GenePOC Strep A
Device Facts
| Record ID | K183366 |
|---|---|
| Device Name | GenePOC Strep A |
| Applicant | Genepoc, Inc. |
| Product Code | PGX · Microbiology |
| Decision Date | Mar 6, 2019 |
| Decision | SESE |
| Submission Type | Traditional |
| Regulation | 21 CFR 866.2680 |
| Device Class | Class 2 |
Indications for Use
The GenePOC™ Strep A assay, performed on the revogene™ instrument, is an automated, qualitative in vitro diagnostic test that utilizes real-time polymerase chain reaction (PCR) for the direct detection of Streptococcus pyogenes (Group A B-hemolytic Streptococcus) nucleic acids from throat swab specimens obtained from patients with signs and symptoms of pharyngitis. The GenePOC™ Strep A assay is intended for use as an aid in the diagnosis of Group A Streptococcus infection.
Device Story
The GenePOC™ Strep A assay is a single-use, qualitative in vitro diagnostic test for detecting Streptococcus pyogenes nucleic acids. It uses a microfluidic cartridge (PIE) and a Sample Buffer Tube (SBT) to process throat swab specimens. The revogene™ instrument automates sample homogenization, cell lysis, DNA extraction, and real-time PCR amplification. An internal Process Control (PrC) verifies sample processing and amplification. The system uses target-specific TaqMan® probes to detect amplified DNA. Results are computed from fluorescent signals and an Early Positive Result Outcome (E-PRO) feature. The device is used in clinical settings by healthcare professionals. Output results (positive, negative, indeterminate, unresolved) are displayed to the user to aid in diagnosing Group A Streptococcus infection, potentially facilitating timely clinical decision-making.
Clinical Evidence
Prospective multicenter study (n=604 evaluable specimens) compared GenePOC Strep A assay to reference culture. Sensitivity: 98.1% (95% CI: 94.4-99.3%); Specificity: 94.7% (95% CI: 92.2-96.4%). Positive Predictive Value: 86.3%; Negative Predictive Value: 99.3%. Analytical studies included precision, reproducibility, limit of detection, cross-reactivity, and interference testing.
Technological Characteristics
The device uses real-time PCR with fluorogenic TaqMan® probes. It consists of a disposable microfluidic cartridge (PIE) and Sample Buffer Tube. The revogene™ instrument automates sample homogenization, lysis, and amplification. The system is factory-calibrated and uses barcode-based specimen identification. Reagents are dried within the cartridge. The assay is designed for use on the revogene™ platform.
Indications for Use
Indicated for patients with signs and symptoms of pharyngitis. Prescription use only. Follow-up culture required if negative result and symptoms persist, or during acute rheumatic fever outbreaks.
Regulatory Classification
Identification
A Streptococcus spp. nucleic acid-based assay is a qualitative in vitro diagnostic device intended to simultaneously detect and identify various Streptococcus spp. nucleic acids extracted directly from clinical specimens. The device detects specific nucleic acid sequences for organism identification. The identification aids in the diagnosis of diseases caused by bacteria belonging to the genus Streptococcus and provides epidemiological information on these diseases. Pathogenic streptococci are associated with infections, such as sore throat, impetigo (an infection characterized by small pustules on the skin), urinary tract infections, rheumatic fever, and kidney disease.
Special Controls
(b) Classification. Class II (special controls). The special controls for this device are: 1) Premarket notification submissions must include detailed device description documentation, including the device components, ancillary reagents required but not provided, and a detailed explanation of the methodology including primer/probe sequence, design, and rationale for sequence selection. 2) Premarket notification submissions must include detailed documentation from the following analytical and clinical performance studies: Analytical sensitivity (Limit of Detection), reactivity, inclusivity, precision, reproducibility, interference, cross reactivity, carry-over, and cross contamination. 3) Premarket notification submissions must include detailed documentation from a clinical study. The study, performed on a study population consistent with the intended use population, must compare the device performance to results obtained from well-accepted reference methods. 4) Premarket notification submissions must include detailed documentation for device software, including, but not limited to, software applications and hardwarebased devices that incorporate software. 5) Premarket notification submissions must include database implementation methodology, construction parameters and quality assurance protocols, as appropriate. 6) The device labeling must include limitations regarding the need for culture confirmation of negative specimens, as appropriate. 7) A detailed explanation of the interpretation of results and acceptance criteria must be included in the device's 21 CFR 809.10(b)(9) compliant labeling. 8) Premarket notification submissions must include details on an end user device training program that will be offered while marketing the device, as appropriate.
*Classification.* Class II (special controls). The special controls for this device are:(1) Premarket notification submissions must include detailed device description documentation, including the device components, ancillary reagents required but not provided, and a detailed explanation of the methodology including primer/probe sequence, design, and rationale for sequence selection. (2) Premarket notification submissions must include detailed documentation from the following analytical and clinical performance studies: Analytical sensitivity (Limit of Detection), reactivity, inclusivity, precision, reproducibility, interference, cross reactivity, carry-over, and cross contamination. (3) Premarket notification submissions must include detailed documentation from a clinical study. The study, performed on a study population consistent with the intended use population, must compare the device performance to results obtained from well-accepted reference methods. (4) Premarket notification submissions must include detailed documentation for device software, including, but not limited to, software applications and hardware-based devices that incorporate software. (5) Premarket notification submissions must include database implementation methodology, construction parameters, and quality assurance protocols, as appropriate. (6) The device labeling must include limitations regarding the need for culture confirmation of negative specimens, as appropriate. (7) A detailed explanation of the interpretation of results and acceptance criteria must be included in the device's 21 CFR 809.10(b)(9) compliant labeling. (8) Premarket notification submissions must include details on an end user device training program that will be offered while marketing the device, as appropriate.
