Distribution and Accumulation of the Alkaloid Peramine in Neotyphodium lolii-Infected Perennial Ryegrass

Through the production of alkaloids and mycotoxins, the endophytic fungus of perennial ryegrass, Neotyphodium lolii, confers resistance to the host grass against several insect pasture pests and causes health problems in livestock. The effects of plant component, plant reproductive development, and plant management (trimming) on the concentration and accumulation of peramine, the alkaloid thought to be most important for conferring resistance against Argentine stem weevil, were investigated. Clones of eight different naturally occurring perennial ryegrass-N. lolii associations were used in the experiment, which was conducted during spring–summer, which corresponded with the height of plant reproductive activity. Clones were either trimmed regularly to maintain them in a vegetative state (V clones) or not trimmed, allowing them to become fully reproductive (R clones). V and R clones of each grass/endophyte association were destructively harvested at times that corresponded roughly with the start, middle, and end of the plant reproductive phase. Harvested material was dissected into a number of plant components that were analyzed for peramine concentration by HPLC. All clippings from the V clones were also analyzed. Peramine concentrations were usually similar in leaf sheaths and leaf blades, but they decreased as leaf age increased. The seed from R clones and younger sheaths and blades of leaves from vegetative tillers contained the highest concentrations, while the root, crown, and dead leaf tissue contained the lowest. During the inflorescence phase, tissues from reproductive tillers generally contained lower concentrations than corresponding tissues from vegetative tillers. Also at this time, mean peramine concentrations were higher in V clones than in R clones. By the end of the plant reproductive phase, at the mature seed stage, mean peramine concentrations in R and V clones were similar. The seed component, which contained approximately 75% of the total peramine present in the plant (at a mean concentration of 63.7 μg/g), probably accounted for much of the recovery in the R clones.     

Peramine and Lolitrem B from Endophyte-Grass Associations Cascade Up the Food Chain

Endophytic fungi in cool-season grass species produce herbivore-toxic alkaloids, which are assumed to harm higher trophic levels along food chains. Previous studies have shown fitness disadvantages for higher trophic levels that feed on aphids that were exclusively reared on perennial ryegrass (Lolium perenne) infected with the endophytic fungus Neotyphodium lolii. However, it is unknown whether the alkaloids produced by the fungus-grass association can be assimilated by plant sap-sucking insects like aphids. Using an ultra high performance liquid chromatography method combined with mass spectrometry, we provide the first evidence that the alkaloids peramine and lolitrem B are present in aphids (Rhopalosiphum padi) and in aphid predators when the aphids are reared on endophyte-infected grass. We conclude that alkaloids can enter the plant sap of the grass and are responsible for longer pupal stages of the ladybird Harmonia axyridis and for fitness disadvantages of aphids and their predators as shown in previous studies.     

Temporal and Spatial Variation in Alkaloid Levels in Achnatherum robustum, a Native Grass Infected with the Endophyte Neotyphodium

The native North American perennial grass Achnatherum robustum (Vasey) Barkworth [= Stipa robusta (Vasey) Scribn.] or sleepygrass is toxic and narcotic to livestock. The causative agents are alkaloidal mycotoxins produced from infections by a systemic and asexual Neotyphodium endophyte. Recent studies suggest that toxicity is limited across the range of sleepygrass in the Southwest USA. We sampled 17 populations of sleepygrass with varying distance from one focal population known for its high toxicity levels near Cloudcroft, NM, USA. For some, we sampled individual plants twice within the same growing season and over successive years (2001–2004). We also determined infection levels in each population. In general, all populations were highly infected, but infection levels were more variable near the focal population. Only infected plants within populations near the Cloudcroft area produced alkaloids. The ergot alkaloid, ergonovine, comprised the bulk of the alkaloids, with lesser amounts of lysergic and isolysergic acid amides and ergonovinine alkaloids. Levels of all alkaloids were positively correlated among individual plants within and between growing seasons. Infected plants that produced no alkaloids in 1 yr did not produce any alkaloids within the same growing season or in other years. Levels of alkaloids in sleepygrass populations declined with distance from the Cloudcroft population, although infection levels increased. Infected plants in populations in northern New Mexico and southern Colorado produced no alkaloids at all despite 100% infectivity. Our results suggest that only specific Neotyphodium haplotypes or specific Neotyphodium–grass combinations produce ergot alkaloids in sleepygrass. The Neotyphodium haplotype or host–endophyte combination that produces toxic levels of alkaloids appears restricted to one locality across the range of sleepygrass. Because of the wide variation in alkaloid levels among populations, interactions between the endophyte and host, and consequences for herbivores, competitors, and pathogens and other components of the community, are likely to vary widely across the geographic range of this native grass.     

