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1.
J Chem Ecol ; 45(10): 879-887, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31686336

RESUMO

Many species of morning glories (Convolvulaceae) form symbioses with seed-transmitted Periglandula fungal endosymbionts, which produce ergot alkaloids and may contribute to defensive mutualism. Allocation of seed-borne ergot alkaloids to various tissues of several Ipomoea species has been demonstrated, including roots of I. tricolor. The goal of this study was to determine if infection of I. tricolor by the Periglandula sp. endosymbiont affects Southern root-knot nematode (Meloidogyne incognita) gall formation and host plant biomass. We hypothesized that I. tricolor plants infected by Periglandula (E+) would develop fewer nematode-induced galls compared to non-symbiotic plants (E-). E+ or E- status of plant lines was confirmed by testing methanol extracts from individual seeds for endosymbiont-produced ergot alkaloids. To test the effects of Periglandula on nematode colonization, E+ and E- I. tricolor seedlings were grown in soil infested with high densities of M. incognita nematodes (N+) or no nematodes (N-) for four weeks in the greenhouse before harvesting. After harvest, nematode colonization of roots was visualized microscopically, and total gall number and plant biomass were quantified. Four ergot alkaloids were detected in roots of E+ plants, but no alkaloids were found in E- plants. Gall formation was reduced by 50% in E+ plants compared to E- plants, independent of root biomass. Both N+ plants and E+ plants had significantly reduced biomass compared to N- and E- plants, respectively. These results demonstrate Periglandula's defensive role against biotic enemies, albeit with a potential trade-off with host plant growth.


Assuntos
Alcaloides de Claviceps/química , Hypocreales/metabolismo , Ipomoea/parasitologia , Tylenchoidea/fisiologia , Animais , Biomassa , Cromatografia Líquida de Alta Pressão , Alcaloides de Claviceps/análise , Ipomoea/química , Ipomoea/metabolismo , Raízes de Plantas/metabolismo , Raízes de Plantas/parasitologia , Tumores de Planta/parasitologia , Sementes/química , Sementes/metabolismo , Solo/parasitologia , Espectrometria de Massas por Ionização por Electrospray , Simbiose
2.
Biotechnol Lett ; 41(12): 1439-1449, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31659576

RESUMO

OBJECTIVE: To enhance ergot alkaloid production of Claviceps purpurea Cp-1 strain by epigenetic modification approach. RESULTS: The chemical epigenetic modifiers were screened to promote ergot alkaloid production of the Cp-1 strain. The histone deacetylase inhibitor suberoylanilide hydroxamic acid (SAHA) was found to significantly enhance the alkaloid productivity of the strain. Particularly, the titers of total ergot alkaloids were gradually increased with the increase of SAHA concentration in the fermentation medium, and the highest production of ergot alkaloids could be achieved at the concentration of 500 µM SAHA. Specially, the titers of ergometrine and total ergot alkaloids were as high as 95.4 mg/L and 179.7 mg/L, respectively, which were twice of those of the control. Furthermore, the mRNA expression levels of the most functional genes in the ergot alkaloid synthesis (EAS) gene cluster were up-regulated under SAHA treatment. It was proposed that SAHA might increase histone acetylation in the EAS gene cluster region in the chromosome, which would loosen the chromosome structure, and subsequently up-regulate the mRNA expression levels of genes involved in the biosynthesis of ergot alkaloids, thereby resulting in the markedly increase in the production of ergot alkaloids. CONCLUSIONS: The ergot alkaloid production by the C. purpurea Cp-1 strain can be effectively increased by the application of histone deacetylase inhibitor. Our work provides a reference for using the chemical epigenetic modifiers to improve SM production in other fungi.


Assuntos
Vias Biossintéticas/genética , Claviceps/genética , Claviceps/metabolismo , Epigênese Genética , Alcaloides de Claviceps/biossíntese , Vias Biossintéticas/efeitos dos fármacos , Claviceps/efeitos dos fármacos , Fermentação , Inibidores de Histona Desacetilases/metabolismo
3.
Inorg Chem ; 58(20): 13771-13781, 2019 Oct 21.
Artigo em Inglês | MEDLINE | ID: mdl-31560525

RESUMO

Cycloclavine is a complex ergot alkaloid containing an unusual cyclopropyl moiety, which has a wide range of biological activities and pharmaceutical applications. The biosynthesis of cycloclavine requires a series of enzymes, one of which is a nonheme FeII/α-ketoglutarate-dependent (aKG) oxidase (Aj_EasH). According to the previous proposal, the cyclopropyl ring formation catalyzed by Aj_EasH follows an unprecedented oxidative mechanism; however, the reaction details are unknown. In this article, on the basis of the recently obtained crystal structure of Aj_EasH (EasH from Aspergillus japonicas), the reactant models were built, and the reaction details were investigated by performing QM-only and combined QM and MM calculations. Our calculation results reveal that the biosynthesis of cyclopropyl moiety involves a radical intermediate rather than a carbocationic or carbanionic intermediate as in the biosynthesis of terpenoid family. The iron(IV)-oxo first abstracts a hydrogen atom from the substrate to trigger the reaction, and then the generated radical intermediate undergoes ring rearrangement to form the fused 5-3 ring system of cycloclavine. On the basis of our calculations, the absolute configuration of the cycloclavine catalyzed by Aj_EasH from Aspergillus japonicus should be (5R,8R,10R), which is different from the product isolated from Ipomoea hildebrandtii (5R,8S,10S). Residues at the active site play an important role in substrate binding, ring rearrangement, and enantioselectivity.


