(261 days)
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No
The description mentions a result algorithm using a series of decision trees to evaluate features of a kinetic luminescent curve. While decision trees are a type of algorithm, the description does not indicate that this algorithm learns or improves from data in a way characteristic of machine learning. The development process described (using known samples and ROC analysis to set a cutoff) is a standard statistical approach, not necessarily indicative of AI/ML. There is no mention of training or inference in the context of a learning model.
No
The device is described as an "in vitro diagnostic test" intended to aid in the prevention and control of MRSA infections by detecting live MRSA cells, not to treat or directly alleviate a condition.
Yes
The "Intended Use / Indications for Use" section explicitly states "The cobas vivoDx MRSA test... is an automated qualitative in vitro diagnostic test for the direct detection of live methicillin-resistant Staphylococcus aureus (MRSA) cells". The term "in vitro diagnostic test" directly indicates its function as a diagnostic device.
No
The device description explicitly mentions the "cobas vivoDx Instrument" which is a piece of hardware used for automated processing, incubation, addition of reagents, measurement, and analysis of the luminescent signal. This indicates the device is not software-only.
Yes, this device is an IVD (In Vitro Diagnostic).
Here's why:
- Intended Use: The very first sentence explicitly states it is an "automated qualitative in vitro diagnostic test".
- Sample Type: It is designed to test "nasal swab samples from patients".
- Purpose: It is intended for the "direct detection of live methicillin-resistant Staphylococcus aureus (MRSA) cells" to "aid in the prevention and control of MRSA infections in healthcare settings". This is a diagnostic purpose performed outside of the body.
- Device Description: The description details a laboratory-based assay using reagents and an instrument to analyze a biological sample.
All of these points align with the definition of an In Vitro Diagnostic device.
N/A
Intended Use / Indications for Use
The cobas vivoDx MRSA test performed on the cobas vivoDx System is an automated qualitative in vitro diagnostic test for the direct detection of live methicillin-resistant Staphylococcus aureus (MRSA) cells in nasal swab samples from patients who are at risk for nasal colonization by MRSA. The test utilizes selective agents and bioparticles (Smarticles technology) to introduce a luciferase gene into targeted bacteria to create an amplified luminescent signal in only viable (live) MRSA cells. The cobas vivoDx MRSA test is intended to aid in the prevention and control of MRSA infections in healthcare settings. It is not intended to diagnose MRSA infections, nor to guide, or monitor treatment. Concomitant cultures are necessary to recover organisms for epidemiological typing or for further susceptibility testing.
The cobas vivoDx MRSA Collection and Transport Kit is used to collect, transport and store human nasal swab specimens for use with the cobas vivoDx MRSA test.
Product codes (comma separated list FDA assigned to the subject device)
QIV, OIV
Device Description
The cobas vivoDx MRSA test, performed on the cobas vivoDx Instrument, is an automated, phenotypic assay for the direct, qualitative detection of MRSA in nasal swab specimens that are collected and transported to the testing laboratory using the cobas vivoDx MRSA Collection and Transport Kit.
Upon receipt in the laboratory, the test operator vortexes the specimen to elute the target organisms (if present) and transfers an aliquot of the transport medium to an MRSA Test Tube containing a dried bead of a selective reagent (cefoxitin) that is solubilized with the sample. The operator then applies a MRSA Reagent Cap to the Test Tube, briefly vortexes the assembled cartridge and loads it onto the cobas vivoDx Instrument for automated processing.
The MRSA Reagent Cap comprises two reagent-filled blisters, one containing Staphylococcus aureus-specific bacteriophage-based Smarticles that encode the enzyme luciferase, and the other, a luminescent substrate. The cobas vivoDx Instrument automatically completes incubation of the cartridge, addition of the Smarticles and substrate to the test sample, as well as measurement and analysis of the luminescent signal. Results are reported as "Positive" (MRSA detected) or "Negative" (no MRSA detected).
Mentions image processing
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Mentions AI, DNN, or ML
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Input Imaging Modality
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Anatomical Site
Nasal swab samples
Indicated Patient Age Range
18 years of age
Intended User / Care Setting
Healthcare settings
Description of the training set, sample size, data source, and annotation protocol
The result algorithm for the cobas vivoDx MRSA test uses a series of decision trees to evaluate features of the kinetic luminescent curve obtained after addition of substrate to the test sample. The algorithm was developed using data from known MRSA positive and negative analytical and clinical samples and the cutoff was selected based on Receiver Operating Curve (ROC) analysis to optimize sensitivity and specificity.
Description of the test set, sample size, data source, and annotation protocol
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Summary of Performance Studies (study type, sample size, AUC, MRMC, standalone performance, key results)
Site-to-Site Reproducibility Study:
Evaluated the between-laboratory reproducibility of the cobas vivoDx MRSA test by testing a panel of samples. Panel members were prepared on each day of testing by serial dilution of bacterial suspensions. Specific sample size and duration of study were redacted as (b) (4). Conducted at (b) (4) sites with one cobas vivoDx system at each site and (b) (4) operators participating at each site. The results obtained were deemed acceptable.
Lot-to-Lot Reproducibility:
Evaluated the reproducibility of the cobas vivoDx MRSA test between reagent lots by testing (b) (4). Results were acceptable.
Reproducibility with Borderline Cefoxitin Susceptible/Resistant S. aureus:
Evaluated reproducibility using isolates of S. aureus that exhibited cefoxitin/oxacillin MICs close to the breakpoints for susceptibility/resistance. The study was conducted at a (b) (4) using (b) (4) reagents. Panel members were prepared fresh daily in simulated nasal matrix and organism concentrations were verified by performing viable counts. Each panel member was tested (b) (4). The study demonstrated acceptable reproducibility for detection of borderline cefoxitin susceptible/resistant S. aureus within and between instruments, reagent lots and days.
