Identification of molecular biomarkers associated with disease progression in the testis of bulls infected with Besnoitia besnoiti

Abstract

Breeding bulls infected with Besnoitia besnoiti may develop sterility during either acute or chronic infection. The aim of this study was to investigate the molecular pathogenesis of B. besnoiti infection with prognosis value in bull sterility. Accordingly, five well-characterized groups of naturally and experimentally infected males were selected for the study based on clinical signs and lesions compatible with B. besnoiti infection, serological results and parasite detection. A broad panel of molecular markers representative of endothelial activation and fibrosis was investigated and complemented with a histopathological approach that included conventional histology and immunohistochemistry. The results indicated the predominance of an intense inflammatory infiltrate composed mainly of resident and recruited circulating macrophages and to a lesser extent of CD3+ cells in infected bulls. In addition, a few biomarkers were associated with acute, chronic or subclinical bovine besnoitiosis. The testicular parenchyma showed a higher number of differentially expressed genes in natural infections (acute and chronic infections) versus scrotal skin in experimental infections (subclinical infection). In subclinical infections, most genes were downregulated except for the CCL24 and CXCL2 genes, which were upregulated. In contrast, the acute phase was mainly characterized by the upregulation of IL-1α, IL-6 and TIMP1, whereas in the chronic phase, the upregulation of ICAM and the downregulation of MMP13, PLAT and IL-1α were the most relevant findings. Macrophages could be responsible for the highest level of gene regulation in the testicular parenchyma of severely affected and sterile bulls, and all these genes could be prognostic markers of sterility.

Changes in serum biomarkers of inflammation in bovine besnoitiosis

“…Another limitation was the small sample size (only three acutely infected bulls with testicular degeneration were evaluated), which may have also contributed to the fact that no markers of sterility were identified. Nevertheless, this study offers robust data and complements a previous study that identified a few molecular markers related to disease progression in testicular tissues [ 13 ]. Moreover, the results obtained in experimentally infected cattle offer valuable data about the kinetics of the acute-phase response during acute and chronic infection, as discussed below.…”

“…In our study, animals with chronic infection, in addition to having values of 70 g/l (Fig. 2 ), also presented an intense inflammatory infiltrate with the presence of lymphocytes, some macrophages, abundant young granulation tissue and an abundance of fibroblasts in the testicular parenchyma [ 13 ].…”

“…In general terms, apart from a suggested correlation of APP levels with acute besnoitiosis severity, this acute response appears to be related to a few upregulated molecular markers (IL-6, IL-1α, PLAT and TIMP1) detected in the testicular parenchyma of acutely infected bulls versus a downregulation of PLAT, IL-1α, IL-6, IL-8 and IL-10 in chronically and subclinically infected bulls [ 13 ]. APPs are predominantly glycoproteins synthesised by hepatocytes in response to IL-6, tumour necrosis factor alpha (TNF-α), and other proinflammatory cytokines such as IL-1α [ 34 ].…”

“…A relevant role of interstitial and recruited macrophages in the testicular parenchyma was recently suggested to contribute to the pathogenesis of infection at the testicular level, and different molecular markers were identified as prognostic indicators of sterility. In sterile bulls, the acute phase was mainly characterised by the upregulation of interleukin-1 alpha (IL-1α), IL-6 and matrix metalloproteinase inhibitor-1 (TIMP1), whereas in the chronic phase, the upregulation of intercellular adhesion molecules (ICAM) and the downregulation of metalloproteinase 13 (MMP13), tissue-type plasminogen activator (PLAT) and IL-1α were observed in the testicular parenchyma [ 13 ]. Since monitoring of disease progression in testicular parenchyma by measuring tissue markers may be difficult to implement as a routine diagnostic procedure, identification of markers in serum samples may be a more straightforward approach.…”

An investigation into bovine besnoitiosis (Besnoitia besnoiti) in an Irish pedigree Aberdeen Angus herd

“…1 Sterility (aspermatogenesis) in males may occur during both the acute and chronic phase of infection, and be caused by necrotising orchitis leading to atrophy and induration of the testes. 27,28 Tissue cysts in the scleral (bulbar) conjunctiva are clinically pathognomonic and develop 6-7 weeks after infection. 1,2,6,11 In endemic situations, cattle mortality rates of <1% are expected.…”