Fractional Separation and Structural Characterization of Chlorophyll and Lignin from Perennial Ryegrass (L. perenne) and Cocksfoot Grass (D. glomerata)

Abstract

One chlorophyll rich fraction and two lignin preparations were separated from perennial grass and cocksfoot grass by sequential three‐stage treatments with 80% ethanol containing 0.2% NaOH, 2.5% H₂O₂?0.2% EDTA containing 1.5% NaOH, and 2.5% H₂O₂?0.2% TAED containing 1.0% NaOH at 75°C for 3 h, respectively, which released 4.6 and 3.6% chlorophyll rich fraction, 2.3 and 5.8%, and 0.9 and 1.0% lignin preparations, except for releasing 8.0 and 10.4%, 79.1 and 77.0%, and 12.9 and 12.5% of the original hemicelluloses, respectively. The lignin fractions obtained from the two different grasses had very similar molecular weights and structural composition. The NMR spectra of the lignin preparations revealed the presence of p‐hydroxyphenyl, guaiacyl, and syringyl structures, and the lignin in chlorophyll rich fraction contained more guaiacyl and syringyl units than p‐hydroxyphenyl unit, whereas the reverse trend was found in the two lignin preparations. The lignin preparations are distinguished with straw and wood lignins by relatively higher contents of p‐hydroxyphenyl unit and lower amounts of condensed units (β‐5 and 5‐5′) and resinol units (β‐β). This difference in distribution of structural units indicated some structural heterogeneity between grass and straw/or wood lignin.     

Accumulation of Hydroxyl Lipids and 4-Hydroxy-2-Hexenal in Live Fish Infected with Fish Diseases

Hydroxy lipids (L-OH) and 4-hydroxy-2-hexenal (HHE) levels as well as other parameters such as lipid level, lipid class, fatty acid composition, and other aldehydes levels in the liver of diseased fish were investigated. Although significant differences in lipid level, lipid class, fatty acid composition, and other aldehyde levels were not always observed between normal and diseased fish, L-OH and HHE levels were significantly higher in the liver of the diseased fish than in that of the normal fish cultured with the same feeds under the same conditions. In the liver of puffer fish (Fugu rubripes) infected with Trichodina, L-OH and HHE levels significantly increased from 25.29 ± 5.04 to 47.70 ± 5.27 nmol/mg lipid and from 299.79 ± 25.25 to 1,184.40 ± 60.27 nmol/g tissue, respectively. When the levels of HHE and other aldehydes in the liver of the normal and diseased puffer fish were plotted, a linear relationship with a high correlation coefficient was observed between HHE and propanal (r ² = 0.9447). Increased L-OH and HHE levels in the liver of the diseased fish and a high correlation between HHE and propanal in the liver of the normal and diseased fish were also observed in flat fish (Paralichthys olivaceus) infected with streptococcus, yellowtail (Seriola quinqueradiata) infected with jaundice, and amberjack (S. purpurascens) infected with Photobacterium damselae subsp. piscicida.     

