Predicate Devices

The input text does not contain any predicate device information, outputting "Not Found".

Reference Devices

Not Found

AI/ML (Artificial Intelligence / Machine Learning)

No
The description details a PCR-based diagnostic test with automated interpretation of melt curve data. While software is used for interpretation, there is no mention of AI or ML algorithms being employed for this process. The interpretation appears to be based on predefined parameters and thresholds related to melt curve analysis.

Therapeutic

No

Explanation: This device is an in vitro diagnostic test designed to detect and identify pathogens and antimicrobial resistance genes from synovial fluid to aid in the diagnosis of joint infection. It provides information for diagnosis but does not directly treat or prevent a disease or condition.

Diagnostic

Yes

Explanation: The "Intended Use / Indications for Use" section explicitly states, "The BioFire Joint Infection (JI) Panel is a multiplexed nucleic-acid-based, in vitro diagnostic test intended for use... as an aid in the diagnosis of specific agents of joint infection." This directly identifies it as a diagnostic device.

SaMD (Software as a Medical Device)

No

The device is an in vitro diagnostic test that requires specific hardware (BioFire FilmArray 2.0 and BioFire FilmArray Torch Systems) and a physical pouch containing reagents to perform the testing. While it includes software for data interpretation, it is not solely software.

IVD (In Vitro Diagnostic)

Yes, this device is an IVD (In Vitro Diagnostic).

Here's why:

Explicit Statement: The "Intended Use / Indications for Use" section explicitly states: "The BioFire Joint Infection (JI) Panel is a multiplexed nucleic-acid-based, in vitro diagnostic test intended for use with BioFire FilmArray 2.0 and BioFire FilmArray Torch Systems..."

This statement alone is sufficient to confirm its classification as an IVD. The rest of the document further supports this by describing its use in testing biological specimens (synovial fluid) to aid in the diagnosis of a medical condition (joint infection).

PCCP (Predetermined Change Control Plan) Authorized

N/A

Overview

Intended Use / Indications for Use

The BioFire Joint Infection (JI) Panel is a multiplexed nucleic-acid-based, in vitro diagnostic test intended for use with BioFire FilmArray 2.0 and BioFire FilmArray Torch Systems for the simultaneous qualitative detection and identification of multiple bacterial and yeast nucleic acids and select antimicrobial resistance genes from synovial fluid obtained from individuals suspected to have a joint infection.

The following organisms are identified using the BioFire JI Panel: Anaerococcus prevotii/vaginalis, Bacteroides fragilis, Candida spp., Candida albicans, Citrobacter, Clostridium perfringens, Cutibacterium avidum/granulosum, Enterobacter cloacae complex, Enterococcus faecalis, Enterococcus faecium, Escherichia coli, Fingoldia magna, Haemophilus influenzae, Kingella kingae, Klebsiella aerogenes, Klebsiella pneumoniae group, Morganella morganii. Neisseria gonorrhoeae, Parvimonas micra, Peptoniphilus, Peptostreptococcus anaerobius, Proteus spp., Pseudomonas aeruginosa, Salmonella spp., Serratia marcescens, Staphylococcus aureus, Staphylococcus lugdunensis, Streptococcus spp., Streptococcus agalactiae, Streptococcus pneumoniae, and Streptococcus pyogenes.

The BioFire JI Panel contains assays for the detection of genetic determinants associated with S. aureus resistance to methicillin (mecA/C) in conjunction with the SCCmec right extremity junction (MREJ), enterococcal resistance to vancomycin (vanA and vanB), and some mechanisms of gram-negative bacterial resistance ß-lactams including penicillins, cephalosporins, monobactams, and carbapenems (blactx.M, blakec, blaNDM, blaOXA-48-like; blavin). Detection of these genetic determinants can aid in the identification of potentially antimicrobial-resistant organisms in synovial fluid samples. The antimicrobial resistance gene or marker detected may or may not be associated with the agent responsible for disease. Negative results for these select antimicrobial resistance gene assays do not indicate susceptibility, as multiple mechanisms of resistance to methicillin, vancomycin, and ß-lactams exist.

The BioFire JI Panel is indicated as an aid in the diagnosis of specific agents of joint infection and results should be used in conjunction with other clinical and laboratory findings. Negative results may be due to infection with pathogens that are not detected by this test, pathogens present below the limit of detection of the assay, or infection that may not be detected in a synovial fluid specimen. Positive results do not rule out co-infection with other organisms. The BioFire JI Panel is not intended to monitor treatment for joint infections.

Product codes

QSN

Device Description

The BioFire Joint Infection (JI) Panel is designed to simultaneously identify 39 different bacteria, yeast, and select genetic determinants of antimicrobial resistance from synovial fluid specimens. The BioFire JI Panel is compatible with BioFire's PCR-based in vitro diagnostic BioFire FilmArray 2.0 and BioFire FilmArray Torch Systems for infectious disease testing. A panel-specific software module (i.e., BioFire JI Panel pouch module software) is used to perform BioFire JI Panel testing on these systems.

A test is initiated by loading Hydration Solution into one port of the BioFire JI Panel pouch and the synovial fluid sample mixed with the provided Sample Buffer into the other port of the BioFire JI Panel pouch and placing it in a FilmArray instrument. The pouch contains all of the reagents required for specimen testing and analysis in a freeze-dried format; the addition of Hydration Solution and Sample/Buffer Mix rehydrates the reagents. After the pouch is prepared, the BioFire Software guides the user through the steps of placing the pouch into the instrument, scanning the pouch barcode, entering the sample identification, and initiating the run.

The FilmArray instrument contains a coordinated system of inflatable bladders and seal points. which act on the pouch to control the movement of liguid between the pouch blisters. When a bladder is inflated over a reagent blister, it forces liquid from the blister into connecting channels. Alternatively, when a seal is placed over a connecting channel it acts as a valve to open or close a channel. In addition, electronically-controlled pneumatic pistons are positioned over multiple plungers in order to deliver the rehydrated reagents into the blisters at the appropriate times. Two Peltier devices control heating and cooling of the pouch to drive the PCR reactions and the melt curve analysis.

Nucleic acid extraction occurs within the FilmArray pouch using mechanical and chemical lysis followed by purification using standard magnetic bead technology. After extracting and purifying nucleic acids from the unprocessed sample, the Film Array performs a nested multiplex PCR that is executed in two stages. During the first stage, the FilmArray performs a single, large volume, highly multiplexed reverse transcription PCR (rt-PCR) reaction. The products from first stage PCR are then diluted and combined with a fresh, primer-free master mix and a fluorescent double stranded DNA binding dye (LC Green Plus, BioFire Diagnostics). The solution is then distributed to each well of the array. Array wells contain sets of primers designed specifically to amplify sequences internal to the PCR products generated during the first stage PCR reaction. The 2nd stage PCR, or nested PCR, is performed in singleplex fashion in each well of the array. At the end of the 2nd stage PCR, the array is interrogated by melt curve analysis for the detection of signature amplicons denoting the presence of specific targets. A digital camera placed in front of the 2nd stage PCR captures fluorescent images of the PCR reactions and software interprets the data.

The FilmArray Software automatically interprets the results of each DNA melt curve analysis and combines the data with the results of the internal pouch controls to provide a test result for each organism on the panel.

Mentions image processing

A digital camera placed in front of the 2nd stage PCR captures fluorescent images of the PCR reactions and software interprets the data.

Mentions AI, DNN, or ML

Not Found

Input Imaging Modality

Not Found

Anatomical Site

Synovial fluid obtained from individuals suspected to have a joint infection.

Indicated Patient Age Range

Not Found

Intended User / Care Setting

Rx - For Prescription Use Only

Description of the training set, sample size, data source, and annotation protocol

Not Found

Description of the test set, sample size, data source, and annotation protocol

The BioFire JI Panel test results are determined by the Melt Detector, which is part of the FilmArray Analysis Software. The Melt Detector and Analysis Software performance was evaluated by comparing FilmArray test results from well-characterized samples (from the clinical evaluation and analytical testing) to expert annotation. Annotations (positive and negative calls) for all melt curves and assay calls were determined by the sponsor.

Summary of Performance Studies (study type, sample size, AUC, MRMC, standalone performance, key results)

1. Analytical Performance - Precision/Reproducibility:

Study Type: Multi-site reproducibility study.
Sample Size: 480 valid runs overall (20 replicates per sample and system at each of three sites).
Key Results: Overall agreement with expected results for the BioFire Joint Infection Panel was 99.94% (95% CI: 99.89-99.97).

2. Analytical Performance - Analytical Reactivity:

Study Type: In silico analysis and wet lab testing.
Sample Size: Over 350 different isolates representing various species, subspecies, strains, serotypes, AMR gene types, and other characterized variants. Each isolate was tested in triplicate at concentrations near LoD or the lowest reportable level.
Key Results: Limitations on assay reactivity were identified for specific bacterial and yeast isolates or sequences and specific AMR gene types or sequences, often associated with single-base sequence variants.

3. Analytical Performance - Exclusivity (Cross-Reactivity):

Study Type: In silico analysis and wet lab testing.
Sample Size: High concentrations of on-panel and off-panel organisms (most bacteria at >1.0E+08 CFU/mL, most yeast at >1.0E+06 CFU/mL, off-panel fungi, viruses, and parasites at highest cultured concentration possible). Each organism tested in triplicate.
Key Results: Summarized observed and predicted cross-reactivities are provided in Table 50 and 49. Cross-reactivity was observed or predicted for Anaerococcus degeneri, hydrogenalis, lactolyticus, murdochii, nagyae, octavius, senegalensis, tetradius (Anaerococcus prevotii/vaginalis assay); Bacteroides xylanisolvens (Bacteroides fragilis assay); Clostridium cadaveris, Clostridium fallax (Clostridium perfringens assay); Enterobacter bugandensis, Enterobacter chengduensis (Enterobacter cloacae complex assay); Escherichia albertii, Escherichia fergusonii, Shigella boydii, dysenteriae, flexneri, sonnei (Escherichia coli assay); Haemophilus aegyptius (Haemophilus influenzae assay); Kingella negevensis (Kingella kingae assay); Cosenzaea myxofaciens (Proteus spp. assay); Staphylococcus argenteus, Staphylococcus schweitzeri (Staphylococcus aureus assay); and CTX-M cross-reactivity with ampC, blaKLU, blaOXY, blaRAHN.

4. Analytical Performance - Interference Testing:

Study Type: Evaluation of potentially interfering substances.
Sample Size: Contrived samples prepared in synovial fluid (SF) matrix, each containing multiple organisms (and AMR genes) at 3× LoD. Tested in triplicate with three reagent lots.
Key Results: No interference observed for all tested endogenous substances (Blood, Cholesterol, C-Reactive Protein, Fibronectin, Lactate, Monosodium urate/Uric Acid, Calcium Phosphate, Calcium Oxalate, Bilirubin, White Blood Cells, Rheumatoid Factor, Type II Collagen), exogenous substances (Acetaminophen, Salicylic Acid, Ibuprofen, Capsaicin Cream, Salicylate Cream, Camphor Balm, Arnica Gel, Nystatin, Fluconazole, Mupirocin, Ceftriaxone, Vancomycin, Clindamycin, Triple antibiotic ointment, Hydrocortisone, Hyaluronic acid, Lidocaine, Cobalt Ions, Chromium Ions, Ultra-High Molecular Weight Polyethylene, Polymethyl methacrylate Bone cement, Iohexol), competitive microorganisms, disinfection/cleaning substances (Reagent Alcohol, Povidone-iodine, Bleach), and sample processing materials (K2-EDTA anticoagulant).

5. Analytical Performance - Specimen Stability:

Study Type: Evaluation of SF sample storage.
Sample Size: Two contrived samples, each with a representative subset of organisms at 3× LoD, tested in 10 replicates immediately after preparation (D0) and after 1 (D1), 3 (D3), 5 (D5), and 7 (D7) days of refrigerated storage.
Key Results: Detection of organisms and AMR genes was observed in 90.0 - 100% of tested replicates. No negative trend observed associated with sample storage over time.

6. Analytical Performance - Fresh vs. Frozen Study:

Study Type: Comparison of fresh and frozen contrived and clinical specimens.
Sample Size: 30 paired fresh and frozen contrived specimens (spiked with two organisms, mostly at 2xLoD, some spanning detection range), and 25 clinical specimens.
Key Results: 100% concordance for all evaluated analytes in contrived specimens. For clinical specimens, 100% concordance for most evaluated analytes, with one exception for Pseudomonas aeruginosa (missed detection near LoD).

7. Analytical Performance - Detection Limit (LoD):

Study Type: LoD estimation and confirmation.
Sample Size: Serial dilutions of contrived samples, with at least 10 replicates each on FilmArray 2.0 and FilmArray Torch (20 total replicates per dilution).
Key Results: Confirmed LoD for each bacterium or yeast provided in Table 53. For resistance genes, data established that amplification and positive assay results are observed at a host concentration similar to the lowest LoD of applicable bacteria.

8. Clinical Studies - Prospective Clinical Study:

Study Type: Multicenter prospective clinical study.
Sample Size: 1544 synovial fluid specimens.
Reference Methods:
- Bacterial and yeast analytes: SoC culture, single PCR followed by quantitative molecular assay including sequencing (tMol).
- AMR genes: Single PCR followed by sequencing, and separate PCR on cultured isolates. Phenotypic AST performed at study sites.
Key Results (Sensitivity/PPA and Specificity/NPA from Table 56):
- Anaerococcus prevotii/vaginalis: PPA 100% (1/1), NPA 100% (1543/1543)
- Clostridium perfringens: PPA - (0/0), NPA 100% (1544/1544)
- Cutibacterium avidum/granulosum: PPA - (0/0), NPA 100% (1544/1544)
- Enterococcus faecalis: PPA 100% (10/10), NPA 99.7% (1529/1534)
- Enterococcus faecium: PPA 100% (1/1), NPA 99.9% (1541/1543)
- Finegoldia magna: PPA 100% (3/3), NPA 99.9% (1540/1541)
- Parvimonas micra: PPA 0% (0/1), NPA 100% (1543/1543)
- Peptoniphilus: PPA 100% (1/1), NPA 99.9% (1542/1543)
- Peptostreptococcus anaerobius: PPA - (0/0), NPA 99.8% (1541/1544)
- Staphylococcus aureus: PPA 93.3% (98/105), NPA 98.5% (1417/1439)
- Staphylococcus lugdunensis: PPA 100% (2/2), NPA 99.8% (1539/1542)
- Streptococcus spp.: PPA 86.4% (38/44), NPA 99.2% (1488/1500)
- Streptococcus agalactiae: PPA 90.9% (10/11), NPA 99.9% (1532/1533)
- Streptococcus pneumoniae: PPA 100% (3/3), NPA 100% (1541/1541)
- Streptococcus pyogenes: PPA 91.7% (11/12), NPA 100% (1532/1532)
- Bacteroides fragilis: PPA - (0/0), NPA 99.9% (1543/1544)
- Citrobacter: PPA 100% (2/2), NPA 100% (1542/1542)
- Enterobacter cloacae complex: PPA 50% (2/4), NPA 99.9% (1538/1540)
- Escherichia coli: PPA 100% (14/14), NPA 99.9% (1529/1530)
- Haemophilus influenzae: PPA 100% (1/1), NPA 99.9% (1542/1543)
- Kingella kingae: PPA 100% (1/1), NPA 99.6% (1537/1543)
- Klebsiella aerogenes: PPA - (0/0), NPA 100% (1544/1544)
- Klebsiella pneumoniae group: PPA 80.0% (4/5), NPA 99.9% (1538/1539)
- Morganella morganii: PPA 100% (1/1), NPA 99.9% (1541/1543)
- Neisseria gonorrhoeae: PPA 100% (2/2), NPA 99.8% (1539/1542)
- Proteus spp.: PPA 100% (4/4), NPA 99.7% (1536/1540)
- Pseudomonas aeruginosa: PPA 100% (2/2), NPA 99.8% (1539/1542)
- Salmonella spp.: PPA - (0/0), NPA 100% (1544/1544)
- Serratia marcescens: PPA 100% (2/2), NPA 99.9% (1541/1542)
- Candida: PPA 57.1% (4/7), NPA 99.9% (1536/1537)
- Candida albicans: PPA 60.0% (3/5), NPA 100% (1539/1539)
- CTX-M: PPA 100% (5/5), NPA 100% (33/33)
- IMP: PPA - (0/0), NPA 100% (38/38)
- KPC: PPA - (0/0), NPA 100% (40/40)
- mecA/C and MREJ (MRSA): PPA 100% (19/19), NPA 95.7% (90/94)
- NDM: PPA - (0/0), NPA 100% (40/40)
- OXA-48-like: PPA 100% (1/1), NPA 100% (33/33)
- vanA/B: PPA 100% (3/3), NPA 100% (14/14)
- VIM: PPA - (0/0), NPA 100% (38/38)

9. Clinical Studies - Archived Specimen Study:

Study Type: Evaluation of preselected archived retrospective synovial fluid specimens.
Sample Size: 134 initially, 97 expected positives and 12 expected negatives analyzed, with a final analysis of 93 confirmed positive analytes in 88 specimens.
Key Results (Sensitivity/PPA and Specificity/NPA from Table 79):
- Cutibacterium avidum/granulosum: PPA 100% (3/3), NPA 100% (4/4)
- Enterococcus faecalis: PPA 100% (8/8), NPA 100% (92/92)
- Enterococcus faecium: PPA 100% (1/1), NPA 100% (100/100)
- Staphylococcus lugdunensis: PPA 100% (8/8), NPA 100% (94/94)
- Streptococcus agalactiae: PPA 93.8% (15/16), NPA 100% (81/81)
- Streptococcus pneumoniae: PPA 100% (1/1), NPA 100% (101/101)
- Streptococcus pyogenes: PPA 100% (3/3), NPA 100% (98/98)
- Enterobacter cloacae complex: PPA 88.9% (8/9), NPA 100% (86/86)
- Escherichia coli: PPA 100% (9/9), NPA 100% (91/91)
- Haemophilus influenzae: PPA 100% (1/1), NPA 100% (6/6)
- Kingella kingae: PPA 100% (1/1), NPA 100% (100/100)
- Klebsiella aerogenes: PPA 100% (1/1), NPA 100% (101/101)
- Klebsiella pneumoniae group: PPA 100% (3/3), NPA 100% (98/98)
- Neisseria gonorrhoeae: PPA 100% (2/2), NPA 100% (100/100)
- Proteus spp.: PPA 100% (3/3), NPA 100% (97/97)
- Pseudomonas aeruginosa: PPA 100% (13/13), NPA 100% (87/87)
- Salmonella spp.: PPA 100% (3/3), NPA 100% (98/98)
- Serratia marcescens: PPA 100% (2/2), NPA 100% (99/99)
- Candida: PPA 100% (2/2), NPA 100% (100/100)
- Candida albicans: PPA 100% (1/1), NPA 100% (101/101)
- CTX-M: PPA 100% (3/3), NPA 100% (32/32)

10. Clinical Studies - Contrived Specimen Testing:

Study Type: Testing of contrived clinical specimens for rare analytes.
Sample Size: 1235 contrived specimens.
Key Results (Overall Sensitivity/PPA and Specificity/NPA from Table 80 & 81):
- Anaerococcus prevotii/vaginalis: PPA 87.4% (83/95), NPA 100% (1125/1125)
- Clostridium perfringens: PPA 84.3% (113/134), NPA 100% (1101/1101)
- Cutibacterium avidum/granulosum: PPA 74.8% (80/107), NPA 100% (1128/1128)
- Enterococcus faecalis: PPA 100% (53/53), NPA 100% (1182/1182)
- Enterococcus faecium: PPA 96.9% (63/65), NPA 99.9% (1169/1170)
- Finegoldia magna: PPA 88.2% (82/93), NPA 100% (1142/1142)
- Parvimonas micra: PPA 70.1% (54/77), NPA 100% (1158/1158)
- Peptoniphilus: PPA 91.9% (57/62), NPA 100% (1173/1173)
- Peptostreptococcus anaerobius: PPA 98.0% (98/100), NPA 100% (1135/1135)
- Staphylococcus lugdunensis: PPA 96.0% (48/50), NPA 99.9% (1184/1185)
- Streptococcus agalactiae: PPA 100% (59/59), NPA 100% (1175/1175)
- Streptococcus pneumoniae: PPA 89.7% (70/78), NPA 99.6% (1152/1157)
- Streptococcus pyogenes: PPA 98.5% (64/65), NPA 100% (1170/1170)
- Bacteroides fragilis: PPA 98.0% (98/100), NPA 100% (1125/1125)
- Citrobacter: PPA 95.7% (67/70), NPA 100% (1165/1165)
- Enterobacter cloacae complex: PPA 100% (50/50), NPA 100% (1185/1185)
- Escherichia coli: PPA 100% (75/75), NPA 100% (1158/1158)
- Haemophilus influenzae: PPA 96.4% (53/55), NPA 100% (1180/1180)
- Kingella kingae: PPA 100% (50/50), NPA 100% (1185/1185)
- Klebsiella aerogenes: PPA 99.0% (99/100), NPA 100% (1135/1135)
- Klebsiella pneumoniae group: PPA 100% (94/94), NPA 100% (1141/1141)
- Morganella morganii: PPA 92.2% (59/64), NPA 100% (1171/1171)
- Neisseria gonorrhoeae: PPA 94.0% (47/50), NPA 99.9% (1178/1179)
- Proteus spp.: PPA 98.1% (52/53), NPA 100% (1182/1182)
- Pseudomonas aeruginosa: PPA 96.8% (121/125), NPA 100% (1105/1105)
- Salmonella spp.: PPA 95.2% (59/62), NPA 100% (1173/1173)
- Serratia marcescens: PPA 96.4% (54/56), NPA 100% (1179/1179)
- Candida: PPA 96.3% (105/109), NPA 100% (1126/1126)
- Candida albicans: PPA 98.1% (52/53), NPA 100% (1182/1182)
- CTX-M: PPA 99.3% (152/153), NPA 100% (544/544)
- IMP: PPA 100% (93/93), NPA 100% (603/603)
- KPC: PPA 100% (79/79), NPA 100% (618/618)
- mecA/C and MREJ (MRSA): PPA 100% (49/49), NPA 86.8% (46/53)
- NDM: PPA 97.1% (66/68), NPA 100% (629/629)
- OXA-48-like: PPA 100% (65/65), NPA 100% (532/532)
- vanA/B: PPA 100% (98/98), NPA 94.7% (18/19)
- VIM: PPA 100% (83/83), NPA 100% (614/614)

Key Metrics (Sensitivity, Specificity, PPV, NPV, etc.)

See "Summary of Performance Studies" section.

Predicate Device(s)

Not Found

Reference Device(s)

Not Found

Predetermined Change Control Plan (PCCP) - All Relevant Information

Not Found

Regulation Number and Section

N/A

Summary

0

Image /page/0/Picture/0 description: The image contains the logo of the U.S. Food & Drug Administration (FDA). The logo consists of two parts: a symbol on the left and the text "FDA U.S. FOOD & DRUG ADMINISTRATION" on the right. The symbol on the left is a stylized representation of a caduceus, a symbol often associated with medicine and healthcare. The text on the right is in a blue font, with "FDA" in a larger, bolder font than the rest of the text.

EVALUATION OF AUTOMATIC CLASS III DESIGNATION FOR BioFire Joint Infection (JI) Panel DECISION SUMMARY

I Background Information:

A De Novo Number

DEN200066

B Applicant

BioFire Diagnostics, LLC

C Proprietary and Established Names

BioFire Joint Infection (JI) Panel

D Regulatory Information

| Product
Code(s) | Classification | Regulation
Section | Panel |
|--------------------|----------------|------------------------------------------------------------------------------------------------------------------------------------------------------------------------|-------------------|
| QSN | Class II | 21 CFR 866.3988 - Device
to detect and identify
microorganism nucleic
acids and resistance
markers from patients with
suspected orthopedic
infection | MI - Microbiology |

II Submission/Device Overview:

A Purpose for Submission:

De Novo request for evaluation of automatic class III designation for BioFire Joint Infection (JI) Panel

B Measurand:

Anaerococcus prevotii/vaginalis, Bacteroides fragilis, Candida spp., Candida albicans, Citrobacter, Clostridium perfringens, Cutibacterium avidum/granulosum, Enterobacter cloacae complex, Enterococcus faecalis, Enterococcus faecium, Escherichia coli, Fingoldia magna, Haemophilus influenzae, Kingella kingae, Klebsiella aerogenes, Klebsiella pneumoniae group, Morganella morganii, Neisseria gonorrhoeae, Parvimonas micra, Peptoniphilus, Peptostreptococcus anaerobius, Proteus spp., Pseudomonas aeruginosa, Salmonella spp., Serratia marcescens,

1

Staphylococcus aureus, Staphylococcus lugdunensis, Streptococcus spp., Streptococcus agalactiae, Streptococcus pneumoniae, Streptococcus pyogenes, CTX-M. IMP. KPC. NDM. OXA-48-like. VIM. mecA/C and MREJ.

C Type of Test:

Qualitative nucleic acid amplification assay

III Indications for Use:

A Indication(s) for Use:

The BioFire Joint Infection (JI) Panel is a multiplexed nucleic-acid-based, in vitro diagnostic test intended for use with BioFire FilmArray 2.0 and BioFire FilmArray Torch Systems for the simultaneous qualitative detection and identification of multiple bacterial and yeast nucleic acids and select antimicrobial resistance genes from synovial fluid obtained from individuals suspected to have a joint infection.

The following organisms are identified using the BioFire JI Panel: Anaerococcus prevotii/vaginalis, Bacteroides fragilis, Candida spp., Candida albicans, Citrobacter, Clostridium perfringens, Cutibacterium avidum/granulosum, Enterobacter cloacae complex, Enterococcus faecalis, Enterococcus faecium, Escherichia coli, Fingoldia magna, Haemophilus influenzae, Kingella kingae, Klebsiella aerogenes, Klebsiella pneumoniae group, Morganella morganii. Neisseria gonorrhoeae, Parvimonas micra, Peptoniphilus, Peptostreptococcus anaerobius, Proteus spp., Pseudomonas aeruginosa, Salmonella spp., Serratia marcescens, Staphylococcus aureus, Staphylococcus lugdunensis, Streptococcus spp., Streptococcus agalactiae, Streptococcus pneumoniae, and Streptococcus pyogenes.

The BioFire JI Panel contains assays for the detection of genetic determinants associated with S. aureus resistance to methicillin (mecA/C) in conjunction with the SCCmec right extremity junction (MREJ), enterococcal resistance to vancomycin (vanA and vanB), and some mechanisms of gram-negative bacterial resistance ß-lactams including penicillins, cephalosporins, monobactams, and carbapenems (blactx.M, blakec, blaNDM, blaOXA-48-like; blavin). Detection of these genetic determinants can aid in the identification of potentially antimicrobial-resistant organisms in synovial fluid samples. The antimicrobial resistance gene or marker detected may or may not be associated with the agent responsible for disease. Negative results for these select antimicrobial resistance gene assays do not indicate susceptibility, as multiple mechanisms of resistance to methicillin, vancomycin, and ß-lactams exist.

The BioFire JI Panel is indicated as an aid in the diagnosis of specific agents of joint infection and results should be used in conjunction with other clinical and laboratory findings. Negative results may be due to infection with pathogens that are not detected by this test, pathogens present below the limit of detection of the assay, or infection that may not be detected in a synovial fluid specimen. Positive results do not rule out co-infection with other organisms. The BioFire JI Panel is not intended to monitor treatment for joint infections.

2

Culture of synovial fluid is necessary to recover organisms for susceptibility testing and epidemiological typing, to identify organisms in the synovial fluid that are not detected by the BioFire JI Panel, and to further identify species in the genus, complex or group results.

B Special Conditions for Use Statement(s):

Rx - For Prescription Use Only

C Special Instrument Requirements:

The BioFire JI Panel is performed on the FilmArray 2.0 or FilmArray Torch systems.

IV Device/System Characteristics:

A Device Description:

The BioFire Joint Infection (JI) Panel is designed to simultaneously identify 39 different bacteria, yeast, and select genetic determinants of antimicrobial resistance from synovial fluid specimens. The BioFire JI Panel is compatible with BioFire's PCR-based in vitro diagnostic BioFire FilmArray 2.0 and BioFire FilmArray Torch Systems for infectious disease testing. A panel-specific software module (i.e., BioFire JI Panel pouch module software) is used to perform BioFire JI Panel testing on these systems.