Limit of Detection (LoD) Study:
Determined the LoD of the cobas vivoDx MRSA test by testing serial dilutions of five different strains of MRSA in simulated nasal matrix. For each strain, at least 20 replicates were tested at each target level with each of three lots of cobas vivoDx MRSA reagents. The concentration of organisms present in each dilution was confirmed by performing viable counts. The LoD for each strain and reagent lot combination, defined as the lowest target level at which 95% of replicates are expected to produce positive results, was estimated by Probit analysis. The highest LoD point estimate was 315 CFU/mL for strain NRS 642 with Reagent Lot #2. The analytical sensitivity for the heteroresistant ATCC 43300 strain was evaluated, with the highest LoD point estimate being 320 CFU/mL.
Analytical Reactivity (Inclusivity) Studies:
- Testing of Alternative PFGE and SCCmec Types: A summary of study results showed initial testing produced (b) (4) positive results.
- Borderline Cefoxitin Susceptibility Panel: Included a panel of S. aureus strains from the CDC & FDA Antimicrobial Resistance Isolate Bank with cefoxitin MICs close to susceptibility/resistance breakpoints. Each strain was initially tested in triplicate in simulated nasal matrix at approximately 1.2 x 10^3 CFU/mL. Additional testing at higher concentrations was performed for strains not yielding 3/3 positive results. 13 of 22 resistant isolates (59.1%) were detected at 1.2 x 10^3 CFU/mL, and the remaining 9 were detected at 1.2 x 10^4 CFU/mL. Of 10 borderline susceptible isolates, 9 (90.0%) were negative even at 1.2 x 10^6 CFU/mL.
Analytical Specificity (Cross-reactivity) Study:
Evaluated (b) (4) strains of bacteria in triplicate at a concentration of (b) (4) CFU/mL. Included 30 strains of MSSA and other nasal flora organisms. No false positive results were obtained with MSSA. Cross-reaction was observed with five other organisms (Listeria monocytogenes, Staphylococcus equorum, three Enterococcus species).
Microbial Interference Study:
Evaluated interference using the same panel of organisms as the Cross-reactivity Study. Each species was tested in triplicate at concentrations between 3.4 x 10^3 and 2.1 x 10^7 CFU/mL in the presence of two MRSA strains at approximately 4X LoD. No interference was observed for MRSA strain NRS 725. False negative results were obtained for MRSA strain NRS 642 in the presence of certain interfering organisms (Citrobacter koseri, Corynebacterium jeikeium, Enterobacter cloacae, Klebsiella pneumoniae) at high levels.
Interfering Substances Study:
Evaluated potential interference by endogenous and exogenous substances. Two MRSA strains were tested at approximately 4X LoD with 24 substances at different concentrations. False negative results were obtained in the presence of topical mupirocin at every concentration. Visibly bloody swabs were excluded from the clinical performance evaluation.
Method Comparison (Matrix comparison):
Compared performance in natural and simulated nasal matrix. Two MRSA strains were diluted in both matrices to approximately the same concentrations. The study demonstrated similar analytical sensitivity in both matrix types, supporting the use of simulated matrix in analytical studies.
Clinical Study:
A prospective Clinical Study was conducted at seven geographically diverse locations in the U.S.
Sample Size: Of 4,198 paired specimens enrolled, 3,963 were evaluable. 210 (5.3%) subjects were MRSA positive based on direct/enriched culture.
Performance against Direct & Enriched Culture (reference):
- Sensitivity: 90.0% (189/210); 85.2-93.4%
- Specificity: 98.6% (3702/3753); 98.2-99.0%
- Positive Predictive Value: 78.8% (189/240)
- Negative Predictive Value: 99.4% (3702/3723)
Performance against Direct Culture alone: - Positive Percent Agreement: 96.7% (176/182); 93.0-98.5%
- Negative Percent Agreement: 98.3% (3717/3781); 97.8-98.7%
Key results indicated that 15/21 false negative cobas vivoDx MRSA test results were MRSA positive only by enriched culture (suggesting low level colonization). 13/51 false positive cobas vivoDx MRSA results compared to the Reference Method were MRSA positive by alternative methods (PCR and/or nonselective direct and enrichment culture). 26/64 false positive cobas vivoDx MRSA results compared to Direct Culture were MRSA positive by alternative methods.
Key Metrics (Sensitivity, Specificity, PPV, NPV, etc.)
Clinical Performance vs. Direct & Enriched Culture (reference):
- Sensitivity: 90.0% (189/210); 85.2-93.4%
- Specificity: 98.6% (3702/3753); 98.2-99.0%
- Positive Predictive Value: 78.8% (189/240)
- Negative Predictive Value: 99.4% (3702/3723)
Clinical Performance vs. Direct Culture alone:
- Positive Percent Agreement: 96.7% (176/182); 93.0-98.5%
- Negative Percent Agreement: 98.3% (3717/3781); 97.8-98.7%
Predicate Device(s): If the device was cleared using the 510(k) pathway, identify the Predicate Device(s) K/DEN number used to claim substantial equivalence and list them here in a comma separated list exactly as they appear in the text. List the primary predicate first in the list.
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Reference Device(s): Identify the Reference Device(s) K/DEN number and list them here in a comma separated list exactly as they appear in the text.
Not Found
Predetermined Change Control Plan (PCCP) - All Relevant Information for the subject device only (e.g. presence / absence, what scope was granted / cleared under the PCCP, any restrictions, etc).
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§ 866.1655 System for detection of microorganisms and antimicrobial resistance using reporter expression.
(a)
Identification. A system for detection of microorganisms and antimicrobial resistance using reporter expression is an in vitro diagnostic device intended for the detection and identification of live microorganisms and the detection of associated antimicrobial drug susceptibility or resistance in specimens from patients at risk of colonization or suspected of infection.(b)
Classification. Class II (special controls). The special controls for this device are:(1) The intended use for the device in the labeling required under § 809.10 of this chapter must include a detailed description of the targets the device detects, the type of results provided to the user, the clinical indications appropriate for test use, and the specific population(s) for which the device is intended.
