A new approach for the identification and the diagnosis of Eimeria media parasite of the rabbit.
In this work, we described a new approach for the isolation of a species-specific probe for the Eimeria media parasite of the rabbit based on the use of the random amplified polymorphic DNA (RAPD) technique. A specific fragment of 800 bp of the studied species was isolated after RAPD and then cloned and DIG-radiolabeled. After dot-blotting, we observed that this probe was specific for E. media. Sequencing of the 3' and 5' ends of this probe enabled the determination of two primers that could be used in a PCR reaction. The amplified product of 750 bp was specific E. media. The use of these primers and of our probe allowed the detection of a very small number of oocysts. With a new protocol of DNA purification, 10 purified oocysts were detected by PCR. The efficiency of the amplification was not changed when two species were mixed. The threshold of detection of oocysts in fecal matter was equal to 30.
Cere
N
N
INRA, Centre de Tours, Nouzilly, France.
Humbert
J F
JF
Licois
D
D
Corvione
M
M
Afanassieff
M
M
Chanteloup
N
N
D000818:Animals; D001483:Base Sequence; D015139:Blotting, Southern / Q000662:veterinary; D003048:Coccidiosis / Q000175:diagnosis / Q000469:parasitology / Q000662:veterinary*; D017931:DNA Primers / Q000737:chemistry; D015342:DNA Probes / Q000235:genetics; D004539:Eimeria / Q000235:genetics / Q000302:isolation & purification*; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction / Q000662:veterinary; D011817:Rabbits / Q000469:parasitology*; D019105:Random Amplified Polymorphic DNA Technique / Q000662:veterinary; D012680:Sensitivity and Specificity; D013045:Species Specificity
D017931:DNA Primers; D015342:DNA Probes