Identification and cultivation of methyllycaconitine degraders from wild ruminants to protect against larkspur poisoning in range cattle

Abstract

Tall larkspur (Delphinium spp.) in the western United States present a serious toxicity danger to rangeland cattle. Consumption of Methyllycaconitine (MLA), the toxic alkaloid in larkspur plants, can cause annual losses of 5-15% of range cattle in grazing pastures with sufficient larkspur. With the wide distribution and abundance of larkspur, wild ruminants in Montana likely encounter tall larkspur while foraging; however, no evidence suggests they are negatively affected by MLA's toxic effects. Therefore, we evaluated: i) whether alkaloids in Delphinium spp., and MLA specifically degraded within ruminal specimens collected from Montana's wild ruminant species over 48 h using in vitro incubations; ii) whether observed degradative activities were abiotic, or mediated by either the fungal or non-fungal (mostly bacterial) residents of the ruminal microbiota in wild ruminant specimens; and iii) if representative microbial isolates individually possessed the ability to degrade MLA within in vitro incubations. Rumen samples were collected from wild ruminant species during the 2019 and 2020 hunting seasons using legal methods by volunteer hunters. In all assays, total alkaloid was measured spectrophotometrically, and MLA by High-Performance Liquid Chromatography Mass Spectrometry (HPLC) from initial and final incubations. Our results demonstrated that, with the exception of white-tailed deer, all wild ruminant species exhibited variable degradative abilities in both total alkaloid (P< 0.001) and MLA (P< 0.001) assays and that such degradation was predominantly mediated by ruminal fungi. Additionally, screening of 15 fungal isolates, representing 10 known genera and 2 isolates of unknown taxonomic identity each obtained from herbivorous hosts, determined all were capable of degrading MLA to some extent. Fungal isolates obtained from wild ruminants exhibited greater degradative activity, with Aestipasuomyces R5 isolated from wild sheep degrading 71% of MLA (P<0.001). Overall, our results indicate that degradation of both total alkaloid and MLA-specifically occurs within the gastrointestinal tract of Montana's wild ruminants and that it is largely influenced by fungal activity. Additionally, fungal strains isolated from wild ruminants are capable of degrading MLA and have the potential to be further used as a direct fed microbial to rangeland cattle as an optimal way to mitigate larkspur toxicosis.