(2) Any device used for specimen collection and transport must be FDA-cleared, approved, or -classified as 510(k) exempt (standalone or as part of a test system) for the collection of the specimen types claimed by this device and for the maintenance of viability of the targeted microorganisms; alternatively, the specimen collection device must be cleared in a premarket submission as a part of this device.
(3) The labeling required under § 809.10(b) of this chapter must include:
(i) A detailed description of the device, including reagents, instruments, ancillary materials, applicable specimen collection and transport device(s) and control elements, and a detailed explanation of the methodology, including all pre-analytical methods for handling and processing of specimens and controls to maintain organism viability;
(ii) Detailed descriptions of the test procedure, including the preparation and maintenance of quality controls and the interpretation of test results;
(iii) Detailed discussion of the performance characteristics of the device for all claimed organisms and specimen types based on analytical studies, including evaluation of analytical sensitivity, inclusivity, cross-reactivity, potentially interfering substances and microorganisms, contamination, specimen stability, precision, and reproducibility;
(iv) Detailed discussion of the performance characteristics of the device observed in a clinical study performed on a population that is consistent with the intended use population in comparison to the results obtained by a reference or comparator method determined to be acceptable by FDA, for microbial detection, identification, and antimicrobial susceptibility testing; and
(v) A limiting statement indicating that a negative test result does not preclude colonization or infection with organisms that do not express detectable levels of the reporter that is identified by the device.
(4) Design verification and validation must include:
(i) A detailed description of the device, including an explanation of the technology, hardware, software, and consumables, as well as an explanation of the result algorithms and method(s) of data processing from signal acquisition to result assignment;
(ii) A detailed description of the impact of any software, including software applications and hardware-based devices that incorporate software, on the device's functions;
(iii) Detailed documentation of the analytical and clinical studies required in paragraphs (b)(3)(iii) and (iv) of this section, including the study protocols containing descriptions of the test methods, prescribed methods of data analysis and acceptance criteria, final study reports, and data line listings;
(iv) Detailed documentation of quality control procedures, including an explanation of how quality control materials were selected, the recommended frequency of testing, methods of control preparation, acceptance criteria for performance and the results from quality control testing performed during the analytical and clinical studies required under paragraphs (b)(3)(iii) and (iv) of this section;
(v) Detailed documentation of studies performed to establish onboard and in-use reagent stability, including the test method(s), data analysis plans, acceptance criteria, final study reports, and data line listings;
(vi) Detailed documentation of studies to establish reagent shelf-life for the assay kit and each applicable specimen collection and transport device, including study protocols containing descriptions of the test method(s), data analysis plans, and acceptance criteria; and
(vii) Documentation of an appropriate end user device training program that will be offered as part of efforts to assure appropriate conduct of the assay and to mitigate the risk associated with false results, including failure to use the device correctly or correctly interpret results.
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EVALUATION OF AUTOMATIC CLASS III DESIGNATION FOR cobas vivoDx MRSA
DECISION SUMMARY
A. DEN Number:
B. Purpose for Submission:
De Novo request for evaluation of automatic class III designation for the cobas vivoDx MRSA test
C. Measurand:
Bioluminescence produced by viable bacterial cells under selective growth conditions
D. Type of Test:
Bioluminescent assay for the determination of antimicrobial susceptibility
E. Applicant:
Roche Molecular Systems, Inc.
F. Proprietary and Established Names:
cobas vivoDx MRSA
G. Regulatory Information:
-
- Regulation section:
21 CFR 866.1655
- Regulation section:
-
- Classification:
Class II
- Classification:
-
- Product code:
QIV
- Product code:
1
-
- Panel:
- 83 Microbiology
H. Indications for Use:
-
- Indications for use:
cobas vivoDx MRSA:
The cobas vivoDx MRSA test performed on the cobas vivoDx System is an automated qualitative in vitro diagnostic test for the direct detection of live methicillin-resistant Staphylococcus aureus (MRSA) cells in nasal swab samples from patients who are at risk for nasal colonization by MRSA. The test utilizes selective agents and bioparticles (Smarticles technology) to introduce a luciferase gene into targeted bacteria to create an amplified luminescent signal in only viable (live) MRSA cells. The cobas vivoDx MRSA test is intended to aid in the prevention and control of MRSA infections in healthcare settings. It is not intended to diagnose MRSA infections, nor to guide, or monitor treatment. Concomitant cultures are necessary to recover organisms for epidemiological typing or for further susceptibility testing.
cobas vivoDx MRSA Collection and Transport Kit:
The cobas vivoDx MRSA Collection and Transport Kit is used to collect, transport and store human nasal swab specimens for use with the cobas vivoDx MRSA test.
-
- Special conditions for use statement(s):
For prescription use only.
- Special conditions for use statement(s):
cobas vivoDx MRSA is only for use with nasal samples collected with cobas vivoDx MRSA Collection and Transport Kits.
Nasal swab specimens for testing with cobas vivoDx MRSA should be processed as soon as possible after collection and should be stored in the cobas vivoDx MRSA Primary Tube for no more than 15 hours at 15-30°C. Analytical studies have shown that there is increased potential for false positive results with specimens containing high levels of MSSA when specimen processing is delaved.
Valid results must be obtained with appropriately prepared Positive and Negative External Controls for results from the cobas vivoDx MRSA test to be displayed by the system. Follow the directions for preparation of External Controls using the recommended strains. Deviation from the recommended control procedures may lead to erroneous results.
In analytical studies, high levels of Staphylococcus equorum, Listeria monocytogenes and the following species of Enterococcus: E. faecium, E. flavescens and E. gallinarum, were found to cross react in the cobas vivoDx MRSA test and produce positive results.
In analytical studies, high levels of Citrobacter koseri, Corynebacterium jeikeium,
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Enterobacter cloacae, and Klebsiella pneumoniae were found to interfere with detection of low levels of MRSA in the cobas vivoDx MRSA test.
Interfering substances that may cause false negative results include but are not limited to:
- Visible blood on the cobas vivoDx MRSA swab or in the primary tube ●
- Mupirocin .
