LuxS-Mediated Signaling inStreptococcus mutansIs Involved in Regulation of Acid and Oxidative Stress Tolerance and Biofilm Formation

Abstract

LuxS-mediated quorum sensing has recently been shown to regulate important physiologic functions and virulence in a variety of bacteria. In this study, the role of luxS of Streptococcus mutans in the regulation of traits crucial to pathogenesis was investigated. Reporter gene fusions showed that inactivation of luxS resulted in a down-regulation of fructanase, a demonstrated virulence determinant, by more than 50%. The LuxS-deficient strain (TW26) showed increased sensitivity to acid killing but could still undergo acid adaptation. Northern hybridization revealed that the expression of RecA, SmnA (AP endonuclease), and Nth (endonuclease) were down-regulated in TW26, especially in early-exponential-phase cells. Other down-regulated genes included ffh (a signal recognition particle subunit) and brpA (biofilm regulatory protein A). Interestingly, the luxS mutant showed an increase in survival rate in the presence of hydrogen peroxide (58.8 mM). The luxS mutant formed less biofilm on hydroxylapatite disks, especially when grown in biofilm medium with sucrose, and the mutant biofilms appeared loose and hive-like, whereas the biofilms of the wild type were smooth and confluent. The mutant phenotypes were complemented by exposure to supernatants from wild-type cultures. Two loci, smu486 and smu487, were identified and predicted to encode a histidine kinase and a response regulator. The phenotypes of the smu486 smu487 mutant were, in almost all cases, similar to those of the luxS mutant, although our results suggest that this is not due to AI-2 signal transduction via Smu486 and Smu487. This study demonstrates that luxS-dependent signaling plays critical roles in modulating key virulence properties of S. mutans.Streptococcus mutans is recognized as the principal etiological agent of dental caries, one of the most prevalent human infectious diseases. The abilities to adhere to and to form biofilms on the tooth surface, to metabolize carbohydrates, and to survive low pH and other environmental insults are believed to be critical to the cariogenicity of this human pathogen (4). Known for its high degree of acid tolerance (aciduricity) and its high capacity to produce acids (acidogenicity), S. mutans has evolved strategies to survive, grow, and carry out glycolysis at extracellular pH values well below those at which demineralization of tooth enamel begins to take place. As an obligate biofilm-forming bacterium (4), S. mutans lives almost exclusively on the tooth surface at high cell density in a high-diversity ecosystem better known as dental plaque. The structure and composition of the plaque are known to be strongly influenced by such factors as the source and availability of nutrients, the pH in the oral cavity, and the ability of its constituents to adapt to fluctuations in environmental conditions (4, 38).In a process termed quorum sensing (QS), which was first discovered in the marine bacterium Vibrio fischeri, bacterial populations coordinate gene expression and, thus, the behavior of the community. It is now appare...

Deficiency of BrpB causes major defects in cell division, stress responses and biofilm formation by Streptococcus mutans

“…mutans strains were allowed to adapt and grow in BHI adjusted to pH 5.0 for 1 h before being subjected to acid killing (Wen & Burne, 2004). The results showed that like wild-type UA159, the BrpB-deficient mutant, JB409, also significantly increased its survival rate after 1 h incubation in BHI adjusted to pH 5.0 (Fig.…”

“…For field emission-scanning electron microscopy (FE-SEM) analysis, Strep. mutans overnight cultures were grown in BMG, BMGS or BMS on HA discs that were placed horizontally in 24-well plates as described elsewhere (Bitoun et al, 2011;Wen & Burne, 2004). After 24 h of growth, the HA discs were removed and washed twice with PBS, pH 7.4, before overnight fixation with 2.5 % glutaraldehyde (Polysciences) at 4 uC.…”

“…To construct the BrpB-deficient mutant, a PCR-ligation-mutagenesis strategy was used as described elsewhere Wen & Burne, 2004). Briefly, the 59 and 39 regions flanking brpB were amplified by PCR using Phusion high-fidelity DNA polymerase (New England Biolabs) with the gene-specific primers shown in Table 2.…”

“…The impact of brpB and brpBA deficiencies on the ability of Strep. mutans to withstand acid and hydrogen peroxide stress was determined by using acid killing and hydrogen peroxide challenge assays as described elsewhere (Wen & Burne, 2004). In brief, planktonic cultures were grown in BHI until early exponential phase (OD 600 0.3) and biofilms cells were grown in BMGS on sterile glass slides in 50 ml Falcon tubes for 24 h (Wen et al, 2010).…”

Mutation of luxS affects motility and infectivity of Helicobacter pylori in gastric mucosa of a Mongolian gerbil model

“…The genomes of many bacterial species, notably Escherichia coli, Salmonella enterica serovar Typhimurium, Shigella flexneri, Proteus mirabilis, Vibrio cholerae, Vibrio vulnificus, Campylobacter jejuni, Porphyromonas gingivalis, Bacillus subtilis, Streptococcus pyogenes, Streptococcus mutans, Clostridium perfringens and Clostridium difficile, include luxS homologues. In several of these, luxS-related AI-2 signals are involved in bacterial characteristics such as biofilm formation (Balestrino et al, 2005;Blehert et al, 2003;Fong et al, 2001;Wen & Burne 2004), flagella and motility (Jeon et al, 2003;Schneider et al, 2002;Stroeher et al, 2003), type III secretion systems (Sperandio et al, 1999), toxin production (Ohtani et al, 2002) and virulence (Lyon et al, 2001; Parsonnet et al, 1991;Stroeher et al, 2003).Helicobacter pylori has been identified as the aetiological agent of chronic active gastritis, peptic ulcer disease (Blaser, 1992;Graham, 1989), gastric adenocarcinoma (Parsonnet et al, 1991) and mucosal-associated lymphoid tissue (MALT) lymphoma (Wotherspoon et al, 1993). H. pylori has flagellar motility and is able to move through the Matsumoto et al, 1997;Yokota et al, 1991).…”