Arbuscular Mycorhizal Fungi Associated with the Olive Crop across the Andalusian Landscape: Factors Driving Community Differentiation

Background

In the last years, many olive plantations in southern Spain have been mediated by the use of self-rooted planting stocks, which have incorporated commercial AMF during the nursery period to facilitate their establishment. However, this was practised without enough knowledge on the effect of cropping practices and environment on the biodiversity of AMF in olive orchards in Spain.

Methodology/Principal Findings

Two culture-independent molecular methods were used to study the AMF communities associated with olive in a wide-region analysis in southern Spain including 96 olive locations. The use of T-RFLP and pyrosequencing analysis of rDNA sequences provided the first evidence of an effect of agronomic and climatic characteristics, and soil physicochemical properties on AMF community composition associated with olive. Thus, the factors most strongly associated to AMF distribution varied according to the technique but included among the studied agronomic characteristics the cultivar genotype and age of plantation and the irrigation regimen but not the orchard management system or presence of a cover crop to prevent soil erosion. Soil physicochemical properties and climatic characteristics most strongly associated to the AMF community composition included pH, textural components and nutrient contents of soil, and average evapotranspiration, rainfall and minimum temperature of the sampled locations. Pyrosequencing analysis revealed 33 AMF OTUs belonging to five families, with Archaeospora spp., Diversispora spp. and Paraglomus spp., being first records in olive. Interestingly, two of the most frequent OTUs included a diverse group of Claroideoglomeraceae and Glomeraceae sequences, not assigned to any known AMF species commonly used as inoculants in olive during nursery propagation.

Conclusions/Significance

Our data suggests that AMF can exert higher host specificity in olive than previously thought, which may have important implications for redirecting the olive nursery process in the future as well as to take into consideration the specific soils and environments where the mycorrhized olive trees will be established.     

Energy Crop and Biotechnology for Biofuel Production

Selection of energy crops is the first priority for large-scale biofuel production in China. As a major topic, it was extensively discussed in the Second International Symposium on Bioen-     

重组人MASP2蛋白在大肠杆菌中的表达和纯化

目的:获得酶原形式的重组人甘露聚糖结合凝集素相关丝氨酸蛋白酶2(MASP2)。方法:在大肠杆菌中诱导表达重组人MASP2全长蛋白,包涵体裂解后,经复性、透析、浓缩、考马斯亮蓝染色、SDS-PAGE及Western印迹,鉴定纯化结果及酶活性。结果:复性后的MASP2蛋白经考马斯亮蓝染色未见杂带。自激活实验表明,当MASP2浓度在1μmool/L以下时,无论在4℃还是37℃,都能较稳定地保持酶原形式;蛋白浓度为3.5μmool/L时只能在4℃保持稳定,37℃发生自激活;蛋白浓度达到12μmool/L后,在4℃时已不能稳定存在。结论:获得了较纯的重组人MASP2蛋白,且具有自激活活性。     

DNA synthesis and distribution in parthenogenetic bovine embryos

Parthenogenetically activated, in vitro-matured bovine oocytes and parthenogenotes obtained at 2 to 4 days post activation were analyzed by 3H-thymidine autoradiography for the timing of the S-phase and for distribution of newly replicated DNA, respectively. Spread pronuclear parthenogenotes revealed that the DNA synthesis in electrically stimulated oocytes commenced at 14 h post activation. At 20 to 24 h, a maximum number of labeled pronuclei was reached (25 to 38%), and DNA synthesis persisted in some parthenogenotes up to 30 h post activation. The DNA labeling detected on semi-thin sections showed that the distribution of newly synthesized DNA in the nuclei of 3- to 16-cell parthenogenotes was mostly irregular or abnormal, documenting that the apparent morphological normalcy of parthenogenotes was in contrast to the data concerning the DNA synthesis and distribution.     

