title: Characterization and annotation of Babesia orientalis apicoplast genome.
abstract: BACKGROUND
  Babesia orientalis is an obligate intraerythrocytic protozoan parasite of the buffalo (Bubalus bubalis, Linnaeus, 1758) transmitted by the tick Rhipicephalus heamaphysaloides. It is the causative agent of water buffalo babesiosis, one of the most important pathogens of water buffalo in central and southern China. As a member of the phylum Apicomplexa, B. orientalis possesses a relatively independent and alga originated organelle the apicoplast. Apicoplasts in other apicomplexa parasites are involved in the biosynthesis of haem, fatty acids, iron-sulphur clusters and isoprenoids. Some of these metabolic pathways were shown to be essential for parasite survival, therefore can serve as potential drug targets.
  
  
  METHODS
  30 pairs of primers were designed based on the full genome sequence of B. orientalis (unpublished data) and by aligning reported apicoplast genomes of Babesia bovis and Theileria parva. Conventional PCRs was performed to obtain overlapped fragments to cover the whole apicoplast genome. Then the apicoplast genome of B.orientalis was sequenced, assembled and aligned with reported apicoplast genomes of B. bovis and T. parva. The obtained apicoplast genome was annotated by using Artemis and comparing with published apicomplexan apicoplast genomes. The SSU and LSU nucleotide sequences generated were used in a phylogenetic analysis using the maximum likelihood implemented in MAGE 6.0.
  
  
  RESULTS
  We have obtained and analyzed the complete genome sequence of the B. orientalis apicoplast. It consisted of a 33.2 kb circular DNA (78.9 % A + T). The apicoplast genome unidirectionally encodes one large and one small subunit ribosomal RNAs, 24 tRNA genes, 4 DNA-dependent RNA polymerase beta subunits (rpoB, rpoC1, rpoC2a and rpoC2b), 17 ribosomal proteins, one EF-Tu elongation factor, 2 Clp protease chaperones, and 14 hypothetical proteins. In addition, it includes two copies of the clpC gene. The structure and organization of the B. orientalis apicoplast genome are most similar to those of the B. bovis apicoplast.
  
  
  CONCLUSIONS
  This is the first report of the complete sequence of the B. orientalis apicoplast genome. This information should be useful in the development of safe and efficient treatment against buffalo babesiosis.
authors.affiliation: State Key Laboratory of Agricultural Microbiology, College of Veterinary Medicine, Huazhong Agricultural University, Wuhan, 430070, Hubei, P.R. China. 543612282@qq.com.
authors.forename: Yuan
authors.initials: Y
authors.lastname: Huang
authors.affiliation: State Key Laboratory of Agricultural Microbiology, College of Veterinary Medicine, Huazhong Agricultural University, Wuhan, 430070, Hubei, P.R. China. helan@mail.hzau.edu.cn.|Key Laboratory of Animal Epidemical Disease and Infectious Zoonoses, Ministry of Agriculture, Huazhong Agricultural University, Wuhan, 430070, Hubei, China. helan@mail.hzau.edu.cn.
authors.forename: Lan
authors.initials: L
authors.lastname: He
authors.affiliation: State Key Laboratory of Agricultural Microbiology, College of Veterinary Medicine, Huazhong Agricultural University, Wuhan, 430070, Hubei, P.R. China. 1057030377@qq.com.
authors.forename: Jinfang
authors.initials: J
authors.lastname: Hu
authors.affiliation: State Key Laboratory of Agricultural Microbiology, College of Veterinary Medicine, Huazhong Agricultural University, Wuhan, 430070, Hubei, P.R. China. 1415144593@qq.com.
authors.forename: Pei
authors.initials: P
authors.lastname: He
authors.affiliation: State Key Laboratory of Agricultural Microbiology, College of Veterinary Medicine, Huazhong Agricultural University, Wuhan, 430070, Hubei, P.R. China. 810056241@qq.com.
authors.forename: Junwei
authors.initials: J
authors.lastname: He
authors.affiliation: State Key Laboratory of Agricultural Microbiology, College of Veterinary Medicine, Huazhong Agricultural University, Wuhan, 430070, Hubei, P.R. China. 399742372@qq.com.
authors.forename: Long
authors.initials: L
authors.lastname: Yu
authors.affiliation: State Key Laboratory of Agricultural Microbiology, College of Veterinary Medicine, Huazhong Agricultural University, Wuhan, 430070, Hubei, P.R. China. 2376496022@qq.com.
authors.forename: Ngabu
authors.initials: N
authors.lastname: Malobi
authors.affiliation: State Key Laboratory of Agricultural Microbiology, College of Veterinary Medicine, Huazhong Agricultural University, Wuhan, 430070, Hubei, P.R. China. 46176322@qq.com.|Key Laboratory of Animal Epidemical Disease and Infectious Zoonoses, Ministry of Agriculture, Huazhong Agricultural University, Wuhan, 430070, Hubei, China. 46176322@qq.com.
authors.forename: Yanqin
authors.initials: Y
authors.lastname: Zhou
authors.affiliation: State Key Laboratory of Agricultural Microbiology, College of Veterinary Medicine, Huazhong Agricultural University, Wuhan, 430070, Hubei, P.R. China. 2088657959@qq.com.|Key Laboratory of Animal Epidemical Disease and Infectious Zoonoses, Ministry of Agriculture, Huazhong Agricultural University, Wuhan, 430070, Hubei, China. 2088657959@qq.com.
authors.forename: Bang
authors.initials: B
authors.lastname: Shen
authors.affiliation: State Key Laboratory of Agricultural Microbiology, College of Veterinary Medicine, Huazhong Agricultural University, Wuhan, 430070, Hubei, P.R. China. zhaojunlong@mail.hzau.edu.cn.|Key Laboratory of Animal Epidemical Disease and Infectious Zoonoses, Ministry of Agriculture, Huazhong Agricultural University, Wuhan, 430070, Hubei, China. zhaojunlong@mail.hzau.edu.cn.
authors.forename: JunLong
authors.initials: J
authors.lastname: Zhao
mesh_terms: D000818:Animals; D063546:Apicoplasts / Q000235:genetics*; D001403:Babesia / Q000235:genetics*; D002020:Buffaloes / Q000469:parasitology; D002681:China; D016000:Cluster Analysis; D016054:DNA, Protozoan / Q000737:chemistry / Q000235:genetics; D004275:DNA, Ribosomal / Q000737:chemistry / Q000235:genetics; D016678:Genome; D058977:Molecular Sequence Annotation; D008969:Molecular Sequence Data; D010802:Phylogeny; D016133:Polymerase Chain Reaction; D012335:RNA, Ribosomal / Q000235:genetics; D012337:RNA, Ribosomal, 18S / Q000235:genetics; D017422:Sequence Analysis, DNA
chemical_list: D016054:DNA, Protozoan; D004275:DNA, Ribosomal; D012335:RNA, Ribosomal; D012337:RNA, Ribosomal, 18S; C065096:RNA, ribosomal, 26S