The complete genome sequence of Lactobacillus bulgaricus reveals extensive and ongoing reductive evolution

Abstract

Lactobacillus delbrueckii ssp. bulgaricus (L. bulgaricus) is a representative of the group of lactic acid-producing bacteria, mainly known for its worldwide application in yogurt production. The genome sequence of this bacterium has been determined and shows the signs of ongoing specialization, with a substantial number of pseudogenes and incomplete metabolic pathways and relatively few regulatory functions. Several unique features of the L. bulgaricus genome support the hypothesis that the genome is in a phase of rapid evolution. (i) Exceptionally high numbers of rRNA and tRNA genes with regard to genome size may indicate that the L. bulgaricus genome has known a recent phase of important size reduction, in agreement with the observed high frequency of gene inactivation and elimination; (ii) a much higher GC content at codon position 3 than expected on the basis of the overall GC content suggests that the composition of the genome is evolving toward a higher GC content; and (iii) the presence of a 47.5-kbp inverted repeat in the replication termination region, an extremely rare feature in bacterial genomes, may be interpreted as a transient stage in genome evolution. The results indicate the adaptation of L. bulgaricus from a plant-associated habitat to the stable protein and lactose-rich milk environment through the loss of superfluous functions and protocooperation with Streptococcus thermophilus.

Fig. 1.

Relationship between GC content at position 3 of coding sequences (GC3) and genomic GC content in 232 eubacterial genomes. The L. bulgaricus value was calculated excluding pseudogenes. The L. bulgaricus data point is circled.

Fig. 2.

Genome atlas of the L. bulgaricus genome. The seven circles (outer to inner) show the following. Circle 1, pseudogenes on positive (red) or negative (blue) strand. Circle 2, IS elements (transposases or ISL4-related hypothetical genes). Elements with fewer than four copies are represented in gray, and elements with more than five copies are represented by separate colors of red (ISL7), purple (ISL4), blue (ISL5), and green (ISL4–5). (See also Table 2.) Circle 3, CDS (excluding pseudogenes and transposases) on positive (red) or negative (blue) strand. Circle 4, rRNA (red) and tRNA (green) genes. Circle 5, [(G-C)/window size (2000)], from less than −0.1 (cyan) to more than +0.1 (red). Circle 6, [(A+T)/window size (500)], from <0.3 (cyan) to >0.7 (red). Circle 7, position on the genome. The genome atlas was constructed by using genewiz software (53).

Fig. 3.

rRNA and tRNA genes as a function of genome size. (A) Relationship between the number of 16S rRNA genes and genome size in 54 firmicutes genomes. (B) Relationship between the number of tRNA genes and genome size in 54 firmicutes genomes. L. bulgaricus (1.9 Mbp, 9 16S rRNA, and 95 tRNA) data points are circled. For comparison: L. acidophilus (2.0 Mbp, 4 16S rRNA, and 61 tRNA), L. johnsonii (2.0 Mbp, 6 16S rRNA, and 79 tRNA), and L. plantarum (3.3 Mbp, 5 16S rRNA, and 70 tRNA).

Fig. 4.

Synteny between L. bulgaricus and L. acidophilus genomes. x axis, position on L. bulgaricus genome (Mbp); y axis, position on L. acidophilus genome (Mbp). 0, replication origin. Colors indicate protein similarity by blast score ratio (25) according to the scale on the right.