Assuntos
Aspergillus/enzimologia , Alcaloides de Claviceps/biossíntese , Alcaloides Indólicos/metabolismo , Ácidos Cetoglutáricos/metabolismo , Oxirredutases/metabolismo , Biocatálise , Alcaloides de Claviceps/química , Alcaloides Indólicos/química , Ácidos Cetoglutáricos/química , Modelos Moleculares , Conformação Molecular , Oxirredutases/química , Teoria Quântica , Estereoisomerismo
4.
J Anim Sci ; 97(9): 3776-3785, 2019 Sep 03.
Artigo em Inglês | MEDLINE | ID: mdl-31359059

RESUMO

Fescue toxicosis is a multifaceted syndrome common in cattle grazing endophyte-infected tall fescue and is detrimental to growth and performance. Recent research has shown that supplementing protein has the potential to enhance growth performance in weaned steers. Therefore, the objective of this study was to evaluate the effect of supplemental CP on physiological parameters in stocker steers experiencing fescue toxicosis. Thirty-six weaned Angus steers (6 mo of age) stratified by weight (196.1 ± 3.6 kg) were assigned to a 2 × 2 factorial arrangement for 56 d: endophyte-free (EF) seed and 14% CP (EF-14; n = 9), EF seed and 18% CP (EF-18; n = 9), endophyte-infected (EI) seed and 14% CP (EI-14; n = 9), and EI seed and 18% CP (EI-18; n = 9). Steer growth and hemodynamic responses were collected weekly during ergot alkaloid exposure. On day 14 of the trial, iButton temperature data loggers were subcutaneously inserted in the lateral neck region to record hourly body temperature for 42 d. Data were analyzed using PROC MIXED of SAS with repeated measures. No differences were observed in DMI, BW, ADG, F:G, or BCS during the treatment period (P > 0.05). Hair shedding scores, rectal temperatures, surface temperatures, and respiration rates were greater in EI steers compared to EF steers regardless of supplemental CP (P < 0.05). However, subcutaneous body temperature was greater in EI-14 steers (37.94 °C) compared to other steer groups (37.60, 37.68, 37.72 ± 0.04 °C for EF-14, EF-18, and EI-18, respectively; P < 0.05). Prolactin concentrations tended to be greater in EF steers when compared to EI steers (P = 0.07). Heart rate and hematocrit were reduced for EI-18 steers compared to other steer groups (P < 0.05). Caudal artery diameter was reduced in EI-18 steers compared to EI-14 steers (2.60 vs. 2.75 ± 0.05 mm, respectively; P < 0.05) and caudal vein diameter was reduced in EI-18 steers (3.20 mm) compared to all other steer groups (3.36, 3.39, 3.50 mm for EF-14, EF-18, and EI-14, respectively; P < 0.05). However, there was no difference observed in systolic or diastolic blood pressure during the treatment period (P > 0.05). Based on the data, exposure to low to moderate levels of ergot alkaloids during the stocker phase had a negative impact on hemodynamic responses and supplemental CP had minimal impact to alleviate symptoms. Therefore, feeding additional protein above established requirements is not expected to help alleviate fescue toxicosis.


Assuntos
Doenças dos Bovinos/prevenção & controle , Bovinos/fisiologia , Suplementos Nutricionais/análise , Endófitos/fisiologia , Lolium/microbiologia , Intoxicação por Plantas/veterinária , Ração Animal/análise , Animais , Peso Corporal , Bovinos/crescimento & desenvolvimento , Dieta/veterinária , Alcaloides de Claviceps/efeitos adversos , Hemodinâmica , Lolium/química , Masculino , Intoxicação por Plantas/prevenção & controle , Prolactina/sangue , Proteínas/análise , Distribuição Aleatória , Sementes/química , Sementes/microbiologia
5.
Toxins (Basel) ; 11(6)2019 06 21.
Artigo em Inglês | MEDLINE | ID: mdl-31234268

RESUMO

Fescue toxicosis is a multifaceted syndrome that elicits many negative effects on livestock consuming ergot alkaloids produced by endophyte-infected tall fescue. The economic losses associated with fescue toxicosis are primarily due to reproductive failure including altered cyclicity, suppressed hormone secretion, reduced pregnancy rates, agalactia, and reduced offspring birth weights. For decades, a multitude of research has investigated the physiological and cellular mechanisms of these reproductive failures associated with fescue toxicosis. This review will summarize the various effects of ergot alkaloids on female reproduction in grazing livestock species.


Assuntos
Alcaloides de Claviceps/toxicidade , Gado , Intoxicação por Plantas/complicações , Reprodução/efeitos dos fármacos , Animais , Encéfalo/efeitos dos fármacos , Encéfalo/fisiologia , Feminino , Festuca , Lactação , Ovário/efeitos dos fármacos , Ovário/fisiologia , Intoxicação por Plantas/veterinária , Gravidez , Útero/efeitos dos fármacos , Útero/fisiologia
6.
J Anim Sci ; 97(7): 3153-3168, 2019 Jul 02.
Artigo em Inglês | MEDLINE | ID: mdl-31051033

RESUMO

The objective of this study was to assess how exposure to ergot alkaloids during 2 stages of gestation alters fetal growth, muscle fiber formation, and miRNA expression. Pregnant ewes (n = 36; BW = 83.26 ± 8.14 kg; 4/group; 9 groups) were used in a 2 × 2 factorial arrangement with 2 tall fescue seed treatments [endophyte-infected (E+) vs. endophyte-free (E-)] fed during 2 stages of gestation (MID, days 35 to 85 vs. LATE, days 86 to 133), which created 4 possible treatments (E-/E-, E+/E-, E-/E+, or E+/E+). Ewes were individually fed a total mixed ration containing E+ or E- fescue seed according to treatment assignment. Terminal surgeries were conducted on day 133 of gestation for the collection of fetal measurements and muscle samples. Data were analyzed as a 2 × 2 factorial with fescue treatment, stage of gestation, and 2-way interaction as fixed effects. Fetuses exposed to E+ seed during LATE gestation had reduced (P = 0.0020) fetal BW by 10% compared with E- fetuses; however, fetal body weight did not differ (P = 0.41) with E+ exposure during MID gestation. Fetuses from ewes fed E+ seed during MID and LATE gestation tended to have smaller (P = 0.058) kidney weights compared with E- fetuses. Liver weight was larger (P = 0.0069) in fetuses fed E- during LATE gestation compared with E+. Fetal brain weight did not differ by fescue treatment fed during MID (P = 0.36) or LATE (P = 0.40) gestation. The percentage of brain to empty body weight (EBW) was greater (P = 0.0048) in fetuses from ewes fed E+ fescue seed during LATE gestation, which is indicative of intrauterine growth restriction (IUGR). Primary muscle fiber number was lower (P = 0.0005) in semitendinosus (STN) of fetuses exposed to E+ during MID and/or LATE gestation compared with E-/E-. miRNA sequencing showed differential expression (P < 0.010) of 6 novel miRNAs including bta-miR-652_R+1, mdo-miR-22-3p, bta-miR-1277_R-1, ppy-miR-133a_L+1_1ss5TG, hsa-miR-129-1-3p, and ssc-miR-615 in fetal STN muscle. These miRNA are associated with glucose transport, insulin signaling, intracellular ATP, hypertension, or adipogenesis. This work supports the hypothesis that E+ tall fescue seed fed during late gestation reduces fetal weight and causes asymmetrical growth, which is indicative of IUGR. Changes in primary fiber number and miRNA of STN indicate that exposure to E+ fescue fed during MID and LATE gestation alters fetal muscle development that may affect postnatal muscle growth and meat quality.