冀中坳陷深县凹陷油气成藏条件及其分布特征

综合利用地质、地震和化验分析等资料,从烃源岩、储集层、生储盖组合、圈闭类型和输导体系5个方面对深县凹陷油气成藏条件进行了系统分析。研究表明,深县凹陷主要发育2套烃源岩,分布在深西—白宋庄和虎北洼槽;储集层主要是新生界碎屑岩储层和古生界基岩储层,发育三种生储盖层组合模式;圈闭类型以构造和潜山圈闭为主,圈闭定型时间早于本区油气生成和运移时间,油气成藏条件较好;输导系统以断裂的垂向疏导为主。受生烃洼槽、二级构造带和断裂分布控制,油气平面分布呈3个条带:西北部何庄-深西潜山油藏聚集带,南部深南背斜油气聚集带,东部榆科背斜油气聚集带。纵向发育6个含油组合,平面上不同部位的含油组合的叠置差异较大。     

黑麦草对土壤中抗生素的降解及其机理研究

抗生素作为一种高效治疗和防治疾病的药物,已经被广泛运用到我国的畜禽养殖行业中.但是,抗生素的滥用现象十分普遍,土壤中残留的高浓度抗生素会对土壤微生物的活性以及生物群落的构成产生很大影响.围绕黑麦草对土壤中残留抗生素的吸收降解进行了一系列的机理研究,结果表明,黑麦草的种植对于土壤中典型抗生素的降解有明显效果,其中对四环素中金霉素的降解率最高,为38％;磺胺类抗生素中磺胺二甲基嘧啶的降解率最低,为19.2％;喹诺酮类各抗生素的降解率在22.3％至35％之间,其中诺氟沙星的降解率最高,洛美沙星最低.同时,黑麦草的种植还使土壤微生物抑制得到了缓解,种植黑麦草的土壤中微生物的呼吸作用强度和氨化作用强度是未种植黑麦草土壤微生物的1.4倍和3.2倍.根系的各方面指标在前期受残留抗生素影响,前期(10～25 d)增长速度比较慢,中期(25～30 d)增长速度快,后期(30～40 d)趋于平稳,这可能与黑麦草的生长周期有关.     

The Use of Isolated Human Lymphocytes in Mycotoxin Cytotoxicity Testing

The cytotoxicity of selected mycotoxins against isolated human lymphocytes was investigated, as a means of detecting mycotoxins in extracts derived from cereal samples. The methodology was based on the ability of viable cells to reduce methyl tetrazolium bromide to a purple formazan dye that could be quantitated by spectrophometric means and hence give a measure of the cytotoxicity of added substances. The results showed that there was good correlation with the occurrence of identified mycotoxins with only a minimum of false positives. For example, of the 13 samples of barley or barley derivatives that were positive for the mycotoxins, fumonisin B₁ (FB1) deoxynivalenol (DON) and ochratoxin A (OTA), all gave positive cytotoxicity responses. Two samples negative for mycotoxins gave no cytotoxicity responses. There was little variation between the results for lymphocytes drawn from the same healthy volunteer on three different occasions. Furthermore, for two of the mycotoxins tested (FB1 and DON) it was possible to correlate general levels of mycotoxins present to the cytotoxic response of the lymphocytes but not for OTA, where it was concluded that interfering substances prevented direct correlation. It was concluded that this method was suited for general application as it could handle relatively high number of samples in a short period of time.     

Neotyphodium coenophialum Mycelial Protein and Herbage Mass Effects on Ergot Alkaloid Concentration in Tall Fescue

