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Current IssueEditorial BoardOnline First ArticlesArchive

volume 49 issue 5 (october 2015) : 687-693,   Doi: 10.18805/ijar.5583

Isolation and identification of Mycobacterium bovis and Mycobacterium tuberculosis from animal tissues by conventional and molecular method

A
Abdul Basit
M
Mubbashir Hussain
S
Sultan Ayaz
M
Muhammad Shahid
K
Kashif Rahim
I
Iqbal Ahmad
R
Riaz Ullah
A
Abeer Hashem
E
Elsayed Abd-Allah
A
Abdulaziz Abdullah Alqarawi
N
Naila Gul
1Department of Microbiology, Kohat University of Science and Technology (KUST), Khyber Pakhtunkhwa Kohat, 26000, Pakistan.
Cite article:- Basit Abdul, Hussain Mubbashir, Ayaz Sultan, Shahid Muhammad, Rahim Kashif, Ahmad Iqbal, Ullah Riaz, Hashem Abeer, Abd-Allah Elsayed, Alqarawi Abdullah Abdulaziz, Gul Naila (2025). Isolation and identification of Mycobacterium bovis and Mycobacterium tuberculosis from animal tissues by conventional and molecular method. Indian Journal of Animal Research. 49(5): 687-693. doi: 10.18805/ijar.5583.

ABSTRACT

Tuberculosis is a worldwide contagious and chronic disease of human as well as domestic animals with zoonotic potential. The Mycobacterium bovis and Mycobacterium tuberculosis are the main cause of tuberculosis. It has worldwide distribution with significant effect on animals and has public health importance. Therefore the present study was conducted to determine the prevalence of Mycobacterium bovis and Mycobacterium tuberculosis among the ruminant of district Kohat, Khyber PakhtunKhwa and also to evaluate the sensitivity and specificity of Microscopy and PCR. A total 200 tissue samples of lungs, lymph nodes and liver from cattle, buffaloes, sheep and goats were collected from Abattoir Kohat. All the tissue were first examined by direct smear microscopy by Ziehl Neelsen staining and then subjected to the PCR for the detection of M. bovis and M. tuberculosis. The overall prevalence of tuberculosis was 6.5% by PCR.Prevalence of tuberculosis was recorded in7.87% of lungs samples followed by 5.26% lymph node. Moreover the prevalence was found 5.2%, 4%, 10.6% and 6.5% in cattle, buffalos, Goats and sheep respectively. Furthermorethe sensitivity and specificity of PCR and microscopy in term of detection of TB was determined that PCR was found less sensitive then microscopy because of other species which was not amplified due to non availability of specific primer and were found positive in microscopy.In conclusion PCR is more reliable diagnostic tool for diagnosis of bovine tuberculosis. It is recommended that PCR based diagnostic reference laboratory may be established at district level and Tuberculosis awareness campaign must be arranged.