Assuntos
Endófitos/fisiologia , Alcaloides de Claviceps/toxicidade , Festuca/química , MicroRNAs/genética , Ovinos/fisiologia , Transcriptoma/efeitos dos fármacos , Animais , Encéfalo/efeitos dos fármacos , Encéfalo/crescimento & desenvolvimento , Ergotaminas/toxicidade , Feminino , Festuca/microbiologia , Desenvolvimento Fetal/efeitos dos fármacos , Peso Fetal/efeitos dos fármacos , Fibras Musculares Esqueléticas/efeitos dos fármacos , Placentação , Gravidez , Sementes/química , Sementes/microbiologia , Ovinos/crescimento & desenvolvimento
7.
Toxins (Basel) ; 11(4)2019 03 31.
Artigo em Inglês | MEDLINE | ID: mdl-30935118

RESUMO

The fate of ergot alkaloids during the milling of durum and subsequent production and cooking of pasta was examined. Durum samples containing varying amounts of ergot sclerotia (0.01⁻0.1% by mass) were milled, and all milling product was analyzed for 10 ergot alkaloids using liquid chromatography with tandem mass spectrometry. Spaghetti was prepared from the semolina obtained during milling. Ergocristine, ergocristinine, and ergotamine were the predominant ergot alkaloids observed in the milling fractions and spaghetti. Approximately 84% of the total ergot alkaloid mass of the whole grain durum resided in the milling product fractions associated with the outer kernel layers (bran, shorts, feeds). No consistent loss of ergot alkaloids was observed during the production or cooking of spaghetti. However, changes in the ratio of R- to S-enantiomers occurred during the milling and cooking of spaghetti. Products containing bran, shorts, and feeds, as well as cooked spaghetti, contained a higher proportion of the less biologically active S-enantiomers. The results of this study emphasize the need to monitor R- and S-enantiomers, and to consider food and feed products, as opposed to whole grain, when assessing any exposure of consumers to ergot alkaloids.


Assuntos
Culinária , Grão Comestível/química , Alcaloides de Claviceps/análise , Contaminação de Alimentos/análise , Manipulação de Alimentos , Triticum , Laboratórios
8.
J Anim Sci ; 97(4): 1874-1890, 2019 Apr 03.
Artigo em Inglês | MEDLINE | ID: mdl-30895321

RESUMO

Tall fescue [Lolium arundinaceum (Scheyreb.) Darbysh] is the primary cool season forage grass in the Southeastern United States. Most tall fescue contains an endophytic fungus (Epichloë coenophiala) that produces ergot alkaloids and upon ingestion induces fescue toxicosis. The objective of this study was to assess how exposure to endophyte-infected (E+; 1.77 mg hd-1 d-1 ergovaline and ergovalinine) or endophyte-free (E-; 0 mg hd-1 d-1 ergovaline and ergovalinine) tall fescue seed fed during 2 stages of gestation (MID, days 35-85/LATE, days 86-133) alters placental development. Thirty-six, fescue naïve Suffolk ewes were randomly assigned to 1 of 4 fescue treatments: E-/E-, E-/E+, E+/E-, or E+/E+. Ewes were individually fed the same amount of E+ or E- seed mixed into total mixed ration during MID and LATE gestation. Terminal surgeries were conducted on day 133 of gestation. Ewes fed E+ fescue seed had elevated (P < 0.001) ergot alkaloid excretion and reduced (P < 0.001) prolactin levels during the periods when fed E+ seed. Ewes switched on day 86 from E- to E+ seed had a 4% reduction (P = 0.005) in DMI during LATE gestation, which translated to a 2% reduction (P = 0.07) in DMI overall. Average daily gain was also reduced (P = 0.049) by 64% for E-/E+ ewes during LATE gestation and tended to be reduced (P = 0.06) by 33% overall. Ewes fed E+ seed during LATE gestation exhibited a 14% and 23% reduction in uterine (P = 0.03) and placentome (P = 0.004) weights, respectively. Caruncle weights were also reduced by 28% (P = 0.003) for E-/E+ ewes compared with E-/E- and E+/E-. Ewes fed E+ seed during both MID and LATE gestation exhibited a 32% reduction in cotyledon (P = 0.01) weights, whereas ewes fed E+ seed only during MID gestation (E+/E-) had improved (P = 0.01) cotyledon weights. The percentage of type A placentomes tended to be greater (P = 0.08) for E+/E+ ewes compared with other treatments. Other placentome types (B, C, or D) did not differ (P > 0.05). Total fetal weight per ewe was reduced (P = 0.01) for ewes fed E+ seed during LATE gestation compared with E-; however, feeding E+ seed during MID gestation did not alter (P = 0.70) total fetal weight per ewe. These results suggest that exposure to ergot alkaloids during LATE (days 86-133) gestation has the greatest impact on placental development by reducing uterine and placentome weights. This, in turn, reduced total fetal weight per ewe by 15% in ewes fed E+ seed during LATE gestation (E-/E+ and E+/E+).


