title: Reverse transcription loop-mediated isothermal amplification for the detection of Porcine reproductive and respiratory syndrome virus.
abstract: Porcine reproductive and respiratory syndrome virus (PRRSV) is an important pathogen of swine. The objective of the current study is to investigate the feasibility of using reverse transcription loop-mediated isothermal amplification (RT-LAMP) for the detection of PRRSV. The RT-LAMP is a recently described DNA amplification technique reported to be simple, inexpensive, fast, and accurate. The RT-LAMP reaction was set up using 2 sets of primers that were designed to detect North American and European strains of PRRSV and performed successfully in a simple heat block. The specificity of the amplified product was demonstrated by restriction analysis. The RT-LAMP was able to detect 5 different PRRSV isolates. However, the limit of detection ranged between 10(2) and 10(4) 50% tissue culture infective dose/ml. The RT-LAMP was further evaluated using serum samples from animals of known infection status. The ability of RT-LAMP to detect PRRSV in serum from acutely infected animals was evaluated with 114 serum samples from 18 experimentally inoculated boars. Forty-nine of these samples tested positive by RT-LAMP, while 94 were positive by reverse transcription polymerase chain reaction (RT-PCR). The diagnostic specificity, evaluated with 100 known negative serum samples, was estimated as 99%. The feasibility of RT-LAMP to detect PRRSV was demonstrated in the current study. The RT-LAMP reaction could be performed in just 1 hr with a simple and inexpensive heat block. However, the sensitivity of this technique was significantly lower than that of RT-PCR.
authors.affiliation: University of Minnesota, Veterinary Diagnostic Laboratory, 1333 Gortner Avenue, St. Paul, MN 55108, USA. rove0010@umn.edu
authors.forename: Albert
authors.initials: A
authors.lastname: Rovira
authors.forename: Juan
authors.initials: J
authors.lastname: Abrahante
authors.forename: Michael
authors.initials: M
authors.lastname: Murtaugh
authors.forename: Claudia
authors.initials: C
authors.lastname: Muñoz-Zanzi
mesh_terms: D000818:Animals; D004279:DNA, Viral / Q000145:classification / Q000302:isolation & purification; D009391:Nephelometry and Turbidimetry; D021141:Nucleic Acid Amplification Techniques / Q000662:veterinary*; D019316:Porcine respiratory and reproductive syndrome virus / Q000302:isolation & purification*; D012367:RNA, Viral / Q000145:classification / Q000302:isolation & purification; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods / Q000662:veterinary*; D012680:Sensitivity and Specificity; D013552:Swine
chemical_list: D004279:DNA, Viral; D012367:RNA, Viral