An interlaboratory comparison of immunohistochemistry and PCR methods for detection of Neospora caninum in bovine foetal tissues.
Seven European laboratories contributed to a multi-centre evaluation of detection techniques for Neospora caninum in bovine foetuses. Six laboratories participated in immunohistochemistry (IHC) testing. All seven laboratories participated in PCR testing, but the results from one laboratory were not included in the analysis, because of contamination problems in the preparation of the samples. A coded panel of tissue sections from 36 infected and non-infected foetuses was used to evaluate the IHC detection of parasites. A coded panel consisting of 44 homogenized foetal brain samples from natural bovine abortion cases and 32 spiked samples were used to evaluate the PCR methods. Inclusion of a duplicate dilution series of spiked samples was used to evaluate detection limits and repeatability. IHC methods had a relatively low sensitivity, but a high specificity. There was considerable variation in IHC results between participating laboratories, which may be partly explained by examination practices that depended on the experience of the operator. In addition, the use of different antibody reagents, different antibody dilutions, and different enzymatic treatments of tissues may have contributed to the observed variation. PCR methods generally had a higher sensitivity than IHC methods and also a high specificity. The agreement between the majority scores of IHC and PCR methods was low. False positive PCR results indicated contamination problems in some instances. Agreement between the PCR results of the various laboratories was better, compared with the IHC results. There appeared to be no clear relationship between the PCR format (i.e. single or nested) and diagnostic sensitivity. Consequently, an improvement of diagnostic performance of PCR might possibly be achieved by optimizing DNA extraction methods.
van Maanen
C
C
Animal Health Service, P.O. Box 9, NL-7400 AA Deventer, The Netherlands.
Wouda
W
W
Schares
G
G
von Blumröder
D
D
Conraths
F J
FJ
Norton
R
R
Williams
D J L
DJ
Esteban-Redondo
I
I
Innes
E A
EA
Mattsson
J G
JG
Björkman
C
C
Fernández-García
A
A
Ortega-Mora
L M
LM
Müller
N
N
Sager
H
H
Hemphill
A
A
D000034:Abortion, Veterinary / Q000175:diagnosis / Q000469:parasitology; D000818:Animals; D001921:Brain / Q000196:embryology / Q000469:parasitology; D002417:Cattle; D002418:Cattle Diseases / Q000175:diagnosis / Q000469:parasitology; D003048:Coccidiosis / Q000175:diagnosis / Q000662:veterinary; D005189:False Positive Reactions; D005333:Fetus / Q000469:parasitology; D007150:Immunohistochemistry / Q000379:methods / Q000592:standards / Q000662:veterinary*; D007753:Laboratories / Q000592:standards*; D018688:Neospora / Q000302:isolation & purification*; D016133:Polymerase Chain Reaction / Q000379:methods / Q000592:standards / Q000662:veterinary*; D012680:Sensitivity and Specificity