New insights into the pneumococcal fratricide: relationship to clumping and identification of a novel immunity factor.

In 1971, Tomasz and Zanati discovered that competent pneumococci have a tendency to form aggregates when pelleted by centrifugation and resuspended in 0.01 N HCl by brief vortexing. Interestingly, no clumping was observed with parallel cultures of non-competent cells treated in the same way. We set out to elucidate the mechanism behind this striking phenomenon, and were able to show that it depends on extracellular DNA that is presumably released by so-called competence-induced cell lysis. Competence-induced cell lysis, which was first described a few years ago, seems to rely on the concerted action of several murein hydrolases. Our results confirmed and extended previous findings by showing that competence-induced aggregation is abolished in a lytA-lytC double mutant, and absolutely requires CbpD and its N-terminal CHAP amidase domain. Furthermore, we discovered a novel competence stimulating peptide (CSP)-induced immunity protein, encoded by the early competence gene comM (spr1762), which protects competent pneumococci against their own lysins. Together, the murein hydrolases and the immunity protein constitutes a CSP-controlled mechanism that allows competent pneumococci to commit fratricide by killing non-competent pneumococci sharing the same ecological niche. Through such predatory behaviour, pneumococci can get access to transforming DNA and nutrients, promote the release of virulence factors, and at the same time get rid of competitors.

Release of DNA into the medium by competent Streptococcus pneumoniae: kinetics, mechanism and stability of the liberated DNA.

The release of chromosomal DNA into culture media has been reported for several naturally transformable bacterial species, but a direct link between competence development and the liberation of DNA is generally lacking. Based on the analysis of strains with mutations in competence-regulatory genes and the use of conditions favouring or preventing competence, we provide evidence that DNA release is triggered by the induction of competence in Streptococcus pneumoniae. Kinetic analyses revealed that whereas competence was maximal 20 min after addition of competence-stimulating peptide, and then decreased, the amount of liberated DNA continued to increase and reached a maximum in stationary phase, when cells are no longer competent for DNA uptake. These data are not consistent with the proposal that release of DNA by a fraction of the population is coordinated with uptake by the remainder. Moreover, we observed that an unidentified DNase was specifically induced or released in competent cultures, and that together with the major pneumococcal endonuclease, EndA, it could degrade released DNA. Nearby complete abolition of release in a mutant lacking both the major autolysin, LytA, and the autolytic lysozyme, LytC, indicated that DNA liberation occurs by LytA-LytC-dependent cell lysis. These observations suggest that competence-dependent DNA release is one facet of a more general phenomenon of sensitization to autolysis that reaches its maximum in stationary phase.

Extracellular-peptide control of competence for genetic transformation in Streptococcus pneumoniae.

Bacteria, which often are subjected to fluctuations in nutrients, temperature, radiation, pH, etc., adapt to the physico-chemical environment they live in by making the appropriate changes in their gene expression patterns. During the last decades it has become increasingly clear that bacteria, in addition, have a "social life", and that changes in gene expression can also be elicited by the presence of other bacteria. Traditionally bacteria have been viewed as solitary organisms that in general do not interact with other bacteria in a coordinated manner. Recent advances in the field of bacterial cell-to-cell communication has proved this to be a misconception, and mounting evidence now show that bacterial group behaviour is ubiquitous in nature. Competence for natural genetic transformation in Streptococcus pneumoniae, which has been studied for more than seventy years, has become a paradigm for intercellular communication and cell density dependent regulation of gene expression in Gram-positive bacteria. There has been rapid progress recently in elucidating the molecular mechanisms behind regulation of natural competence in S. pneumoniae. In this review, we describe the current status of our knowledge of natural competence in this bacterium, with particular emphasis on the early phase of competence induction.