Blood and feces were collected from 1,333 lactating cows in 27 dairy herds of southern Chile between September 2003 and August 2004. Blood samples were collected in 10 ml vacutainer tubes and the sera frozen at -20°C until tested using a commercial ELISA kit (IDEXX Laboratories, Inc.). Duplicate samples were assayed following manufacturer recommendations and the S/P ratio for each sample calculated using an automated ELISA reader (IDEXX). Rectal fecal samples were collected using individual polyethylene sleeves, transported to the lab, and cultured within 24h on Herrold's Egg Yolk Medium (HEYM) and mycobactin J (3 tubes) and HEYM without mycobactin J (1 tube). Prior to culture, 2g of each fecal sample was suspended in 35 ml HPC solution and incubated at 37°C overnight. It was then centrifuged and the pellet re-suspended in an antibiotic solution containing nalidixic acid, vancomycin, and amphotericin B, again incubated at 37°C overnight. A 0.15 ml aliquot of each suspension was used to inoculate all HEYM tubes which were incubated at 37°C for 16 weeks. Colonies resembling M. paratuberculosis and showing mycobactin-dependence were tested by IS900 PCR. M. paratuberculosis was isolated from 10 (37.0%) herds and 54 (4.1%) cows. By ELISA, herd-level prevalence was 74.1% (20 herds with ≥1 ELISA-positive cow) and cow-level prevalence was 6.5% (86). ELISA sensitivity and specificity, relative to a single fecal culture, were 38.9% and 94.9%, respectively. The kappa value for ELISA and culture agreement was 0.263. These results show ELISA accuracy similar to that of other reports and that Johnne's disease is widely distributed in dairy herds in southern Chile. A control program to reduce its prevalence is urgently needed.

Sponsorship

Grant research supported by FONDO SAG, Ministry of Agriculture, Chile