Predicate Devices
- Xpert Xpress Strep A (K172126)
Related Devices
- K173398 — Xpert Xpress Strep A · Cepheid · Apr 26, 2018
- K172402 — ARIES Group A Strep Assay, ARIES Group A Strep Assay Protocol File Kit · Luminex Corporation · Oct 30, 2017
Submission Summary (Full Text)
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# 510(k) SUBSTANTIAL EQUIVALENCE DETERMINATION DECISION SUMMARY ASSAY AND INSTRUMENT COMBINATION TEMPLATE
A. 510(k) Number:
K183366
B. Purpose for Submission:
To obtain a Substantial Equivalence Determination for the GenePOC Strep A assay
C. Measurand:
Conserved regions of the *Streptococcus pyogenes* bacterial genome
D. Type of Test:
Qualitative real-time Polymerase Chain Reaction (PCR)
E. Applicant:
GenePOC Inc.
F. Proprietary and Established Names:
GenePOC Strep A
G. Regulatory Information:
1. Regulation section:
21 CFR 866.2680: *Streptococcus* spp. nucleic acid-based assay
2. Classification:
Class II
3. Product code:
PGX: Groups A, C and G β-Hemolytic *Streptococcus* Nucleic Acid Amplification System
OOI: Real time nucleic acid amplification system
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4. Panel:
83-Microbiology
H. Intended Use:
1. Intended use(s):
The GenePOC Strep A assay, performed on the revogene instrument, is an automated, qualitative *in vitro* diagnostic test that utilizes real-time polymerase chain reaction (PCR) for the direct detection of *Streptococcus pyogenes* (Group A β-hemolytic *Streptococcus*) nucleic acids from throat swab specimens obtained from patients with signs and symptoms of pharyngitis. The GenePOC Strep A assay is intended for use as an aid in the diagnosis of Group A *Streptococcus* infection.
2. Indication(s) for use:
Same as Intended Use
3. Special conditions for use statement(s):
For prescription use only.
(Limitation) Additional follow-up testing by culture is required if the GenePOC Strep A assay result is negative and clinical symptoms persist, or in the event of an outbreak of acute rheumatic fever (ARF).
4. Special instrument requirements:
The GenePOC Strep A assay is indicated for use with the revogene instrument.
I. Device Description:
The GenePOC Strep A assay is a single-use test for qualitative detection of *Streptococcus pyogenes* (group A *Streptococcus*-GAS) nucleic acids from throat swab specimens obtained from patients with signs and symptoms of pharyngitis. The GenePOC Strep A assay kit is comprised of the disposable microfluidic cartridge (PIE) with Strep A reagents, Sample Buffer Tube (SBT), and Disposable Transfer Tool (DTT). These components are used to suspend the sample, extract, amplify, and detect *Streptococcus pyogenes* (*S. pyogenes*) nucleic acid.
A Process Control (PrC) is also incorporated into each PIE to verify sample processing and amplification steps. The PrC allows for the verification of potential inhibitor substances as well as microfluidic, instrument, or reagent failures. The GenePOC Strep A assay is designed to be used on the revogene instrument, which automates sample homogenization, sample dilution, cell lysis, DNA amplification, and detection of the amplified PCR products.
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Each GenePOC Strep A assay kit provides components for twenty-four (24) tests. User intervention is required for sample preparation, transferring throat swab specimen into the SBT, using the DTT to transfer the sample into the PIE, and loading/unloading the PIE into the revogene carousel. Each PIE is a completely integrated closed device in which a sample is dispensed and processed through different microfluidic chambers and channels that allow for the sample processing and subsequent real-time PCR steps.
Upon completion of a run, the results are computed by the revogene instrument from measured fluorescent signals and embedded calculation algorithms. The possible output results are: positive, negative, indeterminate, or unresolved results. The user removes the used cartridges and disposes of them in normal biological waste. Results may be viewed, printed, transferred, and/or stored by the user.
J. Substantial Equivalence Information:
1. Predicate device name(s):
Xpert Xpress Strep A
2. Predicate 510(k) number(s):
K172126
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3. Comparison with predicate:
| Similarities | | |
| --- | --- | --- |
| Item | Device (K183366) | Predicate (K172126) |
| | GenePOC Strep A assay | Xpert Xpress Strep A Assay |
| Regulation | 21 CFR 866.2680 | Same |
| Product Code | PGX | Same |
| Device Class | Class II | Same |
| Intended Use | The GenePOC Strep A assay, performed on the revogene instrument, is an automated, qualitative in vitro diagnostic test that utilizes real-time polymerase chain reaction (PCR) for the direct detection of Streptococcus pyogenes (Group A β-hemolytic Streptococcus) nucleic acids from throat swab specimens obtained from patients with signs and symptoms of pharyngitis. The GenePOC Strep A assay is intended for use as an aid in the diagnosis of Group A Streptococcus infection. | The Xpert Xpress Strep A Assay, performed on the GeneXpert Instrument Systems, is a rapid, qualitative in vitro diagnostic test for the detection of Streptococcus pyogenes (Group A β-hemolytic Streptococcus, Strep A) in throat swab specimens from patients with signs and symptoms of pharyngitis. The Xpert Xpress Strep A Assay utilizes an automated real-time polymerase chain reaction (PCR) to detect Streptococcus pyogenes DNA. |
| Analyte | Group A Streptococcus | Same |
| Measurand | Conserved region of the S. pyogenes genome | Same |
| Specimen Type | Throat swab in Liquid Stuart or Liquid Amies Transport Medium | Throat swab in liquid Amies medium |
| Assay Format | Single-use; Automated DNA extraction, amplification and detection | Same |
| External Controls | Materials available commercially | Same |
| Result | Qualitative, Automated | Same |
| Differences | | |
| --- | --- | --- |
| Item | Device (K183366) | Predicate (K172126) |
| | GenePOC Strep A assay | Xpert Xpress Strep A Assay |
| Instrument System | revogene | GeneXpert Dx, GeneXpert Infinity-48s or GeneXpert Infinity-80 instrument systems. |
K. Standard/Guidance Document Referenced (if applicable):
- CLSI Guideline EP25-A, Evaluation of Stability of In Vitro Diagnostic Reagents; Approved Guideline. Wayne, PA: Clinical and Laboratory Standards Institute; 2009.
