The Xpert® Carba-R Assay, performed on the GeneXpert® Instrument Systems, is a qualitative in vitro diagnostic test designed for the detection and differentiation of the blakes, blaypy, blaxx -8, and blang gene sequences associated with carbapenem-non-susceptibility. The test utilizes automated real-time polymerase chain reaction (PCR).

The Xpert Carba-R Assay is intended as an aid to infection of carbapenem-non-susceptible bacteria that colonize patients in healthcare settings. A negative Xpert Carba-R Assay result does not preclude the presence of other resistance mechanisms.

The Xpert Carba R-Assay is for use with the following sample types:

Pure Colonies
The assay is performed on carbapentible pure colonies of Enterobacteriaceae. Acinetobacter baumannii. or Pseudomonas aeruginosa, when grown on blood agar or MacConkey agar. For testing pure colonies, the Xpert Carba-R Assay should be used in conjunction with other laboratory tests including phenotypic antimicrobial susceptibility testing.

The identification of a blawy, blayin metallo-beta-lactamase gene (i.e., the genes that encode the INP, NDM, and VIM metallo-beta-lactamases, respectively) may be used as an aid to clinicians in determining appropriate therapeutic strategies for patients with known or suspected carbapenem-non- susceptible bacterial infections.

Rectal and Perirectal Swab Specimens
The assay is performed on rectal swab specimens from patients at risk for intestinal colonization with carbapenem-non-susceptible bacteria. Concomitant cultures are necessary to recover organisms for epidemiological typing, antimicrobial susceptibility testing, and for further confirmatory bacterial identification.

The Xpert Carba-R Assay, when performed on rectal and perirectal swab specimens, is not intended to guide or monitor treatment for carbapenem-non-susceptible bacterial infections or to determine infection from carbapenem-nonsusceptible bacteria.

Device Description

The Xpert Carba-R Assay is an automated real-time polymerase chain reaction (PCR) in vitro diagnostic test for qualitative detection of the blakpo, blaymy, blavny, blaoxa-as, and blamp gene sequences from rectal or perirectal swab specimens or isolates of pure cultures of carbapenem-non-susceptibility gram-negative bacteria. The Xpert Carba-R Assay is intended as an aid for infection control for monitoring the spread of carbapenem-non-susceptible organisms in healthcare settings.

The Xpert Carba-R Assay is performed on the Cepheid GeneXpert® Instrument Systems (GeneXpert Dx, GeneXpert Infinity-48, GeneXpert Infinity-48s, and GeneXpert Infinity-80 systems). The GeneXpert Instrument System platform automates sample preparation, amplification and real-time detection.

The GeneXpert Instrument Systems require the use of single-use, disposable cartridges (the Xpert Carba-R cartridges) that hold the PCR reagents and host the PCR process. Because the cartridges are self-contained and specimens never come into contact with working parts of the instrument modules, cross-contamination between samples is minimized.

The Xpert Carba-R Assay cartridges contain reagents for the detection of blaxpy blayDM blaym, blacks, and blandp gene sequences. A Sample Processing Control (SPC) and a Probe Check Control (PCC) are controls utilized by the GeneXpert Instrument System platform. The SPC is present to control for adequate processing of the target bacteria and to monitor the presence of inhibitors in the real-time PCR reaction to reduce the possibility of false negative results. The PCC verifies reagent rehydration, real-time PCR tube filling in the cartridge, probe integrity, and dye stability.

The single-use, multi-chambered fluidic cartridges are designed to complete sample preparation and real-time PCR for the detection of the blakpc, blaNDM, blagxA-48 and blamp gene sequences from rectal or perirectal swab specimens or isolates of pure cultures of carbapenem-non- susceptibility gram-negative bacteria in approximately 50 minutes. The GeneXpert Instrument Systems, comprised of the GeneXpert Dx Systems and the GeneXpert Infinity Systems, have 1 to 80 randomly accessible modules, depending upon the instrument, that are each capable of performing separate sample processing and real-time PCR and RT- PCR tests. Each module contains a syringe drive for dispensing fluids (i.e., the syringe drive activates the plunger that works in concert with the rotary valve in the cartridge to move fluids between chambers), an ultrasonic horn for lysing cells or spores, and a proprietary I-CORE® thermocycler for performing real-time PCR and RT-PCR and detection.

Rectal or perirectal swab specimens or bacterial isolates from culture are placed into a sample reagent. The sample is transferred to the sample chamber of the disposable fluidic cartridge (the Xpert Carba-R cartridge). The user initiates a test from the system user interface and places the cartridge into the GeneXpert instrument platform, which performs hands-off real-time, multiplex PCR for detection of the blakers, blayDM blaym, blacks, and blamp gene sequences. The results are automatically generated at the end of the process in a report that can be viewed and printed.

AI/ML Overview

Here's an analysis of the provided text, focusing on the acceptance criteria and the study that proves the device meets them, structured according to your request.

Device Name: Xpert® Carba-R Assay

1. Acceptance Criteria and Reported Device Performance

The document doesn't explicitly state "acceptance criteria" in a numbered or bulleted list with corresponding performance numbers for each criterion, as would be ideal in a formal acceptance criteria table. However, based on the non-clinical and clinical studies presented, we can infer the acceptance criteria and the device's performance against them, particularly focusing on the clinical performance as that's often tied to regulatory acceptance.

The primary measure of clinical performance for this diagnostic device is Positive Percent Agreement (PPA) and Negative Percent Agreement (NPA) against a reference method. Analytical criteria include Limit of Detection (LoD), inclusivity (analytical reactivity), and analytical specificity (cross-reactivity).

Inferred Acceptance Criteria and Reported Device Performance (with direct performance data as provided):

Criterion Type	Specific Criterion	Acceptance Threshold (Inferred)	Reported Device Performance (Test Set)	Basis for Performance (Study type, section)
Clinical Performance	PPA vs. Reference Method (Prospective)	High (e.g., >95%)	IMP: N/A (0 TP)VIM: N/A (0 TP)NDM: 100% (95% CI: 20.7-100)KPC: 100% (95% CI: 34.2-100)OXA-48: 100% (95% CI: 20.7-100)	Clinical Studies - Prospective Perirectal Specimens (Table 8-11)
	NPA vs. Reference Method (Prospective)	High (e.g., >95%)	IMP: 100% (95% CI: 99.6-100)VIM: 100% (95% CI: 99.6-100)NDM: 100% (95% CI: 99.6-100)KPC: 99.6% (95% CI: 98.9-99.8)OXA-48: 99.9% (95% CI: 99.4-100)	Clinical Studies - Prospective Perirectal Specimens (Table 8-11)
	PPA vs. Reference Method (Contrived)	100%	IMP: 100% (95% CI: 95.4-100)VIM: 100% (95% CI: 95.5-100)NDM: 100% (95% CI: 95.4-100)KPC: 100% (95% CI: 95.4-100)OXA-48: 100% (95% CI: 95.4-100)	Clinical Studies - Contrived Specimen Results (Table 8-14)
	NPA vs. Reference Method (Contrived)	100%	IMP: 100% (95% CI: 98.9-100)VIM: 100% (95% CI: 98.9-100)NDM: 100% (95% CI: 98.9-100)KPC: 100% (95% CI: 98.9-100)OXA-48: 100% (95% CI: 98.9-100)	Clinical Studies - Contrived Specimen Results (Table 8-14)
	Perirectal vs. Rectal Swab Equivalence (PPA)	High (e.g., >90%)	94.7% (95% CI: 75.4-99.1)	Clinical Studies - Perirectal Swab and Rectal Swab Equivalence Study (Table 8-15)
	Perirectal vs. Rectal Swab Equivalence (NPA)	High (e.g., >90%)	97.8% (95% CI: 94.5-99.1)	Clinical Studies - Perirectal Swab and Rectal Swab Equivalence Study (Table 8-15)
Analytical Studies	Limit of Detection (LoD)	Low CFU/swab	See Table 8-2 (ranges from 56 to 1303 CFU/swab depending on target)	Non-Clinical Studies - Analytical Sensitivity (Table 8-2)
	Analytical Reactivity (Inclusivity)	Detection of broad panel	69 of 72 carbapenemase-producing bacterial strains detected (IMP-4 detected at higher conc, 3 strains not detected)	Non-Clinical Studies - Analytical Reactivity (Table 8-3)
	Analytical Specificity (Cross-reactivity)	100% Specificity (no cross-react.)	100% (No cross-reaction with 94 potentially cross-reactive organisms/nucleic acids)	Non-Clinical Studies - Analytical Specificity
	Reproducibility	High Agreement (e.g., >90%)	Overall % Agreement by Sample ranges from 92.4% (VIM Low Pos) to 100% (Neg, Mod Pos, NDM Low Pos)	Non-Clinical Studies - Reproducibility Study (Table 8-16)
	Carry-Over Contamination	No detectable contamination	All 50 positive samples correctly reported DETECTED; all 52 negative samples correctly reported NOT DETECTED.	Non-Clinical Studies - Carry-Over Contamination
	Competitive Interference	No significant inhibition	No inhibitory effect observed at 4x LoD for NDM and IMP. Inhibition observed at original low concentration (2X LoD) for NDM and IMP.	Non-Clinical Studies - Competitive Interference
	Potentially Interfering Substances	No significant interference	22 of 24 substances correctly identified. Interference with Barium Sulfate (>0.1% w/v) and Pepto-Bismol (>0.025% w/v) observed.	Non-Clinical Studies - Potentially Interfering Substances

(Note: "N/A" for PPA signifies no True Positives were observed in the prospective study for IMP and VIM, which is reflected in the CI. The acceptance threshold is inferred based on typical diagnostic performance expectations for such devices.)

