Biochemistry of hemlock (Conium maculatum L.) alkaloids and their acute and chronic toxicity in livestock. A review.

The literature on Conium maculatum biochemistry and toxicology, dispersed in a large number of scientific publications, has been put together in this review. C. maculatum is a weed known almost worldwide by its toxicity to many domestic animals and to human beings. It is an Umbelliferae, characterized by long, hollow stems, reaching up to 2 m height at maturity, producing a large amount of lush foliage during its vegetative growth. Its flowers are white, grouped in umbels formed by numerous umbellules. It produces a large number of seeds that allow the plant to form thick stands in modified soils, sometimes encroaching on cultivated fields, to the extent of impeding the growth of any other vegetation inside the C. maculatum area of growth. Eight piperidinic alkaloids have been identified in this species. Two of them, gamma-coniceine and coniine are generally the most abundant and they account for most of the plant acute and chronic toxicity. These alkaloids are synthesized by the plant from eight acetate units from the metabolic pool, forming a polyketoacid which cyclises through an aminotransferase and forms gamma-coniceine as the parent alkaloid via reduction by a NADPH-dependent reductase. The acute toxicity is observed when animals ingest C. maculatum vegetative and flowering plants and seeds. In a short time the alkaloids produce a neuromuscular blockage conducive to death when the respiratory muscles are affected. The chronic toxicity affects only pregnant animals. When they are poisoned by C. maculatum during the fetuses organ formation period, the offspring is born with malformations, mainly palatoschisis and multiple congenital contractures (MCC; frequently described as arthrogryposis). Acute toxicity, if not lethal, may resolve in the spontaneous recovery of the affected animals provided further exposure to C. maculatum is avoided. It has been observed that poisoned animals tend to return to feed on this plant. Chronic toxicity is irreversible and although MCC can be surgically corrected in some cases, most of the malformed animals are lost. Since no specific antidote is available, prevention is the only way to deal with the production loses caused by this weed. Control with herbicides and grazing with less susceptible animals (such as sheep) have been suggested. C. maculatum alkaloids can be transferred to milk and to fowl muscle tissue through which the former can reach the human food chain. The losses produced by C. maculatum chronic toxicity may be largely underestimated, at least in some regions, because of the difficulty in associate malformations in offspring with the much earlier maternal poisoning.

Ergotism and photosensitization in swine produced by the combined ingestion of Claviceps purpurea sclerotia and Ammi majus seeds.

Poisoning of domestic animals happens frequently in the southeast of Buenos Aires Province (Argentina). Intoxications are produced mainly by the ingestion of plants and mycotoxins, but animals are rarely affected simultaneously by both types of agents. One herd of pigs suffered simultaneous intoxications by ergot alkaloids from Claviceps purpurea sclerotia and furocoumarins from Ammi majus seeds. Pigs were fed a diet composed of wheat (poor quality) or corn and protein and vitamin supplements. This diet was completed with forage sorghum. Nervous signs were first observed 5-7 days after the initiation of feeding the suspect ration. These signs were followed by cutaneous irritation. Snout ulcers, eyelid edema, and conjunctivitis were observed in several piglets. Ten days after the start of feeding the incriminated ration, 8 abortions were observed. Many of the sows that were nursing piglets developed udder edema and teat cracking. Dermal lesions were observed in most of the animals with unpigmented areas in the skin but not in a Duroc-Jersey boar. Removal of the incriminated diet and feeding of another diet prepared with good-quality wheat allowed all the animals to recover in 15 days. The herd experienced normal pregnancies and parturitions, litter sizes, and piglet weights when fed a cleaned portion of the poor-quality wheat. No photosensitization lesions were observed. Examination of impurities in the suspected wheat indicated the presence of 2.2% of A. majus seeds and 0.14% of C. purpurea sclerotia. The quantitative analysis indicated the presence of 3.2 g xanthotoxin and 0.65 g bergaptene/100 g A. majus seeds and 0.73 g ergot alkaloids (expressed as ergonovine) per 100 g of C. purpurea sclerotia. Qualitative analysis demonstrated the presence of ergotamine, ergocristine, and ergonovine. These results indicate that clinical signs and lesions were caused by the ingestion of large quantities of these biologically active compounds.

Fusarium crookwellense-produced zearalenone in maize stubble in the field.

Waste grain and vegetative material (stems and leaves) collected from a maize field several months after harvest was analysed by bi-dimensional thin layer chromatography for the presence of aflatoxins, deoxynivalenol, ochratoxin A, sterigmatocystin, T-2 toxin and zearalenone. Deoxinivalenol (0.7 mg/kg) and T-2 toxin (4.1 mg/kg) were found in the grain and zearalenone (3.0 mg/ kg) was found in the stem and leaf. No other toxins were detected. The stubble was examined for the presence of potentially toxigenic Fusarium species, and F. poae, F. moniliforme var. subglutinans and F. crookwellense were isolated and identified. When these isolates were cultured on cracked corn, only F. crookwellense was found to produce micotoxins and then only zearalenone was detected. As corn stubble is commonly grazed in Argentina and in other countries, these findings identify a further source of mycotoxins that may adversely affect animal health and productivity.

