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Hypoglycin A is a plant-derived protoxin that causes atypical myopathy in equids. In atypical myopathy-affected horses, metabolomic and microbiome studies have reported alterations in metabolic markers and faecal microbiota composition, pointing to a potential disruption of microbial homeostasis. However, in vivo observations are strongly confounded by host-related factors, underscoring the need for controlled in vitro approaches. To address this, we used an in vitro static batch fermentation model simulating the equine colon to investigate the direct effects of hypoglycin A on microbiota composition and activity. Faecal inocula from healthy horses were incubated in control and hypoglycin A-treated fermenters for 48 h, with serial analyses of hypoglycin A concentration, short-chain fatty acids, and 16S rRNA gene profiles. Hypoglycin A remained stable in the nutritive medium in the absence of microbiota, confirming that its degradation in inoculated fermenters was microbiota-dependent. The results showed significant microbial-associated hypoglycin A degradation without evidence of toxic metabolite formation. The analysis of α- and β-diversity revealed both an effect of incubation time, reflecting the natural temporal dynamics of microbial communities under batch fermentation, and a specific impact of hypoglycin A exposure, with certain taxa such as Paraclostridium being affected. This study provides the first in vitro evidence that the equine microbiota contributes to hypoglycin A degradation.

Acer pseudoplatanus is a worldwide-distributed tree which contains toxins, among them hypoglycin A (HGA). This toxin is known to be responsible for poisoning in various species, including humans, equids, Père David’s deer and two-humped camels. We hypothesized that any herbivore pasturing with A. pseudoplatanus in their vicinity may be at risk for HGA poisoning. To test this hypothesis, we surveyed the HGA exposure from A. pseudoplatanus in species not yet described as being at risk. Animals in zoological parks were the major focus, as they are at high probability to be exposed to A. pseudoplatanus in enclosures. We also searched for a toxic metabolite of HGA (i.e., methylenecyclopropylacetyl-carnitine; MCPA-carnitine) in blood and an alteration of the acylcarnitines profile in HGA-positive animals to document the potential risk of declaring clinical signs. We describe for the first instance cases of HGA poisoning in Bovidae. Two gnus (Connochaetes taurinus taurinus) exposed to A. pseudoplatanus in their enclosure presented severe clinical signs, serum HGA and MCPA-carnitine and a marked modification of the acylcarnitines profile. In this study, even though all herbivores were exposed to A. pseudoplatanus, proximal fermenters species seemed less susceptible to HGA poisoning. Therefore, a ruminal transformation of HGA is hypothesized. Additionally, we suggest a gradual alteration of the fatty acid metabolism in case of HGA poisoning and thus the existence of subclinical cases.

Equine atypical myopathy is caused by hypoglycin A (HGA) and methylenecyclopropylglycine (MCPrG), the known protoxins of sycamore maple (Acer pseudoplatanus). Various tissues from five atypical myopathy cases were analyzed but only HGA was found. Whether deamination of MCPrG has already occurred in the intestine as the first stage of metabolization has not been investigated. Activation of the protoxins to methylenecyclopropylacetyl (MCPA)-CoA and methylenecyclopropylformyl (MCPF)-CoA, respectively, occurred mainly in the skeletal muscles, as evidenced by very high concentrations of MCPA-carnitine and MCPF-carnitine in this tissue. Inhibition of the acyl-CoA dehydrogenases of short- and medium-chain as well as branched-chain fatty acids by the toxins led to a strong increase in the corresponding acylcarnitines, again preferentially in skeletal muscles. An accumulation of the long-chain acylcarnitines beyond the level of the control samples could not be detected in the tissues. As a high amount of HGA was always found unmetabolized in the organs, we speculate that targeting the interruption of further metabolization might be a way to stop the progression of intoxication. Inhibition of the mitochondrial branched-chain amino acid aminotransferase, i.e., the first enzyme responsible for the activation of sycamore maple protoxins, could be a therapeutic approach.

Hypoglycin A and methylenecyclopropylglycine are protoxins responsible for atypical myopathy in equids. These protoxins are converted into toxins that inhibit fatty acid β-oxidation, leading to blood accumulation of acylcarnitines and toxin conjugates, such as methylenecyclopropylacetyl-carnitine. The enzymes involved in this activation are also present in some prokaryotic cells, raising questions about the potential role of intestinal microbiota in the development of intoxication. Differences have been noted between the faecal microbiota of cograzers and atypical myopathy-affected horses. However, recent blood acylcarnitines profiling revealed subclinical cases among cograzers, challenging their status as a control group. This study investigates the faecal microbiota of horses clinically affected by atypical myopathy, their cograzers, and a control group of toxin-free horses while analysing correlations between microbiota composition and blood parameters. Faecal samples were analysed using 16S amplicon sequencing, revealing significant differences in α-diversity, evenness, and β-diversity. Notable differences were found between several genera, especially Clostridia_ge, Bacteria_ge, Firmicutes_ge, Fibrobacter, and NK4A214_group. Blood levels of methylenecyclopropylacetyl-carnitine and C14:1 correlated with variations in faecal microbial composition. The theoretical presence of enzymes in bacterial populations was also investigated. These results underscore the critical need to investigate the potential role of intestinal microbiota in this poisoning and may provide insights for developing prevention and treatment strategies.