2. Sample Size and Data Provenance

Test Set Sample Sizes:
- Prospective Clinical Study (Perirectal Swab Specimens): 924 eligible specimens (from an initial 963 enrolled).
- Contrived Clinical Study (Perirectal Swab Matrix): 432 specimens.
- Perirectal vs. Rectal Swab Equivalence Study: 201 eligible specimens (from an initial 207 enrolled).
- Analytical Sensitivity (LoD): 20 replicates for each target (10 carbapenemase-producing organisms) at each concentration level, tested over 4 days, then 10 verification replicates at each estimated LoD.
- Analytical Reactivity (Inclusivity): 72 samples, tested in triplicate at approximately 3X LoD.
- Analytical Specificity (Cross-reactivity): 62 carbapenem-susceptible/non-susceptible bacterial strains, 24 commensal bacterial strains/enteric microorganisms, human cells. Tested in triplicate.
- Carry-Over Contamination: 102 tests (50 high positive, 52 negative).
- Competitive Interference: Replicates of eight for each combination.
- Potentially Interfering Substances: 8 replicates of positive samples and 8 replicates of negative samples tested per substance (24 substances).
- Reproducibility: 11 samples x 4 replicates/day x 6 days x 2 operators x 3 sites = 1584 total tests (though 144 replicates per sample type for analysis).
Data Provenance:
- Prospective Clinical Study: "Five geographically diverse sites" collected specimens. This implies a multi-center study, likely in the US, given the FDA submission. The study involved patient samples ("subjects who were hospitalized or in a long-term care facility"). This is prospective data collection.
- Contrived Clinical Study: Prepared in perirectal matrix, likely laboratory-based data using spiked samples. This is non-clinical/laboratory data.
- Perirectal vs. Rectal Swab Equivalence Study: "one site enrolling fresh prospectively collected rectal and perirectal swab specimens from consented subjects who were hospitalized in-patients." This is prospective data collection, from a single center.
- Other non-clinical studies (Analytical, Reproducibility, etc.) are laboratory-based and generated data, not patient-derived.

3. Number of Experts and Qualifications (for Ground Truth)

The document does not explicitly state the number of experts or their specific qualifications (e.g., "radiologist with 10 years of experience") used for establishing the ground truth.

For the clinical studies, the reference method involved:

Culturing the MacConkey enrichment broth.
Screening for carbapenem-non-susceptible organisms.
Confirmation by disk diffusion method (per CLSI document M02 and M100).
DNA extraction, quantification, and amplification with specific primers.
Confirmation of amplicon size on Agilent 2100 Bioanalyzer.
For positive cases, sending the amplicon to an independent laboratory for reference bi-directional sequencing analysis, which was validated for detection of the five targets.

While "experts" are implicitly involved in performing and interpreting these complex laboratory procedures, their specific number, roles, and detailed qualifications are not detailed.

4. Adjudication Method for the Test Set

The document mentions discordant analysis for the prospective clinical study.

"For specimens with discordant results (the Xpert Carba-R Assay was positive for a target gene but a carbapenem-non-susceptible organism was not isolated by reference culture), discordant analysis was performed using bi-directional sequencing on DNA extracted directly from the MacConkey enrichment broth."
This indicates a process of re-evaluation of discrepancies, but it's not a formal multi-reader, consensus-based adjudication process for initial ground truth establishment. It's a method for clarifying "false positive" results by seeking further molecular evidence in the original sample. No mention of 2+1 or 3+1 for human readers or direct adjudication of AI outputs is present.

5. Multi-Reader Multi-Case (MRMC) Comparative Effectiveness Study

No, a Multi-Reader Multi-Case (MRMC) comparative effectiveness study was not done. This device is a diagnostic assay (molecular test), not an AI system assisting human readers with interpreting images or other data. Therefore, the concept of "how much human readers improve with AI vs without AI assistance" is not applicable here.

6. Standalone Performance

Yes, the studies primarily evaluate the standalone performance of the Xpert Carba-R Assay. The device is a "qualitative in vitro diagnostic test designed for the detection and differentiation of gene sequences." Its performance metrics (PPA, NPA, LoD, specificity, reproducibility) are reported for the device itself, not in conjunction with human interpretation of its results, beyond the clinical correlation implied by its "aid to infection control" indication. It's an "algorithm only" in the sense that it provides a direct qualitative result (DETECTED/NOT DETECTED) based on its PCR analysis.

7. Type of Ground Truth Used

The ground truth for the clinical studies (prospective and contrived) was established by a reference method combination of:

Culture: MacConkey enrichment broth culture followed by disk diffusion method (per CLSI M02/M100) to confirm carbapenem non-susceptibility.
DNA Sequencing: Bi-directional DNA sequence analysis of purified and amplified DNA from isolated colonies or, in discordant cases, directly from the MacConkey enrichment broth.

For the analytical studies, the ground truth was established by precise laboratory preparations (e.g., known concentrations of bacterial strains for LoD, well-characterized strains for reactivity and specificity).

8. Sample Size for the Training Set

The document describes premarket clinical trials and analytical studies for testing and validation. There is no mention of a separate "training set" or "training data" in the context of machine learning or AI models. This device is a molecular diagnostic assay (PCR-based), not an AI algorithm that undergoes data-driven training. The development of such a device involves assay design, optimization, and then extensive analytical and clinical validation, but not "training" in the AI sense.

9. How the Ground Truth for the Training Set was Established

As noted above, there is no mention of a "training set" for an AI model. For the development and validation of the PCR assay itself, ground truth for optimization and development would implicitly have been established through a combination of:

Known Reference Strains: Using well-characterized bacterial strains with confirmed genetic markers.
Synthetic Oligonucleotides: Designing and testing probes and primers against known sequences.
Internal Laboratory Experiments: Repeated testing and refinement to achieve desired assay performance characteristics before formal validation studies.

These are common practices in the development of molecular diagnostic assays, but they are not described as AI-style "training" with a dedicated "training set."

Summary

{0}------------------------------------------------

Image /page/0/Picture/0 description: The image contains the logos of the Department of Health & Human Services and the Food and Drug Administration (FDA). The Department of Health & Human Services logo is on the left, and the FDA logo is on the right. The FDA logo is a blue square with the letters "FDA" in white, followed by the words "U.S. FOOD & DRUG ADMINISTRATION" in blue.

January 9, 2018

Cepheid Jim Kelly Executive Director, Regulatory Affairs 904 Caribbean Drive Sunnyvale, California 94089

Re: K173263

Trade/Device Name: Xpert Carba-R Regulation Number: 21 CFR 866.1640 Regulation Name: Antimicrobial susceptibility test powder Regulatory Class: Class II Product Code: POC, PMY, OOI Dated: October 10, 2017 Received: October 12, 2017

Dear Jim Kelly:

We have reviewed your Section 510(k) premarket notification of intent to market the device referenced above and have determined the device is substantially equivalent (for the indications for use stated in the enclosure) to legally marketed predicate devices marketed in interstate commerce prior to May 28, 1976, the enactment date of the Medical Device Amendments, or to devices that have been reclassified in accordance with the provisions of the Federal Food, Drug, and Cosmetic Act (Act) that do not require approval of a premarket approval application (PMA). You may, therefore, market the device, subject to the general controls provisions of the Act. The general controls provisions of the Act include requirements for annual registration, listing of devices, good manufacturing practice, labeling, and prohibitions against misbranding and adulteration. Please note: CDRH does not evaluate information related to contract liability warranties. We remind you, however, that device labeling must be truthful and not misleading.

If your device is classified (see above) into either class II (Special Controls) or class III (PMA), it may be subject to additional controls. Existing major regulations affecting your device can be found in the Code of Federal Regulations, Title 21, Parts 800 to 898. In addition, FDA may publish further announcements concerning your device in the Federal Register.

Please be advised that FDA's issuance of a substantial equivalence determination does not mean that FDA has made a determination that your device complies with other requirements of the Act or any Federal statutes and regulations administered by other Federal agencies. You must comply with all the Act's requirements, including, but not limited to: registration and listing (21 CFR Part 807); labeling (21 CFR Part 801 and Part 809); medical device reporting of medical device-related adverse events) (21 CFR 803); good manufacturing practice requirements as set forth in the quality systems (QS) regulation (21 CFR Part 820); and if applicable, the electronic product radiation control provisions (Sections 531-542 of the Act); 21 CFR 1000-1050.

{1}------------------------------------------------

K173263

Also, please note the regulation entitled, "Misbranding by reference to premarket notification" (21 CFR Part 807.97). For questions regarding the reporting of adverse events under the MDR regulation (21 CFR Part 803), please go to http://www.fda.gov/MedicalDevices/Safety/ReportaProblem/default.htm for the CDRH's Office of Surveillance and Biometrics/Division of Postmarket Surveillance.

For comprehensive regulatory information about medical devices and radiation-emitting products, including information about labeling regulations, please see Device Advice (https://www.fda.gov/MedicalDevices/DeviceRegulationandGuidance/) and CDRH Learn (http://www.fda.gov/Training/CDRHLearn). Additionally, you may contact the Division of Industry and Consumer Education (DICE) to ask a question about a specific regulatory topic. See the DICE website (http://www.fda.gov/DICE) for more information or contact DICE by email (DICE@fda.hhs.gov) or phone (1-800-638-2041 or 301-796-7100).