Experimental toxicity of verbesina encelioides in sheep and isolation of galegine.

Verbesina encelioides is a widely distributed weed in Argentina. Although it has been suspected as poisonous, there seem to be no previous reports of its toxicity in this country. Its biological activity was evaluated through an experiment in sheep. Four 5-y-old sheep, averaging 33 kg in body weight, were given a single dose of 1.0, 3.2, 5.0 or 6.3 g of dried plant material (17.9% dry matter)/kg body weight orally in a water suspension. Forty-eight hours later, the animals given 5.0 g or 6.3 g showed dullness and lack of appetite. The higher dosed sheep became recumbent a few hours later and died 60 h after dosing without showing further signs. The other animals showed no signs. Necropsy of the dead sheep showed severe lesions in different organs: liver, kidneys, lungs, lymph nodes and digestive tract. Hydrothorax was also observed. The animal receiving 5.0 g/kg was sacrificed after 72 h to determine the degree of recovery or lesions. Microscopically, severe glomerulonephrosis and congestion in the liver, with cellular degeneration and fatty changes were observed. Additionally, hemorrhagic lymph nodes, and hemorrhagic and edematous lungs were noted. No gross nor microscopic lesions were found in the sheep receiving 5.0 g/kg. Mice injected ip with the chromatographic extract died in less than 15 min. Galegine was identified in the plant material. The minimum toxic dose of 5-6 g/kg indicates a high toxicity when compared to other poisonous plants of the southeastern area of Buenos Aires province, Argentina.

Fluoride-induced early teeth wearing in Argentinian cattle.

The relationship between observed early teeth wearing in cattle and the existence of large areas where excess fluoride in underground water prevails in Argentina, but has never been studied. Average values of bone fluoride exceeding 5000 ppm and well water containing up to 10.5 ppm of this element were found in a farm (ETWF) where early teeth wearing is observed, while in a control farm (NETWF), those values were 1480 ppm and 3.0 ppm respectively. Urine samples from animals from the ETWF contained almost 4 times more fluoride than those from the latter. Forage in both farms contained about the same levels of fluoride (15 to 25 ppm). None of the farms was exposed to industrial, volcanic, or geothermal activity. Lactating and adult animals were transferred from one farm to the other. After 30 months, adult animals from the NETWF showed no damage in teeth, while adults from the ETWF continued a rapid teeth wearing. Young animals from the NETWF developed severely damaged permanent teeth, while young animals from the ETWF developed normal teeth. We concluded that the problem exists in the ETWF and being a standard farm, the condition could be extrapolated to other areas.

Influence of temperature on zearalenone production by regional strains of Fusarium graminearum and Fusarium oxysporum in culture.

Zearalenone production by Fusarium graminearum and Fusarium oxysporum was studied under two temperature conditions. Incubation at 25 degrees C for 4 weeks enhanced zearalenone synthesis, improving detection of zearalenone-producing strains of Fusarium oxysporum. Zearalenone production was either totally or partially inhibited when temperature was lowered to 12-14 degrees C during the last 2 weeks of incubation.

Lack of effect of a diet containing zearalenone on spermatogenesis in rams.

Six breeding rams were fed a diet containing 12 mg of zearalenone daily for eight weeks. A control group of six rams was fed a diet free of zearalenone. The weekly production of spermatozoa of both groups was measured during the period of administration of zearalenone and for six weeks after the administration of zearalenone ceased. Semen production was measured in terms of the volume of ejaculate and its concentration, and the motility and abnormalities in the spermatozoa. The feeding of zearalenone had no significant effects on any of these measurements.

Importance of the development of veterinary toxicology in Argentina.

The different types of toxic substances that affect farm animals in Argentina are described. These substances include (a) toxic plants, such as Solanum glaucophyllum, Baccharis coridifolia, Festuca arundinacea; (b) mycotoxins, such as zearalenone, tremorgenic mycotoxins, and ergoalkaloid; (c) trace elements, such as fluorine, copper, molybdenum, and arsenic; and (d) others, such as toxic algae and pesticides. The direct and indirect economic loss of farm animals associated with these toxic substances in Argentina is roughly estimated at 50 million dollars a year. A number of recommendations are made to ameliorate the situation.

[Hyperestrogenism in swine due to natural poisoning with zearalenone]. / Hiperestrogenismo en cerdas por intoxicación natural con zearalenona.

A field case is described in which all prepuberal swine of a group of 20 pigs and 11 sows showed marked estrogenic effects. These consisted of enlarged mammary glands, swelled tumefacient vulva, and greatly enlarged internal reproductive organs. The corn used to feed these animals was found to contain 56 ppm zearalenone. Deoxynivalenol (4.9 ppm) was found in the corn; T-2 toxin, nivalenol, fusarenon-X, diacetoxyscirpenol, aflatoxins and ochratoxins were absent. Identity of Z was confirmed by TLC in four solvent systems, behavior of the suspected spots under UV light of different wavelengths, change of fluorescence from green to blue after spraying with 5% AlCl3 in alcohol and heating at 110 degrees C during 5 minutes, and by its UV spectrum. A zearalenone producing strain of Fusarium oxysporum was isolated from the suspected grain. Histopathology of uterine tissue showed typical changes produced by zearalenone: hyperplasia, hypertrophy, and metaplasia of the myometrium. Feeding of the grain to a prepuberal sow under controlled conditions reproduced all the effect found in the farm animals. This is the first field case of zearalenone poisoning reported in Argentina.

