title: Genetic diversity and drug sensitivity studies on Eimeria tenella field isolates from Hubei Province of China.
abstract: BACKGROUND
  Avian coccidiosis is an intracellular intestinal parasitic disease, caused by intracellular intestinal parasites from the genus Eimeria, among which Eimeria tenella is one of the most pathogenic species and causes great economic losses. Frequent applications of anticoccidial drugs have resulted in the development of drug-resistance in E. tenella. In the present study, we sought to determine the genetic diversity of E. tenella isolates prevalent in chicken farms in Hubei Province of China and examine their sensitivity to three anticoccidial drugs. The results provide useful information for the prevention and control of coccidiosis in this region.
  
  
  METHODS
  Eimeria tenella oocysts were isolated from faecal samples collected from different commercial broiler production farms in Hubei Province, China. After oocyst sporulation and animal inoculation for expansion of the field isolates, DNA and RNA were extracted from excysted sporozoites for molecular characterization. Species identification of field isolates were performed by polymerase chain reaction (PCR) amplification of the internal transcribed spacer 1 (ITS1) region of ribosomal DNA. Random amplified polymorphic DNA (RAPD) was used for population genetic analysis. Subsequently, sequences of the major sporozoite surface antigen (SAG), micronemal protein 2 (MIC-2) and cytochrome b (cytb) genes from genomic DNA, and the Eimeria tenella cation-transport ATPase (EtCat ATPase) gene from cDNA were obtained for genotyping using multi-sequence alignments. Finally, sensitivity of the field isolates to three commonly used anticoccidial drugs (diclazuril, decoquinate and maduramycin) were tested to assess the prevalence of drug resistance in E. tenella in Hubei Province of China.
  
  
  RESULTS
  Analysis of the ITS1 sequences indicated that all the isolates were E. tenella. RAPD analysis and multi-sequence alignments of the SAG, MIC-2, EtCat ATPase and cytb showed genetic diversity among these isolates. Finally, drug sensitivity tests demonstrated that all field isolates were sensitive to diclazuril but resistant to decoquinate (except for the isolates from eastern Hubei) and maduramicin.
  
  
  CONCLUSIONS
  Population genetic analysis indicated that genetic polymorphisms among field isolates were closely related with their regional distributions. Drug sensitivity testing demonstrated that E. tenella isolates in Hubei Province were sensitive to diclazuril, but resistant to maduramycin and decoquinate. The results presented here provide important information for the control and preventions of coccidiosis in the Hubei Province of China.
authors.affiliation: State Key Laboratory of Agricultural Microbiology, College of Veterinary Medicine, Huazhong Agricultural University, Wuhan, 430070, Hubei, People's Republic of China.
authors.forename: Li
authors.initials: L
authors.lastname: Tan
authors.affiliation: State Key Laboratory of Agricultural Microbiology, College of Veterinary Medicine, Huazhong Agricultural University, Wuhan, 430070, Hubei, People's Republic of China.
authors.forename: Yalin
authors.initials: Y
authors.lastname: Li
authors.affiliation: State Key Laboratory of Agricultural Microbiology, College of Veterinary Medicine, Huazhong Agricultural University, Wuhan, 430070, Hubei, People's Republic of China.
authors.forename: Xin
authors.initials: X
authors.lastname: Yang
authors.affiliation: State Key Laboratory of Agricultural Microbiology, College of Veterinary Medicine, Huazhong Agricultural University, Wuhan, 430070, Hubei, People's Republic of China.
authors.forename: Qiyun
authors.initials: Q
authors.lastname: Ke
authors.affiliation: State Key Laboratory of Agricultural Microbiology, College of Veterinary Medicine, Huazhong Agricultural University, Wuhan, 430070, Hubei, People's Republic of China.
authors.forename: Weiqiang
authors.initials: W
authors.lastname: Lei
authors.affiliation: State Key Laboratory of Agricultural Microbiology, College of Veterinary Medicine, Huazhong Agricultural University, Wuhan, 430070, Hubei, People's Republic of China.
authors.forename: Mudassar Niaz
authors.initials: MN
authors.lastname: Mughal
authors.affiliation: State Key Laboratory of Agricultural Microbiology, College of Veterinary Medicine, Huazhong Agricultural University, Wuhan, 430070, Hubei, People's Republic of China. fangrui19810705@163.com.
authors.forename: Rui
authors.initials: R
authors.lastname: Fang
authors.affiliation: State Key Laboratory of Agricultural Microbiology, College of Veterinary Medicine, Huazhong Agricultural University, Wuhan, 430070, Hubei, People's Republic of China.
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, People's Republic of China.
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, People's Republic of China. zhaojunlong@mail.hzau.edu.cn.
authors.forename: Junlong
authors.initials: J
authors.lastname: Zhao
mesh_terms: D000818:Animals; D000981:Antiprotozoal Agents / Q000494:pharmacology*; D002645:Chickens / Q000469:parasitology*; D002681:China / Q000453:epidemiology; D003048:Coccidiosis / Q000453:epidemiology / Q000469:parasitology / Q000662:veterinary*; D021901:DNA, Intergenic / Q000235:genetics; D004351:Drug Resistance; D016786:Eimeria tenella / Q000187:drug effects* / Q000235:genetics*; D014644:Genetic Variation; D010802:Phylogeny
chemical_list: D000981:Antiprotozoal Agents; D021901:DNA, Intergenic
keywords: Decoquinate; Diclazuril; Eimeria tenella; Genetic diversity; Maduramycin