Table 1.
Distribution of granulomatous lesions in cattle naturally infected with M. bovi.
Fig 1.
Extensive lesions suggestive of tuberculosis were observed in lymph nodes from calves naturally infected with M. bovis and compared to lesions found in adult cattle.
(A) Mediastinal lymph node with severe granulomatous lymphadenitis and (B) H&E, 40x showing a granuloma encapsulated by connective tissue and abundant necrosis and mineralization from a 3-year-old Holstein-Friesian dairy cow. (C) Mediastinal lymph node with extensive areas of caseous necrosis and microhemorrhages and (D) H&E, 40x showing a granuloma without connective tissue and abundant necrosis and mineralization from a one-month-old Holstein-Friesian calf.
Fig 2.
Calves had a greater number of granulomatous lesions and a higher bacilli scale when compared to adults.
(A) Average microscopic lesions in cattle from both groups. (B) Granulomas with more than 50 bacilli are frequently seen in calves. (C) A lower average of giant cells is observed in calf lesions. (D) H&E, 40x, showing abundant giant cells (arrows) located between necrosis and fibrous capsule of a stage IV granuloma in adult cattle. (E) H&E, 400x, showing giant cells (arrows) close to an area with calcium deposits within a calf granuloma. Zielh Neelsen Stain (ZN) (F and G 1000x) (F) ZN stain showing a giant cell with intracytoplasmic bacilli in an adult bovine granuloma, and (G) ZN stain from a calf granulomatous lesion, showing a giant cell with abundant bacilli marked with arrows.
Fig 3.
Calcium deposits in granulomatous lesions are independent of fibrous capsule presence in calves.
(A) Masson's trichrome stained section showing a stage IV granuloma surrounded by abundant fibrous tissue (yellow arrow) in a bovine adult. (B) Masson's trichrome stained section showing a stage III-IV granuloma, without a fibrous capsule, but with surrounding disorganized collagen fibers in a calf. White arrow indicates connective tissue of the lymph node capsule as a reference. (C and D) Von Kossa staining, 200x, exhibiting stage IV (adult) and stage III-IV (calf) granulomatous lesions, with calcium deposits dyed in black.
Fig 4.
Histopathological classification of granulomas found in lymph nodes from calves four months of age or younger.
(A) Graph of granuloma stages identified in adults, showing a higher frequency of stages I and IV. (B) Graph showing a higher frequency of stage I granulomas in calves. H&E (C) Stage I: 400X, small diffuse foci of infiltrated inflammatory cells, mainly macrophages, epithelioid macrophages and abundant cellular detritus at the center of the lesion. (D) Stage II: 200X, Larger structure with epithelioid macrophages, multinucleated giant cells, cell debris and necrosis. (E) Stage II-III: 40X, large necrotic areas without borders, absence of connective tissue capsule or lymphocyte accumulation in the periphery. (F) Stage III-IV: 40X, lesions showing extensive necrotic areas, mineralization and cell debris without defined borders, surrounded by macrophages, absence of connective tissue capsule or lymphocytes in the periphery; lesions tend to coalesce and fuse.
Fig 5.
Histopathological classification of granulomas found in lung tissue from calves.
H&E (A.-C 200x, D-E 40x F.-1000x). (A) Stage I. (B) Stage II. (C) Stage II-III. (D) Stage III-IV. (E) Close up of stage II-III. (F) Zielh Neelsen 1000x with abundant extracellular acid-fast bacilli in stage II-III.
Fig 6.
Molecular identification of M. bovis in naturally infected cattle.
(A) Nested PCR amplification products of the 208 bp mpb70 gene. (B) Endpoint PCR of the 206 bp fragment from the M. bovis RD9 gene and the 333 bp fragment from the M. tuberculosis gene. (C) Endpoint PCR of the 268 bp fragment from the M. bovis RD4 gene and the 172 bp fragment from the M. tuberculosis gene MP molecular weight marker is 100 bp; lanes 1–11: bacteriological culture samples; lanes 12–15: negative tissue samples; and lanes 16–18: positive controls (M. bovis BCG and M. tuberculosis H37rv) and negative control.
Table 2.
Bacteriological isolation and molecular genotyping of M. bovis.