1. Evaluation of the BD MAX™ MRSA XT Assay in a Clinical Laboratory
Carly Kubasek, Faris Galambo, Elaine Vendrone, Suzane Silbert, Ray Widen
Department of Esoteric Testing/R&D, Tampa General Hospital, Tampa, FL
2014 CVS
Annual Meeting
T80
Abstract
Methods continued
Results
Background: The BD MAX™ MRSA XT assay (BD Diagnostics, Québec, Canada) performed on the BD MAX™ System (BD Diagnostics, Sparks, MD) is an FDA-cleared assay designed to detect methicillin-resistant Staphylococcus aureus (MRSA) specific DNA in nasal samples. Probes labeled with different fluorophores are used to detect a specific amplicon in the SCCmec right-extremity junction (MREJ), the genes for methicillin resistance mecA and mecC, and sample processing control (SPC) amplicons in three different optical channels of the BD MAX™ System. The objective of this study was to compare the new BD MAX™ MRSA XT assay with the standard PCR MRSA test used in our laboratory: the BD MAX™ MRSA 1st generation assay (BD Diagnostics). Methods: A total of 118 clinical samples were tested. Specimens were collected from the nares of each patient, using an NP ESwab (Copan Diagnostics Inc, USA). First, two aliquots of 200µL each of the same ESwab sample were transferred to two BD MAX Sample Buffer Tubes (SBT). Each SBT was tested separately by the BD MAX™ MRSA assay and by the BD MAX™ MRSA XT assay, according to manufacturer’s recommendations. This allowed direct comparison of results from the same patient. Direct cultures of the samples with discordant results were performed by pipetting 50µL of the remaining ESwab samples to each of the following media: TSA II, BBL™ CHROMagar™ Staph aureus, BBL CHROMagar MRSA II, and TSB w/ 6.5% NaCl (BD Diagnostics). Plates were examined for growth after 20-26h of incubation at 35°C. S. aureus identification was confirmed by latex agglutination tests and by an in-house validated mecA/femA specific PCR. Additionally, a challenge panel comprising multiple control strains was evaluated by the BD MAX™ MRSA XT assay to verify the ability of the assay to correctly identify a variety of MRSA strains and to exclude S. aureus strains with mecA gene dropouts, incorrectly identified as MRSA. Results: Out of 118 clinical samples tested, 34 were MRSA positive by both assays and 2 were MRSA positive by only the BD MAX™ MRSA 1st generation assay. Culture results from these 2 discordant samples revealed the presence of Staphylococcus epidermidis on the TSA II plate and in the TSB w/ 6.5% NaCl. No discordant results were obtained with the BD MAX™ MRSA XT assay for the challenge panel. Conclusion: Our results showed that the new BD MAX™ MRSA XT assay performed on the BD MAX™ System has excellent sensitivity and specificity. In addition to that, the new BD MAX™ MRSA XT assay appeared to have the ability to correctly identify a variety of MRSA strains and excludes the strains of S. aureus with mecA gene dropouts, incorrectly identified by the 1st generation assay as MRSA.
Bacterial Culture:
50 µL of the remaining ESwab samples were plated into each of the following media:
TSA II, BBL™ CHROMagar™ S. aureus, BBL CHROMagar MRSA II, and TSB w/ 6.5% NaCl (BD Diagnostics).
Colony growth was observed after 20-26h of incubation at 35°C.
S. aureus identification was confirmed by latex agglutination tests and by an in-house validated mecA/femA specific PCR.
A challenge panel comprising multiple control strains was evaluated by the BD MAX™ MRSA XT and the Cepheid Xpert® MRSA assays to verify the ability of the assays to correctly identify MRSA.
Table 1. BD MAX™ MRSA 1st Generation, BD MAX™ MRSA XT and Direct Culture Results:
Table 2. Discrepant Results Analysis Among BD MAX™ MRSA 1st Generation, BD MAX™ MRSA XT and MRSA Culture:
Results
BD MAX MRSA 1st G.
