title: Development and evaluation of tailored specific real-time RT-PCR assays for detection of foot-and-mouth disease virus serotypes circulating in East Africa.
abstract: Rapid, reliable and accurate diagnostic methods provide essential support to programmes that monitor and control foot-and-mouth disease (FMD). While pan-specific molecular tests for FMD virus (FMDV) detection are well established and widely used in endemic and FMD-free countries, current serotyping methods mainly rely either on antigen detection ELISAs or nucleotide sequencing approaches. This report describes the development of a panel of serotype-specific real-time RT-PCR assays (rRT-PCR) tailored to detect FMDV lineages currently circulating in East Africa. These assays target sequences within the VP1-coding region that share high intra-lineage identity, but do not cross-react with FMD viruses from other serotypes that circulate in the region. These serotype-specific assays operate with the same thermal profile as the pan-diagnostic tests making it possible to run them in parallel to produce CTvalues comparable to the pan-diagnostic test detecting the 3D-coding region. These assays were evaluated alongside the established pan-specific molecular test using field samples and virus isolates collected from Tanzania, Kenya and Ethiopia that had been previously characterised by nucleotide sequencing. Samples (n=71) representing serotype A (topotype AFRICA, lineage G-I), serotype O (topotypes EA-2 and EA-4), serotype SAT 1 (topotype I (NWZ)) and serotype SAT2 (topotype IV) were correctly identified with these rRT-PCR assays. Furthermore, FMDV RNA from samples that did not contain infectious virus could still be serotyped using these assays. These serotype-specific real-time RT-PCR assays can detect and characterise FMDVs currently circulating in East Africa and hence improve disease control in this region.
authors: The Pirbright Institute, Ash Road, Pirbright, Woking, Surrey GU24 0NF, UK. Electronic address: kasia.bankowska@pirbright.ac.uk.
authors: Katarzyna
authors: K
authors: Bachanek-Bankowska
authors: Department of Microbiology and Parasitology & Southern African Centre for Infectious diseases Surveillance, Sokoine University of Agriculture, Morogoro, Tanzania.
authors: Herieth R
authors: HR
authors: Mero
authors: The Pirbright Institute, Ash Road, Pirbright, Woking, Surrey GU24 0NF, UK.
authors: Jemma
authors: J
authors: Wadsworth
authors: The Pirbright Institute, Ash Road, Pirbright, Woking, Surrey GU24 0NF, UK.
authors: Valerie
authors: V
authors: Mioulet
authors: Tanzania Veterinary Laboratory Agency, P. O. Box 9252, Dar es Salaam, Tanzania.
authors: Raphael
authors: R
authors: Sallu
authors: National Veterinary Institute, Technical University of Denmark, Lindholm, Kalvehave DK-4771, Denmark.
authors: Graham J
authors: GJ
authors: Belsham
authors: Department of Microbiology and Parasitology & Southern African Centre for Infectious diseases Surveillance, Sokoine University of Agriculture, Morogoro, Tanzania.
authors: Christopher J
authors: CJ
authors: Kasanga
authors: The Pirbright Institute, Ash Road, Pirbright, Woking, Surrey GU24 0NF, UK.
authors: Nick J
authors: NJ
authors: Knowles
authors: The Pirbright Institute, Ash Road, Pirbright, Woking, Surrey GU24 0NF, UK.
authors: Donald P
authors: DP
authors: King
mesh_terms: D000351:Africa, Eastern / Q000453:epidemiology; D000818:Animals; D036022:Capsid Proteins / Q000235:genetics; D005002:Ethiopia / Q000453:epidemiology; D005536:Foot-and-Mouth Disease / Q000175:diagnosis* / Q000453:epidemiology / Q000821:virology*; D029544:Foot-and-Mouth Disease Virus / Q000145:classification / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification*; D007630:Kenya / Q000453:epidemiology; D025202:Molecular Diagnostic Techniques / Q000379:methods / Q000592:standards*; D012367:RNA, Viral; D060888:Real-Time Polymerase Chain Reaction / Q000379:methods*; D017422:Sequence Analysis, DNA; D065288:Serogroup; D012703:Serotyping / Q000379:methods / Q000592:standards; D013636:Tanzania / Q000453:epidemiology
chemical_list: D036022:Capsid Proteins; D012367:RNA, Viral; C442501:VP1 protein, Foot-and-mouth disease virus
keywords: East Africa; Foot-and-mouth-disease virus; Real-time RT-PCR; Serotype-specific