Background: There is an increasing demand on laboratories to deliver respiratory viral diagnosis by molecular methods. Different strategies are explored which include – point of care testing which is simple, requires minimal hands on time, is fast and can be done in ward areas and not in centralised laboratories. Tests considered need to be shown to have good performance.
GenMark Diagnostics Inc. (Carlsbad, USA) has developed a respiratory panel assay (RP) for the ePlex system detecting 26 microbes, including 22 virus and 4 (atypical) bacteria in 90 min.
The RP cartridge contains all reagents required to run the RP Panel assay. Lysis and nucleic acid extraction, PCR amplification and hybridization-based electrochemical detection occur inside the cartridge, reducing the hands-on-time to less than 1 min per sample.
The objective of this study was to compare and study the performance of the new GenMark ePlex assay against the in house real-time PCR, a lab developed test (LDT).
Material and methods: 81 nasopharyngeal swabs samples (NPS) in UTM were previously tested by an in-house Real Time PCR. Samples selected contained the following respiratory pathogens: respiratory syncytial virus, influenza A, influenza B, rhinovirus, enterovirus, bocavirus, coronaviruses, metapneumovirus, parainfluenza viruses, Bordetella pertussis and Mycoplasma pneumoniae 32.1% samples were co-infected, even with 4 different organisms. Samples selected were less than 4 months old with only one freeze/thaw cycle. Ct values ranging from 17.11 to 40.39 mean 24.74.
200 μl of each NPS sample was added to the ePlex Sample Buffer device, transferred to the RP cartridge and then inserted on the ePlex device.
Agreement between the original LDT results and the results obtained with the ePlex assay was assessed as detected or not detected.
Additionally 5 successive 1:10 dilutions were performed for 7 different specimens: RSV, influenza A H1N1, influenza B, rhinovirus, bocavirus, Mycoplasma pneumoniae and Bordetella pertussis. Dilutions were tested on both assays to identify and compare the lower limit of detection to the LDT.
Results: Total concordance was observed in 91.73% of cases. Only 10 discrepancies were identified. 7 organisms were detected by the ePlex assay and missed by the LDT and 3 organisms were positive detected by the LDT and negative by the ePlex. Discrepancies were repeated in both assays showing same results.
Total concordance was observed in 80% of dilutions. 1 dilution 10−4 RSV sample was detected by the ePlex and resulted negative for the LDT. As well 6 samples (10−3–10−5 dilutions) were positive for the LDT and negative for the ePlex assay. B. pertussis did not detect the 2 lower dilutions as a different target gene was in use in ePlex assay.
Conclusion: The preliminary evaluation on a small sample set show a very good agreement across a range of pathogens with the GenMark compares in house real-time PCR. The assay was also found to be very simple and easy to perform and would be suitable for a hospital ward or outpatient environment.