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Performance Characterization of the IRIDICA™ BAC SFT Assay* for Detection and Identification of Diverse Bacteria and Candida in Tissues and Body fluids

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Performance Characterization of the IRIDICA™ BAC SFT Assay* for Detection and Identification of Diverse Bacteria and Candida in Tissues and Body fluids Performance characterization of the IRIDICA™ BAC SFT Assay* for detection and identification of diverse bacteria and Candida in tissues and body fluids Mark W. Frinder, David Metzgar, Megan Rounds, Heather E. Carolan, Donna M. Toleno, Rangarajan Sampath, David J. Ecker, Lawrence B. Blyn Ibis Biosciences, an Abbott Company, Carlsbad, CA, USA Color Key Table 2: Potentially interfering substances tested with the 4 core organisms at 3X Objectives: Identifying causal organisms in Results: The BAC SFT Assay was able to detect and identify all IRIDICA detections, matched LOD in synovial Fluid, muscle tissue, and diluent matrices.Data shown reflects IRIDICA detections, unmatched Standard of care detections, missed by IRIDICA tissue and body fluid infections through tested organisms at concentrations of 5 to 1000 CFU/sample, concentration in the final 5ml sample. No interference was observed (all 4 targets and their associated antibiotic resistance markers were successfully culture-based methods is time-consuming and the sensitivity of the assay was comparable between Burkholderia vietnamiensis (1) and challenging. Culture-based methods are tissue, body fluid, and sample diluent matrices (Figure 1). The detected in 3/3 samples). Micrococcus luteus (1) Corynebacterium striatum (1) often rendered ineffective by antibiotic assay was able to detect organisms in the presence of diverse Test Substance Concentration Test Substance Concentration Corynebacterium accolens (2) Propionibacterium acnes (5) Pseudomonas entomophila/putida (1) pre-treatment, the presence of fastidious or tissues or fluids (Table 1), and potentially interfering Bilirubin 171 µmol/L * Doxycycline 67.5 µmol/L Acinetobacter junii (4) Hemoglobin 2 g/L Fluconazole 245 µmol/L uncultureable species, and growth inhibition substances (Table 2). No cross-reactivity to untargeted Triglycerides 37 mmol/L Gentamicin 21 µmol/L Staphylococcus aureus (14) Staphylococcus aureus (2) Klebsiella pneumoniae (1) associated with biofilm encasement. In order organisms, viruses, or human DNA was observed (Table 3). White blood cell count 15000 cells/µl Imipenem 0.2490 mg/mL Staphylococcus sp. (1) Klebsiella oxytoca (1) Staphylococcus lugdunensis (1) Glucose 1.2 mg/mL Metronidazole 701 µmol/L Staphylococcus stimulans (1) to address these issues, a broad-spectrum During the clinical sample study, the assay yielded an overall Staphylococcus epidermidis (3) Escherichia coli (4) Amikacin 136.8 µmol/L Piperacillin 1236 µg/mL Staphylococcus epidermidis (1) Escherichia coli (1) E.coli/Shigella flexneri/S.sonnei (1) Staphylococcus capitis (2) Escherichia vulneris (1) molecular assay based on a robust universal calculated positive percent agreement (PPA) with standard- Amphotericin B 0.0033 mg/mL Vancomycin 69 µmol/L Staphylococcus capitis (1) Enterobacter cloacae complex (2) Ceftazidime 0.1170 mg/mL Dexamethasone 1.53 µmol/L Staphylococcus caprae (1) Enterobacter aerogenes (1) lysis method, broad-spectrum mismatch- of-care microbiology results of 90% in all cases where the Staphylococcus saccharolyticus (1) Ciprofloxacin 30.2 µmol/L Dobutamine 15000 µg/mL Staphylococcus saccharolyticus (1) Proteus mirabilis (1) Pantoea vagans (1) tolerant PCR, and electrospray ionization presence of coinfecting organisms or normal flora did not Clindamycin 89.1 µmol/L Warfarin 32.5 µmol/L Enterococcus faecalis (1) Streptococcus agalactiae (1) Enterococcus faecalis (1) Streptococcus agalactiae (1) *Bilirubin did not pass the initial test at 342 µmol/L, due to a source which contained bacterial/fungal DNA Enterococcus faecium (2) Streptococcus disgalactiae (3) mass spectrometry (PCR/ESI-MS) was prevent comparison, and 95% when common contaminants Enterococcus faecium (1) Streptococcus mitis/pneumoniae (1) contamination. Final passing concentration for Bilirubin was equal to 171 µmol/L. Fungi Enterococcus avium (1) Viridans/anginosus group Streptococcus (2) Enterococcus asini (1) Streptococcus sp. (1) Candida albicans (5) Candida glabrata (2) developed. In the work described here, we were disregarded (Figure 2). Calculated negative percent Streptococcus intermedius (1) Candida albicans (1) Lactobacillus johnsonii (1) CM Jan 29 2015 Fungus detected, no ID provided (2) explore the analytical sensitivity, specificity, agreement (NPA) was 99% when all possible organism Table 3: Potentially cross-reacting organisms.The viruses and microorganisms robustness, reproducibility, and breadth of detections were considered. shown below were tested at targeted concentrations of 100,000 CFU/mL Figure 2: Organism by organism PPA and NPA between culture