Genome sequence of Lactobacillus pentosus KCA1: vaginal isolate from a healthy premenopausal woman

Abstract

The vaginal microbiota, in particular Lactobacillus species, play an important role in female health through modulation of immunity, countering pathogens and maintaining a pH below 4.7. We report the isolation and genome sequence of Lactobacillus pentosus strain KCA1 (formally known as L. plantarum) from the vagina of a healthy Nigerian woman. The genome was sequenced using Illumina GA II technology. The resulting 16,920,226 paired-end reads were assembled with the Velvet tool. Contigs were annotated using the RAST server, and manually curated. A comparative analysis with the available genomes of L. pentosus IG1 and L. plantarum WCFS1 showed that over 15% of the predicted functional activities are found only in this strain. The strain has a chromosome sequence of 3,418,159 bp with a G+C content of 46.4%, and is devoid of plasmids. Novel gene clusters or variants of known genes relative to the reference genomes were found. In particular, the strain has loci encoding additional putative mannose phosphotransferase systems. Clusters of genes include those for utilization of hydantoin, isopropylmalate, malonate, rhamnosides, and genes for assimilation of polyglycans, suggesting the metabolic versatility of L. pentosus KCA1. Loci encoding putative phage defense systems were also found including clustered regularly interspaced short palindromic repeats (CRISPRs), abortive infection (Abi) systems and toxin-antitoxin systems (TA). A putative cluster of genes for biosynthesis of a cyclic bacteriocin precursor, here designated as pentocin KCA1 (penA) were identified. These findings add crucial information for understanding the genomic and geographic diversity of vaginal lactobacilli.

Figure 1. Genome atlas view of the scaffolded contigs of L. pentosus KCA1.

From the outer circle inward: The first ring shows the entire chromosome. The second ring shows the location of the 83 contigs based on L. plantarum WCFS1 genome order/orientation as template. The black arrow-heads indicates the position of some of the genes of interest located in the corresponding contigs described in the text with the locus tag in bracket. The fourth ring shows the local %GC plot and the innermost circle shows the GC-skew with sharp changes occurring at the origin and terminus of replication. The Atlas was constructed using DNA plotter.

Figure 2. Whole genome alignment of L. pentosus KCA1 with L. pentosus IG1.

Constructed with the ACT tool. Red lines indicate orthologous genes in the same orientation. Blue lines indicate orthologous genes in reverse orientation. The large inverted region in L. pentosus IG1 is indicated.

Figure 3. Phylogenetic tree of Lactobacillus species showing the position of L. pentosus KCA1 based on the 16S rRNA gene sequences.

The numbers at the end of each strain indicates the accession number. Sequences were aligned with MUSCLE , and unreliable positions were curated using Gblocks . A maximum likelihood tree was generated by PhyML using the GTR substitution model and allowing 4 rate substitution categories. Confidence values for the branching order were generated by bootstrapping (based on 100 replications). The number at the nodes indicates the bootstrap values. The scale bar indicates 1 nucleotide substitution per 100 nucleotides.

Figure 4. Phylogenetic tree of Lactobacillus species showing the position of L. pentosus KCA1 based on the sequences of the housekeeping gene recA, relative to other plantarum and pentosus strains.

The numbers at the end of each strain indicates the accession number. Sequences were aligned with MUSCLE , and unreliable positions were curated using Gblocks . A maximum likelihood tree was generated by PhyML using the GTR substitution model and allowing 4 rate substitution categories. Confidence values (%) for the branching order were generated by bootstrapping (based on 100 replications). The number at the nodes indicated the bootstrap values. The scale bar indicates 1 nucloetide substitution per 100 nucleotides.

Figure 5. Class V bacteriocin biosynthesis gene cassettes and their organization in three Lactobacillus species including L. pentosus KCA1.

PenA represents the pentocin KCA1 precursor. Adapted from Kawai et al, 2009, AEM 75∶1324–1330.