Sincerely,

Ribhi Shawar -S

Uwe Scherf, M.Sc., Ph.D. Director Division of Microbiology Devices Office of In Vitro Diagnostics and Radiological Health Center for Devices and Radiological Health

Enclosure

{2}------------------------------------------------

Indications for Use

510(k) Number (if known) K173263

Device Name

Xpert Carba-R Assay

Indications for Use (Describe)

The Xpert Carba R-Assay is for use with the following sample types:

Pure Colonies

The assay is performed on carbapentible pure colonies of Enterobacteriaceae. Acinetobacter baumannii. or Pseudomonas aeruginosa, when grown on blood agar or MacConkey agar. For testing pure colonies, the Xpert Carba-R Assay should be used in conjunction with other laboratory tests including phenotypic antimicrobial susceptibility testing.

Rectal and Perirectal Swab Specimens

The assay is performed on rectal swab specimens from patients at risk for intestinal colonization with carbapenem-non-susceptible bacteria. Concomitant cultures are necessary to recover organisms for epidemiological typing, antimicrobial susceptibility testing, and for further confirmatory bacterial identification.

Type of Use (Select one or both, as applicable)
X Prescription Use (Part 21 CFR 801 Subpart D)	Over-The-Counter Use (21 CFR 801 Subpart C)

CONTINUE ON A SEPARATE PAGE IF NEEDED.

This section applies only to requirements of the Paperwork Reduction Act of 1995.

*DO NOT SEND YOUR COMPLETED FORM TO THE PRA STAFF EMAIL ADDRESS BELOW."

The burden time for this collection of information is estimated to average 79 hours per response, including the time to review instructions, search existing data sources, gather and maintain the data needed and review the collection of information. Send comments regarding this burden estimate or any other aspect of this information collection, including suggestions for reducing this burden, to:

Department of Health and Human Services Food and Drug Administration Office of Chief Information Officer Paperwork Reduction Act (PRA) Staff PRAStaff(@fda.hhs.gov

"An agency may not conduct or sponsor, and a person is not required to respond to, a collection of information unless it displays a currently valid OMB number."

{3}------------------------------------------------

8.0 510(k) Summary

As required by 21 CFR Section 807.92(c).

Submitted by:	Cepheid904 Caribbean DriveSunnyvale, CA 90489Phone number: (408) 745-4183Fax number: (408) 744-1479
Contact:	Jim Kelly, Ph.D.
Date of Preparation:	January 04, 2018
Device:
Trade name:	Xpert® Carba-R
Common name:	Xpert Carba-R Assay
Type of Test:	Qualitative nucleic acid amplification test of the blaKPCblaNDM, blaVIM, blaOXA-48, and blaIMP gene sequencesassociated with carbapenem-non-susceptibility in gram-negative bacteria obtained from rectal swab specimens,perirectal swab specimens, and bacterial isolates
Classification:Regulation number:Classification name:Product code:	II866.1640Antimicrobial susceptibility test powderPOC, OOI
ClassificationAdvisory Panel	Microbiology (83)
Prescription Use	Yes
Predicate DeviceAssay:	Cepheid Xpert® Carba-R[510(k) #K160901]

{4}------------------------------------------------

Device Description:

{5}------------------------------------------------

Device Intended Use:

The Xpert® Carba-R Assay, performed on the GeneXpert® Instrument Systems, is a qualitative in vitro diagnostic test designed for the detection and differentiation of the blaker, blaNDM, blaoxs-48, and blaMp gene sequences associated with carbapenemnon-susceptibility. The test utilizes automated real-time polymerase chain reaction (PCR).

The Xpert Carba-R Assay is intended as an aid to infection control in the detection of carbapenem-non-susceptible bacteria that colonize patients in healthcare settings. A negative Xpert Carba-R Assay result does not preclude the presence of other resistance mechanisms.

The Xpert Carba R-Assay is for use with the following sample types:

Pure Colonies

The assay is performed on carbapenem-non-susceptible pure colonies of Enterobacteriaceae, Acinetobacter baumannii, or Pseudomonas aeruginosa, when grown on blood agar or MacConkey agar. For testing pure colonies, the Xpert Carba-R Assay should be used in conjunction with other laboratory tests including phenotypic antimicrobial susceptibility testing.

The identification of a blanp, blaNDM, or blaviM metallo-beta-lactamase gene (i.e., the genes that encode the IMP, NDM, and VIM metallo-beta-lactamases, respectively) may be used as an aid to clinicians in determining appropriate therapeutic strategies for patients with known or suspected carbapenem-non- susceptible bacterial infections.

Rectal and Perirectal Swab Specimens

The assay is performed on rectal and perirectal swab specimens from patients at risk for intestinal colonization with carbapenem-non-susceptible bacteria. Concomitant cultures are necessary to recover organisms for epidemiological typing, antimicrobial susceptibility testing, and for further confirmatory bacterial identification.

Substantial Equivalence:

The Cepheid Xpert Carba-R Assay is substantially equivalent to the Xpert® Carba-R Assay, 510(k) #K160901. Both assays utilize the same GeneXpert cartridge and detect target gene sequences using real-time PCR amplification and fluorogenic target-specific hybridization detection. The performance of the Xpert Carba-R Assay for an expanded indication was determined in a multi-site clinical study in which the performance of the Xpert Carba-R Assay was evaluated relative to culture and reference DNA sequence analysis and in a second separate clinical study using rectal and perirectal swab specimens to demonstrate the equivalency of both sample types. The results of the study demonstrated that the performance of the Xpert Carba-R Assay is substantially equivalent to the predicate device. Please refer to K160901 for information on the

{6}------------------------------------------------

performance of the Xpert Carba-R Assay with rectal swab specimens.

Table 8-1 shows the similarities and differences between the Xpert Carba-R Assay and the predicate device.

Similarities
Item	Device	Predicate Device
	Cepheid Xpert Carba-R Assay	Cepheid Xpert Carba-R AssayK160901
General Intended Use	The Xpert® Carba-R Assay, performedon the GeneXpert® Instrument Systems,is a qualitative in vitro diagnostic testdesigned for the detection anddifferentiation of the blaKPC, blaNDM,blaVIM, blaOXA-48, and blaIMP genesequences associated with carbapenem-non-susceptibility. The test utilizesautomated real-time polymerase chainreaction (PCR).The Xpert Carba-R Assay is intended asan aid to infection control in thedetection of carbapenem-non-susceptible bacteria that colonizepatients in healthcare settings. Anegative Xpert Carba-R Assay resultdoes not preclude the presence of otherresistance mechanisms.	The Xpert® Carba-R Assay, performedon the GeneXpert® Instrument Systems,is a qualitative in vitro diagnostic testdesigned for the detection anddifferentiation of the blaKPC, blaNDM,blaVIM, blaOXA-48, and blaIMP genesequences associated with carbapenem-non-susceptibility. The test utilizesautomated real-time polymerase chainreaction (PCR).The Xpert Carba-R Assay is intended asan aid to infection control in thedetection of carbapenem-non-susceptible bacteria that colonizepatients in healthcare settings. The XpertCarba-R Assay is not intended to guideor monitor treatment for carbapenem-non-susceptible bacterial infections. Anegative Xpert Carba-R Assay resultdoes not preclude the presence of otherresistance mechanisms.
	The Xpert Carba R-Assay is for use withthe following sample types:Pure ColoniesThe assay is performed on carbapenem-non-susceptible pure colonies ofEnterobacteriaceae, Acinetobacterbaumannii, or Pseudomonasaeruginosa , when grown on blood agaror MacConkey agar. For testing purecolonies, the Xpert Carba-R Assayshould be used in conjunction with otherlaboratory tests including phenotypicantimicrobial susceptibility testing.The identification of a blaIMP blaNDMor blaVIM metallo-beta-lactamase gene(i.e., the genes that encode the IMP,NDM, and VIM metallo-beta-	The Xpert Carba R-Assay is for use withthe following sample types:Pure ColoniesThe assay is performed on carbapenem-non-susceptible pure colonies ofEnterobacteriaceae, Acinetobacterbaumannii, or Pseudomonasaeruginosa , when grown on blood agaror MacConkey agar. For testing purecolonies, the Xpert Carba-R Assayshould be used in conjunction with otherlaboratory tests including phenotypicantimicrobial susceptibility testing.
Similarities
Item	Device	Predicate Device
	Cepheid Xpert Carba-R Assay	Cepheid Xpert Carba-R AssayK160901
	as an aid to clinicians in determiningappropriate therapeutic strategies forpatients with known or suspectedcarbapenem-non- susceptible bacterialinfections.
	Rectal and Perirectal Swab SpecimensThe assay is performed on rectal andperirectal swab specimens from patientsat risk for intestinal colonization withcarbapenem-non-susceptible bacteria.Concomitant cultures are necessary torecover organisms for epidemiologicaltyping, antimicrobial susceptibilitytesting, and for further confirmatorybacterial identification.The Xpert Carba-R Assay, whenperformed on rectal and perirectal swabspecimens, is not intended to guide ormonitor treatment for carbapenem-non-susceptible bacterial infections or todetermine infection from carbapenem-non-susceptible bacteria.	Rectal Swab SpecimensThe assay is performed on rectal swabspecimens from patients at risk forintestinal colonization with carbapenem-non-susceptible bacteria.Concomitant cultures are necessary torecover organisms for epidemiologicaltyping, antimicrobial susceptibilitytesting, and for further confirmatorybacterial identification.
Type of test	Same	Qualitative
TechnologicalPrinciples	Same	Fully-automated nucleic acidamplification (DNA); real-timePCR
Test Cartridge	Same	Disposable single-use, multi-chamberedfluidic cartridge
Probes	Same	TaqMan® Probes
Controls	Same	Internal sample processingcontrol (SPC) and probecheck control (PCC)External controls available
Instrument System	Same	GeneXpert Instrument System(includes GeneXpert Dx, Infinity-48,Infinity-48s, and Infinity-80)
Time to obtain testresults	Same	Approximately 50 minutes to results
Interpretation of testresults	Same	Diagnostic software of the GeneXpertInstrument System
Similarities
Item	Device	Predicate Device
Laboratory Users	Same	Operators in CLIA Moderate or HighComplexity labs
Differences
Item	New Device	Predicate Device
Sample Types	Bacterial isolates from culture, rectalswab and perirectal swab specimens	Bacterial isolates from culture andrectal swab specimens
	Cepheid Xpert Carba-R Assay	Cepheid Xpert Carba-R AssayK160901

