Development and evaluation of a loop-mediated isothermal amplification assay for rapid detection of chicken anaemia virus.
OBJECTIVE
Chicken anaemia virus (CAV) causes an economically important viral disease in chickens worldwide. The main aim of this study was to establish a rapid, sensitive and specific loop-mediated isothermal amplification (LAMP) assay for detecting CAV infection.


RESULTS
A set of four specific LAMP primers were designed based on the nucleotide sequence of the CAV VP2 gene, which encodes a nonstructural protein. These were used for the amplification of a specific target region of the VP2 gene. LAMP amplicons were successfully amplified and detected by DNA electrophoresis and by direct naked eye SYBR Green I visualization. A sensitivity test systematically demonstrated that the LAMP assay was superior to a conventional PCR assay with a minimum concentration limit of 100 fg compared to 10 ng for the conventional PCR. The specificity of the LAMP assay for CAV detection is consistent with conventional PCR. Using this established LAMP assay, infected and uninfected clinical samples obtained from an experimental farm were fully verified.


CONCLUSIONS
A novel nucleic acid-based approach of LAMP assay was successfully developed for detecting CAV infection.


CONCLUSIONS
In this study, these results indicate that the developed LAMP assay herein for CAV detection is a time-effective, simple, sensitive and specific test that can be used as an alternative approach in the future for large-scaled diagnosis on the farm of CAV infection.
Huang
C-H
CH
 Institute of Biochemistry, College of life Science, National Chung Hsing University, Taichung, Taiwan.
Lai
G-H
GH
 Institute of Biochemistry, College of life Science, National Chung Hsing University, Taichung, Taiwan.
Lee
M-S
MS
 Department of Medical Research, Tung's Taichung MetroHarbour Hospital, Taichung, Taiwan.
Lin
W-H
WH
 Master Degree Program, PhD Program, School of Pharmacy Undergraduate Program, China Medical University, Taichung, Taiwan.
Lien
Y-Y
YY
 Department of Veterinary Medicine, National Pingtung University of Science and Technology, Pingtung, Taiwan.
Hsueh
S-C
SC
 Department of Veterinary Medicine, National Pingtung University of Science and Technology, Pingtung, Taiwan.
Kao
J-Y
JY
 Institute of Biochemistry, College of life Science, National Chung Hsing University, Taichung, Taiwan.
Chang
W-T
WT
 School of Chinese Medicine Resources, China Medical University, Taichung, Taiwan.
Lu
T-C
TC
 Institute of Biochemistry, College of life Science, National Chung Hsing University, Taichung, Taiwan.
Lin
W-N
WN
 College of Pharmacy, Graduate Institute of Chinese Pharmaceutical Sciences, China Medical University, Taichung, Taiwan.
Chen
H-J
HJ
 Department of Safety, Health and Environmental Engineering, Mingchi University of Technology, Taipei, Taiwan.
Lee
M-S
MS
 School of Chinese Medicine Resources, China Medical University, Taichung, Taiwan.
D000818:Animals; D036022:Capsid Proteins / Q000235:genetics; D017727:Chicken anemia virus / Q000235:genetics / Q000302:isolation & purification*; D002645:Chickens / Q000821:virology*; D017931:DNA Primers / Q000235:genetics; D008099:Liver / Q000821:virology; D021141:Nucleic Acid Amplification Techniques / Q000379:methods*; D016133:Polymerase Chain Reaction / Q000379:methods; D012680:Sensitivity and Specificity
D036022:Capsid Proteins; D017931:DNA Primers; C442521:VP2 protein, Chicken anemia virus