Development of nested duplex PCR assays for detection of pathogen Eimeria species in cattle in Papua, Indonesia

• Published in: IOP conference series. Earth and environmental science 2022-02, Vol.976 (1), p.12008
• Main Authors: Ekawasti, F, Nashrulloh, M F, Nurcahyo, R W, Priyowidodo, D, Prastowo, J, Firdausy, L W
• Format: Article
• Language: English
• Subjects: Calves; Cattle; Deoxyribonucleic acid; Diarrhea; DNA; Eimeria; gastrointestinal; Hemorrhage; moleculer; Oocysts; Polymerase chain reaction; protozoa; Research facilities; Research institutions; Species
• ISSN: 1755-1307; 1755-1315
• DOI: 10.1088/1755-1315/976/1/012008

Eimeria bovis and Eimeria zuernii are highly pathogenic Eimeria species in cattle that are the most prevalent causes of a severe clinical illness characterized by hemorrhagic diarrhea in calves and young cattle with potentially fatal effects over the world. The oocysts of a handful of the known bovine eimeriosis species are difficult to distinguish morphologically. For the specific differentiating evidence of Eimeria species, symptomatic research institutions are increasingly relying on DNA-based technologies. This research offers a duplex polymerase chain reaction (PCR) test based on the internal transcribed spacer-1 (ITS-1) gene that may be used to diagnose E. bovis and E. zuernii in cattle from various locations at the same time. The oocysts were concentrated and purified using a fecal harvesting method. The genomic DNA is extracted according to the instructions included with the kit. Primer pairs specific to each species, as well as a standard optimum annealing temperature of 55°C for these species, were discovered. The samples were amplified in a homogenous way, resulting in a homogeneous band ladder, revealing that the test could distinguish between two highly pathogenic Eimeria species in one tube reaction. This duplex PCR assay can detect a high pathogenic bovine Eimeria simultaneously in a rapid and low cost.