A High-Performance Multiplex Immunoassay for Serodiagnosis of Flavivirus-Associated Neurological Diseases in Horses

Table 4

Comparison of the identification of circulating flaviviruses in equine field sera by three different techniques: flavivirus ELISA, MIA, and VNT.

Positive; sera	ELISA				MIA					Flavivirus; identification		VNT
	Number of field sera	ELISA; pos.	ELISA; doubt.	ELISA; neg.	WNV.sE; pos.	WNV.EDIII; pos.	JEV.EDIII; pos.	TBEV.EDIII; pos.	Neg.			WNV; pos.	JEV; pos	USUV; pos.	TBEV; pos.	Neg.	Flavivirus; identification
WNV	103: Pak. (68) + Mada. (35)	101	2	0	100	99	53	2	3	96	WNV positive	99	36	9/35; (Mada.)	NA	4	96	WNV positive
										4	Undetermined; flavivirus						3	Undetermined; flavivirus
JEV	101 (Japan)	88	2	11	90	29	87	2	11	82	JEV positive	11	96	NA	NA	5	91	JEV positive
										3	WNV positive						1	WNV positive
										5	Undetermined flavivirus						4	Undetermined Flavivirus
TBEV	74 (Austria)	59	3	12	59	1	7	61	13	60	TBEV positive	3	NA	4	62	12	62	TBEV positive
										1	Undetermined; flavivirus
Negative; sera	160 (Camargue + Ireland)			160	2	16	12	0	158	158	Negative
										2	WNV positive

Not analysed (NA).
Positive sera evaluated as false reactives for JEV or WNV if WNV.sE signal was negative.
Bold italic: negative sample and italic: doubtful sample.
The thresholds for ELISA, MIA, and MNT were as defined in Table 3. In the event of cross-reaction with rEDIII beads during MIA, a horse was considered infected with a specific flavivirus if the corresponding bead coupled to rEDIII generated an S/P ratio at least 1.5-fold greater than that generated with the other rEDIII beads. In the event of cross-reaction with MNT, flavivirus identification is determined by the virus with the highest neutralisation capacity and at least a fourfold difference in titres.