Equine atypical myopathy (AM) is seasonal intoxication resulting from the ingestion of seeds and seedlings of the sycamore maple (Acer pseudoplatanus) that contain toxins, among them, hypoglycin A (HGA). Literature mentions several cases of AM among gravid mares and in unweaned foals. The objective of this study was to determine whether HGA and/or its metabolite are present in milk from grazing mares exposed to sycamore maple trees as confirmed by detection of HGA and its metabolite in their blood. Four mare/foal couples were included in the study. Both HGA and its metabolite were detectable in all but one of the milk samples. To our knowledge, this is the first study describing transfer of HGA to the milk. This unprecedented observation could partially explain cases of unweaned foals suffering from AM. However, a transplacental transfer of the toxin cannot be excluded for newborn foals. Besides being a source of contamination for offspring, milk contamination by toxins from fruits of trees of the Sapindaceae family might constitute a potential risk for food safety regarding other species’ raw milk or dairy products.

Equine atypical myopathy (AM) is caused by hypoglycin A (HGA) and methylenecyclopropylglycine (MCPG) intoxication resulting from the ingestion of seeds or seedlings of some Acer tree species. Interestingly, not all horses pasturing in the same toxic environment develop signs of the disease. In other species, it has been shown that the intestinal microbiota has an impact on digestion, metabolism, immune stimulation and protection from disease. The objective of this study was to characterize and compare fecal microbiota of horses suffering from AM and healthy co-grazers. Furthermore, potential differences in fecal microbiota regarding the outcome of diseased animals were assessed. This prospective observational study included 59 horses with AM (29 survivors and 30 non-survivors) referred to three Belgian equine hospitals and 26 clinically healthy co-grazers simultaneously sharing contaminated pastures during spring and autumn outbreak periods. Fresh fecal samples (rectal or within 30 min of defecation) were obtained from all horses and bacterial taxonomy profiling obtained by 16S amplicon sequencing was used to identify differentially distributed bacterial taxa between AM-affected horses and healthy co-grazers. Fecal microbial diversity and evenness were significantly (p < 0.001) higher in AM-affected horses as compared with their non-affected co-grazers. The relative abundance of families Ruminococcaceae, Christensenellaceae and Akkermansiaceae were higher (p ≤ 0.001) whereas those of the Lachnospiraceae (p = 0.0053), Bacteroidales (p < 0.0001) and Clostridiales (p = 0.0402) were lower in horses with AM, especially in those with a poor prognosis. While significant shifts were observed, it is still unclear whether they result from the disease or might be involved in the onset of disease pathogenesis.

In spring, grazing equids may suffer from a severe rhabdomyolysis syndrome named atypical myopathy. This environmental intoxication results from ingestion of toxins contained in Acer pseudoplatanus seedlings. The aim of this study was to investigate the effectiveness of herbicide spraying and mowing to reduce the toxic pressure of sycamore seedlings. In a first experiment, the efficacy of three herbicides to eradicate seedlings was compared to mowing. In a second experiment, the influence of the mowing timing on pasture productivity was determined. In both experiments, sycamore seedling counting, grass height and botanical composition were determined. In experiment 2, the final harvest biomass and its nutritional value were also determined. Herbicides and mowing both reduced the number of seedlings, which nevertheless disappear naturally over time without intervention (i.e., in control areas). As opposed to mowing, herbicide spraying altered the composition of the pasture flora. Both sprayed and mowed seedlings remain toxic until full decomposition. Early mowing (i.e., early April) did not affect the harvest yield. Late mowing (i.e., end of April) reduced the harvest yield but its nutritional value fitted the horses’ need. In conclusion, mowing is the best strategy to reduce the risk of Acer pseudoplatanus poisoning in grazing equids while preserving ecological sustainability and nutritional value of pastures.

