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United States Department of Agriculture

Agricultural Research Service


Location: Poisonous Plant Research

2013 Annual Report

1a.Objectives (from AD-416):
Objective I: Pine Needles 1.1 Determine if isocupressic acid (ICA; the abortifacient compound in pine needles) concentration in pine needles is modulated by the environment. 1.2 Identify the matabolites of isocupressic acid in pine needles that cause abortions in cattle. Determine the biological mechanism, develop diagnostic techniques, and therapeutic procedures. 1.3 Determine factors that influence cattle consumption of pine needles and develop management practices to prevent abortion. Objective II: Broom Snakeweed 2.1 Identify the toxic and abortifacient compounds in broom snakeweed. 2.2 Describe the ecology of broom snakeweed, develop management and control guidelines to reduce incidence of poisoning and abortion in livestock. Objective III: Lupine 3.1 Isolate, identify, and evaluate toxicity and teratogenicity of lupine alkaloids which cause birth defects in calves born to cows that graze these plants. 3.2 Evaluate the role of genotype and environment on lupine alkaloids, and thus the relative toxicity of various species and populations of lupine. 3.3 Determine the physiological mechanism of lupine-induced birth defects and evaluate the maternal and fetal toxicokinetics of alkaloids. 3.4 Evaluate the influence of climate on population cycles of lupine. 3.5 Determine the importance of lupines as nutritional components for cattle during critical times of the year. 3.6 Identify conditions under which cattle graze various lupine species. Objective IV: Veratrum 4.1 Develop models to study the toxicokinetics, including clearance times, and toxicity of steroidal alkaloids in Veratrum californicum.

1b.Approach (from AD-416):
1.1 Data on environmental conditions will be collected at each site using local weather stations. ICA levels and environmental conditions will be correlated to determine if any patterns emerge. Soil samples will be collected at each site for future evaluation. 1.2 Samples of maternal and fetal tissues will be collected for histologic analysis and determination of ICA concentrations using existing ELISA’s and GC/MS methods. Proteomic analyses via LC/MS/MS techniques will be done. 1.3 Pen and field studies using cattle in high, medium and low body condition will be done to determine effects on needle consumption and grazing times. Nutrient supplements will be offered to determine if pine needle consumption will be altered. 2.1 The diterpene acid “fingerprint” of broom snakeweed from various populations in Arizona, New Mexico and Utah will be determined by chemical analysis. Subsequent in vitro and in vivo studies will be done to determine abortifacient activity. 2.2 A grazing study will be conducted to determine if various management practices can be implemented to force cattle to graze snakeweed as a biological control. A clipping study will be conducted to further describe the effects of defoliation on snakeweed and the surrounding plant community. 3.1 Alkaloids will be isolated by chemical methods and identified by chromatography, NMR, mass spectrometry, and elemental analysis. Toxicology will be evaluated using a mouse bioassay and cell lines that express nicotinic acetylcholine receptors. 3.2 A chemical fingerprint of Lupinus sulphureus collected from different locations will be generated using chemical methods. Fingerprints will be analyzed via cluster analysis and phylogenetic analysis will be performed using AFLPs (Amplified Fragment Length Polymorphisms) to determine the genetic relationship of the populations. 3.3 Pregnant goats in late gestation will be used to determine the rate of absorption, distribution and elimination of the teratogenic alkaloids. The pharmacokinetic profiles of the alkaloids will be compared between maternal and fetal systems. 3.4 Established transects will be monitored over the next 5 years to determine the influence of weather patterns on lupine density. Correlations of lupine age, class, density, and trends will be made with seasonal precipitation and temperature. 3.5 Consumption of lupines by cattle on rangelands dominated by low quality forages may be related to nutrient content. Twelve yearling heifers in a field study will be supplemented with different levels of protein to compare lupine ingestion. 3.6 Short-duration and high intensity grazing studies in early, mid, and late summer will be used to determine what role grazing pressure has on lupine intake during different seasons of the year. 4.1 A monogastric model (swine) will be used to determine the kinetics (clearance and metabolism) of a well known teratogenic alkaloid from Veratrum. This pilot project will be a model for testing the clearance of other plant toxins from animal tissues to evaluate food safety of animal products. Clearance rates between the monogastric model and small ruminant model will be compared.

3.Progress Report:
This is the final report for project 5428-31320-005-00D. In collaboration with the Forage and Range Research Laboratory, Logan, UT, scientists at the Poisonous Plant Research Laboratory continued evaluations of replicated plots of various grass species and forage Kochia in Eastern Washington for forage production. Results from the first year of grazing studies on these plots with cattle were evaluated and a second year of grazing studies was conducted. A three year research project was initiated to determine whether the abortifacient compounds in western juniper trees change throughout the year, or from year to year. To this end needles, bark, and berries were collected bimonthly from western juniper trees at three locations across the state of Oregon. Additional work was performed to determine if metabolites of isocupressic acid detected in stomach and thoracic fluids from aborted calves can be used a diagnostic tool for identification of pine needle abortion cases. The replacement project is 5428-32630-012-00D.

