Production and initial characterisation of the xylan-degrading system of Phanerochaete chrysosporium

We report the optimum conditions for the degradation of oat spelt arabinoxylan and a preliminary characterisation of the inducible xylan-degrading system of the lignin-degrading white-rot fungus Phanerochaete chrysosporium. Xylanase activity was optimal at pH 5.0 and 50°C; see attached sheet the maximum reaction velocity (V_max) of the system was 3.86 units (U) mg^–1 protein with arabinoxylan as substrate and the substrate concentration giving half V_max (S_0.5) was 0.52 mg ml^–1. At concentrations of arabinoxylan greater than 15 mg ml^–1 excess substrate inhibition was observed. Xylose at 0.9 mm inhibited activity to the extent of 50%. Xylanase activity increased as a function of the dilution of the enzyme preparation prior to assay. It was resolved into four peaks by using a DEAE-Biogel column; the material in these peaks differed with respect to xylan solubilisation and the formation of reducing sugars. Electrofocusing gels allowed visualisation of several bands of activity corresponding to each peak. The arabinoxylan degradation system of P. chrysosporium is therefore composed of multiple components. Correspondence to: P. Broda     

In vitro survival of murine morulae after quick freezing in the presence of chemically defined macromolecules and different cryoprotectants

We studied the ability of frozen-thawed mouse morulae to develop in vitro when the cryoprotectant proteins were substituted with one of the following nonorganic macromolecules: polyvinyl alcohol (PVA), polyvinyl pyrrolidone (PVP), and ficoll. We also determined how these agents interacted with 3 different cryoprotectants: glycerol (GLY), propylene glycol (PG), and ethylene glycol (EG). The influence of both of the above factors was measured on the basis of post-thaw morphological appearance, the percentage of development to the expanded blastocyst stage and the total cell count. Morulae (n=950) were collected from superovulated mice. Those classified as good or excellent were distributed among the 12 different freezing solutions, obtained by combining the 3 cryoprotectants with the 4 macromolecules (the 3 mentioned above, plus a control of 5% fetal calf serum) in phosphate buffered saline (PBS). Embryos frozen in PVA, PVP and ficoll tended to be a little difficult to recover from the straws. Development to the expanded blastocyst stage was significantly lower (P<0.05) in propylene glycol (43.6%) than in ethylene glycol (79.5%) or in glycerol (76.1%). Polyvinyl alcohol provided a higher survival rate when combined with glycerol (90.3) or ethylene glycol (95.0), but when it was combined with propylene glycol, only 56.5% of embryos survived after thawing. A positive interaction was observed between glycerol and PVA and between ethylene glycol and PVA or ficoll. The results indicate that fetal serum could be successfully substituted for any of the 3 chemically defined macromolecules. However, our findings also suggest that the use of PG as a cryoprotectant should be avoided when mouse morulae are frozen using the quick freezing method.     

Life-history studies of heterophyid trematodes in the Neotropical Region: Ascocotyle (Phagicola) diminuta (Stunkard & Haviland, 1924) and A. (P.) angrense Travassos, 1916

The life-cycle of Ascocotyle (Phagicola) diminuta (Stunkard & Haviland, 1924) was reproduced experimentally, starting from cercariae from naturally infected Littoridina castellanosae and L. parchappei (Hydrobiidae) collected from artificial ponds in the Zoological Garden in Buenos Aires and from Los Ranchos stream, Buenos Aires Province, respectively. Metacercariae were found encysted in the gills of experimentally exposed Cnesterodon decemmaculatus (Poecilidae) and of other naturally infected freshwater fishes. Adults were obtained experimentally in chicks and mice and from a naturally infected egret, Egretta thula. A. (P.) angrense Travassos, 1916 was found parasitising the egret Ixobrychus involucris; it is considered a valid species and the morphological differences between it and A. (P.) diminuta were established. The “Phagicola-form” of the cercaria in the present life-cycle is also known in the genus Pygidiopsis.     

