Development of a non-destructive PCR method for detection of the salivary gland hypertrophy virus (SGHV) in tsetse flies.
A PCR based diagnostic method to detect salivary gland hypertrophy virus (SGHV) in tsetse flies is described. Two sets of primers GpSGHV1F/GpSGHV1R and GpSGHV2F/GpSGHV2R were selected from a virus-specific sequence. Both primer sets can detect specifically the virus in individual tsetse flies by generating an amplicon of 401 bp. Attempts were made to develop a simple and reliable non-destructive virus detection method in live flies. PCR reactions were performed on either crude or purified tsetse DNA from saliva and legs. While saliva can be an indicator for the presence of the virus in flies, the method is laborious. Crude extract from an excised middle leg resulted in a positive PCR reaction equivalent to crude extract from whole fly. However, sensitivity could be significantly increased when purified DNA was used as the template. In conclusion, PCR using a purified DNA template from a single tsetse leg represents an efficient, non-destructive method for virus diagnosis in live tsetse flies.
Abd-Alla
Adly
A
Entomology Unit, FAO/IAEA Agriculture and Biotechnology Laboratory, A-2444 Seibersdorf, Austria. a.m.m.abd-alla@iaea.org
Bossin
Hervé
H
Cousserans
François
F
Parker
Andrew
A
Bergoin
Max
M
Robinson
Alan
A
D000818:Animals; D003587:Cytomegalovirus / Q000235:genetics / Q000302:isolation & purification*; D017931:DNA Primers / Q000235:genetics; D004279:DNA, Viral / Q000235:genetics*; D016133:Polymerase Chain Reaction / Q000379:methods*; D012469:Salivary Glands / Q000821:virology; D014370:Tsetse Flies / Q000821:virology*
D017931:DNA Primers; D004279:DNA, Viral