Gross and microscopic lesions

3. Gross and microscopic lesions • Characteristic, but not pathognomonic • Insensitive diagnostic method (19% positive animals lack lesions) • Could be difficult to differentiate from other pneumonias (Pasteurella, Haemophilus)

6. Seroconversion to Different Agents vs Average Lesion Score

7. Serology • ELISA-based tests currently available • Tween-ELISA • Commercial Tween-ELISA • IDDEX ELISA • Dako ELISA

8. Tween-ELISAS • Antigen is a composite of membrane proteins • Excellent when prepared in-house • Some dispute as to whether it cross reacts with flocculare • Commercial tests give some false positives

9. IDDEX ELISA • Becoming a very common test • Very high sensitivity • Specificity can be a problem:false positives • In negative herd monitoring, positives must be confirmed with the Dako test

10. Dako ELISA • Targets a specific protein: blocking ELISA • Appears to be as sensitive and more specific than other tests • Expense is a concern

11. One site continuous flow

12. One site AI/AO

13. 3 Site / Multisite

14. PCR • Rapid and specific • Does not depend on viable bacteria • Can be done in live (lung lavages) and dead animals • works well from necropsy and lung lavages ……….. application in the field?

15. Nested PCR • May be done in live animals • Good correlation with serology • It is not quantitative • Should be used at the population level

20. Nested PCR Interpretation

21. N-PCR Interpretation • Nasal Mycoplasma are found only when shed from tracheobronchial sites • Early Infection: PCR positive, probably due to non-attaching and dead Mycoplasmas • Late Infection: Positives due to active shedding • Mid-term Infection: Negatives due to attached Mycoplasma not being carried to nose

22. The future • One-step PCR …….. as a diagnostic profiling test • More rapid • least false positives • N-PCR…use for monitoring clean herds