丝素蛋白/羟基磷灰石复合物对磷酸钙骨水泥性能的影响

目的：磷酸钙骨水泥（Calcium phosphate cement,CPC）以其诸多优点正得到了越来越多的应用,但其较差的力学性能表现也限制了它的使用范围。本研究目的在于改善磷酸钙骨水泥的力学性能,同时评估改性后的磷酸钙骨水泥的其他性能。方法：通过丝素蛋白（Silk fibroin,SF）的矿化自组装方法制备丝素蛋白／羟基磷灰石复合物（silk fibroin/hydroxyapitite composite, SF/HA）。按照1％、2％、3％、4％的质量分数加入磷酸钙骨水泥中,与磷酸钙骨水泥组对比。比较内容包括力学强度、抗渍散性能及细胞毒性。结果：以丝素蛋白溶液为液相组的磷酸钙骨水泥强度大约为35MPa。随后随着添加丝素蛋白／羟基磷灰石复合物的质量分数从1％增至3％,磷酸钙骨水泥的强度逐渐增加（P〈0．05）,最高约至45MPa。而当丝素蛋白／羟基磷灰石的质量分数达到4％时,磷酸钙骨水泥的强度较质量分数3％组小幅度下降至43MPa（P〈0．05）。以丝素蛋白溶液作为液相时,磷酸钙骨水泥的抗溃散能力也得到了加强。在MTT法测定细胞活力的对照实验中,无论是加入丝素蛋白溶液或丝素蛋白／羟基磷灰石复合物,都未观察到细胞毒性。结论：在磷酸钙骨水泥中加入3％质量分数的丝素蛋白／羟基磷灰石复合物,能显著提高磷酸钙骨水泥的抗压强度。而丝素蛋白溶液作为液相可改善磷酸钙骨水泥的抗溃散能力。同时,丝素蛋白和丝素蛋白／羟基磷灰石复合物都不表现出细胞毒性。更理想的力学强度和更强的抗溃散能力,大大扩展了磷酸钙骨水泥的应用范围。     

phlD-based genetic diversity and detection of genotypes of 2,4-diacetylphloroglucinol-producing Pseudomonas fluorescens

Diversity within a worldwide collection of 2,4-diacetylphloroglucinol-producing Pseudomonas fluorescens strains was assessed by sequencing the phlD gene. Phylogenetic analyses based on the phlD sequences of 70 isolates supported the previous classification into 18 BOX-PCR genotypes (A-Q and T). Exploiting polymorphisms within the sequence of phlD, we designed and used allele-specific PCR primers with a PCR-based dilution endpoint assay to quantify the population sizes of A-, B-, D-, K-, L- and P-genotype strains grown individually or in pairs in vitro, in the rhizosphere of wheat and in bulk soil. Except for P. fluorescens Q8r1-96, which strongly inhibited the growth of P. fluorescens Q2-87, inhibition between pairs of strains grown in vitro did not affect the accuracy of the method. The allele-specific primer-based technique is a rapid method for studies of the interactions between genotypes of 2,4-diacetylphloroglucinol producers in natural environments.     

The biosynthesis of brain gangliosides. Ganglioside-glycosylating activity in rat brain neuronal perikarya fraction.

Rat brain homogenate and the synaptosmal and neuronal perikarya fractions from 17-day-old rats were compared for their activities in sialosylating endogenous gangliosides and transferring N-acetylneuraminic acid and galactose to several glycolipids in vitro. The sialosylation of endogenous gangliosides and the activities of sialosyltransferases acting either on lactosylceramide or haematoside as acceptors, as well as galactosyltransferase acting on Tay-Sachs ganglioside as acceptor, were between 3-and 12-fold higher in the neuronal perikarya fraction than in whole homgenate on a protein or ganglioside basis. The activities found in the synaptosomal fraction were negligible. No evidence was found to indicate that the low activities in this fraction were due to the presence of inhibitors of the transfer activities or to inacessibility of the substrates to their respective enzymes. These findings, and the time course of labelling of gangliosides of the neuronal perikarya and synaptosomes from rats that received an injection of N-[3H]acetylmannosamine, indicate that the main cellular site of glycosylation of neuronal gangliosides is in the neuronal perikarya.     

Differential Ability of Genotypes of 2,4-Diacetylphloroglucinol-Producing Pseudomonas fluorescens Strains To Colonize the Roots of Pea Plants

Indigenous populations of 2,4-diacetylphloroglucinol (2,4-DAPG)-producing fluorescent Pseudomonas spp. that occur naturally in suppressive soils are an enormous resource for improving biological control of plant diseases. Over 300 isolates of 2,4-DAPG-producing fluorescent Pseudomonas spp. were isolated from the rhizosphere of pea plants grown in soils that had undergone pea or wheat monoculture and were suppressive to Fusarium wilt or take-all, respectively. Representatives of seven genotypes, A, D, E, L, O, P, and Q, were isolated from both soils and identified by whole-cell repetitive sequence-based PCR (rep-PCR) with the BOXA1R primer, increasing by three (O, P, and Q) the number of genotypes identified previously among a worldwide collection of 2,4-DAPG producers. Fourteen isolates representing eight different genotypes were tested for their ability to colonize the rhizosphere of pea plants. Population densities of strains belonging to genotypes D and P were significantly greater than the densities of other genotypes and remained above log 6.0 CFU (g of root)⁻¹ over the entire 15-week experiment. Genetic profiles generated by rep-PCR or restriction fragment length polymorphism analysis of the 2,4-DAPG biosynthetic gene phlD were predictive of the rhizosphere competence of the introduced 2,4-DAPG-producing strains.     