3. Special instrument requirements:
cobas vivoDx system
I. Device Description:
The cobas vivoDx MRSA test, performed on the cobas vivoDx Instrument, is an automated, phenotypic assay for the direct, qualitative detection of MRSA in nasal swab specimens that are collected and transported to the testing laboratory using the cobas vivoDx MRSA Collection and Transport Kit.
Upon receipt in the laboratory, the test operator vortexes the specimen to elute the target organisms (if present) and transfers an aliquot of the transport medium to an MRSA Test Tube containing a dried bead of a selective reagent (cefoxitin) that is solubilized with the sample. The operator then applies a MRSA Reagent Cap to the Test Tube, briefly vortexes the assembled cartridge and loads it onto the cobas vivoDx Instrument for automated processing.
The MRSA Reagent Cap comprises two reagent-filled blisters, one containing Staphylococcus aureus-specific bacteriophage-based Smarticles that encode the enzyme luciferase, and the other, a luminescent substrate. The cobas vivoDx Instrument automatically completes incubation of the cartridge, addition of the Smarticles and substrate to the test sample, as well as measurement and analysis of the luminescent signal. Results are reported as "Positive" (MRSA detected) or "Negative" (no MRSA detected).
J. Standard/Guidance Document Referenced (if applicable):
Not applicable.
K. Test Principle:
The cobas vivoDx MRSA test is an automated, phenotypic assay for the qualitative detection of MRSA in nasal swab specimens from patients who are at risk of colonization. The device uses selective bacteriophage-based Smarticle technology to transduce cells of Staphylococcus aureus that may be present in nasal swab samples with a reporter plasmid that carries the genes for bacterial luciferase (luxA/luxB). The luciferase genes are constitutively expressed in viable S. aureus cells, resulting in continuous production of the luciferase enzyme. When substrate is added, the luciferase catalyzes a reaction that results in emission of light which can be detected by the cobas vivoDx system. In the absence of viable cells of S. aureus, no luciferase activity is observed.
To discriminate methicillin susceptible and resistant S. aureus, after Smarticle transduction,
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the nasal swab sample is incubated in the presence of a fixed concentration of cefoxitin. Methicillin susceptible S. aureus (MSSA) does not grow under these conditions and therefore does not accumulate luciferase and does not emit light upon addition of substrate. In contrast, methicillin resistant S. aureus remains viable, resulting in accumulation of luciferase and emission of a detectable signal upon addition of the luciferase substrate. The results of the test are interpreted automatically by the cobas vivoDx system.
The major components of the cobas vivoDx MRSA test system are summarized in Table 1.
| Test Component | Description |
|---|---|
| cobas vivoDx System | Automated, continuous loading instrument for processing of cobas |
| vivoDx MRSA test cartridges, comprised of the following sub- systems: | |
| Transfer Unit Movement of test cartridges; barcode reading Temperature Control Station Incubation and shaking of test cartridges Dispense Station Dispense of Smarticle-containing liquid reagent Measurement Station Dispense of luciferase substrate and kinetic measurement of luminescence Input/Output Racks For insertion and removal of test cartridges LED Indicator Interface To indicate operating status Embedded Computer Responsible for running the Instrument Control software Tablet Computer Provides the user interface, result display and interfaces with the LIS External Barcode Reader Used for sample log-in | |
| cobas vivoDx MRSA Collection | |
| and Transport Kit | Sterile collection swab (Puritan Hydraflock) and a primary transport |
| tube containing proprietary medium that is designed to support the | |
| viability of MRSA and inhibit growth other organisms that may be | |
| cobas vivoDx MRSA | MRSA Reagent Cap Comprises two separate blister packs containing Smarticles and luciferase substrate that are punctured by the cobas vivoDx system at specific points in the automated workflow Fitted onto the MRSA Test Tube by the operator after addition of sample MRSA Test Tube Contains a dried bead of cefoxitin that is rehydrated with the test sample |
| cobas vivoDx MRSA Control | |
| Media | The same proprietary medium as in the cobas vivoDx MRSA |
| Collection and Transport Kit for use in preparation of Positive and | |
| Negative External Controls |
Table 1. Major components of the cobas vivoDx MRSA test system
LED: Light Emitting Diode; LIS: Laboratory Information System
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External Positive and Negative Controls comprised of cultured cells of known methicillin susceptible and resistant strains of S. aureus must be tested with cobas vivoDx MRSA at a frequency specified by the user. Unless the expected results for both External Controls are obtained, test results from patient samples are not displayed by the cobas vivoDx instrument. Patient samples may be reported as "Positive" (MRSA detected), "Negative" (MRSA not detected), "Invalid" (due to repeated control failure), "Failed" (due to an instrument error) or "Results pending" (display pending receipt of valid External Control results).
L. Performance Characteristics (if/when applicable):
-
- Analytical performance:
- a. Precision/Reproducibility:
Site-to-Site Reproducibility Study
The between-laboratory reproducibility of the cobas vivoDx MRSA test was evaluated by testing a(b) member panel of samples over a period of (b) (4) at sites, using a single lot of reagents and one cobas vivoDx system at each each of (b) site. (b) operators participated at each site.
Because the cobas vivoDx MRSA test is designed to detect viable organisms, panel members were prepared on each day of testing by serial dilution of bacterial suspensions(b) (4)
The MRSA-positive panel members comprised (b) (4)
(Table 2). (b) (4)
Table 2. Strains of MRSA used to evaluate the reproducibility of the cobas vivoDx MRSA test
| Characteristic | MRSA Strain | |
|---|---|---|
| NRS (b) | NRS (b) | |
| PFGE Type | (b) (4)(A) | |
| SCC mec Type | ||
| Cefoxitin MIC (µg/mL) |
PFGE: Pulsed Field Gel Electrophoresis SCCmec: Staphylococcal Cassette Chromosome mec MIC: Minimal Inhibitory Concentration
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Image /page/5/Figure/0 description: The image shows the text '(b) (4)' in the upper left corner and again in the lower left corner. The text '(Table 3). (b) (4)' is in the middle of the image. The text 'These results are acceptable' is in the lower right corner.
