Development, characterization and diagnostic application of a monoclonal antibody specific for a proteinase K resistant Lawsonia intracellularis antigen.

Proliferative enteropathy (PE) is one of the most important infections in pigs caused by Lawsonia intracellularis, an obligate intracellular bacterium. The purpose of the present investigation was to develop monoclonal antibodies with specificity to L. intracellularis useful both for diagnostic purposes (by immunohistochemistry) and for bacterial characterization. Several antibody producing hybridomas were established by fusion of mouse myeloma with spleen cells from BALB/c mice immunized with mucosa scrapings of the intestinal mucosa from a L. intracellularis infected pig. A monoclonal antibody (mAb), Law1-DK, isotyped as IgG2b was selected by indirect immunofluorescence antibody test (IFAT). Histological sections of the intestines from pigs affected by proliferative enteropathy and in vitro grown bacteria in cell culture were tested positive for the presence of L. intracellularis with the mAb. A molecule at 21 kDa was recognized by the mAb in a Western blotting analysis when a whole-cell preparation of L. intracellularis was run on a sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). This antigen was released from L. intracellularis by mild heat treatment and was resistant to proteinase K digestion, suggesting it to be non-protein, e.g., lipopolysaccharide (LPS). This suggestion was supported by its presence in the aqueous phase of a phenol-water extract. The inhibitory effect of periodate oxidation on the antigen-antibody binding confirmed the participation of a carbohydrate epitope. The new mAb was tested highly specific for L. intracellularis by applying in situ hybridization with a L. intracellularis specific probe targeting 16S ribosomal RNA simultaneously with the IFAT.

The influence of diet on Lawsonia intracellularis colonization in pigs upon experimental challenge.

The objective of this investigation was to study if different feeding strategies influence experimental infections of pigs with Lawsonia intracellularis, the causative agent of proliferative enteropathy. In three sequential trials, a total of 144 weaned pigs were fed five different diets all made from a standard diet based on wheat and barley as carbohydrate source and soybean as protein source. The five diets were: a standard diet (fine ground and pelleted), the standard diet fed as fermented liquid feed, the standard diet added 1.8% formic acid, the standard diet added 2.4% lactic acid and a diet similar to the standard diet (made from the same ingredients), but fed coarse ground. Twenty-four pigs on each diet were orally inoculated with L. intracellularis and growth performance and faecal excretion of bacteria were monitored. Twenty-four pigs fed the standard diet were included as not experimentally infected controls. Pigs in the first two trials were sacrificed 4 weeks post-inoculation, whereas animals in the third trial were sacrificed after 5 weeks. Pigs in all experimentally infected groups excreted L. intracellularis. The fermented liquid diet delayed the excretion of L. intracellularis and furthermore, pigs fed the standard diet supplemented with lactic acid had limited pathological lesions when the intestines were examined 4 weeks after inoculation. The growth performance was reduced in pigs experimentally challenged with L. intracellularis, however the prevalence and severity of diarrhea was limited.