Table 8-1: Comparison of Similarities and Differences of the Xpert Carba-R Assay with the Predicate Device

{7}------------------------------------------------

{8}------------------------------------------------

The Xpert Carba-R Assay has the same general intended use as the predicate device and has the same technological characteristics as the predicate device. The differences between the Xpert Carba-R Assay and the predicate device do not raise questions of safety and effectiveness. The clinical study demonstrates that the Xpert Carba-R Assay is acceptable for its intended use with inexperienced laboratory users and is substantially equivalent to the predicate device described above.

Non-Clinical Studies:

Analytical Sensitivity (Limit of Detection) -Perirectal Swabs

The analytical sensitivity or Limit of Detection (LoD) of the Xpert Carba-R Assay was assessed using carbapenemase-producing organisms seeded into pooled negative human perirectal swab matrix. The LoD was determined for two carbapenemase-producing bacteria for each gene analyte, i.e., the genes encoding KPC, NDM, VIM, OXA-48, and IMP. Bacteria were titered by plate counts and spiked onto clean swabs. Swabs were placed into pooled negative perirectal swab matrix and replicates of 20 were evaluated at a minimum of five different concentrations over four days. The LoD for each of the ten carbapenemase-producing organisms was estimated by probit analysis. The LoD is defined as the lowest concentration of target cells (CFU/swab) that can be reproducibly distinguished from negative samples with 95% confidence. The study was performed with two different lots of Xpert Carba-R reagents and the claimed LoD is the higher of the two determinations. The estimated LoDs were verified by preparing and testing 10 replicates from two independent dilutions of each bacterium at each estimated LoD.

The claimed LoD for each pair of carbapenemase-producing organism in perirectal swab matrix are shown in Table 8-2.

{9}------------------------------------------------

Target Gene and Organism	LoD Estimates (Probit) CFU/swab		LOD Claim CFU/swab	Estimated LoD In Sample Reagent CFU/mL	Verification (Positives/20)
	Lot 1	Lot 2
IMP-1 Acinetobacter baumannii	90	118	118	24	19/20
IMP-1 Klebsiella pneumoniae	269	635	635	127	20/20
VIM-1 Klebsiella pneumoniae	901	514	901	180	20/20
VIM-4 Escherichia coli	446	403	446	89	20/20
NDM-1 Klebsiella pneumoniae ATCC BAA-2146	133	113	133	27	20/20
NDM Klebsiella pneumoniae	56	54	56	11	20/20
KPC-3 Klebsiella pneumoniae NCTC 13438	358	292	358	72	20/20
KPC Enterobacter cloacae	1259	1303	1303	261	20/20
OXA-48 Enterobacter cloacae	223	166	223	45	20/20
OXA-48 Escherichia coli	126	137	137	27	20/20

Table 8-2: LoD Estimates and Verification for Organisms Harboring anenemase Genes using the Xpert Carba-R Assay in Perirectal Swah Matriy

Analytical Reactivity (Inclusivity)

The analytical reactivity of the Xpert Carba-R Assay with perirectal swab matrices was evaluated by testing a panel of 72 samples. This panel consisted of 11 blakec (KPC), 11 blavm (VIM), 8 blaoxa-48 (OXA-48), 5 blayDM/blaoxA-181 (NDM/OXA-181), 6 blaoxA-181(OXA-181), 17 blamp (IMP), and one blaxpc/blavIM (KPC/VIM) well-characterized bacterial strains. The strains tested in perirectal swab matrix and their test concentrations are presented in Table 8-3.

For testing in perirectal swab matrix, organisms were seeded into pooled negative perirectal swab matrix. All bacterial strains were tested in triplicate at approximately 3X LoD. Xpert Carba-R Assay target genes were detected in 69 of 72 carbapenemaseproducing bacterial strains although IMP-4 was detected only using a higher concentration (Table 8-3). Xpert Carba-R Assay target DNA sequences were not detected in three bacterial strains as shown in Table 8-3. In one of the three bacterial strains, the IMP-13 gene was not detected by the assay, although it was predicted to be detected by in silico analysis. In two of the other three bacterial strains, the IMP-7 and IMP-14 genes were not predicted to be detected by in silico analysis and were not detected by the assay. See Limitations in the package insert.

The variants detected, and predictions for detecting other subtypes of each resistance gene based on in silico analysis, are presented in Table 8-4.

{10}------------------------------------------------

Strain ID	Organism	Resistance Marker with Variant Information	Concentration Tested in Perirectal Swab Matrix (CFU/mL)
NCTC 13438	Klebsiella pneumoniae	KPC-3	153
31551	Klebsiella pneumoniae	KPC-4	50
ATCC BAA-1705	Klebsiella pneumoniae	KPC-2	130
PA-Col	Pseudomonas aeruginosa	KPC-2	250
KBM18	Enterobacter aerogenes	KPC-2	250
BM9	Klebsiella pneumoniae	KPC-3	330
PA3	Klebsiella pneumoniae	KPC-2	100
CGNC	Serratia marcescens	KPC-2	300
CFVL	Enterobacter cloacae	KPC-2	160
COL	Escherichia coli	KPC-2	147
GR-04/KP-69	Klebsiella pneumoniae	KPC-2, VIM	80
164-3	Klebsiella oxytoca	KPC	70
NCTC 13437	Pseudomonas aeruginosa	VIM-10	500
NCTC 13439	Klebsiella pneumoniae	VIM-1	130
NCTC 13440	Klebsiella pneumoniae	VIM-1	70
758	Pseudomonas aeruginosa	VIM	250
PA-87	Klebsiella pneumoniae	VIM	200
B92A	Pseudomonas aeruginosa	VIM	2000
Coll	Pseudomonas aeruginosa	VIM-2	500
BM19	Serratia marcescens	VIM-2	250
KOW7	Escherichia coli	VIM-4	250
DIH	Klebsiella pneumoniae	VIM-19	250
MSH2014-3	Enterobacter cloacae	VIM	500
NCTC 13443	Klebsiella pneumoniae	NDM-1	80
ATCC BAA-2146	Klebsiella pneumoniae	NDM-1	80
34262	Klebsiella pneumoniae	NDM	80
GEN	Acinetobacter baumannii	NDM-1	130
3047	Enterobacter cloacae	NDM-1	70
7892	Proteus mirabilis	NDM-1	30
CAN	Salmonella spp.	NDM-1	70
EGY	Acinetobacter baumannii	NDM-2	40
15	Escherichia coli	NDM-4	30
405	Escherichia coli	NDM-5	30
CF-ABE	Citrobacter freundii	NDM	30
73999	Pseudomonas aeruginosa	NDM	50
39365	Providencia rettgeri	NDM-1	70
NCTC 13442	Klebsiella pneumoniae	OXA-48	40
OM11	Klebsiella pneumoniae	OXA-48	60
501	Enterobacter cloacae	OXA-48	80
Strain ID	Organism	ResistanceMarker withVariantInformation	ConcentrationTested inPerirectal SwabMatrix(CFU/mL)
DUW	Klebsiella pneumoniae	OXA-48	120
OM22	Escherichia coli	OXA-48	80
BOU	Enterobacter cloacae	OXA-48	80
TUR	Enterobacter cloacae	OXA-48	120
11670	Escherichia coli	OXA-48	100
166643	Klebsiella pneumoniae	OXA-181	20
42194	Klebsiella pneumoniae	OXA-181	20
MSH2014-64	Klebsiella pneumoniae	OXA-181	280
MSH2014-72	Escherichia coli	OXA-181	100
74	Escherichia coli	OXA-181	100
CDC0051	Klebsiella ozaenae a	OXA-181	250
B108A	Klebsiella pneumoniae	NDM, OXA-181	10
C10192-DISCS	Enterobacter aerogenes	NDM, OXA-181	10
KP-OMA3	Klebsiella pneumoniae	NDM, OXA-181	60
1300920	Klebsiella pneumoniae	NDM, OXA-181	15
MSH2014-69	Klebsiella pneumoniae	NDM, OXA-181	20
NCTC 13476	Escherichia coli	IMP-1	250
695	Acinetobacter baumannii	IMP-1	1720
2340	Enterobacter cloacae	IMP-1	250
IMPBMI	Klebsiella pneumoniae	IMP-1	100
Yonsei_1	Acinetobacter baumannii	IMP-1	1000
Yonsei_2	Acinetobacter baumannii	IMP-1	500
6852	Klebsiella pneumoniae	IMP-1	100
MKAM	Pseudomonas aeruginosa	IMP-1	500
70450-1	Pseudomonas aeruginosa	IMP-1	250
3994	Pseudomonas spp.	IMP-10	250
CDC0161	Enterobacter aerogenes a	IMP-4	5.00E+04
5344	Pseudomonas aeruginosa	IMP-2	60
3985	Pseudomonas aeruginosa	IMP-11	2000
4032	Pseudomonas aeruginosa	IMP-6	80
3424	Pseudomonas aeruginosa	IMP-7b, c	1.00E+06
32443	Klebsiella pneumoniae	IMP-13c	1.00E+06
92	Pseudomonas aeruginosa	IMP-14b, c	1.00E+06

Table 8-3: Analytical Reactivity of the Xpert Carba-R Assay in Perirectal Swab Matrix

{11}------------------------------------------------

a. These organisms were not tested as bacterial isolates.

b. IMP-7 and IMP-14 genes (Pseudomonas aeruginosa) were not detected by the assay and were not predicted to be detected by in silico analysis (see Limitations).