Table 3. Summary of results from the cobas vivoDx MRSA Site-to-Site Reproducibility Study
Image /page/5/Figure/2 description: This image shows a table with the title "Agreement by Panel Member (observed range of CFU/mL) 1". The table has columns labeled "Factor", "Site", "Day", "Operator", and "Overall". The cells in the table are filled with the text "(b) (4)".
Lot-to-Lot Reproducibility
The reproducibility of the cobas vivoDx MRSA test between reagent lots was evaluated by testing (b) (4)
Image /page/5/Picture/5 description: I am sorry, but there is nothing visible in the image. As a result, I am unable to provide a description.
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Image /page/6/Figure/0 description: The image shows a table with the title "Agreement by Panel Member (range of CFU/mL) 1". The table has two columns, "Lot" and "Agreement by Panel Member (range of CFU/mL) 1". The first row contains the text "(b) (4)" in both columns. The second row contains the text "Overall" in the first column and "(b) (4)" in the first column.
Table 4. Summary of results from the cobas vivoDx MRSA Lot-to-Lot Reproducibility Study
Reproducibility with Borderline Cefoxitin Susceptible/Resistant S. aureus To challenge the cobas vivoDx MRSA system, an additional Reproducibility Study was conducted using isolates of S. aureus that exhibited cefoxitin/oxacillin MICs close to the breakpoints for susceptibility/resistance. The study was conducted at a (b) (4)
reagents. Panel members were prepared fresh daily in simulated nasal matrix and organism concentrations were verified by performing viable counts (Table 5). Each panel member was tested (b) (4)
The results are summarized in Table 6
and demonstrated acceptable reproducibility for detection of borderline cefoxitin susceptible/resistant S. aureus within and between instruments, reagent lots and days.
Table 5. Strains of borderline cefoxitin susceptible/resistant S. aureus used to evaluate the reproducibility of the cobas vivoDx MRSA test
| Phenotype 1 | Panel Member | Strain 2 | Cefoxitin MIC (µg/mL) | CFU/mL 3 |
|---|---|---|---|---|
| MRSA 4 | (b) (4) | |||
| MSSA 5 |
MRSA: Methicillin Resistant S. aureus; MSSA: Methicillin Susceptible S. aureus; MIC: Minimum Inhibitory Concentration
1 Phenotype based on cefoxitin MIC and mecA status (positive/negative); (D) (4)
2 Obtained from the CDC & FDA Antimicrobial Resistance Isolate Bank
- 3 Observed viable counts ranged from (b) (4) (4) CFU/mL (mean = 1,506 CFU/mL) for MRSA and (b) (4) CFU/mL (mean = 1.4 x 10° CFU/mL) for MSSA
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Table 6. Reproducibility of cobas vivoDx MRSA test performance with borderline cefoxitin susceptible/resistant S. aureus
Image /page/7/Figure/1 description: This image is a table that shows the agreement by panel member and strain. The table has three columns: Factor, MRSA, and MSSA. The rows are Reagent Lot, Instrument, Day, and Overall. The Factor column also includes (b) (4).
MRSA: Methicillin Resistant S. aureus; MSSA: Methicillin Susceptible S. aureus Refer to Table 5 for detailed description of each panel member
1 To help ensure viability, panel members were prepared fresh each day from standardized bacterial suspensions and viable counts were performed to verify their concentration
-
b. Linearity/assay reportable range:
Not applicable. -
c. Traceability, Stability, Expected values (controls, calibrators, or methods):
External Controls
Instructions for preparation of External Positive and Negative Controls using commercially available strains of S. aureus (ATCC 33591 [MRSA] and ATCC 25923 [MSSA]) are provided in the device labeling. Valid results must be obtained with appropriately prepared Positive and Negative External Controls for results from the cobas vivoDx MRSA test to be displayed by the system, although the frequency of control testing is at the discretion of the laboratory based on applicable regulations and accreditation requirements. If either member of a control pair fails, patient test results are withheld until valid External Control results are obtained. If repeated control failure occurs, patient test results are reported as "Invalid."
(b) During the Clinical Study described in Section L(3)(a).(4). pairs of External Quality Control pairs were tested, of which (b) (99.7%) produc lid results. (4)
The following additional studies were also performed to verify the reproducibility of the recommended Quality Control procedures:
Stability of Primary Cultures: Primary culture plates of the recommended External Control strains of S. aureus were (b) (4)
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(b) (4) using two different lots of cobas vivoDx MRSA reagents. All results were as expected (b) (4) Positive and (b) (4) Negative Control results), demonstrating the stability of primary subcultures of the External Control strains when stored at room temperature for up to four days.
Verification of Positive Control Strain: Fresh secondary and tertiary subcultures of MRSA strain ATCC (b) (4) were used to prepare External Positive Controls according to the recommended procedure. The controls were tested usin (b) (4)
All cobas vivoDx MRSA
control and phenotypic test results were as expected (b) (4) in each case). demonstrating the reproducibility of suspensions prepared with ATCC(b) (4) External Positive Controls.
Maintenance of Quality Control Strains: Fresh secondary and tertiary subcultures of the recommended strains were used to prepare Positive and Negative External Controls on each day over a period of (b) (4) The controls were tested using two different lots of cobas MRSA reagents (b) (4)
. All the Negative External Controls produced the expected results (b) (4) of the Positive External Controls, demonstrating the Reproducibility of External Control performance over the period of the study.
Based on the results of these studies, the instructions for use of the cobas vivoDx MRSA test indicate that only fresh secondary and tertiary cultures should be used to prepare external controls. Primary and secondary cultures may be stored a(0) (1) , respectively, prior to subculture.
The instructions for preparation and maintenance of the cobas vivoDx MRSA test External Positive and Negative Controls and associated performance are acceptable.
