PubMed · 4355831
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H A Wood. 1973. Viruses with double-stranded RNA genomes.. https://doi.org/10.1099/0022-1317-20-supplement-61
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This paper describes the production and purification of a group-specific recombinant protein VP7 of African horse sickness virus serotype 3 (AHSV-3) and validation of an I-ELISA for the detection of IgG-antibodies to VP7 in horse sera. Baculovirus-expressed VP7 crystals were purified from infected insect cells. Analytical accuracy of the I-ELISA was examined using sera (n = 38) from an experimentally infected horse, from foals born to vaccinated mares, from guinea-pigs immunized with nine serotypes of AHSV, and from sera of animals infected with other orbiviruses. Compared to traditional serological assays, the I-ELISA was more sensitive in detection of the earliest immunological response in an infected horse and declining levels of maternal immunity in foals. Antibodies to all nine serotypes of AHSV could be detected. Cross-reactivity to related orbiviruses was not observed. Diagnostic accuracy of the I-ELISA was assessed by testing sera from vaccinated horses (n = 358) residing in AHS-enzootic areas and from unvaccinated horses (n = 481) residing in an AHS-free area. Sera were categorised as positive or negative for antibodies to AHSV using virus neutralisation tests. The TG-ROC analysis was used for the selection of the cut-off value. At a cut-off of 11.9 of the high positive control serum (percentage positivity), the I-ELISA specificity was 100%, sensitivity 99.4%, and the Jouden index was 0.99.
Culicoides obsoletus (Meigen) and Culicoides scoticus Downes & Kettle (Diptera: Ceratopogonidae) are sibling species of the Obsoletus group. This group comprises species of biting midges that are suspect vectors of bluetongue virus (family Reoviridae, genus Orbivirus, BTV) and African horse sickness virus (family Reoviridae, genus Orbivirus, AHSV). BTV and AHSV have been isolated several times from females of this group, although it has not been possible to determine the particular species harboring the virus, because of the inability to clearly identify the females of each species based on morphology. Both sexes of C. obsoletus and C. scoticus midges from Catalonia and the Balearics (Spain) were sequenced for the mitochondrial gene cytochrome oxidase subunit I (COI), and these sequences were analyzed to determine intra- and interspecific genetic variability. Species-specific primers for C. obsoletus and C. scoticus were designed and a polymerase chain reaction (PCR)-based diagnostic assay based on the COI gene and using a hemi-nested PCR technique was developed for reliably distinguishing the females of both species. The species-specific PCR diagnostic was compared with morphological discrimination of C. obsoletus and C. scoticus females. The morphologic characters were not fully reliable.