{12}------------------------------------------------

c. IMP-13 gene (Klebsiella pneumoniae): although predicted to be detected by in silico analysis, the IMP-13 gene was not detected by the assay (see Limitations).

Marker(or Traditional Subgroup)	No. of Samples	Wet testing		Not tested but predicted to be detected Based on In Silico Analysis
		Type(s) Detected	Type(s) not Detected
KPC	12	KPC-2, 3, 4	--	KPC-5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16
NDM	18	NDM-1, 2, 4, 5	--	NDM-3, 6, 7, 8, 9
VIM	12	VIM-1, 2, 4, 10, 19	--	VIM-5, 6, 7, 8, 9, 11, 12, 13, 14, 15, 16, 17, 18, 20, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38
OXA-48	19	OXA-48, 181(OXA-48 variant)	--	OXA-162, 163, 204, 232, 244, 245, 247
IMP	17	IMP-1 (9 strains), IMP-2, 4, 6, 10, 11	IMP-7a, 13b, 14a	IMP-3, 8, 9, 13b, 19, 20, 21, 22, 24, 25, 27, 28, 30, 31, 33, 37, 40, 42

Table 8-4: Summary of Variants Detected by Wet Testing or Predicted to be
Detected Based on In Silico Analysis

a. IMP-7 and IMP-14 genes (Pseudomonas aeruginosa) were not detected by the assay and were not predicted to be detected by in silico analysis (see Limitations).

b. IMP-13 gene (Klebsiella pneumoniae) was tested: although predicted to be detected by in silico analysis, the IMP-13 gene was not detected by the assay (see Limitations).

Analytical Specificity (Cross-reactivity)

The analytical specificity of the Xpert Carba-R Assay was evaluated for organisms seeded into perirectal swab matrix. A panel of 62 well-characterized bacterial strains of carbapenem-susceptible bacteria or bacteria with carbapenem non-susceptibility due to genes or mechanisms other than the Xpert Carba-R target genes (Table 8-5 and Table 8-6) and 24 commensal bacterial strains and other enteric microorganisms were also evaluated in the study (Table 8-7). Human cells were also tested in perirectal swab matrix (Table 8-8). Resistance mechanisms were determined by individual PCR assays, DNA sequence analysis, or Check-Points array version CT102.

For perirectal swab matrix samples, 62 strains were tested at concentrations >1 x 100 CFU/mL with the exception of Peptostreptococcus anaerobius that was tested at 5 x105 CFU/mL. Viruses were tested at >1 x 105 TCID50/mL or greater than 2.5 x107 RNA copies/mL. A bladder cell line (human genomic DNA) was tested at 1 x 10° cells/mL. Organisms were diluted into pooled negative perirectal swab matrix and tested in triplicate. None of the 94 potentially cross-reactive organisms and nucleic acids tested was detected with the Xpert Carba-R Assay.

The Xpert Carba-R Assay did not cross react with any of the organisms tested (Table 8-5, Table 8-6, and Table 8-7). The analytical specificity of the assay was 100%.

{13}------------------------------------------------

each Allubloutic
	Ertapenem	ImipenemMeropenem
Susceptible	19	30	24
Intermediate		8	ਪ
Resistant	43	24	34

Table 8-5: Number of Carbapenem-susceptible and Non-susceptible Organisms for each Antibiotic

Organism	Strain ID	Confirmed ResistanceMechanisms	Carbapenemsusceptibility (S/I/R)a
			ETPa	IMPa	MEMa
Escherichia coli	NCTC 13441	CTX-M (-1, -type 15 like); TEM	S	S	S
Klebsiella pneumoniae	NCTC 13465	CTX-M (25)	S	S	S
Enterobacter aerogenes	810	OmpC/OmpF deficient; TEM	R	R	R
Citrobacter freundii	1698	TEM (WT+164S)	S	S	S
Enterobacter cloacae	5557	AmpC (ACT/MIR)	R	R	R
Klebsiella pneumoniae	kpn5	CTX-M-2	R	S	R
Klebsiella pneumoniae	kpn12	TEM; SHV; CTX-M	R	R	R
Escherichia coli	eco1	TEM; CTX-M-2	R	R	R
Escherichia coli	eco2	CTX-M (2); TEM	R	S	S
Enterobacter cloacae	corl	CTX-M (2); TEM	R	R	R
Serratia marcescens	hpp21	CTX-M (2); TEM	S	S	S
Morganella morganii	fer29	CTX-M (2); TEM	S	R	S
Proteus mirabilis	gut25	CTX-M (2); TEM	S	R	S
Salmonella spp.	3209	CTX-M (2); TEM	S	S	S
Shigella flexnerii	3331	CTX-M (2); TEM	S	S	S
Enterobacter cloacae	PA_3	AmpC; CTX-M-15; TEM	S	S	S
Klebsiella pneumoniae	32189	SHV	S	S	S
Klebsiella pneumoniae	32443	CTX-M (1, -type 15 like); SHV	S	S	S
Klebsiella pneumoniae	32598	CTX-M (-1, -type 15 like); SHV;TEM	R	I	R
Klebsiella pneumoniae	33560	CTX-M (15); SHV-11; TEM-1	S	S	S
Klebsiella pneumoniae	33603	SHV-2	R	I	R
Klebsiella pneumoniae	33617	SHV-27	S	S	S
Klebsiella pneumoniae	33643	SHV (-5, -55); TEM	S	S	S
	Confirmed Resistance	Carbapenemsusceptibility (S/I/R)a
Organism	Strain ID	Mechanisms	ETPa	IMPa	MEMa
Klebsiella pneumoniae	34430	SHV; TEM; CTX-M-15	S	S	S
Klebsiella pneumoniae	34680	TEM; CTX-M-2	R	S	R
Klebsiella pneumoniae	34732	CTX-M (15); SHV; TEM	R	S	S
Enterobacter cloacae	PA_174	GX-/Culture+; SHV; TEM	S	S	S
Enterobacter aerogenes	STU 645	SHV (WT+238S+240K)	R	S	R
Enterobacter aerogenes	STU 669	SHV (WT+238S+240K)	R	R	R
Escherichia coli	C3015	AmpC (CMY II); TEM	R	R	R
Enterobacter aerogenes	RI_100	AmpC (DHA); SHV	R	R	R
Klebsiella pneumoniae	B4A	SHV (WT + 238S +240K)	R	R	R
Klebsiella pneumoniae	B13A	SHV (WT + 238S +240K)	R	S	S
Enterobacter cloacae	RI_474	AmpC (ACT/MIR)	R	I	I
Enterobacter amnigenus	B71	AmpC (ACT/MIR)	R	R	R
Klebsiella pneumoniae	DD82A	SHV (WT + 238S + 240K)	R	S	R
Klebsiella pneumoniae	B100	CTX-M (-1, type-15 like); SHV(WT+238S); TEM	R	S	R
Enterobacter cloacae	135B	TEM	S	S	S
Klebsiella pneumoniae	B157	SHV; TEM	R	R	R
Escherichia coli	T2914280	CTX-M (-1, -15); TEM	R	S	R
Providencia stuartii	DD188	TEM (104K + 164S)	R	I	I
Enterobacter cloacae	DD189	AmpC (ACT/MIR)	R	S	S
Escherichia coli	B198B	CTX-M (-1, type -15 like); TEM	R	S	R
Klebsiella pneumoniae	T3019989-1	CTX-M (-1, type-15 like); SHV	R	I	R
Klebsiella pneumoniae	T3019989-2	CTX-M (-1, type-15 like); SHV	R	S	R
Enterobacter cloacae	ENC-THAI14	VEB-1, TEM	S	S	S
Escherichia coli	CB154006	CTX-M (9); TEM	R	I	I
Enterobacter cloacae	S35766	AmpC(ACT/MIR)	S	S	S
Enterobacter cloacae	X1856910	AmpC (ACT/MIR); TEM	R	I	I
Klebsiella pneumoniae	W3758164	CTX-M (-1, -15 like); SHV; TEM.	R	I	R
Klebsiella pneumoniae	X2135758	CTX-M (-1, -15 like); SHV	R	S	S
Klebsiella pneumoniae	W3809535	CTX-M (-1, -15 like); SHV	R	R	R
Pseudomonas aeruginosa	CDC0064	SPM	R	R	R
Organism	Strain ID	Confirmed ResistanceMechanisms	Carbapenemsusceptibility (S/I/R)a
			ETPa	IMPa	MEMa
Serratia marcescens	CDC0099	SME	R	R	R
Serratia marcescens	CDC0121	SME	R	R	R
Serratia marcescens	CDC0122	SME	R	R	R
Serratia marcescens	CDC0123	SME	R	R	R
Serratia marcescens	CDC0124	SME	R	R	R
Serratia marcescens	CDC0130	SME	R	R	R
Serratia marcescens	CDC0131	SME	R	R	R
Enterobacter cloacaegroup	CDC0132	IMI	R	R	R
Enterobacter cloacaecomplex	CDC0164	IMI	R	R	R

