Histopathological Investigations and Molecular Confirmation Reveal Mycobacterium bovis in One-Horned Rhinoceros (Rhinoceros unicorns)

Abstract

Mycobacterium bovis causes tuberculosis in dairy and wild animals. Presence of tuberculosis in animals poses a threat not only to their herd mates but also for public. No reports are available about the clinical, pathological, and molecular investigation of naturally occurring tuberculosis (TB) due to M. bovis in one-horned rhinoceros. One-horned female rhinoceros (Rhinoceros unicorns) at the age of 41 years died in a public park in Pakistan. Postmortem and other investigations were carried out to know the cause of death. The present study describes necropsy, histopathology, and molecular-based confirmation of TB in a captive female rhinoceros that died of this infection. Clinically, the rhinoceros showed nonspecific clinical signs including anorexia, lethargy, dyspnoea, coughing, and sudden death. At necropsy, the trachea exhibited mild congestion and contained catarrhal exudate at the bronchial bifurcation. Macroscopic examination revealed characteristic tubercles on all parenchymatous organs. The lungs showed consolidation, grey hepatization, and contained granulomatous lesions packed with cheesy exudate. Histopathological examination showed severe pneumonic changes in the form of granulomatous inflammation consisting of lymphocytes, multinucleated giant cells, caseous materials, and mineralized foci surrounded by a fibrous capsule. PCR amplicon of 500 bp confirmed the presence of M. bovis in multiple hepatic and pulmonary tissue samples, as well as in uterine exudates. It was concluded that the presence of tuberculosis in rhinoceros may pose potential transmission risk to other animals and the application of practical tools to determine TB status in the rhinoceros is crucial.

Figure 1

Photographs of one-horned female rhinoceros died of M. bovis infection. (a) Showing blood mixed exudate coming out of the vagina (white arrow), (b) uterine pus mixed with tissue debris (blue arrow), (c) small multifocal tubercular lesions (arrowheads) containing creamy white exudate in the lungs, and (d) small multifocal tubercular lesions (arrowheads) on the parietal surface of liver (arrowheads).

Figure 2

Photomicrographs of lungs of one-horned female rhinoceros died of M. bovis infection showing (a) thickened alveolar septa (double-sided arrows), and alveoli filled with fibrino-necrotic edema (fne) fluid along with massive infiltration of inflammatory cells, (b) micronodules with minimum calcified exudate surrounded by fronts of fibroblasts, fibrocytes, monocytes, macrophages (arrows), multinucleated giant cells, and calcified centre (cc) along with inflammatory cells infiltration in bronchioles (if), (c) two calcified (ca) tubercle nodules surrounded by fronts of fibroblasts, fibrocytes, monocytes, macrophages (arrows), and (d) calcified (ca) centre. H & E; magnification: (a)–(c) 100X; (d) -400X.

Figure 3

Photomicrograph of lung tissues of rhinoceros died of M. bovis infection showing (a) extensive fibrosis, presence of bacilli (arrows), and hyperplasia of pneumocytes. H & E; 400X. (b) Confirmation of Mycobacterium bovis by PCR (500 bp) from lungs (5), liver (6), and uterine pus (7). Lane 1 shows DNA marker (50 bp), lane 3 positive samples and lane 2 and 4 show negative control.