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In Streptococcus pneumoniae the competence-stimulating peptide (CSP), encoded by the comC gene, controls competence development and influences biofilm growth. We explored the influence of pherotype, defined by the two major comC allelic variants (comC1 and comC2), on biofilm development and recombination efficiency. Among isolates recovered from human infections those presenting comC1 show a higher capacity to form in vitro biofilms. The influence of pherotype on biofilm growth was confirmed by experiments with isogenic strains differing in their comC alleles. Biofilm architecture evaluated by confocal laser scanning microscopy showed that strains carrying comC1 form biofilms that are denser and thicker than those carrying the comC2 allele. Isogenic strains carrying the comC1 allele yielded more transformants than those carrying the comC2 allele in both planktonic and biofilm growth. Transformation assays with comC knockout strains show that ComD1 needs lower doses of the signaling peptide to reach the same biological outcomes. In contrast to mixed planktonic growth, within mixed biofilms inter-pherotype genetic exchange is less frequent than that occurring between bacteria of the same pherotype. Since biofilms are a major bacterial lifestyle, these observations may explain the genetic differentiation between populations with different pherotypes reported previously. Considering that biofilms have been associated with colonization our results suggest that strains carrying the comC1 allele may be more transmissible and more efficient at persisting in carriage. Both effects may help explain the higher prevalence of the comC1 allele in the pneumococcal population.

Several research fields frequently deal with the analysis of diverse classification results of the same entities. This should imply an objective detection of overlaps and divergences between the formed clusters. The congruence between classifications can be quantified by clustering agreement measures, including pairwise agreement measures. Several measures have been proposed and the importance of obtaining confidence intervals for the point estimate in the comparison of these measures has been highlighted. A broad range of methods can be used for the estimation of confidence intervals. However, evidence is lacking about what are the appropriate methods for the calculation of confidence intervals for most clustering agreement measures. Here we evaluate the resampling techniques of bootstrap and jackknife for the calculation of the confidence intervals for clustering agreement measures. Contrary to what has been shown for some statistics, simulations showed that the jackknife performs better than the bootstrap at accurately estimating confidence intervals for pairwise agreement measures, especially when the agreement between partitions is low. The coverage of the jackknife confidence interval is robust to changes in cluster number and cluster size distribution.

Streptococcus pneumoniae (pneumococcus) is able to form biofilms in vivo and previous studies propose that pneumococcal biofilms play a relevant role both in colonization and infection. Additionally, pneumococci recovered from human infections are characterized by a high prevalence of lysogenic bacteriophages (phages) residing quiescently in their host chromosome. We investigated a possible link between lysogeny and biofilm formation. Considering that extracellular DNA (eDNA) is a key factor in the biofilm matrix, we reasoned that prophage spontaneous activation with the consequent bacterial host lysis could provide a source of eDNA, enhancing pneumococcal biofilm development. Monitoring biofilm growth of lysogenic and non-lysogenic pneumococcal strains indicated that phage-infected bacteria are more proficient at forming biofilms, that is their biofilms are characterized by a higher biomass and cell viability. The presence of phage particles throughout the lysogenic strains biofilm development implicated prophage spontaneous induction in this effect. Analysis of lysogens deficient for phage lysin and the bacterial major autolysin revealed that the absence of either lytic activity impaired biofilm development and the addition of DNA restored the ability of mutant strains to form robust biofilms. These findings establish that limited phage-mediated host lysis of a fraction of the bacterial population, due to spontaneous phage induction, constitutes an important source of eDNA for the S. pneumoniae biofilm matrix and that this localized release of eDNA favors biofilm formation by the remaining bacterial population.

Very diverse research fields frequently deal with the analysis of multiple clustering results, which should imply an objective detection of overlaps and divergences between the formed groupings. The congruence between these multiple results can be quantified by clustering comparison measures such as the Wallace coefficient (W). Since the measured congruence is dependent on the particular sample taken from the population, there is variability in the estimated values relatively to those of the true population. In the present work we propose the use of a confidence interval (CI) to account for this variability when W is used. The CI analytical formula is derived assuming a Gaussian sampling distribution and recurring to the algebraic relationship between W and the Simpson's index of diversity. This relationship also allows the estimation of the expected Wallace value under the assumption of independence of classifications. We evaluated the CI performance using simulated and published microbial typing data sets. The simulations showed that the CI has the desired 95% coverage when the W is greater than 0.5. This behaviour is robust to changes in cluster number, cluster size distributions and sample size. The analysis of the published data sets demonstrated the usefulness of the new CI by objectively validating some of the previous interpretations, while showing that other conclusions lacked statistical support.