(90 days)
The Panther Fusion GI Expanded Bacterial Assay is a multiplex real-time PCR in vitro diagnostic test for the rapid and qualitative detection and differentiation of Yersinia enterocolitica, Vibrio (V. parahaemolyticus, V. vulnificus, V. cholerae), Escherichia coli O157, and Plesiomonas shigelloides. Nucleic acids are isolated and purified from Cary-Blair preserved stool specimens collected from individuals exhibiting signs and symptoms of gastroenteritis.
This assay is intended to aid in the differential diagnosis of Yersinia enterocolitica, Vibrio (V. parahaemolyticus, V. vulnificus, V. cholerae), Escherichia coli O157, and Plesiomonas shigelloides infections. The results of this assay should be used in conjunction with clinical presentation, laboratory findings, and epidemiological information and should not be used as the sole basis for diagnosis, treatment, or other patient management decisions. Positive results do not rule out coinfection with other organisms that are not detected by this test and may not be the sole or definitive cause of patient illness. Negative results in the setting of clinical illness compatible with gastroenteritis may be due to infection by pathogens that are not detected by this test, or non-infectious causes such as ulcerative colitis, irritable bowel syndrome, or Crohn's disease. This assay is designed for use on the Panther Fusion System.
The Panther Fusion GI Expanded Bacterial Assay is a multiplex real-time PCR in vitro diagnostic test for the rapid and qualitative detection and differentiation of Yersinia enterocolitica, Vibrio (V. parahaemolyticus, V. vulnificus, V. cholerae), Escherichia coli O157, and Plesiomonas shigelloides. Nucleic acids are isolated and purified from Cary-Blair preserved stool specimens collected from individuals exhibiting signs and symptoms of gastroenteritis.
The Panther Fusion System fully automates specimen processing, including sample lysis, nucleic acid capture, amplification, and detection for the Panther Fusion GI Expanded Bacterial Assay. Nucleic acid capture and elution takes place in a single tube on the Panther Fusion System. The eluate is transferred to the Panther Fusion System reaction tube containing the assay reagents. Multiplex real-time PCR is then performed for the eluted nucleic acid on the Panther Fusion System.
Sample processing: Prior to processing and testing on the Panther Fusion System, specimens are transferred to an Aptima Multitest tube containing specimen transport media (STM) that lyses the cells, releases target nucleic acid, and protects them from degradation during storage.
Nucleic acid capture and elution: An internal control (IC-B) is added automatically to each specimen via the working Panther Fusion Capture Reagent-B (wFCR-B) to monitor for interference during specimen processing, amplification, and detection caused by reagent failure or inhibitory substances. Specimens are first incubated in an alkaline reagent (FER-B) to enable cell lysis. Nucleic acid released during the lysis step hybridizes to magnetic particles in the wFCR-B. The capture particles are then separated from residual specimen matrix in a magnetic field by a series of wash steps with a mild detergent. The captured nucleic acid is then eluted from the magnetic particles with a reagent of low ionic strength (Panther Fusion Elution Buffer).
Multiplex PCR amplification and fluorescence detection: Lyophilized single unit dose reaction master mix is reconstituted with the Panther Fusion Reconstitution Buffer I and then combined with the eluted nucleic acid into a reaction tube. Panther Fusion Oil reagent is added to prevent evaporation during the PCR reaction. Target-specific primers and probes then amplify targets via polymerase chain reaction while simultaneously measuring fluorescence of the multiplexed targets. The Panther Fusion System compares the fluorescence signal to a predetermined cut-off to produce a qualitative result for the presence or absence of each analyte.
The analytes and the channel used for their detection on the Panther Fusion System are summarized in the table below:
| Analyte | Gene Targeted | Instrument Channel |
|---|---|---|
| Yersinia enterocolitica | InvA (Invasive antigen A) | FAM |
| Vibrio parahaemolyticus | gyrB (Gyrase B) | HEX |
| Vibrio vulnificus | gyrB (Gyrase B) | HEX |
| Vibrio cholerae | ompW (Outer Membrane Protein W) | HEX |
| Escherichia coli O157 | rfbE (Perosamine synthase-O-antigen) | ROX |
| Plesiomonas shigelloides | hugA (Heme utilization gene A) | RED647 |
| Internal Control | Not Applicable | RED677 |
Assay Components: The assay components configuration for the Panther Fusion GI Expanded Bacterial Assay is analogous to the Panther Fusion Respiratory Assays. The reagents required to perform the Panther Fusion GI Expanded Bacterial Assay are packaged and sold separately. There are 7 boxes containing 9 reagents which are required for sample processing. The Panther Fusion GI Expanded Bacterial Assay requires one ancillary kit and one specimen collection kit, neither of which are provided with the assay and can be acquired separately:
- Aptima Assay Fluids Kit (303014)
- Aptima Multitest Swab Specimen Collection Kit (PRD-03546)
Table 1: Reagents Required to Perform the Panther Fusion GI Expanded Bacterial Assay
| Box | Components Description |
|---|---|
| Refrigerated Box | Panther Fusion GI Expanded Bacterial Assay Cartridges |
| Room Temperature Box | Panther Fusion Extraction Reagent-B• Panther Fusion Capture Reagent-B bottles• Panther Fusion Enhancer Reagent-B bottles |
| Refrigerated Box | Panther Fusion Internal Control-B |
| Room Temperature Box | Panther Fusion Reconstitution Buffer I |
| Room Temperature Box | Panther Fusion Elution Buffer |
| Room Temperature Box | Panther Fusion Oil |
| Refrigerated Box | Panther Fusion GI Expanded Bacterial Assay Controls• Panther Fusion GI Expanded Bacterial Positive Control• Panther Fusion Negative Control |
Table 2: Ancillary and Collection Kits Required to Perform the Panther Fusion GI Expanded Bacterial Assay
| Aptima Assay Fluids Kit |
|---|
| Aptima Multitest Swab Specimen Collection Kit |
Instrumentation: The Panther Fusion GI Expanded Bacterial Assay has been designed for and validated on the Panther Fusion system. The Panther Fusion System fully automates specimen processing, including sample lysis, nucleic acid capture, amplification, and detection for the Panther Fusion GI Expanded Bacterial Assay.
N/A
FDA 510(k) Clearance Letter - Panther Fusion GI Expanded Bacterial Assay
Page 1
U.S. Food & Drug Administration
10903 New Hampshire Avenue
Silver Spring, MD 20993
www.fda.gov
Doc ID # 04017.08.00
September 25, 2025
Hologic Inc.
Patricia Villarreal
Regulatory Affairs Specialist
10210 Genetic Center Drive
San Diego, California 92121
Re: K251993
Trade/Device Name: Panther Fusion GI Expanded Bacterial Assay
Regulation Number: 21 CFR 866.3990
Regulation Name: Gastrointestinal Microorganism Multiplex Nucleic Acid-Based Assay
Regulatory Class: Class II
Product Code: PCH, OOI
Dated: July 26, 2025
Received: August 26, 2025
Dear Patricia Villarreal:
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 (the 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. Although this letter refers to your product as a device, please be aware that some cleared products may instead be combination products. The 510(k) Premarket Notification Database available at https://www.accessdata.fda.gov/scripts/cdrh/cfdocs/cfpmn/pmn.cfm identifies combination product submissions. 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.
Additional information about changes that may require a new premarket notification are provided in the FDA guidance documents entitled "Deciding When to Submit a 510(k) for a Change to an Existing Device"
Page 2
(https://www.fda.gov/media/99812/download) and "Deciding When to Submit a 510(k) for a Software Change to an Existing Device" (https://www.fda.gov/media/99785/download).
Your device is also subject to, among other requirements, the Quality System (QS) regulation (21 CFR Part 820), which includes, but is not limited to, 21 CFR 820.30, Design controls; 21 CFR 820.90, Nonconforming product; and 21 CFR 820.100, Corrective and preventive action. Please note that regardless of whether a change requires premarket review, the QS regulation requires device manufacturers to review and approve changes to device design and production (21 CFR 820.30 and 21 CFR 820.70) and document changes and approvals in the device master record (21 CFR 820.181).
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 (reporting of medical device-related adverse events) (21 CFR Part 803) for devices or postmarketing safety reporting (21 CFR Part 4, Subpart B) for combination products (see https://www.fda.gov/combination-products/guidance-regulatory-information/postmarketing-safety-reporting-combination-products); good manufacturing practice requirements as set forth in the quality systems (QS) regulation (21 CFR Part 820) for devices or current good manufacturing practices (21 CFR Part 4, Subpart A) for combination products; and, if applicable, the electronic product radiation control provisions (Sections 531-542 of the Act); 21 CFR Parts 1000-1050.
All medical devices, including Class I and unclassified devices and combination product device constituent parts are required to be in compliance with the final Unique Device Identification System rule ("UDI Rule"). The UDI Rule requires, among other things, that a device bear a unique device identifier (UDI) on its label and package (21 CFR 801.20(a)) unless an exception or alternative applies (21 CFR 801.20(b)) and that the dates on the device label be formatted in accordance with 21 CFR 801.18. The UDI Rule (21 CFR 830.300(a) and 830.320(b)) also requires that certain information be submitted to the Global Unique Device Identification Database (GUDID) (21 CFR Part 830 Subpart E). For additional information on these requirements, please see the UDI System webpage at https://www.fda.gov/medical-devices/device-advice-comprehensive-regulatory-assistance/unique-device-identification-system-udi-system.