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- CLSI Guideline EP05-A3, Evaluation of Precision of Quantitative Measurement Procedures; Approved Guideline-Third Edition. Wayne, PA: Clinical and Laboratory Standards Institute; 2014.
- CLSI Guideline EP17-A2, Evaluation of Detection Capability for Clinical Laboratory Measurement Procedures; Approved Guideline. Second Edition. Wayne, PA: Clinical and Laboratory Standards Institute; 2012.
L. Test Principle:
The revogene automates and integrates nucleic acid extraction, amplification, and detection of the target sequence in complex samples using real-time PCR. A throat swab specimen is collected using a rayon tipped swab. The specimen is transferred into the SBT containing 1.5 ml of sample buffer, and the swab stem is broken. After a 15-second vortex step at maximal speed, the inoculated sample buffer is transferred into the GenePOC Strep A PIE using the DTT. The loaded GenePOC Strep A PIE is placed into the revogene for further sample processing. No operator intervention is necessary once the PIE is loaded onto the revogene instrument.
Each GenePOC Strep A PIE is a completely integrated and self-contained device. Each sample is sequentially transferred by centrifugation from one microfluidic chamber to the next and all reagents specific for the PCR reaction are incorporated and dried within the PCR wells. The stepwise process includes sample homogenization, lysis of cells, sample dilution, and DNA extraction followed by the subsequent real-time PCR steps within one PCR well in the cartridge. An internal PrC is contained in the homogenization chamber and is therefore present in every test to verify critical steps of the analytical process as well as system or reagent failures. The amplified products are detected in real-time from measured fluorescent signals and embedded calculation algorithms. Results may be viewed, printed, transferred, and/or stored by the user using target-specific TaqMan chemistry-based probes. An Early Positive Result Outcome (E-PRO) feature provides positive results if the signal from the target reaches a predetermined threshold before the full PCR cycles have been completed. The results are computed by the system. Results may be viewed, printed, transferred, and/or stored by the user.
M. Performance Characteristics (if/when applicable):
1. Analytical performance:
a. Precision/Reproducibility:
Site-to-Site Reproducibility
The reproducibility of the GenePOC Strep A assay between sites was evaluated in a study performed by two operators at each of three sites over a period of five days. Each operator tested a blinded panel of S. pyogenes positive (two different levels based on multiples of the LoD) and negative samples using the same lot of reagents. Ninety (90) data points were collected for each positive panel member, and (120) data points were collected for the negative panel member. The panels were prepared in Sample Buffer
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(SB) using S. pyogenes-negative throat swab matrix and two S. pyogenes strains (ATCC 12344 and ATCC 19615). No differences in reproducibility between sites or operators were observed for the panel members.
The results of the study demonstrated acceptable reproducibility from site-to-site at target levels close to the limit of detection (LoD) of the assay (Table 1). No unresolved results were observed in this study. Across the Reproducibility Study (site-to-site and within site studies), eight samples were indeterminate (IND) for a rate of $1.6\%$ (8/500). The samples were repeated and yielded a reportable result after repeat for a final IND rate of $0\%$ . Each Positive External Control and Negative External Control tested during the Reproducibility Study yielded the expected results.
Table 1. Summary of Results from the GenePOC Strep A Assay Site-to-Site Reproducibility Study, Stratified by Site and Panel Member.
| Level | Positive/Number (%) | | | |
| --- | --- | --- | --- | --- |
| | Site 1 | Site 2 | Site 3 | Overall |
| Moderate Positive 3X LoD1 | 30/30 (100%) | 30/30 (100%) | 30/30 (100%) | 90/90 (100%) |
| Low Positive 1-2X LoD2 | 30/30 (100%) | 30/30 (100%) | 30/30 (100%) | 90/90 (100%) |
| Negative | 0/40 (0.0%) | 0/40 (0.0%) | 0/40 (0.0%) | 0/120 (0.0%) |
1 LoD: Limit of Detection for S. pyogenes strain ATCC 19615
3X LoD = 3.0 x $10^{3}$ CFU/ml of SB; 1X LoD = 1.0 x $10^{3}$ CFU/ml of SB
2 LoD: Limit of Detection for S. pyogenes strain ATCC 12344
1-2X LoD= 650 CFU/ml of SB; 1X LoD= 333 CFU/ml of SB
# Within Laboratory Precision/Repeatability
Within laboratory precision/repeatability of the GenePOC Strep A assay was evaluated by two operators who tested a panel of samples on a single revogene instrument over a period of 20 days. Eighty (80) replicates were tested for each panel member. The panel members were the same as those used in the Site-to-Site Reproducibility Study, above. The results of the study demonstrated acceptable repeatability and precision from day-to-day with target levels even close to the LoD of the assay (Table 2). One unresolved (UNR) was observed. This sample was repeated for a valid result.
