Evaluation of techniques for the detection of toxigenic Pasteurella multocida strains from pigs.

Abstract

Recently acquired field isolates and archived isolates from our collection of Pasteurella multocida were analyzed for production of dermonecrotic toxin. Detection of the toxin was carried out using a fetal lung feline (FLF) cell line and a commercial enzyme-linked immunosorbent assay (ELISA) kit. The dermonecrotic toxin gene (ToxA) was also detected using a polymerase chain reaction (PCR) technique. Results from the 3 methods were compared. Field isolates (group 1) came from a commercial herd that had clinical signs of atrophic rhinitis. Fifty-six (17.9%) strains were isolated from 312 nasal swabs. Thirty-five of these strains belonged to serotype A and the rest (21/56), although probably serotype D, were not characterized further. All of these strains were toxin negative based on both the ELISA and FLF cell culture results. Five isolates gave faint bands in the PCR reaction, and the rest (51/56) were PCR negative. PCR and ELISA were also performed from the initial swab cultures (mixed cultures); 7 samples gave faint PCR bands, but ELISA results were all negative. Archived strains (group 2) had been isolated from clinical cases of atrophic rhinitis and from cases of pulmonary pasteurellosis. A total of 76 strains were analyzed; 46 were serotype A, and the rest (30) were serotype D. ELISA and FLF cell culture tests were negative for all serotype A strains; however, 3 strains showed faint bands in the PCR reaction. Fourteen serotype D strains showed positive results in both the ELISA and the FLF cell culture tests. PCR from these samples also gave positive results showing a strong band in the gel. However, 4 strains that were ELISA and FLF cell culture negative showed a faint band in the PCR reaction. The 3 methods gave similar results in the detection of the P. multocida dermonecrotic toxin. However, complete agreement among the tests was achieved only when strong PCR bands were considered positive. This is the first report that demonstrates the use of FLF cell line for the detection of toxigenic P. multocida.