Assuntos
Epichloe/química , Alcaloides de Claviceps/toxicidade , Festuca/química , Ovinos/fisiologia , Ração Animal , Animais , Endófitos , Epichloe/fisiologia , Ergotaminas/toxicidade , Feminino , Festuca/microbiologia , Placentação/efeitos dos fármacos , Gravidez , Distribuição Aleatória , Ovinos/crescimento & desenvolvimento , Sudeste dos Estados Unidos , Útero/crescimento & desenvolvimento , Útero/fisiologia
9.
Toxins (Basel) ; 11(4)2019 03 28.
Artigo em Inglês | MEDLINE | ID: mdl-30925708

RESUMO

Analysis of ergot alkaloids remains a topic of importance and the European Food Safety Authority (EFSA) has encouraged laboratories to provide monitoring data for the further evaluation of their occurrence in food and feed. While LC-MS/MS has dominated developments in recent years, LC-FLD is still more widespread, especially in developing countries. To improve the analysis of ergot alkaloids by LC-FLD, we developed an improved protocol introducing lysergic acid diethylamide (LSD) for internal standardization. Several aspects such as the composition and pH of the extraction medium, type of sorbent and conditions applied for solid-phase extraction/clean-up, use of a keeper during final evaporation and the type of syringe filter used for filtration prior to injection were thoroughly investigated. Optimized conditions comprise extraction by ethyl acetate, methanol and 28% aqueous ammonia in combination with basic aluminum oxide for extract clean-up. Use of a keeper was found inappropriate as LC-FLD analysis was significantly affected by co-eluting keeper components. Similar observations were made with some of the investigated syringe filters, where polytetrafluoroethylene (PTFE) proved to be the most suitable. Validation and application of the optimized methodology to real samples provided limits of detection and quantification suitable for the evaluation of relevant ergot alkaloid contaminations in rye and bakery products with superior precision that was facilitated by the introduced internal standard, LSD.


Assuntos
Alcaloides de Claviceps/análise , Farinha/análise , Contaminação de Alimentos/análise , Secale , Cromatografia Líquida , Fluorescência , Dietilamida do Ácido Lisérgico
10.
J Anim Sci ; 97(4): 1891-1902, 2019 Apr 03.
Artigo em Inglês | MEDLINE | ID: mdl-30763439

RESUMO

Previous research has shown that livestock exposed to ergot alkaloids results in decreased vasoactivity of gastrointestinal and peripheral vasculature. Little is known regarding the effect ergot alkaloid exposure during gestation may have on vasculature supporting the fetus. The objective of this study was to evaluate contractile responses of uterine and umbilical arteries collected from ewes consuming ergot alkaloids during gestation. On day 35 of gestation, 36 Suffolk ewes (78.24 ± 9.5 kg) were assigned to endophyte-infected (E+) or endophyte-free (E-) tall fescue seed treatments that were fed either throughout or switched on day 86 of gestation, creating four seed treatments E+E+, E+E-, E-E+, and E-E-. Ewes were fed E+ tall fescue seed to provide 1.77 mg of total ergovaline ⋅ hd-1 ⋅ d-1 with E- ewes receiving the same quantity of E- seed. Gestation was terminated on day 133, and sections of uterine artery and umbilical cord were surgically collected. Only collections from 28 ewes (n = 7/treatment) were of sufficient viability to proceed with the contractility experiments. Arteries were cleaned, sliced into 2-mm cross sections, and suspended in multi-myograph chambers containing 5 mL of continuously oxygenated Krebs-Henseleit buffer. Vessels were exposed to increasing concentrations (5 × 10-8 to 1 × 10-4 M) of norepinephrine, serotonin, ergotamine, and ergovaline (5 × 10-9 to 1 × 10-5M; extract of tall fescue seed) in 15-min intervals. Increasing concentrations of norepinephrine generated a contractile response by the uterine artery (P < 0.05), but no response in the umbilical artery. Increasing concentrations of serotonin resulted in negligible responses in uterine preparations, whereas umbilical artery preparations were responsive (P < 0.05) to serotonin. Ewes receiving E+E+ and E-E+ treatments had decreased vasoactivity in umbilical arteries to serotonin with a dextral shift in concentrations where the response curve initiated (P < 0.05). Interestingly, uterine arteries were not responsive to exposure to ergotamine or ergovaline, whereas umbilical arteries were responsive (P < 0.05). Umbilical arteries collected from ewes receiving E-E- and E+E- were more vasoactive to ergot alkaloids (P < 0.05) than other treatments. These findings indicate that maternal blood supply to the placenta appears protected from negative effects of ergot alkaloids; however, umbilical vasculature is not, and this could adversely influence fetal growth.


Assuntos
Endófitos/química , Alcaloides de Claviceps/toxicidade , Festuca/química , Contaminação de Alimentos , Ovinos/crescimento & desenvolvimento , Ração Animal/análise , Animais , Dieta/veterinária , Endófitos/fisiologia , Ergotamina/toxicidade , Ergotaminas/toxicidade , Feminino , Festuca/microbiologia , Placenta/irrigação sanguínea , Placenta/efeitos dos fármacos , Gravidez , Ovinos/fisiologia , Artérias Umbilicais/efeitos dos fármacos , Artéria Uterina/efeitos dos fármacos , Útero/irrigação sanguínea , Útero/efeitos dos fármacos
11.
Fungal Genet Biol ; 125: 71-83, 2019 04.
Artigo em Inglês | MEDLINE | ID: mdl-30731202