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Table 2. Summary of Results from the Within Laboratory Precision/ Repeatability Study for the GenePOC Strep A Assay
| Level | Positive/Tested (%) |
| --- | --- |
| Moderate Positive | 80/80 |
| 3X LoD1 | (100%) |
| Low Positive | 80/80 |
| 1-2X LoD2 | (100%) |
| Negative | 2/80 |
| | (2.5%) |
1 LoD: Limit of Detection for S. pyogenes strain ATCC 19615
3X LoD: $3.0 \times 10^{3}$ CFU/ml of SB; 1X LoD: $1.0 \times 10^{3}$ CFU/ml of SB
2 LoD: Limit of Detection for S. pyogenes strain ATCC12344
1-2X LoD: 650 CFU/ml of SB
# Lot-to-Lot Reproducibility
The lot-to-lot reproducibility of the GenePOC Strep A assay was evaluated by testing a panel of S. pyogenes positive and negative samples in negative throat matrix and SB with each of three lots of reagents over a period of five days. Ninety (90) replicates per positive panel member were tested, and a total of 120 replicates were tested for the negative panel member. All the panel members were the same as those used in the Reproducibility and Precision/Repeatability Studies above. The results are summarized in Table 3 and show acceptable performance with each lot of reagents.
Table 3. Summary of Results from the GenePOC Strep A assay Lot-to-Lot Reproducibility Study, Stratified by Reagent Lot and Panel Member
| Level | Positive/Tested (%) | | | |
| --- | --- | --- | --- | --- |
| | Lot 1 | Lot 2 | Lot 3 | Overall |
| Moderate Positive | 30/30 | 30/30 | 30/30 | 90/90 |
| 3X LoD1 | (100%) | (100%) | (100%) | (100%) |
| Low Positive | 30/30 | 29/30 | 29/30 | 88/90 |
| 1-2X LoD2 | (100%) | (96.7%) | (96.7%) | (97.8%) |
| Negative | 0/40 | 0/40 | 0/40 | 0/120 |
| | (0.0%) | (0.0%) | (0.0%) | (0.0%) |
1 LoD: Limit of Detection for S. pyogenes strain ATCC 19615
3X LoD: $3.0 \times 10^{3}$ CFU/ml of SB; 1X LoD: $1.0 \times 10^{3}$ CFU/ml of SB
2 LoD: Limit of Detection for S. pyogenes strain ATCC 12344
1-2X LoD: 650 CFU/ml of SB
b. Linearity/assay reportable range:
Not Applicable.
c. Traceability, Stability, Expected values (controls, calibrators, or methods):
Internal process control (PrC):
Each PIE contains a $\mathrm{PrC}$ that controls for amplification inhibition, assay reagents, and sample processing effectiveness.
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A sample in which there is an amplification/detection failure of the PrC and the GAS target DNA is reported as “Unresolved” (UNR) and must be re-tested.
## External Controls
Two External Controls are to be prepared by the end user. GenePOC recommends using a $0.5 \pm 0.05$ McFarland suspension of a *S. pyogenes* commercially available strain (e.g., ATCC 12344) diluted 1/100 in saline as a Positive External Control and a $0.5 \pm 0.05$ McFarland suspension of a *Streptococcus salivarius* commercially available strain (e.g., ATCC 13419) as a Negative External Control. These are recommended, but not required, to allow the user to select the most appropriate for their laboratory quality control program.
## Sample Stability in PIE
The GenePOC Strep A assay PIE (microfluidic cartridge) stability study was validated using positive samples comprised of the *S. pyogenes* (GAS) strain ATCC 12344 (at 3X LoD) and negative samples in presence of *S. pyogenes*-negative throat matrix. The study was conducted using three (3) GenePOC Strep A kit lots, three (3) replicates of *S. pyogenes* and negative samples (one replicate per kit lot), and five (5) revogene instruments. Samples were tested across four (4) time points: 0 minutes (T0), 60 minutes (T60), 120 minutes (T120), and 180 minutes (T180) to validate the PIE stability study at $25^{\circ}\mathrm{C}$ after opening the PIE pouch. No unresolved (UNR) samples were recorded in the PIE stability study. One IND was observed for an initial IND rate of $2.8\%$; however, this sample was repeated and yielded reportable results for a final $0\%$ IND rate. The results of the study support the recommended maximum interval of one hour from the opening of the PIE pouch and sample addition into the GenePOC Strep A assay PIE to the processing on the revogene instrument.
## Specimen Stability
The stability of throat swab specimens collected with rayon swabs in Liquid Stuart transport media was evaluated analytically with the GenePOC Strep A assay. The time periods tested in this study were used to establish specimen stability between: i) throat swab specimen collection and SBT inoculation and ii) SBT inoculation and loading of the GenePOC Strep A PIE prior to initiation of the revogene run. The stability study was conducted by testing positive samples comprised of the *S. pyogenes* (GAS) strain ATCC 12344 (3X LoD) and negative samples in the presence of *S. pyogenes*-negative throat matrix. The cumulative shelf life stability (nested stability) corresponds to the period of time from swab specimen collection up to the initiation of the revogene assay run. The study included three (3) lots GenePOC Strep A assay kits. For each sample type (positive or negative), and for each time point, a total of twelve (12) swabs were inoculated. Study results support throat swab specimen storage at $25 \pm 2^{\circ}\mathrm{C}$ up to two (2) days or $2 - 8^{\circ}\mathrm{C}$ for up to seven (7) days. SBT inoculated with throat swab specimens may be stored up to three (3) days at $25 \pm 2^{\circ}\mathrm{C}$ or $2 - 8^{\circ}\mathrm{C}$ for up to seven (7) days prior testing with the GenePOC Strep A assay.
