Location: Forage-animal Production Research
2022 Annual Report
Objectives
Objective 1: Determine relationships of ergot alkaloids to receptors in animal tissues and subsequent effects on animal physiology, and the implications of these relationships on clearance from animal tissues.
Subobjective 1.A. Evaluate the effect of ergot alkaloid exposure on vascular biogenic amine receptors.
Subobjective 1.B. Characterize the interaction of isoflavones and their metabolites with ergot alkaloids on vasoactivity.
Subobjective 1.C. Determine if there is ergovaline in bovine portal blood and determine site of absorption.
Subobjective 1.D. Develop an oral endotoxin challenge model to determine the effects of fescue-derived alkaloid consumption on intestinal barrier function and innate immunity in cattle.
Objective 2: Develop cost-effective management approaches to alleviate or mitigate the adverse effects of fescue toxicosis on animal physiology and well-being.
Subobjective 2.A. Determine the impact of combining feeding soybean hulls (SBH) and red clover on weight gain performance of steers grazing toxic E+ tall fescue and mitigation of fescue toxicosis.
Subobjective 2.B. Compare vasorelaxation between different sources of isoflavones for goats exhibiting ergot alkaloid-induced vasoconstriction.
Subobjective 2.C. Determine if rumen tryptophan-utilizing bacteria will degrade ergot alkaloids in vivo.
Subobjective 2.D. Assess phenotypic variation of cattle in their susceptibility to fescue toxicosis.
Subobjective 2.E. Evaluate management approaches to cost effectively add value to cull cows and enhance ground beef quality.
Subobjective 2.F. Comparison of supplemental selenium form to ameliorate physiological and gene expression stress response parameters in white blood cell (WBC), pituitary, adrenal, kidney, and liver of growing steers consuming E+ or E- tall fescue seed.
Subobjective 2.G. Assess effects of animal temperament on fescue toxicosis-induced changes in animal growth and immunological responses.
Objective 3: Determine the biological mechanisms used by certain plant secondary metabolites to function as antimicrobials in ruminants and non-ruminants, and assess their impact on animal health, performance, and well-being.
Subobjective 3.A. Elucidate the antimicrobial mechanism of action of the red clover isoflavone, biochanin A, and determine antagonistic, additive or synergistic activity with other antimicrobials.
Subobjective 3.B. Determine the effect of biochanin A on the rates of antibiotic resistance in ruminants.
Subobjective 3.C. Determine the effect of clover phenolic compounds on nitrogen efficiency and weight gain in lambs.
Subobjective 3.D. Evaluate spent brewer’s yeast as a carrier for hops secondary metabolites (prenylated phloroglucinols) in ruminant production.
Approach
Experiments will be conducted to determine mechanisms by which ergot alkaloids interact with receptors in animal tissues and affect their physiology. Saphenous veins collected from cattle at a local abattoir will be used to determine in vitro if inhibition of the phospholipase C and protein kinase C enzymes will alleviate ergot alkaloid induced vasoconstriction of smooth muscle. An in vitro experiment will ascertain if plant secondary metabolites, isoflavones, can mitigate the vasoactivity caused by ergot alkaloids. Endothelium cells from saphenous cells will also be exposed to ergot alkaloids to assess their effects on receptor signaling by ß-arrestin and G proteins. Catheters will be inserted in in the hepatic, portal and mesenteric veins of six rumen-fistulated steers to determine if rumen infused ergovaline is absorbed by the rumen and small intestines, or if the ergopeptine is degraded by rumen microbes. Field experiments will be conducted to evaluate management approaches to mitigate fescue toxicosis. Rumens will be infused with ergot alkaloids at a diet concentration of 0.8 ppm ergovaline and combined with either ground soybean meal, red clover, white clover, or a no isoflavone control. Cross sectional luminal areas of the right carotid artery of each goat will be measured by color Doppler ultrasonography. A grazing experiment with steers will evaluate the effects of feeding soybean hulls and overseeding toxic endophyte-infected tall fescue with red clover on animal weight gain and well-being. Four combinations of with and without the two treatments will be used as treatments to determine cost effectiveness of the treatments and mitigation of fescue toxicosis. Ear notches and phenotype data will be collected from multiple cow herds to detect polymorphisms of certain genes associated with fescue toxicosis and determine if these polymorphisms can be used to predict genetic tolerance to toxic ergot alkaloids. Three management approaches (soybean hulls, chemical seed head suppression, and red clover) to mitigate fescue toxicosis will be compared for adding body condition and weight to cull cows that graze toxic endophyte-infected tall fescue in either the spring or fall. To assess if selenium can ameliorate fescue toxicosis and if ergot alkaloids suppress immune response and alter gene expression in the liver, steers will be fed selenium depleted diets for 28 days and then switched to either inorganic selenium or an inorganic and organically bound selenium treatments for the remaining 98 days of the trial. The steers will also be fed either endophyte-infected or endophyte-free seed for the final 42 days. Jugular blood will be periodically collected and there will be staggered euthanasia of the steers for tissue collection. Effects of biochanin A on rate of antibiotic resistance will be determined by feeding rumen fistulated steers with either 0, 3 or 6 g biochanin A/day/steer and collecting rumen and fecal samples and using metagenomic DNA for quantitative PCR screening for antibiotic resistant genes.