A test is initiated by loading Hydration Solution into one port of the BioFire JI Panel pouch and the synovial fluid sample mixed with the provided Sample Buffer into the other port of the BioFire JI Panel pouch and placing it in a FilmArray instrument. The pouch contains all of the reagents required for specimen testing and analysis in a freeze-dried format; the addition of Hydration Solution and Sample/Buffer Mix rehydrates the reagents. After the pouch is prepared, the BioFire Software guides the user through the steps of placing the pouch into the instrument, scanning the pouch barcode, entering the sample identification, and initiating the run.

The FilmArray instrument contains a coordinated system of inflatable bladders and seal points. which act on the pouch to control the movement of liguid between the pouch blisters. When a bladder is inflated over a reagent blister, it forces liquid from the blister into connecting channels. Alternatively, when a seal is placed over a connecting channel it acts as a valve to open or close a channel. In addition, electronically-controlled pneumatic pistons are positioned over multiple plungers in order to deliver the rehydrated reagents into the blisters at the appropriate times. Two Peltier devices control heating and cooling of the pouch to drive the PCR reactions and the melt curve analysis.

Nucleic acid extraction occurs within the FilmArray pouch using mechanical and chemical lysis followed by purification using standard magnetic bead technology. After extracting and purifying nucleic acids from the unprocessed sample, the Film Array performs a nested multiplex PCR that is executed in two stages. During the first stage, the FilmArray performs a single, large volume, highly

3

multiplexed reverse transcription PCR (rt-PCR) reaction. The products from first stage PCR are then diluted and combined with a fresh, primer-free master mix and a fluorescent double stranded DNA binding dye (LC Green Plus, BioFire Diagnostics). The solution is then distributed to each well of the array. Array wells contain sets of primers designed specifically to amplify sequences internal to the PCR products generated during the first stage PCR reaction. The 2nd stage PCR, or nested PCR, is performed in singleplex fashion in each well of the array. At the end of the 2nd stage PCR, the array is interrogated by melt curve analysis for the detection of signature amplicons denoting the presence of specific targets. A digital camera placed in front of the 2nd stage PCR captures fluorescent images of the PCR reactions and software interprets the data.

The FilmArray Software automatically interprets the results of each DNA melt curve analysis and combines the data with the results of the internal pouch controls to provide a test result for each organism on the panel.

Materials provided in each BioFire Joint Infection Panel kit:

Each kit contains sufficient reagents to test 30 samples (30-test kit; RFIT-ASY-0138):

. Individually-packaged BioFire JI Panel pouches
Single-use (1.0 mL) Sample Buffer ampoules .
Single-use pre-filled (1.5 mL) Hydration Injection Vials (blue) .
Single-use Sample Injection Vials (red) .
Individually-packaged Transfer Pipettes .

Materials required but not provided:

. FilmArray system including:
- FilmArray 2.0 or FilmArray Torch and accompanying software
- FilmArray Pouch Loading Station
· 10% bleach solution

Interpretation of Results

When PCR2 is complete, the FilmArray instrument performs a DNA melting analysis on the PCR products and measures the fluorescence signal generated in each well (for more information see appropriate FilmArray Operator's Manual). The FilmArray Software then performs several analyses and assigns a final assay result. The steps in the analyses are described below.

Analysis of Melt Curves

The FilmArray Software evaluates the DNA melt curve for each well of the PCR2 array to determine if a PCR product was present in that well. If the melt profile indicates the presence of a PCR product, then the analysis software calculates the melting temperature (Tm) of the curve and compares it against the expected Tm range for the assay. If the software

4

determines that the Tm of the curve is within the assay-specific Tm range. the melt curve is called positive. If the software determines that the Tm of the curve is not in the appropriate Tm range, the melt curve is called negative.

Analysis of Replicates

Once positive melt curves have been identified, the software evaluates the replicates for each assay to determine the assay result. For an assay to be called positive, two associated melt curves must be called positive, and both Tms must be similar. Assays that do not meet these criteria are called negative.

Organism and Antimicrobial Resistance Gene Interpretation

Each positive and negative assay result is interpreted by the FilmArray Software to provide results for the identification of specific bacteria and antimicrobial resistance (AMR) genes. For most analytes detected by the BioFire JI Panel, interpretations are based on the result of a single assay. However, results for the AMR genes require interpretation based on more than one assay result, as discussed in the relevant sections below.

Interpretations for Gram-positive Bacteria

The BioFire Joint Infection Panel provides a Detected or Not Detected result for most gram-positive bacteria based on one corresponding assay result. If the assay is positive, the test result will be Detected, and if the assay is negative, the test result will be Not Detected. Detection of organisms for which results are based on the interpretation of more than one assay are described below.

Cutibacterium avidum/granulosum

The BioFire JI Panel contains two assays (Cutibacterium1 and Cutibacterium 2) for the detection of these two Curibacterium species. A positive result for one or both assays will generate a Cutibacterium avidum/granulosum Detected test result. Cutibacterium avidum/granulosum will be reported as Not Detected when both assays are negative.

Staphylococcus aureus

The BioFire JI Panel contains two different assays (Saureus1 and Saureus2) for the detection of Staphylococcus aureus. The FilmArray Software interprets each of these assays independently (as described above) and if one or a combination of the assays is positive, the result will be Staphylococcus aureus Detected. If both assays are negative the result will be Staphylococcus aureus Not Detected.

5

Streptococcus spp.

The BioFire JI Panel contains four assays for the detection of Streptococcus species. Species-specific assays are included for the detection of Streptococcus pyogenes (Spyogenes), Streptococcus agalactiae (Sagalactiae), and Streptococcus pneumoniae (Spneumoniae). The fourth assay is a genus level assay (Streptococcus) designed to react with most Viridans group and other Streptococcus species that are not specifically identified by one of the other assays on the panel. The software integrates the results of all four Streptococcus assays into a Streptococcus spp. result as shown in the table below.

BioFire JI Panel Results	Streptococcus	Sagalactiae	Spneumoniae	Spyogenes	Description
	Assay	Assay	Assay	Assay
Streptococcus spp Not
Detected
Streptococcus agalactiae
Not Detected
Streptococcus pneumoniae
Not Detected
Streptococcus pyogenes
Not Detected	Negative	Negative	Negative	Negative	No Streptococcus species
detected in the sample
Streptococcus spp Detected
Streptococcus agalactiae
Not Detected
Streptococcus pneumoniae
Not Detected
Streptococcus pyogenes
Not Detected	Positive	Negative	Negative	Negative	One or more Streptococcus
species detected in the sample
(not S. agalactiae, S.
pneumoniae, or S. pyogenes)
Streptococcus spp Detected
Streptococcus agalactiae
Detected
Streptococcus pneumoniae
Not Detected
Streptococcus pyogenes
Not Detected	Any Result	Positive	Negative	Negative	S. agalactiae detected in the
sample.
Note: additional Streptococcus
species (not S. pneumoniae or
S. pyogenes) may also be in
the sample.
Streptococcus spp Detected
Streptococcus agalactiae
Not Detected
Streptococcus pneumoniae
Detected
Streptococcus pyogenes
Not Detected	Any Result	Negative	Positive	Negative	S. pneumoniae detected in the
sample.
Note: additional Streptococcus
species (not S. agalactiae or S.
pyogenes) may also be in the
sample.
Streptococcus spp Detected
Streptococcus agalactiae
Not Detected
Streptococcus pneumoniae
Not Detected
Streptococcus pyogenes
Detected	Any Result	Negative	Negative	Positive	S. pyogenes detected in the
sample.
Note: additional Streptococcus
species (not S. agalactiae or S.
pneumoniae) may also be in
the sample.

Table 1. Streptococcus Species Results Reporting

6

Interpretations for Gram-negative Bacteria

The BioFire JI Panel contains assays for the specific detection of several gram-negative aerobic and anaerobic species associated with bone and joint infections. Species are identified individually (Bacteroides fragilis, Escherichia coli, Haemophilus influenzae, Kingella kingae, Klebsiella aerogenes, Morganella morganii, Neisseria gonorrhoeae, Pseudomonas aeruginosa. Serratia marcescens), or as multi-species complex, group, or genus results (Enterobacter cloacae complex, Klebsiella pneumoniae group, Citrobacter, Proteus spp., and Salmonella spp.). Each species, complex, group, or genus result is reported as Detected or Not Detected based on an individual corresponding assay result. If the assay is positive, the result will be Detected; if the assay is negative, the result will be Not Detected.

Interpretations for Antimicrobial Resistance (AMR) Genes

The BioFire JI Panel contains assays for the specific detection of several genetic determinants of resistance to multiple classes of antibiotics found in select gram-positive bacteria (mecA/C and MREJ [MRSA] and vanA/B) or gram-negative bacteria (CTX-M, IMP, KPC, NDM, OXA-48-like, and VIM). Results for the AMR genes are not reported unless an applicable bacterium (Table 2) is also detected, therefore the results are based on multiple assays, as described below.

The results for each of the antimicrobial resistance genes will be listed as either:

i Detected - when an applicable bacterium is detected AND the antimicrobial resistance gene assay(s) are positive.
i Not Detected - when an applicable bacterium is detected AND the antimicrobial resistance gene assay(s) are negative.
i N/A - when all applicable bacteria are Not Detected, regardless of the result for the antimicrobial resistance gene assay(s).

Table 2: Antimicrobial Resistance (AMR) Genes and Applicable Organisms

AMR Gene Result	Applicable Bacteria
mecA/C and MREJ	Staphylococcus aureus
vanA/B	Enterococcus faecalis
Enterococcus faecium

7

AMR Gene Result	Applicable Bacteria
CTX-M	Citrobacter
Enterobacter cloacae complex
Escherichia coli
Klebsiella aerogenes
Klebsiella pneumoniae group
Morganella morganii
Proteus spp.
Pseudomonas aeruginosa
Salmonella spp.
Serratia marcescens
IMP	Citrobacter
Enterobacter cloacae complex
Escherichia coli
Klebsiella aerogenes
Klebsiella pneumoniae group
Morganella morganii
Proteus spp.
Pseudomonas aeruginosa
Salmonella spp.
Serratia marcescens
KPC	Citrobacter
Enterobacter cloacae complex
Escherichia coli
Klebsiella aerogenes
Klebsiella pneumoniae group
Morganella morganii
Proteus spp.
Pseudomonas aeruginosa
Salmonella spp.
Serratia marcescens
NDM	Citrobacter
Enterobacter cloacae complex
Escherichia coli
Klebsiella aerogenes
Klebsiella pneumoniae group
Morganella morganii
Proteus spp.
Pseudomonas aeruginosa
Salmonella spp.
Serratia marcescens
VIM	Citrobacter
Enterobacter cloacae complex
Escherichia coli
Klebsiella aerogenes
Klebsiella pneumoniae group
Morganella morganii
Proteus spp.
Pseudomonas aeruginosa
Salmonella spp.
Serratia marcescens
OXA-48-like	Citrobacter
Enterobacter cloacae complex
Escherichia coli
Klebsiella aerogenes
Klebsiella pneumoniae group
Morganella morganii
Proteus spp.
Salmonella spp.
Serratia marcescens

Each AMR gene result is associated with a single corresponding assay except for the mecA/C and MREJ result, which is dependent on both the mecA/C assay and the MREJ assay. Detection of both Staphylococcus aureus and the mecA/C and MREJ markers is indicative of Methicillin Resistant Staphylococcus Aureus (MRSA).

Run Summary

The Run Summary section of the test report provides information about the sample and the run including: Sample ID, time and date of run, control results, and an overall summary of the test results. Control results are reported as Passed, Failed, or Invalid. The Table 3 below provides additional information for each of the possible control field results.

Table 3: Interpretation of Controls Field on the BioFire JI Panel Test Report

| Control

Result	Explanation	Action
Passed	The run was successfully completed
AND
Both pouch controls were
successful.	None
Report the results provided on the test report.
Failed	The run was completed
BUT
At least one of the pouch controls
(RNA Process Control and/or
PCR2) failed.	Repeat the test using a new pouch.
If the error persists, contact Customer
Technical Support for further instruction.

8

| Control

Result	Explanation	Action
Invalid	The controls are invalid because the
run did not complete.
(Typically this indicates a software
or hardware error).	Note the Run Status field in the Run Details
section of the report. Refer to the appropriate
BioFire operator's manual or contact
Technical Support for further instruction.
Once the error is resolved, repeat the test or
repeat the test using another module, if
available.

Result Summary

The Results Summary section of the test report lists the result for each target tested by the panel. Possible results for each organism are Detected, Not Detected, or Invalid. Possible results for each antimicrobial resistance gene are Detected, Not Detected, N/A, or Invalid. Table 4 below provides an explanation for each interpretation and any follow-up necessary to obtain a final result.

Result	Explanation	Action
Detected	The run was successfully completed
AND
The pouch controls were successful (Passed)
AND
The assay(s) for the organism were POSITIVE	Report
results.
Not Detected	The run was successfully completed
AND
The pouch controls were successful (Passed)
AND
The assay(s) for the organism were NEGATIVE	Report
results.
Invalid	The pouch controls were not successful (Failed)
OR
The run was not successful
(Run Status displayed as: Aborted, Incomplete, Instrument
Error or Software Error)	See Table
3 for
instruction.
N/A
(Antimicrobial
Resistance
Genes only)	The run was successfully completed
AND
The pouch controls were successful (Passed)
AND
The assay(s) for the organism(s) associated with the
antimicrobial resistance gene were NEGATIVE so the
results of the antimicrobial resistance gene are not
applicable to the test results.	Report
results.

Table 4: Reporting of Results and Required Actions

B Principle of Operation

9

The BioFire JI Panel pouch is a closed system disposable that stores all the necessary reagents for sample preparation, reverse transcription, polymerase chain reaction (PCR), and detection in order to isolate, amplify, and detect nucleic acid from multiple bacterial and/or fungal pathogens within a single synovial fluid specimen. After sample collection, the user injects hydration solution and sample combined with sample buffer into the pouch, places the pouch into a FilmArray instrument, and starts a run. The entire run process takes about one hour. Additional detail can be found in the appropriate FilmArray Operator's Manual.

During a run, the FilmArray system:

. Lyses the sample by agitation (bead beading).
. Extracts and purifies all nucleic acids from the sample using magnetic bead technology.
Performs nested multiplex PCR bv: .
- · First performing reverse transcription and a single, large volume. massively-multiplexed reaction (PCR1)
- · Then performing multiple singleplex second-stage PCR reactions (PCR2) to amplify sequences within the PCR1 products
. Uses endpoint melting curve data to detect and generate a result for each target on the BioFire Joint Infection Panel array.

C Instrument Description Information

1. Instrument Name:
  FilmArray 2.0 or FilmArray Torch
1. Specimen Identification:
  Synovial fluid specimens
1. Specimen Sampling and Handling:
  Synovial fluid specimens should be tested as soon as possible after collection. If transport or storage is required, specimens can be held refrigerated for up to 7 days (2-8°C),
1. Calibration:
  N/A
1. Quality Control:
  See section VI.5 for information on internal and external controls.

10

V Standards/Guidance Documents Referenced:

. ISO 14971:2019 Medical devices - Applications of risk management to medical devices
IEC 62366-1:2015, Medical device Application of usability . engineering to medical devices
ISO 62304:2006. Medical device software Software life-cycle . processes - IEC 62304:2006, November 27, 2008
. ISO 15223-1:2016 Medical Devices - Symbols to be used with medical devie labels, labeling and information to be supplied - Part 1: General requirements
. ISO13485:2016/EN ISO 13485:2016. Medical devices - Quality Management System - Requirements for regulatory purposes
ISO 20916:2019 In vitro diagnostic medical devices Clinical . performance studies using specimens from human subjects - Good study practice
. EN 13612:20002, Performance evaluation of in vitro diagnostic medical devices (European Commission)
EN ISO 18113-1:2011. In vitro diagnostic medical devices -. Information supplied by the manufacturer (labeling) - Part 1: Terms, definition, and general requirements
. EN ISO 18113-2:2011, In vitro diagnostic medical devices -Information supplied by the manufacturer (labeling) - Part 2: In vitro diagnostic reagents for professional use
EN ISO 23640: 2015. In vitro diagnostic medical devices -. Evaluation of stability of in vitro diagnostic reagents

Performance Characteristics: VI

A Analytical Performance:

1. Precision/Reproducibility:

A multi-site reproducibility study of the BioFire JI Panel was performed with contrived synovial fluid samples over multiple days at three laboratory locations (sites) on a combination of FilmArray 2.0 and FilmArray Torch systems. Reproducibility represents the run-to-run variability of results under actual use conditions over time and is measured as agreement with the expected result. The study evaluated contrived samples containing a subset of representative organisms and AMR genes at two concentrations (and negative). The study incorporated potential variation introduced by site (three), day (five), operator (at least two per site), instrument module/system, and reagent kit lot (three). Negative results were obtained from samples that were not spiked with the organism or AMR gene.

11

Each of the three sites tested 20 replicates per sample and system for a total of 120 valid runs per sample and 480 valid runs overall.

A summary of results (percent (%) agreement with the expected Detected or Not Detected result) for each atypical bacterium and virus (by site and system) is provided Table 5 below.

| | Concentration
Tested | Expected
Result | Agreement with Expected Result | | | |
|----------------------------------------------------------------------------|----------------------------------------------------------------------|--------------------------------------------|--------------------------------|--------------------------------|------------------------------------------|-------------------------------------|
| Analyte | | | FilmArray
2.0 | FilmArray
Torch | All Sites
[95% CI] | |
| | | Gram Positive Bacteria | | | | |
| Anaerococcus
prevotii/vaginalis | None
(No Analyte) | Not Detected | 240/240
100% | 240/240
100% | 480/480
100%
[99.2%-
100%] | |
| Clostridium perfringens | None
(No Analyte) | Not Detected | 240/240
100% | 240/240
100% | 480/480
100%
[99.2%-
100%] | |
| Cutibacterium
avidum/granulosum | None
(No Analyte) | Not Detected | 240/240
100% | 240/240
100% | 480/480
100%
[99.2%-
100%] | |
| Enterococcus faecalis | None
(No Analyte) | Not Detected | 240/240
100% | 238/240
99.2% | 478/480
99.6%
[98.5%-
99.9%] | |
| | Moderate Positive
3× LoD
3.6E+03 copies/mL | Detected | 60/60
100% | 60/60
100% | 120/120
100%
[97.0%-
100%] | |
| Enterococcus faecium
(ATCC 700221) | Low Positive
1× LoD
1.2E+03 copies/mL | Detected | 60/60
100% | 60/60
100% | 120/120
100%
[97.0%-
100%] | |
| | None
(No Analyte) | Not Detected | 120/120
100% | 120/120
100% | 240/240
100%
[98.5%-
100%] | |
| Finegoldia magna | None
(No Analyte) | Not Detected | 240/240
100% | 240/240
100% | 480/480
100%
[99.2%-
100%] | |
| Parvimonas micra | None
(No Analyte) | Not Detected | 240/240
100% | 240/240
100% | 480/480
100%
[99.2%-
100%] | |
| Peptoniphilus | None
(No Analyte) | Not Detected | 240/240
100% | 240/240
100% | 480/480
100%
[99.2%-
100%] | |
| Peptostreptococcus
anaerobius
(ATCC 27337) | Moderate Positive
3× LoD
4.8E+04 copies/mL | Detected | 60/60
100% | 60/60
100% | 120/120
100%
[97.0%-
100%] | |
| | Concentration
Tested | Expected
Result | Agreement with Expected Result | | | |
| Analyte | | | FilmArray
2.0 | FilmArray
Torch | All Sites
[95% CI] | |
| | Low Positive
1× LoD
1.6E+04 copies/mL | Detected | 60/60
100% | 60/60
100% | 120/120
100%
[97.0%-
100%] | |
| | None
(No Analyte) | Not Detected | 120/120
100% | 120/120
100% | 240/240
100%
[98.5%-
100%] | |
| | Moderate Positive
3× LoD
1.3E+04 copies/mL | Detected | 60/60
100% | 60/60
100% | 120/120
100%
[97.0%-
100%] | |
| Staphylococcus aureus
(ATCC 43300) | Low Positive
1× LoD
4.2E+03 copies/mL | Detected | 60/60
100% | 60/60
100% | 120/120
100%
[97.0%-
100%] | |
| | None
(No Analyte) | Not Detected | 120/120
100% | 119/120
99.2% | 239/240
99.6%
[97.7%-
99.9%] | |
| Staphylococcus
lugdunensis | None
(No Analyte) | Not Detected | 240/240
100% | 240/240
100% | 480/480
100%
[99.2%-
100%] | |
| | Moderate Positive
3× LoD
1.6E+03 copies/mL | Detected | 58/60
96.7% | 60/60
100% | 118/120
98.3%
[94.1%-
99.8%] | |
| Streptococcus spp.
(Streptococcus pneumoniae;
ATCC 6303) | Low Positive
1× LoD
5.3E+02 copies/mL | Detected | 59/60
98.3% | 59/60
98.3% | 118/120
98.3%
[94.1%-
99.8%] | |
| | None
(No Analyte) | Not Detected | 120/120
100% | 120/120
100% | 240/240
100%
[98.5%-
100%] | |
| Streptococcus agalactiae | None
(No Analyte) | Not Detected | 240/240
100% | 240/240
100% | 480/480
100%
[99.2%-
100%] | |
| Streptococcus pneumoniae
(ATCC 6303) | Moderate Positive
3× LoD
1.6E+03 copies/mL | Detected | 58/60
96.7% | 60/60
100% | 118/120
98.3%
[94.1%-
99.8%] | |
| Analyte | Concentration
Tested | Expected
Result | Agreement with Expected Result | | | |
| | | | FilmArray
2.0 | FilmArray
Torch | All Sites
[95% CI] | |
| | Low Positive
1x LoD
5.3E+02 copies/mL | Detected | 59/60
98.3% | 59/60
98.3% | 118/120
98.3%
[94.1%-
99.8%] | |
| | None
(No Analyte) | Not Detected | 120/120
100% | 120/120
100% | 240/240
100%
[98.5%-
100%] | |
| Streptococcus pyogenes | None
(No Analyte) | Not Detected | 240/240
100% | 240/240
100% | 480/480
100%
[99.2%-
100%] | |
| | Concentration
Tested | Expected
Result | Agreement with Expected Result | | | |
| Analyte | | | FilmArray
2.0 | FilmArray
Torch | All Sites
[95% CI] | |
| Bacteroides fragilis
(ATCC 25285) | Moderate Positive
3× LoD
3.3E+03 copies/mL | Detected | 60/60
100% | 60/60
100% | 120/120
100%
[97.0%-
100%] | |
| | Low Positive
1× LoD
1.1E+03 copies/mL | Detected | 60/60
100% | 60/60
100% | 120/120
100%
[97.0%-
100%] | |
| | None
(No Analyte) | Not Detected | 120/120
100% | 120/120
100% | 240/240
100%
[98.5%-
100%] | |
| Citrobacter
(Citrobacter freundii;
ATCC 8090) | Moderate Positive
3× LoD
1.4E+04 copies/mL | Detected | 60/60
100% | 60/60
100% | 120/120
100%
[97.0%-
100%] | |
| | Low Positive
1× LoD
4.7E+03 copies/mL | Detected | 60/60
100% | 60/60
100% | 120/120
100%
[97.0%-
100%] | |
| | None
(No Analyte) | Not Detected | 120/120
100% | 120/120
100% | 240/240
100%
[98.5%-
100%] | |
| Enterobacter cloacae
complex | None
(No Analyte) | Not Detected | 240/240
100% | 240/240
100% | 480/480
100%
[99.2%-
100%] | |
| Eschericia coli
AR-Bank#0150 | High Positive
30× LoD
1.8E+05 copies/mL | Detected | 60/60
100% | 60/60
100% | 120/120
100%
[97.0%-
100%] | |
| | High Positive
10× LoD
6.0E+04 copies/mL | Detected | 60/60
100% | 60/60
100% | 120/120
100%
[97.0%-
100%] | |
| | None
(No Analyte) | Not Detected | 120/120
100% | 120/120
100% | 240/240
100%
[98.5%-
100%] | |
| Haemophilus influenzae
ATCC 10211 | Moderate Positive
3× LoD
2.1E+03 copies/mL | Detected | 60/60
100% | 60/60
100% | 120/120
100%
[97.0%-
100%] | |
| | Low Positive
1× LoD
6.9E+02 copies/mL | Detected | 60/60
100% | 60/60
100% | 120/120
100%
[97.0%-
100%] | |
| | None
(No Analyte) | Not Detected | 120/120
100% | 120/120
100% | 240/240
100%
[98.5%-
100%] | |
| Analyte | Concentration
Tested | Expected
Result | FilmArray
2.0 | FilmArray
Torch | All Sites
[95% CI] | |
| Kingella kingae | None
(No Analyte) | Not Detected | 240/240
100% | 240/240
100% | 480/480
100%
[99.2%-
100%] | |
| Klebsiella aerogenes | None
(No Analyte) | Not Detected | 240/240
100% | 240/240
100% | 480/480
100%
[99.2%-
100%] | |
| Klebsiella pneumoniae
group
(Klebsiella pneumoniae;
AR-Bank#0097) | Moderate Positive
3x LoD
4.8E+04 copies/mL | Detected | 60/60
100% | 60/60
100% | 120/120
100%
[97.0%-
100%] | |
| Klebsiella pneumoniae
group
(Klebsiella pneumoniae;
AR-Bank#0097) | Low Positive
1x LoD
1.6E+04 copies/mL | Detected | 60/60
100% | 60/60
100% | 120/120
100%
[97.0%-
100%] | |
| Klebsiella pneumoniae
group
(Klebsiella pneumoniae;
AR-Bank#0097) | None
(No Analyte) | Not Detected | 120/120
100% | 120/120
100% | 240/240
100%
[98.5%-
100%] | |
| Morganella morganii | None
(No Analyte) | Not Detected | 240/240
100% | 240/240
100% | 480/480
100%
[99.2%-
100%] | |
| Neisseria gonorrhoeae
ATCC 19424 | Moderate Positive
3x LoD
6.6E+03 copies/mL | Detected | 60/60
100% | 60/60
100% | 120/120
100%
[97.0%-
100%] | |
| Neisseria gonorrhoeae
ATCC 19424 | Low Positive
1x LoD
2.2E+03 copies/mL | Detected | 59/60
98.3% | 60/60
100% | 119/120
99.2%
[95.4%-
99.9%] | |
| Neisseria gonorrhoeae
ATCC 19424 | None
(No Analyte) | Not Detected | 120/120
100% | 120/120
100% | 240/240
100%
[98.5%-
100%] | |
| Proteus spp. | None
(No Analyte) | Not Detected | 240/240
100% | 240/240
100% | 480/480
100%
[99.2%-
100%] | |
| Pseudomonas aeruginosa | None
(No Analyte) | Not Detected | 240/240
100% | 240/240
100% | 480/480
100%
[99.2%-
100%] | |
| Salmonella spp. | None
(No Analyte) | Not Detected | 240/240
100% | 240/240
100% | 480/480
100%
[99.2%-
100%] | |
| Serratia marcescens | None
(No Analyte) | Not Detected | 240/240
100% | 240/240
100% | 480/480
100%
[99.2%-
100%] | |
| Yeast | | | | | | |
| Analyte | Concentration
Tested | Expected
Result | Agreement with Expected Result | | | |
| | | | FilmArray
2.0 | FilmArray
Torch | All Sites
[95% CI] | |
| Candida krusei
(ATCC 6258) | Moderate Positive
3× LoD
3.0E+03 CFU/mL | Detected | 60/60
100% | 60/60
100% | 120/120
100%
[97.0%-
100%] | |
| Candida krusei
(ATCC 6258) | Low Positive
1× LoD
1.0E+03 CFU/mL | Detected | 60/60
100% | 60/60
100% | 120/120
100%
[97.0%-
100%] | |
| Candida krusei
(ATCC 6258) | None
(No Analyte) | Not Detected | N/A | | | |
| Candida albicans
(ATCC 90028) | Moderate Positive
3× LoD
1.5E+03 CFU/mL | Detected | 60/60
100% | 60/60
100% | 120/120
100%
[97.0%-
100%] | |
| Candida albicans
(ATCC 90028) | Low Positive
1× LoD
5.0E+02 CFU/mL | Detected | 60/60
100% | 60/60
100% | 120/120
100%
[97.0%-
100%] | |
| Candida albicans
(ATCC 90028) | None
(No Analyte) | Not Detected | 120/120
100% | 120/120
100% | 240/240
100%
[98.5%-
100%] | |
| Analyte | Concentration
Tested | Expected
Result | Agreement with Expected Result | | All Sites
[95% CI] | |
| CTX-M | None
(No Analyte) | Not Detected | 240/240
100% | 240/240
100% | 480/480
100%
[99.2%-
100%] | |
| IMP | None
(No Analyte) | Not Detected | 240/240
100% | 240/240
100% | 480/480
100%
[99.2%-
100%] | |
| KPC
(Klebsiella pneumoniae;
AR-Bank#0097) | Moderate Positive
3× LoD
4.8E+04 copies/mL | Detected | 60/60
100% | 60/60
100% | 120/120
100%
[97.0%-
100%] | |
| | Low Positive
1× LoD
1.6E+04 copies/mL | Detected | 60/60
100% | 60/60
100% | 120/120
100%
[97.0%-
100%] | |
| | None
(No Analyte) | Not Detected | 120/120
100% | 120/120
100% | 240/240
100%
[98.5%-
100%] | |
| mecA/C and MREJ
(MRSA)
(Staphylococcus aureus;
ATCC 43300) | Moderate Positive
3× LoD
1.3E+04 copies/mL | Detected | 60/60
100% | 60/60
100% | 120/120
100%
[97.0%-
100%] | |
| | Low Positive
1× LoD
4.2E+03 copies/mL | Detected | 60/60
100% | 60/60
100% | 120/120
100%
[97.0%-
100%] | |
| | None
(No Analyte) | Not Detected | 120/120
100% | 120/120
100% | 240/240
100%
[98.5%-
100%] | |
| NDM
(E. coli;
AR-Bank#0150) | High Positive
30× LoD
1.8E+05 copies/mL | Detected | 60/60
100% | 60/60
100% | 120/120
100%
[97.0%-
100%] | |
| | High Positive
10× LoD
6.0E+04 copies/mL | Detected | 60/60
100% | 60/60
100% | 120/120
100%
[97.0%-
100%] | |
| | None
(No Analyte) | Not Detected | 120/120
100% | 120/120
100% | 240/240
100%
[98.5%-
100%] | |
| OXA-48-like | None
(No Analyte) | Not Detected | 240/240
100% | 240/240
100% | 480/480
100%
[99.2%-
100%] | |
| vanA/B
(Enterococcus faecium;
ATCC 700221) | Moderate Positive
3× LoD
3.6E+03 copies/mL | Detected | 60/60
100% | 60/60
100% | 120/120
100%
[97.0%-
100%] | |
| | Analyte | Concentration
Tested | Expected
Result | Agreement with Expected Result | | |
| | | | | FilmArray
2.0 | FilmArray
Torch | All Sites
[95% CI] |
| | | Low Positive
1× LoD
1.2E+03 opies/mL | Detected | 60/60
100% | 60/60
100% | 120/120
100%
[97.0%-
100%] |
| | | None
(No Analyte) | Not Detected | 120/120
100% | 120/120
100% | 240/240
100%
[98.5%-
100%] |
| | VIM | None
(No Analyte) | Not Detected | 240/240
100% | 240/240
100% | 480/480
100%
[99.2%-
100%] |
| | Overall Agreement with Expected Results
[95% Confidence Interval] | | | | 18,468/18,480
99.94%
[99.89-99.97] | |

Table 5: Reproducibility of BioFire Joint Infection Panel Results

12

13

14

15

16

17

18

2. Linearity:

Not applicable.

