Toxigenic Endophyte-Infected Tall Fescue and Ergot Alkaloids.

"Fescue toxicosis" and reproductive ergotism present identical toxidromes in late-gestational mares and, likely, other equids. Both toxic syndromes are caused by ergopeptine alkaloids (EPAs) of fungal origin, and they are collectively referred to as equine ergopeptine alkaloid toxicosis (EEPAT). EPAs are produced by either a toxigenic endophyte (Epichloë coenophiala) in tall fescue and/or a nonendophytic fungus (Claviceps purpurea), infecting small grains and grasses. EEPAT can cause hypoprolactinemia-induced agalactia/dysgalactia, prolonged gestation, dystocia, and other reproductive abnormalities in mares, as well as failure of passive transfer in their frequently dysmature/overmature/postmature foals. Prevention relies on eliminating exposures and/or reversing hypoprolactinemia.

Examining the long-term impacts of ergot alkaloids on fetal heifer development and subsequent growth and reproductive performance.

Consumption of toxic endophyte-infected tall fescue (EI) results in poor reproductive performance in domestic livestock. In this study, the objective was to evaluate the effects of ergovaline exposure during mid-gestation (days 93 through 188 of gestation) on dam performance, the growing female fetus, and the subsequent growth and reproductive performance of the gestationally exposed heifer calves. Pregnant Angus and Simmental-Angus cows were blocked by age (2 to 3, to 7, and >7 y), body weight (BW), and breed; and then randomly assigned to graze either novel endophyte-infected tall fescue (EN; <5% infection rate; n = 27 year 1, n = 16 year 2) or toxic EI (99% infection rate; n = 27 year 1, n = 17 year 2). Weekly BW, body condition scores (BCS), hair coat scores, hair shedding scores (HSS), and blood samples for progesterone (P4) analysis were collected from mid-April through July of 2017 (year 1) and 2018 (year 2). Gestation length, birth weight, placental characteristics, heifer calf growth, onset of puberty, ovarian characteristics, and artificial insemination pregnancy rates were measured. Data were analyzed using the MIXED procedure of SAS. Cows grazing EI pastures had reduced average daily gain, reduced BCS, greater HSS, and decreased P4 concentrations compared to cows on EN pasture (P < 0.01). Birth weights were decreased for heifers whose dams were exposed to EI pastures during their second trimester (P < 0.01). Heifer pregnancy rates were not impacted by EI pasture exposure during gestation for either year of the study. However, a treatment-by-year effect was seen for the pregnancy rate for EI-exposed heifers in year 2; EI-exposed heifers in year 2 had increased pregnancy rates at two of the inseminations. Combined, these data reinforce that consumption of toxic EI during gestation can negatively impact both dam and offspring performance. More studies are needed to evaluate more parameters in an effort to elucidate the possible life-long impacts of ergovaline exposure during gestation.

The U.S. livestock industry incurs over one billion dollars of economic loss every year due to fescue toxicosis, caused by consuming ergot alkaloids produced by an endophytic fungus in some grass species. Identifying means to mitigate the negative effects of fescue toxicosis is needed for U.S. beef producers. Effective treatment for this toxicosis is still needed. The objective of this study was to evaluate the effects of ergovaline exposure during mid-gestation on dam performance, the growing female fetus, and the subsequent growth and reproductive performance of the gestationally exposed heifer calves. We identified specific phenotype traits that undergo developmental programming in utero in response to fescue toxicosis. However, measurements of growth and reproductive performance were not altered by ergot exposure.

High deoxynivalenol and ergot alkaloid levels in wheat grain: effects on growth performance, carcass traits, rumen fermentation, and blood parameters of feedlot cattle.

This study was designed to assess the impacts of a mixture of deoxynivalenol (DON) and ergot alkaloids (EAs) on growth performance, rumen function, blood parameters, and carcass traits of feedlot cattle. Forty steers (450 ± 6.0 kg) were stratified by weight and randomly allocated to 1 of 4 treatments; control-low (CON-L), control-high (CON-H) which contained low or high wheat screenings that lacked mycotoxins at the same level as the mycotoxin-low (MYC-L; 5.0 mg/kg DON, 2.1 mg/kg EA), and mycotoxin-high (MYC-H: 10 mg/kg DON, 4.2 mg/kg EA) diets that included wheat screening with mycotoxins. Steers were housed in individual pens for a 112-day finishing trial. Intake was 24.8% lower (P < 0.001) for MYC steers compared to CON steers. As a result, average daily gains of MYC steers were 42.1% lower (P < 0.001) than CON steers. Gain to feed ratio was also lower (P < 0.001) for MYC steers compared to CON steers. Platelets, alanine aminotransferase, globulins, and blood urea nitrogen were lower (P ≤ 0.008), and lymphocytes, glutathione peroxidase activity (GPx), and interleukin-10 (IL-10) were elevated (P ≤ 0.002) in MYC steers compared to CON steers. Hot carcass weights and backfat thickness were reduced (P < 0.001) in MYC steers, resulting in leaner (P < 0.001) carcasses and higher (P < 0.007) meat yield compared to CON steers. Results suggest that a mixture of DON and EAs negatively impacted health, performance, and carcass traits of feedlot steers, with the majority of this response likely attributable to EAs. However, more research is needed to distinguish the relative contribution of each mycotoxin to the specific responses observed.

