Immunoproteomic insights into inflammatory diseases of the critically endangered black rhinoceros (Diceros bicornis)

Black rhinoceros are critically endangered due to poaching in the wild (in situ). Globally, fewer than 200 animals are maintained as an ex situ insurance population. Unfortunately, the ex situ population faces major sustainability challenges from disease syndromes characterized by high inflammatory burdens and diverse manifestations of immunometabolic dysfunction, not known to be present among their wild counterparts. Overlapping ex situ disease phenotypes limit diagnostic specificity and highlight the need to define underlying disease mechanisms. In the present study, using a cohort of presumed clinically healthy and inflammatory black rhinoceros, we generated the first immunoproteomic profile of any endangered mammal species and identified 1,311 immune cell proteins. However, no significant differences were detected among clinical phenotypes. Therefore, we applied unsupervised machine learning approaches to detect molecular features suggestive of healthy versus inflammatory phenotypes. Forty-three proteins associated with inflammatory pathways were differentially expressed in a cohort of samples derived from both presumed healthy and inflammatory phenotypes. Results suggest subclinical disease may be relatively widespread ex situ, and that animals experience temporal fluctuations in inflammatory state over time. Findings implicate neutrophil degranulation and dysregulation of the oral-gut-liver axis as drivers of disease syndromes of ex situ black rhinoceros. The forty-three proteins associated with inflammatory pathways represent candidate inflammatory biomarkers to be assessed for clinical applications in future validation studies. Upon validation, these candidate biomarkers may guide management practices to strengthen long-term population sustainability.