Also, please note the regulation entitled, "Misbranding by reference to premarket notification" (21 CFR 807.97). For questions regarding the reporting of adverse events under the MDR regulation (21 CFR Part 803), please go to https://www.fda.gov/medical-devices/medical-device-safety/medical-device-reporting-mdr-how-report-medical-device-problems.
For comprehensive regulatory information about medical devices and radiation-emitting products, including information about labeling regulations, please see Device Advice (https://www.fda.gov/medical-devices/device-advice-comprehensive-regulatory-assistance) and CDRH Learn (https://www.fda.gov/training-and-continuing-education/cdrh-learn). 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 (https://www.fda.gov/medical-devices/device-advice-comprehensive-regulatory-assistance/contact-us-division-industry-and-consumer-education-dice) for more information or contact DICE by email (DICE@fda.hhs.gov) or phone (1-800-638-2041 or 301-796-7100).
Page 3
Sincerely,
Bryan M. Grabias -S [Digitally signed by Bryan M. Grabias -S Date: 2025.09.25 11:10:47 -04'00']
Bryan Grabias, Ph.D.
Acting Branch Chief
Bacterial Respiratory and Medical Countermeasures Branch
Division of Microbiology Devices
OHT7: Office of In Vitro Diagnostics
Office of Product Evaluation and Quality
Center for Devices and Radiological Health
Enclosure
Page 4
DEPARTMENT OF HEALTH AND HUMAN SERVICES
Food and Drug Administration
Indications for Use
Form Approved: OMB No. 0910-0120
Expiration Date: 07/31/2026
See PRA Statement below.
510(k) Number (if known): K251993
Device Name: Panther Fusion GI Expanded Bacterial Assay
Indications for Use (Describe)
The Panther Fusion GI Expanded Bacterial Assay is a multiplex real-time PCR in vitro diagnostic test for the rapid and qualitative detection and differentiation of Yersinia enterocolitica, Vibrio (V. parahaemolyticus, V. vulnificus, V. cholerae), Escherichia coli O157, and Plesiomonas shigelloides. Nucleic acids are isolated and purified from Cary-Blair preserved stool specimens collected from individuals exhibiting signs and symptoms of gastroenteritis.
This assay is intended to aid in the differential diagnosis of Yersinia enterocolitica, Vibrio (V. parahaemolyticus, V. vulnificus, V. cholerae), Escherichia coli O157, and Plesiomonas shigelloides infections. The results of this assay should be used in conjunction with clinical presentation, laboratory findings, and epidemiological information and should not be used as the sole basis for diagnosis, treatment, or other patient management decisions. Positive results do not rule out coinfection with other organisms that are not detected by this test and may not be the sole or definitive cause of patient illness. Negative results in the setting of clinical illness compatible with gastroenteritis may be due to infection by pathogens that are not detected by this test, or non-infectious causes such as ulcerative colitis, irritable bowel syndrome, or Crohn's disease. This assay is designed for use on the Panther Fusion System.
Type of Use (Select one or both, as applicable)
☒ Prescription Use (Part 21 CFR 801 Subpart D)
☐ Over-The-Counter Use (21 CFR 801 Subpart C)
CONTINUE ON A SEPARATE PAGE IF NEEDED.
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DO NOT SEND YOUR COMPLETED FORM TO THE PRA STAFF EMAIL ADDRESS BELOW.
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Page 5
510(k) SUMMARY
Panther Fusion GI Expanded Bacterial Assay
I. SUBMITTER:
Hologic, Inc.
10210 Genetic Center Drive
San Diego, CA 92121
Contact Person: Patricia Villarreal
Regulatory Affairs Specialist
Patricia.Villarreal@hologic.com
Phone: 562.833.7906
Fax: N/A
Date Prepared: June 26, 2025
II. DEVICE
Proprietary Name: Panther Fusion GI Expanded Bacterial Assay
Classification Name: Gastrointestinal microorganism multiplex nucleic acid-based assay
Regulation Number: 21 CFR 866.3990 and 21 CFR 862.2570
Regulatory Class: Class II
Product Code: PCH and OOI
III. PREDICATE DEVICE
The predicate device for the Panther Fusion GI Expanded Bacterial Assay is the BioFire FilmArray Gastrointestinal (GI) Panel (K160459, K143005 & K140407).
Page 6
IV. DEVICE DESCRIPTION
The Panther Fusion GI Expanded Bacterial Assay is a multiplex real-time PCR in vitro diagnostic test for the rapid and qualitative detection and differentiation of Yersinia enterocolitica, Vibrio (V. parahaemolyticus, V. vulnificus, V. cholerae), Escherichia coli O157, and Plesiomonas shigelloides. Nucleic acids are isolated and purified from Cary-Blair preserved stool specimens collected from individuals exhibiting signs and symptoms of gastroenteritis.
The Panther Fusion System fully automates specimen processing, including sample lysis, nucleic acid capture, amplification, and detection for the Panther Fusion GI Expanded Bacterial Assay. Nucleic acid capture and elution takes place in a single tube on the Panther Fusion System. The eluate is transferred to the Panther Fusion System reaction tube containing the assay reagents. Multiplex real-time PCR is then performed for the eluted nucleic acid on the Panther Fusion System.
Sample processing: Prior to processing and testing on the Panther Fusion System, specimens are transferred to an Aptima Multitest tube containing specimen transport media (STM) that lyses the cells, releases target nucleic acid, and protects them from degradation during storage.
Nucleic acid capture and elution: An internal control (IC-B) is added automatically to each specimen via the working Panther Fusion Capture Reagent-B (wFCR-B) to monitor for interference during specimen processing, amplification, and detection caused by reagent failure or inhibitory substances. Specimens are first incubated in an alkaline reagent (FER-B) to enable cell lysis. Nucleic acid released during the lysis step hybridizes to magnetic particles in the wFCR-B. The capture particles are then separated from residual specimen matrix in a magnetic field by a series of wash steps with a mild detergent. The captured nucleic acid is then eluted from the magnetic particles with a reagent of low ionic strength (Panther Fusion Elution Buffer).
Multiplex PCR amplification and fluorescence detection: Lyophilized single unit dose reaction master mix is reconstituted with the Panther Fusion Reconstitution Buffer I and then combined with the eluted nucleic acid into a reaction tube. Panther Fusion Oil reagent is added to prevent evaporation during the PCR reaction.
Page 7
Target-specific primers and probes then amplify targets via polymerase chain reaction while simultaneously measuring fluorescence of the multiplexed targets. The Panther Fusion System compares the fluorescence signal to a predetermined cut-off to produce a qualitative result for the presence or absence of each analyte.
The analytes and the channel used for their detection on the Panther Fusion System are summarized in the table below:
| Analyte | Gene Targeted | Instrument Channel |
|---|---|---|
| Yersinia enterocolitica | InvA (Invasive antigen A) | FAM |
| Vibrio parahaemolyticus | gyrB (Gyrase B) | HEX |
| Vibrio vulnificus | gyrB (Gyrase B) | HEX |
| Vibrio cholerae | ompW (Outer Membrane Protein W) | HEX |
| Escherichia coli O157 | rfbE (Perosamine synthase-O-antigen) | ROX |
| Plesiomonas shigelloides | hugA (Heme utilization gene A) | RED647 |
| Internal Control | Not Applicable | RED677 |
Assay Components
The assay components configuration for the Panther Fusion GI Expanded Bacterial Assay is analogous to the Panther Fusion Respiratory Assays. The reagents required to perform the Panther Fusion GI Expanded Bacterial Assay are packaged and sold separately. There are 7 boxes containing 9 reagents which are required for sample processing. The Panther Fusion GI Expanded Bacterial Assay requires one ancillary kit and one specimen collection kit, neither of which are provided with the assay and can be acquired separately:
- Aptima Assay Fluids Kit (303014)
- Aptima Multitest Swab Specimen Collection Kit (PRD-03546)
Page 8
Table 1: Reagents Required to Perform the Panther Fusion GI Expanded Bacterial Assay
| Box | Components Description |
|---|---|
| Refrigerated Box | Panther Fusion GI Expanded Bacterial Assay Cartridges |
| Room Temperature Box | Panther Fusion Extraction Reagent-B• Panther Fusion Capture Reagent-B bottles• Panther Fusion Enhancer Reagent-B bottles |
| Refrigerated Box | Panther Fusion Internal Control-B |
| Room Temperature Box | Panther Fusion Reconstitution Buffer I |
| Room Temperature Box | Panther Fusion Elution Buffer |
| Room Temperature Box | Panther Fusion Oil |
| Refrigerated Box | Panther Fusion GI Expanded Bacterial Assay Controls• Panther Fusion GI Expanded Bacterial Positive Control• Panther Fusion Negative Control |
Table 2: Ancillary and Collection Kits Required to Perform the Panther Fusion GI Expanded Bacterial Assay
| Aptima Assay Fluids Kit |
|---|
| Aptima Multitest Swab Specimen Collection Kit |
Instrumentation
The Panther Fusion GI Expanded Bacterial Assay has been designed for and validated on the Panther Fusion system. The Panther Fusion System fully automates specimen processing, including sample lysis, nucleic acid capture, amplification, and detection for the Panther Fusion GI Expanded Bacterial Assay.