3. Analytical Specificity/Interference:

Analytical Reactivity

Analytical reactivity of the BioFire JI Panel assays was evaluated using a combination of in silico analysis of sequences available in public databases and testing of over 350 different isolates representing various species, subspecies, strains, serotypes, AMR gene types, and other characterized variants. Each isolate was tested in triplicate at concentrations near LoD or the lowest reportable level for the analyte.

Limitations on assay reactivity (observed and/or predicted by in silico analysis) with specific bacterial and yeast isolates or sequences and specific AMR gene types or sequences are noted (Table 6). Most limitations are associated with single-base sequence variants under one or more assay primers.

Table 6. Limitations on Analytical Reactivity of BioFire JI Panel Assays

Limitation	Analyte	Strain/Isolate Variant
Minor
(Detected at mecA/C and MREJd,e	MREJ type xvd, xviiie, xixe, xxe
	VIM	VIM types 7, 39, 45, 46, 61, 65, 67
	Rare or Non-relevant Species
	Candida spp.	several Candida species; see Error! Reference source not found.
	Citrobacter	Citrobacter almonaticus, C. farmeri, C.gillenii, C. rodentium, C. sedlakii
	Peptoniphilus spp.	Peptoniphilus coxii, P. duerdenii, P. ivorii, P. koenoeneniae, P. massiliensisf, P. porci, P. olsenii, P. tyrelliase
	Streptococcus spp.	Streptococcus entericus, S.halitosis, S. hyovaginalis, S. pantholopis

19

3 Detection near LoD was impaired for four isolates of A. vaginalis. Sequencing revealed primer mismatches predicted to impar detection. Comparable sequence variants were observed in two A. vaginalis sequences retrieved from public databases. A limitation on reactivity is predicted for approximately 25% of A. prevotii/vaginalis sequences and isolates evaluated.

b Reactivity limitations observed or predicted for sequence variants identified for E. hormacchei ATCC 49162, E. asburiae ATCC 35953 (tested), ATCC 35954 (not tested), ATCC 35955 (not tested), and a small subset of database sequences for E. cloacae, E. hormaechei, E. ludwigii and E. mori with similar or less impactful variants under assay primers. Variant sequences with major or minor reactivity limitations represent less than 2% of sequences for ECC species.

· Petite strains of Candida albicans will not be detected by the Candida albicans-specific assay but will be amplified by the multi-species Candida assay and reported as Candida Detected.

4 Sequence analysis predicts that approximately 40% of MREJ type xv-like sequences will not be detected due to a variant base at the 3' end of an assay primer.

€ MREJ types xviii, xix and xx will not be described in association with methicillin-sensitive isolates, so the meca/C and MREJ (MRSA) Not Detected result with the methicillin-sensitive phenotype of isolates with these MREJ types.

f Not a validly published Peptoniphilus species.

20

| Organism | Isolate ID
(Strain) | Test
Concentration
(copies/mL) | xLoD | Result |
|-----------------------------------------|----------------------------|--------------------------------------|------|--------------------------------------------------|
| Anaerococcus
prevotii | ATCC 9321
(PC 1) | 4.8E+03 | 1x | Anaerococcus prevotii/vaginalis Detecteda |
| | ATCC 14952
(M3) | 1.4E+05 | 3x | |
| | CCUG 72601 | 1.4E+05 | 3x | |
| | VTK 400239 | 1.4E+05 | 3x | |
| | ATCC 51170
(GIFU 12669) | 4.8E+04 | 1x | |
| | DSM 25446
(ph9) | 1.4E+05 | 3x | |
| Anaerococcus
vaginalis | GRE 1654021 | 1.4E+05 | 3x | |
| | GRE 1554051a | 1.4E+06 | 30x | |
| | GRE 1653021 | 4.8E+05 | 10x | |
| | GRE 1757298a | 4.8E+05 | 10x | |
| | VTK 401665a | 1.4E+06 | 30x | |
| | VTK 401672a | 4.8E+05 | 10x | |

Table 7: Angerococcus prevotii/vaginalis Isolates Tested

aVariant sequence with 3' base mismatch to a primer that may impair detection near LoD (10 to 30-fold). Variant sequences with minor detection impairment represent ~25% of total A. prevotii/vaginalis sequences evaluated.

Table 8: Clostridium perfringens Isolates Tested

Organism	Toxinotype	Isolate ID	Test Concentration		Result
			(copies/mL)	X LoD
Clostridium
perfringens	A	ATCC 13124 (S 107)	1.3E+03	1x	Clostridium
perfringens
	A	ATCC 27059 (814)	3.9E+03	3x	Clostridium
perfringens
	C	ATCC 3628 (Strain 51)	3.9E+03	3x	Clostridium
perfringens
	E	ATCC 8009	3.9E+03	3x	Detected
	-	ATCC 9081 (13942)	3.9E+03	3x

		Test Concentration
Organism	Isolate ID	(copies/mL)	X LoD	Result
Cutibacterium
avidum	ATCC 25577 (1689B, VPI 0179)	5.0E+04	1x	Cutibacterium
Detected
	ATCC 49753 (VPI 0575)	1.5E+05	3x
	ATCC 49754 (VPI 0576)	5E+05	10x
	ATCC 49755 (VPI 0589)	1.5E+05	3x
	ATCC 49769 (VPI 0670)	5E+05	10x
	ATCC 25564 (VPI 0507)	5.0E+04	1x
	ATCC 11829 (VPI 0210)	1.5E+05	3x
Cutibacterium
granulosum	ATCC 25746 (D-34)	1.5E+05	3x	Cutibacterium
Detected
	CCUB 14831 (Serovar 3)	1.5E+05	3x
	GRE 1554046	1.5E+05	3x
	GRE 1951015a	5E+05	10x
	GRE 1760015a	5.0E+06	100x	Cutibacterium
Not Detected

Table 9: Cutibacterium avidum/granulosum Isolates Tested

aSequences with predicted impacts on reactivity by in silico analysis

21

Organism	Isolate ID (Strain)	Test Concentration		Result
		(copies/mL)	X LoD
Enterococcus
faecalis	ATCC 51299 (NJ-3)	5.0E+03	1x	Enterococcus
faecalis
Detected
	ATCC 19433 (Tissier)	1.5E+04	3x
	ATCC 49533 (UWH 1936)	1.5E+04	3x
	ATCC 700802 (V583)	1.5E+04	3x
	ATCC BAA-2573 (bMx 0502240)	1.5E+04	3x
	JMI 12536	1.5E+04	3x

Table 10: Enterococcus faecalis Isolates Tested

Table 11: Enterococcus faecium Isolates Tested

Organism	Isolate ID (Strain)	Test Concentration		Result
		(copies/mL)	xLoD
Enterococcus
faecium	ATCC 700221	1.2E+03	1x
	ATCC 19434 (Grumbach serotype 11)	3.6E+03	3x	Enterococcus
faecium
	ATCC 27270 (X3)	3.6E+03	3x
	ATCC 51858 (Vancomycin-dependent #4)	3.6E+03	3x	Detected
	ATCC BAA-2318	3.6E+03	3x
	JMI 475	3.6E+03	3x

Table 12: Finegoldia magna Isolates Tested

| Organism | Isolate ID (Strain) | Test Concentration
(copies/mL) X LoD | Reported Result |
|-----------------------------|----------------------|-----------------------------------------|--------------------------------------|
| Finegoldia
magna | ATCC 15794 (2974) | 3.1E+05 1x | Finegoldia magna
Detected |
| | ATCC 14955 (BU) | 9.3E+05 3x | |
| | ATCC 29328 (WAL2508) | 9.3E+05 3x | |
| | ATCC 53516 (312) | 9.3E+05 3x | |
| | DSM 20362 (168) | 9.3E+05 3x | |
| | GRE 1556006 | 9.3E+05 3x | |

Table 13: Parvimonas micra Isolates Tested

Organism	Isolate ID (Strain)	Test Concentration
		(copies/mL	xLoD	Result
Parvimonas
micra	ATCC 33270 (3024A)	4.8E+03	Ix	Parvimonas
micra
Detected
	CCUG 56809	1.4E+04	3x
	CCUG 57049	1.4E+04	3x
	GRE 1651163	1.4E+04	3x
	GRE 1757098	1.4E+04	3x

Table 14: Peptoniphilus spp. Isolates Tested

Species	Isolate ID (Strain)	Test Concentration		Result
		(copies/mL)	xLoD
Peptoniphilus
assacharolyticus	ATCC 14963 (BAI, UW 228)	4.0E+04	1x
Peptoniphilus allenia	ATCC BAA-1643 (WAL 1768N)	1.2E+05	3x
Peptoniphilus
gorbachii	ATCC BAA-1383 (WAL 10418)]	1.2E+05	3x	Peptoniphilus
Detected
Peptoniphilus
grossensisa	DSM 25475 (ph5)	1.2E+05	3x
Peptoniphilus harei	ATCC BAA-601 (SBH 432)	1.2E+05	3x

22

	DSM 10021 (SBH 064)	1.2E+05	3x
	GRE 1554070	1.2E+05	3x
Peptoniphilus
indolicus	ATCC 29427 (R13)	1.2E+05	3x
	GRE 1556024	1.2E+05	3x
Peptoniphilus
lacrimalis	ATCC 51171 (GIFU 7667)	1.2E+05	3x
	CCUG 47146	1.2E+05	3x
Peptoniphilus
senegalensis	DSM 25694 (JC140)	1.2E+05	3x
Peptoniphilus
tyrrelliaeb	CCUG 59621 (RMA 19911)	8.0E+08
CFU/mL	High
Peptoniphilus
koenoeneniaeb	ATCC BAA-1638 (WAL 20371)	8.0E+08
CFU/mL	High
Peptoniphilus coxii	ATCC BAA-2106 (RMA 16757)	8.0E+08
CFU/mL	High
Peptoniphilus
duerdenii	ATCC BAA-1640 (WAL1998L)	8.0E+08
CFU/mL	High	Peptoniphilus
Not Detected
Peptoniphilus ivorii	ATCC BAA-602 (SBH 093)	8.0E+08
CFU/mL	High
Peptoniphilus
massiliensisa	ATCC BAA-1641 (WAL 18041)	8.0E+08
CFU/mL	High
Peptoniphilus olsenii	ATCC BAA-1384 (WAL 12922)	8.0E+08
CFU/mL	High
Peptoniphilus porci	In silico prediction (not tested)
Other Peptoniphilus
species	Unknown Reactivity
(no sequences/not tested)

ªIsolates tested were characterized by the culture collection as P. allenii, P. grossensis, and P. massiliensis, though none are currently validly published Peptoniphilus species.

b Peptoniphilus koenoeneniae and Peptoniphilus tyrelliae were detected at a concentration >100x LoD.

Table 15: Peptostreptococcus anaerobius Isolates Tested
---------------------------------------------------------

Organism	Isolate ID (Strain)	Test Concentration		Result
		(copies/mL)	xLoD
Peptostreptoc
occus
anaerobius	ACC 27337 (A prevot 4372)	1.6E+04	1x
	ATCC 49031 (MSHD)	4.8E+04	3x	Peptostreptococcus
anaerobius
Detected
	CCUG 37992	4.8E+04	3x
	CCUG 38379	4.8E+04	3x
	CCUG 46594 (GIFU 7800)	4.8E+04	3x

Table 16: Staphylococcus aureus Isolates Tested

| Organism | Isolate ID (Strain) | Test
Concentration | | Result |
|--------------------------|---------------------------|-----------------------|------|--------------------|
| | | (copies/mL) | xLoD | |
| Staphylococcus
aureus | ATCC BAA-2313 (M10/0148) | 4.2E+03 | 1x | S. aureus Detected |
| | ATCC BAA-2312 (M10/0061) | 1.3E+04 | 3x | |
| | ATCC BAA-1700 (HFH-33798) | 1.3E+04 | 3x | |
| | ATCC BAA-1707 (MW2) | 1.3E+04 | 3x | |
| | ATCC BAA-1749 (96:308) | 1.3E+04 | 3x | |
| | ATCC BAA-1759 (N7129) | 1.3E+04 | 3x | |
| | ATCC BAA-1764 (7031) | 1.3E+04 | 3x | |

23

	ATCC BAA-1765 (102-04)	1.3E+04	3x
	NARSA NRS662 (CO-34)	1.3E+04	3x
	NARSA NRS683 (GA-298)	1.3E+04	3x
	NARSA NRS689 (GA-442)	1.3E+04	3x
	NARSA NRS691 (GA-62)	1.3E+04	3x
	NARSA NRS701 (MN-082)	1.3E+04	3x
	NARSA NRS705 (NY-12)	1.3E+04	3x
	NARSA NRS707 (NY-155)	1.3E+04	3x
	NARSA NRS745 (CA-629)	1.3E+04	3x
	BEI NR-46081 (HIP12899)	1.3E+04	3x
	GRE 0860042	1.3E+04	3x
	NARSA NRS648 (CA-347)	1.3E+04	3x
	SHSC Sunl	1.3E+04	3x
	ATCC 4330 (F-182)	4.2E+03	1x
	ATCC 12600	1.3E+04	3x
	ATCC 14154	1.3E+04	3x
Staphylococcus
aureus ssp.
aureus	ATCC 25923 (Seattle 1945)	1.3E+04	3x
	ATCC BAA-39	1.3E+04	3x
	ATCC BAA-42 (HDE288)	1.3E+04	3x
	ATCC BAA-44 (HPV107)	1.3E+04	3x
	ATCC BAA-1717 (TCH1516)	1.3E+04	3x
	ATCC BAA-1720 (MRSA252)	1.3E+04	3x
Staphylococcus
aureus ssp.
anaerobius	ATCC 35844 (MVF-7)	1.3E+04	3x

Table 17: Staphylococcus lugdunensis Isolates Tested

Organism	Isolate ID (Strain)	Test Concentration		Result
		(copies/mL)	xLoD
Staphylococcus
lugdunensis	ATCC 43809 (N860297)	2.6E+03	1x
	ATCC 49576 (LRA 260.05.09)	7.8E+03	3x
	ATCC 700582 (7829)	7.8E+03	3x	S. aureus
	NCTC 7990 (Kelly)	7.8E+03	3x	Detected
	ATCC 700328 (6733)	7.8E+03	3x

Table 18: Isolates Streptococcus spp. Isolates Tested

Organism	Isolate ID (Strain)	Test Concentration		Result
		(copies/mL)	xLoD
Streptococcus agalactiae	See S. agalactiae table
Streptococcus dysgalactiae ssp.
dysgalactiae	ATCC 43078 (NCDO 2023)	7.5E+05	3x	Streptococcus spp.
Detected
Streptococcus dysgalactiae ssp.
equisimilis	ATCC 8543 (LRA 06 11 76)	7.5E+05	3x
Streptococcus bovis/equinus	ATCC 33317 (Pearl 11 NCDO 597)
ATCC 9812 (H 12 B)	7.5E+05	3x
Streptococcus gallolyticus ssp.
gallolyticus	DSM 16831	7.5E+05	3x

in Physics Company T

24

| Streptococcus gallolyticus ssp.

pasteurianus	ATCC 7000338 (RG)	7.5E+05	3x
Streptococcus infantarius ssp.
infantarius	ATCC BAA-102 (NCIMB 700599)	7.5E+05	3x
Streptococcus anginosus	ATCC 33397 (Havill III R. Lancefield
F68A)	7.5E+05	3x
Streptococcus constellatus	ATCC 27513 (VPI 7712)	7.5E+05	3x
Streptococcus intermedius	ATCC 27335 (VPI 3372A)	7.5E+05	3x
Streptococcus salivarius	ATCC 13419 (C699 [S30D])	7.5E+05	3x
Streptococcus salivarius ssp.
thermophilus	ATCC 19258 (NCDO 573)	7.5E+05	3x
Streptococcus vestibularis	ATCC 49124 (MM1)	7.5E+05	3x
Streptococcus australis	ATCC 700641 (AI-1)	7.5E+05	3x
Streptococcus sobrinusa	ATCC 33478 (SL1)	7.5E+05	3x
Streptococcus mutans	ATCC25175 (IFO 13955)	2.50E+05	1x
Streptococcus gordonii	ATCC 10558 (SK3)	7.5E+05	3x
Streptococcus mitis	ATCC 49456 (NS 51: SK142)	7.5E+05	3x
Streptococcus oralisa	ATCC 35037 (PB182; LVG/1)	7.5E+05	3x
Streptococcus oralis ssp. tigurinus	DSM 24864 (AZ 3a)	7.5E+05	3x
Streptococcus pneumoniae	See S. pneumoniae table
Streptococcus pseudopneumoniae	ATCC BAA-960 (CDC-SS-1757)	7.5E+05	3x
Streptococcus pyogenes	See S. pyogenes table
Streptococcus sanguinis	ATCC 10556 (DSS-10)	7.5E+05	3x
Streptococcus cristatus	ATCC 51100 (CR311)	7.5E+05	3x
Streptococcus parasanguinis	ATCC 15912 (SS 898)	7.5E+05	3x
Streptococcus peroris	ATCC 700780 (GTC 848)	7.5E+05	3x
Streptococcus equi ssp. equib	ATCC 33398 (C15)	7.5E+05	3x
Streptococcus equi ssp. zooepidemicusb	ATCC 43079 (NCDO 1358)	7.5E+05	3x
Streptococcus suisa	ATCC 43765 (735)	7.5E+05	3x
Streptococcus sinensis	DSM 14990 (HKU4)	7.5E+05	3x

4In silico analysis identified sequence variation that is predicted to impact reactivity in approximately 8% of S. oralis sequences evaluated and in approximately 2% of the S. sobrinus, S suis, and S. uberis sequences evaluated. b Although the two isolates of S. equi tested were detected near LoD, in silico analysis predicts some impairment of detection for most (97%) S. equi sequences.

Organism	Result
Streptococcus acidominimus
Streptococcus azizii
Streptococcus bovimastitidis	Streptococcus
spp. Detected
Streptococcus caballi
Streptococcus canis
Streptococcus castoreus
Streptococcus criceti
Streptococcus cuniculi
Streptococcus devriesei
Streptococcus didelphis
Streptococcus downei
Streptococcus ferus
Streptococcus halotolerans
Streptococcus henryi
Streptococcus himalayensis
Streptococcus hongkongensis
Streptococcus hyointestinalis
Streptococcus ictaluri
Streptococcus infantis
Streptococcus iniae
Streptococcus intermedius
Streptococcus lactarius
Streptococcus lutetiensis
Streptococcus macacae
Streptococcus marimammalium
Streptococcus marmotae
Streptococcus massiliensis
Streptococcus merionis
Streptococcus milleri
Streptococcus minora
Streptococcus orisasini
Streptococcus orisratti
Streptococcus ovis
Streptococcus parasuis
Streptococcus parauberis
Streptococcus penaeicida
Streptococcus phocae
Streptococcus pluranimalium
Streptococcus plurextorum
Streptococcus porci
Streptococcus porcinus
Streptococcus pseudoporcinus
Streptococcus ratti
Streptococcus respiraculi
Streptococcus ruminantium
Streptococcus thoraltensis
Streptococcus troglodytae
Streptococcus uberisb
Streptococcus urinalis
Streptococcus entericus
Streptococcus halitosis
Streptococcus hyovaginalis
Streptococcus pantholopis

25

26

31n silico analysis identified 1/2 (50%) S. minor sequence variation that is predicted to impact reactivity

bIn silico analysis identified sequence variation that is predicted to impact reactivity in approximately 8% of S. oralis sequences evaluated and in approximately 2% of the S. sobrinus, S suis, and S. uberis sequences evaluated.

Organism	Isolate ID (Strain)	Test Concentration		Result
		(copies/mL)	xLoD
Streptococcus
agalactiae	ATCC 13813 (G19)	1.9E+04	1x	Streptococcus
agalactiae Detected
	ATCC 12403 [D136C(3)]	5.7E+04	3x
	ATCC BAA-2669 (5030-08)	5.7E+04	3x
	CI 2460	5.7E+04	3x
	ATCC BAA-611 (2603 V/R)	5.7E+04	3x
	ATCC 12386 (090R)	5.7E+04	3x

Table 20: Streptococcus agalactiae Isolates Tested

Table 21: Streptococcus pneumoniae Isolates Tested

Organism	Isolate ID (Strain)	Test Concentration		Result
		(copies/mL)	xLoD
Streptococcus
pneumoniae	ATCC 6303 (CIP 104225)	5.3E+02	1x	Streptococcus
pneumoniae
Detected
	ATCC 33400 (SV1)	1.6E+03	3x
	ATCC 700672 (VH14)	1.6E+03	3x
	ATCC 700673 (19A-6 [HUN663])	1.6E+03	3x
	ATCC BAA-1409 (62076)	1.6E+03	3x
	ATCC BAA-341 (SPN1439-106)	1.6E+03	3x

Table 22: Streptococcus pyogenes Isolates Tested

Organism	Isolate ID (Strain)	Test Concentration		Result
		(copies/mL)	xLoD
Streptococcus pyogenes	ATCC 19615 (Bruno)	8.9E+03	1x	Streptococcus
pyogenes Detected
	ATCC 49399 (QC A62)	2.7E+04	3x
	ATCC 12344 (T1)	2.7E+04	3x
	ATCC 12348 (S43)	2.7E+04	3x
	ATCC 700294 (SF370;M1 GAS)	2.7E+04	3x
	ATCC BAA-947 (MGAS 5005)	2.7E+04	3x
	ATCC 12384 (C203)	2.7E+04	3x
	P-03-0543 804 ISOª	8.9E+05	100x	Streptococcus
pyogenes
Not Detected

3Isolate was from a clinical specimen; a deletion in the gene target was identified that prevents amplification/detection

Table 23: Bacteroides fragilis Isolates Tested

Organism	Isolate ID (Strain)	Test Concentration		Result
		(copies/mL)	xLoD

27

| Bacteroides fragilis | ATCC 25285 (VPI 2553) | 1.1E+03 | 1x | Bacteroides fragilis
Detected |
|----------------------|-------------------------------------|---------|----|----------------------------------|
| | ATCC BAA-2283 (2-1-56 FAA) | 3.3E+03 | 3x | |
| | ATCC 29768 (12256/P8) | 3.3E+03 | 3x | |
| | ATCC 29771 (2044 [CDC 1261; M-488]) | 3.3E+03 | 3x | |
| | ATCC 43937 (F1355 [WAL 78-189A]) | 3.3E+03 | 3x | |

Table 24: Citrobacter Isolates Tested

| Organism | Isolate ID (Strain) | Test Concentration
(copies/mL) xLoD | | Result |
|--------------------------|------------------------------------------------|----------------------------------------|------|--------------------------------|
| Citrobacter braakii | ATCC 51113 (CDC 80-58) | 1.4E+04 | 3x | Citrobacter
Detected |
| Citrobacter europaeus | GRE 1953016 | 1.4E+04 | 3x | |
| Citrobacter freundii | ATCC 8090 (ATCC 13316) | 4.7E+03 | 1x | |
| | ATCC 43864 (LRA 117.03.76) | 1.4E+04 | 3x | |
| | AR Bank #0116 | 1.4E+04 | 3x | |
| | AR Bank #0157 | 1.4E+04 | 3x | |
| GRE 1062177 | 1.4E+04 | 3x | | |
| Citrobacter koseri | ATCC 27156 (CDC 3613-63) | 1.4E+04 | 3x | Citrobacter
Detected |
| | ATCC 27028 (14804) | 1.4E+04 | 3x | |
| Citrobacter murliniae | ATCC 51118 (CDC 2970-59) | 1.4E+04 | 3x | Citrobacter
Detected |
| Citrobacter werkmaniiaa | ATCC 51114 (CDC 0876-58) | 1.4E+04 | 3x | Citrobacter
Detected |
| Citrobacter youngae | ATCC 29935 (460-61) | 1.4E+04 | 3x | Citrobacter
Detected |
| Citrobacter amalonaticus | ATCC 25405 (9823) | 8.0E+08 | High | Citrobacter
Not
Detected |
| Citrobacter farmeri | ATCC 51112 (CDC 2991-81) | 8.0E+08 | High | |
| Citrobacter gillenii | ATCC 51117 (CDC 4693-86) | 8.0E+08 | High | |
| Citrobacter rodentium | GRE 1654045 | 8.0E+08 | High | |
| Citrobacter sedlakii | ATCC 51494 | 8.0E+08 | High | |
| Citrobacter cronae | Unknown Reactivity
(no sequence/not tested) | | | |

ª In silico analysis identified sequence variation that is predicted to impact reactivity in 4/6 (50%) C. werkmanii sequences

Table 25. Citrobacter Reactivity Predicted (in silico)

Organism	Result
Citrobacter pasteurii	Citrobacter
Citrobacter portucalensis	Detected

Table 26: Enterobacter cloacae complex Isolates Tested

Organism	Isolate ID (Strain)	Test Concentration		Result
		(copies/mL)	xLoD
Enterobacter asburiaea	GRE 1753006	3.9E+05	3x	Enterobacter
cloacae
Detected
	ATCC 35953 (CDC 1497-78)	1.3E+07	100x	Enterobacter
cloacae

28

				Not Detected
Enterobacter cloacaea	AR Bank #0154	1.3E+05	1x
Enterobacter cloacae ssp.
cloacaea	NCTC 13464	3.9E+05	3x
	ATCC 13047 (CDC 442-68)	3.9E+05	3x
	ATCC 222 (CDC 435)	3.9E+05	3x
Enterobacter cloacae ssp.
dissolvensa	ATCC 23373D-5 gDNA (ICPB ED105)	3.9E+05	3x
Enterobacter hormaechei	ATCC BAA-2082	3.9E+05	3x
	ATCC 700323	3.9E+05	3x
Enterobacter hormaechei ssp.
hormaechei	ATCC 49162	1.3E+07	100x	Enterobacter
cloacae
Detected
Enterobacter hormaechei ssp.
oharea	CCUG 53905T	3.9E+05	3x
Enterobacter hormaechei ssp.
steigerwaltii	CCUG 53904T	3.9E+05	3x
Enterobacter hormaechei ssp.
xiangfangensis	DSM 46348	3.9E+05	3x
Enterobacter kobei	GRE 1753004	3.9E+05	3x
Enterobacter ludwigiia	DSM 16688 (EN-119)	3.9E+05	3x
	CCUG 23050	3.9E+05	3x
Enterobacter mori	DSM 26271 (R18-2)	3.9E+05	3x
Enterobacter roggenkampii	DSM 16690 (EN-117)	3.9E+05	3x

4 Enterobacter asburiae isolate ATCC 35953 has sequence variation under assay primers that impairs detection at 100x LoD and lower. A similar impact on reactivity is predicted for 6/76 (7.9%) Enterobacter asburiae sequences evaluated and impaired amplification and detection is also predicted for a subset of E. cloacae (8/516, 1.6%) and E. ludwigii (2/25, 8.0%) sequences.

bE. hormaechei ssp. hormaechei isolate ATCC 49162 has sequence variation under assay primers that impairs detection at 10x LoD and lower. A similar impact on reactivity is predicted for 10/685 (1.4%) E. hormacchei and 1/8 (12.5%) E. mori sequences evaluated.

