A novel multiplex qPCR targeting 23S rDNA for diagnosis of swine dysentery and porcine intestinal spirochaetosis. Issue 1 (December 2016)

• Record Type: Journal Article
• Authors: Borgström, Anna; Scherrer, Simone; Kirchgässner, Constanze; Schmitt, Sarah; Frei, Daniel; Wittenbrink, Max
• Is Part Of: BMC veterinary research. Volume 13:Issue 1(2017)
• Page Start: 1
• Page End: 8
• Publication Date: 2016-12
• Subjects: Swine dysentery -- Brachyspira hyodysenteriae -- 23S rDNA -- Multiplex Real-time PCR -- Swabs; Veterinary medicine -- Research -- Periodicals; Veterinary medicine -- Periodicals; 636.0890724
• DOI: 10.1186/s12917-016-0939-6
• Languages: English
• ISSNs: 1746-6148

Abstract

Abstract Background A multiplex qPCR targeting a 128 bp region on the 23S rDNA gene was developed for detection ofBrachyspira (B.) hyodysenteriae andB. pilosicoli, the agents of swine dysentery (SD) and porcine intestinal spirochaetosis (PIS), together with a triplet of apathogenicBrachyspira spp. (B. innocens, B. intermedia, B. murdochii ) in porcine feces. The multiplex qPCR was evaluated against a duplex PCR (La et al., J Clin Microbiol 41:3372–5, 2003). Results Using DNA extracted from fecal culture, the multiplex qPCR showed excellent agreement with the duplex PCR (κ = 0.943 and 0.933). In addition, thanks to the three probes whereof one detecting the apathogenicBrachyspria spp., a more diversified overview of the brachyspiral flora in porcine fecal samples can be delivered as a part of the routine diagnostic. The multiplex qPCR with a limit of detection of 5–10 genomic equivalents (GE) per reaction (6 × 102 GE per gram) allows reliable detection ofBrachyspira species directly from fecal swab DNA. In line with this, analysis of 202 fecal swabs in comparison with culture-based qPCR showed a high agreement for the causative agents of SD (B.hyodysenteriae: κ = 0.853, sensitivity 87% specificity 98%). Conclusion The novel multiplex qPCR is robust and has a high analytical sensitivity and is therefore suitable for high-throughput screening of porcine fecal swabs for the causative agents of SD. This assay can therefore be used for the direct proof of the pathogenicB . spp. in fecal swabs within the scope of a monitoring program.