Simultaneous Detection of Streptococcus pneumoniae, S. mitis, and S. oralis by a Novel Multiplex PCR Assay Targeting the
            <i>gyrB</i>
            Gene

Kim, Wonyong; Park, Hee Kuk; Hwang, Woo-Jin; Shin, Hyoung-Shik

doi:10.1128/jcm.02920-12

Cited by 14 publications

(8 citation statements)

References 49 publications

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“…A freely available software from manufacturers, nSolver, was used for quality assessment of the data, and normalization. The RNA counts were normalized against the geometric mean of gyrB and metG [ 65 ], [ 66 ]. The 16S rRNA gene is not optimal for normalization in the NanoString, as the high abundance of this transcript packs the field of view.…”

Section: Methodsmentioning

confidence: 99%

Function of BriC peptide in the pneumococcal competence and virulence portfolio

et al. 2018

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Streptococcus pneumoniae (pneumococcus) is an opportunistic pathogen that causes otitis media, sinusitis, pneumonia, meningitis and sepsis. The progression to this pathogenic lifestyle is preceded by asymptomatic colonization of the nasopharynx. This colonization is associated with biofilm formation; the competence pathway influences the structure and stability of biofilms. However, the molecules that link the competence pathway to biofilm formation are unknown. Here, we describe a new competence-induced gene, called briC, and demonstrate that its product promotes biofilm development and stimulates colonization in a murine model. We show that expression of briC is induced by the master regulator of competence, ComE. Whereas briC does not substantially influence early biofilm development on abiotic surfaces, it significantly impacts later stages of biofilm development. Specifically, briC expression leads to increases in biofilm biomass and thickness at 72h. Consistent with the role of biofilms in colonization, briC promotes nasopharyngeal colonization in the murine model. The function of BriC appears to be conserved across pneumococci, as comparative genomics reveal that briC is widespread across isolates. Surprisingly, many isolates, including strains from clinically important PMEN1 and PMEN14 lineages, which are widely associated with colonization, encode a long briC promoter. This long form captures an instance of genomic plasticity and functions as a competence-independent expression enhancer that may serve as a precocious point of entry into this otherwise competence-regulated pathway. Moreover, overexpression of briC by the long promoter fully rescues the comE-deletion induced biofilm defect in vitro, and partially in vivo. These findings indicate that BriC may bypass the influence of competence in biofilm development and that such a pathway may be active in a subset of pneumococcal lineages. In conclusion, BriC is a part of the complex molecular network that connects signaling of the competence pathway to biofilm development and colonization.

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Section: Methodsmentioning

confidence: 99%

Function of BriC peptide in the pneumococcal competence and virulence portfolio

et al. 2018

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“…RNA was hybridized onto the nCounter chip according to the manufacturer recommendations. RNA counts were normalized to the geometric mean of the housekeeping genes gyrB and metG (Carvalho et al ., ; Kim et al ., ) using manufacturer's software, nSolver. The in vitro and in vivo levels were compared using Student's t ‐test in the GraphPad Prism 6 tool and the heat map was generated using nSolver.…”

Section: Methodsmentioning

confidence: 99%

A novel streptococcal cell–cell communication peptide promotes pneumococcal virulence and biofilm formation

Cuevas

Eutsey

Kadam

et al. 2017

Molecular Microbiology

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Streptococcus pneumoniae (pneumococcus) is a major human pathogen. It is a common colonizer of the human respiratory track, where it utilizes cell-cell communication systems to coordinate population-level behaviors. We reasoned that secreted peptides that are highly expressed during infection are pivotal for virulence. Thus, we used in silico pattern searches to define a pneumococcal secretome, and analyzed the transcriptome of the clinically important PMEN1 lineage to identify which peptide-encoding genes are highly expressed in vivo. In this study, we characterized virulence peptide 1 (vp1), a highly expressed Gly-Gly peptide-encoding gene in chinchilla middle ear effusions. The vp1 gene is widely distributed across pneumococcus as well as encoded in related species. Studies in the chinchilla model of middle ear infection demonstrated that VP1 is a virulence determinant. The vp1 gene is positively regulated by a transcription factor from the Rgg family and its cognate SHP (short hydrophobic peptide). In vitro data indicated that VP1 promotes increased thickness and biomass for biofilms grown on chinchilla middle ear epithelial cells. Further, the wild-type biofilm is restored with the exogenous addition of synthetic VP1. We conclude that VP1 is a novel streptococcal regulatory peptide that controls biofilm development and pneumococcal pathogenesis.

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“…Importantly, gyrB gene sequences are predicted to have faster rates of molecular evolution than the rate for 16S rRNA gene sequences (Yamamoto & Harayama 1995). Therefore, it has been reported as a good phylogenetic marker for identifying some members of Pseudomonas (Yamamoto & Harayama 1998), Acinetobacter (Yamamoto, Bouvet & Harayama 1999), Mycobacterium (Kasai, Ezaki & Harayama 2000), Salmonella, Shigella and Escherichia coli (Fukushima, Kakinuma & Kawaguchi 2002), Aeromonas (Y añez, Catal an & Apr aiz 2003) and Streptococcus (Kim, Park & Hwang 2013). In this study, we developed a quantitative PCR method for S. parauberis targeting the gyrB gene sequences detected by the gyrB-F20/R25 primer pair and probe as shown in Table 1.…”

Section: Discussionmentioning

confidence: 99%

Development of real‐time PCR for detection and quantitation of Streptococcus parauberis

Kim

et al. 2014

Journal of Fish Diseases

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Streptococcus parauberis is an increasing threat to aquaculture of olive flounder, Paralichthys olivaceus Temminck & Schlegel, in South Korea. We developed a real-time polymerase chain reaction (PCR) method using the TaqMan probe assay to detect and quantify S. parauberis by targeting the gyrB gene sequences, which are effective for molecular analysis of the genus Streptococcus. Our real-time PCR assay is capable of detecting 10 fg of genomic DNA per reaction. The intra- and interassay coefficient of variation (CV) values ranged from 0.42-1.95%, demonstrating that the assay has good reproducibility. There was not any cross-reactivity to Streptococcus iniae or to other streptococcal/lactococcal fish pathogens, such as S. agalactiae and Lactococcus garvieae, indicating that the assay is highly specific to S. parauberis. The results of the real-time PCR assay corresponded well to those of conventional culture assays for S. parauberis from inoculated tissue homogenates (r = 0.957; P < 0.05). Hence, this sensitive and specific real-time PCR is a valuable tool for diagnostic quantitation of S. parauberis in clinical samples.

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Simultaneous Detection of Streptococcus pneumoniae, S. mitis, and S. oralis by a Novel Multiplex PCR Assay Targeting the gyrB Gene

Cited by 14 publications

References 49 publications

Function of BriC peptide in the pneumococcal competence and virulence portfolio

Function of BriC peptide in the pneumococcal competence and virulence portfolio

A novel streptococcal cell–cell communication peptide promotes pneumococcal virulence and biofilm formation

Development of real‐time PCR for detection and quantitation of Streptococcus parauberis

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