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A Study On The Incidence Of Zoonotic Tuberculosis To Assess The Associated Risk Factors And Zoonotic Potential Of Bovine Tuberculosis In Lahore

By: Syeda Anum Hadi (2013-VA-04) | Dr. Hassan Mushtaq.
Contributor(s): Dr. Abdul Majeed Akhtar | Professor Dr.Mansur-ud-din Ahmad | Dr. Aamir Gafoor Bajwa.
Material type: materialTypeLabelBookPublisher: 2015Description: 68p.Subject(s): Department of Epidemiology and Public HealthDDC classification: 2321-T Dissertation note: In the review by O’Reiley and his colleagues, Tuberculosis has been defined as a disease that affects the respiratory system foremost and its route of transmission from one animal species to another is by the airborne route along with consumption of un-pasteurized milk (O'Reilly, 1995) (De la Rua-Domenech, 2006) (Thoen et al. 2006). The review states that Mycobacteriumbovis causes tuberculosis in bovines as well as a number of wild animals such as goats, cats, dogs, pigs, buffalo, badgers, possums, deer, bison and non-human primates but most importantly it causes tuberculosis in humans. This makes the disease of significant public health importance due to its zoonotic nature. The study was conducted in two of the largest dairy colonies in Lahore- Rakhchandra and Harbanspura dairy colony. 400 dairy animals (lactating) were selected from the target areas. 200 animals per field were chosen through convenience sampling. The research was divided into two parts. Phase 1 was concerned with screening of animals for bovine tuberculosis through performance of comparative intradermal tuberculin test (CIDTT) and followed by culturing of milk samples from animals that came positive. Phase 2 was concerned with testing of all human subjects who were in contact with the positively screened livestock. Since none of the human subjects showed any of the signs for tuberculosis, no testing of the humans could be performed. The first step to animal testing was concerned with the screening of selected animals with comparative cervical intradermal tuberculin test (CIDTT). This involved the intradermal injection of bovine tuberculin purified protein derivative (PPD) and the subsequent detection of swelling (delayed hypersensitivity) at the site of injection 72 hours later (Anonyms, 2008a). The test was considered positive if the difference between the swellings on the two sites was more than 4mm and it was the mammalian site that showed more swelling. Once the results were read, the dairy farmers were asked a set of questions designed to identify risk-factors for zoonotic tuberculosis. The farmers responded to nearly all the questions that were posed to them. Milk sample was collected from the animals that tested positive. 50 ml of milk was collected from the positive animals. Once collected, the milk bottles were quickly capped and labeled and put in the ice-box before being transported to Provincial Tuberculosis Reference Laboratory in Lahore. Staining followed by culturing of milk samples for the isolation of Mycobacterium bovis was then proceeded with. For the purpose of culturing two types of media were prepared before-hand-Lowenstein Jensen (LJ) media and LJ-pyruvate media. LJ medium allows the growth of Mycobacterium tuberculosis, whereas LJ with pyruvate medium allows the growth of Mycobacterium bovis. Petroff’s method was employed for the processing of milk samples which originally is used for sputum processing (Anonyms, 2009). The process was altered to suit our requirements. Once processed 300ul of pipette tips were used to place 100ul of processed sample on pre-marked slides for ziehl-neelson staining and 120ul on pre-made media slants a total of 4 bottles, 2 each of LJ media and LJ pyruvate media for duplication of results and to act as control. The bottles were checked for growth every week on Monday till 8 weeks of time. At Rakhchandra dairy colony the tuberculin test done on 200 animals revealed only three (3) positive animals. Thus the prevalence of TB in Rakhchandra came out to be 1.5%. Out of 200 animals in Harbanspura dairy colony, six (6) animals showed hypersensitivity reaction and were positive. Prevalence of TB in Harbanspura came out to be 3%. Out of 400 animals tested, 90 were cattle and 310 were buffaloes. Only buffaloes showed hypersensitivity reaction to tuberculin. None of the cows tested came out to be tuberculin positive. In this particular study, the prevalence of TB on the basis of tuberculin test in buffalo was 2.9% where as in cattle it was 0%. When milk was collected and processed from the above mentioned nine (9) animals, the results showed a different picture. None of the cultures showed any signs of growth by 8 weeks of incubation. All nine milk samples after cleaning were stained by ZN staining and observed under microscope for the presence of mycobacterium, none came out positive. The Basic Health Units (BHU) in each of the colony were contacted and it was found that in the last 10 years less than 10 patients