## Swab Specimen Stability-Various Swabs
This study was designed to evaluate the cumulative stability of throat swabs specimens
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when collected and conserved with Rayon Liquid Amies, Polyester Liquid Stuart, Polyester Liquid Amies and the inoculated SBT used with the GenePOC Strep A assay at defined temperature of $25 \pm 2^{\circ}\mathrm{C}$ and $2 - 8^{\circ}\mathrm{C}$. The study was executed by testing *S. pyogenes* strain ATCC 12344 at 3X LoD and negative samples in presence of *S. pyogenes*-negative throat matrix in SB. The study included three (3) to four (4) lots GenePOC Strep A assay kits for each type of swab. For each sample type (positive or negative), and for each time point, a total of twelve (12) swabs were inoculated. Study results support throat swab specimen stability for GenePOC Strep A assay at $25 \pm 2^{\circ}\mathrm{C}$ up to two (2) days or $2 - 8^{\circ}\mathrm{C}$ for up to seven (7) days; SBT inoculated with throat swab specimens may be stored up to three (3) days at $25 \pm 2^{\circ}\mathrm{C}$ or $2 - 8^{\circ}\mathrm{C}$ for up to seven (7) days prior testing with the GenePOC Strep A assay, as confirmed with the rayon swab in Liquid Amies transport media and Polyester swab in Liquid Stuart and Liquid Amies transport media.
## Reagent Stability
The shelf life of the GenePOC Strep A assay reagents was evaluated in a real-time stability study performed on three lots of reagents that were stored under different conditions. GenePOC claims a shelf life of 2 months.
## d. Detection limit:
### Limit of Detection
The Limit of Detection (LoD) of the GenePOC Strep A assay was estimated for three strains of *S. pyogenes* by testing various dilutions of enumerated cell stocks in *S. pyogenes*-negative throat swab matrix. The LoD for each strain was then confirmed by testing a further 20 replicates at the lowest target level that produced $100\%$ positive results. The LoD was defined as the lowest concentration tested at which $\geq 95\%$ of assay replicates produced positive results. For ATCC 19615, ATCC 12344, and ATCC 700942, the LoD was determined to be $1.0 \times 10^{3}$ CFU/ml, 333 CFU/ml, and $1.3 \times 10^{3}$ CFU/ml, respectively. No unresolved (UNR) results were obtained during the study. Eighteen (18) out of 595 samples obtained IND results during the GenePOC Strep A assay LoD determination study for an initial IND rate of $3.0\%$. These samples were repeated and all yielded reportable results for a final IND rate of $0\%$.
### Inclusivity (Analytical Reactivity)
The inclusivity of the GenePOC Strep A assay was evaluated by testing nine additional strains of *S. pyogenes* in negative throat matrix (Table 4). Eight of the nine strains produced 9/9 positive results at a concentration of 999 CFU/ml (3X the LoD for ATCC12344). Strain ATCC 49399 gave one false negative result at this target level but produced 9/9 positive results at $1.67 \times 10^{3}$ CFU/ml, equivalent to 5X LoD for ATCC 12344. These results are acceptable.
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Table 4. Strains of S. pyogenes used to Evaluate the Inclusivity of the GenePOC Strep A assay
| Origin | Strain Number | Emm-Type1 |
| --- | --- | --- |
| USA | ATCC 12384 | 3 |
| Unknown | ATCC 49399 | N/A |
| Sweden | CCUG 33409 | N/A |
| Sweden | CCUG 39158 | N/A |
| Sweden | CCUG 53553 | N/A |
| New Zealand | ATCC 700952 | 92 |
| Unknown | ATCC 700294 | 1 |
| England | ATCC 12357 | 18 |
| Sweden | CCUG 65322 | N/A |
For the LoD Study, the Emm-type for ATCC 19615, ATCC 12344, and ATCC 700942 were reported as 80, 1, and 82.
# e. Analytical specificity:
# Cross-reactivity Study
The analytical specificity of the GenePOC Strep A assay was evaluated by testing a 50-member panel [(42) bacteria, (1) yeast, (6) viruses, (1) human gDNA] with organisms that might be found in throat swab specimens (Table 5). Each panel member was tested in triplicate in SB with S. pyogenes-negative throat matrix. Bacteria and yeast were tested at $\geq 10^{6}$ CFU/ml while viral nucleic acid and human gDNA samples were tested at a concentration $\geq 10^{5}$ cp/ml. No cross-reactivity was observed. No UNR were observed. Three IND were produced for an initial IND rate of $1.9\%$ ; all repeats yielded valid results. These study results are acceptable.
