A rapid and specific real time RT-PCR assay for diagnosis of Toscana XXXX .

@entity1 @entity5422 @entity5827 @entity778
BACKGROUND: To scan a virus (TOSV) belongs to the Phlebovirus genus within the Bunyaviridae family. TOSV is an arbovirus transmitted by sandflies. In Mediterranean countries, TOSV is one of the major viral pathogens involved in @entity5827 and @entity5422 . OBJECTIVES: Development and assessment of a new sensitive and specific real-time RT-PCR assay for TOSV diagnosis. STUDY DESIGN: TOSV-specific primers and probe targeting the S-segment of the genome were designed, based on recent TOSV sequences available in public databases. Sensitivity was assessed using 10-fold serial dilutions of a RNA transcript and serial dilutions of TOSV strains isolated from infected @entity1 beings. Specificity was determined by testing RNA extracts from closely related Phleboviruses. The assay was then used for @entity778 diagnosis in 971 clinical samples and for TOSV detection in 2000 sandflies. RESULTS: The real-time RT-PCR assay exhibited a sensitivity of under 257 copies per reaction for the RNA transcripts and 0.0056 and 0.014 TCID50 of Italian and Spanish TOSV genotypes per reaction, respectively. No other close Phleboviruses were detected. TOSV was identified in 17 clinical samples and in 3 sandflies. CONCLUSIONS: The assay described is a rapid, robust and reliable real-time RT-PCR test for accurate diagnosis of @entity1 @entity778 as well as for the surveillance of TOSV in vector populations.
