The Enigma ML FluAB-RSV assay: a fully automated molecular test for the rapid detection of influenza A, B and respiratory syncytial viruses in respiratory specimens

Figures & data

Table 1. Relative performance characteristics of several commercially available, rapid, multiplex molecular assays for the detection of influenza, RSV and other respiratory viruses.

Assay name and manufacturer	Technology	Targets detected	Sensitivity (95% CI)^†	Specificity (95% CI)^†	Total turnaround time (min)	Hands on time (min)	Specimen types accepted	Run size	Random access	Consumables storage conditions	Regulatory status
Alere™ I Influenza A&B	Isothermal nucleic acid amplification	Influenza A Influenza B	96.6 (90.6–98.8) 84.1 (73.2–91.1)	89.5 (85.9–92.3) 96.3 (93.9–97.8)	15	4	NS	1	Yes	2–8°C	CE-IVD, not currently available in US
Enigma^® ML FluAB-RSV, Enigma Diagnostics, Salisbury, UK	Multiplex RT-PCR	Influenza A Influenza B Respiratory syncytial virus	100 (98.2–100) 100 (98.2–100) 100 (98.2–100)	98.3 (95.5–99.4) 98.7 (96–99.6) 99.4 (97.2–99.9)	95	2	NPS	1–6	Yes	Room temperature	CE-IVD
FilmArray^® Respiratory Panel, BioFire Diagnostics Inc. (now BioMerieux), Salt Lake City, UT, USA	Multiplex nested-PCR Detection of targets using end-point melt curves	Adenovirus Bordetella pertussis Chlamydophila pneumonia Coronavirus HKU1 Coronavirus NL63 Coronavirus OC43 Coronavirus 229E Human metapneumovirus Human Rhinovirus/enterovirus Influenza A Influenza A subtype H1 Influenza A subtype H3 Influenza A subtype 2009 H1 Influenza B Mycoplasma pneumonia Parainfluenza virus 1 Parainfluenza virus 2 Parainfluenza virus 3 Parainfluenza virus 4 Respiratory syncytial virus	88.9 (70.8–97.7) 100 (54.1–100) 100 (NA) 95.8 (78.9–99.9) 95.8 (78.9–99.9) 100 (76.8–100) 100 (73.5–100) 94.6 (87.9–98.2) 92.7 (88.2–95.8) 90 (55.5–99.8) NA NA 88.9 (51.8–99.7) NA 100 (39.8–100) 100 (NA) 87.4 (47.4–99.7) 95.8 (78.9–99.9) 100 (66.4–100) 100 (93.2–100)	98.3 (97.2–99.1) 99.9 (99.5–100) 100 (99.7–100) 99.8 (99.1–100) 100 (99.6–100) 99.6 (99–99.9) 99.8 (99.4–100) 99.2 (98.3–99.7) 94.6 (92.6–96.2) 99.8 (99.2–100) 100 (99.6–100) 100 (99.6–100) 99.6 (99–99.9) 100 (99.6–100) 100 (99.7–100) 99.9 (99.5–100) 99.8 (99.4–100) 98.8 (97.8–99.4) 99.9 (99.5–100) 89.1 (86.8–91.2)	60	2	NPS	1	Yes	Room temperature	CE-IVD FDA approved, CLIA moderate complexity
Liat™ Influenza A/B Assay, IQuum Inc. Marlborough, MA, USA (now Roche)	RT-PCR	Influenza A Influenza B	100 (89.8–100) 100 (88.6–100)	96.8 (94.5–98.1) 94.1 (91.3–96)	25	2–5	NPS	1	Yes	2–8°C	FDA approved, CLIA moderate complexity
Simplexa™ Flu A/B & RSV Direct assay, Focus Diagnostics, Cypress CA, USA	Multiplex RT-PCR using the 3M Integrated Cycler	Influenza A Influenza B Respiratory syncytial virus	97.1 (89.9–99.2) 100 (84.5–100) 98.6 (92.5–99.8)	97.9 (96.4–98.7) 99.9 (99.2–100) 89.5 (84.1–93.3)	70	10	NPS	1–8	No	-20°C	CE-IVD FDA approved, CLIA moderate complexity
Verigene^® Respiratory Virus Plus Assay, Nanosphere, Northbrook, IL, USA	RT-PCR and Nanoparticle probe-based DNA-array hybridization	Influenza A Influenza A/H1 Influenza A/H3 sens Influenza A 2009/H1N1 Influenza B Respiratory syncytial virus A Respiratory syncytial virus B H275Y (Olsetamivir resistance)	98.7 (96.8–99.5) 100 (91–100) 100 (96.6–100) 99.5 (97.3–99.9) 100 (91.8–100) 100 (93.0–100) 100 (93.2–100) NA	93.2 (91.1–94.8) 99.9 (99.4–100) 100 (99.6–100) 100 (99.5–100) 99.7 (99.1–99.9) 100 (99.6–100) 99.9 (99.6–100) NA	155	5	NPA	1	Yes	2–8°C	CE-IVD, FDA approved, CLIA moderate complexity
Xpert^® Flu Assay (Cephied, Sunnyvale, CA, USA)	Real time multiplex RT-PCR	Influenza A Influenza A subtype 2009 H1N1 Influenza B	94.8 (90.4–97.6) 98.5 (92.1–100) 90 (73.5–97.9)	100 (97.9–100) 99.6 (98.1–100) 100 (98.9–100)	77	2	NA/NW, NPS	1	Yes	Room temperature	CE-IVD, FDA approved, CLIA moderate complexity

^†As stated in manufacturer’s package insert.

CLIA: Committee for Laboratory Improvement Act; NA: Nasal aspirate; NPS: Nasopharyngeal swab; NS: Nasal swab; NW: Nasal wash; RSV: Respiratory syncytial virus; RT-PCR: Reverse transcriptase polymerase chain reaction.

Figure 1. The Enigma ML platform in a single module configuration.

Figure 2. The Enigma^® ML cartridge.

Table 2. Performance characteristics of the Enigma ML FluAB/RSV assay using the Luminex xTAG RVP as gold standard comparator.

	True positive (n)	True negative (n)	False positive (n)	False negative (n)	Sensitivity (%) (95% CI)	Specificity (%) (95% CI)	Positive predictive value (%) (95% CI)	Negative predictive value (%) (95% CI)
Influenza A	25	176	3	0	100 (98.2–100)	98.3 (95.5–99.4)	89.3 (84.3–92.8)	100 (98.2–100)
Influenza B	53	149	2	0	100 (98.2–100)	98.7 (96–99.6)	96.4 (92.8–98.2)	100 (98.2–100)
RSV	24	179	1	0	100 (98.2–100)	99.4 (97.2–99.9)	96 (92.4–100)	100 (98.2–100)

RSV: Respiratory syncytial virus; RVP: Respiratory viral panel.