription, thus confirming a general trend toward genome shrinkage. L. seeligeri harbors no plasmid and carries only a single copy of a prophage and no transposon in its genome. The mean G+C content of the L. seeligeri genome is 37.4%, which is close to the average value of all known Listeria strains (6, 9). G/C skew analysis revealed a bidirectional replication mechanism, and the origin of replication (oriC) is located close to the dnaA gene, which is positioned diametrically opposite to the replication terminus. We identified six 16S-23S-5S rRNA operons, all of which are located on the leading strand, two on the right and four on the left replichore. Additionally, a total of 67 tRNA genes were detected. We used MAVID (2) for phylogenetic analysis of the genomes of L. monocytogenes, L. innocua, L. welshimeri, and L. seeligeri and conclude that the “phylogenomic” relationship corresponds exactly to phylogenetic analysis based on either 16S rRNA genes (8, 15) or other additional specific marker genes (8, 13).

The genome sequence of L. seeligeri is the fourth species of genus Listeria to be reported.

Nucleotide sequence accession number.

The genome sequence of L. seeligeri serovar 1/2b (SLCC3954) reported here has been deposited in the EMBL database under accession number FN557490.