Protein phosphatases in higher plants: multiplicity of type 2A phosphatases in Arabidopsis thaliana

Two DNA fragments, AP-1 and AP-2, encoding amino acid sequences closely related to Ser/Thr protein phosphatases were amplified from Arabidopsis thaliana genomic DNA. Fragment AP-1 was used to screen. A. thaliana cDNA libraries and several positive clones were isolated. Clones EP8a and EP14a were sequenced and found to encode almost identical proteins (97% identity). Both proteins are 306 amino acids in length and are very similar (79–80% identity) to the mammalian isotypes of the catalytic subunit of protein phosphatase 2A. Therefore, they have been designated PP2A-1 and PP2A-2. A third cDNA clone, EP7, was isolated and sequenced. The polypeptide encoded (308 amino acids, lacking the initial Met codon) is 80% identical with human phosphatases 2A and was named PP2A-3. The PP2A-3 protein is extremely similar (95% identity) to the predicted protein from a cDNA clone previously found in Brassica napus. Southern blot analysis of genomic DNA using AP-1 and AP-2 probes, as well as probes derived from clones EP7, EP8a and EP14a strongly indicates that at least 6 genes closely related to type 2A phosphatases are present in the genome of A. thaliana. Northern blot analysis using the same set of probes demonstrates that, at the seedling stage, the mRNA levels for PP2A-1, PP2A-3 and the gene containing the AP-1 sequence are much higher than those of PP2A-2 and AP-2. These results demonstrate that a multiplicity of type 2A phosphatases might be differentially expressed in higher plants.     

A pulsing device for packed-bed bioreactors: II. Application to alcoholic fermentation

When the immobilized cells are employed in packed-bed bioreactors several problems appear. To overcome these drawbacks, a new bioreactor based on the use of pulsed systems was developed [1]. In this work, we study the glucose fermentation by immobilized Saccharomyces cerevisiae in a packed-bed bioreactor. A comparative study was then carried out for continuous fermentation in two packed-bed bioreactors, one of them with pulsed flow. The determination of the axial dispersion coefficients indicates that by introducing the pulsation, the hydraulic behaviour is closer to the plug flow model. In both cases, the residence time tested varied from 0.8 to 2.6 h. A higher ethanol concentration and productivity (increases up to 16%) were achieved with the pulsated reactors. The volumes occupied by the CO₂ were 5.22% and 9.45% for fermentation with/without pulsation respectively. An activity test of the particles from the different sections revealed that the concentration and viability of bioparticles from the two bioreactors are similar. From the results we conclude that the improvements of the process are attributable to a mechanical effect rather than to physiological changes of microorganisms.List of Symbols D m²/s dispersion coefficient - K _is l/g inhibition substrate constant - K _ip l/g inhibition ethanol constant - K _s g/l Apparent affinity constant - P g/l ethanol concentration - q _p g/(gh) specific ethanol productivity - Q _p g/(lh) overall ethanol productivity - q _s g/(gh) specific glucose consumption rate - Q _s g/(lh) glucose consumption rate - S g/l residual glucose concentration - S(in0) g/l initial glucose concentration - V _max g/(lh) maximum rate - Y _p/s g/g yield in product     

Pollen wall development in Vigna vexillata I. Characterization of wall layers

Light and electron microscope observations characterized the layers that comprise Vigna vexillata L. pollen walls, and identified the timing of their development. Exine sculpturings form an unusually coarse ektexinous reticulum. The structure of the ektexine is granular; this differs from the columellate/tectate type of structure typical of most angiosperm pollen. The ektexine overlies a homogeneous-to-lamellar, electron-dense endexine, which in turn surrounds a thick, microfibrillar intine. Pollen grains are triporate and operculate, with Zwischenkörper and thickened intine underlying the apertures. The ektexine forms during the tetrad period of microspore development, the endexine and Zwischenkörper during the free microspore stage, and the intine during the bicelled (pollen) stage. Coarsely reticulate exine sculpturings and the granular structure of the patterned exine wall of the pollen grains are features that make this species suitable for detailed studies of pollen wall pattern formation.     