Page 9
V. INDICATIONS FOR USE
The Panther Fusion GI Expanded Bacterial Assay is a multiplex real-time PCR in vitro diagnostic test for the rapid and qualitative detection and differentiation of Yersinia enterocolitica, Vibrio (V. parahaemolyticus, V. vulnificus, V. cholerae), Escherichia coli O157, and Plesiomonas shigelloides. Nucleic acids are isolated and purified from Cary-Blair preserved stool specimens collected from individuals exhibiting signs and symptoms of gastroenteritis.
This assay is intended to aid in the differential diagnosis of Yersinia enterocolitica, Vibrio (V. parahaemolyticus, V. vulnificus, V. cholerae), Escherichia coli O157, and Plesiomonas shigelloides infections. The results of this assay should be used in conjunction with clinical presentation, laboratory findings, and epidemiological information and should not be used as the sole basis for diagnosis, treatment, or other patient management decisions. Positive results do not rule out coinfection with other organisms that are not detected by this test and may not be the sole or definitive cause of patient illness. Negative results in the setting of clinical illness compatible with gastroenteritis may be due to infection by pathogens that are not detected by this test, or non-infectious causes such as ulcerative colitis, irritable bowel syndrome, or Crohn's disease. This assay is designed for use on the Panther Fusion System.
Page 10
VI. COMPARISON OF TECHNOLOGICAL CHARACTERISTICS WITH THE PREDICATE DEVICE
A comparison of the Panther Fusion GI Expanded Bacterial Assay to the predicate BioFire FilmArray Gastrointestinal (GI) Panel (K160459, K143005 & K140407) is summarized in Table 3.
Table 3: Comparison of Similarities and Differences Between the Predicate Device (BioFire FilmArray Gastrointestinal (GI) Panel) and the Subject Device (Panther Fusion GI Bacterial Assay)
| PredicateBioFire FilmArray Gastrointestinal (GI) Panel | SubjectHologic Panther Fusion GI Expanded Bacterial Assay | |
|---|---|---|
| Classification | Class II | Class II |
| Regulation no./Product code | 21CFR 866.3990PCH, OOI | 21CFR 866.3990PCH, OOI |
| Intended use | The FilmArray Gastrointestinal (GI) Panel is a qualitative multiplexed nucleic acid based in vitro diagnostic test intended for use with FilmArray systems. The FilmArray GI Panel is capable of the simultaneous detection and identification of nucleic acids from multiple bacteria, viruses, and parasites directly from stool samples in Cary Blair transport media obtained from individuals with signs and/or symptoms of gastrointestinal infection. The following bacteria (including several diarrheagenic E. coli/Shigella pathotypes), parasites, and viruses are identified using the FilmArray GI Panel:Campylobacter (C. jejuni/C. coli/C. upsaliensis)Clostridium difficile (C. difficile) toxin A/BPlesiomonas shigelloidesSalmonella | The Panther Fusion GI Expanded Bacterial Assay is a multiplex real-time PCR in vitro diagnostic test for the rapid and qualitative detection and differentiation of Yersinia enterocolitica, Vibrio (V. parahaemolyticus, V. vulnificus, V. cholerae), Escherichia coli O157, and Plesiomonas shigelloides. Nucleic acids are isolated and purified from Cary-Blair preserved stool specimens collected from individuals exhibiting signs and symptoms of gastroenteritis.This assay is intended to aid in the differential diagnosis of Yersinia enterocolitica, Vibrio (V. parahaemolyticus, V. vulnificus, V. cholerae), Escherichia coli O157, and Plesiomonas shigelloides infections. The results of this assay should be used in conjunction with clinical |
Page 11
| PredicateBioFire FilmArray Gastrointestinal (GI) Panel | SubjectHologic Panther Fusion GI Expanded Bacterial Assay | |
|---|---|---|
| Intended use (continued) | Vibrio (V. parahaemolyticus/V. vulnificus/V. cholerae), including specific identification of Vibrio choleraeYersinia enterocoliticaEnteroaggregative Escherichia coli (EAEC)Enteropathogenic Escherichia coli (EPEC)Enterotoxigenic Escherichia coli (ETEC) lt/stShiga-like toxin-producing Escherichia coli (STEC) stx1/stx2 (including specific identification of the E. coli O157 serogroup within STEC)Shigella/ Enteroinvasive Escherichia coli (EIEC)CryptosporidiumCyclospora cayetanensisEntamoeba histolyticaGiardia lamblia (also known as G. intestinalis and G. duodenalis)Adenovirus F 40/41AstrovirusNorovirus GI/GIIRotavirus ASapovirus (Genogroups I, II, IV, and V)The FilmArray GI Panel is indicated as an aid in the diagnosis of specific agents of gastrointestinal illness and results are meant to be used in conjunction with other clinical, laboratory, and epidemiological data. Positive results do not rule out coinfection with organisms not included in the FilmArray GI Panel. The agent detected may not be the definite cause of the disease.Concomitant culture is necessary for organism recovery and further typing of | presentation, laboratory findings, and epidemiological information and should not be used as the sole basis for diagnosis, treatment, or other patient management decisions. Positive results do not rule out coinfection with other organisms that are not detected by this test and may not be the sole or definitive cause of patient illness. Negative results in the setting of clinical illness compatible with gastroenteritis may be due to infection by pathogens that are not detected by this test, or non-infectious causes such as ulcerative colitis, irritable bowel syndrome, or Crohn's disease. This assay is designed for use on the Panther Fusion System. |
Page 12
| PredicateBioFire FilmArray Gastrointestinal (GI) Panel | SubjectHologic Panther Fusion GI Expanded Bacterial Assay | |
|---|---|---|
| Intended use (continued) | bacterial agents.This device is not intended to monitor or guide treatment for C. difficile infection.Due to the small number of positive specimens collected for certain organisms during the prospective clinical study, performance characteristics for E. coli O157, Plesiomonas shigelloides, Yersinia enterocolitica, Astrovirus, and Rotavirus A were established primarily with retrospective clinical specimens.Performance characteristics for Entamoeba histolytica, and Vibrio (V. parahaemolyticus, V. vulnificus, and Vibrio cholerae) were established primarily using contrived clinical specimens.Negative FilmArray GI Panel results in the setting of clinical illness compatible with gastroenteritis may be due to infection by pathogens that are not detected by this test or non-infectious causes such as ulcerative colitis, irritable bowel syndrome, or Crohn's disease.A gastrointestinal microorganism multiplex nucleic acid-based assay also aids in the detection and identification of acute gastroenteritis in the context of outbreaks. | |
| Technology | Fully automated nucleic acid amplification, detection and result interpretation (multiplex PCR) | Fully automated nucleic acid amplification, detection and result interpretation (multiplex PCR) |
| Analyte | RNA/DNA | DNA |
| Organism Detected | Campylobacter (C. jejuni/C. coli/C.upsaliensis), Clostridium difficile (C.difficile) toxin A/B, Plesiomonas shigelloides, Salmonella, Vibrio (V. | Yersinia enterocolitica, Vibrio (V. parahaemolyticus, V. vulnificus, V. cholerae), Escherichia coli O157, and Plesiomonas shigelloides |
Page 13
| PredicateBioFire FilmArray Gastrointestinal (GI) Panel | SubjectHologic Panther Fusion GI Expanded Bacterial Assay | |
|---|---|---|
| Organism Detected (continued) | parahaemolyticus/V. vulnificus/V. cholerae) including specific identification of Vibrio cholera, Yersinia enterocolitica, Enteroaggregative Escherichia coli (EAEC), Enteropathogenic Escherichia coli (EPEC), Enterotoxigenic Escherichia coli (ETEC) lt/st, Shiga-like toxin-producing Escherichia coli (STEC) stx1/stx2 (including specific identification of the E. coli O157 serogroup within STEC), Shigella/Enteroinvasive Escherichia coli (EIEC),Cryptosporidium, Cyclospora cayetanensis, Entamoeba histolytica, Giardia lamblia (also known as G. intestinalis and G. duodenalis), Adenovirus F 40/41, Astrovirus, Norovirus GI/GII, Rotavirus A, and Sapovirus (Genogroups I,II, IV, and V). | |
| Specimen Types | Human stool in Cary Blair transport medium | Human stool in Cary Blair transport medium |
| Sample Preparation | Automated sample processing | Automated sample processing |
| Intended Users | Professional use | Professional use |
| Amplification mode | Nested multiplex PCR | Real-time PCR |
| Detection mode | Fluorogenic double stranded DNA binding dye | Fluorogenic target-specific hybridization |
| Instrumentation | BioFire FilmArray Systems | Panther Fusion System |
| Time to Result | Approximately 1 hour | Approximately 2.4 hours |
| Controls | Two controls are included in each reagent pouch to control for sample processing and both stages of PCR and melt analysis. | Internal control in each sample. External control processed at periodic interval |
Page 14
VII. PERFORMANCE DATA
The following performance data (analytical and clinical) were provided in support of the substantial equivalence determination.
Brief Description of Analytical (Non-Clinical) Studies
The following analytical studies (non-clinical) were conducted to support the clearance of the Panther Fusion GI Expanded Bacterial Assay on the Panther Fusion System.
