title: Detection, discrimination and quantitation of 22 bluetongue virus serotypes using real-time RT-PCR with TaqMan MGB probes.
abstract: Bluetongue virus (BTV) is the etiological agent of bluetongue (BT) disease, a noncontagious insect-transmitted disease of international importance. To date, 26 BTV serotypes have been recognized worldwide. Methods to discriminate BTV serotypes in clinical samples are essential to epidemiological surveillance efforts and BTV vaccination programs. The BTV VP2 major outer capsid protein, encoded by genomic segment 2 (Seg-2), is the most highly variable BTV protein and is the primary determinant of the virus serotype. Here, we report the development of rapid and reliable real-time RT-PCR assays to detect and discriminate 22 BTV serotypes on the basis of VP2-encoding genomic sequences. Serotype-specific primers and probes detected only the targeted BTV serotype and displayed no cross-amplification of off-target BTV serotypes or other closely related Reoviridae and Bunyaviridae family members. The real-time RT-PCR assays developed were highly sensitive, and the majority of serotype-specific reactions could detect template when present at ≥10 copies. These BTV serotype-specific real-time RT-PCR assays represent a rapid, sensitive, and reliable method for the identification, differentiation and quantification of 22 BTV serotypes.
authors.affiliation: Department of Basic Veterinary Medicine, College of Veterinary Medicine, Northeast Agricultural University, 59 Mucai Street, Xiangfang District, Harbin, 150030, People's Republic of China, FYF_anna@163.com.
authors.forename: Yufei
authors.initials: Y
authors.lastname: Feng
authors.forename: Tao
authors.initials: T
authors.lastname: Yang
authors.forename: Qingyuan
authors.initials: Q
authors.lastname: Xu
authors.forename: Encheng
authors.initials: E
authors.lastname: Sun
authors.forename: Junping
authors.initials: J
authors.lastname: Li
authors.forename: Shuang
authors.initials: S
authors.lastname: Lv
authors.forename: Haixiu
authors.initials: H
authors.lastname: Wang
authors.forename: Qin
authors.initials: Q
authors.lastname: Zhang
authors.forename: Jikai
authors.initials: J
authors.lastname: Zhang
authors.forename: Donglai
authors.initials: D
authors.lastname: Wu
mesh_terms: D000818:Animals; D001819:Bluetongue / Q000821:virology*; D001820:Bluetongue virus / Q000145:classification* / Q000235:genetics; D005838:Genotype; D060005:Genotyping Techniques / Q000379:methods*; D015345:Oligonucleotide Probes; D060888:Real-Time Polymerase Chain Reaction / Q000379:methods*; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity; D065288:Serogroup; D012756:Sheep
chemical_list: D015345:Oligonucleotide Probes