Polymerase chain reaction directed to Eimeria ITS1 rDNA or single copy orthologues corroborates standard micro-oocyst analysis of intestinal tissue from E. acervulina, E. maxima, or E. tenella-infected chickens

Authors: Jenkins, Mark; Obrien, Celia; Parker, Carolyn; Thompson, Peter; FITZCOY, STEVE - Merck Animal Health; BAUTISTA, DANIEL - Zoetis

Submitted to: Avian Diseases
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 4/28/2022
Publication Date: 7/1/2022
Citation: Jenkins, M.C., Obrien, C.N., Parker, C.C., Thompson, P.C., Fitzcoy, S., Bautista, D. 2022. Polymerase chain reaction directed to Eimeria ITS1 rDNA or single copy orthologues corroborates standard micro-oocyst analysis of intestinal tissue from E. acervulina, E. maxima, or E. tenella-infected chickens. Avian Diseases. 66(2). https://doi.org/10.1637/aviandiseases-D-22-00001.
DOI: https://doi.org/10.1637/aviandiseases-D-22-00001

Interpretive Summary: Avian coccidiosis is an intestinal disease causing over $ 10 billion annual loss to the worldwide poultry industry. The disease is caused by protozoa in the genus Eimeria. Diagnosis of avian coccidiosis usually relies on the submission of moribund chickens to a diagnostic laboratory. A sample of intestinal content is placed on a glass slide and then examined under the microscope for the presence of Eimeria parasites. This technique is laborious and requires a trained microscopist. The present paper describes a molecular technique that examines DNA from the gut scraping for the presence of Eimeria DNA that can be detected using a method called polymerase chain reaction (PCR). PCR was shown to be useful for detecting the parasite in both experimental and natural Eimeria infections and showed a high correlation with standard micro-oocyst counting. The molecular method is objective and does not require a highly trained microscopist to diagnose Eimeria infection. This method should give timely and accurate insight to poultry growers and companies in combatting avian coccidiosis.

Technical Abstract: The purpose of this study was to compare micro-oocyst counts of Eimeria to PCR analysis of intestinal DNA from smears of duodenum, jejunum/ileum, and cecum of chickens infected with E. acervulina, E. maxima, or E. tenella oocysts. Broiler chicks were infected in triplicate with various doses of E. acervulina, E. maxima, or E. tenella oocysts and were necropsied 5-6 days later to recover duodenal, jejunal, or cecal tissue for micro-oocyst count and for DNA recovery. Micro-oocyst counts were done independently by 3 individuals. Micro-oocyst counts and PCR directed to ITS1-rDNA or 2 different single copy orthologues (SCO 4141 and SCO 5995) displayed a linear relationship with oocyst dose for each Eimeria species. A strong correlation was found between mean micro-oocysts counts and all 3 PCR assays for E. acervulina (r = 0.78 - 0.94), E. maxima (r = 0.79 - 0.98), and E. tenella (r = 0.85 - 0.96). There was excellent agreement among different PCR assays- E. acervulina (r = 0.88 - 0.98), E. maxima (r = 0.75 - 0.95), and E. tenella (r = 0.93 - 0.98). Micro-oocyst counts also corroborated findings from ITS1 analysis of DNA recovered from Eimeria-infected broiler chickens submitted to a poultry diagnostic laboratory. These findings indicate that ITS1 PCR and SCO PCR can supplant traditional micro-oocyst counts used in the diagnosis of Eimeria infection in chickens and may provide for an estimate of the relative abundance of each Eimeria species in a natural infection.