Genetic characterization of E2 gene of classical swine fever virus by restriction fragment length polymorphism and phylogenetic analysis.
An RT-nested PCR (RT-nPCR)-based restriction fragment length polymorphism (RFLP) analyses of the E2 gene were developed for genetic subtyping and differentiation of vaccinated and infected classical swine fever virus (CSFV) strains. RT-nPCR identified 96 CSFV-positive samples from 321 clinical specimens from southeastern China during 2003-2008. The PCR products of positive samples were further differentiated using MspI digestion, 23 were identified as the C-strain, 62 as field strains, and 11 as mixture of the vaccine strain and field ones. RFLP with BglI, DdeI, DraI, and PstI were then used for subtyping of the field CSFV isolates. Thirty-eight field isolates phylogenetically classified as subgroup 2.1 based on E2 were divided into 11 subtypes by this RFLP scheme. Both RFLP profiling and sequence-based phylogenetic analysis revealed genetic diversity of CSFV in the field. Three novel substitutions at amino acid positions 17, 93, and 286 were identified in the predominant subtype VI strains isolated in 2008 as compared to other strains including historical subtype VI strains. These results suggest that CSFV in China experienced gradual variations and evolutionary accumulation progress. Thus, the RFLP methods targeting on the CSFV E2 gene are suitable for epidemiological survey in endemic area where the C-strain is applied for vaccination. Combination of the RFLP schemes with sequence-based phylogenetic analysis could provide more detailed information on transmission of CSFV in the region or even its evolution.
Chen
Ning
N
Zhejiang Provincial Key Laboratory of Preventive Veterinary Medicine, Institute of Preventive Veterinary Medicine, Zhejiang University, 268 Kaixuan Road, 310029 Hangzhou, Zhejiang, People's Republic of China.
Li
Dejiang
D
Yuan
Xuemei
X
Li
Xiaoliang
X
Hu
Hongxia
H
Zhu
Binglin
B
Wan
Xiaoyuan
X
Fang
Weihuan
W
D019943:Amino Acid Substitution / Q000235:genetics; D000818:Animals; D002681:China; D006691:Classical Swine Fever / Q000821:virology*; D006692:Classical Swine Fever Virus / Q000145:classification* / Q000235:genetics / Q000302:isolation & purification; D016000:Cluster Analysis; D016172:DNA Fingerprinting; D005838:Genotype; D017720:Molecular Epidemiology; D008969:Molecular Sequence Data; D010802:Phylogeny; D016133:Polymerase Chain Reaction / Q000379:methods*; D012150:Polymorphism, Restriction Fragment Length; D012367:RNA, Viral / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*; D017422:Sequence Analysis, DNA; D014759:Viral Envelope Proteins / Q000235:genetics*; D014773:Virology / Q000379:methods*
D012367:RNA, Viral; D014759:Viral Envelope Proteins; C087086:glycoprotein E2, classical swine fever virus