Table 5. Organisms Tested for Potential Cross-Reaction in the GenePOC Strep A Assay
| Name | Identification | Name | Identification |
| --- | --- | --- | --- |
| Bacteria and Yeasts | | | |
| Acinetobacter Iwoffii | ATCC 15309 | Klebsiella pneumoniae | ATCC 27736 |
| Arcanobacterium haemolyticum | ATCC BAA-1784 | Lactococcus lactis | ATCC 19435 |
| Bacillus cereus | ATCC 14579 | Legionella jordanis | ATCC 33623 |
| Bordetella pertussis | ATCC 9797 | Legionella micdadei (Tatlockia micdadei) | CCUG 31229 |
| Burkholderia cepacia | ATCC 25416 | Legionella pneumophila | ATCC 33152 |
| Corynebacterium diphtheriae | ATCC 13812 | Moraxella catarrhalis | ATCC 25238 |
| Enterococcus faecalis | ATCC 19433 | Neisseria gonorrhoeae | ATCC 43069 |
| Escherichia coli | ATCC 11775 | Neisseria subflava | ATCC 49275 |
| Fusobacterium necrophorum | ATCC 25286 | Parvimonas micra | ATCC 33270 |
| Haemophilus influenza | ATCC 9006 | Pseudomonas aeruginosa | ATCC 35554 |
| Serratia marcescens | ATCC 13880 | Streptococcus dysgalactiae subsp. dysgalactiae | ATCC 9926 |
| Staphylococcus aureus | ATCC 33592 | Streptococcus dysgalactiae subsp. dysgalactiae | ATCC 43078 |
| Staphylococcus epidermidis | ATCC 14990 | Streptococcus gordonii | ATCC 10558 |
| Stenotrophomonas maltophilia | ATCC 13637 | Streptococcus intermedius | ATCC 27335 |
| Streptococcus agalactiae | ATCC 12403 | Streptococcus mitis | NCIMB 13770 |
| Streptococcus anginosus | ATCC 33397 | Streptococcus mutans | ATCC 25175 |
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| Name | Identification | Name | Identification |
| --- | --- | --- | --- |
| Streptococcus anginosus subsp. whileyi | CCUG 39159 | Streptococcus oralis | ATCC 6249 |
| Streptococcus bovis | ATCC 33317 | Streptococcus pneumoniae | ATCC 49619 |
| Streptococcus canis | ATCC 43496 | Streptococcus salivarius | ATCC 13419 |
| Streptococcus constellatus subsp. viborgensis | CCUG 62387 | Streptococcus sanguinis | ATCC 10556 |
| Streptococcus suis | CCUG 7984 | Veillonella parvula | ATCC 10790 |
| | | Candida albicans | ATCC 10231 |
| Viruses and Human DNA | | | |
| Adenovirus Type 1 | ATCC VR-1D | Influenza A/Aichi/2/68/H3N2 | ATCC VR-1680D |
| Influenza B/Taiwan/2/62 | ATCC VR-1735D | Parainfluenza virus 4b | ATCC VR-1377D |
| Rhinovirus Type 17 | ATCC VR-1663D | Human DNA | N/A |
| Adenovirus type 11 (Slobitski) | ATCC VR-12D | | |
# Bioinformatic Analysis
The in silico specificity of the GenePOC Strep A assay was validated through analysis of the assay's primers and probe sequences specific for the detection of S. pyogenes. The in silico study suggested that non-specific amplifications or detection will not occur with the primers and probe of the GenePOC Strep A assay used to target the S. pyogenes sequence. The primers and probes have been proven to be specific to their respective target following Basic Local Alignment Search Tool (BLAST) analysis. No target other than S. pyogenes was found to have a significant level of homology with the S. pyogenes primers and probe. In addition, it was concluded that the S. pyogenes target primers and probe will not interact with the PrC sequence. Further, the PrC primers did not demonstrate cross-reactivity with other organisms.
# Contamination Study
The potential for false-positive results with the GenePOC Strep A assay due to within-run or between-run cross contamination was evaluated by testing an alternating series of $S$ pyogenes "high positive" and negative samples in successive instrument runs. The high positive samples contained $S$ pyogenes at a concentration of $10^{6}$ CFU/ml in the presence of $S$ pyogenes-negative throat matrix. Negative samples comprised $S$ pyogenes-negative throat matrix without GAS. The expected results were obtained for all $S$ pyogenes positive and negative samples (40/40 each). No false positive results were detected in the within-run cross-contamination study ( $n = 80$ ) or in the between-run carry-over study ( $n = 80$ ). No UNR or IND results were obtained throughout the cross-contamination and carry-over study. These results are acceptable.
# f. Assay cut-off:
Thresholds and cut-offs for the GenePOC Strep A assay are embedded within the Assay Definition File that also encodes the instrument settings required to perform the test. Both native (throat swab specimens) and contrived samples were used to set the thresholds of positivity and negativity of the GenePOC Strep A assay. The assay cut-off was determined by testing $n = 509$ native and contrived samples.
# g. Assay interference:
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# Potentially Interfering Substances
The potential for interference with the GenePOC Strep A assay was evaluated with (19) endogenous and exogenous substances that may be present in throat swab specimens (Table 6). Two (2) strains of S. pyogenes (ATCC 12344 and ATCC 19615) were tested at a moderate load (3X LoD, 999 CFU/ml and 3000 CFU/ml of SB, respectively) in the presence of potentially interfering substances and S. pyogenes-negative throat matrix. Negative samples were also tested in the presence of potentially interfering substances to evaluate the impact on the $\mathrm{PrC}$ . The potentially interfering substances were tested at the highest concentration that may be encountered in a throat swab specimen. Results demonstrated that Analgesic/Antipyretic (Tylenol), Nonsteroidal anti-inflammatory drug (Aspirin, Advil), Bronchodilator (Albuterol sulfate), Whole Blood, and Mucin had a potentially inhibitory effect on the detection of $\mathrm{PrC}$ or S. pyogenes when any of these substances was present in SBT at a concentration of $4.3\%$ (w/v or v/v). At $0.4\%$ (w/v), Tylenol, Advil, Albuterol sulfate and Mucin showed no reportable interference with the GenePOC Strep A assay. When tested at for at $0.1\%$ , no interference was observed with Aspirin (w/v) and Whole Blood (v/v). The potential for certain substances to interfere with the GenePOC Strep A assay is noted in the device labeling.
