Development of loop-mediated isothermal amplification (LAMP) method for diagnosis of equine piroplasmosis.
Loop-mediated isothermal amplification (LAMP) is a novel nucleic acid method whereby DNA is amplified with high specificity, efficiency, and rapidity under isothermal conditions using a set of four specifically designed primers and a DNA polymerase with strand displacement activity. In this study, we used LAMP primer sets designed from EMA-1 and Bc 48 genes for detection of Theileria equi and Babesia caballi infections, respectively. These primer sets specifically amplified DNA of the respective parasites. Both primer sets amplified T. equi and B. caballi up to 10(-6) dilution of 10-fold serially diluted samples. Furthermore, DNA extracted from blood collected from a horse experimentally infected with T. equi was amplified by a T. equi LAMP primer set from days 2 to 35 post-infection, demonstrating the high sensitivity of these primers. Of 55 samples collected from China, 81.8% and 56.3% were positively detected by LAMP for T. equi and B. caballi infections, respectively. In contrast, 91.8% and 45.9% of the 37 samples collected from South Africa were LAMP positive for T. equi and B. caballi, respectively. These results suggest that LAMP could be a potential diagnostic tool for epidemiological studies of equine piroplasmosis.
Alhassan
Andy
A
National Research Center for Protozoan Diseases, Obihiro University of Agriculture and Veterinary Medicine, Obihiro, Hokkaido 080-8555, Japan.
Thekisoe
Oriel M M
OM
Yokoyama
Naoaki
N
Inoue
Noboru
N
Motloang
Makhosazana Y
MY
Mbati
Peter A
PA
Yin
Hong
H
Katayama
Yoshinari
Y
Anzai
Toru
T
Sugimoto
Chihiro
C
Igarashi
Ikuo
I
D000818:Animals; D001403:Babesia / Q000302:isolation & purification; D001404:Babesiosis / Q000175:diagnosis / Q000662:veterinary*; D017931:DNA Primers / Q000737:chemistry; D016054:DNA, Protozoan / Q000737:chemistry*; D003937:Diagnosis, Differential; D006734:Horse Diseases / Q000175:diagnosis*; D006736:Horses; D021141:Nucleic Acid Amplification Techniques / Q000379:methods / Q000592:standards / Q000662:veterinary*; D016133:Polymerase Chain Reaction / Q000379:methods / Q000592:standards / Q000662:veterinary; D012680:Sensitivity and Specificity; D013045:Species Specificity; D016794:Theileria / Q000302:isolation & purification; D013801:Theileriasis / Q000175:diagnosis*; D017282:Tick-Borne Diseases / Q000175:diagnosis / Q000662:veterinary*
D017931:DNA Primers; D016054:DNA, Protozoan