Abstract Background Locoregional tumor control and prolonged survival for dogs with genitourinary carcinoma (CGUC) reportedly are achievable using treatment with radiotherapy (RT) with or without adjunctive chemotherapy and nonsteroidal anti-inflammatory drugs (NSAIDs). Objectives To characterize event-free and overall survival after treatment of CGUC using NSAIDs, mitoxantrone (MTX), and a standardized RT protocol (57 Gy in 20 fractions). Animals Fifty-one client-owned dogs treated between 2008 and 2017. Methods Dogs were retrospectively categorized into treatment groups: (a) first-line concurrent chemoradiotherapy (≥1 dose of MTX started within 1 month of RT); (b) first-line chemotherapy (MTX administered for >1 month before RT without tumor progression); (c) RT as a salvage procedure (MTX, surgery or both with subsequent locoregional tumor progression before RT). Treatment-induced toxicoses, event-free survival (EFS), and overall survival times (OSTs) were recorded. The influence of demographics, staging, and treatment-related factors on survival was assessed using Cox proportional hazards modeling. Results Median EFS and OST for all dogs were 260 and 510 days with no significant differences among groups 1 (n = 39), 2 (n = 4), and 3 (n = 8). Both EFS and OST were shorter in dogs with moderate to severe clinical signs (P < .001 and P < .001, respectively); OST was shorter in dogs with prostatic involvement (P = .02). Permanent urinary incontinence developed in 16 dogs (31%) at a median of 70 days postirradiation; other toxicoses were mild and self-limiting. Conclusions and Clinical Importance Mild clinical signs and lack of prostate involvement were associated with favorable prognosis for survival. Client education regarding the risk of urinary incontinence is warranted.

Background Growing evidence suggests that soil microbes can improve plant fitness under drought. However, in potato, the world’s most important non-cereal crop, the role of the rhizosphere microbiome under drought has been poorly studied. Using a cultivation independent metabarcoding approach, we examined the rhizosphere microbiome of two potato cultivars with different drought tolerance as a function of water regime (continuous versus reduced watering) and manipulation of soil microbial diversity (i.e., natural (NSM), vs. disturbed (DSM) soil microbiome). Results Water regime and soil pre-treatment showed a significant interaction with bacterial community composition of the sensitive (HERBST) but not the resistant cultivar (MONI). Overall, MONI had a moderate response to the treatments and its rhizosphere selected Rhizobiales under reduced watering in NSM soil, whereas Bradyrhizobium , Ammoniphilus , Symbiobacterium and unclassified Hydrogenedensaceae in DSM soil. In contrast, HERBST response to the treatments was more pronounced. Notably, in NSM soil treated with reduced watering, the root endophytic fungus Falciphora and many Actinobacteriota members ( Streptomyces , Glycomyces , Marmoricola , Aeromicrobium , Mycobacterium and others) were largely represented. However, DSM soil treatment resulted in no fungal taxa and fewer enrichment of these Actinobacteriota under reduced watering. Moreover, the number of bacterial core amplicon sequence variants (core ASVs) was more consistent in MONI regardless of soil pre-treatment and water regimes as opposed to HERBST, in which a marked reduction of core ASVs was observed in DSM soil. Conclusions Besides the influence of soil conditions, our results indicate a strong cultivar-dependent relationship between the rhizosphere microbiome of potato cultivars and their capacity to respond to perturbations such as reduced soil moisture. Our study highlights the importance of integrating soil conditions and plant genetic variability as key factors in future breeding programs aiming to develop drought resistance in a major food crop like potato. Elucidating the molecular mechanisms how plants recruit microbes from soil which help to mitigate plant stress and to identify key microbial taxa, which harbour the respective traits might therefore be an important topic for future research.

In 2014, atypical myopathy (AM) was linked to Acer pseudoplatanus (sycamore maple) in Europe. The emergence of this seasonal intoxication caused by a native tree has raised many questions. This manuscript aims at answering the five most frequently asked questions (FAQs) regarding (1) identification of toxic trees; reduction of risk at the level of (2) pastures and (3) equids; (4) the risk associated with pastures with sycamores that have always been used without horses being poisoned and (5) the length of the risk periods. Answers were found in a literature review and data gathered by AM surveillance networks. A guide is offered to differentiate common maple trees (FAQ1). In order to reduce the risk of AM at pasture level: Avoid humid pastures; permanent pasturing; spreading of manure for pasture with sycamores in the vicinity and avoid sycamore maple trees around pasture (FAQ2). To reduce the risk of AM at horse level: Reduce pasturing time according to weather conditions and to less than six hours a day during risk periods for horses on risk pasture; provide supplementary feeds including toxin-free forage; water from the distribution network; vitamins and a salt block (FAQ3). All pastures with a sycamore tree in the vicinity are at risk (FAQ4). Ninety-four percent of cases occur over two 3-month periods, starting in October and in March, for cases resulting from seeds and seedlings ingestion, respectively (FAQ5).