Table 8-6: Cross-reactivity Panel

Xpert Carba-R 510(k)

{14}------------------------------------------------

{15}------------------------------------------------

a. S//R = Susceptible/Intermediate/Resistant, ETP = Ertapenem, IMP = Imipenem, MEM = Meropenem

Strain ID	Organism	Concentration Tested(CFU/mL unless otherwise specified)
ATCC 25922	Escherichia coli	2.67E+06
ATCC 29212	Enterococcus faecalis	3.15E+06
ATCC 700603	Klebsiella pneumoniae	5.20E+06
ATCC 35218	Escherichia coli	2.47E+06
ATCC 25923	Staphylococcus aureus	4.53E+06
ATCC 27853	Pseudomonas aeruginosa	3.17E+06
ATCC 9689	Clostridium difficilea	1.80E+07
ATCC 700621	Enterobacter cloacae	8.95E+06
ATCC 9756	Enterococcus faecium	6.54E+06
ATCC 13182	Klebsiella oxytoca	4.76E+06
ATCC BAA-747	Acinetobacter baumannii	2.27E+06
ATCC 33128	Citrobacter freundii	2.01E+06
ATCC 49948	Morganella morganii	8.19E+06
ATCC 51331	Stenotrophomonas maltophilia	3.15E+06
ATCC 27028	Citrobacter koseri	5.05E+06
ATCC 49809	Providencia stuartii	3.01E+06
ATCC 49037	Peptostreptococcus anaerobiusa	5.00E+05
CCUG 29780 / ATCC 12401	Streptococcus agalactiae	5.21E+06
ATCC 15703	Bifidobacterium adolescentisa	1.10E+08
ATCC 51697	Enterobacter aerogenes	3.19E+06

Table 8-7: Cross-reactivity Panel (Commensal and Other Enteric Microorganisms)

{16}------------------------------------------------

Strain ID	Organism	Concentration Tested(CFU/mL unless otherwisespecified)
ATCC 43071	Proteus mirabilis	1.78E+06
CCUG 34787	Acinetobacter spp.	2.40E+06
CCUG 418	Citrobacter freundii	2.95E+06
CCUG 33629	Corynebacterium diphtheriae	4.48E+06
CCUG 17874	Helicobacter pylori	1.61E+06
CCUG 33548	Listeria monocytogenes	4.77E+06
CCUG 6325	Providencia alcalifaciens	4.91E+06
CCUG 43594 / ATCC 33560	Campylobacter jejunia	3.27E+06
MRVP/ZeptoMetrix	Adenovirus B Type 7A/NYa	1.40E+05 TCID50/mL
MRVP/ZeptoMetrix	Enterovirus Type 71/NYa	4.40E+05 TCID50/mL
Clinical Sample - Cepheid	Norovirus GIIa	2.5 x 107 RNA copies/mL

a. These organisms were tested in perirectal swab matrix only.

Table 8-8: Cell Line Representing Human Genomic DNA

Organism Name	Source
Bladder Cell Carcinoma(hgDNA)	ATCC HTB-4

Competitive Interference

A competitive interference study was performed to test whether a high titer of one or more carbapenemase-producing organisms would interfere with the detection of a second target carbapenemase-producing organism that was present at a low titer. High titer samples were formulated at concentrations of 5 x 10° CFU/swab and low titer targets were formulated at approximately 2X LoD for the respective strain in perirectal swab matrix. One carbapenemase-producing bacterial strain for each gene analyte, i.e., the genes encoding KPC, NDM, VIM, OXA-48, and IMP, was used in this study. Each carbapenemase-producing bacterial strain type was tested at low titers in conjunction with a high titer of each of the other one or two carbapenemase-producing bacterial strain types (Table 8-9). Samples were tested in replicates of eight.

An inhibitory effect was observed for two of the five targets (NDM and IMP) when a low concentration of each target was present in combination with a high concentration of one or two other targets for samples tested in perirectal swab matrix. The two targets (NDM and IMP) were tested at a higher concentration (4X LoD) in combination with a high concentration of one or two other targets for samples in perirectal swab matrix. No inhibitory effect was observed for the two targets (NDM and IMP) at 4x LoD in the presence of clinically relevant co-infections for the Xpert Carba-R Assay.

The competitive inhibitory effect on the Carba-R targets (NDM, IMP, VIM and OXA-48) is addressed in the Limitations section in the package insert.

{17}------------------------------------------------

Combination
High KPC/High NDM/Low VIM
High KPC/High NDM/Low OXA
High KPC/High NDM/Low IMP
High VIM/High OXA/Low KPC
High VIM/High OXA/Low NDM
High VIM/High OXA/Low IMP
High IMP/Low KPC
High IMP/Low NDM
High IMP/Low VIM
High IMP/Low OXA
High OXA/Low VIM
High VIM/Low OXA
High KPC/Low NDM
Negative

Table 8-9: Combinations of Carbapenemase-producing Bacteria Tested with the Xpert Carba-R Assav

Potentially Interfering Substances

The performance of the Xpert Carba-R Assay was evaluated with 24 potentially interfering substances that may be present in perirectal swab specimens. Potentially interfering substances (IS) solutions were prepared and tested at concentrations specified in

{18}------------------------------------------------

Table 8-10. Positive and negative samples were included in this study. Positive samples consisted of a mix of five carbapemase-producing organisms harboring KPC, NDM, VIM, IMP-1 and OXA-48 gene sequences seeded into pooled negative perirectal swab matrix at approximately 3X LoD. Eight replicate positive samples were tested per substance. Negative samples consisted of pooled negative perirectal swab matrix not seeded with carbapenemase-producing organisms. Eight replicate negative samples were tested per substance to determine the effect on the performance of the sample processing control (SPC). Controls consisted of positive and negative samples with no interfering substances added. The effect of each potentially interfering substance on positive and negative replicates was evaluated by comparing target cycle threshold (Ct) values generated in the presence of the substance to Ct values from controls lacking the substance. The positive and negative replicate samples for 22 potentially interfering substances were correctly identified using the Xpert Carba-R Assay. Interference with the Xpert Carba-R Assay may be observed with barium sulfate at > 0.1% w/v and Pepto-Bismol at > 0.025% w/v in tests with perirectal swab matrix samples. See Limitations in the package insert.

{19}------------------------------------------------

Substance/Class	Active Ingredient	Concentration Tested
Non-steroidal anti-inflammatory medication	Naproxen	0.25% w/v
Imaging compound	Barium sulfate	0.25% and 0.1% w/v
Antibiotic (oral)	Cephalexin	0.25% w/v
Antibiotic (oral)	Ciprofloxacin	0.25% w/v
Condom with spermicidal lubricant	Nonoxynol-9	1 condoma
Creams/ointment/suppositories	Hydrocortisone	0.25% w/v
Laxative	Sennosides	0.25% w/v
Lipids	Stearic acid/Palmitic acid/Cholesterol (fecal fat)	0.25% w/v
Anti-diarrheal medication	Loperamide hydrochloride/bismuth subsalicylate (Imodium)	0.25% w/v
Anti-diarrheal medication	Loperamide hydrochloride/bismuth subsalicylate (Kaopectate)	0.25% w/v
Topical cream	K-Y Jelly	0.25% w/v
Antacids	Calcium carbonate/aluminum hydroxide/magnesium hydroxide/simethicone (Milk of Magnesia)	0.25% w/v
Enemas	Mineral oil	0.25% w/v
Antibiotic (topical)	Polymixin B/ Neomycin/Bacitracin (Neosporin)	0.25% w/v
Anti-fungal/anti-itch Vaginal	Nystatin	0.25% w/v
Antacid	Famotidine (Pepcid)	0.25% w/v
Anti-diarrheal medication	Loperamide hydrochloride/bismuth subsalicylate (Pepto-Bismol)	0.25%, 0.1%, 0.05%, 0.025%, 0.01% w/v
Topical cream	Petroleum jelly	0.25% w/v
Anti-hemorrhoid creams/ointments	Phenylephrine (Preparation H)	0.25% w/v
Acid reducer; antacid	Oemprazole (Prilosec)	0.25% w/v
Enemas	Saline-enema	0.25% w/v
Antacid	Cimetidine (Tagamet)	0.25% w/v
Anti-fungal/anti-itch Vaginal	Benzocaine, resorcinol (Vagisil)	0.25% w/v
Moist towelettes	Benzalkonium chloride, ethanol (Wet Ones)	1 pieceb

	Table 8-10: Potentially Interfering Substances Tested

a. One condom added to 40 mL swab matrix.

b. One piece (5 inch x 7-1/2 inch) added to 40 mL swab matrix.