Organism	Isolate ID (Strain)	Test Concentration		Result
		(copies/mL)	xLoD
E. coli	CDC AR Bank #0150	6.0E+03	1x	E. coli
Detected
	CDC AR Bank #0137	1.8E+04	3x	E. coli
Detected
	CDC AR Bank #0162	1.8E+04	3x
	CDC AR Bank #0086	1.8E+04	3x
	CDC AR Bank #0061	1.8E+04	3x
	ATCC 11775 (9001 U 5/41)	1.8E+04	3x
	GRE 1256018	1.8E+04	3x
	Zeptometrix 0801905 (Z136)	1.8E+04	3x

Table 27: E. coli Isolates Tested

Table 28: Haemophilus influenzae Isolates Tested

| Organism | Serotype | Biotype | Isolate ID (Strain) | Test
Concentration | | Result |
|-----------------------------------|----------|---------|----------------------------------------|-----------------------|------|-----------------------------------|
| | | | | (copies/mL) | xLoD | |
| Haemophilus
influenzae | b | I | ATCC 10211 (AMC 36-A-1 [572]) | 6.9E+02 | 1x | Haemophilus
influenzae |
| Haemophilus
influenzae | a | | ATCC 9006 (AMC 36-A-3 [610, PCM 2436]) | 2.1E+03 | 3x | Haemophilus
influenzae |

29

c		ATCC 49699 (C 9007)	2.1E+03	3x	Detected
d	-	ATCC 9008 (AMC 36-A-6 [611])	2.1E+03	3x
e		ATCC 8142 (595 Murray Biotype IV: AMC 36-A-7 [595])	2.1E+03	3x
f		ATCC 700223 (GA 1264)	2.1E+03	3x
Non-typeable	II	ATCC 33391 (680 Biotype II)	2.1E+03	3x
	V	ATCC 51997 (INT 1 Biotype V)	2.1E+03	3x
Unknown	-	BF Clinical Isolate 006433-PBC-1-0029-ISO-1a	7.5E+09 CFU/mL	High	Haemophilus influenzae Not Detected

4Isolate was obtained from a clinical specimen; a deletion in the gene target was identified that prevents amplification/detection.

Table 29: Kingella kingae Isolates Tested

| Organism | Isolate ID (Strain) | Test
Concentration | | Result |
|-----------------|----------------------|-----------------------|------|------------------------|
| | | (copies/mL) | xLoD | |
| Kingella kingae | ATCC 23330 (4177/66) | 3.4E+03 | 1x | |
| | ATCC 23331 | 1.0E+04 | 3x | Kingella Kingae |
| | CCUG 63569 | 1.0E+04 | 3x | |
| | CCUG 50167A | 1.0E+04 | 3x | Detected |
| | CCUG 44801 | 1.0E+04 | 3x | |

Table 30: Klebsiella aerogenes Isolates Tested

| Organism | Isolate ID (Strain) | Test
Concentration
(copies/mL) | | Result |
|----------------------|--------------------------|--------------------------------------|------|---------------------------------|
| Klebsiella aerogenes | AR Bank #0074 | 7.5E+03 | 1x | Kingella Kingae |
| Klebsiella aerogenes | AR Bank #0062 | 2.3E+04 | 3x | Detected |
| Klebsiella aerogenes | AR Bank #0161 | 2.3E+04 | 3x | Detected |
| Klebsiella aerogenes | ATCC 13048 (NCDC 819-56) | 2.3E+04 | 3x | Detected |
| Klebsiella aerogenes | ATCC 29751 (MULB-250)a | 7.5E+05 | 100x | Kingella Kingae
Not Detected |
| Klebsiella aerogenes | GRE 1254066 | 2.3E+04 | 3x | Kingella Kingae
Detected |

3Klebsiella aerogenes isolate ATCC 29751 has sequence variation under the assay primers that impairs detection at 100x LoD and lower. A similar impact on reactivity is predicted for 9/193 (4.7%) Klebsiella aerogenes sequences evaluated.

Table 31: Klebsiella pneumoniae group Isolates Tested

Organism	Isolate ID (Strain)	Test Concentration		Result
		(copies/mL)	xLoD
Klebsiella pneumoniae	AR Bank #0097	1.6E+04	1x	Klebsiella
pneumoniae
Detected
	AR Bank #0079	4.8E+04	3x
	AR Bank #0107	4.8E+04	3x
	AR Bank #0075	4.8E+04	3x

30

	JMI 766	4.8E+04	3x
	AR Bank #0040	4.8E+04	3x
	AR Bank #0068	3.1E+04	1.9x
	AR Bank #0080	4.5E+04	2.5x
Klebsiella pneumoniae ssp.	ATCC 11296	4.8E+04	3x
ozaenae	AR Bank #0051	4.8E+04	3x
Klebsiella pneumoniae ssp.
pneumoniae	ATCC 13883	4.8E+04	3x
Klebsiella pneumoniae ssp.
rhinosclermatis	ATCC 13884 (R-70)	4.8E+04	3x
Klebsiella quasipneumoniae	DSM 28211	4.8E+04	3x
Klebsiella quasipneumoniae
ssp. similipneumoniae	DSM 28212	4.8E+04	3x
Klebsiella variicola	ATCC BAA-830 (F2R9)	4.8E+04	3x

Table 32: Morganella morganii Isolates Tested

| Organism | Isolate ID (Strain) | Test
Concentration | | Result |
|--------------------------|--------------------------|-----------------------|------|------------|
| | | (copies/mL) | xLoD | |
| Morganella morganii | AR Bank #0057 | 6.6E+03 | 3x | |
| Morganella morganii ssp. | ATCC 25830 (M11) | 2.2E+03 | 1x | Morganella |
| morganii | ATCC 33791 (Potter) | 6.6E+03 | 3x | morganii |
| Morganella morganii ssp. | ATCC 49948 (CDC 9103-85) | 6.6E+03 | 3x | Detected |
| sibonii | ATCC51207 (CDC 8246-91) | 6.6E+03 | 3x | |

Table 33: Neisseria gonorrhoeae Isolates Tested

| Organism | Isolate ID (Strain) | Test
Concentration | | Result |
|-----------------------|----------------------------|-----------------------|------|------------------------------------------|
| | | (copies/mL) | xLoD | |
| Neisseria gonorrhoeae | ATCC 19424 (B 5025) | 2.2E+03 | 1x | Neisseria
gonorrhoeae
Detected |
| | NCTC 6820 (Gono 4) | 6.6E+03 | 3x | |
| | ATCC 19088 (CH-6) | 6.6E+03 | 3x | |
| | ATCC 700825 (FA1090) | 6.6E+03 | 3x | |
| | Zeptometrix 0801482 (Z017) | 6.6E+03 | 3x | |
| | NCTC 13817a | 8.0E+08
CFU/mL | High | Neisseria
gonorrhoeae
Not Detected |

41solate (also described as WHO-U strain) carries an atypical variant of the gene target (suspected horizontal transfer with homologous gene in N. meningitidis) that is not amplified/detected by the assay.

Table 34: Proteus spp.ª Isolates Tested

Organism	Isolate ID (Strain)	Test Concentration		Result
		(copies/mL)	xLoD
Proteus salimentorum	In silico prediction (not tested)
Proteus columbae	In silico prediction (not tested)
Proteus hauseri	ATCC 13315 (Lehmann)	1.6E+04	3x	Proteus spp.
Detected
Proteus hauseri	ATCC 700826 (CDC 1732-80)	1.6E+04	3x
Proteus mirabilis	ATCC 35659 (LRA 08 01 73)	5.2E+03	1x

31

	ATCC 29906 (CDC PR 14)	1.6E+04	3x
	AR Bank #0156	5.2E+04	10x
	AR Bank #0159	5.2E+04	10x
	GRE1254053	1.6E+04	3x
Proteus penneri	ATCC 33519 (CDC 1808-73)	1.6E+04	3x
Proteus penneri	ATCC 35197 (CDC 1655-67)	1.6E+04	3x
Proteus terrae	DSM 29910 (N5/687)	1.6E+04	3x
Proteus terrae ssp. cibarius	DSM 100173 (JS9)	1.6E+04	3x
Proteus vulgaris	ATCC 29905 (CDC PR1)	1.6E+04	3x
Proteus vulgaris	ATCC 27973 (CDC 1787-64-SC1)	1.6E+04	3x

aThe Proteus genus now also includes the species P. cibi and P. faecis. Reactivity with these species has not been evaluated.

Table 35: Pseudomonas aeruginosa Isolates Tested

| Organism | Isolate ID (Strain) | Test
Concentration | | Result |
|------------------------|---------------------------|-----------------------|------|-------------------------------------------|
| | | (copies/mL) | xLoD | |
| Pseudomonas aeruginosa | CDC AR Bank #0092 | 1.30E+04 | 1x | Pseudomonas
aeruginosa
Detected |
| | ATCC 27853 (Boston 41501) | 3.9E+04 | 3x | |
| | CDC AR Bank #0090 | 3.9E+04 | 3x | |
| | CDC AR Bank #0100 | 3.9E+04 | 3x | |
| | CDC AR Bank #0054 | 3.9E+04 | 3x | |
| | CUSM PS28 | 3.9E+04 | 3x | |
| | NCTC 13437 | 3.9E+04 | 3x | |
| | CDC AR Bank #0111 | 3.9E+04 | 3x | |
| | CDC AR Bank #0064 | 3.9E+04 | 3x | |
| | CDC AR Bank #0103 | 1.3E+05 | 10x | |
| | ATCC 9027a | 1.3E+06 | 100x | |
| | ATCC 25619b | 8.0E+08
CFU/mL | High | Pseudomonas
aeruginosa
Not Detected |

ª Pseudomonas aeruginosa isolate ATCC 9027 has sequence variation under assay primers that impairs detection at 10× LoD and lower. Detection was observed in all replicates at 100× LoD (~1.3E+06 copies/mL). Similar impacts on reactivity are predicted for approximately 50/1524 (3.3%) P. aeruginosa sequences evaluated.

b Pseudomonas aeruginosa isolate ATCC 25619 has sequence variation under assay primers that prevents amplification and detection.

Table 36: Salmonella spp. Isolates Tested

Organism	Isolate ID (Strain)	Serovar	Test Concentration		Result
			(copies/mL)	xLoD
Salmonella bongori	SGSC 3100/SarC11 (RKS3041)	-	4.8E+03	3x
Salmonella bongori	NCTC 10946 (BR 1859 66:z41:-)	Brookfield	4.8E+03	3x
Salmonella bongori	ATCC 43975 (1224.72)	-	4.8E+03	3x
Salmonella enterica ssp.
arizonae	ATCC 13314 (DC5.CIP 8230)	-	4.8E+03	3x	Salmonella spp.
Detected
Salmonella enterica ssp.
diarizonae	SGSC 3069 (RKS2979; SarC8)	-	4.8E+03	3x
Salmonella enterica ssp.
enterica	CDC AR Bank#0407	Concord	1.6E+03	1x
Salmonella enterica ssp.
enterica	ATCC 700720 (LT2)	Typhimirium	4.8E+03	3x

32

	ATCC BAA-708	Enteritidis	$4.8E+03$	3x
	SGSC 2210 (SARA30)	Heidelberg	$4.8E+03$	3x
	ATCC BAA-710 (G4639)	Montevideo	$4.8E+03$	3x
	CDC AR Bank #0127	Senftenberg	$4.8E+03$	3x
	ATCC 700931D-5 (Ty2)	Typhi	$4.8E+03$	3x
Salmonella enterica ssp.
houtenae	SGSC 3074 (RKS3015)	-	$4.8E+03$	3x
Salmonella enterica ssp. indica	SGSC 3116 (RKS2995)	-	$1.6E+04$	10x
Salmonella enterica ssp.
salamae	SGSC 3047 (RKS2993)	-	$4.8E+03$	3x

Table 37: Serratia marcescens Isolates Tested

Organism	Isolate ID (Strain)	Test Concentration		Result
		(copies/mL)	xLoD
Serratia marcescens	API 1411137	3.3E+04	3x	Serratia
marcescens
	API 1512393	3.3E+04	3x
	CDC AR Bank #0091	3.3E+04	3x
	JMI 697	3.3E+04	3x
Serratia marcescens ssp.
marcescens	ATCC 13880 (BS 303)	1.1E+04	1x	Detected
Serratia marcescens ssp.
sakuensis	ATCC BAA-885 (KRED)	3.3E+04	3x

Table 38: Candida spp. Isolates Tested

Organism	Isolate ID (Strain)	Test Concentration		Result
		(copies/mL)	xLoD
Candida albicans	See Table 41
Candida dubliniensis	ATCC MYA-646 (CBS 7987)	3.0E+03	3x
	ATCC MYA-578 (H12)	3.0E+03	3x
Candida glabrata	ATCC 15545 (NRRL YB-4025)	3.0E+03	3x
	ATCC 2001 (CBS 138)	3.0E+03	3x
	CI-953	3.0E+03	3x	Candida spp.
Detected
Candida krusei
(Issatchenkia orientalis)	ATCC 6258	1.0E+03	1x
	ATCC 28870 (CBS 2052)	3.0E+03	3x
Candida orthopsilosis	ATCC 96139 (MCO457)	3.0E+03	3x
	ATCC 28475 (CBS 2915)	3.0E+03	3x
Candida parapsilosis	ATCC 22019 (CBS 604)	3.0E+03	3x
	ATCC 750	3.0E+03	3x
Candida tropicalis	ATCC 66029 (AmMS 227)	3.0E+03	3x
Candida metapsilosis	ATCC96143 (MCO429)	1.0E+04	10x
Candida sojae	NRRL Y-17909	1.0E+04	10x
Candida sphaerica	GRE 1951001	1.0E+05	100x
Candida inconspicua	ATCC 16783 (CBS 180)	1.0E+05	100x
	AR Bank #0381	1.0E+05	100x
Candida auris	AR Bank #0385	1.0E+05	100x
	GRE 1756004	8.0E+06	8000x	Candida spp.
Detected

33

| Candida lusitaniae | ATCC 42720 (45090) | 1.0E+05 | 100x | Candida spp.
Not Detected |
|------------------------|------------------------------|---------|----------|------------------------------|
| Candida nivariensis | ATCC 34449 (IFO 1019) | 8.0E+06 | 8000x | |
| Candida Intermedia | CCUG 56432 | 8.0E+08 | 795,000x | |
| Candida kefyr | ATCC 14439 | 8.0E+06 | 8000x | |
| Candida norvegensis | ATCC 204093 | 8.0E+06 | 8000x | Candida spp. |
| Candida utilis | GRE 0856055 | 8.0E+06 | 8000x | Detected |
| Candida haemoluniia | ATCC 22023 | 8.0E+06 | 8000x | |
| Candida viswanathii | AR Bank #0393 | 3.9E+08 | 389,000x | |
| Candida guilliermondii | ATCC 22981 | 4.3E+08 | 430,000x | |
| Candida ciferrii | ATCC 38290 (Tu 62304-2) | 5.7E+08 | 570,000x | |
| Candida colliculosa | ATCC 584433 (CBS 5295) | 8.0E+06 | 8000x | |
| Candida holmii | ATCC 10662 (NRRL Y-866) | 8.0E+06 | 8000x | |
| Candida lipolytica | DSM 70627 | 8.0E+06 | 8000x | Candida spp.
Not Detected |
| Candida rugosa | ATCC 18944 (NRRL YB-423-12) | 8.0E+06 | 8000x | Candida spp.
Not Detected |
| Candida thermophila | ATCC 10571 (NRRL Y-1496) | 8.0E+06 | 8000x | |
| Candida famata | ATCC 58401 | 8.0E+06 | 8000x | |
| | ATCC 4144 (D.R. 1658 No. 14) | 8.9E+08 | 893,000x | |

aSpecies may be detected at high concentration (>100x LoD).

Table 39: Candida spp. Predicted Reactivity (In silico)

Organism	Result
Candida duobushaemulonis
Candida fabianii
(Cyberlindner fabianii)	Candida spp.
Not Detected
Candida fermentati
(Myerozyma carribica)
Candida jadinii
Candida pelliculosa
(Wickerhamomyces anomalus)

Table 40: Candida albicans Isolates Tested

| Organism | Isolate ID (Strain) | Test
Concentration | | Result |
|------------------|-----------------------|-----------------------|------|------------------|
| | | (copies/mL) | xLoD | |
| Candida albicans | ATCC 90028 (NCCLS 11) | $5.0E+02$ | 1x | |
| Candida albicans | ATCC 10231 (3147) | $1.5E+03$ | 3x | Candida albicans |
| Candida albicans | ATCC 11006 | $1.5E+03$ | 3x | |
| Candida albicans | ATCC 14053 (NIH 3172) | $1.5E+03$ | 3x | Detected |
| Candida albicans | ATCC 22972 (M 97) | $1.5E+03$ | 3x | |

The following tables describe the reactivity of the AMR genes assays with different AMR gene types in various host bacteria. Results are shown for the isolates tested as well as

34

predictions of reactivity with untested AMR gene types based on in silico analysis of sequences retrieved from public databases.

| CTX-M Type | Organism | Isolate ID | Test concentration
(copies/mL) | Result |
|-----------------------------------|---------------------------|---------------|-----------------------------------|----------------|
| CTX-M-3 | E. coli | NCTC 13452 | 4.1E+05 | |
| CTX-M-3 | S. flexneri | AR Bank #0421 | 4.5E+03 | |
| CTX-M-14 | K. pneumoniae | AR-Bank #079 | 4.5E+03 | |
| CTX-M-14 | C. freundii | GRE 1062177 | 4.5E+03 | |
| CTX-M-15 | K. pneumoniae | AR Bank #0075 | 4.5E+03 | |
| CTX-M-15 | K. pneumoniae | AR-Bank #0040 | 4.5E+03 | |
| CTX-M-15 | M. morganii | AR-Bank #0057 | 4.5E+03 | |
| CTX-M-15 | S. enterica ssp. enterica | AR-Bank #0407 | 1.5E+03 | CTX-M Detected |
| CTX-M-22 | P. mirabilis | GRE 1254053 | 4.5E+03 | |
| CTX-M-55 | E. coli | AR Bank #0346 | 4.5E+03 | |
| CTX-M-2 | K. pneumoniae | AR Bank #0107 | 4.5E+03 | |
| CTX-M-124 | K. ascorbatea | AR Bank #0144 | 4.5E+03 | |
| CTX-M-8 | E. coli | NCTC 13463 | 4.5E+03 | |
| CTX-M-9 | E. cloacae | NCTC 13464 | 4.5E+03 | |
| CTX-M-25 | K. pneumoniae | NCTC 13465 | 4.5E+03 | |
| In Silico Reactivity Predictionsb | | | | |
| Detected | | Not Detected | Unknown Reactivity | |
| CTX-M-1 – CTX-M-69 | CTX-M-71 – CTX-M-73 | CTX-M-74 | CTX-M-70 | CTX-M-140 |
| CTX-M-76 – CTX-M-112 | CTX-M-114 – CTX-M-117 | CTX-M-75 | CTX-M-119 | CTX-M-143 |
| CTX-M-121 – CTX-M-127 | CTX-M-129 – CTX-M-132 | CTX-M-113 | CTX-M-120 | CTX-M-145 |
| CTX-M-134 | CTX-M-136 – CTX-M-139 | CTX-M-151 | CTX-M-128 | CTX-M-149 |
| | CTX-M-141 – CTX-M-142 | | CTX-M-133 | CTX-M-153 |
| | CTX-M-144 | | CTX-M-135 | CTX-M-154 |
| | CTX-M-146 – CTX-M-148 | | | |
| | CTX-M-150 | | | |
| | CTX-M-152 | | | |
| | CTX-M-155 – CTX-M-229 | | | |

Table 41: Isolates Tested Containing the blaCTX-M gene and In Silico Predicted Reactivity for CTX-M Types

31solate was tested only to evaluate CTX-M assay reactivity, the species is not detected by the panel.

bA subset of CTX-M sequences (K. pneumoniae | AR-Bank #0080 | 3.9 E+03 | |
| IMP-8 | E. cloacae | AR Bank #0502 | 3.9 E+03 | |
| IMP-8 | K. pneumoniae | GRE 1062084 | 3.9 E+03 | |
| IMP-13 | K. pneumoniae | Zeptometrix 0801904 | 3.9 E+03 | |
| IMP-14 | P. aeruginosa | AR Bank #0092 | 1.3 E+03 | |
| In Silico Reactivity Predictions | | | | |
| | Detecteda | Not Detected | Unknown Reactivity | |
| | IMP-1 – IMP-30 | IMP-31 | IMP-36 | |
| | IMP-51 – IMP-56 | IMP-35 | IMP-47 | |
| | IMP-32 – IMP-34 | IMP-46 | IMP-50 | |
| | IMP-58 – IMP-64 | | IMP-57 | |
| | IMP-37 – IMP-45 | | IMP-65 | |
| | IMP-66 – IMP-84 | | | |
| | IMP-48 – IMP-49 | | | |

4 pproximately 10% of IMP sequences of various types have mismatches to the assay primer(s) that may impact detection.

Table 43: Isolates Tested Containing the blaKPC gene and In Silico Predicted Reactivity
for KPC Typesa

| KPC Type | Organism | Isolate ID | Test concentration
(copies/mL) | Result |
|----------|-----------------------|---------------|-----------------------------------|--------------|
| KPC-2 | C. freundii | AR Bank #0116 | 1.1 E+03 | |
| KPC-2 | P. aeruginosa | CUDM PS28 | 1.1 E+03 | |
| KPC-2 | S. marcescens | JMI 697 | 1.1 E+03 | |
| KPC-3 | K. pneumoniae | AR Bank #0097 | 3.6 E+02 | KPC Detected |
| KPC-3 | E. coli | AR Bank #0061 | 1.1 E+03 | |
| KPC-4 | Klebsiella pneumoniae | JMI 697 | 1.1 E+03 | |
| KPC-5 | P. aeruginosa | AR Bank #0090 | 1.1 E+03 | |
| KPC-6 | P. mirabilis | AR Bank #0155 | 1.1 E+03 | |
| KPC-11 | K. pneumoniae | AR Bank #0525 | 1.1 E+03 | |
| Unknown | E. hormaechei | BAA-2082 | 1.1 E+03 | |

a In silico analyses predict reactivity with all KPC types (KPC-1 - KPC-46).

Table 44: Isolates Tested Containing mecA/C" and MREJ (MRSA) sequences and In Silico Predicted Reactivity for MREJ Types

36

| Organism | Isolate ID (Strain) MREJ Type | Test concentration
(copies/mL) | Result |
|-------------------------------------------------------------------------------|------------ | Methicillin-
resistant
Staphylococcus
aureus
(MRSA) | NARSA NRS705 (NY-12) | SCCmec Type II | 1.10E+04 MREJ
(MRSA)
Detected |
| | NARSA NRS701 (MN-082) | | ATCC (TCH1516) | | 1.3E+04 | | | NARSA NRS683 (GA-298) | | NARSA NRS662 (CO-34) | | NARSA NRS707 (NY-155) | | ATCC BAA-1707 (MW2) | | NARSA NRS691 (GA-62) | | NARSA NRS648 (CA-347) | | NARSA NRS689 (GA-442) | | ATCC 30137) | SCCmec Type IV | 6.0E+03 | | | BEI NR-46081 (HIP12899) | | ATCC Kansas) | SCCmec Type II | 4.20E+03 | | | ATCC BAA-1720 | | NARSA NRS745 (CA-629) | | ATCC BAA-2312 | | ATCC BAA-2313 | | ATCC BAA-38 | | NARSA NRS686 | | ATCC BAA-44 | | ATCC BAA-42 | | ATCC BAA-39 | | ATCC BAA-40 | | GRE 1062264 | | ATCC BAA-2096 | | GRE 1055015 | | GRE 0860042 | | GRE 1052034 | | GRE 1151100 | | GRE 0960006 | | GRE 1055017 | | GRE 0759163 | | GRE 1057114 | Methicillin-
sensitive
Staphylococcus
aureus
(MSSA) | GRE 1062373 | MREJ Type xvb | 1.20E+04 MREJ
(MRSA) |
| | GRE 1062292 | | ATCC BAA-2421c MREJ
(MRSA)
Detected |
| | Rennes 1060728 DAC | SCCmec Type/
-------------------|---------------------------|-----------------------------------|-------------------------------------------------|
| mecA/C and
| | 1.3E+04 | |
BAA-1717
|
| | 1.3E+04 | |
| SCCmec Type IV | 1.3E+04 | |
| | 1.3E+04 | |
| | 1.3E+04 | |
| | 1.3E+04 | |
| SCCmec Type II or IV | 1.3E+04 | |
| | 1.3E+04 | |
BAA-1700 (HFH-
|
| | 9.7E+03 | |
43300 (F182
|
| | 1.3E+04 | |
| SCCmec Type IV or V | 1.10E+04 | |
| SCCmec Type XI | 1.3E+04 | |
| | 4.2E+03 | |
| MREJ Type i | 1.10E+04 | |
| | 1.3E+04 | |
| MREJ Type ii | 1.3E+04 | |
| | 1.3E+04 | |
| MREJ Type iii | 1.3E+04 | |
| MREJ Type iv | 1.3E+04 | |
| | 1.3E+04 | |
| MREJ Type v | 1.3E+04 | |
| MREJ Type vi | 1.3E+04 | |
| MREJ Type vii | 1.3E+04 | |
| MREJ Type ix | 4.20E+04 | |
| MREJ Type xi | 1.3E+04 | |
| MREJ Type xii | 1.3E+04 | |
| MREJ Type xiii | 1.3E+04 | |
| MREJ Type xiv | 1.3E+04 | |
| MREJ Type xvii | 1.3E+04 | |
| mecA/C and
| MREJ Type xviiii | 8.00E+08 CFU/mL | Not Detected |
| SCCmec Type IIc | 1.3E+04 | mecA/C and
| Empty SCCmec cassette | 4.2E+05 | mecA/C and |

37

| Organism | Isolate ID (Strain) | SCCmec Type/
MREJ Type | Test concentration
(copies/mL) | Result |
|---------------|---------------------|-------------------------------------------------|-----------------------------------|--------------------------------|
| | GRE 1062519 | MREJ Type xixd | 8.9E+09 CFU/mL | MREJ
(MRSA)
Not Detected |
| | | In Silico Reactivity Predictions for MREJ Types | | |
| | Detectede | | Not Detected | Unknown
Reactivity |
| MREJ Type i | MREJ Type vii | MREJ Type xvi | MREJ Type xvb | MREJ Type viii |
| MREJ Type ii | MREJ Type ix | MREJ Type xvii | MREJ Type xviii | MREJ Type x |
| MREJ Type iii | MREJ Type xi | MREJ Type xxi | MREJ Type xixd | |
| MREJ Type iv | MREJ Type xii | | MREJ Type xxd | |
| MREJ Type v | MREJ Type xiii | | | |
| MREJ Type vi | MREJ Type xiv | | | |

41n silico analysis predicts that more than 99.9% of the mecA and mecC sequences evaluated will be detected.

bApproximately 40% of the MREJ type xv - like sequence variation that is prediction that is predicted to substantially impair or prevent detection by the MREJa assay. However, no limitations on detection are predicted for ~60% of MREJ type xv - like sequences evaluated. The prevalence of MREJ type xv, with or without the sequence variation, is currently unknown.