REFERENCES

  1. Aliyu, M.,Adamu,M.J.Y., and Bilyaminu Y.A. (2009). Current Prevalence of Tuberculosis Lesions among Slaughtered Cattle in Northeastern States of Nigeria. Revue d’elevage et de medecineveterinaire des pays tropicaux, 62: 13-16.
  2. Al-SaqurM, Al-Thwani, A.N.and Al-Attar, I.M. (2009). Detection of Mycobacteria spp in cow’s milk using conventional methods and PCR. Iraqi Journal of Veterinary Sciences, 23: 259-262.
  3. Ashford,D.A., Whitney,E., Raghunathan,P and Cosivi,O. (2001). Epidemiology of selected mycobacteria that infect humans and other animals. Rev. sci. tech. Off. int. Epiz., 20: 325-337.
  4. Bekele, M and Belay, I. (2011). Evaluation of routine meat inspection procedure to detect bovine tuberculosis suggestive lesions in Jimma Municipal Abattoir, South West Ethiopia. Global Veterinaria, 6: 172-179.
  5. Collins, C.H and Grange, J.M. (1994). The bovine tubercule bacillus” Journal of Applied Bacteriology, 55: 13-29.
  6. Corner L.A. (1994) “Post mortem diagnosis of Mycobacterium bovis in cattle. Veterinary Microbiology, 40: 53-63.
  7. Ewnetu, L., Melaku, A. and Birhanu, A. (2012). Bovine Tuberculosis Prevalence in Slaughtered Cattle at Akaki Municipal Abattoir, Based on Meat Inspection Methods. Global Veterinaria 9: 541-545.
  8. Ghumman, M.A., ManzoorA.W., NazS., Ahmad R. Ahmad R. (2013). Prevalence of Tuberculosis in Cattle and Buffalo at Various Livestock Farms in Punjab. International Journal of Veterinary Medicine, Article ID 145084, DOI: 10.5171/ 2013.145084.
  9. Hena,S.A., Tanimomo,B.K., Tarhyel,R., Ngbede,E.O., Oluwasina,O.O., Jagtap,G.P., Dhavale,M.C., and Dey, U. (2012). A tentative case of tuberculosis detected at postmortem examination of animal carcasses slaughtered at some local slaughter slabs in north-western Nigeria. Scientific Journal of Agricultural, 1: 27-32.
  10. Iqbal, M.J., Ali, Z.U. and Ali, S.S. (2012). Agroclimatic modeling for estimation of wheat production in the Punjab province, Pakistan. Proceeding of Pakistan academy of Science, 49: 241- 249.
  11. Ifrahim, M. (2001). Epidemiological studies on tuberculosis in cattle and buffalo population in villages around Faisalabad. (M.Sc. thesis), Department of Veterinary Microbiology, University of Agriculture, Faisalabad.
  12. Javed,M.T., Usman,M., Irfan, M.,Cagiola, M. (2006). A study on tuberculosis in buffaloes: some epidemiological aspects, along with haematological and serum protein changes. VeterinarskiArhiv, 76: 193-206.
  13. KassaG.M., AbebeF., WorkuY., LegesseM., MedhinG., BjuneG., and Ameni G. (2012). Tuberculosis in Goats and Sheep in Afar Pastoral Region of Ethiopia and Isolation of Mycobacterium tuberculosis from Goat. Veterinary Medicine International, 1-8.
  14. Khan.F.A., Chaudhry, Z.I., Ali, M.I., Khan, S., Mumtaz, N and Ahmad, I. (2010). Detection of Mycobacterium avium subsp. Paratuberculosis in tissue samples of cattle and buffaloes. Tropical Animal Health and Production. 42: 633-638.
  15. Khan,I.A., and Khan A. (2007). Prevalence and risk factors of bovine tuberculosis in Nili-Ravi buffaloes in the Punjab, Pakistan. Italian Journal of Animal Science, 6: 817-820.
  16. Khan,I. A., KhanA., Mubarak A.and Ali S. (2008). Factors Affecting Prevalence Of Bovine Tuberculosis In Nili Ravi Buffaloes. Pakistan Veterinary Journal, 28: 155-158.
