Optimization of a Fluorescence-Based Assay for Large-Scale Drug Screening against Babesia and Theileria Parasites.
A rapid and accurate assay for evaluating antibabesial drugs on a large scale is required for the discovery of novel chemotherapeutic agents against Babesia parasites. In the current study, we evaluated the usefulness of a fluorescence-based assay for determining the efficacies of antibabesial compounds against bovine and equine hemoparasites in in vitro cultures. Three different hematocrits (HCTs; 2.5%, 5%, and 10%) were used without daily replacement of the medium. The results of a high-throughput screening assay revealed that the best HCT was 2.5% for bovine Babesia parasites and 5% for equine Babesia and Theileria parasites. The IC50 values of diminazene aceturate obtained by fluorescence and microscopy did not differ significantly. Likewise, the IC50 values of luteolin, pyronaridine tetraphosphate, nimbolide, gedunin, and enoxacin did not differ between the two methods. In conclusion, our fluorescence-based assay uses low HCT and does not require daily replacement of culture medium, making it highly suitable for in vitro large-scale drug screening against Babesia and Theileria parasites that infect cattle and horses.
National Research Center for Protozoan Diseases, Obihiro University of Agriculture and Veterinary Medicine, Inada-Cho, Obihiro, Hokkaido, Japan; Department of Internal Medicine and Infectious Diseases, Faculty of Veterinary Medicine, Mansoura University, Mansoura, Egypt.
Mohamed Abdo
MA
Rizk
National Research Center for Protozoan Diseases, Obihiro University of Agriculture and Veterinary Medicine, Inada-Cho, Obihiro, Hokkaido, Japan; Department of Biochemistry and Chemistry of Nutrition, Faculty of Veterinary Medicine, Mansoura University, Mansoura, Egypt.
Shimaa Abd El-Salam
SA
El-Sayed
National Research Center for Protozoan Diseases, Obihiro University of Agriculture and Veterinary Medicine, Inada-Cho, Obihiro, Hokkaido, Japan.
Mohamed Alaa
MA
Terkawi
Department of Internal Medicine and Infectious Diseases, Faculty of Veterinary Medicine, Mansoura University, Mansoura, Egypt.
Mohamed Ahmed
MA
Youssef
Department of Biochemistry and Chemistry of Nutrition, Faculty of Veterinary Medicine, Mansoura University, Mansoura, Egypt.
El Said El Shirbini
el Sel S
El Said
Department of Biochemistry and Chemistry of Nutrition, Faculty of Veterinary Medicine, Mansoura University, Mansoura, Egypt.
Gehad
G
Elsayed
Department of Internal Medicine and Infectious Diseases, Faculty of Veterinary Medicine, Mansoura University, Mansoura, Egypt.
Sabry
S
El-Khodery
Department of Internal Medicine and Infectious Diseases, Faculty of Veterinary Medicine, Mansoura University, Mansoura, Egypt.
Maged
M
El-Ashker
National Research Center for Protozoan Diseases, Obihiro University of Agriculture and Veterinary Medicine, Inada-Cho, Obihiro, Hokkaido, Japan; Department of Animal Medicine and Infectious Diseases, Faculty of Veterinary Medicine, Minoufiya University, Sadat City, Minoufiya, Egypt.
Ahmed
A
Elsify
Department of Parasitology, Faculty of Veterinary Medicine, South Valley University, Qena, Egypt.
Mosaab
M
Omar
Department of Animal Medicine and Infectious Diseases, Faculty of Veterinary Medicine, Minoufiya University, Sadat City, Minoufiya, Egypt.
Akram
A
Salama
National Research Center for Protozoan Diseases, Obihiro University of Agriculture and Veterinary Medicine, Inada-Cho, Obihiro, Hokkaido, Japan.
Naoaki
N
Yokoyama
National Research Center for Protozoan Diseases, Obihiro University of Agriculture and Veterinary Medicine, Inada-Cho, Obihiro, Hokkaido, Japan.
Ikuo
I
Igarashi
D000818:Animals; D000977:Antiparasitic Agents / Q000737:chemistry / Q000494:pharmacology*; D001403:Babesia / Q000187:drug effects* / Q000254:growth & development; D002417:Cattle / Q000469:parasitology; D002478:Cells, Cultured; D004353:Drug Evaluation, Preclinical / Q000379:methods*; D006400:Hematocrit; D006736:Horses / Q000469:parasitology; D008856:Microscopy, Fluorescence; D016794:Theileria / Q000187:drug effects* / Q000254:growth & development
D000977:Antiparasitic Agents