Livestock grazing endophyte-infected tall fescue (Festuca arundinacea Schreb.) pastures often suffer from ergot poisoning. The endophyte,Neotyphodium coenophialum (Morgan-Jones and Gams) Glenn, Bacon, and Hanlin, comb. nov., also provides drought-tolerant, insect-resistant, and disease-resistant qualities to the plant. Therefore, producers are faced with a biological dilemma of risking loss of pasture by using endophyte-free tall fescue pasture or animal losses with endophyte-infected tall fescue pasture. One potential solution is to breed endophyte-infected tall fescue with lower levels of alkaloids. However, breeding could select for plants that are antagonistic to the endophyte, resulting in reduced plant vigor as a consequence of disruption of the mutualistic association between the organisms. The objectives of this study were to determine the relationship between plant mass and endophyte mycelial proteins versus ergot alkaloid concentration. To examine the endophyte effect on mycelial mass, tissue culture regenerants from tall fescue genotype PDN2 were infected with endophyte isolates EDN11, EDN12, and EDN2 to eliminate confounding effects of multiple plant genotypes. Crosses with PDN11 as the maternal parent and plant genotypes DN2, DN12, and DN15 as paternal parents were used to produce a population of different plant genotypes, all containing the same endophyte. Fungal mycelial protein was extracted from lyophilized tall fescue leaf sheath tissue and immunochemically quantified with monoclonal antibodies specific toN. coenophialum proteins. Ergot alkaloid concentration was also immunochemically measured. Quantities of mycelial protein and ergot alkaloids were calculated by regressing experimental values against standards of each. There was no correlation between herbage mass and alkaloid concentration or fungal protein quantity and alkaloid concentration when different fungal isolates were inserted into the same plant genotype. Coefficients of determination (r ²) were low (0.31 and 0.17) between leaf sheath and leaf blade alkaloid concentrations and endophyte protein when different plant genotypes contained the same endophyte isolate. Likewise,r ² values were low between leaf sheath and leaf blade alkaloid concentrations and herbage mass. These data suggest that little or no antagonism occurred to the endophyte among plants low in alkaloid concentration.     

安塞油田杏河区长6油藏分布规律

安塞油田的油气分布规律一直受到研究者的关注。前人认为该地区油气分布主要受控于沉积相,区内发育的鼻状隆起对油气分布是否有控制作用也存在争议。随着勘探开发程度的提高,这些问题有了一些新的看法。以杏河区为例,通过对沉积相、储层物性、构造及其与油藏分布关系的研究发现,不仅沉积相是其油气分布的控制因素,而且储层物性的差异性也是形成圈闭遮挡的重要条件,该区长6发育的鼻状隆起对长61、长62、长63油藏油气的分布有不同程度的控制作用,对长61油藏控制程度弱,而对长62、长63有重要的控制作用。     

Perennial Electron Mobility in Quantum Transition State and Molecular Processes of Oxygenation

The controlled conditions have established the molecular features in various types of oxygenation. The molecular induction period as molecular properties, is clear from the molecule to molecule processes under influence of time and temperature effects graphically represented, emphasizing quantum transition state (unitized). The perennial electron mobility (resonance, i.e. regulated, or random) in molecules at close approach, has eliminated confusions arising out of existing MO theories and made possible the new theories for generation of electron energy and its transfer towards molecular activation and molecular Khan-London forces in elucidation for the behaviour of quantum giant molecule in variety of molecular physical states, or for that of quantum pseudo-molecules, i. e., quantum transition state.     

Growth and Mycotoxin Production by Chaetomium globosum Is Favored in a Neutral pH

Chaetomium globosum is frequently isolated in water-damaged buildings and produces two mycotoxins called chaetoglobosins A and C when cultured on building material. In this study, the influence of ambient pH on the growth of C. globosum was examined on an artificial medium. This fungus was capable of growth on potato dextrose agar ranging in pH from 4.3 to 9.4 with optimal growth and chaetoglobosin C production occurring at a neutral pH. In addition, our results show that sporulation is favored in an acidic environment.     

