Validation of ELISA for the detection of African horse sickness virus antigens and antibodies.

The mortality rate in susceptible populations of horses during an epizootic of African horse sickness (AHS) may be in excess of 90%. Rapid and reliable assays are therefore essential for the confirmation of clinical diagnoses and to enable control strategies to be implemented without undue delay. One of the major objectives of a recent European Union funded project was the validation of newly developed diagnostic assays which are rapid, sensitive, highly reproducible and inexpensive, for the detection of African horse sickness virus (AHSV) antigens and antibodies. The Laboratorio de Sanidad y Produccion Animal (LSPA) in Algete, Spain was charged with the responsibility of co-ordinating and supplying samples of viruses and antisera to the participating laboratories in Spain, France and the United Kingdom. The panels comprised 76 antigen samples for assay by indirect sandwich ELISAs and 53 serum samples for antibody detection by either indirect or competitive ELISAs. Results generated by ELISA for each laboratory were analysed in LSPA in terms of their relative sensitivities and specificities. There was a good agreement between the ELISAs used for either antigen or antibody detection. The participating groups agreed that any field sample giving a doubtful result would always be retested by ELISA and an alternative assay.

Clinical features of the 1992 outbreak of equine viral arteritis in Spain.

During 1992, a widespread outbreak of Equine viral arteritis (EVA) occurred at a riding establishment near Barcelona, Spain. A total of 31 out of 186 horses on the premises displayed clinical signs, most frequently, fever, depression, mild ventral and limb oedema and a vesicular-erosive stomatitis, with hypersalivation, petechiations and small ulcerations. Affected horses developed illness of varying severity with only a few exhibiting a severe form of the disease and no mortality was recorded. Haematological and blood biochemical examination the most severely affected horses revealed a thrombocytopenia, slight leucocytosis with neutrophilia, lymphopenia and eosinopenia, an increase in plasma fibrinogen and a small rise in serum proteins and indirect bilirubin values. Diagnosis was confirmed by demonstration of seroconversion to equine arteritis virus in acute and convalescent phase sera. Attempted isolation of the virus from citrated blood samples proved unsuccessful.

Ion-exchange microchromatography and thiobarbituric acid colorimetry for the measurement of canine glycated hemoglobins.

Nonenzymatic glycation of hemoglobin is a slow, continuous, and irreversible process which takes place during the whole lifespan of the erythrocyte. When hemolytic diseases are ruled out, the levels of glycated hemoglobins reflect the time-averaged serum glucose concentration for the preceding weeks. Canine hemoglobin also binds physiologically to intraerythrocytic glucose to form a glycated fraction which provides information on the animal's long-term glycemic status. This study describes an overall evaluation of ion-exchange microchromatography and thiobarbituric acid (TBA) colorimetry for the measurement of canine glycated hemoglobins. The intra- and inter-assay coefficients of variation (CVs) found were less than 5% in normal and diabetic canine samples, and both assays proved linear over the analytical range tested, which was wide enough to include the expected clinical values. Under our laboratory's conditions, the reference range for HbA(1) was 5.82 +/- 0.62% and for HbA(1)c was 2.35 -/+ 0.47%. Sample stability was lower using the ion-exchange procedure, with increases in HbA(1) observed after 4 days in whole blood and hemolysates stored at room temperature, after 12 days in whole blood stored at 4 degrees C, and after 7 days in hemolysates stored at 4 degrees C and -20 degrees C. In the case of TBA colorimetry, whole blood was stable for at least 21 days at room temperature and at 4 degrees C, and hemolysates were stable for 18 days at room temperature, at least 21 days at 4 degrees C, and up to 3 months at -20 degrees C.

Current status of the diagnosis and control of African horse sickness.

African horse sickness (AHS) is an infectious, non-contagious, highly fatal viral disease of Equidae, transmitted by arthropod vectors of the genus Culicoides, and endemic in Africa south and east of the Sahara. The disease is caused by a virus of the Reoviridae family, genus Orbivirus, and 9 serotypes have been recognized. Recent outbreaks of AHS in the Iberian peninsula and Northern Africa emphasize the need for accurate diagnosis and rapid implementation of control measures. In this paper, the epizootiological factors, clinical signs and necropsy findings of AHS are discussed, and an updated review of diagnostic laboratory methods together with a description of the main control measures is given.

African horse sickness in Spain.

The aetiology, pathogenesis and epizootiology of African horse sickness (AHS) are reviewed with special reference to recent outbreaks in the Iberian peninsula. AHS is a highly fatal insect-borne viral disease of Equidae. It is caused by an Orbivirus (family Reoviridae) and nine serotypes are recognised. Outbreaks occurred in central Spain in 1987 and in southern regions of the Iberian peninsula in 1988, 1989 and 1990. All were associated with serotype 4 of the virus, whereas other occurrences of AHS outside Africa have all been caused by serotype 9. The clinical picture in the outbreaks was mainly of the acute (pulmonary) form except in 1988 when the subacute (cardiac) form of disease predominated. Several hundred horses died or were destroyed as a result of the outbreaks. Further spread was contained by a combination of slaughter of sick animals, movement controls, and vaccination which was extended over an increasingly wide area in successive years. The 1987 outbreak is believed to be associated with infected zebras imported from Africa. Possible explanations for the recurrence of disease in Spain in successive years are considered to include (a) the climatic conditions in Southern Spain, which could permit continuous vector activity, (b) the relative clinical resistance of mules and donkeys, which may permit subclinical circulation of the virus, (c) incomplete population immunity among horses due to possible gaps in the vaccination strategy.

Antibodies in horses, mules and donkeys following monovalent vaccination against African horse sickness.

A total of 256 sera collected from three species of domesticated equidae in four different Spanish provinces were examined 1-4 months after the administration of attenuated monovalent African horse sickness virus (AHSV) serotype 4 vaccine. Approximately 10% of the sera were negative by ELISA, virus neutralization, agar gel immuno-diffusion and complement fixation tests. Similar negative reactions were recorded with sera from two ponies after experimental primary vaccination. The rapid rise in antibodies in sera from these two ponies, after a second dose of vaccine, suggested they would probably have been immune to challenge. It is therefore suggested that the apparent absence of antibodies against AHSV in some animals after primary vaccination may not necessarily indicate a total lack of protection.