Molecular and clinical analyses of the gene encoding the collagen-binding adhesin of Streptococcus mutans.
ABSTRACT Streptococcus mutans is a known pathogen of dental caries and its major cell surface antigens have been widely investigated. Recently, an approximately 120 kDa Cnm protein with binding properties to type I collagen was identified, and its encoding gene (cnm) cloned and sequenced. In the present study, we sequenced cnm from 47 different clinical S. mutans strains and found that the nucleotide alignment of the collagen-binding domain was well conserved. We devised a PCR method for identifying the cnm gene, examined the prevalence of cnm-positive S. mutans strains in various mother-child groups, and assessed the significance of such strains for transmission and dental caries. The detection rate of cnm-positive strains was significantly lower in strains isolated from Japanese children in the 2000s (8.0 %) as compared to those isolated in the 1980s (15.8 %) (P<0.05). Furthermore, the presence of S. mutans possessing cnm in salivary specimens collected from 55 S. mutans-positive mother-child pairs was 40 and 32.7 % in the mothers and children, respectively. The frequency of cnm-positive children whose mothers were also positive was 72 %, which was significantly higher than that of cnm-positive children with negative mothers (P<0.0001, odds ratio 17.5). In addition, clinical parameters indicating dental caries were significantly increased in children with cnm-positive S. mutans in saliva (n=13), as compared to those with cnm-negative S. mutans (n=15) and S. mutans-negative children (n=20) (P<0.01). These results indicate that cnm-positive S. mutans strains are closely correlated with dental caries, while vertical transmission in cnm-positive mother-child pairs was also demonstrated.
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ABSTRACT: Cnm, a collagen- and laminin-binding protein present in a subset of S. mutans strains, mediates binding to extracellular matrices (ECM), intracellular invasion and virulence in the Galleria mellonella model. Antibodies raised against Cnm were used to confirm expression and the cell surface localization of Cnm in the highly invasive OMZ175 strain. Sequence analysis identified two additional genes (cnaB and cbpA) encoding putative surface proteins immediately upstream of cnm. Inactivation of cnaB and cbpA in OMZ175, individually or in combination did not decrease the ability of this highly invasive and virulent strain to bind to different ECM proteins, invade human coronary artery endothelial cells (HCAEC), or to kill G. mellonella. Similarly, expression of cnaB and cbpA in the cnm(-) strain UA159 revealed that these genes did not enhance Cnm-related phenotypes. However, integration of cnm in the chromosome of UA159 significantly increased its ability to bind to collagen and laminin, invade HCAEC, and kill G. mellonella. Moreover, the presence of antibodies against Cnm nearly abolished the ability of OMZ175 to bind to collagen and laminin and invade HCAECs, and significantly protected G. mellonella against OMZ175 infection. We concluded that neither CnaB nor CbpA are necessary for the expression of Cnm-related traits. We also provided definitive evidence that Cnm is an important virulence factor and a suitable target for the development of novel preventive and therapeutic strategies to combat invasive S. mutans strains. This article is protected by copyright. All rights reserved.Molecular oral microbiology. 09/2013;
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ABSTRACT: Enterococcus faecium NRRL B-2354 is a surrogate microorganism used in place of pathogens for validation of thermal processing technologies and systems. We evaluated the safety of strain NRRL B-2354 based on its genomic and functional characteristics. The genome of E. faecium NRRL B-2354 was sequenced and found to comprise a 2,635,572 bp chromosome and a 214,319 bp mega-plasmid. A total of 2,639 coding sequences were identified, including 45 genes unique to this strain. Hierarchical clustering of the NRRL B-2354 genome with 126 other E. faecium genomes as well as pbp5 locus comparisons and Multi-Locus Sequence Typing (MLST) showed that the genotype of this strain is most similar to commensal, or community-associated, strains of this species. E. faecium NRRL B-2354 lacks antibiotic resistance genes, and both NRRL B-2354 and its clonal relative ATCC 8459 are sensitive to clinically-relevant antibiotics. This organism also lacks or contains non-functional copies of enterococcal virulence genes including acm, cyl, the ebp operon, esp, gelE, hyl, and IS16, and associated phenotypes. It does contain scm, sagA, efaA, and the pilA, although these genes were either not expressed or their roles in enterococcal virulence are not well understood. Compared with the clinical strains TX0082 and 1,231,502, E. faecium NRRL B-2354 was more resistant to acidic conditions (pH 2.4) and high temperatures (60°C) and was able to grow in 8% ethanol. These findings support the continued use of E. faecium NRRL B-2354 in thermal process validation of food products.Applied and environmental microbiology 01/2014; · 3.69 Impact Factor
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ABSTRACT: Mutans streptococci are a group of bacteria significantly contributing to tooth decay. Their genetic variability is however still not well understood. Genomes of 6 clinical S. mutans isolates of different origins, one isolate of S. sobrinus (DSM 20742) and one isolate of S. ratti (DSM 20564) were sequenced and comparatively analyzed. Genome alignment revealed a mosaic-like structure of genome arrangement. Genes related to pathogenicity are found to have high variations among the strains, whereas genes for oxidative stress resistance are well conserved, indicating the importance of this trait in the dental biofilm community. Analysis of genome-scale metabolic networks revealed significant differences in 42 pathways. A striking dissimilarity is the unique presence of two lactate oxidases in S. sobrinus DSM 20742, probably indicating an unusual capability of this strain in producing H2O2 and expanding its ecological niche. In addition, lactate oxidases may form with other enzymes a novel energetic pathway in S. sobrinus DSM 20742 that can remedy its deficiency in citrate utilization pathway.Using 67 S. mutans genomes currently available including the strains sequenced in this study, we estimates the theoretical core genome size of S. mutans, and performed modeling of S. mutans pan-genome by applying different fitting models. An "open" pan-genome was inferred. The comparative genome analyses revealed diversities in the mutans streptococci group, especially with respect to the virulence related genes and metabolic pathways. The results are helpful for better understanding the evolution and adaptive mechanisms of these oral pathogen microorganisms and for combating them.BMC Genomics 06/2013; 14(1):430. · 4.40 Impact Factor