SIsolate carries a mecA gene variant that is amplified by the mecA/C assay but is nonfunctional. Reporting based on genotype will not match the phenotype.

dIsolates with MREJ Types xix and xx have been described as methicillin sensitive.

*Approximately 1% of MREJ sequences of various types have mismatches to the assay primer(s) that may impact detection.

Table 45: Isolates Tested Containing blaNDM gene and In Silico Predicted Reactivity for NDM Types

| NDM Type | Organism | Isolate ID | Test concentration
(copies/mL) | Result |
|----------------------------------|----------------------|---------------|-----------------------------------|--------------|
| NDM-1 | C. freundii | AR Bank #0157 | 3.0 E+04 | NDM Detected |
| | E. cloacae | AR-Bank #0038 | 3.0 E+04 | |
| | M. morganii | AR-Bank #0057 | 3.0 E+04 | |
| | P. mirabilis | AR Bank #0159 | 3.0 E+04 | |
| | P. aeruginosa | AR Bank #0246 | 3.0 E+04 | |
| | S. enterica | AR Bank #0127 | 3.0 E+04 | |
| NDM-2 | A. baumaniia | GRE 1153064 | 3.0 E+04 | |
| NDM-5 | E. coli | AR Bank #0150 | 9.9E+03 | |
| NDM-6 | E. coli | AR Bank #0137 | 1.9E+04 | |
| NDM-7 | K. pneumoniae | AR Bank #0138 | 3.0 E+04 | |
| | | AR Bank #0068 | 3.0 E+04 | |
| In Silico Reactivity Predictions | | | | |
| Detectedb | | | Unknown Reactivity | |
| NDM-1 – NDM-13 | NDM-32 | NDM-14 | NDM-33-39 | |

38

| NDM Type | Organism | Isolate ID | Test concentration
(copies/mL) | Result |
|-----------------|----------|-----------------|-----------------------------------|--------|
| NDM-15-NDM-23 | NDM-40 | NDM-24 - NDM-26 | | |
| NDM-27 - NDM-29 | | NDM-30 - NDM-31 | | |

31solate was tested only to evaluate NDM assay reactivity, the species is not detected by the panel. b Less than 1% of NDM sequences of various types have mismatches to the assay primer(s) that may impact detection.

Table 46: Isolates Tested Containing blaOXA-48-like gene and In Silico Predicted Reactivity for OXA-48-like Types

| OXA-48-like Type | Organism | Isolate ID | Test
concentration | Result | |
|----------------------------------|---------------|---------------|-----------------------|----------------------|---------|
| OXA-48 | K. aerogenes | AR Bank #0074 | 3.1 E+02 | | |
| OXA-48-like | S. marcescens | API 1411137 | 9.3E+02 | | |
| OXA-162 | K. pneumoniae | GRE 1355030 | 9.3E+02 | OXA-48-like Detected | |
| OXA-181 | K. pneumoniae | AR Bank #0051 | 9.3E+02 | | |
| OXA-232 | K. pneumoniae | AR Bank #0075 | 9.3E+02 | | |
| In Silico Reactivity Predictions | | | | | |
| | Detected | | | Not Detecteda,b,c | |
| OXA-48 | OXA-244 | OXA-515 | OXA-54 | OXA-439 | OXA-551 |
| OXA-48-like | OXA-245 | OXA-519 | OXA-163 | OXA-517 | OXA-552 |
| OXA-162 | OXA-252 | OXA-546 | OXA-247 | OXA-535 | OXA-553 |
| OXA-181 | OXA-370 | OXA-547 | OXA-405 | OXA-538 | OXA-567 |
| OXA-199 | OXA-484 | OXA-566 | OXA-416 | OXA-548 | OXA-731 |
| OXA-204 | OXA-505 | | OXA-436 | OXA-549 | |
| OXA-232 | OXA-514 | | OXA-438 | OXA-550 | |

4Non-OXA-48-like types (e.g., OXA-23-like, OXA-40/240like, OXA-51-like, OXA-58-like, OXA-143a-like and OXA-143-like) will not be detected

bOXA-48-like types with altered carbapenem hydrolysis activity will not be detected.

·OXA-48-like types with altered carbapenem hydrolysis activity will not be detected.

Table 47: Isolates Tested Containing vanA/B genes

| van Type | Organism | Isolate ID | Test concentration
(copies/mL) | Result |
|----------|-------------|---------------|-----------------------------------|-----------------|
| vanA | E. faecium | ATCC 700221 | 1.2 E+03 | |
| | | JMI 475 | 3.6E+03 | vanA/B Detected |
| | | ATCC BAA-2318 | 3.6E+03 | |
| | E. faecalis | JMI 12536 | 3.6E+03 | |
| | | ATCC BAA-2573 | 3.6E+03 | |
| vanB | E. faecium | ATCC 51858 | 3.6E+03 | |
| | E. faecalis | ATCC 700802 | 1.5E+04 | |
| | | ATCC 51575 | 1.5E+04 | |
| | | ATCC BAA-2365 | 1.5E+04 | |

39

| van Type | Organism | Isolate ID | Test concentration
(copies/mL) | Result |
|----------|----------|------------|-----------------------------------|--------|
| | | ATCC 51299 | 5.0E+03 | |

Table 48: Isolates Tested Containing blaVIM-like gene and In Silico Predicted Reactivity for VIM Types

| VIM Type | Organism | Isolate ID | Test
concentration
(copies/mL) | Result | |
|----------------------------------|-----------------|---------------|--------------------------------------|--------------|--|
| VIM-1 | E. cloacae | AR Bank #0154 | 3.1E+03 | | |
| VIM-2 | P. aeruginosa | AR Bank #0100 | 9.3E+03 | VIM Detected | |
| VIM-4 | P. aeruginosa | AR Bank #0054 | 9.3E+03 | | |
| VIM-7 | E. coli | GRE 1256018 | 9.3E+03 | | |
| VIM-10 | P. aeruginosa | NCTC 13437 | 9.3E+03 | | |
| VIM-11 | P. aeruginosa | AR Bank #0239 | 9.3E+03 | | |
| VIM-27 | K. pneumoniae | AR Bank #0040 | 9.3E+03 | | |
| In Silico Reactivity Predictions | | | | | |
| Detecteda | Not Detected | | Unknown Reactivity | | |
| VIM-1 - VIM-6 | VIM-47 - VIM-60 | VIM-7 | VIM-61 | VIM-21 | |
| VIM-8 - VIM-20 | VIM-52 - VIM-64 | VIM-39 | VIM-65 | VIM-22 | |
| VIM-23 - VIM-38 | VIM-66 | VIM-45 | VIM-67 | | |
| VIM-40 - VIM-44 | | VIM-46 | | | |

ªApproximately 3% of VIM sequences of various types have mismatches to assay primer(s) that may impact detection.

Exclusivity

The potential for non-specific amplification and detection (cross-reactivity) by the BioFire II Panel assays was evaluated by in silico analysis of available sequences and by testing high concentrations of on-panel and off-panel organisms (and antimicrobial resistance genes). Each organism was tested in triplicate with most bacteria tested at a concentration >1.0E+08 CFU/mL and most yeast tested at a concentration >1.0E+06 CFU/mL. Off-panel fungi, viruses, and parasites were tested at the highest cultured concentration possible.

The on-panel and off-panel organisms tested are listed in Table 50 below. Testing included species and AMR genes that are genetically related to the species or AMR genes detected by the panel (same genus or otherwise related) as well as unrelated organisms that may be found in synovial fluid as pathogens or contaminants (e.g. skin microorganisms, viruses. etc.). All observed or predicted cross-reactivities are indicated. Erroneous results due to cross-reactivity with organisms that were not evaluated or due to cross-reactivity with emerging or novel sequences are also possible.

Table 49. Summary of Observed and Predicted Cross-Reactivity of BioFire JI Panel Assavs

40

BioFire JI Panel Result	Cross-Reactive Organism
Anearococcus prevottii/vaginalis	Anaerococcus degeneri
	Anaerococcus hydrogenalis
	Anaerococcus lactolyticus
	Anaerococcus murdochii
	Anaerococcus nagyae
	Anaerococcus octavius
	Anaerococcus senegalensis
	Anaerococcus tetradius
Bacteroides fragilis	Bacteroides xylanisolvens
	Clostridium cadaveris
Clostridium perfringens	Clostridium fallax
	Enterobacter bugandensisb
Enterobacter cloacae complex	Enterobacter chengduensisb
	Escherichia albertii
	Escherichia fergusonii
	Shigella boydii
Escherichia coli	Shigella dysenteriae
	Shigella flexneri
	Shigella sonnei
Haemophilus influenzae	Haemophilus aegyptius
Kingella kingae	Kingella negevensis
Proteus spp.	Cosenzaea (Proteus) myxofaciens
Staphylococcus aureusb
(and mecA/C and MREJ (MRSA))	Staphylococcus argenteusb
	Staphylococcus schweitzerib
	AMR Genes Derived from Similar Lineages
CTX-Mc	ampC, blaKLU, blaOXY, blaRAHN
vanA/B	vanM

Enterobacter bugandensis and E. chengduensis are recently identified species that are very closely-related to ECC species. Both are indicated as cross-reactive with the Enterobacter cloacae complex assay because their designation as ECC members is currently uncertain.

b Staphylococcus aureus, S. argenteus and S. schweitzeri are closely-related members of the Staphylococcus aureus complex.

CTX-M cross-reactivity with ancestral blakky genes and other related beta-lactamases is predicted to be inefficient and will only occur at high concentrations. The cross-reactive product will only be reported as CTX-M Detected if an applicable gram-negative bacterial species is also detected in the sample.

Table 50. On-Panel and Off-Panel Organisms Tested for Evaluation of BioFire JI Panel Analytical Specificity (Organisms detected or predicted to be detected at high concentration are shown in bold. Grey shading indicates cross-reactivity.)

41

		ON PANEL
		Gram Positive Bacteria
Anaerococcus vaginalis	Peptoniphilus
koenoeneniae	Streptococcus equinus	Streptococcus
oligofermentans
Clostridium perfringens	Peptoniphilus lacrimalis	Streptococcus gallolyticus
(ssp. gallolyticus &
pasteruianus)	Streptococcus peroris
Cutibacterium avidum	Peptoniphilus massiliensisa	Streptococcus gordonii	Streptococcus pneumoniae
Cutibacterium granulosum	Peptoniphilus senegalensis	Streptococcus infantarius	Streptococcus
psseudopneumoniae
Enterococcus faecalis	Peptoniphilus tyrelliae
Streptococcu equis	Peptonipilus allenii	Streptococcus pyogenes
Enterococcus faecium		Peptoniphilus harei	Streptococcus salivarius
(ssp. salivarius &
thermophilus)
Finegoldia magna	Streptococcus agalactiae	Peptoniphilus indolicus	Streptococcus vestibularis
Parvimonas micra	Streptococcus alactolyticus	Peptostreptococcus
anaerobius	Staphylococcus argenteus
Peptoniphilus
asaccharolyticus	Streptococcus anginosus	Peptoniphilus olseniia	Staphylococcus aureus
Peptoniphilus coxia	Streptococcus bovis	Streptococcus australis	Streptococcus oralis
Peptoniphilus duerdeniia	Streptococcus constellatus	Streptococcus intermedius	Streptococcus
parasanguinis
Peptoniphilus gorbachii	Streptococcus cristatus	Streptococcus mitis	Streptococcus sanguinis
Peptoniphilus grossensis	Streptococcus downei	Streptococcus mutans	Streptococcus suis
Peptoniphilus ivoriia	Streptococcus dysgalactiae
(ssp. dysgalactiae &
equismilis)	Staphylococcus
lugdunensis
		Gram Negative Bacteria
Bacteroides fragilis	Citrobacter sedlakiib	Enterobacter mori	Proteus hauseri
Citrobacter braakii	Citrobacter werkmanii	Escherichia coli	Proteus mirabilis
Citrobacter europaeus	Citrobacter youngae	Haemophilus influenzae	Proteus penneri
Citrobacter farmerib	Enterobacter asburiae	Kingella kingae	Proteus vulgaris
Citrobacter freundii	Enterobacter cloacae	Klebsiella aerogenes	Pseudomonas aeruginosa
Citrobacter gilleniib	Enterobacter hormaechei	Klebsiella pneumoniae	Salmonella bongori
Citrobacter koseri	Enterobacter kobei	Morganella morganii	Salmonella enterica
Citrobacter murlinae	Enterobacter ludwigii	Neisseria gonorrhoeae	Serratia marascens
Citrobacter rodentiumb
		Antimicrobial Resistance Genes
mecA/C and MREJ
(MRSA)	blaCTX-M	blaKPC	BlaOXA-48-like
vanA/B	blaIMP	blaNDM	BlaVIM
		Yeast and Fungi
Candida albicans	Candida (Meyerozma)
guilliermondiic,d	Candida metapsilosis	Candida sojae
Candida aurisc	Candida holmiic	Candida nivariensis
(Nakaseomyces
nivariensisa)	Candida sphaerica
(Kluyveromyces lactis)n
Candida
(Trichomonascus) ciferriic	Candida intermediac	Candida (Pichia)
norvegensisc	Candida thermophilac,e
Candida colliculosa
(Torulaspora delbrueckii)	Candida kefyr
(Kluyveromyces
marxianus)c	Candida orthopsilosis	Candida tropicalis
Candida dubliniensis	Candida krusei
(Issatchenkia orientalis)	Candida parapsilosis	Candida utilis
(Cyberlindnera jadinii)c
Candida famata
(Debaryomyces hansenii)c	Candida (Yarrowia)
lipolyticac	Candida (Diutina) rugosac	Candida viswanathiic
Candida glabrata
(Nakaseomyces glabrataa)	Candida (clavispora)
lusitaniaec
		OFF PANEL
		Gram Positive Bacteria
Abaerococcus
senegalensisf	Clostridium fallaxg	Enterococcus pseudoavium	Peptostreptococcus
stomatis
Actinomyces (Schaalia)
odontolyticus	Clostridium ramosum	Enterococcus
saccharolyticus	Propionibacterium
freudenreichii
Actinomyces israelii	Clostridium septicum	Filifactor alocis	Rhodococcus equi
Actinomyces naeslundii	Clostridium sordellii	Gallicola barnesae	Sarcina (Clostridium)
ventriculi
Aerococcus sanquinicola	Clostridium sphenoides	Gemella haemolysans	Slackia
heliotrinitrireducens
Aerococcus urinae	Clostridium sporogenes	Helcococcus kunzii	Staphylococcus argenteus
Aidiproprionbacterium
acidipropionici	Clostridium tertium	Gemella morbllorum	Staphylococcus caprae
Aidiproprionbacterium
jensenii	Clostridium tetani	Gamella sanguinis	Staphylococcus
saprophyticus
Anaerococcus degenerif	Corynebacterium
diphtheriae	Gordonia bronchialis	Staphylococcus capitis
Anaerococcus
hydrogenalisf	Corynebacterium jeikeium	Granulicatella adiacens	Staphylococcus carmosus
Anaerococcus lactolyticusf	Corynebacterium
pseudodiphtheriticum	Lactobacillus casei	Staphylococcus cohnii
Anaerococcus murdochiif	Enterococcus hirae	Lactobacillus salivarius	Staphylococcus epidermidis
Anaerococcus nagyaef	Enterococcus raffinosus	Lactococcus garvieae	Staphylococcus equorun
Anaerococcus octaviusf	Lactobacillus rhamnosus	Lactococcus lactis	Staphylococcus
haemolyticus
Anaerococcus pacaensis	Enterococcus mundtii	Listeria monocytogenes	Staphylococcus pasteuri
Anaerococcus tetradiusf	Cutibacterium
(Proprionibacterium)
acidifaciens	Lysinibacillus sphaericus	Staphylococcus hominis
Atopobium parvulum	Cutibacterium
(Proprionibacterium) acnes	Macrococcus caseolyticus	Staphylococcus intermediu
Bifidobacterium bifidum	Cutibacterium
(Proprionibacterium)
namnetense	Micrococcus luteus	Staphylococcus lutrae
Bifidobacterium dentium	Corynebacterium striatum	Murdochiella
asaccharolytica	Staphylococcus
pseudointermedius
Blautia producta	Corynebacterium
urealyticum	Mycobacterium kansasii	Staphylococcus
saprophyticus
Brevibacterium linens	Enterococcus avium	Mycobacterium abscessus	Staphylococcus schleiferi
Clostridioides
(Clostridium) dificile	Enterococcus casseliflavus	Peptococcus niger	Staphylococcus
schweitzeri h
Clostridium botulinum	Enterococcus cecorum	Nocardia brasiliensis	Staphylococcus warneri
Clostridium butyricum	Enterococcus durans	Mycobacterium marinum	Staphylococcus xylosus
Clostridium cadaveris §	Enterococcus gallinarum	Mycobacterium
tuberculosis	Vagococcus fluvialis
Clostridium
clostridioforme
	Gram Negative Bacteria
Actinobacillus arthritidis	Edwardsiella tarda	Massilia timonae	Pseudomonas otitidis
Acidaminococcus
fermentans	Eikenella corrodens	Megasphaera elsdenii	Pseudomonas
pertucinogena
Acinetobacter
nosocomialis	Enterobacter bugandensis l	Megasphaera indica	Pseudomonas protegens
Acinetobacter schindleri	Enterobacter cancerogenus	Megasphaera massiliensis	Pseudomonas putida
Aeromonas hydrophila	Enterobacter chengduensis l	Moraxella catarrhalis	Pseudomonas stutzeri
Aggregatibacter
actinomycetemcomitans	Escherichia albertii m	Moraxella lacunata	Ralstonia pickettii
Bacteroides dorei	Escherichia coli	Neisseria cinerea	Raoultella ornithinolytic
Bacteroides caccae	Escherichia fergusonii m	Neisseria flava	Raoultella planticola
Bacteroides eggerthii	Escherichia hermannii	Neisseria f;avescens	Serratia ficaria
Bacteroidesforsytus	Escherichia vulneris	Neisseria lactamica	Serratia fonticola
Bacteroides helcogenes	Fusobacterium nucleatum	Neisseria meningitidis	Serratia liquifaciens
Bacteroides stercoris	Haemophilus aegyptius n	Neisseria mucosa	Serratia odorifera
Bacteroides
thetaiotaomicron	Haemophilus ducreyi	Neisseria perflava	Serratia plymuthica
Bacteroides uniformis	Haemophilus haemolyticus	Neisseria sicca	Serratia proteamaculens
Bacteroides vulgatis	Haemophilus
parahaemoluticus	Neisseria subflava	Serratia rubidaea
Bacteroides xylanisolvens l	Haemophilus
parainfluenzae	Pantoea agglomerans	Shewanella algae
Bacteroides ovatus	Haemophilus parasuis	Parabacteroides distasonis	Shewanella denitrificans
Bordetella flabilis	Haemophilus quentini	Parabacteroides merdae	Shewanella putrefaciens
Borrelia burgdorferi	Haemophilus sputorum	Pasteruella multocida	Shigella boydii m
Brucella abortus	Hafnia alvei	Photorhabdus asymbiotica	Shigella dysenteriam i
Brucella melitensis	Hafnia paralveis	Pluralibacter
(Enterobacter( gergoviae	Shigella flexneri m
Brucella suis	Kingella denitrificans	Porphyromonas gingivalis	Shigella sonnei m
Burkholderia mallei	Kingella negevensis o	Prevotella intermedia	Shimwellia blattae
Burkholderia multivorans	Kingella oralis	Prevotella melaninogenica	Stenotrophomonas
maltophilia
Burkholderia pseudomallei	Klebsiella grimontii	Prevotella nigrescens	Stenotrophomonas
rhizophila
Campylobacter jejuni	Klebsiella michiganensis	Providencia rettgeri	Trabulsiella guamensis
Cedecea davisae	Klebsiella oxytoca	Providencia stuartii	Veillonella atypica
Citrobacter amalonaticusl	Klebsiella
quasipneumoniae	Pseudomonas alcaligenes	Veillonella dispar
Coszenaea ( Proteus )
myxofaciensk k	Klebsiella variicola	Pseudomonas chlororaphis	Veillonella parvula
Cronobacter malonaticus	Kluyvera intermedia	Pseudomonas fluorescens	Veillonella rogosae
Cronobacter muytjensii	Kosakonia (Enterobacter)
sacchari	Pseudomonas luteola	Vibrio vulnificus
Cronobacter sakazakii	Lelliotia (Enterobacter)
amnigena	Pseudomonas mendocina	Yersinia enterocolitica
Cronobacter turicensis	Lelliotia (Enterobacter)
nimipressuralis	Pseudomonas nitroreducens
Cronobacter zurichensis
(Siccibacter turicensis)	Leclercia adecarboxylata	Pseudomonas oryzihabitans
Mycoplasma and Intracellular Bacteria
Chlamydia trachomatis	Mycoplasma fermentans	Mycoplasma hominis	Mycoplasma penetrans
Mycoplasma arthritidis	Mycoplasma genitalium	Mycoplasma orale	Ureaplasma urealyticum
Antimicrobial Resistance Genes
vanMP	blaoxyq	OmpC	SME
AmpCq	blaRAHNq	OmpK	SPM
blaKLUA/blaKLUCq	CMY (II)	SHV	TEM
Yeast and Fungi
Aspergillus candidus	Coccidioides immitis	Histoplasma capsulatum	Talaromyces (Penecillium,
marneffei
Aspergillus clavatus	Cryptococcus gattii	Malassexia fufur	Saccharomyces cerevisiae
Aspergillus fumigatus	Cryptococcus neoformans	Malassexia globose	Schizosaccharomyces
pombe
Aspergillus terreus	Exophiala dermatitidis	Neosartorya fischeri	Sporothrix schenckii
Blastomyces dematitidis	Exophiala xenobiotica	Penicillium chrysogenum
Parasites
Chryptosporidium parvum	Entamoeba histolytica
Viruses
Chikungunya Virus	Hepatitis C Virus (HCV)	Human T-cell
Lymphotropic Virus
(HTLV)	Rubella Virus
Dengue Virus	Herpes Simplex Virus 1
(HSV-1)	Parvovirus B19	Varicella Zoster Virus
(VZV)
Epstein Barr Virus (EBV)	Herpes Simplex Virus 2
(HSV-2)	Measles Virus	West Nile Virus
Hepatitis A virus (HAV)	Human Immunodeficiency
Virus (HIV)	Mumps Virus	Zika Virus
Hepatitis B Virus (HBV)

42

43

44

aThe Peptoniphilus assay may not react with several Peptoniphilus species; see Analytical Reactivity section above.

"The Citrobacter assay may not react with several Citrobacter species; see Analytical Reactivity section above.

·Several of the Candida species detected at the high concentrations tested in this study may not be detected at lower concentrations; see the Analytical Reactivity section.

4C. guilliermondii is also classified as Candida fermentatis

eC. thermophila is also classified as Candida (Hansenula) parapolymorpha.

1Various Anaerococcus species are detected as Anaerococcus prevotii/vaginalis due to cross

reactivity. The efficiency of the cross-reactivity varies by species.

8 Clostridium cadaveris and Clostridium fallax are detected as Clostridium perfringens due to crossreactivity. Sequence analysis predicts a similar risk of cross-reactivity at high concentrations for C. baratii, C. disporicum and C. grantii.

45

bStaphylococcus argenteus and Staphylococcus schweitzeri are detected as Staphylococcus aureus (all three species are part of the S. aureus complex) due to cross-reactivity. mecA/C and MREJ (MRSA) was also detected in the S. argenteus isolate.

Bacteroides xylanisolvens is detected as Bacteroides fragilis due to cross-reactivity. The Citrobacter assay may not react with several Citrobacter species; see Analytical Reactivity section above.

Cosenzaea myxofaciens (formerly Proteus myxofaciens) is detected as Proteus spp. due to crossreactivity.

Enterobacter bugandensis (tested) and Enterobacter chengduensis (not tested, in silico prediction only) are detected as Enterobacter cloacae complex due to cross-reactivity.

mEscherichia albertii, Escherichia fergusonii and Shigella species are detected as Escherichia coli due to cross-reactivity.

"Haemophilus aegyptius (formerly described as H. influenzae biogroup aegyptius) is detected as Haemophilus influenzae due to cross-reactivity.

· Kingella negevensis is detected as Kingella kingae due to cross-reactivity.

PvanM is detected as vanA/B (not tested, in silico prediction only) due to cross-reactivity 9The CTX-M assay cross-reacts weakly with the blaoxy gene carried in an isolate of Klebsiella michiganensis (reported as N/A because and applicable bacterium is not detected by the panel). Based on sequence analysis, the CTX-M assay is predicted to cross-react weakly with the blaoxy gene, blaRABs gene (found primarily in Rahnella and Leminorella species), blaki.u genes (isolated primarily from Kluyvera species), and some variants of ampC (not observed when tested at high concentration in this study).

Interference Testing

Potentially interfering substances that could be present in synovial fluid specimens or that may be introduced during specimen collection and testing were evaluated for their effect on the BioFire JI Panel performance. Substances included endogenous substances that may be found in specimens at normal or elevated levels, various commensal or infectious microorganisms, medications, a variety of sample processing substances and substances used to clean. decontaminate, or disinfect work areas. The effect of interfering substances has only been evaluated for those listed in Table 51. Interference from substances that were not evaluated could lead to erroneous results.

For this study, contrived samples were prepared in synovial fluid (SF) matrix, with each sample containing multiple organisms (and AMR genes) at low levels (3× the limit of detection (LoD)). The subset of organisms included in the samples represent all organism types and AMR genes detected by the panel (aerobic and anaerobic gram-positive and gramnegative bacteria, including fastidious species. with AMR genes for methicillin-resistance and vancomycin resistance in the gram-positive bacteria and for extended-spectrum beta lactamase and carbapenemase activity in gram-negative bacteria, as well as Candida yeast species). Since the functions of the test that could be affected by interference from various substances would impact detection of the different types of organisms and AMR genes similarly, testing with a representative subset is effective for evaluating interference for the full panel. Testing was performed with analytes at concentrations near LoD in order to identify the effects of even low-level interference on analyte detection.

46

Each contrived sample was tested first as a 'no substance' or 'no interference' positive control followed by testing of the same sample after addition of a substance or microorganism. Substances were added to the contrived samples (or to negative SF, to test the impact of substance alone) at concentrations equal to or greater than the levels expected in clinical SF specimens, and microorganisms were added at the highest possible concentration to evaluate the 'worst case' scenario for interference. Control and test samples were tested in triplicate with three reagent lots.