Table 6. Substances Evaluated for Potential Interference with the GenePOC Strep A Assay
| Substance | Test Concentration (in SBT) |
| --- | --- |
| Endogenous Substances | |
| Human Saliva | 4.3% (v/v) |
| Whole Blood | 4.3% (v/v) |
| Mucin | 4.3% (v/v) |
| Exogenous Substances | |
| Tylenol | 4.3% (w/v) |
| Aspirin | 4.3% (w/v) |
| Advil | 4.3% (w/v) |
| Albuterol sulfate | 4.3% (w/v) |
| Zicam | 4.3% (w/v) |
| Hall Menthoy-Lyptus | 4.3% (w/v) |
| Cepacol Extra Strength (Cough lozenge) | 4.3% (w/v) |
| Sucrets Complete (Cough lozenge) | 4.3% (w/v) |
| Crest Pro-Health Advanced (toothpaste) | 0.56% (w/v) |
| Crest Scope Classic (Mouthwash) | 4.3% (v/v) |
| Listerine Ultraclean (Mouthwash) | 4.3% (v/v) |
| NyQuil Complete | 4.3% (v/v) |
| Dimetapp (Cough Syrup) | 4.3% (v/v) |
| Robitussin Cough Control Extra Strength | 4.3% (v/v) |
| Robitussin Total Cough, Cold & Flu Extra Strength Nighttime | 4.3% (v/v) |
| Benadryl Allergy Elixir (Antihistamines) | 4.3% (v/v) |
# Microbial Interference
The potential for interference with the GenePOC Strep A assay was investigated using (30) non-targeted microorganisms genetically close to the assay analytes, flora potentially
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found in the mouth/throat/respiratory tract, and the most frequent throat and upper respiratory tract infection causative agents. Two (2) S. pyogenes strains, ATCC 12344 and ATCC 19615, were tested at 3X LoD (999 CFU/ml and 3000 CFU/ml of SB, respectively) in triplicate in the presence of potentially interfering non-targeted microorganisms and S. pyogenes-negative throat matrix. Negative samples were also tested in the presence of potentially interfering non-targeted microorganisms to evaluate the impact on the PrC. None of the thirty (30) organisms present at $10^{6}$ CFU/ml of SB for bacteria and yeast and $10^{5}$ copies/ml of SB for viruses interfered with the detection of PrC or with S. pyogenes strain ATCC 19615. A combinatory effect of Streptococcus mutans, Streptococcus oralis, Streptococcus salivarius and Moraxella catarrhalis at $\geq 10^{6}$ CFU/ml of SB (pool), or the presence of Streptococcus dysgalactiae subsp. dysgalactiae at $\geq 10^{6}$ CFU/ml of SB, may have an inhibitory effect on the detection of S. pyogenes ATCC 12344. When each bacterium from the pool (Streptococcus mutans, Streptococcus oralis, Streptococcus salivarius and Moraxella catarrhalis) was tested individually at the same load ($1 \times 10^{6}$ CFU/ml of SB) in presence of S. pyogenes ATCC 12344, none interfered with the detection of S. pyogenes. After additional testing, S. dysgalactiae subsp. dysgalactiae strain ATCC 43078 no longer showed interference with S. pyogenes ATCC 12344 at a load of $1 \times 10^{5}$ CFU/ml of SB. These results are noted in the device labeling, where appropriate.
2. Comparison studies:
a. Method comparison with predicate device:
Not Applicable.
b. Matrix comparison:
Not Applicable.
3. Clinical studies:
a. Clinical Sensitivity:
The performance of the GenePOC Strep A assay was evaluated in a prospective multicenter study that was conducted at geographically diverse clinical trial sites. These included: (7) collection/testing sites [(5) US sites and (2) Canadian sites] and (1) dedicated reference method center (across the US and Canada). Throat swab specimens were collected from patients with signs and symptoms of pharyngitis. Samples were tested with both the Reference Method (culture) and the GenePOC Strep A assay.
The reference culture procedure was performed within 48 hours of specimen collection and comprised inoculation of BAP with one swab of a dual throat swab set and incubation of plates under anaerobic conditions for up to 48 hours. Culture plates that did not exhibit $\beta$-hemolytic colonies after 48 hours were recorded as negative for Group A Streptococcus. Colonies morphologically resembling S. pyogenes were isolated on a new
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BAP and identified by catalase, gram-stain, Latex agglutination, and Matrix-Assisted Laser Desorption/Ionization Time-Of-Flight Mass Spectrometry (MALDI-TOF).
Seven hundred and sixty-seven (767) specimens were initially enrolled in the study. Of these, 163 were excluded from the analysis of performance due to: failure to comply with the revogene testing procedures—delay in pouch opening or transfer to SBT (3), exceeded the delay in shipment (13), did not meet inclusion criteria (23), did not have complete testing results (27), and failure to comply with the reference culture protocol (97). A total of 604 specimens were used to evaluate performance of the assay (Table 7). For UNR samples, there was an initial UNR rate of $0.5\%$ (3/610) with a final UNR rate of $0.2\%$ (1/610) after re-testing. For IND samples, there was an initial IND rate of $1.1\%$ (7/610) with a final rate of $0.8\%$ (5/610) after re-testing.
Table 7. GenePOC Strep A Clinical Performance vs Reference Culture
| | Reference Culture | | | |
| --- | --- | --- | --- | --- |
| | | Positive | Negative | Total |
| GenePOC Strep A assay | Positive | 151 | 241 | 175 |
| | Negative | 32 | 426 | 429 |
| | Total | 154 | 450 | 604 |
| Sensitivity | | 151/154 = 98.1% (95% CI: 94.4-99.3%) | | |
| Specificity | | 426/450 = 94.7% (95% CI: 92.2-96.4%) | | |
| Positive Predictive Value | | 151/175 = 86.3% (95% CI: 80.4-90.6%) | | |
| Negative Predictive Value | | 426/429 = 99.3% (95% CI: 98.0-99.8%) | | |
95% CI: Two-sided 95% score confidence interval
1 17/24 specimens were positive by an alternative PCR/bi-directional sequencing assay; 1/24 samples was inconclusive.