who were suspected to have tuberculosis were referred to District Health Quarter (DHQ). Even though a higher percentage (44.44%) of farmers in Harbanspura was recorded to have some knowledge about the zoonotic aspect of tuberculosis as compare to those in Rakhchandra (22.22%), yet a higher number of tuberculin positive animals was found in Harbanspura (6 versus 3). The economic status of farmers in Harbanspura was comparatively higher with 33.33% of farmers earning more than 1 lakh rupees per month, whereas in Rakhchandra this figure stood at 27.78%. This might be a mere chance of co-incidence but it also implies the unwillingness of farmers to apply biosecurity measures at their farms. Lack of willingness to take such precautionary steps places the farmers and their animals in great peril, since in the last six months alone 66.67% of the farmers in Harbanspura had purchased at least one animal, which is enough to bring disease in an un-infected herd. Only 27.78% of farmers in Rakhchandra had purchased animals on the other hand. Also only 77.78% of farmers in Harbanspura would clean the dung from the farms twice a day whereas 100% Rakhchandra farmers would cleanup twice a day. The tuberculin positive animals were found to be spending most time of their day in filthy places. Their sheds were not cleaned regularly. Heaps of dung and ground wet with urine was observed on every visit. It exposed animals to numerous infections and 11.11% of animals in Harbanspura and 44.44% of animals in Rakhchandra were suffering from unidentified chronic illnesses. Farmers said that they preferred to sell such animals to butchers (85.8% combined percentage), rather than burying after culling (3.7% combined percentage). Deworming was not considered a mode of disease prevention amongst the farmers since only 22.22% of all farmers bothered to deworm their animals. The animals were seen to not having a score of above 2.5 when their body scoring was done (Scale 1-5). The one blissful factor discovered was the habit of nearly all farmers (92.59%) preferred to boil milk before consumption. Even dairy products were made from boiled milk (81.48%). This single factor could be the reason why the farmers consuming otherwise contaminated milk was still in such a glowing healthy condition. The study allowed us to get a measure of the status of disease in lactating animals and to investigate the conditions that prevail in the two dairy colonies. It showed a difference in the prevalence of disease in Harbanspura and Rakhchandra famous for providing milk to Lahore city. This was scrutinized through a detailed analysis of farmer habits and environment of animals in both the fields. This study would permit upcoming researchers to have an up-to-date status of tuberculosis in the dairy colonies.
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In the review by O’Reiley and his colleagues, Tuberculosis has been defined as a disease that affects the respiratory system foremost and its route of transmission from one animal species to another is by the airborne route along with consumption of un-pasteurized milk (O'Reilly, 1995) (De la Rua-Domenech, 2006) (Thoen et al. 2006). The review states that Mycobacteriumbovis causes tuberculosis in bovines as well as a number of wild animals such as goats, cats, dogs, pigs, buffalo, badgers, possums, deer, bison and non-human primates but most importantly it causes tuberculosis in humans. This makes the disease of significant public health importance due to its zoonotic nature.
The study was conducted in two of the largest dairy colonies in Lahore- Rakhchandra and Harbanspura dairy colony. 400 dairy animals (lactating) were selected from the target areas. 200 animals per field were chosen through convenience sampling. The research was divided into two parts. Phase 1 was concerned with screening of animals for bovine tuberculosis through performance of comparative intradermal tuberculin test (CIDTT) and followed by culturing of milk samples from animals that came positive. Phase 2 was concerned with testing of all human subjects who were in contact with the positively screened livestock. Since none of the human subjects showed any of the signs for tuberculosis, no testing of the humans could be performed.
The first step to animal testing was concerned with the screening of selected animals with comparative cervical intradermal tuberculin test (CIDTT). This involved the intradermal injection of bovine tuberculin purified protein derivative (PPD) and the subsequent detection of swelling (delayed hypersensitivity) at the site of injection 72 hours later (Anonyms, 2008a). The test was considered positive if the difference between the swellings on the two sites was more than 4mm and it was the mammalian site that showed more swelling. Once the results were read, the dairy farmers were asked a set of questions designed to identify risk-factors for zoonotic tuberculosis. The farmers responded to nearly all the questions that were posed to them.
