Proceedings of the 10th International Colloquium on Paratuberculosis

Environmental samples were collected from manure, slurry, and manure scrapers when present.. All
samples collected (n=32) were submitted for culture (6) and PCR. PCR was performed by bead
grinding extraction (5) followed by testing using a commercial kit (Paratuberculosis Adiavet,
Adiagene).
The fields fertilized with <strong>the</strong> contaminated manure/slurry in autumn/winter period were repeatedly
sampled, keeping fixed <strong>the</strong> sampling area, by <strong>the</strong> collection <strong>of</strong>:
• 10 grass samples, in <strong>the</strong> end <strong>of</strong> April;
• 9 field-dried hay samples, before baling, in <strong>the</strong> end <strong>of</strong> May;
• 9 field-dried hay samples, at <strong>the</strong> beginning <strong>of</strong> <strong>the</strong> use for animal feeding, in July-August.
From <strong>the</strong> herd no.. 4, hay samples were not collected, because <strong>the</strong> grass was not used for animal
feeding.
The forage (nearly 800 g <strong>of</strong> fresh grass per sample, or 400 g <strong>of</strong> dried hay per sample) was submitted
to an extraction procedure, by leaving it overnight, after agitation, in sterile distilled water, enough to
wash it completely (nearly 500-1000 ml). From 100 <strong>of</strong> <strong>the</strong> extract liquid we performed culture and from
10 ml we performed bead grinding extraction followed by PCR as done for <strong>the</strong> environmental samples.
RESULTS
The prevalence <strong>of</strong> ELISA positive and clinical cases in <strong>the</strong> 10 herds are shown in Table 1.. The withinherd
sero-prevalence ranged from 4% to 38%. In herd 7, were <strong>the</strong> maximum sero-prevalence value
was recorded (38%), <strong>the</strong> clinical case prevalence was higher than 5%.
Table 1.. Herd dimension and Paratuberculosis diffusion in <strong>the</strong> selected herds
Herd Code Lactating cows ELISA positive cows Sero-prevalence Clinical case incidence
1 24 1 4 % 0 %
2 122 11 9 % 2-5 %
3 88 13 15 % 0-2 %
4 100 14 14 % 0-2 %
5 48 8 17 % 0-2 %
6 75 13 17 % 0-2 %
7 29 11 38 % >5 %
8 48 5 10 % 0 %
9 69 4 6 % 2-5 %
10 53 7 13 % 0-2 %
The results <strong>of</strong> <strong>the</strong> culture and PCR tests performed on manure and slurry are shown in Table 2.
Table 2.. Results <strong>of</strong> <strong>the</strong> tests (PCR and culture) performed on environmental samples
Slurry Manure Scraper
Herd No. PCR Culture No. PCR Culture No. PCR Culture
Code
Positive Positive Positive Positive Positive Positive
1 1 0 0 2 0 0 1 0 0
2 3 3 3 1 1 1 1 1 1
3 1 1 1 1 1 1 1 1 1
4 1 1 0 1 1 1
5 2 2 2 1 1 1 1 1 1
6 1 1 1 1 1 1
7 1 1 1 1 1 0
8 1 0 1 1 0 1 1 1 1
9 1 1 1 1 1 0 1 1 1
10 2 2 0 1 1 0 1 1 0
Total 14 12 (86%) 10 (71%) 9 6 (67%) 4 (44%) 9 8 (89%) 7 (78%)
The environmental samples had high contamination levels, particularly in <strong>the</strong> case <strong>of</strong> liquid slurry and
scrapers.
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