Analytical Sensitivity - Limit of Detection (LoD)
The analytical sensitivity (Limit of Detection or LoD) of the Panther Fusion GI Expanded Bacterial Assay was determined by testing dilutions of preserved negative Cary-Blair stool processed with STM (negative CBS matrix) spiked with bacterial cultures of Yersinia (2 strains), Vibrio (3 strains), Plesiomonas (2 strains), and STEC O157 (2 strains). A minimum of 24 replicates were tested with each of the 3 reagent lots. The LoD for each analyte was determined by Probit analysis for each reagent lot and was confirmed with an additional 24 replicates using a single reagent lot in single analyte and multi- analyte configuration. Analytical sensitivity is defined as the lowest concentration at which ≥95% of all replicates tested positive, as summarized in Table 4.
Table 4. Analytical Sensitivity
| Strain | LoD Concentration (CFU/mL)ᵃ | |
|---|---|---|
| Aptima Multitest Tube | Preserved Stool | |
| Yersinia enterocolitica, 33114 | 91 | 1,820 |
| Yersinia enterocolitica, 1375, O:8 | 94 | 1,880 |
| Vibrio parahaemolyticus, EB101 | 90 | 1,800 |
| Vibrio vulnificus, B9629 | 10 | 200 |
| Vibrio cholerae, 8021 | 33 | 660 |
| STEC O157:H7, EDL 931 | 53 | 1,060 |
| O157:NM, CDC 92-3073 | 357 | 7,140 |
| Plesiomonas shigelloides, CDC 3085-55 | 65 | 1,300 |
| Plesiomonas shigelloides, GNI 14 | 34 | 680 |
CFU = colony forming units.
ᵃ Analyte concentrations in Aptima Multitest tube are ~ 20X dilute compared to preserved stool (~150μL preserved stool in ~3 mL
Page 15
Inclusivity/Reactivity – Wet Testing
The inclusivity/reactivity of the Panther Fusion GI Expanded Bacterial Assay was determined by testing bacterial strains in negative CBS matrix. Each strain was tested in triplicate at 3X LoD with 1 reagent lot in single or multi-analyte configuration. For strains not detected at 3X LoD, additional testing at higher concentrations was performed until 100% positivity was observed. Table 5 shows the lowest concentration of each strain at which 100% positivity was observed.
Table 5. Inclusivity/Reactivity Summary for the GI Expanded Bacterial Assay Analytes
| Organism | ATCC# or Source | Strain/ serovar/ serotype/ antigenic properties | Test Concentration (3X LoD) (CFU/mL) | |
|---|---|---|---|---|
| Aptima Multitest Tube | Preserved Stool | |||
| Yersinia enterocolitica | BEI NR-207 | CDC 497-70, O:8 | 282 | 5,640 |
| BEI NR-212 | NCTC 11175, O:3 | 282 | 5,640 | |
| 23715 | Billups-1803-68, O:8 | 282 | 5,640 | |
| 49397 | 1375, O:8c | 282 | 5,640 | |
| NCTC 10463 | P 77, O:5, 27 | 282 | 5,640 | |
| CCUG 4588 | Type 2, O:9 | 282 | 5,640 | |
| CCUG 8050 | N/A | 282 | 5,640 | |
| CCUG 8232 | Type 5, O:1, 2, 3 | O:2, 3 | O:3/XI | 282 | 5,640 | |
| CCUG 8234 | Type 4 | 282 | 5,640 | |
| 55075 | O:9 | 282 | 5,640 | |
| 27729 | WA, Type 1, O:8 | 282 | 5,640 | |
| Vibrio parahaemolyticus | BEI NR-21990 | 48057, O4: K12 | 270 | 5,400 |
| BEI NR-21992 | KXV 755, O4: K41 | 270 | 5,400 | |
| BAA-242 | VP250, O1:KUT | 270 | 5,400 | |
| 27969 | FC 1011 | 270 | 5,400 | |
| BAA-241 | VP232, O4:K68 | 270 | 5,400 | |
| 33845 | 117 [CDC KC830] | 270 | 5,400 | |
| 43996 | NCTC 10884 [70/116655] | 270 | 5,400 | |
| 33846 | 205 [9302] | 270 | 5,400 | |
| 49529 | MDL 3875-7-83, O4:K12 | 270 | 5,400 | |
| CCUG 34902 | N/A | 270 | 5,400 | |
| CCUG 67711 | N/A | 270 | 5,400 | |
| Vibrio vulnificus | 33847 | 279 [11590] | 270 | 5,400 |
| 33817 | 329 [CDC B3547], Biotype 2 | 33 | 660 | |
| BAA-86 | CDC 9505-95 | 33 | 660 | |
| CCUG 38297 | N/A | 33 | 660 |
Page 16
| Organism | ATCC# or Source | Strain/ serovar/ serotype/ antigenic properties | Test Concentration (3X LoD) (CFU/mL) | |
|---|---|---|---|---|
| Aptima Multitest Tube | Preserved Stool | |||
| Vibrio vulnificus (continued) | CCUG 47321 | N/A | 33 | 660 |
| 29306 | CDCA1402 [P. Baumann 328] | 33 | 660 | |
| 43382 | VVL1 | 33 | 660 | |
| 29307 | CDCA8694 | 33 | 660 | |
| CCUG 38297 | N/Aᵇ | 55 | 1,110 | |
| Vibrio cholerae | BEI NR-147 | N16961, O:1 | 99 | 1,980 |
| BEI NR-148 | CVD 101, O:1 | 99 | 1,980 | |
| BEI NR-149 | Nanking 32/123, O:2 | 99 | 1,980 | |
| BEI NR-152 | Nanking 32/124 (NCTC 8042), O:7 | 99 | 1,980 | |
| 14033 | NCTC 8457 [R. Hugh 1092], O1, Inaba | 99 | 1,980 | |
| 9459 | AMC 20-A-10 [R. Hugh 583], Inaba | 99 | 1,980 | |
| CCUG 2573 | NAG/NCV | 99 | 1,980 | |
| CCUG 2569 | NAG/NCV | 99 | 1,980 | |
| CCUG 4070 | Non O-1 | 99 | 1,980 | |
| CCUG 21589 | 18 | 99 | 1,980 | |
| CCUG 56875 | N/A | 99 | 1,980 | |
| CCUG 53725 | O1/O139 | 99 | 1,980 | |
| CCUG14542 | NA | 99 | 1,980 | |
| 9458 | AMC 20-A-41 [R. Hugh 582], Ogawa | 99 | 1,980 | |
| 25870 | 569B | 99 | 1,980 | |
| STEC O157: H7 | 43890 | CDC C984 [CDC 3526-87], H7 | 159 | 3,180 |
| 43895 | CDC EDL 933, H7 | 159 | 3,180 | |
| 43894 | CDC EDL 932, H7 | 159 | 3,180 | |
| 700927 | EDL 933, H7:K- | 159 | 3,180 | |
| STEC O157: NM | 700375 | CDC 94-G7771, NM | 1,197 | 23,940 |
| 700377 | CDC 92-3099, NM | 1,197 | 23,940 | |
| 700378 | CDC 92-3073, NM | 1,197 | 23,940 | |
| AR Bank # 427ᵃ | N/A | 1,197 | 23,940 | |
| AR Bank # 428ᵃ | N/A | 1,197 | 23,940 | |
| AR Bank # 429ᵃ | N/A | 1,197 | 23,940 | |
| AR Bank # 430ᵃ | N/A | 1,197 | 23,940 | |
| Plesiomonas shigelloides | 14030 | CDC 16408 [Ferguson and Henderson C27, RH 864], O:17 | 195 | 3,900 |
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| Organism | ATCC# or Source | Strain/ serovar/ serotype/ antigenic properties | Test Concentration (3X LoD) (CFU/mL) | |
|---|---|---|---|---|
| Aptima Multitest Tube | Preserved Stool | |||
| Plesiomonas shigelloides (continued) | 51903 | GNI 14ᶜ | 195 | 3,900 |
| 51572 | CIP 69.35 [2886] | 195 | 3,900 | |
| CCUG 7041A | O17: H2 | 195 | 3,900 | |
| CCUG 9221 | O17 | 195 | 3,900 | |
| CCUG 14309 | O17: H2 | 195 | 3,900 | |
| CCUG 14597 | N/A | 195 | 3,900 |
CFU = colony forming units.
ᵃ These strains were evaluated using the higher LoD of the 2 serotypes which is the NM serotype.
ᵇ For this strain 100% positivity was observed at ~ 5X LoD. In silico analysis showed 100% homology to amplification region.
ᶜ Strains used to establish LoD.
Inclusivity/Reactivity - In silico Analysis
The inclusivity of the Panther Fusion GI Expanded Bacterial Assay was evaluated using in silico inclusivity analysis for each analyte. In silico analysis was performed using analyte sequences available in the NCBI database and the whole genome shotgun sequence database. For each analyte, corresponding oligonucleotide sequences (primers and probes) were evaluated against the database sequences. Any sequences with insufficient lengths (not covering the entire amplicon region) were excluded from the analysis.