Specimen Stability
The stability of nasal swab specimens in cobas vivoDx MRSA Collection and Transport Tubes was evaluated by testing simulated samples that were seeded at clinically relevant concentrations with either MRSA, methicillin susceptible S. aureus (MSSA) or Staphylococcus epidermidis and which were then stored for different durations. The results of the study demonstrated that nasal swab specimens for use with the cobas vivoDx MRSA test may be transported and stored in cobas vivoDx MRSA Collection and Transport Tubes for up to 15 hours when held at 15-30°C.
The results of the study also showed that, when specimen processing is delayed and high levels of MSSA are present, there is increased potential for false positive results to occur. This is noted as a Limitation in the device labeling (refer to Section H2).
Reagent Stability
cobas vivoDx MRSA reagent kits comprised of cobas vivoDx MRSA Reagent Caps and Test Tubes, should be stored at 2-8°C. The cobas vivoDx MRSA Collection and Transport Kit and Control Media should be stored at 15-30℃. Stability testing of the
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cobas vivoDx MRSA reagents to establish expiration dating under their respective storage conditions is on-going.
Open Kit Stability
cobas vivoDx MRSA kits containing cobas vivoDx MRSA Reagent Caps and Test Tubes were shown to produce the expected results with simulated samples and controls when stored at 2-8°C for up to 7 days after opening.
- d. Detection Limit:
The Limit of Detection of the cobas vivoDx MRSA test was determined by testing serial dilutions of five different strains of MRSA in simulated nasal matrix (refer to Section L(2)(b)). For each strain, at least 20 replicates were tested at each target level with each of three lots of cobas vivoDx MRSA reagents. The concentration of organisms present in each dilution was confirmed by performing viable counts. The LoD for each strain and reagent lot combination, defined as the lowest target level at which 95% of replicates are expected to produce positive results, was estimated by Probit analysis (Table 7). The highest LoD point estimate was 315 CFU/mL for strain NRS 642 with Reagent Lot #2. This value was used as the basis for establishing appropriate target levels for subsequent Analytical Studies.
| MRSA Strain 1 | PFGE Type | SCCmec Type | Reagent Lot | LoD (CFU/mL) 2 |
|---|---|---|---|---|
| ATCC BAA-44 | Iberian | (b) (4) | (b) (4) | 17 (b) (4) |
| (b) (4) | ||||
| NRS 642 | 100 | (b) (4) | 315 (b) (4) | |
| (b) (4) | ||||
| NRS 651 | 200 | (b) (4) | 108 (b) (4) | |
| (b) (4) | ||||
| NRS 685 | 500 | (b) (4) | 78 (b) (4) | |
| (b) (4) | ||||
| NRS 725 | 300 | (b) (4) | 69 (b) (4) | |
| (b) (4) |
Table 7. Summary of results from the LoD Study for the cobas vivoDx MRSA test
PFGE: Pulsed Field Gel Electrophoresis; SCCmec: Staphylococcal Cassette Chromosome mec Note: The highest predicted LoD for each strain is shaded
1 The cefoxitin MIC of each strain included in the study was >16 ug/mL
2 LoD point estimate with 95% confidence intervals
3 The LoD point estimate for strain NRS 642 with Reagent Lot #2 was used to determine appropriate target levels for subsequent Analytical Studies
4 Confidence intervals are broad due to a non-monotonic increase in hit rate with organism concentration
In addition to the five strains of MRSA shown in Table 7, the analytical sensitivity of the cobas vivoDx MRSA test for the heteroresistant ATCC 43300 strain was also
10
evaluated. The highest LoD point estimate for this strain was 320 (95% confidence interval: 190-772) CFU/mL with Reagent Lot #1.
- e. Analytical Reactivity:
The inclusivity of the cobas vivoDx MRSA test was evaluated in two separate Analytical Reactivity Studies as described below:
Testing of Alternative PFGE and SCCmec Types
A summary of the results of the study is provided in Table 8. On initial testing, the cobas vivoDx MRSA test produced(b) (4)
11
Table 8. Results of the cobas vivoDx MRSA Inclusivity Study, stratified by SCCmec and PFGE type
| SCCmec Type | PFGE Type | Number | Positive 1 | % Positive |
|---|---|---|---|---|
| I | Iberian | (b) (4) | ||
| II | USA 100 | |||
| USA 200 | ||||
| USA 600 | ||||
| III | N/A | |||
| ST239 | ||||
| IV | Iberian | |||
| USA 300 | ||||
| USA 300-114 | ||||
| USA 400 | ||||
| USA 500 | ||||
| USA 700 | ||||
| USA 800 | ||||
| USA 1000 | ||||
| USA 1100 | ||||
| IVa | USA 400 | |||
| WA-MRSA | ||||
| V | USA 700 | |||
| USA 1000 | ||||
| VI | USA 600 | |||
| IX (mecC) | N/A | |||
| Total | ||||
| (4) |
Borderline Cefoxitin Susceptibility Panel
(0)
The second Analytical Reactivity Study included a panel of strains of S. aureus obtained from the CDC & FDA Antimicrobial Resistance Isolate Bank that exhibited cefoxitin MICs which were close to the respective breakpoints for susceptibility and resistance (mecA | Total
Tested | Number of Strains
Positive at Target Level (CFU/mL) 1 | | | |
|------------------------|--------------------------|-------------|-----------------|----------------------------------------------------------|-----------|-----------|-----------|
| Resistant | 8 | Positive | 8 | 1.2 x 103 | 1.2 x 104 | 1.2 x 105 | 1.2 x 106 |
| | 16 | Positive | 10 | 4 | 42 | Not done | Not done |
| | >16 | Positive | 4 | 6 | 1 | Not done | Not done |
| Susceptible | 4 | Positive | 1 | 3 | 0 | 0 | 0 |
| | 4 | Negative | 9 | 0 | 0 | 0 | 03 |
Cefoxitin breakpoints: 8 ug/mL, Resistant
1 Lowest level at which 3/3 replicates reported as positive by the cobas vivoDx MRSA test
2 One strain that was positive at 1.2 x 104 CFU/mL also had 2/3 positive results at 1.2 x 103 CFU/mL
3 One strain produced 2/3 positive results at 1.2 x 106 CFU/mL
f. Analytical Specificity:
Cross-reactivity
e analytical specificity of the cobas vivoDx MRSA test was evaluated by testing (b) (4) strains of bacteria in triplicate at a concentration of (b) (4)
CFU/mL, depending on the strain. The list of species tested is shown in Table 10 and included 30 strains of Methicillin Susceptible S. aureus (MSSA) with cefoxitin MICs (b) (4) as well as species that are phylogenetically related to S. aureus and other organisms that may be found as part of the nasal flora. One yeast species was also tested at a concentration of (b) (4) . No false positive results were obtained with any of the strains of MSSA, although crossreaction was observed with five other organisms, including Listeria monocytogenes. Staphylococcus equorum and three different species of Enterococcus (Table 11). In each case, the expected results were obtained when testing of the cross-reactive species was repeated at a lower concentration. The potential for false positive results with organisms other than MRSA is included as a Limitation in the device labeling (refer to Section H2).