RESUMO

Epichloë festucae forms mutualistic symbiotic interactions with grasses of the Lolium and Festuca genera. Protection from insect and mammalian herbivory are the best-documented host benefits of these associations. The two main classes of anti-mammalian alkaloids synthesized by E. festucae are the ergot alkaloids and indole diterpenes, of which ergovaline and lolitrems are the principal terminal products. Synthesis of both metabolites require multiple gene products encoded by clusters of 11 genes located at the subtelomeric regions of chromosomes I and III respectively. These loci are essentially unexpressed in axenic culture but among the most highly expressed genes in planta. We show here that heterochromatin 1 protein (HepA) is an important component of the regulatory machinery that maintains these loci in a silent state in culture. Deletion of this gene led to derepression of eas and ltm gene expression under non-symbiotic culture conditions. Although there was no obvious culture phenotype, RNAseq analysis revealed that around 1000 genes were differentially expressed in the ΔhepA mutant compared to wild type with just one-third upregulated. Inoculation of the ΔhepA mutants into seedlings of Lolium perenne led to a severe host interaction phenotype characterized by a reduction in tiller length but an increase in tiller number. Hyphae within the leaves of these associations were much more abundant in the intercellular spaces of the leaves and aberrantly colonized the vascular bundles. This physiological change was accompanied by a dramatic change in the transcriptome with around 900 genes differentially expressed, with two thirds of these upregulated. This major physiological change was accompanied by a decrease in ltm gene expression and loss of the ability to synthesize lolitrems. These results show that HepA has an important role in controlling the chromatin state of these sub-telomeric secondary metabolite genes, including their symbiosis-specific regulation.


Assuntos
Proteínas Cromossômicas não Histona/genética , Epichloe/genética , Alcaloides de Claviceps/genética , Simbiose/genética , Diterpenos/metabolismo , Endófitos/genética , Endófitos/crescimento & desenvolvimento , Epichloe/crescimento & desenvolvimento , Epigênese Genética , Alcaloides de Claviceps/biossíntese , Proteínas Fúngicas/genética , Regulação Fúngica da Expressão Gênica/genética , Heterocromatina/genética , Hifas/genética , Hifas/crescimento & desenvolvimento , Lolium/genética , Lolium/microbiologia
12.
J Anim Sci ; 97(4): 1456-1467, 2019 Apr 03.
Artigo em Inglês | MEDLINE | ID: mdl-30772895

RESUMO

Fescue toxicosis is a multifaceted syndrome common in cattle grazing endophyte-infected tall fescue. The objective of this study was to evaluate the impact of the slick hair trait on physiological and reproductive parameters in heifers experiencing fescue toxicosis. Angus × Senepol heifers (n = 31) were blocked by weight (393.5 ± 17.3 kg) and phenotype relative to hair coat at birth, and randomly fed novel endophyte fescue (EN) or endophyte-infected fescue (EI) haylage in a total mixed ration for 91 d. Weekly measurements were collected to monitor heifer growth and response during ergot alkaloids exposure. Following 28 d of treatment, estrus was synchronized and heifers were inseminated. Ovary mapping and AI pregnancy rate were examined via transrectal ultrasonography. Blood samples were taken for genotyping: slick (S) or wildtype (W). Data were analyzed using repeated measures in PROC MIXED of SAS including fescue treatment (EN vs. EI), genotype (S vs. W), and sample collection time as main effects. Body condition scores were decreased for W heifers compared with S heifers (5.48 vs. 5.66, respectively; P < 0.0001). Surface temperature was greater for EI-W heifers (37.2 °C) compared with other groups (36.4, 36.6, 36.7 °C for EN-S, EN-W, EI-S, respectively; P < 0.05). Serum PRL concentrations were reduced for EI heifers compared with EN heifers (133.5 vs. 163.1 ng/mL, respectively; P < 0.05). The average number of 2 to 4 mm follicles were greater in EI-W heifers (13.8 follicles) compared with other groups (12.2, 10.6, and 11.1 for EN-S, EN-W, and EI-S, respectively; P < 0.0001). However, the average number of preovulatory follicles (≥9 mm) were reduced in EI-W heifers (0.52 follicles) compared with other heifer groups (0.94, 0.88, and 0.85 ± 0.04 for EN-S, EN-W, and EI-S, respectively; P < 0.05). Ovulatory follicle size was smaller in EI-W heifers compared with EN-W heifers (9.14 vs. 11.57 mm, respectively; P = 0.05). Corpus luteum area was reduced in EI-W heifers (235.1 mm2) compared with other heifer groups (297.2, 272.7, and 276.8 mm2 for EN-S, EN-W, and EI-S, respectively; P < 0.05). Concentrations of P4 were greater for EN heifers compared with EI heifers (2.7 vs. 1.8 ng/mL, respectively; P < 0.05). Pregnancy was not established in EI-W heifers (0%) compared with other heifer groups (37.5%, 57.1%, and 62.5% for EN-S, EN-W, and EI-S, respectively; P < 0.05). Overall, the slick hair mutation appears to aid in offsetting the physiological symptoms associated with fescue toxicosis and helps to improve reproductive performance.


Assuntos
Bovinos/fisiologia , Endófitos/fisiologia , Alcaloides de Claviceps/efeitos adversos , Festuca/microbiologia , Reprodução/efeitos dos fármacos , Ração Animal/análise , Animais , Peso Corporal/efeitos dos fármacos , Corpo Lúteo/fisiologia , Dieta/veterinária , Estro/efeitos dos fármacos , Feminino , Cabelo , Folículo Ovariano/fisiologia , Ovário/fisiologia , Fenótipo , Gravidez , Distribuição Aleatória
13.
J Anim Sci ; 97(2): 631-643, 2019 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-30476104