Based on in silico analysis of all sequences available up to May 30, 2023 in the databases, the Panther Fusion GI Expanded Bacterial Assay is predicted to detect 99.9% of 1,054 Yersinia Enterocolitica, 99.5% of 1,337 Vibrio parahaemolyticus, 99.1% of 1,180 Vibrio vulnificus, 98.0% of 1,189 Vibrio cholerae, 100% of 2,004 STEC O157, and 91.5% of 47 Plesiomonas shigelloides sequences evaluated.
Analytical Specificity: Cross Reactivity and Microbial Interference - Wet Testing
Analytical specificity (cross-reactivity) and microbial interference for the Panther Fusion GI Expanded Bacterial Assay were evaluated in the presence of non-targeted microorganisms that are either phylogenetically related to the assay analytes or potentially found in clinical specimens. Panels consisting of 109 bacteria, viruses, parasites, and yeast listed in Table 6 were tested in negative CBS matrix in the absence and in the presence of GI Expanded Bacterial Assay analytes at 3X LoD. Except where noted, bacteria, yeast, and parasites were
Page 18
evaluated at 10⁶ CFU/mL or 10⁶ rRNA copies/mL or 10⁶ cells/mL; viruses were evaluated at 10⁵ TCID₅₀/mL. If cross-reactivity or interference was observed in the initial testing, then the organism was tested at lower concentrations until the expected result was observed. No cross-reactivity or microbial interference was observed with any of the organisms tested on the Panther Fusion GI Expanded Bacterial Assay at the indicated concentrations.
Table 6. Microorganisms Tested for Cross-Reactivity and Microbial Interference
| Microorganism | Test Concentration | Microorganism | Test Concentration |
|---|---|---|---|
| Arcobacter cryaerophilus | 10⁶ CFU/mL | Entercoccus faecalis | 10⁶ CFU/mL |
| Neisseria gonorrhoeae | 10⁶ CFU/mL | Enterobacter aerogenes | 10⁶ CFU/mL |
| Streptococcus pyogenes | 10⁶ CFU/mL | Enterobacter cloacae | 10⁶ CFU/mL |
| Trabulsiella guamensis | 10⁶ CFU/mL | Escherichia fergusonii | 10⁶ CFU/mL |
| Faecalibacterium prausnitzii | 10⁶ rRNA copies /mL | Escherichia hermanii | 10⁶ CFU/mL |
| Escherichia coli (non-shigatoxigenic) | 10⁶ CFU/mL | Escherichia vulneris | 10⁶ CFU/mL |
| Giardia lamblia BG-Aᵃ | 10⁶ copies/mL | Gardnerella vaginalis | 10⁶ CFU/mL |
| Cyclosporaᵃ | 10⁶ copies/mL | Helicobacter pylori | 10⁶ CFU/mL |
| Cryptosporidiumᵃ | 10⁶ copies/mL | Klebsiella oxytoca | 10⁶ CFU/mL |
| Norovirus (Noro GII)ᵃ | 10⁶ copies/mL | Klebsiella ozaenae | 10⁶ CFU/mL |
| Astrovirusa | 10⁶ copies/mL | Klebsiella pneumoniae | 10⁶ CFU/mL |
| Sapovirus (GII)ᵃ | 10⁵ copies/mL | Lactobacillus acidophilus | 10⁶ CFU/mL |
| Enterovirus (Ent V)ᵃ | 10⁵ copies/mL | Lactobacillus crispatus | 10⁶ CFU/mL |
| Rhinovirusᵃ | 10⁵ copies/mL | Lactococcus lactis | 10⁶ CFU/mL |
| Coronavirus 229E | 10⁵ TCID₅₀/mL | Listeria grayi | 10⁶ CFU/mL |
| Coxsakeivirus Type B4 | 10⁵ TCID₅₀/mL | Listeria monocytogenes | 10⁶ CFU/mL |
| Adenovirus Type 7A | 10⁵ TCID₅₀/mL | Morganella morganii | 10⁶ CFU/mL |
| Rotavirusᵃ | 10⁵ copies/mL | Peptostreptococcus anaerobius | 10⁶ CFU/mL |
| Anaerococcus tetradius | 10⁶ CFU/mL | Peptostreptococcus micros | 10⁶ rRNA copies /mL |
| Abiotrophia defectivia | 10⁶ CFU/mL | Photobacterium damselae | 10⁶ CFU/mL |
| Acinetobacter baumannii | 10⁶ CFU/mL | Prevotella bivia | 10⁶ CFU/mL |
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| Microorganism | Test Concentration | Microorganism | Test Concentration |
|---|---|---|---|
| Acinetobacter lwoffii | 10⁶ CFU/mL | Prevotella melaninogenica | 10⁶ CFU/mL |
| Aeromonas hydrophila | 10⁶ CFU/mL | Proteus mirabilis | 10⁶ rRNA copies /mL |
| Alcaligenes faecalis | 10⁶ CFU/mL | Proteus penneri | 10⁶ CFU/mL |
| Campylobacter upsaliensis | 10⁶ CFU/mL | Proteus vulgaris | 10⁶ CFU/mL |
| Anaerococcus vaginalis | 10⁶ CFU/mL | Providencia alcalifaciens | 10⁶ CFU/mL |
| Arcobacter butzleri | 10⁶ CFU/mL | Providencia rettgeri | 10⁶ CFU/mL |
| Bacillus cereus | 10⁶ CFU/mL | Providencia stuartii | 10⁶ CFU/mL |
| Bacteriodes fragilis | 10⁶ CFU/mL | Pseudomonas aeruginosa | 10⁶ CFU/mL |
| Bacteroides thetaiotaomicron | 10⁶ CFU/mL | Pseudomonas fluorescens | 10⁶ CFU/mL |
| Bacteroides vulgatus | 10⁶ CFU/mL | Serratia liquefaciens | 10⁶ CFU/mL |
| Bifidobacterium adolescentis | 10⁶ CFU/mL | Serratia marcescens | 10⁶ CFU/mL |
| Bifidobacterium longum | 10⁶ rRNA copies /mL | Staphylococcus aureus | 10⁶ CFU/mL |
| Campylobacter fetus | 10⁶ CFU/mL | Staphylococcus epidermidis | 10⁶ CFU/mL |
| Campylobacter hyointestinalis | 10⁶ CFU/mL | Stenotrophomonas maltophilia | 10⁶ CFU/mL |
| Campylobacter rectus | 10⁶ CFU/mL | Streptococcus anginosus | 10⁶ CFU/mL |
| Campylobacter sputorum | 10⁶ CFU/mL | Streptococcus dysgalactiae | 10⁶ CFU/mL |
| Candida albicans | 10⁶ CFU/mL | Yersinia bercovieri | 10⁶ CFU/mL |
| Citrobacter freundii | 10⁶ CFU/mL | Yersinia pseudotuberculosis | 10⁶ CFU/mL |
| Citrobacter koseri | 10⁶ CFU/mL | Yersinia rohdei | 10⁶ CFU/mL |
| Clostridium difficile | 10⁶ CFU/mL | Campylobacter lari | 10⁶ CFU/mL |
| Clostridium perfringens | 10⁶ CFU/mL | Entamoeba histolytica | 10⁴ cells/mL |
| Clostridium ramosum | 10⁶ CFU/mL | Megasphaeara elsdenii | 10⁶ CFU/mL |
| Clostridium sordellii | 10⁶ CFU/mL | Chlamydia trachomatis | 10⁵ IFU/mL |
| Clostridium tertium | 10⁶ CFU/mL | Leptotrichia buccalis | 10⁶ CFU/mL |
| Collinsella aerofaciens | 10⁶ CFU/mL | Cytomegalovirus | 10⁵ TCID₅₀/mL |
| Corynebacterium genitalium | 10⁶ CFU/mL | Salmonella enterica | 10⁶ CFU/mL |
| Cronobacter sakazakii | 10⁶ CFU/mL | Campylobacter jejuni | 10⁶ CFU/mL |
| Edwardsiella tarda | 10⁶ CFU/mL | Shigella sonnei | 10⁶ CFU/mL |
| Egglerthella lenta | 10⁶ rRNA copies /mL | STEC - stx1 | 10⁶ CFU/mL |
| STEC - stx2 | 10⁶ CFU/mL | Vibrio mimicus | 10⁶ CFU/mL |
| Vibrio fluvialis | 10⁶ CFU/mL | Yersinia frederiksenii | 10⁶ CFU/mL |
| Vibrio furnissii | 10⁶ CFU/mL | Yersinia kristensenii | 10⁶ CFU/mL |
| Vibrio metschnikovii | 10⁶ CFU/mL | Vibrio alginolyticusᵇ | 10⁴ CFU/mL |
CFU = colony forming units, IFU = inclusion forming units, rRNA copies = ribosomal ribonucleic acid copies, TCID₅₀ = Median Tissue Culture Infectious Dose.
ᵃ In vitro transcripts were used to evaluate cross-reactivity and microbial interference as cultured virus or whole genome purified nucleic acid are not readily available.
ᵇ Cross reactivity was observed at concentrations ≥10⁵ CFU/mL.