  17. Muhammad A., Ifrahim,M., AshrafM., Rehman S.U., and Khan H.A. (2012). Epidemiological studies on tuberculosis in buffalo population in villages around Faisalabad. The Journal of Animal and Plant Sciences, 22: 246 – 249.
  18. Ndugga, K.L., Cleeff, M.V., Juma, E., Kimwomi, J., Githuui,W., oskam, L., Schuittema, A., Soolingen, D.V., Nganga, L., Kibuga, D., Odhiambo, J.and Klatser, P. (2004), Comparison of PCR with routine procedure for diagnosis of tuberculosis in population with high prevalence of tuberculosis and human immunodeficiency virus. Journal of Clinical Microbiology, 42, 1012-1015.
  19. Quinn,P.J., Carter,M.E., Markey, B.K. and Carter, S.G.R. (1994). Clinical veterinary microbiology. (Wolfe, London), 144-155.
  20. Ramadan, H. H., El-Gohary, A.H.N., Mohamed, A.A. and Nasr, E.A. (2012). Detection of Mycobacterium Bovis and Mycobacterium Tuberculosis from Clinical Samples by Conventional and Molecular Techniques in Egypt. Global Veterinaria, 9 : 648-654.
  21. Shirtaye, J. E., Getahun, B., Alemayehu, T., (2006). A prevalence study of bovine tuberculosis by using abattoir meat inspection and tuberculin skin testing data, histopathological and IS1610 PCR examination of tissues with tuberculous lesions in cattle in Ethiopia. Veterinary Medicine, 51: 512-522.
  22. Shitaye, J.E., Getahun, B., Alemayehu, T., SkoricM., Treml, F., Fictum, P., Vrbas,V., Pavlik, I. (2006). A prevalence study of bovine tuberculosis by using abattoir meat inspection and tuberculin skin testing data, histopathological and IS 6110 PCR examination of tissues with tuberculosis lesions in cattle in Ethiopia. VeterinarniMedicina, 51: 512–522.
  23. Teklue, A., Asseged, B., Yimer, E., Gebeyehu, M.,Woldesenbet, Z. (2004). Tuberculosis lesions not detected by routine abattoir inspection: the experience of the Hossana municipal abattoir, southern Ethiopia. Rev. Sci. Tech. Off. Int. Epiz., 23: 957-964.
  24. Tenguria, R.K., Khan, F.N., Pandey, S.Q.A. (2011). Epidemiological study of zoonotic Tuberculosis complex (ZTBC). World Journal of Science and Technology, 1: 31-56.
  25. Thoen, C.O., Lobue, P.A,.Enarson, D.A., Kaneene, J.B., and de Kantor, I.N. (2009). A re-emerging disease in animals and humans. Veterinariaitaliana, 45: 135-181.
  26. Tschopp, R., Aseffa, A., Schelling, E., Berg, S., Hailu, E., Gadisa, E., Habtamu, M., Argaw, K. and Zinsstag, J. (2010). Bovine Tuberculosis at the wildlife livestock-Human interface in Hamer Woreda, South Omo, Southern Ethiopia. PLoS ONE, 5: 1-7.
  27. Tschopp, R., Bobosha, K., Aseffa, A., Schelling, E., Habtamu, M., Iwnetu, R., Hailu, E., Firdessa, R., Hussein, J., Young, D., and Zinsstag, J. (2011). Bovine tuberculosis at a cattle-small ruminant-human interface in Meskan, Gurage region,Central Ethiopia. BMC Infectious Diseases. 11:318.
  28. Victor, T., Dutoit, R., and Vanhelden, P.D. (1992). Purification of Sputum Samples through Sucrose Improves Detection of Mycobacterium tuberculosis by Polymerase Chain Reaction. Journal Of Clinical Microbiology. 1514-1517.
Published In
Indian Journal of Animal Research
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    volume 49 issue 5 (october 2015) : 687-693,   Doi: 10.18805/ijar.5583

