الفهرس 
IntroductionOnly 14 pages are availabe for public view
 |  | from | 174 |  |  |
| | | from | 174 | | |
Abstract
Paratuberculosis or PTB or Johne‘s disease is a chronic mammalian disease,
it is caused by Mycobacterium avium subsp. paratuberculosis (Map). Johne‘s
disease is considered as one of the most serious problems affecting the world‘s
ruminant industry due to significant impact on global economy. It resembles some
clinical and pathological aspects of Crohn‘s disease in humans. The classical
method of diagnosis of PTB takes long incubation period, high costs, and possible
contamination, so the aim of this study was to develop fast and sensitive diagnosis
technique for detection of Map in faecal and milk samples of the same animal
among the Egyptian farms, using different primers and different PCR assays and
comparing between them to find the most sensitive method for detecting the
infection. In addition, we evaluated the effect of Map infection on cattle milk
production.
For this, a total sample of 300 consisting of 150 cattle faeces and 150
individual cow‘s milk were collected randomly from the cattle with suspected and
health status. The samples were assayed using IS900 and F57 primers. We found
that the real-time PCR assay is more sensitive than conventional PCR since it
could detect 1.1x10-8 μg/μl pure DNA while the conventional PCR detected
1.1x10-6 μg/μl using IS900 primer, and we also found that the using F57 primer in
conventional PCR assay have high sensitivity than IS900 primer since it could
detect 1.1x10-13 μg/μl pure DNA. But in clinical samples real-time PCR showed
high sensitive in detection of Map DNA, it detected 42% from total 150 faecal
samples and the conventional PCR detect 31.33% by using IS900 primer set. And
in milk samples real-time detected Map DNA in 5.3%, while the conventional PCR
detected 4.67 % of total 150 milk samples, and by comparing the sensitivity of two
different primers on clinical samples they have shown the same sensitivity. Where
F57 primer detected 28.67% of total 150 faecal samples and 3.33% of total 150
milk samples, showing the same sensitivity with IS900 primer in detecting Map
infection in faecal and milk samples.
The samples were divided according to the healthy statues of the animals and the
results was (34.6%), (32.05%) and (39.7%) of total 77 diseased animals using
IS900, F57 conventional PCR and IS900 real-time PCR, respectively and this
results have been shown that no significant difference in the detection sensitivity in
98
diseased animals between of two primers (IS900 and F57) and different PCR
assays (real-time and conventional PCR) using IS900. But in apparently healthy
animals the results was (27.4%), (26.03%) and (45.21%) of total 73 samples using
IS900, F57 conventional PCR and IS900 real-time PCR, respectively which
appeared a significant different in the sensitivity of the detection of IS900 realtime
PCR for Map than IS900 conventional PCR but no difference between IS900
primer set and F57 Primer set.
The percentage of infection among the three governorates showed that the highest
percentage was in El-Behera then Beni-Suef and the lowest percentage was in El-
Dakahlia.
In the milk samples the detection limit of the two PCR assays showed no
significant difference; also no difference between the two primers that wwere used.
But the high significant difference in detecting the infection was shown between
the faecal and milk samples.
And by taking the history of milk production of the animals we found that the
symptomic animals showed decrease in their milk production that ranged from
2.14% to 69.4%, while asymptomic animals ranged from 2% to 25%. The high
reduction in milk production was found in El-Behera then in Beni-Suef and then
El-Dakahlia.