JI Panel
Substance	Concentration Tested	Testing Outcome
	Endogenous Substances
Blood	30% v/v	No Interference
Cholesterol	4 mg/mL	No Interference
C-Reactive Protein	0.17 mg/mL	No Interference
Fibronectin	3 mg/mL	No Interference
Lactate	5.7 mg/mL	No Interference
Monosodium urate/Uric Acid	0.235 mg/mL	No Interference
Calcium Phosphate	16 mg/mL	No Interference
Calcium Oxalate	7.9 µg/mL	No Interference
Bilirubin	0.4 mg/mL	No Interference
White Blood Cells	3.0E+07 cells/mL	No Interference
Rheumatoid Factor	1,800 IU/mL	No Interference
Type II Collagen	10.1 µg/mL	No Interference
	Exogenous Substances
Acetaminophen	156 µg/mL	No Interference
Salicylic Acid	28.6 µg/mL	No Interference
Ibuprofen	219 µg/mL	No Interference
Capsaicin Cream (0.1% capsaicin)	0.5% (m/v)	No Interference
Salicylate Cream (30% methyl
salicylate)	0.5% (m/v)	No Interference
Camphor Balm (11% camphor)	0.5% (m/v)	No Interference
Arnica Gel (7% Arnica
montana)	1.0% v/v	No Interference
Nystatin	5000 Units/mL	No Interference
Fluconazole	25.5 µL	No Interference
Mupirocin	1.5 µg/mL	No Interference
Ceftriaxone	840 µg/mL	No Interference
Vancomycin	120 µg/mL	No Interference
Clindamycin	51 µg/mL	No Interference
Triple antibiotic ointment (10000 U
polymyxin B, 3.5 mg neomycin, 500
U bacitracin)	0.5% (m/v)	No Interference
Hydrocortisone	8.3 mg/mL	No Interference
Hyaluronic acid	16 mg/mL	No Interference
Lidocaine	23 mg/mL	No Interference
Cobalt Ions	20 µg/mL	No Interference
Chromium Ions	50 µg/mL	No Interference
Ultra-High Molecular Weight
Polyethylene	1 mg/mL	No Interference
Substance	Concentration Tested	Testing Outcome
Polymethyl methacrylate Bone
cement	1% m/v	No Interference
Iohexol	250 mg/mL	No Interference
	Competitive Microorganisms
Streptococcus pyogenes	7.56E+08 CFU/mL	No Interference
Eschericia coli	8.1E+08 CFU/mL	No Interference
Finegoldia magna	8.8E+07 CFU/mL	No Interference
Candida albicans	7.9E+07 CFU/mL	No Interference
Cutibacterium acnes	1.1E+07 cells/mL	No Interference
Staphylococcus epidermidis	8.8E+08 CFU/mL	No Interference
Cornebacterium striatum	7.8E+08 CFU/mL	No Interference
Cryprococcus neoformans	1.0E+07 CFU/mL	No Interference
Parvovirus B19	7.0E+04 IU/mL	No Interference
Chikungunya virus	2.2E+07 genomic
equivalents/mL	No Interference
	Disinfection/Cleaning Substances
Reagent Alcohol	1.0% v/v	No Interference
Povidone-iodine	1.0% v/v	No Interference
Bleach	1.0% v/v
(600 ppm chlorine)	No Interference
	Sample Processing Materials
K2-EDTA anticoagulant	0.99 ug/mL
No Interference

Table 51: Evaluation of Potentially Interfering Substances on the BioFire JI Panel

47

Notably, bleach is a strong oxidizer capable of damaging nucleic acids. When bleach has been evaluated for interference in other sample types associated with different BioFire FilmArray panels (e.g. nasopharyngeal swab in transport medium, cerebrospinal fluid (CSF), blood culture, stool in Cary Blair transport medium), the oxidizing effects of this disinfectant on results has varied based on the properties of the sample type, the concentration of bleach, and the amount of time the bleach was incubated with the sample. In some cases, such as with CSF, the damaging effect of bleach on the organisms and nucleic acids in the sample were apparent at low bleach concentration and short incubation times. In other cases, no impact on analyte detection was observed, even after 24-hour incubation or bleach concentration up to 5% (3000 ppm). In this study, bleach was tested at 1% (600 ppm) in SF following 15-minute and 24-hour incubation times and no effects on low-level analyte amplification and detection were observed. Nevertheless, to maintain sample integrity, caution should be taken to prevent the direct mixing of bleach with SF samples prior to testing.

4. Assay Reportable Range:

Not applicable.

1. Traceability, Stability, Expected Values (Controls, Calibrators, or Methods):

Process Controls

Two process controls are included in each pouch:

48

RNA Process Control: The RNA Process Control assay targets an RNA transcript from the yeast Schizosaccharomyces pombe. The yeast is present in the pouch in a freeze-dried form and becomes rehydrated when sample is loaded. The control material is carried through all stages of the test process, including lysis, nucleic acid purification, reverse transcription, PCR1, dilution. PCR2. and DNA melting. A positive RNA Process Control result indicates that all steps carried out in the BioFire JI Panel pouch were successful.

PCR2 Control: The PCR2 Control assay detects a DNA target that is dried into wells of the array along with the corresponding primers. A positive result indicates that 2nd stage PCR was successful.

Both control assays must be positive for the test run to pass. If either control fails, the Controls field of the test report will display "Failed" and all results will be listed as "Invalid". If the controls fail, the sample should be retested using a new pouch.

External Controls

External controls should be used in accordance with laboratory protocols and the appropriate accrediting organization requirements, as applicable. Previously characterized positive samples or negative samples spiked with well-characterized organisms can be used as external positive controls.

Specimen Stability

Diagnostic testing of SF specimens is intended to be performed as soon as possible after collection of the specimen, however, transport to the testing lab and subsequent storage of a specimen is sometimes required. A study was performed to confirm that accurate BioFire JI Panel test results can be obtained from SF samples stored refrigerated for up to seven days. Two contrived samples were prepared in a pooled SF matrix, with each sample containing a representative subset of organisms at a concentration near (3×) the established limit of detection (LoD) (as well as 'native' analytes present in the pooled SF matrix at an unknown concentration). The subset of spiked organisms included aerobic and anaerobic gram-positive and gram-negative bacteria (some harboring antimicrobial resistance (AMR) genes), as well as two species of Candida yeast.

Each contrived sample was tested in ten replicates immediately after preparation (D0, no storage) and the remaining sample volume was stored under standard refrigerated conditions for subsequent testing. Ten replicates were tested for each sample after one (D1), three (D3), five (D5) and seven (D7) days of refrigerated storage. Prolonged room temperature storage of SF prior to testing with the BioFire JI Panel will not be recommended and was not evaluated.

49

Detection of organisms and AMR genes was observed in 90.0 - 100% of the no storage control (D0) sample replicates tested and there was no trend toward inaccurate test results associated with sample storage over time.

		Analyte Detection
Analyte	Strain	No
Storage
(D0)	Day 1
(D1)	Day 3
(D3)	Day 5
(D5)	Day 7
(D7)
Gram Positive Bacteria
Clostridium
perfringens	ATCC 13124	9/10	9/10	9/10	9/10	9/10
Enterococcus
faecium	ATCC 700221	10/10	10/10	11/11b	10/10	9/10
vanA/B	ATCC 700221	10/10	10/10	11/11b	10/10	10/10
Staphylococcus
aureus	ATCC43300	10/10	10/10	10/10	10/10	10/10
mecA/C and MREJ	ATCC43300	10/10	10/10	10/10	10/10	10/10
Streptococcus spp.
Streptococcus
pneumoniae	ATCC 6303	10/10	10/10	11/11b	10/10	10/10
Gram Negative Bacteria
Bacteroides fragilis	ATCC 25285	10/10	10/10	9/11b	10/10	9/10
Escherichia coli
NDM	AR-BANK #0150	10/10	10/10	11/11b	10/10	10/10
Haemophilus
influenzae	ATCC 10211	10/10	10/10	11/11b	10/10	10/10
Kingella kingae	ATCC 23330	10/10	10/10	10/10	10/10	10/10
Klebsiella
pneumoniae
KPC	AR-BANK #0097	10/10	10/10	10/10	10/10	10/10
Neisseria
gonorrhoeae	ATCC 19424	10/10	10/10	10/10	10/10	10/10
Candida
Candida albicans	ATCC 90028	10/10	10/10	10/10	10/10	10/10
Candida
(Candida krusei)	ATCC 6258	10/10	10/10	10/11b	10/10	10/10

Table 52: BioFire Joint Panel Results for Stored Synovial Fluid Specimensa
----------------------------------------------------------------------------	--	--

ªAdditional analytes (E. faecalis, S. aureus, mecA/C and MREJ (MRSA) and S. lugdunensis) present in the pooled synovial fluid matrix were also detected.

6 An extra replicate of Sample 2 was tested due to a suspected pouch anomaly/error. Unexpected rosults were reported for 3 analytes (Bacteroides fragilis, NDM and Candida) in replicate 7 of Sample 2 stored for three days (D3).

Analysis of the Cp values across the evaluated analytes did not indicate a negative trend across the entire period of the stability study.

50

Fresh vs. Frozen Study

Table 53: Summary of Limit of Detection (LoD) for BioFire JI Panel Bacteria and Yeast

52

For resistance genes, data were collected to demonstrate detection of each AMR gene at a concentration corresponding to the lowest LoD of the applicable bacteria. LoD estimate dilution series data and/or LoD confirmation testing data for host organisms were used to demonstrate detection at the applicable bacterial LoD concentrations. Data from this testing establish that amplification and positive assay results for the AMR genes are observed at a host concentration that is the same as or similar to the lowest LoD of the applicable bacteria.

	Table 54: AMR Gene Detection at Lowest Applicable LoD
--	-------------------------------------------------------	--	--

| BioFire JI AMR
Target | Applicable
Organism | LoD Concentration |
|--------------------------|------------------------|-------------------|
| | | Molecular (DNA) |
| CTX-M | Salmonella enterica | 1.6E+03 copies/mL |

53

| BioFire JI AMR
Target | Applicable
Organism | LoD Concentration
Molecular (DNA) |
|--------------------------|---------------------------|--------------------------------------|
| IMP | Pseudomonas
aeruginosa | 1.3E+03 copies/mL |
| KPC | Klebsiella pneumoniae | 1.6E+03 copies/mL |
| mecA/C and MREJ | Staphylococcus aureus | 4.2E+03 copies/mL |
| NDM | Escherichia coli | 6.0E+03 copies/mL |
| NDM | Morganella morganii | 2.2E+03 copies/mL |
| NDM | Salmonella enterica | 1.6E+03 copies/mL |
| OXA-48-like | Klebsiella aerogenes | 7.5E+02 copies/mL |
| vanA/B | Enterococcus faecium | 1.2E+03 copies/mL |
| vanA/B | Enterococcus faecalis | 1.0E+03 copies/mL |
| VIM | ATCC 15794 | 2.6E+03 copies/mL |

7. Assay Cut-Off:

The BioFire Joint Infection Panel is part of BioFire Diagnostics' (BFDX) FilmArray system. The FilmArray system is designed to interpret the test data and automatically report the test results to the operator. The FilmArray system uses the results of the Melt Detector to determine each test result. The Melt Detector is part of the FilmArray Analysis Software and assigns a positive or negative result to each reaction on the array through analysis of the melt data collected during the test. These positive and negative results are combined in the FilmArray Analysis Software (using the replicate, assay and interpretation rules) to report the presence or absence of each pathogen in the panel.

The purpose of this study was to validate the use of this Melt Detector with current optimization parameters with the BioFire Bone and Joint Panel. To evaluate the Melt Detector performance, the observed sensitivity and specificity rates for the individual melt curves and assay calls are reported. These sensitivity and specificity rates are determined by comparing the FilmArray test results obtained from well-characterized samples, collected as part of the clinical evaluation and analytic testing of the Bone and Joint Panel, to expert annotation. Annotations (positive and negative calls) for all melt curves and assay calls were determined by the sponsor.

For individual melt curves, the observed sensitivity and specificity, as compared to expert annotation, of the Melt Detector is 99.59% and 99.98%. respectively. For the Analysis Software, the observed sensitivity and specificity, as compared to the expert annotation, of the assay calls are 99.49% for sensitivity and 99.97% specificity. The validation results met the predefined acceptance criteria of >95% accuracy as compared to expert annotation.

54

8. Carry-Over:

A formal carry-over study in support of this regulatory submission for the BioFire JI Panel was not performed, since carry-over studies with high positive samples followed by negative samples have been performed for other FDA-cleared FilmArray Panels (i.e. FilmArray RP, BCID, and GI Panels) for both the FilmArray 2.0 and the FilmArray Torch systems, and no carry-over has been observed.

B Comparison Studies:

1. Method Comparison:
  Not Applicable.
1. Matrix Comparison:
  Not applicable.

C Clinical Studies:

1. Clinical Sensitivity:
  The clinical performance of the BioFire Joint Infection Panel was established during a multicenter study conducted at thirteen geographically distinct study sites in the U.S. and in Europe over approximately two years from May 2018 to March 2020. A total of 1591 synovial fluid specimens were acquired for the prospective clinical study. A total of 47 synovial fluid specimens were excluded from the final data analysis. The most common reasons for specimen exclusion was the specimen was found to not meet the inclusion criteria after the specimen had been enrolled, a valid JI Panel test result was not obtained, or the study site was unable to complete the Case Report Form (CRF). The final data set consisted of 1544 specimens, of which 771 (49.9%) were frozen before testing. No difference in performance between fresh and frozen specimens was observed when results were compared, therefore the data from both specimen types have been combined for all analyses.

55

		Overall	Site 1	Site 2	Site 3	Site 4
Sex	Male	878
(56.9%)	184
(52.3%)	46
(52.3%)	108
(53.7%)	65
(73.0%)	60
(58.8%)	45
(51.7%)	108
(54.8%)	39
(59.1%)	88
(60.3%)	88
(64.7%)	10
(58.8%)	27
(55.1%)	10
(71.4%)
	Female	666
(43.1%)	168
(47.7%)	42
(47.7%)	93
(46.3%)	24
(27.0%)	42
(41.2%)	42
(48.3%)	89
(45.2%)	27
(40.9%)	58
(39.7%)	48
(35.3%)	7
(41.2%)	22
(44.9%)	4
(28.6%)
Age	$Gram Positive Bacteria
Anaerococcus prevotii/vaginalis	1/1	100	-	1543/1543	100	99.8-100%
Clostridium perfringens	0/0	-	-	1544/1544	100	99.8-100%
Cutibacterium avidum/granulosum	0/0	-	-	1544/1544	100	99.8-100%
Enterococcus faecalisa	10/10	100	72.2-100%	1529/1534	99.7	99.2-99.9%
Enterococcus faeciumb	1/1	100	-	1541/1543	99.9	99.5-100%
Finegoldia magnac	3/3	100	43.9-100%	1540/1541	99.9	99.6-100%
Parvimonas micrad	0/1	0	-	1543/1543	100	99.8-100%
Peptoniphiluse	1/1	100	-	1542/1543	99.9	99.6-100%
Peptostreptococcus anaerobiusf	0/0	-	-	1541/1544	99.8	99.4-99.9%
Staphylococcus aureusg	98/105	93.3	86.9-96.7%	1417/1439	98.5	97.7-99.0%
Staphylococcus lugdunensish	2/2	100	34.2-100%	1539/1542	99.8	99.4-99.9%
Streptococcus spp.i	38/44	86.4	73.3-93.6%	1488/1500	99.2	98.6-99.5%
Streptococcus agalactiaej	10/11	90.9	62.3-98.4%	1532/1533	99.9	99.6-100%
Streptococcus pneumoniae	3/3	100	43.9-100%	1541/1541	100	99.8-100%
Streptococcus pyogenesk	11/12	91.7	64.6-98.5%	1532/1532	100	99.7-100%
Gram Negative Bacteria
Bacteroides fragilisl	0/0	-	-	1543/1544	99.9	99.6-100%
Citrobacter	2/2	100	34.2-100%	1542/1542	100	99.8-100%
Enterobacter cloacae complexm	2/4	50	15-85.0%	1538/1540	99.9	99.5-100%
Escherichia colin	14/14	100	78.5-100%	1529/1530	99.9	99.6-100%
Haemophilus influenzaeo	1/1	100	-	1542/1543	99.9	99.6-100%
Kingella kingaep	1/1	100	-	1537/1543	99.6	99.2-99.8%
Klebsiella aerogenes	0/0	-	-	1544/1544	100	99.8-100%
Klebsiella pneumoniae groupq	4/5	80.0	37.6-96.4%	1538/1539	99.9	99.6-100%
Morganella morganiir	1/1	100	-	1541/1543	99.9	99.5-100%
Neisseria gonorrhoeaes	2/2	100	34.2-100%	1539/1542	99.8	99.4-99.9%

Table 56: BioFire Joint Infection Panel Prospective Clinical Performance Summary

57

Analyte	Sensitivity/PPA			Specificity/NPA
	TP/
(TP + FN)	%	95%CI	TN/
(TN + FP)	%	95%CI
Proteus spp.t	4/4	100	51.0-100%	1536/1540	99.7	99.3-99.9%
Pseudomonas aeruginosau	2/2	100	34.2-100%	1539/1542	99.8	99.4-99.9%
Salmonella spp.	0/0	-	-	1544/1544	100	99.8-100%
Serratia marcescensv	2/2	100	34.2-100%	1541/1542	99.9	99.6-100%
Yeast
Candidaw	4/7	57.1	25.0-84.2%	1536/1537	99.9	99.6-100%
Candida albicansx	3/5	60.0	23.1-88.2%	1539/1539	100	99.8-100%

a E. faecalis was detected in all five FP specimens using an additional comparator method

b E. faecium was detected in both FP specimens using an additional molecular method

& F. magna was detected in the single FP specimen using an additional molecular method

d P. micra was detected in the single FN specimen using an additional molecular method

e P. was detected in the single FP specimen using an additional molecular method

t P. anaerobius was detected in all three FP specimens using an additional molecular method

8 S. aureus was detected in 57 FN specimens using an additional molecular testing of one of the remaining two FN specimens and its isolate identified it as S. argenteus. S. aureus was detected in 19/22 FP specimens using an additional molecular method.

h S. lugdunensis was detected in all three FP specimens using an additional molecular method

1 Streptococcus spp. was detected in 4/7 FN specimens and in all 12 FP specimens using an additional method

I S. agalactiae was detected in the single FN specimen using an additional molecular method

k The single FN specimen was negative for S. pygogenes when tested by additional molecular methods

1 B. fragilis was detected in the single FP specimen using an additional molecular method

m E. cloacae complex was detected in 1/2 FN specimens using an additional molecular method

n E. coli was detected in the single FP specimen using an additional molecular method

0H. influenzae was detected in the single FP specimen using an additional molecular method

P K. kingae was detected in all six FP specimens using an additional molecular methods

9 K. pneumoniae group was detected in the single FN specimen using an additional molecular method

T M. morganii was detected in both FP specimens using an additional molecular method

8 N. gonorrhoeae was detected in all three FP specimens using an additional molecular method

1 Proteus spp. was detected in all four FP specimens using an additional molecular method

u P. aeruginosa was detected in all three FP specimens using an additional molecular method

" S. marcescens was detected in the single FP specimen using an additional molecular method

w Candida was detected in 2/3 FN specimens and in the single FP specimen using an additional mothod

Candida albicans was detected in 1/2 FN specimens using an additional molecular method

BioFire JI Panel Genus and Group level organism assay performance is stratified by species for Anaerococcus prevotii/vaginalis, Peptoniphilus, Streptococcus spp., Citrobacter, Enterobacter cloacae complex, Klebsiella pneumoniae group, Proteus spp., and Candida in Table 50. Note: multiple organisms from a group may be detected in a single specimen, therefore the "Total" values in these tables may not match the performance values presented above, which are reported per specimen.

Table 57. Sensitivity of the BioFire JI Panel Species Inclusive Assays Stratified by Species

Species	BioFire JI Panel Sensitivity
Anaerococcus prevotii/vaginalis
A. vaginalis	1/1 (100%)
Peptoniphilus spp.
P. asaccharolyticus	1/1 (100%)
Streptococcus spp.
S. agalactiae	10/11 (90.9%)
S. anginosus	1/1 (100%)
S. anginosus group	1/1 (100%)
S. constellatus	1/1 (100%)
S. dysgalactiae	7/8 (85.7%)
S. gallolyticus	0/1 (0%)
S. gordonii	2/2 (100%)
S. mitis	1/1 (100%)
S. oralis	1/1 (100%)
S. pneumoniae + S. pyogenes	1/1 (100%)
S. pneumoniae	2/2 (100%)
S. pyogenes	10/11 (90.9%)
S. salivarius/vestibularis group	0/1 (0%)
Viridans streptococci	1/2 (50%)
Total Streptococcus species	38/44 (86.4%)
Citrobacter spp.
C. freundii	1/1 (100%)
C. koseri	1/1 (100%)
Total Citrobacter species	2/2 (100%)
Enterobacter coacae complex
E. cloacae	1/2 (50%)
E. cloacae complex	1/2 (50%)
Total E. cloacae species	2/4 (50%)
Klebsiella pneumoniae group
K. pneumoniae	4/5 (80%)
Proteus spp.
P. mirabilis	4/4 (100%)
Candida spp.
C. albicans	3/5 (60.0%)
C. parapsilosis	1/2 (50%)
Total Candida species	4/7 (57.1%)

58

Antimicrobial Resistance Genes

AMR gene results are reported only when one or more applicable bacteria that may carry the gene are also detected in the sample. If no applicable bacteria are detected, the AMR gene results are reported as Not Applicable (N/A). The results are summarized for each AMR gene in Table 51.

59

| Analyte | TP/
(TP + FN) | % | 95%CI | TN/
(TN + FP) | % | 95%CI |
|------------------------|------------------|-----|-----------|------------------|------|------------|
| CTX-M | 5/5 | 100 | 56.6-100% | 33/33 | 100 | 89.6-100% |
| IMP | 0/0 | - | - | 38/38 | 100 | 90.8-100% |
| KPC | 0/0 | - | - | 40/40 | 100 | 91.2-100% |
| mecA/C and MREJ (MRSA) | 19/19 | 100 | 83.2-100% | 90/94 | 95.7 | 89.6-98.3% |
| NDM | 0/0 | - | - | 40/40 | 100 | 91.2-100% |
| OXA-48-like | 1/1 | 100 | - | 33/33 | 100 | 89.6-100% |
| vanA/B | 3/3 | 100 | 43.9-100% | 14/14 | 100 | 78.5-100% |
| VIM | 0/0 | - | - | 38/38 | 100 | 90.8-100% |

Table 58: BioFire Joint Infection Panel Prospective Clinical Performance Summary - AMR Genes

" Three out of the four FP specimens evidence of mecA/C AMR gene was observed in the isolates using independent molecular method with sequencing. Additionally, SOC testing confirmed an AST phenotype of methicillin resistant for all three isolates. The mecA/c and MREJ AMR genes were found in the remaining independent molecular methods.

AMR gene results are reported only when one or more applicable bacteria that may carry the gene are also detected in the sample. If no applicable bacteria are detected, the AMR gene results are reported as Not Applicable (N/A). The results are summarized for each AMR gene in Table 59 through Table 74. Note: the 'Performance Summary' tables below do not include specimens for which an applicable bacteria was not reported (i.e. the AMR gene was reported as N/A); these specimens are instead accounted for in the 'Distribution of Clinical Specimens' tables below.

| S. aureus

mecA/C and MREJ		SoC: S. aureus PCR/seq: mecA/C
JI Panel
Result		Org+/ Res+	Org+/ Res-	Org -	Total
	Org+/ Res+	15	3	5	23
	Org+/ Res-	0	73	17	90
	Org -	0	7	1393	1400
	Total	15	83	1415	1513a
Performance		Agreement	%	95%CI
Org+ / Res+		15/15	100%	79.6-100%
Org+/ Res-		73/83	88.0%	79.2-93.3%
Org -		1393/1415	98.4%	97.7-99.0%%
Interpretation		PPA	NPA	Prevalence
MRSA		15/15
(100%)	1490/1498
(99.5%)	23/1513
(1.5%%)
MSSA		73/83
(88.0%)	1413/1430
(98.8%)	90/1513
(5.9%)
S. aureus		91/98
(92.9%)	1393/1415
(98.4%)	113/1513
(7.5%)

Table 59: Distribution of mecA/C and MREJ in Clinical Specimens

aThirty-one (31) specimens excluded from molecular analysis for mecA/C and MREJ (MRSA) due to volume constraints either initially or following a failure during comparator testing

Table 60. Stratification of mecA/C and MREJ by Applicable Host Organism

60

| Applicable Bacteria Result
(JI Panel) | N | Positive Percent
Agreement | | Negative Percent
Agreement | |
|------------------------------------------|----|-------------------------------|---------------|-------------------------------|----------------|
| | | % | 95% CI | % | 95% CI |
| Staphylococcus aureus | 98 | 100%
(19/19) | 83.2-
100% | 95.7%
(90/94) | 89.6-
98.3% |

		Table 61. Distribution of VIM in Clinical Specimens
--	--	-----------------------------------------------------	--	--	--

VIM		SoC: Applicable Bacteria PCR/seq: VIM	Org+ / Res+	Org+ / Res-	Org -
JI Panel
Result	Org+/ Res+	0	0	0	0
	Org+/ Res-	0	28a	10	38
	Org -	0	3	1472	1475
Total		0	31	1482	1513b
Performance		Agreement	%	95%CI
Org+/Res+		0/0	-	-
Org+/Res-		28/31	90.3%	75.1-96.7%
Org -		1472/1482	99.3%	98.8-99.6%

4Two specimens had co-detection of Escherichia coli with Proteus spp.; one specimen had co-detection of Klebsiella pneumoniae group with Morganella morganii; one specimen had co-detection of Klebsiella pneumoniae group with Morganella morganii and Proteus spp.

bThirty-one (31) specimens were excluded from molecular analysis for VIM due to volume constraints either initially or following a failure during comparator testing

Table 62. Stratification of VIM Clinical Performance by Applicable Host Organism

61

| Applicable Bacteria Result
(JI Panel) | Positive Percent
Agreement | | Negative Percent
Agreement | |
|--------------------------------------------|-------------------------------|--------|-------------------------------|---------------|
| | % | 95% CI | % | 95% CI |
| Overall (any applicable bacteria Detected) | (0/0) | - | 100%
(38/38a) | 90.8-
100% |
| Citrobacter | (0/0) | - | 100%
(2/2) | 34.2-
100% |
| Enterobacter cloacae
complex | (0/0) | - | 100%
(4/4) | 51.0-
100% |
| Escherichia coli | (0/0) | - | 100%
(15/15) | 79.6-
100% |
| Klebsiella aerogenes | (0/0) | - | (0/0) | - |
| Klebsiella pneumoniae
group | (0/0) | - | 100%
(5/5) | 56.6-
100% |
| Morganella morganii | (0/0) | - | 100%
(3/3) | 43.9-
100% |
| Proteus spp. | (0/0) | - | 100%
(8/8) | 67.6-
100% |
| Pseudomonas aeruginosa | (0/0) | - | 100%
(4/4) | 51.0-
100% |
| Salmonella spp. | (0/0) | - | (0/0) | - |
| Serratia marcescens | (0/0) | - | 100%
(2/2) | 34.2-
100% |

aTwo specimens had co-detection of Escherichia coli with Proteus spp .; one specimen had co-detection of Klebsiella pneumoniae group with Morganella morganii; one specimen had co-detection of Klebsiella pneumoniae group with Morganella morganii and Proteus spp.

Table 63. Distribution of CTX-M in Clinical Specimens

CTX-M		Org+ / Res+	Org+ / Res-	Org -	Total
JI Panel
Result	Org+ / Res+	5	0	0	5
	Org+ / Res-	0	23a	10	33
	Org -	0	3	1472	1475
Total		5	26	1482	1513b
Performance		Agreement	%	95%CI
Org+ / Res+		5/5	100%	56.6-100%
Org+ / Res-		23/26	88.5%	71.0-96.0%
Org -		1472/1482	99.3%	98.8-99.6%

aTwo specimens had co-detection of Escherichia coli with Proteus spp ; one specimen had co-detection of Klebsiella pneumoniae group with Morganella morganii; one specimen had co-detection of Klebsiella pneumoniae group with Morganella morganii and Proteus spp.

bThirty-one (31) specimens excluded from tMol analysis for CTX-M due to volume constraints either initially or following a failure during comparator testing

Table 64. Stratification of CTX-M Clinical Performance by Applicable Host Organism

62

| Applicable Bacteria Result
(JI Panel) | Positive Percent
Agreement | | Negative Percent
Agreement | |
|--------------------------------------------|-------------------------------|---------------|-------------------------------|---------------|
| | % | 95% CI | % | 95% CI |
| Overall (any applicable bacteria Detected) | 100
(5/5) | 56.6-
100% | 100%
(33/33a) | 89.6-
100% |
| Citrobacter | (0/0) | - | 100%
(2/2) | 34.2-
100% |
| Enterobacter cloacae
complex | (0/0) | - | 100%
(4/4) | 51.0-
100% |
| Escherichia coli | 100
(2/2) | 34.2-
100% | 100%
(13/13) | 77.2-
100% |
| Klebsiella aerogenes | (0/0) | - | (0/0) | - |
| Klebsiella pneumoniae
group | 100
(3/3) | 43.9-
100% | 100%
(2/2) | 34.2-
100% |
| Morganella morganii | (0/0) | - | 100%
(3/3) | 43.9-
100% |
| Proteus spp. | (0/0) | - | 100%
(8/8) | 67.6-
100% |
| Pseudomonas aeruginosa | (0/0) | - | 100%
(4/4) | 51.0-
100% |
| Salmonella spp. | (0/0) | - | (0/0) | - |
| Serratia marcescens | (0/0) | - | 100%
(2/2) | 34.2-
100% |

"Two specimens had co-detection of Escherichia coli with Proteus spp .; one specimen had codetection of Klebsiella pneumoniae group with Morganella morganii; one specimen had codetection of Klebsiella pneumoniae group with Morganella morganii and Proteus spp.

