MicroRNAs as serum biomarker for Senecio brasiliensis poisoning in cattle
Graphical Abstract
Introduction
Poisonous plants and their secondary metabolites are responsible for many cases of poisoning in cattle worldwide. It is established that Senecio spp. is one of the most frequent plant-related poisonings in animal production. It is an important problem in countries such as Brazil (Molossi et al., 2021) and Uruguay (García et al., 2020), as well as in Europe (Cortinovis and Caloni, 2013, Cortinovis and Caloni, 2015a), Australia, and North America (Gardner et al., 2006). The Senecio genus (Compositae family) is composed of more than 1200 species distributed worldwide, 25 of which have been confirmed to be poisonous. These plants contain a series of pyrrolizidine alkaloids (PAs) which become cumulative hepatotoxins after they are bioactivated to pyrrole metabolites in the liver (Cortinovis and Caloni, 2015a).
The clinical, anatomopathological, and histopathological profiles of Senecio spp. are quite characteristic in acute and subacute intoxications. Liver function tests and histological studies of liver biopsies are used to establish a prognosis in animals when there is mistrust of seneciosis (Barros et al., 2007, Giaretta et al., 2014b). Although hepatic biopsy is considered the optimal test for diagnosing seneciosis, it is an invasive method that must be performed by an experienced professional, and tissue processing is time-consuming, thus delaying clinical decision (Barros et al., 2007, Emanuelli et al., 2016).
An important aspect of public health is the transfer of toxins to humans through the ingestion of meat, milk, and other derivatives. The PAs can be eliminated in milk (Hoogenboom et al., 2011). A study showed that goats' milk poisoned with Crotalaria spectabilis, which contain PAs, induced toxicity in rats (MEDEIROS et al., 1999). The mutagenic and teratogenic effects of Senecio spp. alkaloids in milk should be considered, although there are no reports of diseases caused in humans due to their consumption (Cheeke, 1988). This problem becomes more specific when poisoned cattle do not present clinical signs.
In this context, microRNAs (miRNA) detection has aroused interest by playing an important role in veterinary medicine (Chodkowska et al., 2017, Do et al., 2021, Dong et al., 2017, Farrell et al., 2015, Mukiibi et al., 2020, Shaughnessy et al., 2020). The miRNAs are small 19–23 nucleotide non-coding RNA molecules that modulate approximately 30% of mRNA, with fundamental roles in proliferation, differentiation, and apoptosis mechanisms (Ambros, 2004, Chi et al., 2009). According to information from the largest miRNA database (miRBase), up to now, 1025 miRNA have been described in cattle (Griffiths-Jones, 2006). Currently, miRNAs have already been studied as biomarkers for the rapid diagnosis of mastitis in dairy cows (Li et al., 2015b) and as markers for hepatic function in anabolic use (Becker et al., 2011b), early gestation (Ioannidis and Donadeu, 2017), milk quality (Wang et al., 2016a), early detection of Mycobacterium avium (Britton et al., 2016), among other applications. A feature of great interest in using miRNA as diagnostic biomarkers is their presence in biological fluids such as serum, urine, milk, and saliva in a stable manner, allowing a less invasive detection (Sempere, 2012, Wang et al., 2016b).
Thus, the study of new forms of diagnosis is necessary and of great interest to the area. Therefore, this work aimed to identify a miRNA profile in cattle serum intoxicated to Senecio brasiliensis. The study of miRNA in the serum of animals poisoned by Senecio spp. may improve the knowledge of the disease, organs involved, degree of poisoning, and prognosis, allowing a greater understanding of the clinical signs.
Section snippets
Animals
This study analyzed naturally exposed animals that were assisted by veterinary doctors of the university (Adriano T. Ramos and André L.F. Goetten). Animals were kept in native pasture, not receiving ration. No experimental protocol with animals were performed, they received medical attention and diagnosis was done afterwards.
Toxicosis event
The cattle consisted of Aberdeen Angus cows (36−48 months old), reared in Curitibanos, SC, Brazil. The extensive grazing of beef cattle in these areas, is the most used
Animals
Animals presenting at least one histopathological alteration were considered intoxicated (20 animals), and those with no hepatic damage were grouped as control (eight healthy animals).
Serum miRNA expression
Relative expression of miR-30a, mir-378, mir-21, mir-122, and miR-885 were higher in intoxicated animals than in healthy animals (Fig. 1 B-E, G). No changes were observed in let-7f and miR-181b levels (Fig. 1 A, F).
Diagnostic value of the miR-30a, miR-378, miR-21, miR-885 and miR-122 in serum
Receiver operating characteristic (ROC) analyses were performed to explore the diagnostic value of
Discussion
Seneciosis is one of the most frequent plant-related poisonings in animal production that result in hepatotoxicity (Cortinovis and Caloni, 2015b, Molossi et al., 2021). Poisonous plants like Senecio spp. can induce the death of animals. However, its effects often remain unmeasured and are not easily identified. James et al. (2019) reports that big economic losses are associated with undiagnosed intoxications resulting in low animal performance, immunodeficiency, reduction in reproductive
Conclusion
In summary, our study analyzed miRNAs expressed differently between intoxicated and healthy animals and detected five upregulated miRNAs in the serum animals, allowing significant correlations. Furthermore, the sensitivity and specificity of the miRNAs by ROC analysis indicated that the miR-122, miR-885, and, especially, miR-21 signatures have potential application for detecting cattle poisoning by Senecio spp. Although the present study has some limitations, including the small sample size, it
CRediT authorship contribution statement
Evelyn Winter: Conceptualization, Writing – original draft, Supervision, Project administration. Julia Cisilotto: Investigation. André L.F. Goetten: Investigation. Ângela Veiga: Investigation. Adriano T. Ramos: Investigation. Francielli C. Zimermann: Investigation. Carolina Reck: Investigation. Tânia B. Creczynski-Pasa: Resources, Writing – review & editing.
Declaration of Competing Interest
The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.
Acknowledgements
This study was supported by FAPESC (Fundação de Amparo à Pesquisa e Inovação do Estado de Santa Catarina) grants ID 2017TR1702, by CNPq (Conselho Nacional de Desenvolvimento Científico e Tecnológico), Brazil, which provided grant to Tânia B. Creczynski-Pasa, as well as by CAPES (Coordenação de Aperfeiçoamento de Pessoal de Nível Superior), Brazil, which provided PNPD-fellowships for Júlia Cisilotto.
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