FIG 1: Phylogenetic tree (Clustal method) showing the relationships between the nine serotypes of African horse sickness virus (AHsv) based on amino acid sequence analysis of the protein NS3. sP Spain, M Morocco, us United States

After 14C‐labelled cortisol infusion in ponies and pigs, faecal samples were collected. Extraction of 0.5 g faeces with 5 ml 80–90 % methanol yielded the highest radioactivity in the supernatant. Most of the metabolites were ether soluble. After high performance liquid chromatography (HPLC), the presence of immunoreactive metabolites was demonstrated by measuring each HPLC fraction using enzyme immunoassays for cortisol, corticosterone and 11‐oxoaetiocholanolone. Only the assay for 11‐oxoaetiocholanolone revealed peaks with co‐eluting radioactivity. For biological validation of the test system, adrenocorticotrophic hormone (ACTH) and dexamethasone were injected intravenously successively in both species (n = 6). Cortisol concentration in blood and the 11‐oxoaetiocholanolone immunoreactive substances in faeces were determined. In horse faeces, basal values of 2.3–35.2 nmol/kg were measured. After ACTH administration, an increase (more than 200 % above basal values) of these metabolites was seen about 1 day after ACTH administration. After dexamethasone injection the levels decreased, reaching minimum concentrations 2 days after administration. In pigs, an increase in these metabolites was measured in only three animals after ACTH; dexamethasone did not cause a decrease. The stability of the samples after defecation was tested by storing samples from cows, horses and pigs at room temperature. It was shown that there was a significant increase in the concentration of measured cortisol metabolites in bovine, equine and porcine faeces after storage for 1 h, 4 h and 24 h, respectively. In frozen samples this effect was diminished after thawing samples at 40°C; thawing the samples at 95°C prevented an increase in immunoreactive substances.

Skin biopsies were taken from four body sites (head, thorax, flank and perineum) of three male entire Beagles and the primary hair follicles were isolated. Culture conditions were established to keep the hair follicles growing for up to 7 days. Additionally, hair follicles were incubated in supplemented medium (containing insulin, transferrin, glutamine and sodium selenite) with or without the addition of testosterone (T) (1, 10 or 100 ng/ml) or oestradiol-17beta (E2beta) (0.01, 0.1 or 1 ng/ml), respectively and the daily growth of hair follicles was measured. In vitro daily growth of hair follicles from the thorax was stimulated by the low concentration of both hormones, but the growth of those from the flank was inhibited by the high concentration of both hormones. Hair follicles from the head were positively influenced by the lowest concentration of T and the medium concentration of E2beta. The daily growth of hair follicles from the perineum was not significantly influenced by either hormone.