Climate Effects on Ergot and Ergot Alkaloids Occurrence in Italian Wheat.

In recent years, there has been an intensification of weather variability worldwide as a result of climate change. Some regions have been affected by drought, while others have experienced more intense rainfall. The incidence and severity of moldy grain and mycotoxin contamination during the growing and harvesting seasons have increased as a result of these weather conditions. Additionally, torrential rains and wet conditions may cause delays in grain drying, leading to mold growth in the field. In July 2023, a wheat field in Lecco (Lombardy, Italy) was affected by torrential rains that led to the development of the Claviceps fungi. In the field, dark sclerotia were identified on some ears. Wheat ears, kernels, and sclerotia were collected and analyzed by LC-MS/MS at IZSLER, Food Chemical Department, in Bologna. The wheat ears, kernels, and sclerotia were analyzed for 12 ergot alkaloids (EAs) according to (EU) Regulation 2023/915 (ergocornine/ergocorninine; ergocristine/ergocristinine; ergocryptine/ergocryptinine; ergometrine/ergometrinine; ergosine/ergosinine; ergotamine/ergotaminine), after QuEChERS (Z-Sep/C18) purification. The analyzed sclerotia showed significant differences in total alkaloid content that vary between 0.01 and 0.5% (w/w), according to the results of the 2017 EFSA scientific report. EAs detected in sclerotia were up to 4951 mg/kg, in wheat ears up to 33 mg/kg, and in kernels were 1 mg/kg. Additional mycotoxins, including ochratoxin A, deoxynivalenol, zearalenone, fumonisins, T2-HT2 toxins, and aflatoxins, were investigated in wheat kernels after purification with immunoaffinity columns (IAC). The analysis revealed the presence of deoxynivalenol in wheat kernels at a concentration of 2251 µg/kg. It is expected that climate change will increase the frequency of extreme weather events. In order to mitigate the potential risks associated with mycotoxin-producing fungi and to ensure the protection of human health, it is suggested that official controls be implemented in the field.

Simultaneous determination of total ergot alkaloids in wheat flour by Orbitrap mass spectrometry.

The optimization screening methods for total ergot alkaloids in wheat extracts involve transforming them into a single compound, which is then analyzed via high-resolution Orbitrap mass spectrometry (Orbitrap MS). Orbitrap MS provides highly sensitive and accurate mass measurements, enhancing the selectivity and sensitivity of the analysis. Various hydrolysis and reduction methods have been investigated, and the use of superhydrides has emerged as the most effective method for transforming ergopeptine alkaloids. This study also focused on the epimerization of ergot alkaloids, particularly the differences between R- and S-epimers and their impact on the mass spectra. We validated our method by assessing the linearity, sensitivity, recovery, matrix effects, repeatability, and stability. The limits of detection and quantitation were set at 0.43 and 1.30 µg LSA/kg wheat, respectively. The proposed method offers a robust analytical approach for screening and quantifying total ergot alkaloids in wheat samples, addressing important concerns about their presence in food and feed.

First Synthesis of Ergotamine-13CD3 and Ergotaminine-13CD3 from Unlabeled Ergotamine.

Ergot alkaloids (EAs) formed by Claviceps fungi are one of the most common food contaminants worldwide, affecting cereals such as rye, wheat, and barley. To accurately determine the level of contamination and to monitor EAs maximum levels set by the European Union, the six most common EAs (so-called priority EAs) and their corresponding epimers are quantified using high-performance liquid chromatography coupled with tandem mass spectrometry (HPLC-MS/MS). The quantification of EAs in complex food matrices without appropriate internal standards is challenging but currently carried out in the standard method EN 17425:2021 due to their commercial unavailability. To address the need for isotopically labeled EAs, we focus on two semi-synthetic approaches for the synthesis of these reference standards. Therefore, we investigate the feasibility of the N6-demethylation of native ergotamine to yield norergotamine, which can subsequently be remethylated with an isotopically labeled methylating reagent, such as iodomethane (13CD3-I), to yield isotopically labeled ergotamine and its C8-epimer ergotaminine. Testing the isotopically labeled ergotamine/-inine against native ergotamine/-inine with HPLC coupled to high-resolution HR-MS/MS proved the structure of ergotamine-13CD3 and ergotaminine-13CD3. Thus, for the first time, we can describe their synthesis from unlabeled, native ergotamine. Furthermore, this approach is promising as a universal way to synthesize other isotopically labeled EAs.

