Use of a Multiplex Molecular Beacon Platform for Rapid Detection of Methicillin and Vancomycin Resistance in
            <i>Staphylococcus aureus</i>

Sinsimer, Daniel; Leekha, Surbhi; Park, Steven; Marras, Salvatore A. E.; Koreen, Larry; Willey, Barbara M.; Naidich, Steve; Musser, Kimberlee A.; Kreiswirth, Barry N.

doi:10.1128/jcm.43.9.4585-4591.2005

Cited by 33 publications

(20 citation statements)

References 26 publications

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“…A conserved sequence in the 16S rRNA was selected to count the total bacteria. 16 All primers were commercially synthesized (Bioneer, Seoul, Korea).…”

Section: Methodsmentioning

confidence: 99%

Changes in salivary periodontal pathogens after orthodontic treatment: An in vivo prospective study

Kim

Jung

Cho

et al. 2015

The Angle Orthodontist

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Objective: To analyze the initial changes in salivary levels of periodontal pathogens after orthodontic treatment with fixed appliances. Materials and Methods: The subjects consisted of 54 adult patients. The Simplified Oral Hygiene Index, Plaque Index, and Gingival Index were measured as periodontal parameters. Both the plaque and gingival indexes were obtained from the central and lateral incisors and first molars of both arches. Whole saliva and periodontal parameters were obtained at the following four time points: immediately before debonding (T1), 1 week after debonding (T2), 5 weeks after debonding (T3), and 13 weeks after debonding (T4). Repeated measures analysis of variance was used to determine salivary bacterial levels and periodontal parameters among the four time points after quantifying salivary levels of Aggregatibacter actinomycetemcomitans (Aa), Fusobacterium nucleatum (Fn), Porphyromonas gingivalis (Pg), Prevotella intermedia (Pi), Tannerella forsythia (Tf), and total bacteria using the real-time polymerase chain reaction. Results: All periodontal parameters were significantly decreased immediately after debonding (T2). The salivary levels of total bacteria and Pg were decreased at T3, while Pi and Tf levels were decreased at T4. However, the amount of Aa and Fn remained at similar levels in saliva during the experimental period. Interestingly, Aa and Fn were present in saliva at higher levels than were Pg, Pi, and Tf. Conclusion: The higher salivary levels of Aa and Fn after debonding suggests that the risk of periodontal problems cannot be completely eliminated by the removal of fixed orthodontic appliances during the initial retention period, despite improved oral hygiene. (Angle Orthod. 2016;86:998-1003.)

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“…A conserved sequence in the 16S rRNA was selected to count the total bacteria. 16 All primers were commercially synthesized (Bioneer, Seoul, Korea).…”

Section: Methodsmentioning

confidence: 99%

Changes in salivary periodontal pathogens after orthodontic treatment: An in vivo prospective study

Kim

Jung

Cho

et al. 2015

The Angle Orthodontist

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“…Isolates were also screened for the presence of arginine catabolic mobile element (ACME), which is known to be associated with USA300 (12). Detection of Panton-Valentine leukocidin in these strains was performed by real-time PCR with molecular beacons targeting the lukF-PV gene (51). Detection of ACME was accomplished by conventional PCR targeting two genes from the ACME operon, arcA and opp3A.…”

Section: Methodsmentioning

confidence: 99%

Diverse Enterotoxin Gene Profiles among Clonal Complexes of Staphylococcus aureus Isolates from the Bronx, New York

