Development of a Molecular Assay for the Determination of Eimeria tenella Oocyst Viability

Coccidiosis is a cosmopolitan disease with major impacts on the commercial poultry industry. The causative agents, Apicomplexan parasites of the genus Eimeria , infect epithelial cells of the intestine causing diarrhea, secondary infections, and negatively impacting meat or egg production. The infective parasite stage, the oocyst, is shed in feces and must sporulate over several days in the environment to reach infectivity. The number of viable, sporulated oocysts ingested is proportional to the severity of disease. The self-limiting and highly immunogenic nature of infection by Eimeria spp. make live vaccination an effective means of coccidiosis control. High prevalence of drug resistance and consumer demand for poultry products with no “human medically important antibiotics” or raised without antibiotics indicates an increasing role for vaccination in the control of coccidiosis going forward. Paramount to vaccine efficacy is the ability to administer precise numbers of viable oocysts to stimulate the development of immunity without causing disease. Unfortunately, no rapid and accurate method for determination of oocyst viability is presently available. Time-consuming and expensive live-infection trials are the current gold standard. Work completed for this study demonstrated the development of a molecular assay for the determination of Eimeria tenella Tyzzer, 1929 oocyst viability. The assay used the quantification of specific biomolecules to assess changes in gene expression in response to heat stimulation that indicates viability of a sample of oocysts. Data showed strong predictive value for infectivity of an oocyst sample as confirmed by oocyst output (fecundity) in infection trials. Use of the assay would be low cost compared to the planning, resources, execution, and data collection required for infection trials. Additionally, the assay was shown to offer results in hours versus days for the live infection trials. Development of this first-generation assay paves the way for further development of assays for the determination of viability in mixed species samples (multivalent vaccines), for vaccine quality control at hatcheries, and with application in environmental monitoring (“early warning” programs) and on-farm diagnostics.