title: Molecular evidence of infections with Babesia gibsoni parasites in Japan and evaluation of the diagnostic potential of a loop-mediated isothermal amplification method.
abstract: Detection and analysis of Babesia gibsoni infection were performed with whole-blood samples collected between July 2002 and July 2003 from 945 and 137 dogs from the Aomori and Okinawa Prefectures of Japan, respectively, by PCR and loop-mediated isothermal amplification (LAMP). On the basis of the criterion for positivity by PCR, 3.9% (37 of 945) and 10.9% (15 of 137) of the dogs had B. gibsoni DNA. All 37 positive animals from Aomori Prefecture were male Tosa dogs (Japanese mastiff). The 15 dogs from Okinawa Prefecture with positive PCR assay results were of various breeds, ages, and sexes. The 18S ribosomal DNA (18S rDNA) sequences from all samples showed 100% homology to each other and to published B. gibsoni sequences. The limits of detection of B. gibsoni parasitemia by the PCR and LAMP methods with an 18S rDNA-based primer set were 0.0005% each. A comparison of the PCR and LAMP methods with microscopic examination for the detection of B. gibsoni infections in blood samples from 945 field dogs in Aomori Prefecture and 137 field dogs in Okinawa Prefecture showed that 37 and 15 dogs, respectively, were positive by the PCR and LAMP methods and that 16 and 12 dogs, respectively, were positive by light microscopic examination. All samples found to be positive by microscopic examination were also positive by the PCR and LAMP methods. The results of the PCR and LAMP methods agreed for samples with positive results by either method. Moreover, nonspecific reactions were not observed by the LAMP method. These results suggest that the LAMP method provides a useful tool for the detection of B. gibsoni infections in dogs.
authors.affiliation: Department of Veterinary Parasitology, School of Veterinary Medicine and Animal Sciences, Kitasato University, Towada, Aomori 034-8628, Japan. ikadai@vmas.kitasato-u.ac.jp
authors.forename: Hiromi
authors.initials: H
authors.lastname: Ikadai
authors.forename: Hiroko
authors.initials: H
authors.lastname: Tanaka
authors.forename: Nona
authors.initials: N
authors.lastname: Shibahara
authors.forename: Aya
authors.initials: A
authors.lastname: Matsuu
authors.forename: Masami
authors.initials: M
authors.lastname: Uechi
authors.forename: Naoyuki
authors.initials: N
authors.lastname: Itoh
authors.forename: Sugao
authors.initials: S
authors.lastname: Oshiro
authors.forename: Noboru
authors.initials: N
authors.lastname: Kudo
authors.forename: Ikuo
authors.initials: I
authors.lastname: Igarashi
authors.forename: Takashi
authors.initials: T
authors.lastname: Oyamada
mesh_terms: D000818:Animals; D001403:Babesia / Q000302:isolation & purification; D001404:Babesiosis / Q000175:diagnosis* / Q000662:veterinary*; D004283:Dog Diseases / Q000175:diagnosis*; D004285:Dogs; D021141:Nucleic Acid Amplification Techniques / Q000379:methods*; D016133:Polymerase Chain Reaction; D017422:Sequence Analysis, DNA