Varshney

Mediavilla

Robiou

et al. 2009

Appl Environ Microbiol

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101

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Staphylococcal enterotoxins (SE) can cause toxin-mediated disease, and those that function as superantigens are implicated in the pathogenesis of allergic diseases. The prevalence of 19 enterotoxin genes was determined by PCR in clinical S. aureus strains derived from wounds (108) and blood (99). We performed spa typing and multilocus sequence typing (MLST) to determine clonal origin, and for selected strains staphylococcal enterotoxin B (SEB) production was measured by enzyme-linked immunosorbent assay. Strains carried a median of five SE genes. For most SE genes, the prevalence rates among methicillinresistant and methicillin-sensitive S. aureus isolates, as well as wound-and blood-derived isolates, did not differ. At least one SE gene was detected in all except two S. aureus isolates (>99%). Complete egc clusters were found in only 11% of S. aureus isolates, whereas the combination of sed, sej, and ser was detected in 24% of clinical strains. S. aureus strains exhibited distinct combinations of SE genes, even if their pulsed-field gel electrophoresis and MLST patterns demonstrated clonality. USA300 strains also showed considerable variability in SE content, although they contained a lower number of SE genes (mean, 3). By contrast, SE content was unchanged in five pairs of serial isolates. SEB production by individual strains varied up to 200-fold, and even up to 15-fold in a pair of serial isolates. In conclusion, our results illustrate the genetic diversity of S. aureus strains with respect to enterotoxin genes and suggest that horizontal transfer of mobile genetic elements encoding virulence genes occurs frequently.As a commensal, Staphylococcus aureus colonizes the nasal mucosa of 20 to 40% of humans (54), and as a pathogen it causes pyogenic diseases and toxin-mediated diseases (38). S. aureus produces many different virulence factors, including enterotoxins (SEs), which can cause defined toxic shock syndromes (4). The characterization of some of these toxins led to the discovery of superantigens (41), which bind to major histocompatibility complex class II molecules and V␤ chains of T-cell receptors, resulting in the activation of large numbers of T cells (20 to 30%) and massive cytokine production (10, 18). These superantigen-induced "cytokine storms" are responsible for the toxic effects seen in staphylococcal entertoxin B (SEB)-and toxic shock syndrome toxin (TSST)-associated shock syndromes in S. aureus infections (13, 40, 47). To date, 19 SEs have been identified based on sequence homologies, and studies have reported enterotoxin genes in up to 80% of all S. aureus strains (4, 21). Although many new enterotoxins have been identified, i.e., seg ser and seu (33,37,44,49), their precise functions have not been characterized yet. The majority of experimental work with SEs is still done with SEB, toxic shock syndrome toxin 1, and SEA (27, 31), because these toxins are commercially available. Most SEs are located on mobile elements in bacterial genomes such as plasmids or pathogenicity islands and can thu...

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“…Finally, a universally conserved region of mecA was selected as an internal control for SCCmec, as well as to identify examples of SCC elements lacking mecA. Additional primers and molecular beacons used in this study included (i) a primerprobe set targeting the J1 region of SCCmec subtype IVa and (ii) a primer-probe set specific for the protein A (spa) gene of S. aureus (70), used to differentiate S. aureus from CoNS and to confirm MB-PCR results for methicillin-susceptible S. aureus (MSSA) strains.…”

Section: Methodsmentioning

confidence: 99%

Multiplex Real-Time PCR for Rapid Staphylococcal Cassette Chromosome mec Typing

et al. 2009

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Rapid identification and typing of methicillin (meticillin)-resistant Staphylococcus aureus (MRSA) is important for understanding the molecular epidemiology and evolution of MRSA and offers many advantages for controlling transmission in both health care and community settings. We developed a rapid molecular beacon real-time PCR (MB-PCR) assay for staphylococcal cassette chromosome mec (SCCmec) typing. The design of this system is based on the established definition of SCCmec types, namely, the combination of the mec class complex with the ccr allotype. The assay consists of two multiplex panels, the combination of which results in two targets (mec class, ccr) for each SCCmec type. MB-PCR panel I targets mecA, ccrB2, mecI, and the ⌬mecR1-IS1272 junction (mec class B); it can definitively identify SCCmec types II and IV. MB-PCR panel II detects ccrC, ccrB1, ccrB3, ccrB4, and the ⌬mecR1-IS431 junction (mec class C2) and is therefore capable of identifying SCCmec types I, III, V, and VI in combination with panel I. The method can also detect the recently described novel SCCmec type VIII (ccrAB4 with mec class A). Our assay demonstrated 100% concordance when applied to 162 MRSA strains previously characterized by traditional SCCmec typing schemes. Four geographically and temporally diverse S. aureus collections were also successfully classified by our assay, along with 1,683 clinical isolates comprising both hospital-and community-associated MRSA and methicillin-susceptible S. aureus strains. As many as 96 isolates can be classified easily within 3 to 4 h, including DNA isolation, PCR cycling, and analysis. The assay is rapid, robust, sensitive, and cost-effective, allowing for high-throughput SCCmec typing of MRSA isolates.

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Use of a Multiplex Molecular Beacon Platform for Rapid Detection of Methicillin and Vancomycin Resistance in Staphylococcus aureus

Cited by 33 publications

References 26 publications

Changes in salivary periodontal pathogens after orthodontic treatment: An in vivo prospective study

Changes in salivary periodontal pathogens after orthodontic treatment: An in vivo prospective study

Diverse Enterotoxin Gene Profiles among Clonal Complexes of Staphylococcus aureus Isolates from the Bronx, New York

Multiplex Real-Time PCR for Rapid Staphylococcal Cassette Chromosome mec Typing

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