{20}------------------------------------------------

Carry-Over Contamination

A study was conducted to demonstrate that single-use, self-contained GeneXpert cartridges prevent carry-over contamination in negative samples run following very high positive samples. The study consisted of a negative sample processed in the same GeneXpert module immediately following a very high positive sample. The high positive sample is composed of inactivated E. coli cells containing a plasmid with an insert consisting of a synthetic oligonucleotide of the amplicon sequences from the five Xpert Carba-R target analyte genes (KPC, NDM, VIM, IMP and OXA-48 targets). Positive cells were diluted in pooled perirectal swab matrix to a concentration of 1 x 10° CFU/mL. The testing scheme was repeated 25 times on two GeneXpert modules for a total of 102 tests (25 high positive samples per module and 26 negative samples per module) for the perirectal swab matrix. All 50 positive samples correctly reported all Xpert Carba-R targets as DETECTED. All 52 negative samples correctly reported all Xpert Carba-R targets as NOT DETECTED.

Clinical Studies

Clinical Performance -Perirectal Swab Specimens

Performance characteristics of the Xpert Carba-R Assay perirectal swab specimens were determined in a multi-site investigational study. The positive percent agreement (PPA) and negative percent agreement (NPA) of the Xpert Carba-R Assay was evaluated relative to a reference method of culture (MacConkey enrichment broth) and PCR/bidirectional DNA sequence analysis.

Five geographically diverse sites prospectively collected paired perirectal swab specimens from subjects who were hospitalized or in a long-term care facility. Highly soiled perirectal swab specimens, according to the directions in Section 9 of package insert (Sample Preparation and Storage) were excluded from the study. Due to low prevalence of each of the Xpert Carba-R Assay target genes in the absence of an outbreak, contrived specimens were also included in the study.

One swab of the pair was used for Xpert Carba-R Assay testing. The second swab was inoculated into MacConkey enrichment broth and used for reference method testing. A reference culture laboratory determined the presence of carbapenem non-susceptible organisms by culturing the MacConkey enrichment broth from each of the specimens. The MacConkey enrichment broth was screened for the presence of carbapenem-non-susceptible organisms initially by plating the broth on MacConkey agar plates with a meropenem disk. For specimens that exhibited growth of gram-negative bacteria around the meropenem disk, confirmation of carbapenem non-susceptibility was determined on isolated colonies by using the disk diffusion method (per CLSI document M02) as well as CLSI document M100. DNA extracted from the carbapem non-susceptible isolates was purified, quantified, and amplified using primers specific to all five target genes; amplified regions included more bases than the regions amplified by the Xpert Carba-R Assay. The production of the appropriate size amplification product was confirmed on Agilent 2100 Bioanalyzer (Agilent Technologies, Santa Clara, CA).

{21}------------------------------------------------

If bands shown on the Bioanalyzer corresponded to the expected size of the amplicon from any of the five target genes detected by the Xpert Carba-R Assay, the amplicon for the isolate was sent to an independent laboratory for reference bi-directional sequencing analysis, which was validated for detection of the five targets in the Xpert Carba-R Assay. If no bands were shown on the Bioanalyzer for any of the five target genes, the isolate was not sent for sequence analysis and the reference method result was considered negative for the five target genes.

Prospective Specimen Results Obtained with the Xpert Carba-R Assay in Comparison to the Reference Method

A total of 963 prospective perirectal swab specimens were initially enrolled in this clinical study, of which 947 were eligible for inclusion. From the 947 eligible specimens, 924 specimens were included in the final dataset after exclusions based on protocol deviations (including 10 Stenotrophomonas maltophilia, one Pseudomonas putida and one Pseudomonas stutzeri organisms that were excluded due to their intrinsic resistance to the carbapenems tested).

When tested with prospective perirectal swab specimens, the Xpert Carba-R Assay demonstrated a PPA of 100% for the three assay targets (blayDM, blagge and blaoxA-48) relative to the reference method. The NPA for the blakpc, blaNDM, blaoxA-48, and blaIMP gene sequences ranged from 99.6% to 100% relative to the reference method (Table 8-11).

For specimens with discordant results (the Xpert Carba-R Assay was positive for a target gene but a carbapenem-non-susceptible organism was not isolated by reference culture), discordant analysis was performed using bi-directional sequencing on DNA extracted directly from the MacConkey enrichment broth. Discrepant testing results are footnoted in Table 8-11.

Specimen Type	Target	N	TP	FP	TN	FN	PPA	NPA
							(95% CI)	(95% CI)
Perirectalc	IMP	924	0	0	924	0	N/A	100%(99.6-100)
	VIM	924	0	0	924	0	N/A	100%(99.6-100)
	NDM	924	1	0	923	0	100%(20.7-100)	100%(99.6-100)
	KPC	924	2	4a	918	0	100%(34.2-100)	99.6%(98.9-99.8)
	OXA-48	924	1	1b	922	0	100%(20.7-100)	99.9%(99.4-100)

Table 8-11: Xpert Carba-R Performance vs. Reference Culture + Sequencing -Prospective Perirectal Specimens

N = Number, TP = True Positive, FP = False Positive, TN = True Negative, FN = False Negative

a. Testing results by sequencing: 4 of 4 were KPC negative.

b. Testing results by sequencing: 1 of 1 was OXA-48 negative.

c. Of the 924 prospective perirectal swab specimens evaluated in the study, 891 specimens did not yield a culture isolate. From the remaining 33 specimens, 31 carbapenem-non-susceptible organisms were recovered by the Reference Culture in addition to two carbapenem susceptible organisms (Pseudomonas aeruginosa).

{22}------------------------------------------------

Performance of the Xpert Carba-R Assay on the prospective perirectal specimens is shown in Table 8-12 by species. Only organisms for which at least one positive specimen was collected are included in Table 8-12.

Speciesa	Target	N	TP	TN	FN	PPA(95% CI)	NPA(95% CI)
Enterobacteraerogenes	IMP	1	0	1	0	NA	100%(20.7-100)
	VIM	1	0	1	0	NA	100%(20.7-100)
	NDM	1	0	1	0	NA	100%(20.7-100)
	KPC	1	1	0	0	100%(20.7-100)	NA
	OXA-48	1	0	1	0	NA	100%(20.7-100)
Klebsiellapneumoniae	IMP	3	0	3	0	NA	100%(43.9-100)
	VIM	3	0	3	0	NA	100%(43.9-100)
	NDM	3	1	2	0	100%(20.7-100)	100%(34.2-100)
	KPC	3	1	2	0	100%(20.7-100)	100%(34.2-100)
	OXA-48	3	1	2	1	96.2%(20.7-100)	100%(34.2-100)

Table 8-12. Xpert Carba-R Performance vs. Reference Culture + Sequencing by Organism type - Prospective Perirectal Snecimens

a. Acinetobacter baumannii (1), and Pseudomonas aeruginosa (28) were recovered but did not contain target sequences by the Reference Method.

Multiple targets were detected by the Xpert Carba-R Assay in one prospective specimen. The details are provided in Table 8-13, along with the discrepant sequencing result.

Table 8-13. Prospective Perirectal Specimens with Multiple Targets Detected

Specimen	Targets Detectedby Xpert Carba-RAssay	TargetsDetected byReferenceSequencing	Discrepant Testing Results -Targets Detected by ReferenceSequencing
g	NDM, OXA-48	NDM, OXA-48	NA

Contrived Specimen Results Obtained with the Xpert Carba-R Assay in Comparison to the Reference Method

A total of 432 contrived specimens prepared in perirectal matrix were also tested as part of the clinical study.

In addition to Enterobacteriaceae, Pseudomonas aeruginosa, and Acinetobacter baumannii groups tested in the contrived study, 5 other non-Enterobacteriaceae strains were also evaluated: Pseudomonas stutzeri (1), Pseudomonas oryzihabitans (1), Pseudomonas putida (2), and Empedobacter brevis (1).

{23}------------------------------------------------

When tested with contrived specimens, the Xpert Carba-R Assay demonstrated a PPA and NPA of 100% for the blakpc, blaNDM, blayM, blagxA-48, and blaIMP gene sequences was 100% relative to the reference method (Table 8-14).

Specimens
Matrix	Target	N	TP	FP	TN	FN	PPA(95% CI)	NPA(95% CI)
Perirectal	IMP	432	80	0	352	0	100%(95.4-100)	100%(98.9-100)
	VIM	432	82	0	350	0	100%(95.5-100)	100%(98.9-100)
	NDM	432	80	0	352	0	100%(95.4-100)	100%(98.9-100)
	KPC	432	80	0	352	0	100%(95.4-100)	100%(98.9-100)
	OXA-48	432	80	0	352	0	100%(95.4-100)	100%(98.9-100)

Table 8-14. Xpert Carba-R Performance vs. Reference Method - Contrived Snecimens

Perirectal Swab and Rectal Swab Equivalence Study

To demonstrate equivalence of perirectal swab specimens and rectal swab specimens, a study was conducted at one site enrolling fresh prospectively collected rectal and perirectal swab specimens from consented subjects who were hospitalized in-patients.

Paired swab sets provided in the Cepheid Specimen Collection Device were used to collect specimens from each subject. One paired swab set was used to collect the perirectal swab specimen and a second paired swab set was used to collect the rectal swab specimen. The perirectal swab specimen was collected first followed by the rectal swab specimen from the same subject. One swab from each paired swab set was used for Xpert Carba-R Assay testing. The second swab from each paired swab set was used for culture and susceptibility testing when either or both the perirectal or rectal swab specimen(s) were positive for one or more target(s) by the Xpert Carba-R Assay. No culture was performed if perirectal and rectal swab specimens were both negative by the Xpert assay.

