RT Journal Article SR Electronic T1 Comparative genomics reveals signature regions used to develop a robust and sensitive multiplex TaqMan real-time qPCR assay to detect the genus Dickeya and Dickeya dianthicola JF bioRxiv FD Cold Spring Harbor Laboratory SP 847590 DO 10.1101/847590 A1 Shefali Dobhal A1 Gamze Boluk A1 Brooke Babler A1 Michael J. Stulberg A1 John Rascoe A1 Mark Nakhla A1 Toni A. Chapman A1 Alex B. Crockford A1 Michael Melzer A1 Anne M. Alvarez A1 Mohammad Arif YR 2019 UL http://biorxiv.org/content/early/2019/11/20/847590.abstract AB Aims Dickeya species are high consequence plant pathogenic bacteria listed among the quarantine pathogens of the European Union; associated with potato disease outbreaks and subsequent economic losses worldwide. Early, accurate, and reliable detection of Dickeya spp. is needed to prevent establishment and further dissemination of this pathogen. Therefore, a multiplex TaqMan qPCR was developed for sensitive detection of Dickeya spp. and specifically, D. dianthicola.Methods and Results A signature genomic region for the genus Dickeya (mglA/mglC) and unique genomic region for D. dianthicola (alcohol dehydrogenase) were identified using a whole genome based comparative genomics approach. The developed multiplex TaqMan qPCR was validated using extensive inclusivity and exclusivity panels, and naturally/artificially infected samples to confirm broad range detection capability and specificity. Both sensitivity and spiked assays showed detection limit of 10 fg DNA.Conclusion The developed multiplex assay is sensitive and reliable to detect Dickeya spp. and D. dianthicola with no false positives or false negatives. It was able to detect mixed infection from naturally and artificially infected plant materials.Significance and Impact The developed assay will serve as a practical tool for screening of propagative material, monitoring the presence and distribution, and quantification of target pathogens in a breeding program. The assay also has applications in routine diagnostics, biosecurity and microbial forensics.