Biogenic Separation,Accumulation and Cellular Distribution of Cu,Co, and Ni in Medicago sativa under Idealized Conditions

The limits of uptake of Co, Ni, and Cu by the common metallophyte, Medicago sativa, were assessed using hydroponic growth and metal uptake experiments. The influence of the growth substrate metal concentration (500 and 1000 ppm) and exposure time, i.e., the time plants were exposed to the metal solution (24, 48, or 72 h) was investigated. The combined roots and shoots of Medicago sativa accumulated up to 2.2 wt-% Co, 2.0 wt-% Ni, and 3.5 wt-% Cu, when exposed to aqueous solutions containing 1000 ppm Co for 48 h, 1000 ppm Ni for 72 h, and 1000 ppm Cu for 72 h, respectively. The distribution of the sequestered metals was assessed using proton induced that X-ray emission spectroscopy (μ-PIXE), which indicated that translocation mechanism was most likely xylem loading. However, the rate of translocation of the metal from the roots to the plant stem was different for each metal, suggesting differing mechanisms for each. Collectively, these results suggest the separation and removal of the heavy metals Cu, Co, and Ni from contaminated substrates using Medicago sativa is a viable technology.     

Chloroquine-Induced Accumulation of Autophagosomes and Lipids in the Endothelium

Chloroquine (CQ) is an antimalarial drug known to inhibit autophagy flux by impairing autophagosome–lysosome fusion. We hypothesized that autophagy flux altered by CQ has a considerable influence on the lipid composition of endothelial cells. Thus, we investigated endothelial responses induced by CQ on human microvascular endothelial cells (HMEC-1). HMEC-1 cells after CQ exposure were measured using a combined methodology based on label-free Raman and fluorescence imaging. Raman spectroscopy was applied to characterize subtle chemical changes in lipid contents and their distribution in the cells, while the fluorescence staining (LipidTox, LysoTracker and LC3) was used as a reference method. The results showed that CQ was not toxic to endothelial cells and did not result in the endothelial inflammation at concentrations of 1–30 µM. Notwithstanding, it yielded an increased intensity of LipidTox, LysoTracker, and LC3 staining, suggesting changes in the content of neutral lipids, lysosomotropism, and autophagy inhibition, respectively. The CQ-induced endothelial response was associated with lipid accumulation and was characterized by Raman spectroscopy. CQ-induced autophagosome accumulation in the endothelium is featured by a pronounced alteration in the lipid profile, but not in the endothelial inflammation. Raman-based assessment of CQ-induced biochemical changes offers a better understanding of the autophagy mechanism in the endothelial cells.     

Identification and Functional Characterization of Acyl-ACP Thioesterases B (GhFatBs) Responsible for Palmitic Acid Accumulation in Cotton Seeds

In spite of increasing use in the food industry, high relative levels of palmitic acid (C16:0) in cottonseed oil imposes harmful effects on human health when overconsumed in the diet. The limited understanding of the mechanism in controlling fatty acid composition has become a significant obstacle for breeding novel cotton varieties with high-quality oil. Fatty acyl–acyl carrier protein (ACP) thioesterase B (FatBs) are a group of enzymes which prefer to hydrolyze the thioester bond from saturated acyl-ACPs, thus playing key roles in controlling the accumulation of saturated fatty acids. However, FatB members and their roles in cotton are largely unknown. In this study, a genome-wide characterization of FatB members was performed in allotetraploid upland cotton, aiming to explore the GhFatBs responsible for high accumulations of C16:0 in cotton seeds. A total of 14 GhFatB genes with uneven distribution on chromosomes were identified from an upland cotton genome and grouped into seven subfamilies through phylogenetic analysis. The six key amino acid residues (Ala, Trys, Ile, Met, Arg and Try) responsible for substrate preference were identified in the N-terminal acyl binding pocket of GhFatBs. RNA-seq and qRT-PCR analysis revealed that the expression profiles of GhFatB genes varied in multiple cotton tissues, with eight GhFatBs (GhA/D-FatB3, GhA/D-FatB4, GhA/D-FatB5, and GhA/D-FatB7) having high expression levels in developing seeds. In particular, expression patterns of GhA-FatB3 and GhD-FatB4 were positively correlated with the dynamic accumulation of C16:0 during cotton seed development. Furthermore, heterologous overexpression assay of either GhA-FatB3 or GhD-FatB4 demonstrated that these two GhFatBs had a high substrate preference to 16:0-ACP, thus contributing greatly to the enrichment of palmitic acid in the tested tissues. Taken together, these findings increase our understanding on fatty acid accumulation and regulation mechanisms in plant seeds. GhFatBs, especially GhA-FatB3 and GhD-FatB4, could be molecular targets for genetic modification to reduce palmitic acid content or to optimize fatty acid profiles in cotton and other oil crops required for the sustainable production of healthy edible oil.     