2 1/3 specimens were positive by an alternative PCR/bi-directional sequencing assay; 2/3 samples were inconclusive.
The performance of the GenePOC Strep A assay at each clinical site in comparison to the reference culture method is shown in Table 8. The GenePOC Strep A assay sensitivity and specificity for the six hundred and four (604) prospective specimens were $98.1\%$ (151/154) and $94.7\%$ (426/450) respectively. Overall, performance was determined to be acceptable.
Table 8. Performance of the GenePOC Strep A Assay in Comparison to the Reference Culture Method, Stratified by Clinical Site
| (Site) | Culture Positive (%) | Sensitivity % (95% CI) | Specificity % (95% CI) |
| --- | --- | --- | --- |
| 1 | 16/69 (23.2%) | 16/16 = 100% (95% CI: 80.6% - 100%) | 52/53 = 98.1% (95% CI: 90.1% - 99.7%) |
| 2 | 18/67 (26.9%) | 18/18 = 100% (95% CI: 82.4% - 100%) | 44/49 = 89.8% (95% CI: 78.2% - 95.6%) |
| 3 | 2/42 (4.8%) | 2/2 = 100% (95% CI: 34.2% - 100%) | 39/40 = 97.5% (95% CI: 87.1% - 99.6%) |
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| 4 | 59/123 (48.0%) | 58/59 = 98.3% (95% CI: 91.0% - 99.7%) | 58/64 = 90.6% (95% CI: 81.0% - 95.6%) |
| --- | --- | --- | --- |
| 5 | 30/178 (16.9%) | 28/30=93.3% (95% CI: 78.7%-98.2%] | 146/148=98.6% (95% CI: 95.2%-99.6%) |
| 6 | 13/49 (26.5%) | 13/13=100% (95% CI: 77.2%-100%) | 32/36=88.9% (95% CI: 74.7%-95.6%) |
| 7 | 16/76 (21.1%) | 16/16=100% (95% CI: 80.6%-100%) | 55/60=91.7% (95% CI: 81.9%-96.4%) |
b. Clinical specificity:
Refer to Section M(3)(a), above.
c. Other clinical supportive data (when a. and b. are not applicable):
Not applicable.
4. Clinical cut-off:
Not applicable.
5. Expected values/Reference range:
The performance of the GenePOC Strep A assay was evaluated in a prospective clinical study. Collection/testing for S. pyogenes with the GenePOC Strep A assay was performed at (7) geographically diverse clinical sites. A separate site was used as a dedicated Reference Method Center. The overall prevalence of S. pyogenes (Group A Streptococcus) in throat swab specimens was $28.9\%$ (175/604) as determined by the GenePOC Strep A assay and $25.5\%$ (154/604) as determined by culture. The prevalence of S. pyogenes as determined by the GenePOC Strep A assay (Table 9) and the Reference Method (Table 10), is shown below stratified by the age of the subjects.
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Table 9. Prevalence of S. pyogenes Positive Subjects by Age (GenePOC Strep A Assay)
| Age | Number | GenePOC Strep A Assay Positive | % Prevalence |
| --- | --- | --- | --- |
| Age Class | | | |
| >24 months-12 years | 278 | 106 | 38.1% |
| 13-21 years | 80 | 15 | 18.8% |
| 22-64 years | 235 | 51 | 21.7% |
| ≥65 years | 11 | 3 | 27.3% |
| Total | 604 | 175 | 29.0% |
Table 10. Prevalence of S. pyogenes Positive Subjects by Age (Reference Method)
| Age | Number | Reference Method Positive | % Prevalence |
| --- | --- | --- | --- |
| Age Class | | | |
| >24 months-12 years | 278 | 94 | 33.8% |
| 13-21 years | 80 | 14 | 17.5% |
| 22-64 years | 235 | 44 | 18.7% |
| ≥65 years | 11 | 2 | 18.2% |
| Total | 604 | 154 | 25.5% |
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17
N. Instrument Name:
revogene
O. System Descriptions:
1. Modes of Operation:
Does the applicant’s device contain the ability to transmit data to a computer, webserver, or mobile device?
Yes ☐ X or No ☐
Does the applicant’s device transmit data to a computer, webserver, or mobile device using wireless transmission?
Yes ☐ or No ☐ X
2. Software:
FDA has reviewed applicant’s Hazard Analysis and software development processes for this line of product types:
Yes ☐ X or No ☐
3. Specimen Identification:
Barcodes are used to identify patient specimens. The GenePOC Strep A assay’s SBT and PIE are both pre-labeled with a unique barcode to identify both specimen and assay. The instrument has two barcode readers to identify reagents and patient specimens. It provides traceability of the sample ID to the PIE ID, SBT ID, and assay ID.
4. Specimen Sampling and Handling:
User intervention is required for discharging the patient sample into the SBT, transferring the sample into the microfluidic cartridge using the DTT, and for loading the microfluidic cartridge into the revogene. All further specimen handling is automated.
5. Calibration:
The system is factory calibrated by the manufacturer. The calibration is verified annually. Upon the verification, maintenance is performed if required.
6. Quality Control:
Refer to Section M(1)(c).
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P. Other Supportive Instrument Performance Characteristics Data Not Covered In The "Performance Characteristics" Section above:
Not applicable.
Q. Proposed Labeling:
The labeling supports the finding of substantial equivalence for this device.
R. Conclusion:
The submitted information in this premarket notification is complete and supports a substantial equivalence decision.
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