Hiperestrogenismo en cerdas por intoxicación natural con zearalenona / Hyperestrogenism in swine due to natural poisoning with zearalenone

Se describe un caso de intoxicación de cerdas prepúberes con zearalenona (Z) al consumir una dieta conteniendo 56 ppm Z. En el maíz contaminado se encontró una cepa de Fusarium oxysporum que produjo Z en cultivo sobre ese grano. Se indicaron la evidencias de distinta índole que permitieron confirmar que los efecto observados fueron producidos por dicha micotoxina

Hiperestrogenismo en cerdas por intoxicación natural con zearalenona. / [Hyperestrogenism in swine due to natural poisoning with zearalenone]

A field case is described in which all prepuberal swine of a group of 20 pigs and 11 sows showed marked estrogenic effects. These consisted of enlarged mammary glands, swelled tumefacient vulva, and greatly enlarged internal reproductive organs. The corn used to feed these animals was found to contain 56 ppm zearalenone. Deoxynivalenol (4.9 ppm) was found in the corn; T-2 toxin, nivalenol, fusarenon-X, diacetoxyscirpenol, aflatoxins and ochratoxins were absent. Identity of Z was confirmed by TLC in four solvent systems, behavior of the suspected spots under UV light of different wavelengths, change of fluorescence from green to blue after spraying with 5

AlCl3 in alcohol and heating at 110 degrees C during 5 minutes, and by its UV spectrum. A zearalenone producing strain of Fusarium oxysporum was isolated from the suspected grain. Histopathology of uterine tissue showed typical changes produced by zearalenone: hyperplasia, hypertrophy, and metaplasia of the myometrium. Feeding of the grain to a prepuberal sow under controlled conditions reproduced all the effect found in the farm animals. This is the first field case of zearalenone poisoning reported in Argentina.

Hiperestrogenismo en cerdas por intoxicación natural con zearalenona / Hyperestrogenism in swine due to natural poisoning with zearalenone

Se describe un caso de intoxicación de cerdas prepúberes con zearalenona (Z) al consumir una dieta conteniendo 56 ppm Z. En el maíz contaminado se encontró una cepa de Fusarium oxysporum que produjo Z en cultivo sobre ese grano. Se indicaron la evidencias de distinta índole que permitieron confirmar que los efecto observados fueron producidos por dicha micotoxina (AU)

[Identification of the mycotoxin zearalenone in Argentina]. / Identificación de la micotoxina zearalenona en la Argentina.

Zearalenone, an estrogenic toxin produced by several Fusarium sp., was detected by two chromatographic methods in grain and food samples from several farms of th Buenos Aires Province. Suspected zearalenone spots were identified by comparing their Rf against that of the pure substance, by their change of behaviour under ultraviolet light of different wavelengths, by comparing the Rf against a standard preparation after thin layer chromatography in four different solvent systems, and by color reactions obtained after spraying reagents on the thin layer chromatographic plates. Sixteen of 55 samples were positive, with concentrations of the toxin ranging from 200 ppb to 750 ppb. Only Fusarium graminearum was isolated from eleven of these samples, indicating that the fungus was inactivated by heat or antifungal treatments.

Identificación de la micotoxina zearalenona en la Argentina / [Identification of the mycotoxin zearalenone in Argentina]

Zearalenone, an estrogenic toxin produced by several Fusarium sp., was detected by two chromatographic methods in grain and food samples from several farms of th Buenos Aires Province. Suspected zearalenone spots were identified by comparing their Rf against that of the pure substance, by their change of behaviour under ultraviolet light of different wavelengths, by comparing the Rf against a standard preparation after thin layer chromatography in four different solvent systems, and by color reactions obtained after spraying reagents on the thin layer chromatographic plates. Sixteen of 55 samples were positive, with concentrations of the toxin ranging from 200 ppb to 750 ppb. Only Fusarium graminearum was isolated from eleven of these samples, indicating that the fungus was inactivated by heat or antifungal treatments.

Identificación de la micotoxina zearalenona en la Argentina. / [Identification of the mycotoxin zearalenone in Argentina]

Zearalenone, an estrogenic toxin produced by several Fusarium sp., was detected by two chromatographic methods in grain and food samples from several farms of th Buenos Aires Province. Suspected zearalenone spots were identified by comparing their Rf against that of the pure substance, by their change of behaviour under ultraviolet light of different wavelengths, by comparing the Rf against a standard preparation after thin layer chromatography in four different solvent systems, and by color reactions obtained after spraying reagents on the thin layer chromatographic plates. Sixteen of 55 samples were positive, with concentrations of the toxin ranging from 200 ppb to 750 ppb. Only Fusarium graminearum was isolated from eleven of these samples, indicating that the fungus was inactivated by heat or antifungal treatments.