Comparative genomics of Bifidobacterium animalis subsp. lactis reveals a strict monophyletic bifidobacterial taxon

Abstract

Strains of Bifidobacterium animalis subsp. lactis are extensively exploited by the food industry as health-promoting bacteria, although the genetic variability of members belonging to this taxon has so far not received much scientific attention. In this article, we describe the complete genetic makeup of the B. animalis subsp. lactis Bl12 genome and discuss the genetic relatedness of this strain with other sequenced strains belonging to this taxon. Moreover, a detailed comparative genomic analysis of B. animalis subsp. lactis genomes was performed, which revealed a closely related and isogenic nature of all currently available B. animalis subsp. lactis strains, thus strongly suggesting a closed pan-genome structure of this bacterial group.

Fig 1

Comparative genomic analysis of B. animalis subsp. lactis Bl12 with other fully sequenced B. animalis subsp. lactis strains. Panel a represents a circular genome atlas of B. animalis subsp. lactis Bl12 (circle 1) with mapped orthologues (defined as reciprocal best BLASTp hits with more than 30% identity over at least 80% of both protein lengths) in nine publicly available B. animalis subsp. lactis genomes (circles 2 through 10). Circle 11 illustrates B. animalis subsp. lactis DSM10140 G+C% deviation, followed by circle 12, which highlights B. animalis subsp. lactis DSM10140 GC skew (G-C/G+C). Panel b shows a graphical representation of the COG families of B. animalis subsp. lactis Bl12 and other Bifidobacterium species. Each COG family is identified by a one-letter abbreviation: A, RNA processing and modification; B, chromatin structure and dynamics; C, energy production and conversion; D, cell cycle control and mitosis; E, amino acid metabolism and transport; F, nucleotide metabolism and transport; G, carbohydrate metabolism and transport; H, coenzyme metabolism; I, lipid metabolism; J, translation; K, transcription; L, replication and repair; M, cell wall/membrane/envelope biogenesis; N, cell motility; O, posttranslational modification, protein turnover, and chaperone functions; P, inorganic ion transport and metabolism; Q, secondary structure; T, signal transduction; U, intracellular trafficking and secretion; Y, nuclear structure; V, defense mechanisms; Z, cytoskeleton; R, general functional prediction only; S, function unknown.

Fig 2

(a) Comparison of the miaA locus in B. animalis subsp. lactis Bl12 with corresponding loci in various other B. animalis subsp. lactis strains. Each arrow indicates an ORF. The length of the arrow is proportional to the length of the predicted ORF. Corresponding genes are marked with the same color. The putative function of the protein is indicated above each arrow. The percent amino acid identity is indicated. (b) Nucleotide alignment of the portion of the miaA encompassing the identified SNP. (c) Amino acid alignment of the portion of the Mia protein around the identified nonsynonymous mutation.

Fig 3

Genomic diversity of the Bifidobacterium animalis subsp. lactis species. Panel a displays a Venn diagram of homologues shared between type strain B. animalis subsp. lactis Bl12 and other fully sequenced bifidobacterial species. Panel b shows a Venn diagram representation of shared homologues between B. animalis subsp. lactis Bl12 and the nine publicly available B. animalis subsp. lactis genomes. Panel c depicts a phylogenetic supertree based on the sequences of identified core proteins shared by the analyzed Bifidobacterium genomes.

Fig 4

Dot plot comparison based on genomic sequence alignments of B. animalis subsp. lactis Bl12, the nine publicly available B. animalis subsp. lactis strains, and B. animalis subsp. animalis ATCC 25527.

Fig 5

Restriction profiles of B. animalis subsp. lactis strains by in silico prediction. The optical map was generated by in silico digestion with NotI and visualized through Geneious software. The strains analyzed are Bl12 (lane A), BB12 (lane B), BLC1 (lane C), CNCM I-2494 (lane D), DSM10140 (lane E), AD011 (lane F), and B. animalis subsp. animalis ATCC 25527 (lane G); regions of variability are highlighted.

Fig 6

Putative SNPs in B. animalis subsp. lactis Bl12 and the nine publicly available B. animalis subsp. lactis genomes. Panel a shows a heat map of the 47 SNPs listed by Barangou et al. (14) mapped on B. animalis subsp. lactis Bl12 and the nine publicly available B. animalis subsp. lactis genomes. Each color represents a base as indicated. The dendrogram shows genome clustering produced by hierarchical clustering based on the heat map data. Panel b depicts a phylogenetic supertree based on B. animalis subsp. lactis Bl12 and the nine publicly available B. animalis subsp. lactis nucleotide sequences corresponding to 20-bp regions spanning the verified SNPs listed by Barrangou et al. (14). The different B. animalis subsp. lactis groups are highlighted.

Fig 7

The distribution of the number of total genes (a) and core genes (b) found upon sequential addition of n genomes. In panel a, power law fit to the pan-genome size is shown as solid curve. In panel b, an exponential regression to core genome data is shown as a solid curve.