The detection of African horse sickness virus antigens and antibodies in young equidae

Abstract

SUMMARYFour ponies were each inoculated with a different serotype of African horse sickness virus (AHSV) which had been passaged through cell culture in order to achieve attenuation. Three of the ponies died suddenly after showing mild clinical signs, the fourth pony remained clinically normal and was killed at day 38. Infectious AHSV was isolated from blood samples collected at intervals from all four ponies. Positive antigen ELISA reactions were only observed with blood samples from two of the ponies on the two days preceding death. Specific AHSV antibodies were detected by ELISA in serum samples from the other two ponies although one eventually died. African horse sickness viral antigens were detected by ELISA in post-mortem tissue samples collected from all four ponies. No infectious virus could be detected in tissue samples taken post-mortem from the pony which survived African horse sickness (AHS) infection. In the event of a suspected outbreak of AHS it is recommended that sera and heparinized blood should be tested for specific antibodies and AHSV antigen respectively. When available, post-mortem tissues, including spleen, heart, lung and liver, should also be tested for AHSV antigen. Although the ELISA used for the detection of AHSV antigen is highly sensitive and specific, negative ELISA results should be confirmed by virus isolation attempts.

African horse sickness

“…Traditionally this has depended on the isolation of the infectious virus from whole blood collected in anticoagulant (preferably EDTA) during the febrile stage of infection [49]. Because susceptible horses usually die following an acute infection virus may also be readily recovered and identified after death directly from tissues such as spleen, lung, lymph nodes and salivary glands [37,46]. Whole blood should be washed and lysed as soon as possible after collection to remove the anticoagulant and any antibody that might be present in the serum component.…”

“…Group specific antibody against AHSV can be detected using a several diagnostic assays that are directed primarily toward the VP7, including complement fixation [14], agar gel immunodiffusion [4], immunofluorescence [27] and ELISA [44,46,68]. An indirect ELISA based on the detection of antibody against segment 10 (NS3) has also been described for AHSV serotype 4 [69].…”

“…As a result both can occur simultaneously in the same locations and even in the same animal. Fortunately, rapid, sensitive and specific ELISA's are available to enable the detection of the antigen and antibody of both AHSV and EEV [24,[44][45][46]134], and if used in conjunction with each other can provide a rapid and efficient differential diagnosis.…”

A competitive ELISA for the detection of group-specific antibody to equine encephalosis virus

“…Competitive ELISAs have been used successfully for many years for the detection of antibodies to different Orbivirus species, including AHSV (Hamblin et al, 1990;Hamblin et al, 1992), BTV (Anderson, 1984) and EHDV (Thevasagayam et al, 1995). The assay described here measures the competition between an EEV specific guinea pig antiserum and a test serum for a pre-titrated EEV antigen.…”

“…The procedure used was based on the C-ELISA described previously for AHSV (Hamblin et al, 1990;Hamblin et al, 1992). Briefly 50μl/well of EEV antigen, optimally diluted in 0.05 M carbonate-bicarbonate buffer, pH 9.6 (Sigma, Gillingham, UK) was passively adsorbed onto ELISA plates (Dynex Technologies, East Grinstead, UK) overnight at 4°C.…”