Mycotoxicoses in Veterinary Medicine: Fusarium Toxins, Grass Staggers, and Neothyphodium Toxins.

Fungi are a large group of eukaryotic microorganisms that can readily adapt to diverse environments and occur in almost all climatic zones and continents. Although some fungi are inevitable in the environment for the decay and recycling of organic material, many species are known to produce secondary metabolites, and these mycotoxins, when ingested with food or feed materials, can adversely affect animal and human health. Among the toxigenic fungi, Fusarium species are recognized as so-called field fungi, invading crops and producing mycotoxins predominantly before harvest. Fusarium produces a wide array of mycotoxins, causing different plant diseases. Fusariosis causes significant economic losses in a wide range of crops. Fusarium secondary metabolites, particularly trichothecenes, are potent toxins in mammalian species and cause diverse adverse effects in humans and animals. Other prominent Fusarium toxins with entirely different chemical structures are zearalenone and its derivatives and fumonisins. With an entirely different life cycle, toxins of endophytes belonging to the genus Epichloë and Neothyphodium coenophialum and Neothyphodium lolii comprise an animal health risk, particularly for grazing animals. This review aimed to summarize the adverse effects of selected Fusarium and Epichloë toxins, with a special emphasis on their occurrence in roughages and their mechanisms of action, and describe their effect on animal health and welfare and the potentially related public health risks.

A Major Facilitator Superfamily Transporter Contributes to Ergot Alkaloid Accumulation but Not Secretion in Aspergillus leporis.

Ergot alkaloids are fungal natural products with important roles in agriculture and medicine. We used heterologous expression and gene knockout approaches to investigate potential roles for the product of a major facilitator superfamily transporter gene (easT) recently found in an ergot alkaloid biosynthetic gene cluster in Aspergillus leporis. A strain of Aspergillus fumigatus previously engineered to accumulate lysergic acid, but which did not convert the precursor agroclavine to lysergic acid efficiently or secrete lysergic acid well, was chosen as an expression host for easT. Expression of easT in this strain resulted in accumulation of significantly more pathway intermediates but no detectable lysergic acid. Secretion of ergot alkaloids was reduced in the easT-expressing strain. EasT localized to discrete vesicle-like structures in the cytosol of A. fumigatus, with no localization detected in the plasma membrane. When easT was knocked out in A. leporis, accumulation of lysergic acid amides was reduced relative to the wild type. There was no negative effect on secretion of ergot alkaloids in the knockout mutant. The data indicate that easT encodes a product that contributes to accumulation of ergot alkaloids, perhaps by transporting intermediates between cellular compartments, but does not have a significant role in secreting ergot alkaloids.

All-in-one 2LabsToGo system for analysis of ergot alkaloids in whole rye.

Ergot alkaloids, naturally occurring mycotoxins of Claviceps fungi, pose health risks. This necessitates accurate analysis methods to ensure food safety. This study explored the open-source miniaturized all-in-one 2LabsToGo system to analyze ergot alkaloids in whole rye samples. It is suited for sustainable atline analysis as it combines all planar chromatography tasks, allowing low-cost quality control in milling plants. The LOD and LOQ of ergocristine were determined to be 0.4 and 1.2 ng/zone, respectively. Detectability of ergot alkaloids was proven to be below the current maximum limit of 500 µg/kg for rye milling products. The repeatability (%RSD) was 4.1 % and the coefficient of determination of the analytical response (R2) was 0.9918 for ergocristine. The mean recovery rate of ergot alkaloids in spiked whole rye grain was close to 100 %. Results of screening whole rye for ergot alkaloids were successfully verified by comparison with those obtained by conventional status quo HPTLC instrumentation.

Development, in-house validation and application of a method using high-performance liquid chromatography with fluorescence detection (HPLC-FLD) for the quantification of 12 ergot alkaloids in compound feeds.

