title: First molecular characterization of Anaplasma marginale in cattle and Rhipicephalus (Boophilus) microplus ticks in Cebu, Philippines.
abstract: Anaplasma marginale has been detected in the Philippines only by peripheral blood smear examination and serological methods. This study generally aimed to molecularly detect and characterize A. marginale in cattle and ticks in Cebu, Philippines. A total of 12 bovine blood samples and 60 Rhipicephalus (Boophilus) microplus ticks were collected on the Cebu Island in 2011. 16S rRNA-based screening-PCR and DNA sequencing revealed 8 cattle (66.7%) and 8 ticks (13.3%) to be positive for A. marginale, and 1 tick (1.7%) to be positive for A. centrale. Selected positive DNA samples were further characterized based on 16S rRNA (longer sequence), Msp5, Msp1α,gltA and groEL genes for phylogenetic analyses. Sequence identities of partial DNA fragments of A. marginale from the Philippines revealed 99.1-100% (16S rRNA, gltA, groEL and Msp5) and 94.3-97.6% (Msp1α) identities to the closest isolates from other countries. Moreover, sequence analysis of the Msp1α, gene showed 3 variants, including a case of co-infection with 2 variants. Phylogenetic analyses based on Msp1α and Msp5 genes revealed that Philippine A. marginale isolates formed a monophyletic lineage, and were phylogenetically related to Brazilian and Chinese isolates. On the other hand, a highly specific and sensitive nested PCR based on groEL, with a detection limit of 2 copies/PCR, was developed to detect A. marginale in the Philippines. This study reported the first molecular detection and characterization of A. marginale in cattle and R. microplus in Cebu, Philippines.
authors.affiliation: Department of Veterinary Clinical Sciences, Obihiro University of Agriculture and Veterinary Medicine, Obihiro, Hokkaido 080-8555, Japan.
authors.forename: Adrian Patalinghug
authors.initials: AP
authors.lastname: Ybañez
authors.forename: Thillaiampalam
authors.initials: T
authors.lastname: Sivakumar
authors.forename: Rochelle Haidee Daclan
authors.initials: RH
authors.lastname: Ybañez
authors.forename: Jowarren Catingan
authors.initials: JC
authors.lastname: Ratilla
authors.forename: Zandro Obligado
authors.initials: ZO
authors.lastname: Perez
authors.forename: Shirleny Reyes
authors.initials: SR
authors.lastname: Gabotero
authors.forename: Hassan
authors.initials: H
authors.lastname: Hakimi
authors.forename: Shin-ichiro
authors.initials: S
authors.lastname: Kawazu
authors.forename: Kotaro
authors.initials: K
authors.lastname: Matsumoto
authors.forename: Naoaki
authors.initials: N
authors.lastname: Yokoyama
authors.forename: Hisashi
authors.initials: H
authors.lastname: Inokuma
mesh_terms: D042322:Anaplasma marginale / Q000235:genetics*; D000712:Anaplasmosis / Q000453:epidemiology*; D000818:Animals; D001483:Base Sequence; D002417:Cattle; D002418:Cattle Diseases / Q000453:epidemiology* / Q000382:microbiology* / Q000469:parasitology*; D017931:DNA Primers / Q000235:genetics; D008969:Molecular Sequence Data; D010679:Philippines / Q000453:epidemiology; D010802:Phylogeny; D012336:RNA, Ribosomal, 16S / Q000235:genetics; D048494:Rhipicephalus / Q000382:microbiology*; D017422:Sequence Analysis, DNA; D013045:Species Specificity; D013984:Tick Infestations / Q000453:epidemiology / Q000662:veterinary*
chemical_list: D017931:DNA Primers; D012336:RNA, Ribosomal, 16S