    Isolation and identification of Mycobacterium bovis and Mycobacterium tuberculosis from animal tissues by conventional and molecular method

    A
    Abdul Basit
    M
    Mubbashir Hussain
    S
    Sultan Ayaz
    M
    Muhammad Shahid
    K
    Kashif Rahim
    I
    Iqbal Ahmad
    R
    Riaz Ullah
    A
    Abeer Hashem
    E
    Elsayed Abd-Allah
    A
    Abdulaziz Abdullah Alqarawi
    N
    Naila Gul
    1Department of Microbiology, Kohat University of Science and Technology (KUST), Khyber Pakhtunkhwa Kohat, 26000, Pakistan.
    Cite article:- Basit Abdul, Hussain Mubbashir, Ayaz Sultan, Shahid Muhammad, Rahim Kashif, Ahmad Iqbal, Ullah Riaz, Hashem Abeer, Abd-Allah Elsayed, Alqarawi Abdullah Abdulaziz, Gul Naila (2025). Isolation and identification of Mycobacterium bovis and Mycobacterium tuberculosis from animal tissues by conventional and molecular method. Indian Journal of Animal Research. 49(5): 687-693. doi: 10.18805/ijar.5583.

    ABSTRACT

    Tuberculosis is a worldwide contagious and chronic disease of human as well as domestic animals with zoonotic potential. The Mycobacterium bovis and Mycobacterium tuberculosis are the main cause of tuberculosis. It has worldwide distribution with significant effect on animals and has public health importance. Therefore the present study was conducted to determine the prevalence of Mycobacterium bovis and Mycobacterium tuberculosis among the ruminant of district Kohat, Khyber PakhtunKhwa and also to evaluate the sensitivity and specificity of Microscopy and PCR. A total 200 tissue samples of lungs, lymph nodes and liver from cattle, buffaloes, sheep and goats were collected from Abattoir Kohat. All the tissue were first examined by direct smear microscopy by Ziehl Neelsen staining and then subjected to the PCR for the detection of M. bovis and M. tuberculosis. The overall prevalence of tuberculosis was 6.5% by PCR.Prevalence of tuberculosis was recorded in7.87% of lungs samples followed by 5.26% lymph node. Moreover the prevalence was found 5.2%, 4%, 10.6% and 6.5% in cattle, buffalos, Goats and sheep respectively. Furthermorethe sensitivity and specificity of PCR and microscopy in term of detection of TB was determined that PCR was found less sensitive then microscopy because of other species which was not amplified due to non availability of specific primer and were found positive in microscopy.In conclusion PCR is more reliable diagnostic tool for diagnosis of bovine tuberculosis. It is recommended that PCR based diagnostic reference laboratory may be established at district level and Tuberculosis awareness campaign must be arranged.