Progress Report
Sub-objective 1.A. Evaluate the effect of ergot alkaloid exposure on vascular biogenic amine receptors. All proposed experiments have been conducted. Additional experiments looking at intracellular calcium flux were deemed necessary and are ongoing. Some work has been published.
Sub-objective 1.B. Characterize the interaction of isoflavones and their metabolites with ergot alkaloids on vasoactivity. The tissue culture work associated with this sub-objective is complete and data are being analyzed.
Sub-objective 1.C. Determine if there is ergovaline in bovine portal blood and determine site of absorption. Work has been completed and manuscript is in preparation.
Sub-objective 1.D. Develop an oral endotoxin challenge model to determine the effects of fescue-derived alkaloid consumption on intestinal barrier function and innate immunity in cattle. This work has been fully met and a manuscript has been published.
Sub-objective 2.A. Determine the impact of combining feeding soybean hulls (SBH) and red clover on weight gain performance of steers grazing toxic endophyte positive (E+) tall fescue and mitigation of fescue toxicosis A manuscript has been published on the impacts of overseeding red clover in mitigative fescue toxicosis in grazing steers.
Sub-objective 2.B. Compare vasorelaxation between different sources of isoflavones for goats exhibiting ergot alkaloid-induced vasoconstriction. Work has been completed and manuscript was published.
Sub-objective 2.C. Determine if rumen tryptophan-utilizing bacteria will degrade ergot alkaloids in vivo. All experiments have been completed for this sub-objective. Collected data have been analyzed and the resulting manuscripts are in preparation and is expected to be submitted in 2023.
Sub-objective 2.D. Assess phenotypic variation of cattle in their susceptibility to fescue toxicosis. The new phenotype being developed with the Cow Manager system is ongoing.
Sub-objective 2.E. Evaluate management approaches to cost effectively add value to cull cows and enhance ground beef quality. Onset of study was delayed. Feeding portion of the study is ongoing and laboratory analyses expected to be completed in 2023.
Sub-objective 2.F. Comparison of supplemental selenium form to ameliorate physiological and gene expression stress response parameters in white blood cell (WBC), pituitary, adrenal, kidney, and liver of growing steers consuming E+ or endophyte negative (E-) tall fescue seed. Work has been completed. Results have been presented and manuscript is currently under review.
Sub-objective 2.G. Assess effects of animal temperament on fescue toxicosis-induced changes in animal growth and immunological responses. Work has been completed. Results have been presented and manuscript is currently under review.
Sub-objective 3.A. Elucidate the antimicrobial mechanism of action of the red clover isoflavone, biochanin A, and determine antagonistic, additive or synergistic activity with other antimicrobials. Work was prioritized on the polybiochanin A molecule. The manuscript is in preparation.
Sub-objective 3.B. Determine the effect of biochanin A on the rates of antibiotic resistance in ruminants. Samples were collected and analyzed for the number of antibiotic resistant bacteria. Isolates of predominant antibiotic resistant bacteria were sequenced and identified. Steers on a high grain diet carried a substantial sub-population of tetracycline-resistant bacteria in the rumen. Supplementation with biochanin A resulted in a significant reduction (approx. 99.9%) in the number of tetracycline-resistant bacteria. Results were presented, manuscript in preparation.
Sub-objective 3.C. Determine the effect of clover phenolic compounds on nitrogen efficiency and weight gain in lambs. Isolates have been obtained for in vitro testing of biochanin A and other clover phenolic compounds. Three feeding trials with lambs have been conducted to examine nitrogen metabolism by lambs and examine the effects of biochanin A on growth and carcass characteristics.
Sub-objective 3.D. Evaluate spent brewer’s yeast as a carrier for hops secondary metabolites (prenylated phloroglucinols) in ruminant production. Unit scientists wrote a grant proposal with collaborators that was not funded. As a contingency, the hypothesis was tested in vitro and the results published.
Accomplishments
1. Conducted agonist therapy with serotonin to mitigate fescue toxicosis. ARS researchers in Lexington, Kentucky, have previously demonstrated that cattle consuming ergot alkaloids that cause fescue toxicosis have lowered levels of circulating serotonin. A major pathology associated with fescue toxicosis is persistent vasoconstriction and we have demonstrated that is caused by ergot alkaloids binding with serotonin receptors in the vascular smooth muscle and interfering with subsequent receptor function. Agonist therapy is a technique that displaces less desirable compounds at the receptor level with compounds that are less harmful and has proven effective treatment for dependence on neuromodulator drugs such as opioids. In vitro vascular research was conducted by ARS researchers in Lexington, Kentucky, to determine if a controlled increase in serotonin can compete with ergot alkaloids at the receptor and reduce the sustained vasoconstriction caused by ergot alkaloids. Results suggested that there is competition between serotonin and ergot alkaloids at the receptor level. The impact that elevating circulating serotonin levels could have, is a decrease in available binding sites for toxic ergot alkaloids and this could mitigate fescue toxicosis. This work could provide another tool for producers to use to mitigate fescue toxicosis.
