Rapid detection of methicillin-resistant Staphylococcus aureus in screening samples by relative quantification between the mecA gene and the SA442 gene

Bode, L.; Wunnik, Perry van; Vaessen, Norbert; Savelkoul, Paul H. M.; Smeets, Leo C.

doi:10.1016/j.mimet.2012.02.014

Cited by 17 publications

(10 citation statements)

References 19 publications

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“…Recently however, multiple studies have compared the use of broth enrichment in combination with real-time PCR assays to the combination of broth enrichment followed by culture methods. These studies have shown high sensitivities when using broth enrichment and subsequent real-time PCR, showing the reliability of the combined methods (21,24,25). Based on the published data in combination with the data here, we suggest that recommendations on the use of broth enrichment in combination with real-time PCR assays be included in the Dutch guidelines on the laboratory detection of MRSA.…”

Section: Discussionsupporting

confidence: 60%

“…Using this molecular screening approach, 95.2% of the samples could be reported negative based on the real-time PCR assays. Both assays had shown good sensitivities and negative predictive values in earlier studies, making this screening approach very useful for detection of MRSA-negative samples (15,21).…”

Section: Discussionmentioning

confidence: 92%

“…The surplus value of the GeneOhm MRSA assay was demonstrated because MRSA was cultured in 79.4% of all samples that tested positive by the GeneOhm MRSA assay, while the percentage was much lower when samples were cultured directly following the SMM panel (24%). Nevertheless, it should be stressed that positive results of both real-time PCR assays should always be confirmed using culture, since false-positive interpretations can be observed in the SMM panel and the GeneOhm MRSA assay because of a mixture of MRCNS and MSSA or a SCCmec-cassette with a mecA dropout, respectively (21)(22)(23).…”

Section: Discussionmentioning

confidence: 99%

“…Bode et al applied this strategy in their method and focused on the difference in C T values between SA442 and mecA (dC T , the C T value of SA442 minus the C T value of mecA) (21). By introducing this dC T , they showed 97.5% sensitivity (39/40) and 96.6% specificity (1,701/1,760) when a dC T between Ϫ2 and ϩ8 was assessed.…”

Section: Discussionmentioning

confidence: 99%

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A Rapid and High-Throughput Screening Approach for Methicillin-Resistant Staphylococcus aureus Based on the Combination of Two Different Real-Time PCR Assays

et al. 2014

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b Methicillin-resistant Staphylococcus aureus (MRSA) is an important pathogen that has been responsible for major nosocomial epidemics worldwide. For infection control programs, rapid and adequate detection of MRSA is of great importance. We developed a rapid and high-throughput molecular screening approach that consists of an overnight selective broth enrichment, followed by mecA, mecC, and S. aureus-specific (SA442 gene) real-time PCR assays, with subsequent confirmation using a staphylococcal cassette chromosome mec element (SCCmec)-orfX-based real-time PCR assay (GeneOhm MRSA assay) and culture. Here, the results of the screening approach over a 2-year period are presented. During this period, a total of 13,387 samples were analyzed for the presence of MRSA, 2.6% of which were reported as MRSA positive. No MRSA isolates carrying the mecC gene were detected during this study. Based on the results of the real-time PCR assays only, 95.2% of the samples could be reported as negative within 24 h. Furthermore, the performance of these real-time PCR assays was evaluated using a set of 104 assorted MRSA isolates, which demonstrated high sensitivity for both the combination of mecA and mecC with SA442 and the BD GeneOhm MRSA assay (98.1% and 97.1%, respectively). This molecular screening approach proved to be an accurate method for obtaining reliable negative results within 24 h after arrival at the laboratory and contributes to improvement of infection control programs, especially in areas with a low MRSA prevalence.

