Phytotoxicity on cotton ex-plants of an 18.5 kDa protein from culture filtrates of Verticillium dahliae

A phytotoxic protein that evokes the typical symptoms of Verticillium wilt disease in seedlings of Gossypium hirsutum L. (Upland cotton) was isolated from culture filtrates of Verticillium dahliae. The protein was purified by ammonium sulfate precipitation, Sephadex-G100 fractionation, and native PAGE. The 18.5 kDa protein, designated VD18.5, appears to be a single subunit protein with an isoelectric point between 3 and 5. VD18.5 induces symptoms of leaf dehydration, chlorosis, necrosis and stem discoloration in seedlings of the disease susceptible cotton cultivar Siokra 1–4. The LD₅₀ of VD18.5 on protoplasts of Siokra 1–4 was 18 μg mL⁻¹. VD18.5 had no noticeable effect on Pima S-7, which is a disease resistant cultivar. Phytotoxic activity was partially destroyed at high temperature and was abolished by digestion with proteinase K. Mass spectrometry fingerprinting and protein sequence data from VD18.5 yielded no significant matches when submitted to the Mascot search engine and NCBI non-redundant protein databases, respectively. These results suggest that VD18.5 is a novel protein that may be involved in the development of some of the symptoms associated with Verticillium wilt disease in the cotton plant.     

Endophytic-Host Selectivity of Discula umbrinella on Quercus alba and Quercus rubra Characterized by Infection, Pathogenicity and Mycelial Compatibility

The fungal endophytic–host relationships of Discula umbrinella and two oak species, Quercus alba and Quercus rubra, were characterized on the basis of endophytic infection, pathogenicity, and mycelial compatibility. Isolates of D. umbrinella were cultured from leaves of Q. alba and Q. rubra collected from a hardwood forest located in Patuxent Wildlife Research Center in Laurel, Maryland, USA. Endophytic infection was observed on sterile leaf discs and living 2-month-old seedlings of Q. alba and Q. rubra. Fungal-host reciprocal inoculations revealed the presence of conidiomata on both hosts but conidiomata production was more abundant on Q. alba. Isolates from Q. rubra produced milder disease symptoms on both oak species. Mycelial compatibility studies identified seven different MCG groups. MCG groups 1–3 contained isolates from both oak species whereas MCG groups 4–7 contained host specific isolates. Field studies monitored the seasonal appearance of the sexual fruiting structures, perithecia, as a possible source of new genetic variation that might alter host specificity/pathogenicity of the D. umbrinella isolates on Q. alba and Q. rubra hosts. Only 1–2% of the leaves contained perithecia throughout the sampling period (April–September). Isolates collected from Q. alba differed from those collected from Q. rubra in endophytic infection, pathogenic response, and perithecia production. The results of this study suggest that the endophyte–host relationship is one of host selective preference for Q. alba, but that the endophyte has the ability to maintain the endophytic/pathogenic life cycle on the less preferred host species, Q. rubra.     

Soil conditions and land use intensification effects on soil microbial communities across a range of European field sites

Intensive land use practices necessary for providing food and raw materials are known to have a deleterious effect on soil. However, the effects that such practices have on soil microbes are less well understood. To investigate the effects of land use intensification on soil microbial communities we used a combined T-RFLP and pyrosequencing approach to study bacteria, archaea and fungi in spring and autumn at five long term observatories (LTOs) in Europe; each with a particular land use type and contrasting levels of intensification (low and high). Generally, due to large gradients in soil variables, both molecular methods revealed that soil microbial communities were structured according to differences in soil conditions between the LTOs, more so than land use intensity. Moreover, variance partitioning analysis also showed that soil properties better explained the differences in microbial communities than land use intensity effects. Predictable responses in dominant bacterial, archaeal and fungal taxa to edaphic conditions (e.g. soil pH and resource availability) were apparent between the LTOs. Some effects of land use intensification at individual field sites were observed. However, these effects were manifest when land use change affected soil conditions. Uniquely, this study details the responses of different microbial groups to soil type and land use intensification, and their relative importance across a range of European field sites. These findings reinforce our understanding of drivers impacting soil microbial community structure at both field and larger geographic scales.     

Response of the soil fungal community to multi-factor environmental changes in a temperate forest

Both environmental and climatic changes are known to influence soil microbial biomes in terrestrial ecosystems. However, there are limited data defining the interactive effects of multi-factor environmental disturbances, including N-deposition, precipitation, and air temperature, on soil fungal communities in temperate forests. A 3-year outdoor pot experiment was conducted to examine the temporal shifts of soil fungal communities in a temperate forest following N-addition, precipitation and air temperature changes. The shifts in the structure and composition of soil fungal communities were characterized by denaturing gradient gel electrophoresis and DNA sequencing. N-addition regimen induced significant alterations in the composition of soil fungal communities, and this effect was different at both higher and lower altitudes. The response of the soil fungal community to N-addition was much stronger in precipitation-reduced soils compared to soils experiencing enhanced precipitation. The combined treatment of N-addition and reduced precipitation caused more pronounced changes in the lower altitude versus those in the higher one. Certain fungal species in the subphylum Pezizomycotina and Saccharomycotina distinctively responded to N fertilization and soil water control at both altitudes. Redundancy discrimination analysis showed that changes in environmental factors and soil physicochemical properties explained 43.7% of the total variability in the soil fungal community at this forest ecosystem. Variations in the soil fungal community were significantly related to the altitude, soil temperature, total soil N content (TN) and pH value (P < 0.05). We present evidence for the interactive effects of N-addition, water manipulation and air temperature to reshape soil fungal communities in the temperate forest. Our data could provide new insights into predicting the response of soil micro-ecosystem to climatic changes.     

