1. RAW MILK CONSUMPTION : A CAUSE FOR CROHN'S DISEASE ????BY
Dr.Bharathy.S
M.V.Sc Scholar
Department of Veterinary
Public Health and Epidemiology
Madras Veterinary College

2. CROHN’S DISEASE - CAUSES
Crohn's disease, a chronic granulomatous, incurable ileocolitis of humans, has significant similarity to the Paratuberculosis in animals.
Current concepts:
-Dysfunction of the immune system
-Genetic disorder
-Environmental factor
-Escherichia coli.
-Mycobacterium avium subspecies paratuberculosis
(MAP)

3. MAP and Crohn’s disease
The pathological changes in intestines of Crohn’s disease is similar to Johne’s disease in cattle.
MAP cultured from Crohn’s disease affected individuals intestine.
MAP is shed subclinically and clinically infected animals into manure and in milk.
Subclinical shedders are the main problem and act as a source for new infections.

4. Objectives To Detect MAP by PCR in raw milk samples.
To understand the excretory pattern of MAP in milk.

5. Materials and methods for identification of MAP DNA in raw milk
Sample collection:
45 lactating cow and 32 lactating does .
From organized farm and Madras Veterinary College clinics .
10ml of samples collected equally from all the quarters of each lactating animals.
Samples were kept at 4 ˚C until further processing.

6. DNA Extraction from milk sample
10 ml of milk sample in sterile tube
Centrifuged 3000rpm for 10 min
Hardened fat layer,
was aspirated and discarded
Save 0.5 ml sediment
DNA extraction by commercial kit

7. Polymerase chain reaction
IS900 PCR was performed
Forward primer: 5’-CCGCTAATTGAGAGATGCGATTGG-3’
Reverse primer: 5’AATCAA CTCCAGCAGCAGCGCGGCCTCG-3’
Events
Temperature
(˚C)
Time
(minutes)
Initial denaturation 94 3
Denaturation 1
Annealing 55
Extension 72
Final extension 7
Total -35 cycles

8. PCR Products Were Visualized By 1.5 % Agarose Gel .
Goat milk samples
M L L L L4 L L6
229bp
M bp DNA ladder
L1 to L6 - Goat milk samples
Cow milk samples
L1 L M L L4 L5 L6 L L8
L Negative control
L Positive control
M bp DNA ladder
L3 To L Cow milk samples
229bp

9. RESULTS BASED ON PCR
Based on species wise
84.42%
11.11%

10. Conclusion In this study 12 (15.58%) showed IS900 PCR positivity.
IS900 PCR-positive results indicates presence of subclinical MAP infections in dairy animals .
In our study percentage were slightly higher.
Results from these studies indicate that transmission of MAP from animals to humans via raw milk consumption.
So there is a need to minimize the potential threat of this organism as a zoonotic agent of Crohn’s disease??????

11. What can be done???
Reduce public health risk by means of strengthening collaboration.
Prohibit the use of milk derived from animals with clinical Johne’s disease.
Further optimization of pasteurization time and temperature.
Control measures.
Awareness.