Comparative Clinical Pathology

, Volume 28, Issue 2, pp 549–555 | Cite as

Comparative diagnosis of Mycobacterium avium subspecies paratuberculosis in the tissues of clinical and subclinical sheep of paratuberculosis endemic farm

  • G. G. SonawaneEmail author
  • B. N. Tripathi
Original Article


Paratuberculosis (Johne’s disease) is the chronic infectious granulomatous enteritis of ruminants caused by Mycobacterium avium subspecies paratuberculosis. Due to the lack of definite diagnostic tests, the early detection of the disease is difficult. The present study was undertaken to assess comparative efficacy of diagnostic methods routinely used in the identification and confirmation of Mycobacterium avium subspecies paratuberculosis (MAP) in the tissue samples of naturally infected clinical and subclinical sheep. The ileum and mesenteric lymph node (MLN) tissues were collected from 20 paratuberculous sheep from organized farm of Rajasthan. These animals were further classified as paucibacillary (PB) (n = 8) or multibacillary (MB) (n = 12) on the basis of histopathological findings and mycobacterial loads. The ileum and MLN sections from PB and MB and uninfected control groups were tested by indirect immunoperoxidase technique (IPT), Ziehl Neelsen’s (ZN) method, bacterial culture, IS900 PCR and 251 gene PCR. In PB sheep, IPT and ZN were positive in 87.5 and 50% of intestinal sections and 75 and 37.5% of MLN sections, respectively. In MB sheep, IPT and ZN had equal sensitivity. The overall sensitivity of IPT was found to be superior (95%) than ZN method (80%) in demonstration of acid-fast bacteria in intestinal tissues. On the bacterial cultural examination of the intestinal and MLN tissues using Herrold’s egg yolk medium, MAP was isolated in eight (66.6%) MB and in four (50%) PB sheep. Among 20 sheep, IS900 PCR detected 13 (65.0%) and 251 gene PCR detected 16 (80%) sheep as positive for MAP. In the PB sheep, the culture, ZN test and PCR sensitivity was found low in comparison to MB sheep. The sensitivity of IPT in PB sheep was better than other tests in detecting the MAP infection, which underlines its utility in confirmation of subclinical and clinical cases of paratuberculosis in sheep. The sensitivity of 251 gene PCR assay was found better than IS900 gene PCR and has potential to be used as screening test for clinical and subclinical paratuberculosis in sheep flocks.


Comparative diagnosis Paratuberculosis Sheep Endemic farm 



We are thankful to the Director IVRI, Izatnagar, Bareilly (UP) and the Director, CSWRI, Avikanagar, Rajasthan for providing necessary help and facility to carry out this study.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.

Ethical statement

All animal experimental procedures were performed with approval of Institute Animal Ethics Committee (IAEC), ICAR-CSWRI, Avikanagar constituted by Committee for the Purpose of Control and Supervision of Experiments on Animals (CPCSEA) India. All applicable institutional guidelines for the care and use of animals were followed.

Informed consent

Informed consent was obtained from all individual participants included in the study.

Statement on human and animal rights

All institutional and national guidelines for the care and use of laboratory animals were followed.


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Copyright information

© Springer-Verlag London Ltd., part of Springer Nature 2018

Authors and Affiliations

  1. 1.ICAR-Central Sheep and Wool Research Institute, AvikanagarDistt-TonkIndia
  2. 2.ICAR-National Centre on EquinesHisarIndia

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