RESUMO

The goal of this study was to test the hypothesis that sodium selenite (inorganic Se, ISe), SEL-PLEX (organic forms of Se, OSe), vs. a 1:1 blend (MIX) of ISe and OSe in a basal vitamin-mineral (VM) mix would differentially alter pituitary transcriptome profiles in growing beef steers grazing an endophyte-infected tall fescue (E+) pasture. Predominately Angus steers (BW = 183 ± 34 kg) were randomly selected from fall-calving cows grazing E+ pasture and consuming VM mixes that contained 35 ppm Se as ISe, OSe, or MIX forms. Steers were weaned, depleted of Se for 98 d, and subjected to summer-long common grazing of a 10.1 ha E+ pasture containing 0.51 ppm ergot alkaloids. Steers were assigned (n = 8 per treatment) to the same Se-form treatments on which they were raised. Selenium treatments were administered by daily top-dressing 85 g of VM mix onto 0.23 kg soyhulls, using in-pasture Calan gates. As previously reported, serum prolactin was greater for MIX (52%) and OSe (59%) steers vs. ISe. Pituitaries were collected at slaughter and changes in global and selected mRNA expression patterns determined by microarray and real-time reverse transcription PCR analyses, respectively. The effects of Se treatment on relative gene expression were subjected to one-way ANOVA. The form of Se affected the expression of 542 annotated genes (P < 0.005). Integrated pathway analysis found a canonical pathway network between prolactin and pro-opiomelanocortin (POMC)/ACTH/α-melanocyte-stimulating hormone (α-MSH) synthesis-related proteins and that mitochondrial dysfunction was a top-affected canonical pathway. Targeted reverse transcription-PCR analysis found that the relative abundance of mRNA encoding prolactin and POMC/ACTH/α-MSH synthesis-related proteins was affected (P < 0.05) by the form of Se, as were (P ≤ 0.05) mitochondrial dysfunction-related proteins (CYB5A, FURIN, GPX4, and PSENEN). OSe steers appeared to have a greater prolactin synthesis capacity (more PRL mRNA) vs. ISe steers through decreased dopamine type two receptor signaling (more DRD2 mRNA), whereas MIX steers had a greater prolactin synthesis capacity (more PRL mRNA) and release potential by increasing thyrotropin-releasing hormone concentrations (less TRH receptor mRNA) than ISe steers. OSe steers also had a greater ACTH and α-MSH synthesis potential (more POMC, PCSK2, CPE, and PAM mRNA) than ISe steers. We conclude that form of Se in VM mixes altered expression of genes responsible for prolactin and POMC/ACTH/α-MSH synthesis, and mitochondrial function, in pituitaries of growing beef steers subjected to summer-long grazing an E+ pasture.


Assuntos
Bovinos/genética , Endófitos/fisiologia , Alcaloides de Claviceps/análise , Festuca/química , Selênio/farmacologia , Vitaminas/farmacologia , Hormônio Adrenocorticotrópico/biossíntese , Hormônio Adrenocorticotrópico/genética , Ração Animal/análise , Animais , Bovinos/fisiologia , Festuca/microbiologia , Masculino , Minerais/farmacologia , Mitocôndrias/metabolismo , Hipófise/metabolismo , Prolactina/biossíntese , Prolactina/genética , RNA Mensageiro/metabolismo , Estações do Ano , Selenito de Sódio/farmacologia , Transcriptoma , alfa-MSH/biossíntese , alfa-MSH/genética
14.
J Anim Sci ; 96(11): 4912-4922, 2018 Nov 21.
Artigo em Inglês | MEDLINE | ID: mdl-30476153

RESUMO

Ergot alkaloids from endophyte-infected (Epichloë coenophiala) tall fescue (Lolium arundinaceum) induce vasoconstriction. Previous work has shown that serotonin receptor subtype, 5HT2A, is present in bovine ruminal (R) and mesenteric (M) vasculature, plays a role in vasoconstriction, and could be influenced by ergot alkaloids. To determine the influence of ergot alkaloids on 5HT2A, the vasoactivity of an agonist selective for 5HT2A, (4-bromo-3,6-dimethoxybenzocyclobuten-1-yl) methylamine HCl (TCB-2), was evaluated using bovine ruminal and mesenteric arteries and veins (RA, RV, MA, MV) that were exposed to ergovaline (ERV) prior to or during the TCB-2 additions. Ruminal and mesenteric blood vessel segments were collected, cleaned, and cut into 2- to 3-mm cross-sections. Vessel segments were incubated in Krebs-Henseleit buffer containing 0, 0.01 or 1 µM ERV for 2 h prior to TCB-2 dose response or exposed to ERV concentrations simultaneously during TCB-2 dose response. For the dose response portion of the study, vessels were suspended in a multimyograph containing 5 mL of continuously oxygenated Krebs-Henseleit buffer and equilibrated to 1 g tension for 90 min. Vessels were exposed to increasing concentrations of TCB-2 every 15 min and contractile response data were normalized as a percentage of the maximum contractile response induced by 120 mM KCl reference. Analysis of variance was evaluated separately for each vessel and each ERV exposure experiment using the mixed models procedure of SAS for effects of TCB-2 and ERV concentrations. All blood vessels with previous ERV exposure had significantly lower contractile responses to TCB-2 (P < 0.01). All blood vessels with simultaneous exposure to 1 µM ERV had higher (P < 0.01) contractile responses at lower concentrations of TCB-2. Simultaneous ERV addition at 1 × 10-4 M TCB-2 did not affect contractility of RV, MA, MV (P > 0.05), but decreased contractility of RA (P < 0.01). These results indicate that ergopeptine alkaloid exposure influences contractility of bovine ruminal and mesenteric blood vessels through serotonin receptor subtype 5HT2A by acting as both an agonist and antagonist. Additional work is needed to determine if ergot alkaloids like ERV simply occupy receptor binding sites competitively, or influence receptor internalization to cause the observed divergent responses.


Assuntos
Bovinos/fisiologia , Epichloe/fisiologia , Ergotaminas/metabolismo , Festuca/química , Receptor 5-HT2A de Serotonina/metabolismo , Vasoconstrição/efeitos dos fármacos , Animais , Endófitos/química , Alcaloides de Claviceps/metabolismo , Festuca/microbiologia , Masculino , Artérias Mesentéricas/efeitos dos fármacos
15.
Mycotoxin Res ; 34(4): 297-305, 2018 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-30117108

RESUMO

The Ascomycete fungus Claviceps gigantea infects maize kernels and synthetizes several alkaloids, mostly dihydrolysergamides. There is limited information on the damage these toxins cause in mammals, despite reports from infested areas with 90% presence of the fungus sclerotia. With this background, it was decided to determine the biological activity of chemical compounds present in sclerotia of C. gigantea in rabbits 38 days after weaning. Sclerotia of C. gigantea were collected in fields with high incidence of the disease, ground and analysed for nutrients. Experimental diets were prepared with four treatments, where sclerotial powder was added, substituting for alfalfa flour in increasing proportions [C. gigantea/alfalfa flour (0:100, 5:95, 15:85 and 25:75)]. Total ergot alkaloid content was analysed by high-performance liquid chromatography. Male juvenile rabbits were utilised and distributed in completely randomised design with four replications. Initial weight was recorded in each animal, and experimental diet was offered. In this study, weight of animals, feed consumption and feed conversion were evaluated in individual animals. Blood samples were taken for haemograms, and finally euthanasia was practiced. The consumption of C. gigantea had a negative effect on body weight and feed consumption. The necropsies showed anomalies proportional to the consumption of feed contaminated with the fungus.


