Abstract
Streptococcus mutans is a key pathogenic bacterium in the oral cavity and a primary contributor to dental caries. The S. mutans Cid/Lrg system likely contributes to tolerating stresses encountered in this environment as cid and/or lrg mutants exhibit altered oxidative stress sensitivity, genetic competence, and biofilm phenotypes. It was recently noted that the cidB mutant had two stable colony morphologies: a “rough” phenotype (similar to wild type) and a “smooth” phenotype. In our previously published work, the cidB rough mutant exhibited increased sensitivity to oxidative stress, and RNAseq identified widespread transcriptomic changes in central carbon metabolism and oxidative stress response genes. In this current report, we conducted Illumina‐based genome resequencing of wild type, cidB rough, and cidB smooth mutants and compared their resistance to oxidative and acid stress, biofilm formation, and competence phenotypes. Both cidB mutants exhibited comparable aerobic growth inhibition on agar plates, during planktonic growth, and in the presence of 1 mM hydrogen peroxide. The cidB smooth mutant displayed a significant competence defect in BHI, which was rescuable by synthetic CSP. Both cidB mutants also displayed reduced XIP‐mediated competence, although this reduction was more pronounced in the cidB smooth mutant. Anaerobic biofilms of the cidB smooth mutant displayed increased propidium iodide staining, but corresponding biofilm CFU data suggest this phenotype is due to cell damage and not increased cell death. The cidB rough anaerobic biofilms showed altered structure relative to wild type (reduced biomass and average thickness) which correlated with decreased CFU counts. Sequencing data revealed that the cidB smooth mutant has a unique “loss of read coverage” of ~78 kb of DNA, corresponding to the genomic island TnSMU2 and genes flanking its 3′ end. It is therefore likely that the unique biofilm and competence phenotypes of the cidB smooth mutant are related to its genomic changes in this region.
Highlights
Dental caries is considered a microbial shift disease within the human oral cavity (Marsh, 2006; Mira, Simon‐Soro, & Curtis, 2017; Sanz et al, 2017), Streptococcus mutans is highly associated with caries formation (Garcia et al, 2017; Loesche, 1986)
We explore the physiological differences between the cidB rough and cidB smooth mutants in relation to the presence or loss of the TnSMU2 region, respectively
Our characterization of each cidB mutant variant has both confirmed the role of cidB in S. mu‐ tans physiology and reinforces a possible link between this gene and TnSMU2
Summary
Dental caries is considered a microbial shift disease within the human oral cavity (Marsh, 2006; Mira, Simon‐Soro, & Curtis, 2017; Sanz et al, 2017), Streptococcus mutans is highly associated with caries formation (Garcia et al, 2017; Loesche, 1986). The presence of eDNA is import‐ ant for S. mutans biofilm formation and regulated cell death (Perry, Cvitkovitch, & Lévesque, 2009) Oral streptococci such as S. mutans can utilize eDNA for genetic exchange via natural competence. Opp‐mediated uptake is blocked by peptide‐rich media, but ComS has been demonstrated to participate in competence signaling without leav‐ ing the cell (Underhill et al, 2018) While both CSP and XIP induce competence in alternative ways, addition of synthetic forms of ei‐ ther peptide has been shown to stimulate a comX response (Son, Ahn, Guo, Burne, & Hagen, 2012). The S. mutans cidAB transcriptional unit encodes two proteins (CidA and CidB) which have been shown to affect a multitude of virulence traits including biofilm development and oxidative stress response (Ahn, Rice, Oleas, Bayles, & Burne, 2010). This work addresses the physiological role(s) that cidB and TnSMU2 play in S. mutans UA159, and reinforces a previously established regulatory connection (Ahn & Rice, 2016) between these two loci
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