The effect of sample degradation and RNA stabilization on classical swine fever virus RT-PCR and ELISA methods.
Classical swine fever (CSF), also known as hog cholera, is a highly contagious viral infection of swine caused by a member of the genus pestivirus of the family, Flaviviridae. The need for accurate laboratory diagnosis of CSF is particularly important as it is more reliable than clinical diagnosis. CSF is endemic in many tropical countries where the climate is characterized by high ambient temperature and humidity. This study details the effect of sample quality on CSF antigen-capture ELISA (AC-ELISA) and reverse transcriptase-polymerase chain reaction (RT-PCR) methods. RT-PCR assessment of AC-ELISA-positive spleen samples stored in a conventional glycerol/saline buffer demonstrated that the RT-PCR was detrimentally affected by poor sample quality. To provide a more accurate representation of this effect, a 14 days study was performed to determine the effect of tropical ambient conditions on CSF virus-positive spleen samples stored in two transport media; glycerol/saline and a proprietary RNA preservation solution (RNAlater). A protective effect was demonstrated in both assays with RNAlater as samples were positive in both assays until day 14 post-exposure. Samples stored in glycerol/saline were negative at RT-PCR at day 3 post-exposure although AC-ELISA was still positive at day 14 post-exposure.
Blacksell
Stuart D
SD
CSIRO Australian Animal Health Laboratory, P.O. Bag 24, Geelong, Victoria 3200, Australia. stuart@tropmedres.ac
Khounsy
Syseng
S
Westbury
Harvey A
HA
D000818:Animals; D001483:Base Sequence; D006691:Classical Swine Fever / Q000175:diagnosis; D006692:Classical Swine Fever Virus / Q000235:genetics* / Q000302:isolation & purification*; D017931:DNA Primers / Q000235:genetics; D004279:DNA, Viral / Q000235:genetics; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D020871:RNA Stability; D012367:RNA, Viral / Q000235:genetics / Q000302:isolation & purification; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*; D013552:Swine; D014773:Virology / Q000379:methods
D017931:DNA Primers; D004279:DNA, Viral; D012367:RNA, Viral