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Malignant catarrhal fever in a Barbary sheep (Ammotragus lervia)

1993). The first amplification reaction used 100 ng of the primer sets 556 (5′-AGTCTGGGTATATGAATCCAGATGGCTCTC-3′) and 755 (5′-AAGATAAGCACCAGTTATGCATCTGATAAA-3′) in a 25 μl volume containing 1 μl DNA, 300mM Tris-HCl, pH 8·5, 75mM ammonium sulphate, magnesium chloride, 10 per cent dimethyl sulphoxide, 100μM of each of the four nucleotides, and 5 units of Taq DNA polymerase (AmpliTap; Perkin Elmer). This reaction included denaturing of DNA at 95°C for five minutes, followed by 30 cycles at 94°C for 30 seconds, 60°C for 30 seconds and 72°C for 30 seconds. An aliquot of 2·5 μl of primary amplification product specified by primer pair 556/755 was transferred directly to the second reaction mixture. This was amplified with primers 556 and 555 (5′-TTCGG GGTAGTGGCGAGCGAAGGCTTC-3′) for a further 25 cycles at 94°C for 30 seconds, 60°C for 30 seconds, 72°C for three seconds and one extension cycle at 72°C for five minutes (Mirangi and Kangee 1997). Nucleotide sequence analysis was performed on heminested PCR products. The 238 base pair (bp) PCR product was purified (Wizard PCR Prep DNA Purification System; Prometa) and sequenced with an Applied Biosystems Automatic Sequencer with one of the PCR primers. For the 235 bp sequence the primer sets 556/555 were used. The molecular analysis was performed by means of the Pileup software, a part of Genetics Computer Group (Wisconsin Sequence Analysis Package). The most prominent lesions found on postmortem examination were observed in the respiratory and alimentary tracts. The lungs were severely congested and petechial haemorrhages and oedema were present in the trachea and lung. In the oral cavity multifocal erosions involving the tongue, gingiva and the hard and soft palates were found. The abomasal mucosa was congested and multifocal ulcerations, 1 to 2 cm in diameter were observed. The contents of the small and large intestines were bloody and there was submucosal petechiation. The liver was friable but normal in colour. The spleen was mildly enlarged. The kidneys consisted of multifocal irregular pale-grey brown foci in the renal cortex. The urinary bladder mucosa contained numerous petechial and ecchymotic haemorrhages of varying size. The gross appearance of the heart and great vessels was clinically normal. Haemorrhages were present in the parenchyma of both testicles. Lymph nodes throughout the body were enlarged oedematous, and on the cut surface congested petechial haemorrhages were seen. Histopathology of brain, liver, spleen, kidney and lymph nodes showed necrotising vasculitis and perivascular infiltrations by lymphocytes and macrophages. These lesions are consistent with MCF (Liggitt and De Martini 1980). In the brain a non-suppurative meningoencephalitis was present. Lesions of the alimentary tract were characterised by mucosal hyperemia, with diffuse proliferation of lymphoblastic cells in the mucosa and submucosa oedema. There were also massive haemorrhages into the intratubular spaces of both testicles. A 238 bp fragment of viral genomic DNA was detected using hemi-nested PCR in the blood samples of the Barbary sheep and in eight of 20 domestic sheep. Sequences analysis of the 235 bp of the viral genome of the nine positive animals revealed two genetic variants: V1 and V2 (Fig 1). The virus that was detected in the Barbary sheep was identical to the V2 that was detected in only one of eight positive sheep. The remaining seven positive sheep harboured V1 showing two nucleotide differences in positions 111 and 112 (Fig 1) (accession number L05908, S64565). There is evidence that infections with related, yet distinct gammaherpesviruses may be very common in ruminants, but most of these infections seem to run a subclinical course. They may induce a natural state of immunity against potentially harmful heterologous viruses (Metzler 1991). There is Veterinary Record (2004) 155, 463-465