Assuntos
Ração Animal/microbiologia , Claviceps/química , Dieta/métodos , Alcaloides de Claviceps/toxicidade , Contaminação de Alimentos , Envenenamento/patologia , Animais , Peso Corporal , Cromatografia Líquida de Alta Pressão , Modelos Animais de Doenças , Alcaloides de Claviceps/análise , Masculino , Coelhos , Desmame
16.
Mycotoxin Res ; 34(4): 279-287, 2018 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-30117109

RESUMO

Ergot alkaloids are toxins produced by some species of fungi in the genus Claviceps, that may infect rye and triticale and, in a minor degree, other types of cereals. In this study, a new UHPLC-FLD method for the quantification of the six major ergot alkaloids as well as their corresponding epimers was developed. The sample preparation was done by a solid-liquid extraction with acetonitrile and clean-up via freeze-out. The method was fully validated and then applied to 39 samples (wheat, rye, triticale, and barley) harvested in Luxembourg in 2016. Samples were sieved (1.9 × 20 mm) prior to analysis in order to remove sclerotia, hosting the alkaloids. However, 23 samples still contained at least one ergot alkaloid > LOQ and concentrations of the sum of the 6 ergot alkaloids ranged from 0.3 to 2530.1 µg/kg. Interestingly, the highest concentrations were measured in wheat and not in rye or triticale, suggesting that all kinds of cereals should be included in monitoring programs. The outcome of this study allowed giving a first overview of ergot alkaloid concentrations in cereals harvested in Luxembourg, and the measured concentrations were in similar ranges than in other parts of the world (e.g., Canada, France, Germany).


Assuntos
Cromatografia Líquida de Alta Pressão/métodos , Grão Comestível/química , Alcaloides de Claviceps/análise , Alcaloides de Claviceps/isolamento & purificação , Análise de Alimentos/métodos , Contaminação de Alimentos/análise , Espectrometria de Fluorescência/métodos , Luxemburgo
17.
J Anim Sci ; 96(11): 4812-4822, 2018 Nov 21.
Artigo em Inglês | MEDLINE | ID: mdl-30102353

RESUMO

Fescue toxicosis is a syndrome that impairs growth and reproduction in cattle grazing endophyte-infected tall fescue [Lolium arundinaceum [(Schreb.].) Darbysh)] in the United States, resulting in approximately $1 billion in annual economic loss in species that utilize this forage resource. Approximately 90% of tall fescue contains an endophytic fungus (Epichloë coenophiala) that produces ergot alkaloids. Ergot alkaloids cause vasoconstriction and reduced blood flow to the extremities; however, it remains unknown how blood flow to the reproductive organs is affected in cattle. Therefore, the objective of this study was to determine if ergot alkaloids from endophyte-infected tall fescue reduce blood flow to the reproductive organs, thus hindering reproductive function. Angus heifers (n = 36) naïve to ergot alkaloids were placed in Calan gates and randomly assigned to receive either endophyte-infected fescue seed (E+) or noninfected fescue seed (E-; control) in a total mixed ration for 63 d. Weekly measurements were taken to monitor heifer growth and response to ergot alkaloid exposure. Reproductive measurements, including ovarian structures, uterine and ovarian vessel diameter, and hormone concentrations were determined after heifers were synchronized using the standard CO-Synch + 7 d CIDR protocol to ensure all measurements were collected at the same stages of the estrous cycle (0, 4, 10, and 17 d). Data were analyzed using repeated measures in PROC MIXED of SAS. Average daily gain was decreased for the E+ group (0.8 kg/d) compared to control heifers (1.0 kg/d). Body condition scores tended to be greater in control heifers compared to the E+ group (P = 0.053). Additionally, hair coat and hair shedding scores were greater in E+ heifers compared to controls (P < 0.05). Heart rate, rectal temperature, respiration rate, and blood pressure did not differ between treatments (P > 0.05). Vasoconstriction was observed in the caudal artery, but not the caudal vein, in heifers consuming the E+ fescue seed (P < 0.05). No differences were observed in antral follicle counts, corpus luteum area or circulating progesterone concentrations in E+ heifers compared to controls (P > 0.05). There was a significant decrease in the diameter of arteries and veins servicing the ovary and uterus on day 10 and 17 of the estrous cycle. Reduction in blood flow to the reproductive organs during critical times in the estrous cycle may contribute to the reduced ovarian function and pregnancy rates associated with fescue toxicosis.


Assuntos
Bovinos/fisiologia , Epichloe/fisiologia , Alcaloides de Claviceps/farmacologia , Festuca/química , Reprodução/efeitos dos fármacos , Vasoconstrição/efeitos dos fármacos , Animais , Endófitos , Alcaloides de Claviceps/química , Ergotaminas , Ciclo Estral/efeitos dos fármacos , Feminino , Festuca/microbiologia , Frequência Cardíaca/efeitos dos fármacos , Hemodinâmica/efeitos dos fármacos , Ovário/irrigação sanguínea , Gravidez , Distribuição Aleatória , Útero/irrigação sanguínea
18.
Toxins (Basel) ; 10(8)2018 08 01.
Artigo em Inglês | MEDLINE | ID: mdl-30071666