    REFERENCES

    1. Aliyu, M.,Adamu,M.J.Y., and Bilyaminu Y.A. (2009). Current Prevalence of Tuberculosis Lesions among Slaughtered Cattle in Northeastern States of Nigeria. Revue d’elevage et de medecineveterinaire des pays tropicaux, 62: 13-16.
    2. Al-SaqurM, Al-Thwani, A.N.and Al-Attar, I.M. (2009). Detection of Mycobacteria spp in cow’s milk using conventional methods and PCR. Iraqi Journal of Veterinary Sciences, 23: 259-262.
    3. Ashford,D.A., Whitney,E., Raghunathan,P and Cosivi,O. (2001). Epidemiology of selected mycobacteria that infect humans and other animals. Rev. sci. tech. Off. int. Epiz., 20: 325-337.
    4. Bekele, M and Belay, I. (2011). Evaluation of routine meat inspection procedure to detect bovine tuberculosis suggestive lesions in Jimma Municipal Abattoir, South West Ethiopia. Global Veterinaria, 6: 172-179.
    5. Collins, C.H and Grange, J.M. (1994). The bovine tubercule bacillus” Journal of Applied Bacteriology, 55: 13-29.
    6. Corner L.A. (1994) “Post mortem diagnosis of Mycobacterium bovis in cattle. Veterinary Microbiology, 40: 53-63.
    7. Ewnetu, L., Melaku, A. and Birhanu, A. (2012). Bovine Tuberculosis Prevalence in Slaughtered Cattle at Akaki Municipal Abattoir, Based on Meat Inspection Methods. Global Veterinaria 9: 541-545.
    8. Ghumman, M.A., ManzoorA.W., NazS., Ahmad R. Ahmad R. (2013). Prevalence of Tuberculosis in Cattle and Buffalo at Various Livestock Farms in Punjab. International Journal of Veterinary Medicine, Article ID 145084, DOI: 10.5171/ 2013.145084.
    9. Hena,S.A., Tanimomo,B.K., Tarhyel,R., Ngbede,E.O., Oluwasina,O.O., Jagtap,G.P., Dhavale,M.C., and Dey, U. (2012). A tentative case of tuberculosis detected at postmortem examination of animal carcasses slaughtered at some local slaughter slabs in north-western Nigeria. Scientific Journal of Agricultural, 1: 27-32.
    10. Iqbal, M.J., Ali, Z.U. and Ali, S.S. (2012). Agroclimatic modeling for estimation of wheat production in the Punjab province, Pakistan. Proceeding of Pakistan academy of Science, 49: 241- 249.
    11. Ifrahim, M. (2001). Epidemiological studies on tuberculosis in cattle and buffalo population in villages around Faisalabad. (M.Sc. thesis), Department of Veterinary Microbiology, University of Agriculture, Faisalabad.
    12. Javed,M.T., Usman,M., Irfan, M.,Cagiola, M. (2006). A study on tuberculosis in buffaloes: some epidemiological aspects, along with haematological and serum protein changes. VeterinarskiArhiv, 76: 193-206.
    13. KassaG.M., AbebeF., WorkuY., LegesseM., MedhinG., BjuneG., and Ameni G. (2012). Tuberculosis in Goats and Sheep in Afar Pastoral Region of Ethiopia and Isolation of Mycobacterium tuberculosis from Goat. Veterinary Medicine International, 1-8.
    14. Khan.F.A., Chaudhry, Z.I., Ali, M.I., Khan, S., Mumtaz, N and Ahmad, I. (2010). Detection of Mycobacterium avium subsp. Paratuberculosis in tissue samples of cattle and buffaloes. Tropical Animal Health and Production. 42: 633-638.
    15. Khan,I.A., and Khan A. (2007). Prevalence and risk factors of bovine tuberculosis in Nili-Ravi buffaloes in the Punjab, Pakistan. Italian Journal of Animal Science, 6: 817-820.
    16. Khan,I. A., KhanA., Mubarak A.and Ali S. (2008). Factors Affecting Prevalence Of Bovine Tuberculosis In Nili Ravi Buffaloes. Pakistan Veterinary Journal, 28: 155-158.
    17. Muhammad A., Ifrahim,M., AshrafM., Rehman S.U., and Khan H.A. (2012). Epidemiological studies on tuberculosis in buffalo population in villages around Faisalabad. The Journal of Animal and Plant Sciences, 22: 246 – 249.
    18. Ndugga, K.L., Cleeff, M.V., Juma, E., Kimwomi, J., Githuui,W., oskam, L., Schuittema, A., Soolingen, D.V., Nganga, L., Kibuga, D., Odhiambo, J.and Klatser, P. (2004), Comparison of PCR with routine procedure for diagnosis of tuberculosis in population with high prevalence of tuberculosis and human immunodeficiency virus. Journal of Clinical Microbiology, 42, 1012-1015.
    19. Quinn,P.J., Carter,M.E., Markey, B.K. and Carter, S.G.R. (1994). Clinical veterinary microbiology. (Wolfe, London), 144-155.
    20. Ramadan, H. H., El-Gohary, A.H.N., Mohamed, A.A. and Nasr, E.A. (2012). Detection of Mycobacterium Bovis and Mycobacterium Tuberculosis from Clinical Samples by Conventional and Molecular Techniques in Egypt. Global Veterinaria, 9 : 648-654.
    21. Shirtaye, J. E., Getahun, B., Alemayehu, T., (2006). A prevalence study of bovine tuberculosis by using abattoir meat inspection and tuberculin skin testing data, histopathological and IS1610 PCR examination of tissues with tuberculous lesions in cattle in Ethiopia. Veterinary Medicine, 51: 512-522.
    22. Shitaye, J.E., Getahun, B., Alemayehu, T., SkoricM., Treml, F., Fictum, P., Vrbas,V., Pavlik, I. (2006). A prevalence study of bovine tuberculosis by using abattoir meat inspection and tuberculin skin testing data, histopathological and IS 6110 PCR examination of tissues with tuberculosis lesions in cattle in Ethiopia. VeterinarniMedicina, 51: 512–522.
    23. Teklue, A., Asseged, B., Yimer, E., Gebeyehu, M.,Woldesenbet, Z. (2004). Tuberculosis lesions not detected by routine abattoir inspection: the experience of the Hossana municipal abattoir, southern Ethiopia. Rev. Sci. Tech. Off. Int. Epiz., 23: 957-964.
    24. Tenguria, R.K., Khan, F.N., Pandey, S.Q.A. (2011). Epidemiological study of zoonotic Tuberculosis complex (ZTBC). World Journal of Science and Technology, 1: 31-56.
    25. Thoen, C.O., Lobue, P.A,.Enarson, D.A., Kaneene, J.B., and de Kantor, I.N. (2009). A re-emerging disease in animals and humans. Veterinariaitaliana, 45: 135-181.
    26. Tschopp, R., Aseffa, A., Schelling, E., Berg, S., Hailu, E., Gadisa, E., Habtamu, M., Argaw, K. and Zinsstag, J. (2010). Bovine Tuberculosis at the wildlife livestock-Human interface in Hamer Woreda, South Omo, Southern Ethiopia. PLoS ONE, 5: 1-7.
    27. Tschopp, R., Bobosha, K., Aseffa, A., Schelling, E., Habtamu, M., Iwnetu, R., Hailu, E., Firdessa, R., Hussein, J., Young, D., and Zinsstag, J. (2011). Bovine tuberculosis at a cattle-small ruminant-human interface in Meskan, Gurage region,Central Ethiopia. BMC Infectious Diseases. 11:318.
    28. Victor, T., Dutoit, R., and Vanhelden, P.D. (1992). Purification of Sputum Samples through Sucrose Improves Detection of Mycobacterium tuberculosis by Polymerase Chain Reaction. Journal Of Clinical Microbiology. 1514-1517.
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