title: GeXP analyzer-based multiplex reverse-transcription PCR assay for the simultaneous detection and differentiation of eleven duck viruses.
abstract: BACKGROUND
  Duck viral pathogens primarily include the avian influenza virus (AIV) subtypes H5, H7, and H9; duck hepatitis virus (DHV); duck tembusu virus (DTMUV); egg drop syndrome virus (EDSV); duck enteritis virus (DEV); Newcastle disease virus (NDV); duck circovirus (DuCV); muscovy duck reovirus (MDRV); and muscovy duck parvovirus (MDPV). These pathogens cause great economic losses to China's duck breeding industry.
  
  
  RESULTS
  A rapid, specific, sensitive and high-throughput GeXP-based multiplex PCR assay consisting of chimeric primer-based PCR amplification with fluorescent labeling and capillary electrophoresis separation was developed and optimized to simultaneously detect these eleven viral pathogens. Single and mixed pathogen cDNA/DNA templates were used to evaluate the specificity of the GeXP-multiplex assay. Corresponding specific DNA products were amplified from each pathogen. Other pathogens, including duck Escherichia coli, duck Salmonella, duck Staphylococcus aureus, Pasteurella multocida, infectious bronchitis virus, and Mycoplasma gallisepticum, did not result in amplification products. The detection limit of GeXP was 10(3)copies when all twelve pre-mixed plasmids containing the target genes of eleven types of duck viruses were present. To further evaluate the reliability of GeXP, 150 clinical field samples were evaluated. Comparison with the results of conventional PCR methods for the field samples, the GeXP-multiplex PCR method was more sensitive and accurate.
  
  
  CONCLUSIONS
  This GeXP-based multiplex PCR method can be utilized for the rapid differential diagnosis of clinical samples as an effective tool to prevent and control duck viruses with similar clinical symptoms.
authors.affiliation: Department of Biotechnology, Guangxi Key Laboratory of Animal Vaccines and Diagnostics, Guangxi Veterinary Research Institute, 51 Youai North Road, Nanning, 530001, China. zhangyanfang409@126.com.
authors.forename: Yan-Fang
authors.initials: YF
authors.lastname: Zhang
authors.affiliation: Department of Biotechnology, Guangxi Key Laboratory of Animal Vaccines and Diagnostics, Guangxi Veterinary Research Institute, 51 Youai North Road, Nanning, 530001, China. xiezhixun@126.com.
authors.forename: Zhi-Xun
authors.initials: ZX
authors.lastname: Xie
authors.affiliation: Department of Biotechnology, Guangxi Key Laboratory of Animal Vaccines and Diagnostics, Guangxi Veterinary Research Institute, 51 Youai North Road, Nanning, 530001, China. xie3120371@163.com.
authors.forename: Li-Ji
authors.initials: LJ
authors.lastname: Xie
authors.affiliation: Department of Biotechnology, Guangxi Key Laboratory of Animal Vaccines and Diagnostics, Guangxi Veterinary Research Institute, 51 Youai North Road, Nanning, 530001, China. dengxianwen@126.com.
authors.forename: Xian-Wen
authors.initials: XW
authors.lastname: Deng
authors.affiliation: Department of Biotechnology, Guangxi Key Laboratory of Animal Vaccines and Diagnostics, Guangxi Veterinary Research Institute, 51 Youai North Road, Nanning, 530001, China. xzqman2002@sina.com.
authors.forename: Zhi-Qin
authors.initials: ZQ
authors.lastname: Xie
authors.affiliation: Department of Biotechnology, Guangxi Key Laboratory of Animal Vaccines and Diagnostics, Guangxi Veterinary Research Institute, 51 Youai North Road, Nanning, 530001, China. 2004-luosisi@163.com.
authors.forename: Si-Si
authors.initials: SS
authors.lastname: Luo
authors.affiliation: Department of Biotechnology, Guangxi Key Laboratory of Animal Vaccines and Diagnostics, Guangxi Veterinary Research Institute, 51 Youai North Road, Nanning, 530001, China. lhuang405@126.com.
authors.forename: Li
authors.initials: L
authors.lastname: Huang
authors.affiliation: Department of Biotechnology, Guangxi Key Laboratory of Animal Vaccines and Diagnostics, Guangxi Veterinary Research Institute, 51 Youai North Road, Nanning, 530001, China. huangjiaoling728@126.com.
authors.forename: Jiao-Ling
authors.initials: JL
authors.lastname: Huang
authors.affiliation: Department of Biotechnology, Guangxi Key Laboratory of Animal Vaccines and Diagnostics, Guangxi Veterinary Research Institute, 51 Youai North Road, Nanning, 530001, China. tingtingzeng1986@163.com.
authors.forename: Ting-Ting
authors.initials: TT
authors.lastname: Zeng
mesh_terms: D000818:Animals; D017925:Circovirus / Q000235:genetics / Q000302:isolation & purification; D003937:Diagnosis, Differential; D004372:Ducks; D009980:Influenza A virus / Q000235:genetics / Q000302:isolation & purification; D025202:Molecular Diagnostic Techniques / Q000379:methods / Q000662:veterinary; D060885:Multiplex Polymerase Chain Reaction / Q000379:methods* / Q000662:veterinary; D009522:Newcastle disease virus / Q000235:genetics / Q000302:isolation & purification; D011201:Poultry Diseases / Q000175:diagnosis / Q000821:virology*; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D014777:Virus Diseases / Q000175:diagnosis* / Q000662:veterinary* / Q000821:virology; D014780:Viruses / Q000145:classification / Q000235:genetics / Q000302:isolation & purification*