Development and validation of real-time PCR assays specific to four species of Eimeria

Damer Blake (1) , Zonghua Qin , Jianping Cai , Adrian Smith (2)

1 Microbiology
2 Immunology

The development of quantitative real-time PCR assays specific to Eimeria acervulina, Eimeria maxima, Eimeria necatrix and Eimeria tenella is described and validated. PCR templates adopted include a fragment of a gene encoding a microneme protein and previously characterised species-specific RAPD sequences. The sensitivity of each assay allowed the consistent detection of between one and ten parasite genomes, equivalent to between one or two sporulated oocysts or a fraction of a single mature schizont, unaffected by the presence of chicken (host) or other Eimeria species DNA. Regression coefficients in excess of 0.99 over linear ranges of at least six orders of magnitude, together with comparable PCR efficiencies, demonstrated the robust reproducibility of each assay and suggest that two or more may be successfully multiplexed. The species-specific assays described here, combined with a previously published generic Eimeria species real-time PCR, provide valuable components in a ‘tool box' to accurately quantify the presence of specific Eimeria species in environmental or within-host phases of the lifecycle with little specialist knowledge. The application of these assays may benefit chicken husbandry, veterinary practice, quality control of live vaccine production and scientific research.

Mots clés: Life Sciences

Damer Blake, Zonghua Qin, Jianping Cai, Adrian Smith. Development and validation of real-time PCR assays specific to four species of Eimeria. Avian Pathology, 2008, 37 (01), pp.89-94. ⟨10.1080/03079450701802248⟩. ⟨hal-00540107⟩