Comparison of Methods for Virus Detection in Allium spp

• Published in: Journal of phytopathology Vol. 149; no. 11; pp. 731 - 737
• Main Authors: Dovas, C. I, Hatziloukas, E, Salomon, R, Barg, E, Shiboleth, Y, Katis, N. I
• Format: Journal Article
• Language: English
• Published: Oxford UK Blackwell Science Ltd 01-12-2001; Blackwell
• Subjects: Allexivirus; Allium; Allium viruses; Biological and medical sciences; Enzyme-linked immunosorbent assay; Fundamental and applied biological sciences. Psychology; Generalities. Techniques. Transmission, epidemiology, ecology. Antiviral substances, control; immunocapture-reverse transcriptase (RT)-polymerase chain reaction (PCR); leaf extracts; Leek yellow stripe virus; Microbiology; one-step RT-PCR; Onion yellow dwarf virus; Phytopathology. Animal pests. Plant and forest protection; Plant viruses and viroids; reverse transcriptase polymerase chain reaction; RNA-directed DNA polymerase; viruses; Monocotyledones; RNA-directed DNA polymerase; Enzyme; Transferases; Sample; Primer; Geographical origin; Method; Optimization; Virus; Polymerase chain reaction; Onion yellow dwarf virus; Liliaceae; Nucleotidyltransferases; Sensitivity; Sample preparation; Angiospermae; Plant origin; Allium porrum; Spermatophyta; Potyvirus; Detection; Species; Potyviridae
• ISSN: 0931-1785; 1439-0434
• DOI: 10.1046/j.1439-0434.2001.00705.x

Reverse transcriptase (RT)‐polymerase chain reaction (PCR) and immunocapture (IC)‐RT‐PCR protocols were developed and optimized for the sensitive detection of Onion yellow dwarf virus, Leek yellow stripevirus and allexiviruses infecting Allium species. Polyvalence of the designed primers was successfully demonstrated, using samples of different plant species and geographic origins. Different sample preparation procedures were evaluated for their suitability to provide appropriate PCR templates. IC‐PCR, RT‐PCR with plant tissue extracts, and RT‐PCR with total RNA, proved to be 10²–10⁴ times more sensitive than double‐antibody sandwich‐enzyme‐linked immunosorbent assay (ELISA). Furthermore, a ‘one step’ IC‐RT‐PCR assay was developed using plant leaf extract as template source, which proved to be 102 times more sensitive than ELISA, and convenient for testing large numbers of leaf samples.