Detection of Cryptosporidium parvum oocysts in faeces: comparison of conventional coproscopical methods and the polymerase chain reaction.
Conventional and coproscopical methods were compared with the polymerase chain reaction (PCR) for the detection of Cryptosporidium oocysts in bovine faeces. Oocysts were not detected in samples seeded with 10,000 oocysts following formol ether sedimentation and examination using auramine phenol (AP) or by immunofluorescent (IF) staining. When oocysts were concentrated using sucrose flotation the threshold of detection was 4000 oocysts per gram for both staining methods. Following salt flotation 4000 oocysts per gram could be reliably detected by AP staining but the detection limit was increased to 6000 oocysts per gram using IF staining. The recovery of oocysts was significantly less than expected for all techniques. A specific PCR coupled with immunomagnetic particle separation (IMS) of samples detected five oocysts per ml of diluted faeces, which corresponds to 80-90 oocysts per gram. Even allowing for the dilution of formed faecal samples, required for IMS, this represents an increase in sensitivity of several orders of magnitude over the conventional corpodiagnostic methods.
Webster
K A
KA
Central Veterinary Laboratory, New Haw, Addlestone, UK.
Smith
H V
HV
Giles
M
M
Dawson
L
L
Robertson
L J
LJ
D000818:Animals; D001483:Base Sequence; D002417:Cattle; D002469:Cell Separation / Q000379:methods; D002499:Centrifugation, Density Gradient; D016785:Cryptosporidium parvum / Q000302:isolation & purification*; D017931:DNA Primers; D005243:Feces / Q000469:parasitology*; D005455:Fluorescent Antibody Technique; D007202:Indicators and Reagents; D008969:Molecular Sequence Data; D010270:Parasite Egg Count / Q000379:methods / Q000662:veterinary; D016133:Polymerase Chain Reaction / Q000379:methods / Q000662:veterinary*
D017931:DNA Primers; D007202:Indicators and Reagents