13
(b) (4)
14
| Cross-reactive Species | CFU/mL | ||
|---|---|---|---|
| Cross-reactive | |||
| (b) (4) | Not Cross-reactive1 | No Interference2 | |
| Enterococcus faecium | 3.7 x 105 | 6.1 x 104 | |
| Enterococcus flavescens | 3.6 x 105 | 3.4 x 104 | |
| Enterococcus gallinarum | 1.1 x 105 | 1.1 x 105 | |
| Listeria monocytogenes | 7.0 x 104 | 4.3 x 104 | |
| Staphylococcus equorum | 4.6 x 103 | 3.8 x 103 |
Table 11. Species found to cross-react in the cobas vivoDx MRSA test
1 Highest level tested at which no cross-reaction was observed
2 Highest level tested at which no interference was observed (i.e., no false negative results obtained with low levels of MRSA) [see below]
Microbial Interference
The potential for interference with cobas vivoDx MRSA test was evaluated using the same panel of organisms as for the Cross-reactivity Study described above (Table 10), including those that initially produced false positive results (Table 11). Each species was tested in triplicate at a concentration of between 3.4 x 103 and 2.1 x 10' CFU/mL in the presence of each of two strains of MRSA at approximately 4X LoD (i.e. ~ 1.2 x 102 CFU/mL). No interference in the detection of MRSA strain NRS 725 was observed in the presence of any other any species; however, false negative results were obtained when testing was performed with MRSA strain NRS 642 (Table 12), although in each case, the expected positive results for MRSA were produced when the interfering organism was tested at a lower level. The potential for false negative results in the presence of high levels of the species listed in Table 12 is noted as a Limitation in the device labeling (refer to Section H2).
Table 12. Species that interfered with detection of MRSA strain NRS 642 by cobas vivoDx MRSA test
| Interfering Species | CFU/mL (MRSA positive/tested) | |
|---|---|---|
| Interference 1 | No Interference | |
| Citrobacter koseri | (b) (4) | 1.5 x 106 (3/3) |
| Corynebacterium jeikeium | (b) (4) | 1.9 x 105 (3/3) |
| Enterobacter cloacae | (b) (4) | 1.1 x 106 (3/3) |
| Klebsiella pneumoniae | (b) (4) | 3.0 x 104 (3/3) |
1 As determined by reporting of false negative results for MRSA
2 Highest level tested at which no interference observed (i.e., no false negative results obtained with low levels of MRSA)
For the five species that were found to cross-react in the cobas vivoDx MRSA test when present in high concentration, additional testing was performed to assess the potential for interference with the detection of low levels of MRSA (~1.2 x 102 CFU/mL) when the organisms were at levels below the threshold for cross-reactivity. The expected MRSA positive and negative assay results were obtained when the cross-reactive organisms were tested at the levels indicated in Table 11.
15
Interfering Substances
The potential for interference with the cobas vivoDx MRSA test by endogenous and exogeneous substances that may be present in nasal swab specimens was evaluated. Two strains of MRSA were tested at approximately 4X LoD (b) (4) CFU/mL) in the presence and absence of 24 substances at different concentrations. The highest concentration each substance at which no interference was observed is shown in Table 13. Eight substances did not interfere at any of the concentrations tested. However, false negative results were obtained in the presence of the topical antibacterial agent mupirocin at every concentration that was evaluated. The remaining 15 substances were shown to interfere when present at high concentration but the inhibitory affect was alleviated at lower levels. Because of the potential for interference, subjects with visibly bloody swabs (n = 23) were excluded from the clinical performance evaluation described in Section L(3)(a). The potential for false negative results with the cobas vivoDx MRSA in the presence of mupirocin and blood is noted as a Limitation in the device labeling (refer to Section H2).
| Substance Type | Substance | Highest Quantity at which No
Interference Observed | |
|--------------------------------|------------------------------|-------------------------------------------------------|--------------------------|
| | | % Swab Capacity | Concentration
(mg/mL) |
| Antibacterial/antiviral | Bepanthen 1 | (b) (4) | 40.4 |
| | Mupirocin | | Not applicable 2 |
| | Relenza 1 | | 73.1 |
| | Tobramycin and Dexamethasone | | 4.9 |
| Decongestant/allergy
relief | Afrin | | 39.7 |
| | Afrin Menthol | | 36.1 |
| | Azelastine HCL | | 38.3 |
| | Beconase AQ | | 2.3 |
| | ClearLife 1 | | 74.4 |
| | Dristan | | 34.6 |
| | Flonase | | 38.8 |
| | Flunisolide 1 | | 81.4 |
| | Nasacort | | 38.5 |
| | NasalCrom 1 | | 76.3 |
| | Nasonex | | 39.9 |
| | Neo-Synephrine | | 37.7 |
| | Nostrilla | | 37.5 |
| | Otrivine 1 | | 76.7 |
| | Rhinocort 1 | | 85.0 |
| | Saline Nasal Spray | | 19.4 |
| | Zicam | | 47.5 |
| Endogenous | Bovine Mucin 1,3 | | 77.9 |
| | Human Blood 4 | | 21.3 |
| Topical Analgesic | Anbesol | | 42.2 |
Table 13. Substances evaluated for potential interference with the cobas vivoDx MRSA test
(b) (4)
16
-
g. Assay Cut-off:
The result algorithm for the cobas vivoDx MRSA test uses a series of decision trees to evaluate features of the kinetic luminescent curve obtained after addition of substrate to the test sample. The algorithm was developed using data from known MRSA positive and negative analytical and clinical samples and the cutoff was selected based on Receiver Operating Curve (ROC) analysis to optimize sensitivity and specificity. -
- Comparison studies:
- a. Method comparison with predicate device:
Not applicable.