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Coinfection/Competitive Interference
Competitive interference in the Panther Fusion GI Expanded Bacterial Assay was evaluated in triplicate using pairs of assay analytes at low/high concentrations in negative CBS matrix. The low concentration analyte was tested at 3X LoD against a high concentration analyte at 10⁶ CFU/mL. Additionally, analytes were also tested in the absence of a second analyte. If less than 100% positivity was observed for the low concentration analyte, the high concentration analyte was diluted until a concentration was reached where 100% positivity was achieved for the low concentration analyte. The highest concentration of competing analyte at which the low concentration analyte maintained a 100% positivity is shown in Table 7. When the analytes were tested at high concentration, all results for other analytes maintained expected positivity; no competitive interference was observed.
Table 7. Summary of Coinfection Results
| Analyte 1 | Analyte 2 | Yersinia % Pos | Vibrio % Pos | STEC O157 % Pos | Plesiomonas % Pos | ||
|---|---|---|---|---|---|---|---|
| Name | 3X LoD (CFU/mL)ᵃ | Name | High Conc (CFU/mL)ᵇ | ||||
| Negative | NA | Negative | NA | 0% | 0% | 0% | 0% |
| None | 0 | 100% | 0% | 0% | 0% | ||
| Vibrioᵇ | 10⁴ | 100% | 100% | 0% | 0% | ||
| Yersinia | 282 | STEC O157 | 10⁶ | 100% | 0% | 100% | 0% |
| Plesiomonas | 10⁶ | 100% | 0% | 0% | 100% | ||
| None | 0 | 0% | 100% | 0% | 0% | ||
| Yersinia | 10⁶ | 100% | 100% | 0% | 0% | ||
| Vibrio | 282 | STEC O157 | 10⁶ | 0% | 100% | 100% | 0% |
| Plesiomonas | 10⁶ | 0% | 100% | 0% | 100% |
CFU = colony forming units, Pos = positive.
ᵃ Analyte concentration in Aptima Multitest tube.
ᵇ Less than 100% positive results were observed for analyte 1 with Vibrio at ≥10⁵ CFU/mL.
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Interference
Potential inhibitory effects of endogenous and exogenous substances that may be present in a specimen were evaluated in the Panther Fusion GI Expanded Bacterial Assay. Clinically relevant concentrations of potentially interfering substances were added to negative CBS matrix and tested in the absence and in the presence of GI Expanded Bacterial Assay analytes at 3X LoD. Tests were performed in triplicate. The substances and test concentrations are shown in Table 8. No impact on the performance of the Panther Fusion GI Expanded Bacterial Assay was observed for any of the substances at the concentrations tested.
Page 22
Table 8. Substances Tested for Interference
| Substance Type | Generic Name | Active Ingredient(s) | Test Concentrationᵃ' ᵇ' ᶜ |
|---|---|---|---|
| Antibiotics | Amoxicillin | Amoxicillin | 0.7 μg/mL |
| Ampicillin | Ampicillin | 0.9 μg/mL | |
| Doxycycline | Doxycycline | 0.2 μg/mL | |
| Metronidazole | Metronidazole | 1.5 μg/mL | |
| Neosporin® | Polymyxin B sulfate, bacitracin zinc, neomycin sulfate | 1.3% w/v | |
| Anti-microbial and anti-fungal | BZK Antiseptic Towelettes | Benzalkonium chloride | 1.3% v/v |
| Nystatin | Nystatin | 1.3% v/v | |
| Dulcolax® suppository | Bisacodyl | 75 ng/mL | |
| Colace® | Docusate sodium | 3.0 μg/mL | |
| Fleet® mineral oil enema | Mineral oil | 1.3% v/v | |
| Laxatives and stool softeners | Ex-Lax® | Sennosides | 0.8 μg/mL |
| Miralax® | Polyethylene glycol 3350 | 0.1 mg/mL | |
| Milk of Magnesia | Magnesium hydroxide, Aluminum hydroxide | 1.3% v/v | |
| Visicol® | Sodium phosphate | 53 ng/mL | |
| Anti-diarrheal | Imodium | Loperamide hydrochloride | 0.1 μg/mL |
| Anti-Itch | Vagisil® | Benzocaine | 1.3% w/v |
| Preparation H® | Hydrocortisone | 1.3% w/v | |
| Phenylephrine hydrochloride (for hemorrhoids) | Phenylephrine hydrochloride | 0.4 ng/mL | |
| Anti-inflammatory | Mesalazine (Rx only, for Crohns disease/ ulcerative colitis) | Salicylic acid | 0.4 μg/mL |
| Aleve® | Naproxen sodium | 4.5 μg/mL | |
| Antacid | Pepto-Bismol® | Bismuth subsalicylate | 1.3% v/v |
| Tums® | Calcium carbonate | 55 μg/mL | |
| Radiopaque contrast material | Barium sulfate | Barium sulfate | 0.1 mg/mL |
| Lubricants and skin protectants | K-Y® Personal Lubricant Jelly Glycerin | Glycerin | 1.3% w/v |
| Vaseline® Original 100% Pure Petroleum Jelly White | Petrolatum | 1.3% w/v | |
| Desitin® | Zinc oxide | 1.3% w/v |
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| Substance Type | Generic Name | Active Ingredient(s) | Test Concentrationᵃ' ᵇ' ᶜ |
|---|---|---|---|
| Spermicide | Options Conceptrol®Vaginal Contraceptive Gel | Nonoxynol-9 | 1.3% w/v |
| Endogenous | Cholesterol | Cholesterol | 50 μg/mL |
| Fatty acids | Palmitic acid | 16 μg/mL | |
| Fatty acids | Stearic acid | 34 μg/mL | |
| Triglycerides, total (Fecal fat, Intralipid) | Triglycerides | 1.3% v/v | |
| Human Bile | Bilirubin, conjugated | 5.0 μg/mL | |
| Urine | Human Urine | 1.3% v/v | |
| Human Whole Blood | Blood hemoglobin | 1.3% v/v | |
| Mucin | Purified mucin protein | 0.05% w/v |
ᵃ Analyte concentration in Aptima Multitest tube.
ᵇ v/v: volume by volume.
ᶜ w/v: weight by volume.
Stool specimens prepared in various preservative media were evaluated for potential impact on the Panther Fusion GI Expanded Bacterial Assay performance. The preservative media evaluated include 7 different types of Cary-Blair transport media from different vendors and preservative media containing fixatives shown in Table 9. All media were tested with GI Expanded Bacterial Assay analytes at 3X LoD. Comparable performance was seen with all Cary-Blair media. Interference was observed when specimens were processed in media containing fixative.
Page 24
Table 9. Stool Preservative Media Tested for Interference
Cary-Blair Media
- Culture & Sensitivity (C&S) Medium
- Cary Blair Transport Medium w/ Indicator
- Para-Pak® C&S
- Para-Pak® Enteric Plus
- Cardinal Health™ C&S Stool Transport Vial
- Protocol Cary Blair Medium
- Enteric Transport Media (ETM)
Fixative Media (interference was observed)
- Fisher® 10% Buffered Formalin
- Para-Pak® 10% Buffered Formalin
- Para-Pak® LV-PVA
Carryover Contamination
Panther Fusion GI Bacterial Assay and GI Expanded Bacterial Assay belong to the same family of assays that both utilize Cary Blair Stool as the sample type and follow identical assay processing steps. Carryover contamination was evaluated using Panther Fusion GI Bacterial Assay as a representative assay and demonstrated a 0% carryover rate.
Within Laboratory Precision/Repeatability
Panther Fusion GI Expanded Bacterial Assay within laboratory precision was evaluated with a 5-member panel consisting of assay analytes in negative CBS matrix. The 5-member panel included 1 negative, 2 single analyte (Yersinia), and 2 multi-analyte (with Vibrio, STEC O157, and Plesiomonas) panel members. The panels were tested by 3 operators on 2 runs per day, using 3 reagent lots on 3 Panther Fusion Systems over 9 days.
The panel members are described in Table 10, along with a summary of the agreement with the expected results, mean Ct, variability analysis between reagent lots, operators, instruments, days, between and within runs, and overall (total).
