Development of a one step real time RT-PCR assay to detect and quantify Dugbe virus

A one-step real time quantitative RT-PCR (qRT-PCR) assay was developed to detect all published Dugbe virus (DUGV) genomes of the Nairovirus genus. Primers and probes were designed to detect specific sequences on the most conserved regions of the S segment. The limit of detection of the assay was 10 copies per reaction which is an improvement of 3 log10 FFU/mL over the sensitivity of conventional RT-PCR. The specificity of the primers and probe was confirmed with the closely related Nairoviruses CCHFV and Hazara virus, and on the non-related viruses Coronavirus and Influenza A virus. This qRT-PCR assay was used to screen nucleic acids extracted from 498 ticks collected in the Republic of Chad. One sample was found positive suggesting that DUGV is present in this part of the world. The molecular assay developed in this study is sensitive, specific and rapid and can be used for research and epidemiological studies.

► A one-step real time quantitative RT-PCR assay was developed to detect published Dugbe virus genomes of the Nairovirus genus.
► Primers and probes were designed to detect specific sequences on the most conserved regions of the S segment.
► The limit of detection was 10 copies per reaction improving of 3 log10 FFU/mL the conventional RT-PCR.
► The specificity of the primers and probe was confirmed with the closely related Nairoviruses CCHFV and Hazara virus.
► 498 ticks collected in the Republic of Chad were screened: one sample was found positive.