- b. Matrix comparison:
To provide a sufficient quantity of S. aureus-negative material for testing, a simulated throat matrix comprised of human DNA and porcine mucin was used for most of Analytical Studies with the cobas vivoDx MRSA test. A comparison study was therefore conducted to verify that the device performed similarly in the presence of natural and simulated nasal matrix. The study was performed with two strains of MRSA that were each diluted in both matrices to approximately the same concentrations prior to testing. The results of the study are summarized in Table 14 and demonstrate that the analytical sensitivity of the cobas vivoDx MRSA test for each strain was approximately the same in both types of matrix. Because of this, the use of simulated matrix in Analytical Studies to characterize the performance of the cobas vivoDx MRSA test was determined to be acceptable.
| MRSA
Strain | PFGE
Type | SCCmec
Type | Cefoxitin
MIC (µg/mL) | Target Level
(Multiple of LoD) | Positive/Tested (%) | |
|----------------|--------------|----------------|--------------------------|-----------------------------------|---------------------|-----------|
| | | | | | Natural | Simulated |
| NRS 659 | USA 300 | IV | >16 | (b) (4) | | |
| NRS 663 | USA 100 | II | >16 | (b) (4) | | |
Table 14. Summary of results from comparison of cobas vivoDx MRSA test performance with natural and simulated nasal swab matrix
LoD: Limit of Detection; PFGE: Pulsed Field Gel Electrophoresis; SCCmec: Staphylococcal Cassette Chromosome mec: MIC Minimal Inhibitory Concentration
17
3. Clinical studies:
a. Clinical Sensitivity:
The performance of the cobas vivoDx MRSA test was evaluated in a prospective Clinical Study that was conducted at seven geographically diverse locations in the U.S. Subjects aged >18 years of age who were eligible for MRSA screening according to the policies of each respective institution were enrolled under informed consent. Each subject provided two nasal swabs: one for the cobas vivoDx MRSA test and one for reference culture. The order of collection of the swabs was alternated between patients.
Of 4.198 paired specimens enrolled in the study. 3.963 were considered evaluable. Forty-two (42) pairs of specimens were excluded for the following reasons: bloody swab (23), subject previously enrolled (15) or met predefined exclusion criteria (3). subject identity unknown (1). The results from a further 193 subjects were excluded for failure to obtain valid reference and/or cobas vivoDx MRSA test results (specimen not processed or processing delayed for the reference method (94), vivoDx system failure (28), laboratory error (26), delay in cobas vivoDx MRSA testing (21), absence of cobas vivoDx MRSA controls (11), cobas vivoDx MRSA result invalid (7), insufficient sample (4), specimens not tested or lost (2)).
Of the 3,963 subjects with evaluable results, 182 (4.6%) were positive for MRSA by direct culture on chromogenic medium. A further 28 subjects were positive (b)
(4) (D) (4) resulting in a total of 210 (5.3%) MRSA positive subjects. The identity of suspected colonies of MRSA on(b) (4) was confirmed following (b) (4)
The performance of the cobas vivoDx MRSA test in comparison to direct/enriched reference culture and direct culture alone is shown in Table 15. The performance stratified by study site is shown in Table 16.
18
| Direct & Enriched Culture (reference) | |||||
|---|---|---|---|---|---|
| Positive | Negative | Total | |||
| cobas vivoDx | |||||
| MRSA | Positive | 189 | 512 | 240 | |
| Negative | 211 | 3702 | 3723 | ||
| Total | 210 | 3753 | 3963 | ||
| Sensitivity | 90.0% (189/210); 85.2-93.4% | ||||
| Specificity | 98.6% (3702/3753); 98.2-99.0% | ||||
| Positive Predictive Value | 78.8% (189/240) | ||||
| Negative Predictive Value | 99.4% (3702/3723) | ||||
| Direct Culture alone | |||||
| Positive | Negative | Total | |||
| cobas vivoDx | |||||
| MRSA | Positive | 176 | 643 | 240 | |
| Negative | 6 | 3717 | 3723 | ||
| Total | 182 | 3781 | 3963 | ||
| Positive Percent Agreement | 96.7% (176/182); 93.0-98.5% | ||||
| Negative Percent Agreement | 98.3% (3717/3781); 97.8-98.7% |
Table 15. Performance of the vivoDx MRSA test in comparison to culture
(b) (4)
19
Table 16. Performance of the cobas vivoDx MRSA compared to culture, stratified by study site
| Study Site | Direct & Enriched
Culture (reference) | | Direct Culture Alone | |
|------------|------------------------------------------|------------------------|----------------------|------------------------|
| | Sensitivity | Specificity | PPA | NPA |
| (b) (4) | | | | |
| All Sites | 90.0%
(189/210) 1 | 98.6%
(3702/3753) 2 | 96.7%
(176/182) | 98.3%
(3717/3781) 3 |
PPA: Positive Percent Agreement; NPA: Negative Percent Agreement
1 15/21 subjects with false negative cobas vivoDx MRSA test results were MRSA positive only by enriched culture, suggesting the presence of low level colonization (