Sphingosine-1-Phosphate Receptor Type 4 (S1P4) Is Differentially Regulated in Peritoneal B1 B Cells upon TLR4 Stimulation and Facilitates the Egress of Peritoneal B1a B Cells and Subsequent Accumulation of Splenic IRA B Cells under Inflammatory Conditions

Background: Gram-negative infections of the peritoneal cavity result in profound modifications of peritoneal B cell populations and induce the migration of peritoneal B cells to distant secondary lymphoid organs. However, mechanisms controlling the egress of peritoneal B cells from the peritoneal cavity and their subsequent trafficking remain incompletely understood. Sphingosine-1-phosphate (S1P)-mediated signaling controls migratory processes in numerous immune cells. The present work investigates the role of S1P-mediated signaling in peritoneal B cell trafficking under inflammatory conditions. Methods: Differential S1P receptor expression after peritoneal B cell activation was assessed semi‑quantitatively using RT-PCR in vitro. The functional implications of differential S1P₁ and S1P₄ expression were assessed by transwell migration in vitro, by adoptive peritoneal B cell transfer in a model of sterile lipopolysaccharide (LPS)‑induced peritonitis and in the polymicrobial colon ascendens stent peritonitis (CASP) model. Results: The two sphingosine-1-phosphate receptors (S1PRs) expressed in peritoneal B cell subsets S1P₁ and S1P₄ are differentially regulated upon stimulation with the TLR4 agonist LPS, but not upon PMA/ionomycin or B cell receptor (BCR) crosslinking. S1P₄ deficiency affects both the trafficking of activated peritoneal B cells to secondary lymphoid organs and the positioning of these cells within the functional compartments of the targeted organ. S1P₄ deficiency in LPS-activated peritoneal B cells results in significantly reduced numbers of splenic innate response activator B cells. Conclusions: The S1P-S1PR system is implicated in the trafficking of LPS-activated peritoneal B cells. Given the protective role of peritoneal B1a B cells in peritoneal sepsis, further experiments to investigate the impact of S1P₄-mediated signaling on the severity and mortality of peritoneal sepsis are warranted.     

给水管网中含铝水垢对微量金属元素蓄积释放特性综述

该文探讨了含铝沉积物对微量金属元素的蓄积作用,以及原水水质、水处理药剂、溶液pH和水流状态等因素改变时对含铝沉积物结构稳定性和微量金属元素释放特性的影响。为减少输配水过程中微量金属元素的释放,保障管网稳定运行和居民饮水健康,建议通过水质调节控制管网中含铝沉积物的生成量。该研究给出了常见微量金属元素的最大允许蓄积量,并指出了今后的研究方向。     

海拉尔盆地乌尔逊凹陷油气成藏期次及成藏过程分析

研究表明乌尔逊凹陷油气表现为三期成藏：距今约125-120Ma的伊敏组早期、距今约120-90Ma的伊敏组末期以及距今约89-85Ma的青元岗早期。其中伊敏组末期应为凹陷的主要成藏期。不同成藏期次油气成熟度不同,早期油气成熟度较低,以区域不整合为主要运移通道,运移距离较远,可在远离烃源灶的圈闭中聚集成藏;中后期由于凹陷断裂活动,油气横向运移距离受到限制,以垂向运移为主,同时,不整合与断裂裂系相互配合也可使油气呈“阶梯状运移”,此阶段油气主要在烃源灶附近的圈闭中聚集。