1. A rodent model to identify teratogenic compounds in plants. Mimosa tenuiflora is a perennial tree or shrub native to Brazil, and is also distributed throughout tropical portions of South and Central America and Mexico. The plant causes deformed offspring (teratogenesis) when eaten by pregnant livestock, however, the toxin(s) is not known. Collaborative work by ARS scientists at Logan, UT was undertaken with scientists from Brazil to identify the teratogenic compounds using a rat model. Various purified compounds and mixtures of purified compounds were evaluated for their ability to cause fetal deformation and/or to inhibit fetal movement. A mixture of two purified compounds did cause birth defects; however, clear-cut teratogenic relationships were not established. Consequently, the mechanism by which Mimosa tenuiflora produces birth defects remains unknown. Identification of the compound(s) responsible for causing birth defects will allow for the quantitation of risk for specific plant populations so that better management recommendations can be provided to Brazilian livestock producers to prevent livestock losses.

2. The actions of piperidine and pyridine alkaloids on fetal movement in pregnant goats. Plants containing alkaloids that affect normal fetal development in livestock are found on US rangelands. The developing fetus can be poisoned when the pregnant female consumes plants which cause fetal deformities. The Nicotiana and Lupinus genera contain a variety of toxic piperidine and pyridine alkaloids. However, there is little information available on the actions of these alkaloids on fetal development. ARS scientists at Logan, Utah determined that the piperidine alkaloids anabasine and lobeline are more effective than the pyridine alkaloid myosmine at inhibiting fetal movement in a goat model. Unexpectedly, the pyridine alkaloid myosmine was more effective at inhibiting fetal movement than lobeline. These results suggest that the alkaloids tested are agonists at fetal muscle-type nicotinic receptors and that they have the potential to reduce or inhibit fetal movement in goats to cause fetal deformities. A more thorough understanding of the mechanism of action of the plant toxins will aid in the prevention and treatment of birth defects in livestock.

3. Characterization of the alkaloid profiles of seven Sophora and Dermatophyllum species. Species of Sophora are known to contain quinolizidine alkaloids that are toxic and potentially teratogenic. Recently several Sophora species have been reclassified as Dermatophyllum species. ARS scientists in Logan, Utah characterized the quinolizidine alkaloid composition in seven Sophora and Dermatophyllum species throughout their geographical distribution using field collections and herbarium specimens. Several species were found to contain quinolizidine alkaloids, including the teratogen anagyrine. None of the species contained the neurotoxin swainsonine as had been suggested. The results from these studies provide valuable information regarding the potential risk of Sophora and Dermatophyllum species to cause birth defects in livestock species. Characterization of the alkaloid profiles in these species provides valuable information to evaluate potential risks associated with these plants. This knowledge will result in management recommendations to help livestock producers to prevent livestock losses associated with these plants. Additionally, the chemotaxonomic data from these studies support the recent reclassification of the Dermatophyllum species.

4. A grazing strategy to reduce impact of lupine induced crooked calf syndrome. ARS scientists at Logan, Utah determined that intermittent grazing of lupine could potentially mitigate or substantially reduce the impact of lupine to cause crooked calf syndrome. Pregnant cows were fed lupine in a ten day on 5 day off regiment for 30 days and compared with cows fed lupine daily for 30 days. Those cows fed lupine for 30 days uninterrupted gave birth to severely deformed calves all of which were humanely euthanized. Those cows fed lupine on the intermittent regiment produced calves that were marginally affected i.e. slight contractures which resolved spontaneously or were normal. This method confirms that removing cattle from lupine exposure, even for a short period, will allow calves to regain normal fetal movement and will prevent the most severe contracture birth defects. This method has been implemented by some cattle ranchers in Washington State with great success allowing producers to utilize these productive high risk pastures and reduce the impact from lupine.

Review Publications
Green, B.T., Lee, S.T., Welch, K.D., Pfister, J.A., Panter, K.E. 2013. Fetal-muscle type nicotinic acetylcholine receptor activation in TE-671 cells, and inhibition of fetal movement in a day 40 pregnant goat model by optical isomers of the piperidine alkaloid coniine. Journal of Pharmacology and Experimental Therapeutics. 344:1-13.

Green, B.T., Lee, S.T., Welch, K.D., Pfister, J.A., Panter, K.E. 2013. Piperidine, pyridine alkaloid inhibition of fetal movement in a day 40 pregnant goat model. Food and Chemical Toxicology. 58:8-13.

Lee, S.T., Cook, D., Molyneux, R.J., Davis, T.Z., Gardner, D.R. 2013. Alkaloid profiles of Dermatophyllum arizonicum, Dermatophllum gypsophilum, Dermatophyllum secundiflorum, Styphnolobium affine, and Styphnolobium japonicum previously classified as Sophora species. Biochemical Systematics and Ecology. 49:87-93.

Panter, K.E., Gay, C.C., Clinesmith, R., Platt, T.E. 2013. Management practices to reduce lupine-induced Crooked Calf Syndrome in the Northwest. Rangelands. 35(2): 12-6.

Welch, K.D., Cook, D., Gardner, D.R., Parsons, C., Pfister, J.A., Panter, K.E. 2013. A comparison of the abortifacient risk of western juniper trees in Oregon. Rangelands. 35(1):40-4.

Welch, K.D., Gardner, D.R., Pfister, J.A., Panter, K.E., Zieglar, J., Hall, J. 2012. A comparison of the metabolism of the abortifacient compounds from Ponderosa Pine needles in conditioned versus naive cattle. Journal of Animal Science. 90(12): 4611-7.

Welch, K.D., Panter, K.E., Stegelmeier, B.L., Lee, S.T., Gardner, D.R., Cook, D. 2012. Veratrum-induced placental dysplasia in sheep. International Journal of Poisonous Plant Research. 2:54-62.

Last Modified: 4/20/2014
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