Page 25
Table 10. Ct Variability Analysis Summary
| Panel | Description | Analyte | Agreement %ᵃ | Mean Ct | Between Lots | Between Instruments | Between Operators | Between Days | Between Runs | Within Run | Total | |||||||
|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
| SD | CV (%) | SD | CV (%) | SD | CV (%) | SD | CV (%) | SD | CV (%) | SD | CV (%) | SD | CV (%) | |||||
| 1 | Negative (Internal Control) | 162/162 | 100 | 34.6 | 0.07 | 0.2 | 0.08 | 0.23 | 0.04 | 0.12 | 0.00 | 0.00 | 0.00 | 0.00 | 0.48 | 1.39 | 0.5 | 1.43 |
| 2 | Low Pos (1.5X LoD) | Yersinia | 162/162 | 100 | 33.7 | 0.03 | 0.08 | 0.09 | 0.26 | 0.00 | 0.00 | 0.00 | 0.00 | 0.00 | 0.00 | 0.41 | 1.23 | 0.42 |
| 3 | Mod Pos (1.5X LoD) | Yersinia | 162/162 | 100 | 33.7 | 0.12 | 0.35 | 0.07 | 0.21 | 0.01 | 0.04 | 0.00 | 0.00 | 0.17 | 0.52 | 0.23 | 0.69 | 0.32 |
| Vibrio | 162/162 | 100 | 32.7 | 0.07 | 0.21 | 0.12 | 0.37 | 0.00 | 0.00 | 0.00 | 0.00 | 0.19 | 0.57 | 0.2 | 0.6 | 0.3 | ||
| 4 | Low Pos (1.5X LoD) | STEC O157 | 162/162 | 100 | 32.4 | 0.02 | 0.08 | 0.04 | 0.13 | 0.00 | 0.00 | 0.00 | 0.00 | 0.11 | 0.34 | 0.28 | 0.87 | 0.31 |
| Plesiomonas | 162/162 | 100 | 31.3 | 0.02 | 0.08 | 0.06 | 0.2 | 0.00 | 0.00 | 0.03 | 0.1 | 0.00 | 0.00 | 0.21 | 0.68 | 0.22 | ||
| Vibrio | 162/162 | 100 | 33.8 | 0.08 | 0.25 | 0.05 | 0.14 | 0.00 | 0.00 | 0.00 | 0.00 | <0.01 | 0.03 | 0.25 | 0.73 | 0.26 | ||
| 3 | Mod Pos (3X LoD) | STEC O157 | 162/162 | 100 | 33.1 | 0.05 | 0.17 | <0.01 | 0.03 | 0.01 | 0.03 | 0.06 | 0.17 | 0.00 | 0.00 | 0.19 | 0.56 | 0.2 |
| Plesiomonas | 162/162 | 100 | 28 | 0.11 | 0.39 | 0.32 | 1.15 | 0.00 | 0.00 | 0.00 | 0.00 | 0.12 | 0.42 | 0.14 | 0.51 | 0.39 |
Ct = cycle threshold, CV = coefficient of variation, Mod = Moderate, N = sample size, Pos = positive, SD = standard deviation.
ᵃ Agreement to expected panel positivity result.
Reproducibility
Panther Fusion GI Expanded Bacterial Assay reproducibility was evaluated at 3 US sites using 1 negative panel member and 4 panel members positive for 1 or 3 targets. Testing was performed for 5 days by 6 operators (2 at each site) using 1 lot of assay reagents. Each run included 3 replicates of each panel member.
A negative panel member was created using a matrix comprised of stool specimens negative for all assay targets preserved in Cary-Blair media processed into STM. Positive panel members were created by spiking 1.5X LoD (low positive) or 3X LoD (moderate positive) concentrations of the target analytes into the negative matrix.
The agreement with expected results was 100% for all panel members components for Yersinia, Vibrio, STEC O157, and Plesiomonas (Table 11).
Page 26
Table 11. Agreement of Panther Fusion GI Expanded Bacterial Assay Results with Expected Results
| Description | Analyte | N | % (95% CI) |
|---|---|---|---|
| Neg | Internal Control | 89/89 | 100 (95.9-100) |
| Low Posᵃ | Yersiniaᶜ | 90/90 | 100 (95.9-100) |
| Vibrioᶜ | 90/90 | 100 (95.9-100) | |
| STEC O157 | 90/90 | 100 (95.9-100) | |
| Plesiomonasᶜ | 90/90 | 100 (95.9-100) | |
| Mod Posᵇ | Yersiniaᶜ'ᵈ | 90/90 | 100 (95.9-100) |
| Vibrioᶜ | 90/90 | 100 (95.9-100) | |
| STEC O157 | 90/90 | 100 (95.9-100) | |
| Plesiomonasᶜ | 90/90 | 100 (95.9-100) |
CI = score confidence interval, Mod = moderate, N = sample size, Neg = negative, Pos = positive.
ᵃ Low Pos = All targets are 1.5X LoD.
ᵇ Mod Pos = All targets are 3X LoD.
ᶜ Yersinia enterocolitica, Vibrio parahaemolyticus, STEC O157, and Plesiomonas shigelloides strains were used to build the positive panels.
ᵈ One (1) false positive Vibrio result was obtained for a moderate positive Yersinia panel member.
Signal variability was measured as %CV of the Ct values. The total signal variability was ≤1.61% (SD ≤0.55) for all panel components (Table 12). For the sources of variation except the 'within run' factor, %CV values were ≤1.03% for all panel components. The signal variability was ≤1.01% (SD ≤0.33) for the Panther Fusion GI Expanded Bacterial Assay positive controls (Table 13).
Page 27
Table 12. Signal Variability of the Panther Fusion GI Expanded Bacterial Assay by Target and Concentration
| Description | Analyte | N | Mean Ct | Between Site | Between Operator/Runᶜ | Between Day | Within Run | Total | |||||
|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
| SD | CV (%) | SD | CV (%) | SD | CV (%) | SD | CV (%) | SD | CV (%) | ||||
| Low Posᵃ | Yersinia | 90 | 34.7 | 0.17 | 0.50 | 0.21 | 0.61 | 0.09 | 0.27 | 0.44 | 1.25 | 0.52 | 1.51 |
| Vibrio | 90 | 33.7 | 0.16 | 0.49 | 0.08 | 0.25 | 0.00 | 0.00 | 0.26 | 0.77 | 0.32 | 0.95 | |
| STEC O157 | 90 | 32.4 | 0.17 | 0.53 | 0.13 | 0.41 | 0.00 | 0.00 | 0.30 | 0.92 | 0.37 | 1.14 | |
| Plesiomonas | 90 | 33.9 | 0.16 | 0.47 | 0.06 | 0.17 | 0.00 | 0.00 | 0.32 | 0.94 | 0.36 | 1.06 | |
| Mod Posᵇ | Yersinia | 90 | 33.8 | 0.35 | 1.03 | 0.19 | 0.58 | 0.07 | 0.21 | 0.37 | 1.08 | 0.55 | 1.61 |
| Vibrio | 90 | 32.7 | 0.20 | 0.60 | 0.09 | 0.26 | 0.11 | 0.35 | 0.22 | 0.68 | 0.33 | 1.01 | |
| STEC O157 | 90 | 31.4 | 0.24 | 0.75 | 0.08 | 0.27 | 0.07 | 0.21 | 0.26 | 0.81 | 0.36 | 1.16 | |
| Plesiomonas | 90 | 33.2 | 0.22 | 0.67 | 0.12 | 0.37 | 0.00 | 0.00 | 0.26 | 0.78 | 0.36 | 1.09 |
Ct = cycle threshold, CV = coefficient of variation, Mod = moderate, N = sample size, Pos = positive, SD = standard deviation.
Note: The analysis was performed using the SAS MIXED procedure, which applies a lower boundary of 0 to all variance components in the model by default. If a variance component is 0, SD, and %CV are displayed as 0.00
ᵃ Low Pos = All targets are 1.5X LoD.
ᵇ Mod Pos = All targets are 3X LoD.
ᶜ Between Operator may be confounded with Between Run; therefore, Between Operator and Between Run estimates are combined in Between Operator/Run.
Table 13. Signal Variability of the Panther Fusion GI Expanded Bacterial Assay Positive Controls
| Control | Analyte | N | Mean Ct | Between Site | Between Operator | Between Day | Within Day | Total | |||||
|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
| SD | CV (%) | SD | CV (%) | SD | CV (%) | SD | CV (%) | SD | CV (%) | ||||
| Pos | Yersinia | 30 | 32.7 | 0.22 | 0.66 | 0.00 | 0.00 | 0.00 | 0.00 | 0.25 | 0.75 | 0.33 | 1.01 |
| Vibrio | 30 | 33.4 | 0.00 | 0.00 | 0.00 | 0.00 | 0.00 | 0.00 | 0.29 | 0.86 | 0.29 | 0.86 | |
| STEC O157 | 30 | 31.5 | 0.11 | 0.35 | 0.00 | 0.00 | 0.07 | 0.23 | 0.26 | 0.83 | 0.29 | 0.93 | |
| Plesiomonas | 30 | 32.0 | 0.05 | 0.16 | 0.08 | 0.23 | 0.12 | 0.37 | 0.24 | 0.74 | 0.29 | 0.87 |
Ct = cycle threshold, CV = coefficient of variation, N = sample size, Pos = positive, SD = standard deviation.
Note: The analysis was performed using the SAS MIXED procedure, which applies a lower boundary of 0 to all variance components in the model by default. If a variance component is 0, SD, and %CV are displayed as 0.00.
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Brief Description of Clinical Studies
A multicenter study was conducted using remnant stool specimens in Cary-Blair preservative medium collected as part of routine patient care at 10 US clinics from pediatric or adult patients suspected of acute gastroenteritis. All specimens were tested with the Panther Fusion GI Expanded Bacterial Assay and with comparator assays: a PCR plus bidirectional sequencing (run in duplicate) for STEC O157 and an FDA-cleared Nucleic Acid Amplification Test (NAAT) for all other targets. An alternate FDA-cleared NAAT was used for discordant resolution testing, if applicable. Positive (PPA) and negative (NPA) percent agreement, with corresponding 2-sided 95% Score CIs, were calculated relative to comparator results, by target and by specimen category.