内生真菌对高寒草地紫花针茅凋落物分解的影响

禾草内生真菌对于提高宿主植物对生物和非生物胁迫的耐性发挥着积极作用,然而,禾草内生真菌共生体凋落物分解对于草地生态系统养分循环的影响仍不清楚。本研究以紫花针茅为研究对象,比较带内生真菌（E+）与不带内生真菌（E-）植株凋落物分解过程中重量及全氮、木质素和纤维素含量,以及木质素∶N、纤维素∶N与全氮、木质素和纤维素残留率的变化,以期揭示内生真菌在紫花针茅凋落物分解过程中发挥的作用。主要结果如下：E+紫花针茅凋落物分解速率高于E-,具有更短的分解周期;随着时间的延长,紫花针茅凋落物木质素含量和木质素∶N由E+显著高于E-逐渐变为二者之间差异不显著,纤维素含量、纤维素∶N和纤维素残留率则逐渐变为E+显著低于E-;另外,随着分解时间延长,E+和E-凋落物全氮含量呈上升趋势,而全氮残留率呈现出下降-上升-下降的趋势,且后期E+显著低于E-。因此,内生真菌不同程度地促进了紫花针茅凋落物全氮、木质素和纤维素的降解。     

疯草内生真菌Undifilum oxytropis次级代谢产物分析

为阐明疯草内生真菌Undifilum oxytropis次级代谢产物的化学组成,为其生物活性研究和开发利用提供依据,以本实验室保存的疯草内生真菌Undifilum oxytropis为研究菌株,采用察氏液体培养基对该菌发酵培养30d,收集菌丝体和发酵液,按照植物化学成分传统预试方法进行预试。在此基础上用不同极性有机溶剂梯度萃取得到菌丝体和发酵液石油醚相、氯仿相、乙酸乙酯相和正丁醇相,经薄层色谱检测及气相检测方法对其次级代谢产物进行初步分析。系统预试表明,Undifilum oxytropis发酵产物中含有生物碱、蛋白质、酚类与鞣质、黄酮类、醌类、挥发油和油脂等物质;薄层色谱检测发现,Undifilum oxytropis发酵产物中至少含有5种化合物,且菌丝体和发酵液正丁醇相均有苦马豆素及其类似物存在;气相色谱分析Undifilum oxytropis发酵液中苦马豆素含量为5.17×10~(-3)g/L。疯草内生真菌Undifilum oxytropis发酵产物含有苦马豆素等多种次级代谢产物。     

Relative abundance and activity of melanized hyphae in different soil ecosystems

Here we report on the frequency of melanized fungal hyphae in 323 soils, covering different land use types. The proportion of total hyphae that was melanized averaged 61%. Arable fields with loamy sand, heathlands and city parks on sandy soils had the highest percentage of melanized hyphae. In addition to the frequency determinations, a microcosm study was performed on the role of melanized hyphae in two different ecosystems: an ex-arable field and a forest. Melanized hyphae appeared to be part of the active hyphae in the forest soil but not in the ex-arable soil. In conclusion, our results indicate that (1) melanized hyphae represent a large proportion of the total fungal biomass in soils and that (2) their function might differ between ecosystems.     

Fungal endophyte Phomopsis liquidambari B3 enriches the diversity of nodular culturable endophytic bacteria associated with continuous cropping of peanut

Previously, the application of the fungal endophyte Phomopsis liquidambari B3 exhibited notable effect in alleviating soil obstacles caused by continuous cropping of peanut. It was supposed that P. liquidambari B3 could enhance the efficiency of nodulation and nitrogen fixation by enriching the diversity of nodular endophytic bacteria under continous monocropping system. To verify this hypothesis, a pot experiment was conducted under natural conditions. Four treatments were set as follow: discontinuous cropping soil (NS), continuous cropping soil (CK), continuous cropping soil inoculate with actived P. liquidambari B3 (CSB3), and continuous cropping soil inoculate with sterilized P. liquidambari B3 (CSIB3). As a result, a total of 120 isolates were obtained. Our study clearly declared that the addition of fungal endophyte effectively enriched the genetic diversity and the community composition in CSB3. In contrast, a low genetic diversity level and simplex community structure were exhibited in CK and CSIB3. Meanwhile, the positive percentage rate and corresponding community composition of the plant growth-promoting isolates in NS and CSB3 were larger and more abundant than the other. This is the first time to describe the effects of fungal endophytes on the diversity of the nodular culturable endophytic bacteria associated with continuous cropping of peanut.