THE SITE OF SYNTHESIS OF GANGLIOSIDES IN THE CHICK OPTIC SYSTEM

Abstract– In the retinas of 1-day-old chickens that received an intraocular injection of N-[³H]acetylmannosamine the labelling of N-acetylneuraminic acid and CMP-N-acetylneuraminic acid increased for at least 8 h and that of gangliosides for at least 24 h after injection. In the optic tectum contralateral to the injected eye at 8 h after the intraocular injection, the labelling of gangliosides exceeded the labelling of gangliosides in the ipsilateral tectum by approx 20-fold. In the contralateral tectum the highest concentration of labelled gangliosides was in subfractions enriched in synaptosomes and synaptic plasma membranes. No significant contralateral ipsilateral differences were found in the acid soluble substances of the tectum. In the optic tectum, labelled gangliosides appeared earlier in the neuronal perikarya than in synaptosomes when the injection was intracranial. Conversely, when the injection was intraocular the labelling appeared earlier in the synaptosomes than in the neuronal perikarya. The radioactivity pattern of the optic tectum gangliosides resembled the pattern of retina gangliosides when N-[³H]acetylmannosamine was injected intraocularly, but when N-[³H]acetylmannosamine was given intracerebrally the radioactivity pattern resembled that of optic tectum gangliosides. Intraocular injection of colchicine or vinblastine did not affect the labelling of retinal gangliosides from N-[³H]acetylmannosamine injected into the same eye but prevented the appearance of labelled gangliosides in the optic tectum. In vitro the ganglioside glycosylating activity of optic tectum synaptosomes and synaptic plasma membranes was between 6 and 10-fold lower than that found in the optic tectum neuronal perikarya. These findings support the notion that the main subcellular site of synthesis of neuronal gangliosides is in the neuronal perikarya, from which they are translocated to the nerve endings.     

Adult Rat Retina Interneurons Synthesize GD3: GD3 Expression by These Cells Is Regulated by Cell-Cell Interactions

GD3, a ganglioside of the lactosyl series, is prevalent in rat retina neuronal cells. We studied here whether rat retina neurons synthesize their own surface GD3 or if they acquire it from Müller glia cells. We analyzed the activity of GD3 synthase and the in vivo labeling of gangliosides from N-[3H]acetylmannosamine in adult rat retinas after selective destruction of Müller glia cells with the gliotoxic alpha-D,L-aminoadipate (AAA). Immunostaining of rat retina sections and western blot analysis with an antivimentin antibody confirmed the gliotoxic effect of AAA. Neither GD3 synthase activity nor the in vivo labeling of GD3 and other gangliosides was significantly affected by AAA, indicating that neuronal cells synthesize their own GD3. We next analyzed the regulation of the expression of GD3 by these neurons in culture. About 80% of freshly dissociated cells from retina of 4-day-old rats (R4) immunoexpress surface GD3. After 3 days in dispersed cell culture conditions, GD3 expression was under the limit of detection in 80% of neuronal cells, indicating a failure of these cells to maintain the expression of surface GD3 in these experimental conditions. Most flat Müller glia-derived cells present in these cultures were GD3 positive. Surface GD3 was detected in approximately 60% of neuronal cells dissociated from R4 tissue that was developed in vitro as an organ culture for 3 days. Likewise, approximately 50% of neurites that had grown out from R4 retinal explants within 3 days in culture and whose neuronal character was indicated by immunoexpression of growth-associated protein GAP-43 were GD3 positive. These findings suggest that the tissue organization and/or specific interactions modulate GD3 expression in neuronal cells. Under dispersed-cell culture conditions, c-pathway gangliosides (GQ1c and GT1c), which are built up from the sialylation of GD3 and later completion of the oligosaccharide backbone, were detected in approximately 60% of neuronal cells, suggesting a maintenance of production of GD3 as an intermediate for gangliotetraosyl gangliosides.