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International Journal of Current Microbiology and Applied Sciences (IJCMAS)
IJCMAS is now DOI (CrossRef) registered Research Journal. The DOIs are assigned to all published IJCMAS Articles.
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National Academy of Agricultural Sciences (NAAS)
NAAS Score: *5.38 (2020)
[Effective from January 1, 2020]
For more details click here

ICV 2018: 95.39
Index Copernicus ICI Journals Master List 2018 - IJCMAS--ICV 2018: 95.39
For more details click here

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Original Research Articles

PRINT ISSN : 2319-7692
Online ISSN : 2319-7706
Issues : 12 per year
Publisher : Excellent Publishers
Email : editorijcmas@gmail.com / submit@ijcmas.com
Editor-in-chief: Dr.M.Prakash
Index Copernicus ICV 2018: 95.39
NAAS RATING 2020: 5.38

Int.J.Curr.Microbiol.App.Sci.2020.9(5): 1731-1738
DOI: https://doi.org/10.20546/ijcmas.2020.905.194


Diagnosis of Paratuberculosis by Fecal Culture Followed  by Nested-Polymerase Chain Reaction
Anand Mohan*, P. Das and N. Kushwaha
Biological Product Division, Indian Veterinary Research Institute, Izatnagar, India
*Corresponding author
Abstract:

Culture of clinical samples is the confirmatory test for the diagnosis of paratuberculosis but, it is time consuming due to its long incubation period, further it requires a skilled personnel to monitor the growth. Use of molecular techniques such as PCR is advantageous over the long incubation period in culturing of samples; more than eight months of incubation is needed to declare a sample to be negative. Two cattle farms (Cattle rehabilitation center and organized dairy farm) with different organizational setup were screened by culture of fecal samples. Ziehl-Neelsen (ZN) staining and nested-PCR were implied to confirm the growths on these samples after two months of inoculation on Herrold’s Egg Yolk Medium (HYM). Overall, 33.67% samples confirmed the growth of Map bacilli and 25.00% samples confirmed the growth of acid-fast bacilli. In cattle rehabilitation center 50.00% samples confirmed the growth of Map bacilli and 35.63% shown to have acid-fast bacilli, whereas in the organized dairy farm it was 15.00% and 13.57%, respectively. Confirmation of growth of bacilli in samples belongs to age groups 0 - < 3 yrs and 3 - < 6 yrs were indicated, approximately same level of infection (c2; p>0.05), but significantly higher level of infection were confirmed in age group of ≥ 6 yrs (c2; p<0.05). Appreciable colonies were identified after the incubation of 7 to 9 months on cultures positive in both ZN stain and nested-PCR, but cultures negative in nested-PCR were failed to produce colonies even after incubation of one year. Study confirmed that variation in level of paratuberculosis in these two cattle farm with possible organizational set up influences. Present study concludes that, use of nested-PCR in conjunction with fecal culture reduce long incubation period to confirm Map bacilli and high percentage of paratuberculosis positive animals in old age group in both farms. 


Keywords: Mycobacterium avium subspecies paratuberculosis, Nested-PCR, Ziehl-Neelsen staining
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How to cite this article:

Anand Mohan, P. Das and N. Kushwaha. 2020. Diagnosis of Paratuberculosis by Fecal Culture Followed by Nested-Polymerase Chain Reaction.Int.J.Curr.Microbiol.App.Sci. 9(5): 1731-1738. doi: https://doi.org/10.20546/ijcmas.2020.905.194