BD MAX
MRSA XT
Direct Culture MRSA
Positive 36 34 33
Negative 82 84 85
Tests/
Samples
MRSA 1st G.
PCR
MRSA XT PCR
Direct Culture
Enrichment Culture
femA PCR
mecA PCR
Sample 14 + -
Sample 27*
Sample 29*
Figure 1. BD MAX™ Instrument Operation
One manual
pipetting step – 200 µL aliquot from ESwab sample transferred to a Sample Buffer Tube
*Samples 27 (Ct=36) and 29 (Ct=35) were negative for MRSA by direct and enriched culture, positive for mecA gene but negative for femA. Possibly cassette dropouts from a MRSA mixed with a MRCONS.
Table 3. Challenge Panel Results:
CONCLUSIONS
Load reagents
and specimens onto the BD MAX™ rack ID
Strain Description
MREJ Type
BD MAX MRSA XT
Cepheid Xpert MRSA
3097
MSSA
(mecA dropout) ii - + 1
MRSA i
2800 9
iii 11 iv 16 v
2937 vi 19
vii
131 ix
2952
xiii
797
xiv
ATCC
BAA- 2312
(mecC)
xxi
ATCC 29213 NA
ATCC 14990
MSSE
The BD MAX™ MRSA XT is a new automated real-time PCR assay that is able to detect MRSA in nares samples in a single walk away sample in – answer out test.
The first generation BD MAX™ MRSA and the BD MAX™ MRSA XT assay displayed 98.3% overall agreement.
Two samples were positive on the first generation BD MAX™ MRSA and negative on the BD MAX™ MRSA XT. Direct and enriched cultures, and femA PCR were negative for MRSA as well. Positive results for mecA gene were detected suggesting the possibility of a mecA dropout.
The resolved sensitivity and specificity for the BD MAX™ MRSA XT were 100%.
The BD MAX™ MRSA XT assay correctly identified all the targets in a challenge panel that include a variety of MREJ types, along with an isolate that contained mecC rather than mecA. Importantly the BD MAX™ MRSA XT correctly excluded a mecA dropout MSSA isolate.
The data indicates that the BD MAX™ MRSA XT assay provides highly accurate and rapid detection of MRSA in nares samples used in screening patients for colonization with MRSA. The BD MAX MRSA XT assay can potentially assist with isolation steps to help reduce post procedure or nosocomial infections.
Introduction
Create worklist
Methicillin-resistant Staphylococcus aureus (MRSA) is a leading cause of healthcare-associated infections. MRSA colonization is associated with increased risk of disease, high cost and poor clinical outcomes. Early identification of patients colonized with MRSA and subsequent prevention of patient-to-patient spread through infection control measures are considered important interventions to control MRSA.
Traditional culture-based methods for detecting MRSA depend on isolation of pure colonies, followed by identification of S. aureus and confirmation of methicillin resistance either by oxacillin susceptibility testing, detection of the mecA gene or detection of penicillin binding protein PBP2a encoded by the mecA gene. The BD MAX™ MRSA XT assay is a new automated molecular assay that detects the SCCmec right-extremity junction (MREJ) and the mecA and mecC methicillin resistance genes.
Place racks and
cartridges on BD MAX™
Methods
A total 118 clinical samples were collected from the nares of patients for routine MRSA testing, using a NP ESwab (Copan Diagnostics). Note* Use of the ESwab with the BD MAX MRSA XT and BD MAX MRSA assay is outside of package insert claims; however, it’s use was validated at Tampa General Hospital.
BD MAX™ MRSA XT and MRSA Assay (1st Generation)
200 µL of ESwab sample were transferred to two BD MAX Sample Buffer Tubes (SBT)
The Sample Buffer Tubes were vortexed on a multi-tube vortexer for 60 seconds.
The SBTs were transferred to two BD MAX racks, one for each assay.
Unitized Reagent Strips containing assay-specific reagent tubes were placed in BD MAX racks.
The entire assembly was placed in the BD MAX instrument along with BD MAX PCR cartridges.
Close door to initiate run then release results through LIS interface
This study was supported by BD Diagnostics (Sparks, MD).