Milk sample was collected from the animals that tested positive. 50 ml of milk was collected from the positive animals. Once collected, the milk bottles were quickly capped and labeled and put in the ice-box before being transported to Provincial Tuberculosis Reference Laboratory in Lahore. Staining followed by culturing of milk samples for the isolation of Mycobacterium bovis was then proceeded with. For the purpose of culturing two types of media were prepared before-hand-Lowenstein Jensen (LJ) media and LJ-pyruvate media. LJ medium allows the growth of Mycobacterium tuberculosis, whereas LJ with pyruvate medium allows the growth of Mycobacterium bovis.
Petroff’s method was employed for the processing of milk samples which originally is used for sputum processing (Anonyms, 2009). The process was altered to suit our requirements. Once processed 300ul of pipette tips were used to place 100ul of processed sample on pre-marked slides for ziehl-neelson staining and 120ul on pre-made media slants a total of 4 bottles, 2 each of LJ media and LJ pyruvate media for duplication of results and to act as control. The bottles were checked for growth every week on Monday till 8 weeks of time.
At Rakhchandra dairy colony the tuberculin test done on 200 animals revealed only three (3) positive animals. Thus the prevalence of TB in Rakhchandra came out to be 1.5%. Out of 200 animals in Harbanspura dairy colony, six (6) animals showed hypersensitivity reaction and were positive. Prevalence of TB in Harbanspura came out to be 3%. Out of 400 animals tested, 90 were cattle and 310 were buffaloes. Only buffaloes showed hypersensitivity reaction to tuberculin. None of the cows tested came out to be tuberculin positive. In this particular study, the prevalence of TB on the basis of tuberculin test in buffalo was 2.9% where as in cattle it was 0%. When milk was collected and processed from the above mentioned nine (9) animals, the results showed a different picture. None of the cultures showed any signs of growth by 8 weeks of incubation. All nine milk samples after cleaning were stained by ZN staining and observed under microscope for the presence of mycobacterium, none came out positive.
The Basic Health Units (BHU) in each of the colony were contacted and it was found that in the last 10 years less than 10 patients who were suspected to have tuberculosis were referred to District Health Quarter (DHQ).
Even though a higher percentage (44.44%) of farmers in Harbanspura was recorded to have some knowledge about the zoonotic aspect of tuberculosis as compare to those in Rakhchandra (22.22%), yet a higher number of tuberculin positive animals was found in Harbanspura (6 versus 3). The economic status of farmers in Harbanspura was comparatively higher with 33.33% of farmers earning more than 1 lakh rupees per month, whereas in Rakhchandra this figure stood at 27.78%. This might be a mere chance of co-incidence but it also implies the unwillingness of farmers to apply biosecurity measures at their farms. Lack of willingness to take such precautionary steps places the farmers and their animals in great peril, since in the last six months alone 66.67% of the farmers in Harbanspura had purchased at least one animal, which is enough to bring disease in an un-infected herd. Only 27.78% of farmers in Rakhchandra had purchased animals on the other hand. Also only 77.78% of farmers in Harbanspura would clean the dung from the farms twice a day whereas 100% Rakhchandra farmers would cleanup twice a day. The tuberculin positive animals were found to be spending most time of their day in filthy places. Their sheds were not cleaned regularly. Heaps of dung and ground wet with urine was observed on every visit. It exposed animals to numerous infections and 11.11% of animals in Harbanspura and 44.44% of animals in Rakhchandra were suffering from unidentified chronic illnesses. Farmers said that they preferred to sell such animals to butchers (85.8% combined percentage), rather than burying after culling (3.7% combined percentage). Deworming was not considered a mode of disease prevention amongst the farmers since only 22.22% of all farmers bothered to deworm their animals. The animals were seen to not having a score of above 2.5 when their body scoring was done (Scale 1-5). The one blissful factor discovered was the habit of nearly all farmers (92.59%) preferred to boil milk before consumption. Even dairy products were made from boiled milk (81.48%). This single factor could be the reason why the farmers consuming otherwise contaminated milk was still in such a glowing healthy condition.
The study allowed us to get a measure of the status of disease in lactating animals and to investigate the conditions that prevail in the two dairy colonies. It showed a difference in the prevalence of disease in Harbanspura and Rakhchandra famous for providing milk to Lahore city. This was scrutinized through a detailed analysis of farmer habits and environment of animals in both the fields. This study would permit upcoming researchers to have an up-to-date status of tuberculosis in the dairy colonies.

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