RESUMO

As contamination with cereal ergot has been increasing in western Canada, this study evaluated impacts of feeding a mycotoxin binder (Biomin® II; BB) on nutrient digestibility, alkaloid recovery in feces, and lamb growth performance. Forty-eight ram lambs (25.9 ± 1.4 kg) were randomly assigned to one of four barley-based diets: Control (C), no added alkaloids, Control + BB fed at 30 g/head per day (CBB); Ergot, 2564 ppb total R + S epimers (E); Ergot + BB, 2534 ppb R + S epimers (EBB). Lambs were fed ab libitum for up to 11 weeks until slaughter at >46 kg live weight. Both average daily gain (ADG) and gain/feed ratio were greater (p < 0.01) for lambs fed C and CBB diets as compared with those containing added ergot, although dry matter intake was not affected by dietary ergot or BB. Serum prolactin concentrations were two times higher in EBB- compared with E-fed lambs (p < 0.05), although both were lower than in C or CBB (p < 0.001) lambs. Rectal temperatures were greater in lambs receiving dietary ergot (p ≤ 0.001) than in C- and CBB-fed lambs. In a digestibility study using eight ram lambs, treatment with BB increased neutral detergent fiber (NDF) digestibility (p = 0.01). Nitrogen retention (g) was greater (p < 0.05) for lambs receiving C or CBB compared with ergot-contaminated diets. Feces of EBB lambs had 38.5% greater (p < 0.001) recovery of alkaloids compared with those fed E. Based on sparing of prolactin, BB may reduce impacts of ergot alkaloids by increasing their excretion in feces. Accordingly, concentrations of dietary alkaloids, which would not harm sheep, would be increased by feeding BB.


Assuntos
Digestão/efeitos dos fármacos , Alcaloides de Claviceps/farmacocinética , Fezes/química , Micotoxinas/farmacocinética , Substâncias Protetoras/administração & dosagem , Ração Animal , Animais , Dieta/veterinária , Grão Comestível , Alcaloides de Claviceps/química , Contaminação de Alimentos , Hordeum , Masculino , Micotoxinas/química , Nutrientes/administração & dosagem , Prolactina/sangue , Substâncias Protetoras/química , Ovinos
19.
Appl Environ Microbiol ; 84(19)2018 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-30076193

RESUMO

Ergot alkaloids are specialized fungal metabolites with potent biological activities. They are encoded by well-characterized gene clusters in the genomes of producing fungi. Penicillium camemberti plays a major role in the ripening of Brie and Camembert cheeses. The P. camemberti genome contains a cluster of five genes shown in other fungi to be required for synthesis of the important ergot alkaloid intermediate chanoclavine-I aldehyde and two additional genes (easH and easQ) that may control modification of chanoclavine-I aldehyde into other ergot alkaloids. We analyzed samples of Brie and Camembert cheeses, as well as cultures of P. camemberti, and did not detect chanoclavine-I aldehyde or its derivatives. To create a functioning facsimile of the P. camembertieas cluster, we expressed P. camemberti easH and easQ in a chanoclavine-I aldehyde-accumulating easA knockout mutant of Neosartorya fumigata The easH-easQ-engineered N. fumigata strain accumulated a pair of compounds of m/z 269.1288 in positive-mode liquid chromatography-mass spectrometry (LC-MS). The analytes fragmented in a manner typical of the stereoisomeric ergot alkaloids rugulovasine A and B, and the related rugulovasine producer Penicillium biforme accumulated the same isomeric pair of analytes. The P. camemberti eas genes were transcribed in culture, but comparison of the P. camemberti eas cluster with the functional cluster from P. biforme indicated 11 polymorphisms. Whereas other P. camembertieas genes functioned when expressed in N. fumigata, P. camembertieasC did not restore ergot alkaloids when expressed in an easC mutant. The data indicate that P. camemberti formerly had the capacity to produce the ergot alkaloids rugulovasine A and B.IMPORTANCE The presence of ergot alkaloid synthesis genes in the genome of Penicillium camemberti is significant, because the fungus is widely consumed in Brie and Camembert cheeses. Our results show that, although the fungus has several functional genes from the ergot alkaloid pathway, it produces only an early pathway intermediate in culture and does not produce ergot alkaloids in cheese. Penicillium biforme, a close relative of P. camemberti, contains a similar but fully functional set of ergot alkaloid synthesis genes and produces ergot alkaloids chanoclavine-I, chanoclavine-I aldehyde, and rugulovasine A and B. Our reconstruction of the P. camemberti pathway in the model fungus Neosartorya fumigata indicated that P. camemberti formerly had the capacity to produce these same ergot alkaloids. Neither P. camemberti nor P. biforme produced ergot alkaloids in cheese, indicating that nutritionally driven gene regulation prevents these fungi from producing ergot alkaloids in a dairy environment.


Assuntos
Alcaloides de Claviceps/biossíntese , Penicillium/metabolismo , Cromatografia Líquida de Alta Pressão , Ergolinas/metabolismo , Proteínas Fúngicas/genética , Proteínas Fúngicas/metabolismo , Genoma Fúngico , Espectrometria de Massas , Penicillium/genética
20.
Nat Prod Rep ; 35(12): 1347-1382, 2018 07 19.
Artigo em Inglês | MEDLINE | ID: mdl-30024006

RESUMO

Covering: 1969 to 2018 Azepinoindole natural products can be broadly classified as being of monoterpenoid or non-monoterpenoid origin. The non-monoterpenoid azepinoindoles have not received as much attention in the literature as their more revered monoterpenoid counterparts. In this review, an overview of all non-monoterpenoid azepinoindoles is provided. Various biological and chemical aspects are discussed, including their isolation, biosynthesis and the elegant total synthesis studies that have been inspired by these alkaloids.


Assuntos
Alcaloides Indólicos/química , Alcaloides Indólicos/metabolismo , Antibacterianos/química , Antibacterianos/farmacologia , Benzazepinas/química , Benzazepinas/isolamento & purificação , Benzodioxóis/isolamento & purificação , Técnicas de Química Sintética , Alcaloides de Claviceps/síntese química , Alcaloides Indólicos/isolamento & purificação , Alcaloides Indólicos/farmacologia , Lignanas/biossíntese , Lignanas/isolamento & purificação , Estrutura Molecular , Monoterpenos , Quinazolinas/síntese química , Quinazolinas/química , Quinazolinas/isolamento & purificação
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