Introduction: Ergot alkaloids (EAs) are toxic substances naturally produced by Claviceps fungi. These fungi infest a wide range of cereals and grasses. When domestic animals are exposed to EAs through contaminated feeds, it is detrimental to them and leads to significant economic losses. For that reason, it is important to monitor feed for the presence of EAs, especially with methods enabling their determination in processed materials. Material and Methods: Ergot alkaloids were extracted with acetonitrile, and dispersive solid phase extraction (d-SPE) was used for clean-up of the extracts. After evaporation, the extracts were reconstituted in ammonium carbonate and acetonitrile and subjected to instrumental analysis using high-performance liquid chromatography with fluorescence detection. The developed method was validated in terms of linearity, selectivity, repeatability, reproducibility, robustness, matrix effect, limits of quantification and detection and uncertainty. The EA content of 40 compound feeds was determined. Results: All the assessed validation parameters fulfilled the requirements of Regulation (EU) 2021/808. At least one of the monitored alkaloids was determined in 40% of the samples. The EAs with the highest incidence rate were ergocryptine, ergometrinine and ergocornine. The total concentrations of EAs ranged from under the limit of quantification to 62.3 µg kg-1. Conclusion: The results demonstrated that the developed method was suitable for simultaneously determining twelve EAs in compound feed and could be used for routine analysis.

Brote de "pie de festuca" en un rodeo de cría de Salta, Argentina / Outbreak of fescue foot in a livestock in Salta, Argentina

Festuca arundinacea es una gramínea perenne de uso forrajero muy difundida mundialmente. Aquí se describe un brote de “pie de festuca” en el Noroeste Argentino que afectó a un rodeo de cría con una incidencia del 5%. Los signos clínicos fueron claudicación, edema en los miembros, gangrena en las extremidades y exungulación. La pastura donde los animales permanecían correspondía a un lote naturalizado de festuca siendo el nivel de infestación con Neotyphodium coenophialum del 71%. El cuadro de pie de festuca es muy conocido pero poco informado en Argentina. En el brote el diagnostico se confirmó por los antecedentes clínicos y epidemiológicos sumado al elevado grado de infestación del hongo en la pastura. La toxicidad de la festuca es un problema mundialmente distribuido y en Argentina ha sido más estudiado en la región centro del país. Los antecedentes de toxicidad de esta especie son escasos y aislados en el Noroeste Argentino.

Festuca arundinacea is a perennial grass which use as forage is widespread worldwide. This report describes a “fescue foot” outbreak in the Northwest of Argentina. Beef cattle herd were affected and the incidence was 5%. The clinical signs were claudication, edema in the limbs, gangrene in the extremities and exungulation. The grazing areas where the animals remained corresponded to a natural batch of fescue. Neotyphodium coenophialum infestation level was 71%. The clinical presentation of fescue root is well described but barely reported in Argentina. In the outbreak presented here diagnosis was confirmed by the clinical and epidemiological antecedents as well as the high level of infestation of the pasture. The toxicity of fescue is a worldwide distributed problem. In Argentina, the disease is well known in the central region of the country. Scarce information is available regarding the toxicity of this species for northwestern Argentina.

An Autopsy Case of Postpartum Acute Myocardial Infarction Associated with Postpartum Ergot Alkaloids Administration in Old-Aged Pregnant Women

Cases of acute myocardial infarction (AMI) that occur during pregnancy or postpartum are rarely reported. Ergot derivatives are known to induce the spasmodic contraction of coronary arteries. Administration of ergot derivatives can cause AMI, even in normal healthy people. In several reported cases, ergot derivatives triggered severe AMI during the postpartum period. Here, we report the case of a forty-year-old woman who was successfully impregnated by artificial fertilization and died after treatment with ergot derivatives. The autopsy revealed AMI with severe coronary atherosclerosis. This is the first case that reports aggravation of pre-existent severe coronary atherosclerosis after postpartum infusion of ergot derivtives.

Forensic Consideration of Postpartum Acute Myocardial Infarction Associated with Ergot Derivatives Administration in Old Aged Woman: A Case Report / 대한법의학회지

Cases of acute myocardial infarction (AMI) that occurs during pregnancy or postpartum are rarely reported. Ergot derivatives were known to induce spasmodic contraction of coronary arteries. Administration of ergot derivatives can cause AMI even in normal healthy people. Several cases are also reported that ergot derivatives may trigger severe AMI in postpartum period. Here, we report a case of a forty-year-old pregnant woman who was succeeded to be pregnant by artificial fertilization died after treatment with ergot derivatives. Examinations of autopsy have revealed AMI with severe coronary atherosclerosis. This case is the first report which shows aggravation of preexistent severe coronary atherosclerosis after postpartum infusion of ergot derivatives.