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Section: Discussionsupporting

confidence: 60%

Section: Discussionmentioning

confidence: 92%

Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

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A Rapid and High-Throughput Screening Approach for Methicillin-Resistant Staphylococcus aureus Based on the Combination of Two Different Real-Time PCR Assays

et al. 2014

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“…With the acquisition of mecA gene they showed high resistance to all penicillins and cephalosporin's (beta-lactams) (Lowy, 1998). The mecA gene is located on a mobile genomic island called Staphylococcal cassette chromosome mec (SCCmec) (Bode et al, 2012). Nuc gene is located on a plasmid (pIP1607) which encodes the thermo-stable nuclease of S. aureus (Brakstad et al, 1992).…”

Section: S Taphylococcus Aureus Isolates Including Methicillin Resistmentioning

confidence: 99%

Prevalence of MRSA Containing SCCmec Types Isolated from a Tertiary Care Hospital, Rawalpindi

Arshad¹,

Akhtar²,

Irum³

2015

J. Inf. Mol. Biol.

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| With the changes in epidemiology of methicillin-resistant Staphylococcus aureus (MRSA), accurate information on the scope and magnitude of MRSA infections is needed. This study was conducted to investigate the SCCmec types among MRSA strains collected from a hospital in Rawalpindi. Non-duplicate isolates of Staphylococci were isolated (n=28) from pus specimens (n=100). All strains were tested for methicillin resistance using Cefoxitin disc (30 µg) on Muller-Hinton agar. The phenotypically confirmed isolates (n=22) were screened for the presence of mecA, nuc genes and SCCmec types through multiplex PCR. Both mecA and nuc genes were detected in all Staphylococcus aureus strains, whereas, two SCCmec types III and I were detected in (77.27%) and (31.81%) of isolates, respectively along with subtypes. Antimicrobial susceptibility revealed Vancomycin as the most effective drug against all clinical isolates of MRSA. It is concluded that prevalence of MRSA is quite high in and around Rawalpindi area and thus requires necessary interventions for its effective treatment and control.

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Development and first evaluation of a novel multiplex real-time PCR on whole blood samples for rapid pathogen identification in critically ill patients with sepsis

Groep

Bos²,

Savelkoul

et al. 2018

Eur J Clin Microbiol Infect Dis

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Molecular tests may enable early adjustment of antimicrobial therapy and be complementary to blood culture (BC) which has imperfect sensitivity in critically ill patients. We evaluated a novel multiplex real-time PCR assay to diagnose bloodstream pathogens directly in whole blood samples (BSI-PCR). BSI-PCR included 11 species- and four genus-specific PCRs, a molecular Gram-stain PCR, and two antibiotic resistance markers. We collected 5 mL blood from critically ill patients simultaneously with clinically indicated BC. Microbial DNA was isolated using the Polaris method followed by automated DNA extraction. Sensitivity and specificity were calculated using BC as reference. BSI-PCR was evaluated in 347 BC-positive samples (representing up to 50 instances of each pathogen covered by the test) and 200 BC-negative samples. Bacterial species-specific PCR sensitivities ranged from 65 to 100%. Sensitivity was 26% for the Gram-positive PCR, 32% for the Gram-negative PCR, and ranged 0 to 7% for yeast PCRs. Yeast detection was improved to 40% in a smaller set-up. There was no overall association between BSI-PCR sensitivity and time-to-positivity of BC (which was highly variable), yet Ct-values were lower for true-positive versus false-positive PCR results. False-positive results were observed in 84 (4%) of the 2200 species-specific PCRs in 200 culture-negative samples, and ranged from 0 to 6% for generic PCRs. Sensitivity of BSI-PCR was promising for individual bacterial pathogens, but still insufficient for yeasts and generic PCRs. Further development of BSI-PCR will focus on improving sensitivity by increasing input volumes and on subsequent implementation as a bedside test.Electronic supplementary materialThe online version of this article (10.1007/s10096-018-3255-1) contains supplementary material, which is available to authorized users.

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Rapid detection of methicillin-resistant Staphylococcus aureus in screening samples by relative quantification between the mecA gene and the SA442 gene

Cited by 17 publications

References 19 publications

A Rapid and High-Throughput Screening Approach for Methicillin-Resistant Staphylococcus aureus Based on the Combination of Two Different Real-Time PCR Assays

A Rapid and High-Throughput Screening Approach for Methicillin-Resistant Staphylococcus aureus Based on the Combination of Two Different Real-Time PCR Assays

Prevalence of MRSA Containing SCCmec Types Isolated from a Tertiary Care Hospital, Rawalpindi

Development and first evaluation of a novel multiplex real-time PCR on whole blood samples for rapid pathogen identification in critically ill patients with sepsis

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