The CSFV DNAChip: a novel diagnostic assay for classical swine fever virus.
A novel assay, the CSFV DNAChip, was developed to clearly and rapidly discriminate three genotypes of classical swine fever virus (CSFV). Total RNA was extracted from clinical samples and then subjected to a one-step reverse-transcription polymerase chain reaction (RT-PCR) using Cy3-labeled primers from the 5' non-coding region (NCR) of CSFV. Amplicons were hybridized to the CSFV DNAChip and fluorescence scanning was performed for detection of CSFV. A cut-off fluorescence intensity value of 5000 was determined by two-graph receiver operating curve (TG-ROC) analysis. The limit of detection values for the developed DNA chip assay were 0.313ng/μL for amplicon concentration and 1TCID50/100μL for virus titer. Using the developed DNA chip, 157 field samples (91 CSFV-positive and 66 CSFV-negative) were investigated. The genotypes determined by the CSFV DNAChip agreed completely with those determined by nucleotide sequence analysis of the viral genome. The developed CSFV DNAChip will be helpful in implementing a CSFV eradication strategy, as it provides a rapid and accurate diagnostic assay that can discriminate easily among CSFV genotypes.
Viral Disease Division, Animal and Plant Quarantine Agency, Anyang, Gyeonggi-do, 430-757, Republic of Korea.
Yong Kwan
YK
Kim
Viral Disease Division, Animal and Plant Quarantine Agency, Anyang, Gyeonggi-do, 430-757, Republic of Korea.
Seong-In
SI
Lim
Viral Disease Division, Animal and Plant Quarantine Agency, Anyang, Gyeonggi-do, 430-757, Republic of Korea.
Yoon-Young
YY
Cho
Viral Disease Division, Animal and Plant Quarantine Agency, Anyang, Gyeonggi-do, 430-757, Republic of Korea.
Jae-Young
JY
Song
Median Diagnostics Inc, Chuncheon, Gangwon-do, 200-883, Republic of Korea.
JoonBae
J
Kim
Viral Disease Division, Animal and Plant Quarantine Agency, Anyang, Gyeonggi-do, 430-757, Republic of Korea. Electronic address: andj67@korea.kr.
Dong-Jun
DJ
An
D000818:Animals; D006691:Classical Swine Fever / Q000175:diagnosis* / Q000821:virology*; D006692:Classical Swine Fever Virus / Q000145:classification / Q000235:genetics / Q000302:isolation & purification*; D005453:Fluorescence; D005838:Genotype; D060005:Genotyping Techniques / Q000379:methods; D025202:Molecular Diagnostic Techniques / Q000379:methods*; D009693:Nucleic Acid Hybridization; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods*; D012367:RNA, Viral / Q000235:genetics / Q000302:isolation & purification; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012680:Sensitivity and Specificity; D013552:Swine; D014730:Veterinary Medicine / Q000379:methods*
D012367:RNA, Viral
Classical swine fever; DNAChip; Genotyping