Table 65. Distribution of IMP in Clinical Specimens

IMP		SoC: Applicable Bacteria PCR/seq: IMP
		Org+/ Res+	Org+/ Res-	Org -	Total
JI Panel
Result	Org+/ Res+	0	0	0	0
	Org+/ Res-	0	28a	10	38
	Org -	0	3	1472	1475
Total		0	31	1482	1513b
Performance		Agreement	%	95%CI
Org+/ Res+		0/0	-	-
Org+/ Res-		28/31	90.3%	75.1-96.7%
Org -		1472/1482	99.3%	98.8-99.6%

4Two specimens had co-detection of Escherichia coli with Proteus spp ; one specimen had co-detection of Klebsiella pneumoniae group with Morganella morganii; one specimen had co-detection of Klebsiella pneumoniae group with Morganella morganii and Proteus spp.

bThirty-one (31) specimens excluded from molecular analysis for IMP due to volume constraints either initially or following a failure during comparator testing

Table 66. Stratification of IMP Clinical Performance by Applicable Host Organism

63

| Applicable Bacteria Result
(JI Panel) | Positive Percent
Agreement | 95% CI | Negative Percent
Agreement | 95% CI |
|--------------------------------------------|-------------------------------|--------|-------------------------------|---------------|
| Overall (any applicable bacteria Detected) | (0/0) | - | 100%
(38/38a) | 90.8-
100% |
| Citrobacter | (0/0) | - | 100%
(2/2) | 34.2-
100% |
| Enterobacter cloacae
complex | (0/0) | - | 100%
(4/4) | 51.0-
100% |
| Escherichia coli | (0/0) | - | 100%
(15/15) | 79.6-
100% |
| Klebsiella aerogenes | (0/0) | - | (0/0) | - |
| Klebsiella pneumoniae
group | (0/0) | - | 100%
(5/5) | 56.6-
100% |
| Morganella morganii | (0/0) | - | 100%
(3/3) | 43.9-
100% |
| Proteus spp. | (0/0) | - | 100%
(8/8) | 67.6-
100% |
| Pseudomonas aeruginosa | (0/0) | - | 100%
(4/4) | 51.0-
100% |
| Salmonella spp. | (0/0) | - | (0/0) | - |
| Serratia marcescens | (0/0) | - | 100%
(2/2) | 34.2-
100% |

*Two specimens had co-detection of Escherichia coli with Proteus spp.; one specimen had co-detection of Klebsiella pneumoniae group with Morganella morganii; one specimen had co-detection of Klebsiella pneumoniae group with Morganella morganii and Proteus spp.

	OXA-48-like	SoC: Applicable Bacteria PCR/seq: OXA-48-like
		Org+ / Res+	Org+ / Res-	Org -	Total
JI Panel
Result	Org+ / Res+	1	0	0	1
	Org+ / Res-	0	25a	8	33
	Org -	0	3	1476	1479
	Total	1	28	1484	1513b
Performance		Agreement	%	95%CI
Org+ / Res+		1/1	100%	-
Org+ / Res-		25/28	89.3%	72.8-96.3%
Org -		1476/1484	99.5%	98.9-99.7%

Table 67. Distribution of OXA-48-like in Clinical Specimens

4Two specimens had co-detection of Escherichia coli with Proteus spp ; one specimen had co-detection of Klebsiella pneumoniae group with Morganii; one specimen had co-detection of Klebsiella pneumoniae group with Morganella morganii and Proteus spp.

bThirty-one (31) specimens excluded from molecular analysis for OXA-48-like due to volume constraints either initially or following a failure during comparator testing.

Table 68. Stratification of OXA-48-like Clinical Performance by Applicable Host Organism

64

| Applicable Bacteria Result
(JI Panel) | Positive Percent
Agreement | | Negative Percent
Agreement | |
|--------------------------------------------|-------------------------------|--------|-------------------------------|---------------|
| | % | 95% CI | % | 95% CI |
| Overall (any applicable bacteria Detected) | 100
(1/1) | - | 100%
(33/33a) | 89.6-
100% |
| Citrobacter | -
(0/0) | - | 100%
(2/2) | 34.2-
100% |
| Enterobacter cloacae
complex | -
(0/0) | - | 100%
(4/4) | 51.0-
100% |
| Escherichia coli | -
(0/0) | - | 100%
(15/15) | 79.6-
100% |
| Klebsiella aerogenes | -
(0/0) | - | -
(0/0) | - |
| Klebsiella pneumoniae
group | 100
(1/1) | - | 100%
(4/4) | 51.0-
100% |
| Morganella morganii | -
(0/0) | - | 100%
(3/3) | 43.9-
100% |
| Proteus spp. | -
(0/0) | - | 100%
(8/8) | 67.6-
100% |
| Salmonella spp. | -
(0/0) | - | -
(0/0) | - |
| Serratia marcescens | -
(0/0) | - | 100%
(2/2) | 34.2-
100% |

ªTwo specimens had co-detection of Escherichia coli with Proteus spp ; one specimen had co-detection of Klebsiella pneumoniae group with Morganella morganii; one specimen had co-detection of Klebsiella pneumoniae group with Morganella morganii and Proteus spp.

		SoC: Applicable Bacteria PCR/seq: vanA/B
	vanA/B	Org+ / Res+	Org+ / Res-	Org -	Total
JI Panel
Result	Org+/ Res+	1	0	2	3
	Org+/ Res-	0	9	5	14
	Org -	0	0	1496	1496
Total		1	9	1503	1513a
Performance		Agreement	%	95%CI
Org+/ Res+		1/1	100%	-
Org+ / Res-		9/9	100%	70.1-100%
Org -		1496/1503	99.5%	99.0-99.8%

Table 69. Distribution of vanA/B in Clinical Specimens

"Thirty-one specimens excluded from molecular analysis for vanA/B due to volume constraints either initially or following a failure during comparator testing

Table 70. Stratification of vanA/B Clinical Performance by Applicable Host Organism

| Applicable Bacteria Result
(JI Panel) | Positive Percent
Agreement | | Negative Percent
Agreement | |
|--------------------------------------------|-------------------------------|----------|-------------------------------|---------------|
| | % | 95% CI | % | 95% CI |
| Overall (any applicable bacteria Detected) | 100
(3/3) | 43.9-100 | 100%
(14/14) | 78.5-
100% |
| Enterococcus faecalis | -
(0/0) | - | 100%
(14/14) | 78.5-
100% |
| Enterococcus faecium | 100 | 43.9-100 | - | - |

65

(3/3)	(0/0)
-------	-------

NDM		SoC: Applicable Bacteria PCR/seq: NDM
		Org+ / Res+	Org+ / Res-	Org -	Total
JI Panel
Result	Org+ / Res+	0	0	0	0
	Org+ / Res-	0	29a	11	40
	Org -	0	3	1488	1491
Total		0	32	1499	1531b
Performance		Agreement	%	95%CI
Org+ / Res+		0/0	-	-
Org+ / Res-		29/32	90.6%	75.8-96.8%
Org -		1488/1499	99.3%	98.7-99.6%

Table 71. Distribution of NDM in Clinical Specimens

aTwo specimens had co-detection of Escherichia coli with Proteus spp ; one specimen had co-detection of Klebsiella pneumoniae group with Morganii; one specimen had co-detection of Klebsiella pneumoniae group with Morganella morganii and Proteus spp.

bThirteen (13) specimens were excluded from molecular analysis for NDM due to volume constraints during comparator testing

Table 72. Stratification of NDM Clinical Performance by Applicable Host Organism

| Applicable Bacteria Result
(JI Panel) | | Positive Percent
Agreement | | Negative Percent
Agreement | |
|--------------------------------------------|-------|-------------------------------|------------------|-------------------------------|--|
| | % | 95% CI | % | 95% CI | |
| Overall (any applicable bacteria Detected) | (0/0) | - | 100%
(40/40)a | 91.2-
100% | |
| Citrobacter | (0/0) | - | 100%
(2/2) | 34.2-
100% | |
| Enterobacter cloacae
complex | (0/0) | - | 100%
(4/4) | 51.0-
100% | |
| Escherichia coli | (0/0) | - | 100%
(15/15) | 79.6-
100% | |
| Klebsiella aerogenes | (0/0) | - | (0/0) | - | |
| Klebsiella pneumoniae
group | (0/0) | - | 100%
(5/5) | 56.6-
100% | |
| Morganella morganii | (0/0) | - | 100%
(3/3) | 43.9-
100% | |
| Proteus spp. | (0/0) | - | 100%
(8/8) | 67.6-
100% | |
| Pseudomonas aeruginosa | (0/0) | - | 100%
(5/5) | 56.6-
100% | |
| Salmonella spp. | (0/0) | - | (0/0) | - | |
| Serratia marcescens | (0/0) | - | 100%
(3/3) | 43.9-
100% | |

aTwo specimens had co-detection of Escherichia coli with Proteus spp ; one specimen had codetection of Klebsiella pneumoniae group with Morganella morganii; one specimen had co-detection of Klebsiella pneumoniae group with Morganella morganii and Proteus spp.

66

			SoC: Applicable Bacteria PCR/seq: KPC
KPC		Org+ / Res+	Org+ / Res-	Org -	Total
	Org+ / Res+	0	0	0	0
JI Panel	Org+ / Res-	0	29a	11	40
Result	Org -		2	1488	1491
	Total		31	1499	15316
	Performance		Agreement	0/0	95%CI
	Org+ / Res+		0/1	0%	Car
	Org+ / Res-		29/31	93.5%	79.3-98.2%
	Org -		1488/1499	99.3%	98.7-99.6%

Table 72 Dictribution of VDC in Clinical Specie

"Two specimens had co-detection of Escherichia coli with Proteus sp .; one specimen had co-detection of Klebsiella pneumoniae group with Morganii; one specimen had co-detection of Klebsiella pneumoniae group with Morganella morganii and Proteus spp.

bThirteen (13) specimens were excluded from molecular analysis for KPC due to volume constraints during comparator testing

Table 74. Stratification of KPC Clinical Performance by Applicable Host Organism

| Applicable Bacteria Result
(JI Panel) | Positive Percent
Agreement | | Negative Percent
Agreement | |
|--------------------------------------------|-------------------------------|--------|-------------------------------|---------------|
| | % | 95% CI | % | 95% CI |
| Overall (any applicable bacteria Detected) | (0/0) | - | 100%
(40/40)a | 91.2-
100% |
| Citrobacter | (0/0) | - | 100%
(2/2) | 34.2-
100% |
| Enterobacter cloacae
complex | (0/0) | - | 100%
(4/4) | 51.0-
100% |
| Escherichia coli | (0/0) | - | 100%
(15/15) | 79.6-
100% |
| Klebsiella aerogenes | (0/0) | - | (0/0) | - |
| Klebsiella pneumoniae
group | (0/0) | - | 100%
(5/5) | 56.6-
100% |
| Morganella morganii | (0/0) | - | 100%
(3/3) | 43.9-
100% |
| Proteus spp. | (0/0) | - | 100%
(8/8) | 67.6-
100% |
| Pseudomonas aeruginosa | (0/0) | - | 100%
(5/5) | 56.6-
100% |
| Salmonella spp. | (0/0) | - | (0/0) | - |
| Serratia marcescens | (0/0) | - | 100%
(3/3) | 43.9-
100% |

The BioFire JI Panel AMR gene reporting in the specimen was also compared to phenotypic antimicrobial susceptibility testing (AST) methods performed on organism isolates recovered from those specimens. The results presented in Table 75 through Table 78 are only for specimens with concordant (true positive) results, and are further stratified by each applicable host organism recovered from that specimen. Note that antimicrobial resistance, particularly extended-spectrum ß-lactamase (ESBL) activity and carbapenem resistance, may be due to mechanisms other than the presence of the AMR genes detected by the BioFire JI

67

Panel; conversely, detection of these genes may not always indicate an antimicrobial resistance phenotype. Additionally, discordant results between mecA/C and MREJ (MRSA) detection in a SF specimen by the BioFire JI Panel and the observed methicillin (oxacillin/cefoxitin) resistance of cultured Staphylococcus aureus isolates may be due to polymicrobial Staphylococcus aureus cultures containing a mixture of resistant and sensitive organisms.

Table 75. CTX-M Performance (compared to phenotypic AST methods for ESBL activity on cultured isolate(s) from SF specimens)

| Organism Identified by SOC and
Detected by BioFire JI Panel | N
ESBL | N
Non-ESBL | Positive Percent
Agreement
% | Positive Percent
Agreement
95% CI | Negative Percent
Agreement
% | Negative Percent
Agreement
95% CI |
|----------------------------------------------------------------|-----------|---------------|------------------------------------|-----------------------------------------|------------------------------------|-----------------------------------------|
| Overall
(any
applicable bacteria Detected) | 7 | 24 | 71.4% (5/7) | 35.9-91.8% | 100% (24/24) | 86.2-100% |
| Citrobacter | 0 | 2 | (0/0) | - | 100%
(2/2) | 34.2-100% |
| Enterobacter cloacae
complex | 0 | 2 | (0/0) | - | 100%
(2/2) | 34.2-100% |
| Escherichia coli | 2 | 12 | 100% (2/2) | 34.2-100% | 100
(12/12) | 75.8-100% |
| Klebsiella aerogenes | 0 | 0 | (0/0) | - | (0/0) | - |
| Klebsiella pneumoniae
group | 4 | 0 | 75.0% (3/4) | 30.1-95.4% | (0/0) | - |
| Morganella morganii | 0 | 1 | (0/0) | - | 100%
(1/1) | - |
| Proteus spp. | 1 | 3 | 0
(0/1) | - | 100%
(3/3) | 43.9-100% |
| Pseudomonas aeruginosa | 0 | 2 | (0/0) | - | 100%
(2/2) | 34.2-100% |
| Salmonella spp. | 0 | 0 | (0/0) | - | (0/0) | - |
| Serratia marcescens | 0 | 2 | (0/0) | - | 100%
(2/2) | 34.2-100% |

Table 76. Carbapenem Resistance Genes Performance (as compared to phenotypic AST methods for carbapenem resistance on cultured isolate(s) from SF specimens).

68

| Organism Identified
by SOC and
Detected by JI
Panel | N | | IMP | | KPC | | NDM | | OXA-48-like | | VIM | | Overall
(any
resistance
gene) | |
|--------------------------------------------------------------|---|----|-------------|-----------------|-------------|-----------------|-------------|-----------------|---------------|-----------------|-------------|-----------------|----------------------------------------|-----------------|
| | R | S | PPA | NPA | PPA | NPA | PPA | NPA | PPA | NPA | PPA | NPA | PPA | NPA |
| Overall
(any
applicable bacteria
Detected) | 1 | 30 | 0%
(0/1) | 100%
(30/30) | 0%
(0/1) | 100%
(30/30) | 0%
(0/1) | 100%
(30/30) | 100%
(1/1) | 100%
(28/28) | 0%
(0/1) | 100%
(30/30) | 100%
(1/1) | 100%
(30/30) |
| Citrobacter | 0 | 2 | -
(0/0) | 100%
(2/2) | -
(0/0) | 100%
(2/2) | -
(0/0) | 100%
(2/2) | -
(0/0) | 100%
(2/2) | -
(0/0) | 100%
(2/2) | -
(0/0) | 100%
(2/2) |
| Enterobacter cloacae
complex | 0 | 2 | -
(0/0) | 100%
(2/2) | -
(0/0) | 100%
(2/2) | -
(0/0) | 100%
(2/2) | -
(0/0) | 100%
(2/2) | -
(0/0) | 100%
(2/2) | -
(0/0) | 100%
(2/2) |
| Escherichia coli | 0 | 14 | -
(0/0) | 100%
(14/14) | -
(0/0) | 100%
(14/14) | -
(0/0) | 100%
(14/14) | -
(0/0) | 100%
(14/14) | -
(0/0) | 100%
(14/14) | -
(0/0) | 100%
(14/14) |
| Klebsiella aerogenes | 0 | 0 | -
(0/0) | -
(0/0) | -
(0/0) | -
(0/0) | -
(0/0) | -
(0/0) | -
(0/0) | -
(0/0) | -
(0/0) | -
(0/0) | -
(0/0) | -
(0/0) |
| Klebsiella
pneumoniae
group | 1 | 3 | 0%
(0/1) | 100%
(3/3) | 0%
(0/1) | 100%
(3/3) | 0%
(0/1) | 100%
(3/3) | 100%
(1/1) | 100%
(3/3) | 0%
(0/1) | 100%
(3/3) | 100%
(1/1) | 100%
(3/3) |
| Morganella morganii | 0 | 1 | -
(0/0) | 100%
(1/1) | -
(0/0) | 100%
(1/1) | -
(0/0) | 100%
(1/1) | -
(0/0) | 100%
(1/1) | -
(0/0) | 100%
(1/1) | -
(0/0) | 100%
(1/1) |
| Proteus spp. | 0 | 4 | -
(0/0) | 100%
(4/4) | -
(0/0) | 100%
(4/4) | -
(0/0) | 100%
(4/4) | -
(0/0) | 100%
(4/4) | -
(0/0) | 100%
(4/4) | -
(0/0) | 100%
(4/4) |
| Pseudomonas
aeruginosa | 0 | 2 | -
(0/0) | 100%
(2/2) | -
(0/0) | 100%
(2/2) | -
(0/0) | 100%
(2/2) | N/A | N/A | -
(0/0) | 100%
(2/2) | -
(0/0) | 100%
(2/2) |
| Salmonella spp. | 0 | 0 | -
(0/0) | -
(0/0) | -
(0/0) | -
(0/0) | -
(0/0) | -
(0/0) | -
(0/0) | -
(0/0) | -
(0/0) | -
(0/0) | -
(0/0) | -
(0/0) |
| Serratia marcescens | 0 | 2 | -
(0/0) | 100%
(2/2) | -
(0/0) | 100%
(2/2) | -
(0/0) | 100%
(2/2) | -
(0/0) | 100%
(2/2) | -
(0/0) | 100%
(2/2) | -
(0/0) | 100%
(2/2) |

Table 77. mecA/C and MREJ (MRSA) Performance (compared to phenotypic AST methods for methicillin (oxacillin/cefotoxitin) resistance on cultured isolates from SF specimens.

| Organism Identified by SOC and
Detected by BioFire JI Panel | N | | Positive Percent
Agreement | | Negative Percent
Agreement | |
|----------------------------------------------------------------|----|----|-------------------------------|------------|-------------------------------|-----------|
| | R | S | % | 95% CI | % | 95% CI |
| Staphylococcus aureus | 22 | 76 | 81.8
(18/22) | 61.5-92.7% | 100%
(76/76) | 95.2-100% |

Table 78. vanA/B Performance (as compared to phenotypic AST methods for vancomycin resistance on cultured isolates from SF specimens.

69

| Organism Identified by SOC and
Detected by BioFire JI Panel | N | | | Positive Percent
Agreement | Negative Percent
Agreement | | |
|----------------------------------------------------------------|---|----|-------|-------------------------------|-------------------------------|------------|--|
| | R | S | 0/0 | 95% CI | % | 95% CI | |
| Overall
(any
applicable bacteria Detected) | 0 | 11 | (0/0) | | 90.9%
(10/11) | 62.3-98.4% | |
| Enterococcus faecalis | 0 | 10 | (0/0) | | 100%
(10/10) | 72.2-100% | |
| Enterococcus faecium | 0 | | (0/0) | - | 0%
(0/1) | | |

Archived Specimen Study

Many analytes on the BioFire Joint Infection (JI) Panel were of low prevalence during the prospective study and were not encountered in large enough numbers to adequately demonstrate system performance. To supplement the results of the prospective clinical study, an evaluation of preselected archived retrospective synovial fluid specimens was performed at BioFire.

A total of 134 frozen archived specimens were obtained from external laboratories for testing in this evaluation: 107 specimens were expected to contain a single analyte of interest, 14 specimens were expected to contain two analytes of interest, and 13 specimens were expected to be negative for all analytes of interest. Twenty-five (25) specimens were excluded from performance analysis due to low volume (23), because they were found to be the wrong specimen type (1), or because they were discovered to be a duplicated specimen. The remaining 97 expected positives and 12 expected negatives were further analyzed.

Prior to testing with the BioFire JI Panel, the composition/integrity of the laboratoryidentified analytes in archived specimens was first confirmatory molecular methods. Confirmation testing verified the presence of 93 out of 109 expected analytes (93/109; 85.3%) in a total of 88 of the 97 expected positive specimens. Specimens with unconfirmed (or unexpected) analytes were excluded from performance calculations for that particular analyte.

Table 79: BioFire Joint Infection Panel Performance Summary for Confirmed Archived Specimens

		Sensitivity/PP A		Specificity/NPA
Analyte	TP/
(TP + FN)	%	95%CI	TN/
(TN + FP)	%	95%CI
	Gram Positive Bacteria
Cutibacterium avidum/granulosum	3/3	100	43.9-100%	4/4	100	51.0-100%
Enterococcus faecalis	8/8	100	67.6-100%	92/92	100	96.0-100%
Enterococcus faeciu	1/1	100		100/100	100	96.3-100%
Staphylococcus lugdunensis	8/8	100	67.6-100%	94/94	100	96.1-100%
Streptococcus agalactiae	15/16	93.8	71.7-98.9%	81/81	100	95.5-100%
Streptococcus pneumoniae	1/1	100		101/101	100	96.3-100%
Streptococcus pyogenes	3/3	100	43.9-100%	98/98	100	96.2-100%
	Gram Negative Bacteria

70

Analyte	Sensitivity/PPA			Specificity/NPA
	TP/
(TP + FN)	%	95%CI	TN/
(TN + FP)	%	95%CI
Enterobacter cloacae complex	8/9	88.9	56.5-98.0	86/86	100	95.7-100%
Escherichia coli	9/9	100	70.1-100%	91/91	100	95.9-100%
Haemophilus influenzae	1/1	100	-	6/6	100	61.0-100%
Kingella kingae	1/1	100	-	100/100	100	96.3-100%
Klebsiella aerogenes	1/1	100	-	101/101	100	96.3-100%
Klebsiella pneumoniae group	3/3	100	43.9-100%	98/98	100	96.2-100%
Neisseria gonorrhoeae	2/2	100	34.2-100%	100/100	100	96.3-100%
Proteus spp.	3/3	100	43.9-100%	97/97	100	96.2-100%
Pseudomonas aeruginosa	13/13	100	77.2-100%	87/87	100	95.8-100%
Salmonella spp.	3/3	100	43.9-100%	98/98	100	96.2-100%
Serratia marcescens	2/2	100	34.2-100%	99/99	100	96.3-100%
Candida	2/2	100	34.2-100%	100/100	100	96.3-100%
Candida albicans	1/1	100	-	101/101	100	96.3-100%
CTX-M	3/3	100	43.9-100%	32/32	100	89.3-100%

Contrived Specimen Testing

Some analytes were of insufficient prevalence in the prospective and archived specimen evaluations to adequately demonstrate system performance. Therefore, contrived clinical specimens were created to evaluate the performance of the BioFire JI Panel assays for these rare analytes. Note that results for mecA/C and MREJ (MRSA) was not rare in the prospective study, but was included in this study for the evaluation of the rare antimicrobial gene mecC. Contrived specimens (N=1235) were spiked using residual clinical samples that were pre-screened and characterized as negative for the analytes of interest. Specimens were spiked with a variety of different isolates/strains for each organism at concentrations that spanned the detection range of each assay such that approximately 50% of specimens were spiked at a near-LoD test level (i.e. within ~2-fold of the assay LoD). Due to changes in the methods used for organism quantification over the course of the study, specimens were also spiked with analytes at levels below the established LoD for each assay.

Different isolates of organisms were used from those used in analytical testing when possible. Samples positive for one analyte served as negatives for other analytes. Eighty-one (81) negative (unspiked) samples were also randomized with the spiked specimens to facilitate specimen blinding.

The results of the 1235 specimens tested in this study are summarized in Table 80 below

Table 80. BioFire Joint Infection Panel Performance Summary for Contrived Specimen Testing

71

| Analyte | Level Tested | TP/
(TP + FN) | Sensitivity/PPA
% | 95%CI | TN/
(TN + FP) | Specificity/NPA
% | 95%CI |
|------------------------------------|--------------|------------------|----------------------|------------|------------------|----------------------|------------|
| Gram Positive Bacteria | | | | | | | |
| Anaerococcus prevotii/vaginalisa,b | ≥ LoD | 83/93 | 89.2 | 81.3-94.1% | 1125/1125 | 100 | 99.7-100% |
| | Overall | 83/95 | 87.4 | 79.2-92.6% | | | |
| Clostridium perfringensc | ≥ LoD | 92/102 | 90.2 | 82.9-94.6% | 1101/1101 | 100 | 99.7-100% |
| | Overall | 113/134 | 84.3 | 77.2-89.5% | | | |
| Cutibacterium avidum/granulosumd | ≥ LoD | 74/82 | 90.2 | 81.9-95.0% | 1128/1128 | 100 | 99.7-100% |
| | Overall | 80/107 | 74.8 | 65.8-82.0% | | | |
| Enterococcus faecalis | ≥ LoD | 51/51 | 100 | 93.0-100% | 1182/1182 | 100 | 99.7-100% |
| | Overall | 53/53 | 100 | 93.2-100% | | | |
| Enterococcus faeciume | ≥ LoD | 63/65 | 96.9 | 89.5-99.2% | 1169/1170f | 99.9 | 99.5-100% |
| | Overall | 63/65 | 96.9 | 89.5-99.2% | | | |
| Finegoldia magnag | ≥ LoD | 78/87 | 89.7 | 81.5-94.5% | 1142/1142 | 100 | 99.7-100% |
| | Overall | 82/93 | 88.2 | 80.1-93.3% | | | |
| Parvimonas micrah | ≥ LoD | 52/57 | 91.2 | 81.1-96.2% | 1158/1158 | 100 | 99.7-100% |
| | Overall | 54/77 | 70.1 | 59.2-79.2% | | | |
| Peptoniphilusi | ≥ LoD | 56/61 | 91.8 | 82.2-96.4% | 1173/1173 | 100 | 99.7-100% |
| | Overall | 57/62 | 91.9 | 82.5-96.5% | | | |
| Peptostreptococcus anaerobiusj | ≥ LoD | 91/91 | 100 | 95.9-100% | 1135/1135 | 100 | 99.7-100% |
| | Overall | 98/100 | 98.0 | 93.0-99.4% | | | |
| Staphylococcus lugdunensisk | ≥ LoD | 46/48 | 95.8 | 86.0-98.8% | 1184/1185f | 99.9 | 99.5-100% |
| | Overall | 48/50 | 96.0 | 86.5-98.9% | | | |
| Streptococcus agalactiae | ≥ LoD | 58/58 | 100 | 93.8-100% | 1175/1175 | 100 | 99.7-100% |
| | Overall | 59/59 | 100 | 93.9-100% | | | |
| Streptococcus pneumoniael | ≥ LoD | 70/76 | 92.1 | 83.8-96.3% | 1152/1157f | 99.6 | 99.0-99.8% |
| | Overall | 70/78 | 89.7 | 81.0-94.7% | | | |
| Streptococcus pyogenesm | ≥ LoD | 64/65 | 98.5 | 91.8-99.7% | 1170/1170 | 100 | 99.7-100% |
| | Overall | 64/65 | 98.5 | 91.8-99.7% | | | |
| Gram Negative Bacteria | | | | | | | |
| Bacteroides fragilisn | ≥ LoD | 95/95 | 100 | 96.1-100% | 1125/1125 | 100 | 99.7-100% |
| | Overall | 98/100 | 98.0 | 93.0-99.4% | | | |
| Citrobactero | ≥ LoD | 67/69 | 97.1 | 90.0-99.2% | 1165/1165 | 100 | 99.7-100% |
| | Overall | 67/70 | 95.7 | 88.1-98.5% | | | |
| Enterobacter cloacae complex | ≥ LoD | 48/48 | 100 | 92.6-100% | 1185/1185 | 100 | 99.7-100% |
| | Overall | 50/50 | 100 | 92.9-100% | | | |
| Escherichia coli | ≥ LoD | 75/75 | 100 | 95.1-100% | 1158/1158 | 100 | 99.7-100% |
| | Overall | 75/75 | 100 | 95.1-100% | | | |
| Haemophilus influenzaep | ≥ LoD | 52/53 | 98.1 | 90.1-99.7% | 1180/1180 | 100 | 99.7-100% |
| | Overall | 53/55 | 96.4 | 87.7-99.0 | | | |
| Kingella kingae | ≥ LoD | 48/48 | 100 | 92.6-100% | 1185/1185 | 100 | 99.7 -100% |
| | Overall | 50/50 | 100 | 92.9-100% | | | |
| Klebsiella aerogenesq | ≥ LoD | 97/97 | 100 | 96.2-100% | 1135/1135 | 100 | 99.7-100% |
| | Overall | 99/100 | 99.0 | 94.6-99.8% | | | |
| Klebsiella pneumoniae group | ≥ LoD | 93/93 | 100 | 96.0-100% | 1141/1141 | 100 | 99.7-100% |
| | Overall | 94/94 | 100 | 96.1-100% | | | |
| Morganella morganiir | ≥ LoD | 59/63 | 93.7 | 84.8-97.5% | 1171/1171 | 100 | 99.7-100% |
| | Overall | 59/64 | 92.2 | 83.0-96.6% | | | |
| Neisseria gonorrhoeaes | ≥ LoD | 46/48 | 95.8 | 86.0-98.8% | 1178/1179f | 99.9 | 99.5-100% |
| Analyte | Level Tested | Sensitivity/PPA | | | Specificity/NPA | | |
| | | TP/
(TP + FN) | % | 95%CI | TN/
(TN + FP) | % | 95%CI |
| Proteus spp.t | Overall | 47/50 | 94.0 | 83.8-97.9% | | | |
| | ≥ LoD | 52/52 | 100 | 93.1-100% | 1182/1182 | 100 | 99.7-100% |
| Pseudomonas aeruginosau | Overall | 52/53 | 98.1 | 90.1-99.7% | | | |
| | ≥ LoD | 117/119 | 98.3 | 94.1-99.5% | 1105/1105 | 100 | 99.7-100% |
| Salmonella spp.v | Overall | 121/125 | 96.8 | 92.1-98.7% | | | |
| | ≥ LoD | 57/60 | 95.0 | 86.3-98.3% | 1173/1173 | 100 | 99.7-100% |
| Serratia marcescensw | Overall | 59/62 | 95.2 | 86.7-98.3% | | | |
| | ≥ LoD | 53/54 | 98.1 | 90.2-99.7% | 1179/1179 | 100 | 99.7-100% |
| Yeast | | | | | | | |
| Candidax | Overall | 54/56 | 96.4 | 87.9-99.0% | | | |
| | ≥ LoD | 102/105 | 97.1 | 91.9-99.0% | 1126/1126 | 100 | 99.7-100% |
| Candida albicansy | Overall | 105/109 | 96.3 | 90.9-98.6% | | | |
| | ≥ LoD | 50/51 | 98.0 | 89.7-99.7% | 1182/1182 | 100 | 99.7-100% |
| | Overall | 52/53 | 98.1 | 90.1-99.7% | | | |

72

"Sequence variation in A. vaginalis isolates result in impaired detection near the LoD of the assay, See Table 52.

bTen Anaerococcus prevotii/vaginalis FN were observed at or above LoD and two FN were observed below LoD.

