PREMARKET EVALUATION OF THE BD MAXTM CLOSTRIDIUM DIFFICILE PCE ASSAY Raymond Widen, Carly Kubasek, Dominic Uy, John Markantonis and Suzane Silbert Department of Esoteric Testing/R&D, Tampa General Hospital, Tampa, FL 2013 CVS Annual Meeting T50 Table 3. Analytical sensitivity. Tested using dilutions of a positive sample comparing LOD for GeneXpert and BD MAXTM C. difficile PCR BD MAXTM Instrument Operation Results Abstract Materials and Methods Background: C. difficile is problematic in health care institutions as an iatrogenic infection relating to overgrowth of the organism following antimicrobial therapy and as a potential nosocomially transmitted agent due to contamination of surfaces with bacterial spores. The BD MAXTM is a relatively new sample in answer out system for PCR testing that can be used with FDA cleared IVD assays and for laboratory developed tests (LDT). Objectives: To evaluate the performance of the RUO BD MAXTM Clostridium difficile toxin B PCR assay on the BD MAX instrument. Methods: For this study we evaluated a combination of residual refrigerated/frozen and fresh stool samples submitted for C. difficile toxin detection using the Genexpert C. difficile assay which was our test of record at TGH. A total of 137 samples were tested with both assays using the methods described in the manufacturer’s kit instructions. Results: We found excellent overall agreement between the Genexpert and BD MAXTM assays for C.difficile toxin B. 95 samples were negative with both methods, 37 were positive with both methods, 4 were positive with Genexpert only and 1 was positive only on the BD MAXTM. The 4 Genexpert positive/BD MAXTM negative samples had been refrigerated and then frozen for several months and all had high Ct values on initial testing (cycles 34 and higher). The sample that was positive by BD MAXTM/Genexpert was a fresh sample with Ct 36 on the BD MAXTM. The BD MAXTM RUO was tested with a panel of agents commonly associated with diarrhea and was found to demonstrate no reactivity with any of these agents. Conclusions: The results suggest that the BD MAXTM C. difficile toxin PCR will represent a sensitive and specific alternative for detection of C. difficile infection once it receives FDA IVD clearance. One manual pipetting step – use 10µL loop to transfer stool sample to a specimen tube Table 1. Evaluation of C. difficile toxin B gene detection by Genexpert and BD MAXTM PCR assays Results BD MAXTM Dilution GeneXpert BD MAXTM Undiluted Positive 1/10 1/100 1/1,000 1/10,000 1/100,000 Negative 1/1,000,000 Positive Negative 37 4 1 95 Xpert C. difficile Load reagents and specimens onto the BD MAXTM rack CONCLUSIONS Table 2. Specificity challenge results. Each of the agents in the table below was prepared in specificity challenge mixes and tested with the BD MAXTM C. difficile PCR assay. Create worklist The BD MAXTM C. difficile toxin B gene PCR assay was compared with the Cepheid GeneXpert toxin B PCR. Both assays had similar performance in detecting C. difficile toxin B genes in stool samples. There were 4 samples positive with the GeneXpert and negative with the BD MAXTM. These 4 samples had been frozen and thawed and all had cycle numbers on the GeneXpert at 35 or higher. The sample that was positive on the BD MAXTM was a fresh sample and was positive at 37 cycles. 3. The BD MAXTM assay was challenged with organisms likely to be in samples referred for testing for C. difficile. No cross reactivity was observed with any of the challenge organisms, including other Clostridium species. 4. The BD MAXTM represents an effective option for detection of C. difficile with good sensitivity and specificity. We tested the system as an RUO premarket study. It is now FDA cleared as an in vitro diagnostic test. The system allows for simple sample in – answer out processing of up to 24 samples in about 2 hours. Organism ATCC # BD MAX E. coli 35218 NEG E. faecium pt isolate Specificity 1 E. cloacae 13047 E. faecalis 51299 S. marcescens 8100 K. pneumoniae 700603 C. albicans 10321 Ps. aeruginosa 27853 Specificity 2 B. fragilis 23765 S. aureus 977 S. epidermidis 14990 S. sonnei 25931 S. Enteritica 6539 Specificity 3 V. Parahemolyticus 17802 C. jejuni 33291 Adenovirus 41 VR-930 Coxsackie A15 and Echovirus 27 VR-104 VR -1069 NEG NEG Specificity 4 L. monocytogenes C. perfringens, C. sordelii C. sporogenes 13124 9714 3584 NEG NEG NEG Place racks and cartridges on BD MAXTM 1 loose/liquid stool container was received for C. difficile PCR testing by the current standard of care procedure – Cepheid GeneXpert C. difficile. The sample was processed according to package insert instructions. The BD MAXTM C. difficile assay was performed as follows: The residual stool sample was retrieved. Using a 10µL loop the sample was mixed and then transferred to a labeled BD MAXTM sample prep tube. This was repeated for each sample to be tested. The BD MAXTM sample prep tube was vortexed on a multiposition vortex mixer for 60 seconds. The sample prep tubes were transferred to the BD MAX carriers and an extraction cartridge was placed in the rack for each sample. The reagent tube was placed into the processing strip. The entire assembly was placed in the BD MAXTM instrument along with BD MAXTM PCR reaction cards. Sample identification was loaded into the BD MAXTM software and the run started. Close door to initiate run then release results through LIS interface