The lactic acid bacterium Streptococcus thermophilus is widely used for the manufacture of yogurt and cheese. This dairy species of major economic importance is phylogenetically close to pathogenic streptococci, raising the possibility that it has a potential for virulence. Here we report the genome sequences of two yogurt strains of S. thermophilus . We found a striking level of gene decay (10% pseudogenes) in both microorganisms. Many genes involved in carbon utilization are nonfunctional, in line with the paucity of carbon sources in milk. Notably, most streptococcal virulence-related genes that are not involved in basic cellular processes are either inactivated or absent in the dairy streptococcus. Adaptation to the constant milk environment appears to have resulted in the stabilization of the genome structure. We conclude that S. thermophilus has evolved mainly through loss-of-function events that remarkably mirror the environment of the dairy niche resulting in a severely diminished pathogenic potential. Supplementary information The online version of this article (doi:10.1038/nbt1034) contains supplementary material, which is available to authorized users.

Streptococcus thermophilus is a major dairy starter used for the manufacture of yoghurt and cheese. The access to three genome sequences, comparative genomics and multilocus sequencing analyses suggests that this species recently emerged and is still undergoing a process of regressive evolution towards a specialised bacterium for growth in milk. Notably, S. thermophilus has maintained a well-developed nitrogen metabolism whereas its sugar catabolism has been subjected to a high level of degeneracy due to a paucity of carbon sources in milk. Furthermore, while pathogenic streptococci are recognised for a high capacity to expose proteins at their cell surface in order to achieve cell adhesion or to escape the host immune system, S. thermophilus has nearly lost this unique feature as well as many virulence-related functions. Although gene decay is obvious in S. thermophilus genome evolution, numerous small genomic islands, which were probably acquired by horizontal gene transfer, comprise important industrial phenotypic traits such as polysaccharide biosynthesis, bacteriocin production, restriction-modification systems or oxygen tolerance.

In addition to the previously characterized pyruvate oxidase PoxB, the Lactobacillus plantarum genome encodes four predicted pyruvate oxidases (PoxC, PoxD, PoxE, and PoxF). Each pyruvate oxidase gene was individually inactivated, and only the knockout of poxF resulted in a decrease in pyruvate oxidase activity under the tested conditions. We show here that L. plantarum has two major pyruvate oxidases: PoxB and PoxF. Both are involved in lactate-to-acetate conversion in the early stationary phase of aerobic growth and are regulated by carbon catabolite repression. A strain devoid of pyruvate oxidase activity was constructed by knocking out the poxB and poxF genes. In this mutant, acetate production was strongly affected, with lactate remaining the major end product of either glucose or maltose fermentation. Notably, survival during the stationary phase appeared to be dramatically improved in the poxB poxF double mutant.Acetate is the major fermentation end product of the lactic acid bacterium Lactobacillus plantarum when cultivated under aerobic conditions and sugar limitation. It is produced at the expense of lactate as glucose becomes depleted and cells enter the stationary phase of growth. The pathway for lactate-toacetate conversion under these conditions has been shown to involve three enzymatic steps (2,6,12,20): oxidation of lactate to pyruvate by the NAD-dependent D-and L-lactate dehydrogenases (LDH), oxidative decarboxylation of pyruvate to acetyl-phosphate (acetylϳP) by pyruvate oxidase (POX), and dephosphorylation of acetylϳP to acetate by acetate kinase (ACK). This last step produces ATP, which is believed to provide the cells with the additional energy needed for survival in the stationary phase. Acetate itself could also be involved in increased survival by maintaining the pH homeostasis (12,20). Concerning applications, the maintenance of a high viability in the stationary phase under aerobic conditions could be relevant in the development of long-shelf-life probiotic dairy products containing L. plantarum (14,29). Besides its implication in cell survival, acetate is also an important flavor compound of fermented products (e.g., sourdoughs) in which L. plantarum plays a major role (4, 5). Therefore, a better understanding of the pathways involved in acetate production in this species could contribute to the improvement of fermentation processes and products.Previously, it has been established that the oxidative decarboxylation of pyruvate catalyzed by POX is a key step in the lactate-to-acetate conversion pathway (12,27). A null mutant for the gene encoding PoxB, the major POX of L. plantarum, shows a decrease in acetate production up to 80% compared to the parent strain, depending on the growth conditions (12).This LDH-POX-ACK pathway is under control of two environmental factors: sugar and oxygen availability. Regulation takes place essentially at the level of POX activity, which is induced by oxygen or hydrogen peroxide and repressed by glucose (12,19,20,27). In the presence of excess glucose, P...

Streptococcus thermophilus is a major dairy starter used for the manufacture of yoghurt and cheese. The access to three genome sequences, comparative genomics and multilocus sequencing analyses suggests that this species recently emerged and is still undergoing a process of regressive evolution towards a specialised bacterium for growth in milk. Notably, S. thermophilus has maintained a well-developed nitrogen metabolism whereas its sugar catabolism has been subjected to a high level of degeneracy due to a paucity of carbon sources in milk. Furthermore, while pathogenic streptococci are recognised for a high capacity to expose proteins at their cell surface in order to achieve cell adhesion or to escape the host immune system, S. thermophilus has nearly lost this unique feature as well as many virulence-related functions. Although gene decay is obvious in S. thermophilus genome evolution, numerous small genomic islands, which were probably acquired by horizontal gene transfer, comprise important industrial phenotypic traits such as polysaccharide biosynthesis, bacteriocin production, restriction-modification systems or oxygen tolerance.

No abstract