2. Spent brewer’s yeast kills methane- and ammonia-producing rumen microorganisms. Methane and ammonia are byproducts of rumen fermentation that do not promote animal growth, and methane is a key contributor to anthropogenic climate disruption. Emerging research has shown that humulones and lupulones, molecules that are found in the cones of the hops plant have potential to decrease methane and ammonia by selective inhibition of rumen microorganisms. Spent yeast is produced during the beer-brewing process and contains humulones and lupulones in concentrations that vary by beer style but is generally discarded as waste. When ARS researchers in Lexington, Kentucky, added spent craft brewer’s yeast to fermentations with rumen microbes the production of methane and ammonia were suppressed. The hop acid concentrations in spent yeast obtained from six beer styles produced at a brewery. There was a strong correlation between the quantities of hop acids in the spent yeast and the reduction of methane and ammonia. Notably, two of the yeast samples inhibited methane production to a greater degree than an antibiotic that was classically used for that purpose. These results suggest that spent brewer’s yeast, a brewery co-product, has potential to improve ruminant growth while reducing greenhouse gas emission.
3. Determined that legume supplements in goat feed can reduce vasoconstriction associated with fescue toxicosis. A variety of legumes are commonly utilized in livestock production to improve diet quality. Legumes also contain vasodilatory isoflavones that have been shown to alleviate fescue toxicosis in goats and grazing cattle. However, these legume species can vary in both isoflavone concentration and composition. A pen study was conducted by ARS researchers in Lexington, Kentucky, to determine if isoflavone supplementation via red clover, white clover, or soybean meal could mitigate vasoconstriction associated with fescue toxicosis in goats. Rumen fistulated wether goats were assigned to each legume treatment at equal levels of supplementation by weight. Goats were subjected to a fescue toxicosis challenge with toxic tall fescue seed, and the carotid artery area was monitored using Doppler ultrasonography. All isoflavone treatments were able to partially alleviate vasoconstriction. Red clover, with the greatest concentration of isoflavones, was the most effective (+40% artery area). These results demonstrate that red clover, white clover, and soybean meal supplementation could be used to reverse the vasoconstriction associated with fescue toxicosis in goats despite differences in isoflavone concentration and composition. The impact of this research is a legume-derived phytochemical that can be applied in ruminants consuming toxic tall fescue to reverse fescue toxicosis and improve animal health and productivity.
Review Publications
Bryant, R.W., Burns, E.E., Feidler-Cree, C., Carlton, D., Flythe, M.D., Martin, L.J. 2021. Spent craft brewer’s yeast reduces production of methane and ammonia by bovine rumen microbes. Frontiers in Animal Science. 2. Article 720646. https://doi.org/10.3389/fanim.2021.720646.
Valente, E.E., Harmon, D.L., Klotz, J.L. 2021. Influence of prolonged serotonin and ergovaline pre-exposure on vasoconstriction ex vivo. Toxins. 14(1):9. https://doi.org/10.3390/toxins14010009.
Klotz, J.L. 2022. Global impact of ergot alkaloids. Toxins. 14(3). Article 186. https://doi.org/10.3390/toxins14030186.
Harlow, B.E., Flythe, M.D., Goodman, J.P., Ji, H., Aiken, G.E. 2022. Isoflavone containing legumes mitigate ergot alkaloid-induced vasoconstriction in goats (Capra hircus). Animals. 12(6). Article 750. https://doi.org/10.3390/ani12060750.
Loos, C.M., Urschel, K.L., Vanzant, E.S., Oberhaus, E.L., Bohannan, A.D., Klotz, J.L., Mcleod, K.M. 2022. Effects of bromocriptine on glucose and insulin dynamics in normal and insulin dysregulated horses. Frontiers in Veterinary Science. 9. Article 889888. https://doi.org/10.3389/fvets.2022.889888.
Harlow, B.E., Flythe, M.D., Hamilton, T.A., Ji, H., Schrick, F., Aiken, G.E. 2021. Effects of overseeding red clover in endophyte-infected tall fescue pastures on steer physiology and performance. Applied Animal Science. 37(6):748-757. https://doi.org/10.15232/aas.2021-02152.
Harlow, B.E., Flythe, M.D., Klotz, J.L., Harmon, D.L., Aiken, G.E. 2021. Effect of biochanin A on the rumen microbial community of Holstein steers consuming a high fiber diet and subjected to a subacute acidosis challenge. PLoS ONE. 16(7). Article e0253754. https://doi.org/10.1371/journal.pone.0253754.
Zou, J., Reddivari, L., Shi, Z., Li, S., Wang, Y., Bretin, A., Ngo, V.L., Flythe, M.D., Pellizzon, M., Chassaing, B., Gewirtz, A. 2021. Inulin fermentable fiber ameliorates type I diabetes via IL22 and short-chain fatty acids in experimental models. Cellular and Molecular Gastoenterology and Hepatology. 12(3):983-1000. https://doi.org/10.1016/j.jcmgh.2021.04.014.