·Ten Clostridium perfringens FN were observed at or above LoD and 11 FN were observed below LoD.

dEight Cutibacterium avidum/granulosum FN were observed at or above LoD and 19 FN were observed below LoD.

eBoth Enterococcus faecium FN were observed at or above LoD.

TFP results due to background contamination in the matrix used for spiking.

8Nine Finegoldia magna FN were observed at or above LoD and two FN were observed below LoD.

below Parvimonas micra FN were observed at or above LoD and 18 FN were observed below LoD.

Five Peptoniphilus FN were observed at or above LoD.

Both Peptostreptococcus anaerobius FN were observed below LoD.

k Both Staphylococcus lugdunensis FN were observed at or above LoD.

'Six Streptococcus pneumoniae FN were observed at or above LoD and two FN were observed below LoD.

mThe Streptococcus pyogenes FN was observed above LoD.

"Both Bacteroides fragilis FN were observed below LoD.

oTwo Citrobacter FN were observed at or above LoD and one FN was observed below LoD.

POne Haemophilus influenzae FN was observed above LoD and one FN was observed below LoD.

9The Klebsiella aerogenes FN was observed below LoD.

Four Morganella morganii FN were observed at or above LoD and one FN was observed below LoD.

STwo Neisseria gonorrhoeae FN were observed at or above LoD and one FN was observed below LoD.

tThe Proteus spp. FN was observed below LoD.

"Two Pseudomonas aeruginosa FN were observed at or above LoD and two FN were observed below LoD.

·Three Salmonella spp. FN were observed at or above LoD.

"One Serratia marcescens FN was observed at or above LoD and one FN was observed below LoD.

*Three Candida FN were observed at or above LoD and one FN was observed below LoD.

The Candida albicans FN was observed above LoD.

Table 81. BioFire Joint Infection Panel Performance Summary for Contrived Specimen Testing - AMR Genes

Analyte	Level Tested	Sensitivity/PPA			Specificity/NPA
CTX-Ma	≥ LoD	149/150	99.3	96.3-99.9%	544/544	100	99.3-100%
	Overall	152/153	99.3	96.4-99.9%

73

Analyte	Level Tested	Sensitivity/PPA			Specificity/NPA
		TP/
(TP + FN)	%	95%CI	TN/
(TN + FP)	%	95%CI
IMP	≥ LoD	90/90	100	95.9-100%
IMP	Overall	93/93	100	96.0-100%	603/603	100	99.4-100%
KPC	≥ LoD	77/77	100	95.2-100%
KPC	Overall	79/79	100	95.4-100%	618/618	100	99.4-100%
mecA/C and MREJ (MRSA)b,c	≥ LoD	48/48	100	92.6-100%
mecA/C and MREJ (MRSA)b,c	Overall	49/49	100	92.7-100%	46/53d	86.8	75.2-93.5%
NDMe	≥ LoD	66/67	98.5	92.0-99.7%
NDMe	Overall	66/68	97.1	89.9-99.2%	629/629	100	99.4-100%
OXA-48-like	≥ LoD	64/64	100	94.3-100%
OXA-48-like	Overall	65/65	100	94.4-100%	532/532	100	99.3-100%
vanA/B	≥ LoD	96/96	100	96.2-100%
vanA/B	Overall	98/98	100	96.2-100%	18/19d	94.7	75.4-99.1%
VIM	≥ LoD	79/79	100	95.4-100%
VIM	Overall	83/83	100	95.6-100%	614/614	100	99.4-100%

"The CTX-M FN was observed above the host organism's LoD.

bResults were reported as N/A for the resistance marker because the host organism was reported as Not Detected. "Two different strains of Staphylococcus aureus containing mecC were used for spiking 50 contrived specimens. dFP results due to background contamination in the matrix used for spiking.

"One NDM FN was observed at or above the host organism's LoD and one FN was observed below the host organism's LoD.

12. Clinical Specificity:

See Clinical Sensitivity section above.

13. Other Clinical Supportive Data (When 1. and 2. Are Not Applicable):

Not applicable.

D Clinical Cut-Off:

Not applicable.

E Expected Values/Reference Range:

In the prospective clinical evaluation of the BioFire JI Panel, 1544 synovial fluid (SF) specimens were collected and tested at 13 study sites across the United States and Europe over approximately two years (May 2018 to March 2020). Expected values (as determined by the BioFire JI Panel) are stratified by enrollment site in Table 82.

Table 82. Expected Value (EV) as Determined by BioFire JI Panel: Summary by Site for SF Specimens Collected During the BioFire JI Panel Prospective

74

| | | Overall
(N=1544) | | Site 1
(N=352) | | Site 2
(N=88) | | Site 3
(N=201) | | Site 4
(N=89) | | Site 5
(N=102) | | Site 6
(N=87) | | Site 7
(N=197) | | Site 8
(N=66) | | Site 9
(N=146) | | |
|----------------------------------------|-----|---------------------|----|-------------------|---|------------------|----|------------------------|---|------------------|----|-------------------|----|------------------|---|-------------------|---|------------------|----|-------------------|---|----|
| BioFire JI Panel
Result | | | | | | | | | | | | | | | | | | | | | | |
| | # | EV | # | EV | # | EV | # | EV | # | EV | # | EV | # | EV | # | EV | # | EV | # | EV | | |
| | | | | | | | | Gram Positive Bacteria | | | | | | | | | | | | | | |
| Anaerococcus
prevotii/vaginalis | 1 | 0.1% | 0 | 0% | 0 | 0% | 1 | 0.5% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | | |
| Clostridium
perfringens | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | | |
| Cutibacterium
avidum/granulos
um | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | | |
| Enterococcus
faecalis | 15 | 1.0% | 4 | 1.1% | 0 | 0% | 6 | 3.0% | 0 | 0% | 0 | 0% | 1 | 1.1% | 1 | 0.5% | 1 | 1.5% | 0 | 0% | | |
| Enterococcus
faecium | 3 | 0.2% | 1 | 0.3% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 1 | 1.5% | 0 | 0% | | |
| Finegoldia
magna | 4 | 0.3% | 0 | 0% | 0 | 0% | 2 | 1.0% | 0 | 0% | 0 | 0% | 0 | 0% | 1 | 0.5% | 1 | 1.5% | 0 | 0% | | |
| Parvimonas
micra | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | | |
| Peptoniphilus | 2 | 0.1% | 0 | 0% | 0 | 0% | 1 | 0.5% | 0 | 0% | 0 | 0% | 1 | 1.1% | 0 | 0% | 0 | 0% | 0 | 0% | | |
| Peptostreptococc
us anaerobius | 3 | 0.2% | 1 | 0.3% | 0 | 0% | 1 | 0.5% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | | |
| Staphylococcus
aureus | 120 | 7.8% | 19 | 5.4% | 8 | 9.1% | 16 | 8.0% | 4 | 4.5% | 13 | 12.7% | 20 | 23.0% | 6 | 3.0% | 3 | 4.5% | 12 | 8.2% | | |
| Staphylococcus
lugdunensis | 5 | 0.3% | 0 | 0% | 0 | 0% | 3 | 1.5% | 0 | 0% | 0 | 0% | 1 | 1.1% | 0 | 0% | 0 | 0% | 1 | 0.7% | | |
| Streptococcus
spp. | 50 | 3.2% | 6 | 1.7% | 6 | 6.8% | 3 | 1.5% | 3 | 3.4% | 7 | 6.9% | 9 | 10.3% | 4 | 2.0% | 3 | 4.5% | 4 | 2.7% | | |
| Streptococcus
agalactiae | 11 | 0.7% | 0 | 0% | 2 | 2.3% | 1 | 0.5% | 1 | 1.1% | 0 | 0% | 1 | 1.1% | 2 | 1.0% | 1 | 1.5% | 2 | 1.4% | | |
| Streptococcus
pneumoniae | 3 | 0.2% | 1 | 0.3% | 0 | 0% | 0 | 0% | 0 | 0% | 1 | 1.0% | 1 | 1.1% | 0 | 0% | 0 | 0% | 0 | 0% | | |
| Streptococcus
pyogenes | 11 | 0.7% | 2 | 0.6% | 1 | 1.1% | 0 | 0% | 2 | 2.2% | 0 | 0% | 1 | 1.1% | 0 | 0% | 2 | 3.0% | 0 | 0% | | |
| | | | | | | | | Gram Negative Bacteria | | | | | | | | | | | | | | |
| Bacteroides
fragilis | 1 | 0.1% | 1 | 0.3% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | | |
| Citrobacter | 2 | 0.1% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 1 | 1.1% | 1 | 0.5% | 0 | 0% | 0 | 0% | | |
| Enterobacter
cloacae complex | 4 | 0.3% | 1 | 0.3% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 1 | 1.1% | 1 | 0.5% | 0 | 0% | 1 | 0.7% | | |
| Escherichia coli | 15 | 1.0% | 3 | 0.9% | 1 | 1.1% | 4 | 2.0% | 0 | 0% | 1 | 1.0% | 5 | 5.7% | 0 | 0% | 0 | 0% | 1 | 0.7% | | |
| Haemophilus
influenzae | 2 | 0.1% | 1 | 0.3% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 1 | 1.1% | 0 | 0% | 0 | 0% | 0 | 0% | | |
| Kingella kingae | 7 | 0.5% | 1 | 0.3% | 0 | 0% | 1 | 0.5% | 0 | 0% | 1 | 1.0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | | |
| | | Overall
(N=1544) | | Site 1
(N=352) | | Site 2
(N=88) | | Site 3
(N=201) | | Site 4
(N=89) | | Site 5
(N=102) | | Site 6
(N=87) | | Site 7
(N=197) | | Site 8
(N=66) | | Site 9
(N=146) | | |
| BioFire JI Panel
Result | # | EV | # | EV | # | EV | # | EV | # | EV | # | EV | # | EV | # | EV | # | EV | # | EV | # | EV |
| Klebsiella
aerogenes | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | | |
| Klebsiella
pneumoniae
group | 5 | 0.3% | 1 | 0.3% | 0 | 0% | 1 | 0.5% | 0 | 0% | 1 | 1.0% | 3 | 3.4% | 0 | 0% | 0 | 0% | 0 | 0% | | |
| Morganella
morganii | 3 | 0.2% | 0 | 0% | 0 | 0% | 1 | 0.5% | 0 | 0% | 0 | 0% | 2 | 2.3% | 0 | 0% | 0 | 0% | 0 | 0% | | |
| Neisseria
gonorrhoeae | 5 | 0.3% | 2 | 0.6% | 0 | 0% | 0 | 0% | 2 | 2.2% | 0 | 0% | 1 | 1.1% | 0 | 0% | 0 | 0% | 0 | 0% | | |
| Proteus spp. | 8 | 0.5% | 1 | 0.3% | 0 | 0% | 5 | 2.5% | 0 | 0% | 0 | 0% | 2 | 2.3% | 0 | 0% | 0 | 0% | 0 | 0% | | |
| Pseudomonas
aeruginosa | 5 | 0.3% | 1 | 0.3% | 0 | 0% | 2 | 1.0% | 0 | 0% | 0 | 0% | 0 | 0% | 1 | 0.5% | 0 | 0% | 1 | 0.7% | | |
| Salmonella spp. | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | | |
| Serratia
marcescens | 3 | 0.2% | 1 | 0.3% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 1 | 1.1% | 0 | 0% | 0 | 0% | 1 | 0.7% | | |
| AMR Genes | | | | | | | | | | | | | | | | | | | | | | |
| CTX-M | 5 | 0.3% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 1 | 1.0% | 4 | 4.6% | 0 | 0% | 0 | 0% | 0 | 0% | | |
| IMP | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | | |
| KPC | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | | |
| mecA/C and
MREJ (MRSA) | 23 | 1.5% | 4 | 1.1% | 3 | 3.4% | 0 | 0% | 0 | 0% | 0 | 0% | 4 | 4.6% | 0 | 0% | 1 | 1.5% | 6 | 4.1% | | |
| NDM | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | | |
| OXA-48-like | 1 | 0.1% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 1 | 1.1% | 0 | 0% | 0 | 0% | 0 | 0% | | |
| vanA/B | 3 | 0.2% | 1 | 0.3% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 1 | 1.5% | 1 | 0.7% | | |
| VIM | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | | |
| Yeast | | | | | | | | | | | | | | | | | | | | | | |
| Candida | 5 | 0.3% | 0 | 0% | 1 | 1.1% | 3 | 1.5% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 1 | 0.7% | | |
| Candida albicans | 3 | 0.2% | 0 | 0% | 0 | 0% | 2 | 1.0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | 1 | 0.7% | | |

75

76

| BioFire JI Panel
Result | # | Site 10
(N=136) | EV | # | Site 11
(N=17) | EV | # | Site 12
(N=49) | EV | # | Site 13
(N=14) | EV |
|------------------------------------|---|--------------------|----|-------|-------------------|-------|---|-------------------|----|---|-------------------|----|
| Anaerococcus
prevotii/vaginalis | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | | | | |
| Clostridium
perfringens | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | | | | |
| Cutibacterium
avidum/granulosum | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | | | | |
| Enterococcus
faecalis | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | | | | |
| Enterococcus
faecium | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | | | | |
| Finegoldia magna | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | | | | |
| Parvimonas micra | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | | | | |
| Peptoniphilus | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | | | | |
| Peptostreptococcus
anaerobius | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | | | | |
| Staphylococcus
aureus | 4 | 2.9% | 3 | 17.6% | 9 | 18.4% | 3 | 21.4% | | | | |
| Staphylococcus
lugdunensis | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | | | | |
| Streptococcus spp. | 2 | 1.5% | 1 | 5.9% | 1 | 2.0% | 1 | 7.1% | | | | |
| Streptococcus
agalactiae | 1 | 0.7% | 0 | 0% | 0 | 0% | 0 | 0% | | | | |
| Streptococcus
pneumoniae | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | | | | |
| Streptococcus
pyogenes | 0 | 0% | 1 | 5.9% | 1 | 2.0% | 1 | 7.1% | | | | |
| Bacteroides fragilis | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | | | | |
| Citrobacter | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | | | | |
| Enterobacter
cloacae complex | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | | | | |
| Escherichia coli | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | | | | |
| Haemophilus
influenzae | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | | | | |
| Kingella kingae | 0 | 0% | 0 | 0% | 4 | 8.2% | 0 | 0% | | | | |
| Klebsiella
aerogenes | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | | | | |
| Klebsiella
pneumoniae group | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | | | | |
| Morganella
morganii | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% | | | | |

77

| BioFire JI Panel
Result | Site 10
(N=136) | | Site 11
(N=17) | | Site 12
(N=49) | | Site 13
(N=14) | |
|----------------------------|--------------------|------|-------------------|----|-------------------|------|-------------------|------|
| | # | EV | # | EV | # | EV | # | EV |
| Neisseria
gonorrhoeae | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% |
| Proteus spp. | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% |
| Pseudomonas
aeruginosa | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% |
| Salmonella spp. | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% |
| Serratia
marcescens | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% |
| CTX-M | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% |
| IMP | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% |
| KPC | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% |
| mecA/C and MREJ
(MRSA) | 2 | 1.5% | 0 | 0% | 2 | 4.1% | 1 | 7.1% |
| NDM | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% |
| OXA-48-like | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% |
| vanA/B | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% |
| VIM | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% |
| Candida | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% |
| Candida albicans | 0 | 0% | 0 | 0% | 0 | 0% | 0 | 0% |

78

In addition, the observed multiple detections (as determined by the BioFire JI Panel) during the prospective clinical evaluation are presented in Table 15. At least one organism was detected in a total of 242 SF specimens (15.7% positivity rate: 242/1544). Polymicrobial detections of up to seven organisms were observed.

| BioFire JI Panel
Organism Result | Expected Value (as Determined
by Testing of 1544 Prospective SF
Specimens) | |
|-------------------------------------|----------------------------------------------------------------------------------|--------------------------------|
| | Number
Detected
and Reported | % of Total
(% of Positives) |
| Detected (at least one
result) | 242 | 15.7%
(100%) |
| One analyte result | 226 | 14.6%
(93.4%) |
| Two analyte results | 12 | 0.8%
(5.0%) |
| Three analyte results | 2 | 0.1%
(0.8%) |
| More than three organism
results | 2a | 0.1%
(0.8%) |

Table 83: Expected Values Multiple Detections as Determined by the BioFire JI Panel for the BioFIre JI Panel Clinical Evaluation (Mav 2018 - March 2020)

ªOne specimen had six organisms and one specimen had seven organisms observed

The BioFire JI Panel reported a total of 16 specimens with discernible detection of multiple organisms (1.0% of all specimens, 16/1544; and 6.6% of positive specimens, 16/242). The different types of co-detections (categorized by gram stain classification) as reported by the BioFire JI Panel are presented in Table 98 below. The resulting co-detection analyte combinations are presented in Table 99. This table also indicates the number of specimens with false positive (FP) results for each co-detection combination, as well as the specific analytes that were discrepant. FP results were determined by comparison only to the primary comparator method (e.g., standard of care (SOC) culture for organisms, and molecular comparator for the antimicrobial resistance (AMR) genes, irrespective of host organism SOC culture results).

Table 84: Expected Values (Co-detection Types as Determined by the BioFIre JI Panel for the BioFire JI Panel Prospective Clinical Evaluation

79

| BioFire JI Panel Co-

Detection Type	Positive Specimens (N =16)
	#	EV
Gram Positive + Gram
Positive	4	25.0%
Gram Positive + Gram
Negative	9	56.3%
Gram Positive + Yeast	0	0%
Gram Negative + Gram
Negative	2	12.5%
Gram Negative + Yeast'	1	6.3%
Gram Positive + Gram
Negative + Yeast	0	0%

Table 85: Co-detection Combinations as Determined by the BioFire JI Panel, Prospective Study

80

| Analyte
1 | Analyte
2 | Analyte
3 | Analyte
4 | Analyte
5 | Analyte
6 | Analyte 7 | AMR
Gene(s) | Total
Specimens
with Co-
Detection
Combination | #
Specimens
with False
Positive
Co-
Detections* | False Positive
Analyte(s) |
|---------------------------|-------------------------------------------|---------------------------|----------------|-----------------------------------------|-----------------------------------------|------------------|-----------------------------------------|------------------------------------------------------------|----------------------------------------------------------------|--------------------------------------------------------------|
| E. faecalis | F. magna | K.
pneumoniae
group | M.
morganii | Peptoniphilus | Proteus spp. | P.
anaerobius | - | 1 | 1 | F. magna, M.
morganii,
Peptoniphilus, P.
anaerobius |
| E. cloacae
complex | E. faecalis | H.
influenzae | K. Kingae | Streptococcus
spp., S.
pneumoniae | Streptococcus
s spp., S.
pyogenes | - | - | 1 | 1 | H. influenzae, K.
kingae |
| A.
prevotii/vaginalis | F. magna | Streptococcus
s spp. | - | - | - | - | - | 1 | 0 | - |
| E. coli | E. faecium | Proteus spp. | - | - | - | - | vanA/B | 1 | 1 | E. faecium, Proteus
spp. |
| B. fragilis | P.
anaerobius | - | - | - | - | - | - | 1 | 1 | B. fragilis, P.
anaerobius |
| Candida, C.
albicans | E. cloacae
complex | - | - | - | - | - | - | 1 | 1 | E. cloacae complex |
| E. faecalis | Proteus spp. | - | - | - | - | - | - | 1 | 0 | - |
| E. faecalis | S. aureus | - | - | - | - | - | mecA/C and
MREJ
(MRSA) | 1 | 0 | - |
| E. coli | Proteus spp. | - | - | - | - | - | - | 1 | 0 | - |
| E. coli | S. aureus | - | - | - | - | - | CTX-M,
mecA/C, and
MREJ
(MRSA) | 1 | 1 | S. aureus |
| K. kingae | S. aureus | - | - | - | - | - | - | 1 | 1 | K. kingae, S. aureus |
| K.
pneumoniae
group | M. morganii | - | - | - | - | - | - | 1 | 1 | K. pneumoniae group,
M. morganii |
| Proteus spp. | S.
lugdunensis | - | - | - | - | - | - | 1 | 1 | Proteus spp., S.
lugdunensis |
| S.
marcescens | S. aureus | - | - | - | - | - | - | 1 | 1 | S. marcescens |
| S. aureus | Streptococcus
s spp., S.
agalactiae | - | - | - | - | - | - | 1 | 0 | - |
| S. aureus | Streptococcus
s spp., S.
pyogenes | - | - | - | - | - | - | 1 | 0 | - |
| | | | | | | | Total Co-Detections | 16 | 11 | 21/43ª |
| | | | | | | | Total Double Detections | 12 | 8 | 13/24 |
| | | | | | | | Total Quintuple Detections | 2 | 1 | 2/6 |
| | Total Sextuple Detections | | | | | | | 1 | 1 | 2/6 |

8 Determined by comparison to SOC culture for organisms, and molecular methods for AMR genes, irrespective of host organism SOC culture results

b. Of the 21 discrepant analytes (out of 43 total analytes), 20 (95.2%) were confirmed as being present in the specimen during discrepancy investigation: 2/20 (10.0%) were identified from additional laboratory testing performed as SOC and 18/20 (90.0%) were observed using an independent molecular method (59.9%) were detected by qMol, 17/187 (9.1%) were detected using an additional molecular method, and the remaining 3/187 (1.6%) were identified in SOC culture.

Other Supportive Performance Characteristics Data: F

Not applicable.

81

VII Proposed Labeling:

The labeling supports the decision to grant the De Novo request for this device.

Identified Risks and Mitigations:
Identified Risks to Health	Mitigation Measures
Risk of false test results leading to improper
patient management	Use of certain specimen collection devices
identified in special control (1).
Certain labeling information identified in
special control (2), including limitations,
warnings, device descriptions, explanation of
procedures, and performance information
identified in special controls (3)(iii) and
(3)(iv).
Certain design verification and validation
identified in special control (3), including
documentation of certain analytical studies and
clinical studies and device descriptions.
Failure to correctly interpret test results leading
to misdiagnosis and associated risk of false test
results	Certain labeling information identified in
special control (2), including limitations,
warnings, device descriptions, explanation of
procedures, and performance information
identified in special controls (3)(iii) and
(3)(iv).
Certain design verification and validation
identified in special control (3), including
documentation of certain analytical studies
and clinical studies and device descriptions.
Failure to correctly operate the device leading
to false test results or incorrect interpretation
of test results	Use of certain specimen collection devices
identified in special control (1).
Certain labeling information identified in
special control (2), including limitations,
warnings, device descriptions, explanation of
procedures, and performance information
identified in special controls (3)(iii) and
(3)(iv).
Certain design verification and validation
identified in special control (3), including
documentation of certain analytical studies
and clinical studies and device descriptions.

VIII Identified Risks and Mitigations:

IX Benefit/Risk Assessment:

A Summary of the Assessment of Benefit:

The benefit of the assay is aiding in the accurate diagnosis of specific agents of joint infections in conjunction with other clinical and laboratory findings. The BioFire Joint Infection Panel

82

simultaneously detects and identifies nucleic acids from multiple different pathogens as well as eight antimicrobial resistance genes in a platform that provides results in about an hour which is a significant improvement over standard microbiological methods. Aiding in the diagnosis of specific agents of joint infection and identification of genes associated with antimicrobial resistant strains may identify patients for which treatment may be appropriate, including, but not limited to, antibiotic therapy and revision surgery, if the infection is tied to a prosthetic joint. Appropriate treatment of prosthetic joint infection can lead to alleviation of symptoms associated with infection and restoration of joint function.

B Summary of the Assessment of Risk:

The risks associated with the device, when used as intended, are those related to the risk of false test results, the failure to correctly interpret test results, and failure to correctly operate the device.

The risk of a false positive test result is improper patient management, including inappropriate administration of unnecessary antibiotics or anti-fungal medications. Inappropriate administration of antibiotics is associated with toxicity, allergic reactions, and other adverse outcomes including secondary infections such as C. difficile colitis.

The risk of a false negative test result is delaved identification of the cause of the disease in the patient, which could lead to improper patient management, including administration of unnecessary treatment and/or delay or discontinuation of appropriate treatment. An undiagnosed infection or delayed diagnosis could result in increased morbidity and mortality.

Failure to correctly operate the device can lead to false test results. Failure to correctly interpret test results can lead to erroneous results (i.e., false positives, false negatives), with the same risks discussed above.

C Patient Perspectives:

Not applicable.

D Summary of the Assessment of Benefit-Risk:

General controls are insufficient to mitigate the risks associated with the device. However, the clinical benefits outweigh the risks for the proposed assay, considering the mitigations of the risks provided by the special controls established for this device, as well as general controls. The required special controls will help ensure that errors will be uncommon and will facilitate accurate assay implementation and interpretation of results. The clinical performance observed in the clinical trial suggests that errors will be uncommon and that the assay will provide substantial benefits to patients in the diagnosis of joint infection and when used in conjunction with other clinical and diagnostic findings.

The risk of false test results (both positive) is mitigated by the intended use clearly stating that the assay results are intended to be used with other clinical and laboratory findings which include standard of care culture to identify organisms and antimicrobial susceptibility testing. The risk of false results is also mitigated by the inclusion of performance characteristics. from analytical and clinical studies in the labeling.

83

Risks of failure to correctly interpret the test results are mitigated through the inclusion in the labeling of a detailed description of what the device detects, the specimen type for which testing is indicated, the type of results provided to the user in the intended use statement, as well as a detailed explanation of the interpretation of results. Finally, the risk of failure to correctly operate the device is mitigated by the inclusion of detailed directions for use in the package insert, such that the operator can successfully use the instrument.

The clinical performance observed in the clinical studies suggests that errors will be uncommon and that the assay will provide substantial benefits to patients in the diagnosis of joint infection when used in conjunction with other clinical and diagnostic findings.

Given the combination of the device's indications for use, labeling, the required general controls, and the special controls established for this device, the benefits outweigh the risks.

X Conclusion:

The De Novo request is granted and the device is classified under the following:

Product Code(s): QSN Device Type: Device to detect and identify microorganism nucleic acids and resistance markers from patients with suspected orthopedic infection Class: II Regulation:21 CFR 866.3988