and IRIDICA for 201 (bacteria and fungi), or 100,000 TCID /mL or copies/mL for viruses, as deidentified waste human body fluid and tissue specimens. Both culture and the BAC SFT coverage of the method, and compare its 50 Color Key Campylobacter coli calculated for the final 5ml sample volume. Assay utilize single reagents to detect many or all targets. Therefore, the presence of Bacteroidetes Campylobacter jejuni Samples Samples with Detection Core LOD organisms (LOD [CFU/sample] Helicobacter pylori detected organisms can mask the detection of other organisms through competitive performance to that of traditional for synovial fluid / muscle tissue / diluent) Bundle Tested Bacteroides fragilis * Tested Detections Rate Analytical LOD organisms (CFU/sample) Epsilon- interference. For the analysis presented here, results were considered negative if and culture-based methods in a collection of Breadth of coverage (500 CFU/sample) proteobacteria Bartonella henselae Aspergillus flavus and Clavispora lusitaniae 3 0 0% Challenge organisms (500 CFU/sample) Bartonella quintana only if negative for all analytes, while positive samples were considered positive for all Alpha- Brucella neotomae Caulobacter segnis, Aspergillus niger, and Pedobacter heparinus 3 0 0% clinical waste specimens. proteobacteria detected analytes and excluded from analyses of other analytes. In order to account for Shewanella oneidensi, Streptomyces griseus, and Candida kefyr 3 0 0% Corynebacterium diphteriae (640) detections that had no valid comparator, any positive result by either technology in a Corynebacterium jeikeium (160) Proteo- Burkholderia cepacia Influenza, Parainfluenza, Adenovirus 3, and Adenovirus 31 3 0 0% Arcanobacterium haemolyticum Fusobacterium Beta- Bordetella pertussis sample that was negative for that analyte but positive for another by the other Microbacterium sp. nucleatum bacteria proteobacteria Eikenella corrodens Parvovirus B19, Coronavirus, and Herpes simplexvirus 1 3 0 0% Micrococcus luteus Fusobacteria Neisseria meningitidis technology was assigned to the “additional detection” category in the final comparison. Methods: The limit of detection (LOD) of the Mycobacterium chelonae Nocardia nova These additional detections are not shown in this figure, but are listed in the IRIDICA™ BAC SFT Assay (08N39-10, Propionibacterium acnes Actinobacteria Rothia dentocariosa Gamma- Table 4: Assay reproducibility at 3x LOD. The 4 core organisms were spiked at 3X sample-by-sample results shown in Tables 5 and 6 for the corresponding data subsets. CE-IVD)* was tested for three bacteria and Corynebacterium urealyticum proteobacteria Pseudomonas aeruginosa (40) LOD into blood, synovial fluid, muscle tissue, and diluent matrices. Two of the Stenotrophomonas maltophilia Acinetobacter baumannii (320) Table 6. Results from orthopedic specimens, by sample one Candida species in muscle tissue, Acinetobacter lwoffii core organisms were bundled (Candida albicans and Staphylococcus aureus). Moraxella catarrhalis IRIDICA-Positive Detections in Site Matched Positive Detections Additional IRIDICA Detections Additional Culture Detections Bacillus cereus (160) Haemophilus influenzae (40) Panel members were tested by 3 operators using 3 lots of Assay Strips paired Culture-Negative Specimens Clostridium difficile (160) Pasteurella multocida synovial fluid, and the sample diluent used Klebsiella pneumoniae, Staphylococcus aureus (mecA +) Clostridium perfringens Klebsiella pneumoniae (KPC +) with 3 sets of IRIDICA instruments over 5 days. Results reflect the number of 1 Ankle None None None (80/160/80) Veillonella dispar (40/160/80) Staphylococcus simulans Staphylococcus epidermidis (640) Klebsiella oxytoca (160) samples yielding the expected result / the number of samples tested. to resuspend tissue specimens (sterile Aeromonas hydrophila (40) 2 Arm Staphylococcus aureus None None None Staphylococcus haemolyticus Escherichia coli (160) Staphylococcus lugdunensis Clostridiales Firmicutes Enterobacter cloacae (640) Variable (Identity) # of Detections/ # of Tests Reproducibility 3 Arm Streptococcus sp., S. aureus None None None buffer). Based on the resulting Staphylococcus saprophyticus Proteus mirabilis (40) 4 Elbow Streptococcus intermedius None None Staphylococcus aureus Listeria monocytogenes Serratia marcescens (160) Bacillales Operator 1 73 / 75 97.3% Citrobacter freundii 5 Foot Proteus mirabilis, S. aureus None Serratia proteamaculans Enterococcus faecalis demonstration of equivalence, the LODs of a Vibrio parahaemolyticus Enterobacter aerogenes Operator 2 72 / 75 96.0% 6 Foot Staphylococcus aureus None None None Mycoplasma bovis Morganella morganii Candida albicans, Lactobacillales Providencia stuartii Operator 3 73 / 75 97.3% 7 Foot None None None further 19 bacteria and Candida species Proteus vulgaris Streptococcus dysgalactiae Enterococcus faecium (vanA +, vanB +) Streptococcus pneumoniae (1.25) Salmonella enterica Lot/Instrument
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