Optimized PCR assay for detection of white spot syndrome virus (WSSV).
A rapid PCR assay for detection of white spot syndrome virus (WSSV) was developed based on the nested PCR procedure described by Lo et al. (1996) and outlined as the recommended PCR diagnostic assay in the Manual of Diagnostic Tests for Aquatic Animals published by the Office of International Epizootics (OIE, 2009). The optimized procedure incorporated the second step primers used in the nested WSSV PCR. By adjusting the annealing temperature and shortening the cycling times, this modified assay is substantially faster and as sensitive as the recommended OIE protocol. The modified PCR test was compared directly to the two-step nested PCR protocol and a modified nested procedure. The sensitivity of the published assay was determined by template dilutions of semi-purified WSSV virions that had been quantitated using real-time PCR for detection of WSSV. Various isolates were tested using the modified procedure, to ensure that the assay was able to detect WSSV from different geographical locations.
Department of Veterinary Science and Microbiology, University of Arizona, Tucson, AZ 85721, USA. lmn@email.arizona.edu
Linda M
LM
Nunan
Donald V
DV
Lightner
D000818:Animals; D017931:DNA Primers / Q000235:genetics; D004279:DNA, Viral / Q000235:genetics; D033561:Penaeidae / Q000821:virology*; D016133:Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity; D013997:Time Factors; D014773:Virology / Q000379:methods*; D046848:White spot syndrome virus 1 / Q000235:genetics / Q000302:isolation & purification*
D017931:DNA Primers; D004279:DNA, Viral