Seed protein and isozyme variations in Triticum tauschii (Aegilops squarrosa)

Sixty Triticum tauschii (Aegilops squarrosa, 2n=2x=14, DD) accessions were evaluated for the variability of high-molecular-weight (HMW) glutenins, gliadins and isozymes of seed esterase, -amylase and glucose-phosphate isomerase. Wide variability was observed for HMW-glutenins and gliadins. The implications of unique HMW-glutenin alleles for quality parameters are discussed. Isozyme evaluations indicated more variability for the Est-D ^t 5 locus as compared to the Est-D5 of bread-wheat. The polymorphism for -Amy-D ^t 1 was less than that of -Amy-D1. Similar to the bread-wheat situation, Gpi-D ^t 1 showed no polymorphism. The variability observed with the traits evaluated can be readily observed in T. turgidum x T. tauschii synthetic hexaploids (2n=6x=42, AABBDD) suggesting that T. tauschii accessions may be a rich source for enhancing the genetic variability of T. aestivum cultivars.     

Resistance to the cereal cyst nematode (Heterodera avenae Woll.) transferred from the wild grassAegilops ventricosa to hexaploid wheat by a “stepping-stone” procedure

Transfer of resistance toHeterodera avenae, the cereal cyst nematode (CCN), by a stepping-stone procedure from the wild grassAegilops ventricosa to hexaploid wheat has been demonstrated. The number of nematodes per plant was lower, and reached a plateau much earlier, in the resistant introgression line H93-8 (1–2 nematodes per plant) than in the recipient H10-15 wheat (14–16 nematodes per plant). Necrosis (hypersensitive reaction) near the nematode, little cell fusion, and few, often degraded syncytia were observed in infested H93-8 roots, while abundant, well-formed syncytia were present in the susceptible H10-15 wheat. Line H93-8 was highly resistant to the two Spanish populations tested, as well as the four French races (Fr1-Fr4), and the British pathotype Hall, but was susceptible to the Swedish pathotypes HgI and HgIII. Resistance was inherited as though determined by a single quasi-dominant factor in the F₂ generations resulting from crosses of H93-8 with H10-15 and with Loros, a resistant wheat carrying the geneCre1 (syn.Ccn1). The resistance gene in H93-8 (Cre2 orCcn2) is not allelic with respect to that in Loros. RFLPs and other markers, together with the cytogenetical evidence, indicate that theCre2 gene has been integrated into a wheat chromosome without affecting its meiotic pairing ability. Introduction ofCre2 by backcrossing into a commercial wheat backgroud increases grain yield when under challenge by the nematode and is not detrimental in the absence of infestation.     

Melatonin controls cell proliferation and modulates mitochondrial physiology in pancreatic stellate cells

Journal of Physiology and Biochemistry - We have investigated the effects of melatonin on major pathways related with cellular proliferation and energetic metabolism in pancreatic stellate cells....     

Non-linear loss of suitable wine regions over Europe in response to increasing global warming

Evaluating the potential climatic suitability for premium wine production is crucial for adaptation planning in Europe. While new wine regions may emerge out of the traditional boundaries, most of the present-day renowned winemaking regions may be threatened by climate change. Here, we analyse the future evolution of the geography of wine production over Europe, through the definition of a novel climatic suitability indicator, which is calculated over the projected grapevine phenological phases to account for their possible contractions under global warming. Our approach consists in coupling six different de-biased downscaled climate projections under two different scenarios of global warming with four phenological models for different grapevine varieties. The resulting suitability indicator is based on fuzzy logic and is calculated over three main components measuring (i) the timing of the fruit physiological maturity, (ii) the risk of water stress and (iii) the risk of pests and diseases. The results demonstrate that the level of global warming largely determines the distribution of future wine regions. For a global temperature increase limited to 2°C above the pre-industrial level, the suitable areas over the traditional regions are reduced by about 4%/°C rise, while for higher levels of global warming, the rate of this loss increases up to 17%/°C. This is compensated by a gradual emergence of new wine regions out of the traditional boundaries. Moreover, we show that reallocating better-suited grapevine varieties to warmer conditions may be a viable adaptation measure to cope with the projected suitability loss over the traditional regions. However, the effectiveness of this strategy appears to decrease as the level of global warming increases. Overall, these findings suggest the existence of a safe limit below 2°C of global warming for the European winemaking sector, while adaptation might become far more challenging beyond this threshold.