Lawsonia intracellularis is a Gram-negative obligately intracellular bacterium (McOrist et al., 1995) which affects the digestive tract, and particularly the caudal portion of the small intestine -the ileum -of sensitive hosts. In dogs, changes characteristic of the presence of this intracellular bacterium (in the past called Campylobacter-like organisms) were reported in two cases only: in the mucous membrane of ileum (Collins and Libal, 1983) and in stomach mucosa (Leblanc et al., 1993). The techniques used to detect the pathogen in histological sections of affected tissues included silver staining according to Warthin-Starry, immunofluorescence and electron microscopy.This paper describes a methodology used in the case of a dog suffering from the IBD (Husník et al. 2003) where the DNA of the intracellular bacteria L. intracellularis was detected using the nested PCR including sequencing the DNA fragment obtained, and where IgG antibodies against L. intracellularis in the blood serum were demonstrated.
MATERIAL AND METHODSIn the tests, specimens taken from a two and a half years old German smooth-coated cocker spaniel suffering from the inflammatory bowel disease were used. Details on the diseased dog have been given elsewhere (Husník et al., 2003).Collection of specimens 1. Biopsy specimens. A total of eight mucous membrane specimens from the stomach and intestines were taken: two from the stomach and the duodenum each, and one from ileum, caecum, colon ascendens and colon descendens. The material was obtained in three endoscopic examinations on Day 40 (5 June), Day 217 (29 November) and 237 (19 December) following the first examination. The biopsy specimens for histopathological examinations were fixed in 10% neutral formalin. ABSTRACT: A nested polymerase chain reaction (PCR) assay and serological examinations were used to detect the presence of Lawsonia intracellularis in a two and a half years old German smooth-coated cocker spaniel with clinical symptoms of chronic diarrhoea and histologically proven inflammatory bowel disease. Fourteen rectal swabs taken over a period of two weeks and eight biopsy specimens taken over a period of six months were used for laboratory examinations. Using the nested PCR, the DNA of L. intracellularis was found in a total of 2 cases, i.e. one rectal swab and one biopsy specimen of the duodenum six months later. The species specificity of the nested PCR product was confirmed by sequencing. The presence of specific IgG antibodies against L. intracellularis was demonstrated by the IFAT in five samples of blood serum taken over a period of seven months.
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