A total of 1,548 prospective specimens and 251 retrospective specimens were enrolled in the study; 94 specimens were excluded from the performance analyses (for example, duplicate individuals, invalid Panther Fusion or comparator results for all targets). An additional 189 contrived specimens were assessed to supplement the prospective and retrospective data for all targets. Of the 1,919 specimens tested in valid Panther Fusion GI Expanded Bacterial Assay runs, 36 (1.9%) had initial invalid results. Upon retest, 25 of the 36 specimens yielded valid results, for a total of 11 (0.6%) specimens with final invalid results. The final data set consisted of 1,894 evaluable specimens (1,523 prospective specimens, 182 retrospective specimens, and 189 contrived specimens); not all were evaluable for all analytes. Demographic information for the 1,705 evaluable prospective and retrospective specimens is provided in Table 14.
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Table 14. Summary of Subject Demographics
| Total N (%) | Prospective N (%) | Retrospective N (%) | |
|---|---|---|---|
| Total Specimens | 1,705 | 1,523 | 182 |
| Sex | |||
| Female | 898 (52.1) | 793 (52.1) | 95 (52.2) |
| Male | 817 (47.9) | 730 (47.9) | 87 (47.8) |
| Age Group | |||
| 0 to 28 days | 7 (0.4) | 7 (0.5) | 0 (0) |
| 29 days to <2 years | 74 (4.3) | 67 (4.4) | 7 (3.8) |
| 2 to 5 years | 55 (3.2) | 50 (3.3) | 5 (2.7) |
| 6 to 11 years | 68 (4.0) | 66 (4.3) | 2 (1.1) |
| 12 to 17 years | 73 (4.3) | 71 (4.7) | 2 (1.1) |
| 18 to 21 years | 47 (2.8) | 44 (2.9) | 3 (1.6) |
| 22 to 64 years | 825 (48.4) | 724 (47.5) | 101 (55.5) |
| ≥65 years | 556 (32.6) | 494 (32.4) | 62 (34.1) |
N = population size
Performance characteristics for detection of Yersinia, Vibrio, STEC O157, and Plesiomonas are shown in Table 15 through Table 18.
Table 15. Clinical Performance - Yersinia spp.
| Specimen Origin | N | TP | FP | TN | FN | Prevalenceᵃ (%) | PPA % (95% CI)ᵇ | NPA % (95% CI)ᵇ |
|---|---|---|---|---|---|---|---|---|
| Prospective (Fresh) | 1,507 | 10 | 5ᶜ | 1,487 | 1ᵈ | 0.7 | 90.9 (62.3, 98.4) | 99.4 (98.9, 99.7) |
| Retrospective (Frozen) | 182 | 15 | 3ᵉ | 164 | 0 | N/Aᶠ | 100 (79.6, 100) | 98.2 (94.9, 99.4) |
| Contrived (Frozen) | 189 | 63 | 0 | 126 | 0 | N/Aᶠ | 100 (94.3, 100) | 100 (97.0, 100) |
CI = confidence interval, FN = false negative, FP = false positive, N = sample size, NPA = negative percent agreement, PPA = positive percent agreement, TN = true negative, TP = true positive.
ᵃ Study prevalence reported based on comparator testing.
ᵇ Score CI.
ᶜ 5 of 5 discordant false positive prospective specimens were positive for Yersinia by the alternate NAAT.
ᵈ The discordant false negative prospective specimen was negative for Yersinia by the alternate NAAT.
ᵉ The 3 discordant false positive retrospective specimens were positive for Yersinia by the alternate NAAT.
ᶠ Calculation of prevalence is not applicable.
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Table 16. Clinical Performance - Vibrio spp.
| Specimen Origin | N | TP | FP | TN | FN | Prevalenceᵃ (%) | PPA % (95% CI)ᵇ | NPA % (95% CI)ᵇ |
|---|---|---|---|---|---|---|---|---|
| Prospective (Fresh) | 1,507 | 1 | 0 | 1,505 | 1ᶜ | 0.1 | 50.0 (9.5, 90.5) | 100 (99.7, 100) |
| Retrospective (Frozen) | 182 | 9 | 6ᵈ | 167 | 0 | N/Aᶠ | 100 (70.1, 100) | 96.5 (92.6, 98.4) |
| Contrived (Frozen) | 189 | 63 | 1ᵉ | 125 | 0 | N/Aᶠ | 100 (94.3, 100) | 99.2 (95.6, 99.9) |
CI = confidence interval, FN = false negative, FP = false positive, N = sample size, NPA = negative percent agreement, PPA = positive percent agreement, TN = true negative, TP = true positive.
ᵃ Study prevalence reported based on comparator testing.
ᵇ Score CI.
ᶜ The discordant false negative prospective specimen was positive for Vibrio by the alternate NAAT.
ᵈ All 6 discordant false positive retrospective specimens were positive for Vibrio by the alternate NAAT.
ᵉ The discordant false positive contrived specimen was negative for Vibrio by the alternate NAAT.
ᶠ Calculation of prevalence is not applicable.
Table 17. Clinical Performance – STEC O157
| Specimen Origin | N | TP | FP | TN | FN | Prevalenceᵃ (%) | PPA % (95% CI)ᵇ | NPA % (95% CI)ᵇ |
|---|---|---|---|---|---|---|---|---|
| Prospective (Fresh) | 1,522 | 1 | 2ᶜ | 1,519 | 0 | 0.1 | 100 (20.7, 100) | 99.9 (99.5, 100) |
| Retrospective (Frozen) | 182 | 3 | 1ᵈ | 178 | 0 | N/Aᵍ | 100 (43.9, 100) | 99.4 (96.9, 99.9) |
| Contrived (Frozen) | 189 | 62 | 1ᵉ | 125 | 1ᶠ | N/Aᵍ | 98.4 (91.5, 99.7) | 99.2 (95.6, 99.9) |
CI = confidence interval, FN = false negative, FP = false positive, N = sample size, NPA = negative percent agreement, PPA = positive percent agreement, TN = true negative, TP = true positive.
ᵃ Study prevalence reported based on comparator testing.
ᵇ Score CI.
ᶜ 1 of 2 discordant false positive prospective specimens was negative for STEC O157 by the alternate NAAT. The other discordant was positive for O157 but negative for stx1/stx2 by the alternate NAAT.
ᵈ The discordant false positive retrospective specimen was positive for STEC O157 by the alternate NAAT.
ᵉ The discordant false positive contrived specimen was negative for STEC O157 by the alternate NAAT.
ᶠ The discordant false negative contrived specimen was not retested by the alternate NAAT.
ᵍ Calculation of prevalence is not applicable.
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Table 18. Clinical Performance - Plesiomonas
| Specimen Origin | N | TP | FP | TN | FN | Prevalenceᵃ (%) | PPA % (95% CI)ᵇ | NPA % (95% CI)ᵇ |
|---|---|---|---|---|---|---|---|---|
| Prospective (Fresh) | 1,507 | 1 | 1ᶜ | 1,505 | 0 | 0.1 | 100 (20.7, 100) | 99.9 (99.6, 100) |
| Retrospective (Frozen) | 182 | 8 | 1ᵈ | 173 | 0 | N/Aᶠ | 100 (67.6, 100) | 99.4 (96.8, 99.9) |
| Contrived (Frozen) | 189 | 62 | 0 | 126 | 1ᵉ | N/Aᶠ | 98.4 (91.5, 99.7) | 100 (97.0, 100) |
CI = confidence interval, FN = false negative, FP = false positive, N = sample size, NPA = negative percent agreement, PPA = positive percent agreement, TN = true negative, TP = true positive.
ᵃ Study prevalence reported based on comparator testing.
ᵇ Score CI.
ᶜ The discordant false positive prospective specimen was positive for Plesiomonas by the alternate NAAT.
ᵈ The discordant false positive retrospective specimen was positive for Plesiomonas by the alternate NAAT.
ᵉ The discordant false negative contrived specimen was not retested by the alternate NAAT.
ᶠ Calculation of prevalence is not applicable.
No coinfections were detected by the Panther Fusion GI Expanded Bacterial Assay or by the comparator methods in prospective and retrospective specimens.
VIII. CONCLUSIONS
The analytical and clinical study results demonstrate that the Panther Fusion GI Expanded Bacterial Assay on the Panther Fusion system performs comparably to the predicate device that is currently marketed for the same intended use. Hardware and software verification and validation demonstrate that the Panther Fusion GI Expanded Bacterial Assay on the Panther Fusion system will perform as intended.
§ 866.3990 Gastrointestinal microorganism multiplex nucleic acid-based assay.
(a)
Identification. A gastrointestinal microorganism multiplex nucleic acid-based assay is a qualitativein vitro diagnostic device intended to simultaneously detect and identify multiple gastrointestinal microbial nucleic acids extracted from human stool specimens. The device detects specific nucleic acid sequences for organism identification as well as for determining the presence of toxin genes. The detection and identification of a specific gastrointestinal microbial nucleic acid from individuals exhibiting signs and symptoms of gastrointestinal infection aids in the diagnosis of gastrointestinal infection when used in conjunction with clinical evaluation and other laboratory findings. A gastrointestinal microorganism multiplex nucleic acid-based assay also aids in the detection and identification of acute gastroenteritis in the context of outbreaks.(b)
Classification. Class II (special controls). The special controls are set forth in FDA's guideline document entitled: “Class II Special Controls Guideline: Gastrointestinal Microorganism Multiplex Nucleic Acid-Based Assays for Detection and Identification of Microorganisms and Toxin Genes from Human Stool Specimens.” For availability of the guideline document, see § 866.1(e).