Genome Sequence of Lactobacillus amylovorus GRL1112

Abstract

Lactobacillus amylovorus is a common member of the normal gastrointestinal tract (GIT) microbiota in pigs. Here, we report the genome sequence of L. amylovorus GRL1112, a porcine feces isolate displaying strong adherence to the pig intestinal epithelial cells. The strain is of interest, as it is a potential probiotic bacterium.

Recently, a new abundant Lactobacillus species was identified in the intestine of piglets and found to possess probiotic properties, like antimicrobial activity against enteric pathogens in both in vitro and in vivo trials (9, 10, 14). While first classified as Lactobacillus sobrius, subsequent taxonomic studies revealed it to belong to the Lactobacillus amylovorus group that is abundant among Lactobacillus species originating from porcine intestine (7). L. amylovorus is an anaerobic, Gram-positive, nonmotile, non-spore-forming, and rod-shaped bacterium that grows in short chains. It shows features typical of homofermentative Lactobacillus species, like production of large quantities of lactic acid and small amounts of acetic acid but no gas from glucose. It requires a number of vitamins for growth. L. amylovorus does not exhibit oxidase-, catalase-, or nitrate-reducing activities (13). According to taxonomic studies, L. amylovorus is a member of the L. acidophilus group. Regarding the cell surface structures of lactobacilli, L. amylovorus has also been suggested to be one of the dominant S-layer-carrying Lactobacillus species in pigs (8).

As the outermost paracrystalline structures, S-layers are considered to protect the cell from hostile environmental agents and to aid in maintaining cellular integrity (2). Several Lactobacillus S-layer proteins (Slp), like those from L. brevis, L. crispatus, and L. acidophilus, have also been shown to confer adherence to host epithelial cells and extracellular matrix components (4, 6, 15).

L. amylovorus strain GRL1112 was originally isolated from porcine feces and found to be an abundant colonizer of the gastrointestinal tract (GIT) in pigs (11). Being an Slp carrier, GRL1112 also displays a strong adherence to the pig intestinal epithelial cells and has been further studied for its potential probiotic properties (11).

The GRL1112 genome was sequenced (14-times coverage) using a 454 pyrosequencer with GS FLX chemistry (Roche). The contig order was confirmed, and gaps were filled by sequencing PCR fragments from the genomic DNA template using ABI 3730 and BigDye chemistry (Applied Biosystems). Genomic data were processed using the Staden Package (16) and gsAssembler (Roche). Coding sequences (CDSs) were predicted using a combination of GeneMark and Glimmer3 (5, 12) followed by manual curation of the start sites. The remaining intergenic regions were reanalyzed for missed CDSs by using BlastX (1). Annotation transfer was performed based on a BlastP search, followed by PGAAP analysis (http://www.ncbi.nlm.nih.gov/genomes/static/Annotation_pipeline_procedures.txt) and manual verification. Orthologous groups between the different lactobacillar proteomes were identified using EDGAR (3).

The genome of L. amylovorus GRL1112 contains a single circular chromosome of 2.07 million base pairs (Mbp) and two circular plasmids of 25 kbp and 33 kbp (the plasmid sequences are not included in this article). The overall G+C content of the chromosome is 38.2%, whereas two plasmids have G+C contents of 36.3% and 31.8%. The chromosome contains 2,054 CDSs, 60 tRNA genes, and 4 rRNA operons. Out of the predicted CDSs, a putative function was assigned to 1,441 (70.2%), whereas 613 (29.8%) of the CDSs were annotated as hypothetical proteins, and of those 317 were novel. Based on the orthologous grouping, about 470 (22.8%) of the CDSs encoded by GRL1112 have no detectable homologs in any of the complete Lactobacillus genomes published to date.

Nucleotide sequence accession number.

The genome sequence of L. amylovorus GRL1112 was deposited in GenBank under accession number CP002338.