Bi-directional DNA sequencing was performed on DNA extracted from isolated colonies that manifested carbapenem-non-susceptibility by the CLSI disk diffusion method or from MacConkey broth with meropenem disk if the culture result was negative and the Xpert Carba-R Assay result was positive.

A total of 207 specimens were initially enrolled in this clinical study, all of which were eligible for inclusion. Of the 207 eligible specimens were included in the final dataset used for the analyses. Six swab specimens (4 perirectal swab specimens and 2 rectal swab specimens) were excluded due to indeterminate results from the Xpert Carba-R Assay.

Of the 201 specimens included in the data analyses, 92 (45.8%) were collected from female subjects and 109 (54.2%) from male subjects. Overall 45.8% (92/201) specimens were collected from subjects between 21 and 65 years of age and 54.2%

{24}------------------------------------------------

(109/201) were from subjects >65 years of age.

The performance (PPA and NPA) of the Xpert Carba-R Assay using perirectal swab specimens was determined relative to the results of the Xpert Carba-R Assay using rectal swab specimens from the same subject. The PPA and NPA estimates are shown in

Table 8-15. Relative to the Xpert Carba-R Assay rectal swab specimen result, the perirectal swab specimens demonstrated an overall PPA and NPA of 94.7% (95%CI: 75.4-99.1) and 97.8% (95%CI: 94.5-99.1), respectively.

Table 8-15. Xpert Carba-R Assay – Perirectal Swab Specimens vs Rectal Swab
Specimens

Xpert Carba-R Assay – Rectal Swab Specimens
		Positive	Negative	Total
Xpert Carba-R Assay –Perirectal SwabSpecimens	Pos	18a	4b	22
	Neg	1c	178	179
	Total	19	182	201
	PPA		94.7% (95%CI: 75.4-99.1)
	NPA		97.8% (95%CI: 94.5-99.1)

4 For one specimen, Xpert testing on the rectal swab was positive for KPC and OXA-48 and on the perirectal swab was positive for OXA-48 only. The specimen was culture negative for both rectal and perirectal swabs. Sequence results from the MacConkey broths were negative for the perirectal swab and OXA-48 positive for the rectal swab.

b 2 of 4 were culture positive for both rectal and perirectal swabs, sequence results from isolates were both OXA-48 positive, 1 of 4 was culture negative for both rectal and perirectal swabs, sequence result the rectal sequence result was not available due to isolate not saved, the perirectal isolate was interpreted as carbapenem susceptible and per protocol sequencing was not required.

Culture negative for both rectal and perirectal swabs, sequence results from MacConkey broths were both OXA-48 Positive.

{25}------------------------------------------------

Reproducibility Study

Reproducibility of the Xpert Carba-R Assay was evaluated using of a panel of 11 samples, prepared in pooled negative perirectal swab matrix. Two operators at each of the three study sites tested one panel of 11 samples in replicates of four per day over six testing days (11 samples x 2 replicates x 2 times/day x 6 days x 2 operators x 3 sites). Three lots of Xpert Carba-R Assay cartridges were used at each of the 3 testing sites. The Xpert Carba-R Assay was performed according to the Xpert Carba-R Assay procedure. Results are summarized in Table 8-16.

Matrix
Sample	Site 1			Site 2			Site 3			% TotalAgreementby Sample
	Op 1	Op 2	Site	Op 1	Op 2	Site	Op 1	Op 2	Site
Neg	100%(24/24)	100%(24/24)	100%(48/48)	100%(24/24)	100%(24/24)	100%(48/48)	100%(24/24)	100%(24/24)	100%(48/48)	100%(144/144)
IMPMod Pos	100%(24/24)	100%(24/24)	100%(48/48)	100%(24/24)	100%(24/24)	100%(48/48)	100%(24/24)	100%(24/24)	100%(48/48)	100%(144/144)
IMPLow Pos	95.8%(23/24)	91.7%(22/24)	93.8%(45/48)	100%(24/24)	100%(24/24)	100%(48/48)	100%(24/24)	91.7%(22/24)	95.8%(46/48)	96.5%(139/144)
VIMMod Pos	100%(24/24)	100%(24/24)	100%(48/48)	100%(24/24)	100%(24/24)	100%(48/48)	100%(24/24)	100%(24/24)	100%(48/48)	100%(144/144)
VIMLow Pos	100%(24/24)	91.7%(22/24)	95.8%(46/48)	91.7%(22/24)	91.7%(22/24)	91.7%(44/48)	95.8%(23/24)	83.3%(20/24)	89.6%(43/48)	92.4%(133/144)
NDMMod Pos	100%(24/24)	100%(24/24)	100%(48/48)	100%(24/24)	100%(24/24)	100%(48/48)	100%(24/24)	100%(24/24)	100%(48/48)	100%(144/144)
NDMLow Pos	100%(24/24)	100%(24/24)	100%(48/48)	100%(24/24)	100%(24/24)	100%(48/48)	87.5%(21/24)	100%(24/24)	93.8%(45/48)	97.9%(141/144)
KPCMod Pos	100%(24/24)	100%(24/24)	100%(48/48)	100%(24/24)	100%(24/24)	100%(48/48)	100%(24/24)	100%(24/24)	100%(48/48)	100%(144/144)
KPCLow Pos	91.7%(22/24)	91.7%(22/24)	91.7%(44/48)	91.7%(22/24)	95.8%(23/24)	93.8%(45/48)	100%(24/24)	91.7%(22/24)	95.8%(46/48)	93.8%(135/144)
OXA-48Mod Pos	100%(24/24)	100%(24/24)	100%(48/48)	100%(24/24)	100%(24/24)	100%(48/48)	100%(24/24)	100%(24/24)	100%(48/48)	100%(144/144)
OXA-48Low Pos	87.5%(21/24)	87.5%(21/24)	87.5%(42/48)	100%(24/24)	95.8%(23/24)	97.9%(47/48)	95.8%(23/24)	95.8%(23/24)	95.8%(46/48)	93.8%(135/144)

Table 8-16: Summary of Reproducibility Results - % Agreement, Perirectal Swab
Matrix

The reproducibility of the Xpert Carba-R Assay was also evaluated in terms of the fluorescence signal expressed in Ct values for each target detected. The mean, standard deviation (SD), and coefficient of variation (CV) between-sites, between-lots, betweendays, between-operators, and within-assays for each panel member are presented in

Table 8-17.

Table 8-17: Summary of Reproducibility Data for Perirectal Swab Matrix

Sample	AssayChannel(Analyte)	Na	MeanCt	BetweenSite		BetweenLot		BetweenDay		Between-Operator		Within-Assay		Total
				SD	CV(%)	SD	CV(%)	SD	CV(%)	SD	CV(%)	SD	CV(%)	SD	CV(%)
Neg	SPC	144	32.7	0.0	0.0	0.2	0.6	0.0	0.0	0.2	0.5	0.4	1.2	0.5	1.4
IMPMod Pos	IMP	144	33.7	0.0	0.0	0.1	0.2	0.0	0.0	0.2	0.5	0.5	1.5	0.5	1.6
IMPLow Pos	IMP	142	36.0	0.2	0.5	0.0	0.0	0.1	0.3	0.2	0.5	0.8	2.1	0.8	2.3

{26}------------------------------------------------

Sample	Assay		Mean	BetweenSite		BetweenLot		BetweenDay		Between-Operator		Within-Assay		Total
	Channel(Analyte)	Na	Ct	SD	CV(%)	SD	CV(%)	SD	CV(%)	SD	CV(%)	SD	CV(%)	SD	CV(%)
VIMMod Pos	VIM	144	31.2	0.1	0.2	0.1	0.3	0.0	0.1	0.2	0.5	0.4	1.3	0.5	1.5
VIMLow Pos	VIM	142	35.0	0.0	0.0	0.6	1.6	0.0	0.0	0.6	1.7	1.4	4.1	1.6	4.7
NDMMod Pos	NDM	144	33.2	0.0	0.0	0.0	0.0	0.2	0.5	0.2	0.5	0.4	1.2	0.5	1.4
NDMLow Pos	NDM	143	35.7	0.2	0.5	0.0	0.0	0.2	0.6	0.0	0.0	0.9	2.4	0.9	2.5
KPCMod Pos	KPC	144	34.6	0.0	0.0	0.3	1.0	0.0	0.0	0.2	0.5	0.4	1.3	0.6	1.7
KPCLow Pos	KPC	143	36.4	0.0	0.0	0.5	1.3	0.1	0.4	0.0	0.0	0.7	2.0	0.9	2.4
OXA-48Mod Pos	OXA-48	144	34.4	0.1	0.2	0.2	0.6	0.0	0.0	0.2	0.5	0.5	1.5	0.6	1.7
OXA-48Low Pos	OXA-48	144	36.4	0.0	0.0	0.0	0.0	0.4	1.2	0.0	0.0	1.0	2.7	1.1	2.9

a. Results with non-zero Ct values out of 144.

Conclusions

The results of the nonclinical analytical and clinical performance studies summarized above demonstrate that the Xpert Carba-R Assay is safe and effective for its intended use and is substantially equivalent to the predicate device.

Regulation Number and Section

§ 866.1640 Antimicrobial susceptibility test powder.

(a)
Identification. An antimicrobial susceptibility test powder is a device that consists of an antimicrobial drug powder packaged in vials in specified amounts and intended for use in clinical laboratories for determining in vitro susceptibility of bacterial pathogens to these therapeutic agents. Test results are used to determine the antimicrobial agent of choice in the treatment of bacterial diseases.(b)
Classification. Class II (performance standards).