多年卧孔菌属三个中国新记录种

报道了多年卧孔菌属Perenniporia 3个中国新记录种。竹生多年卧孔菌P. bambusicola采自云南省,孔口表面橙色且仅生长于竹子上;树状多年卧孔菌P. dendrohyphidia采自广西,具树状菌丝;灰黄多年卧孔菌P. isabellina采自云南省,担子果平伏,孔口灰黄色,具树状骨架菌丝,且担孢子较大。根据采集的标本材料提供了它们的详细描述和显微结构图。     

多年卧孔菌属五个中国新记录种

报道了多年卧孔菌属Perenniporia5个中国新记录种。非洲多年卧孔菌Perenniporia africana采自安徽省,孔口表面浅黄色至赭色,且骨架菌丝不分枝;下延多年卧孔菌P.decurrata采自云南省,担子果盖形,具较小孔口和担孢子;椭圆孢多年卧孔菌P.ellipsospora采自云南省,担子果平伏,孔口圆形至多角形,担孢子不平截;硬多年卧孔菌P.inflexibilis采自福建省,担子果盖形,孔口表面灰白至浅褐色,担孢子无色至浅黄色;黄多年卧孔菌P.xantha采自海南省,孔口较小,表面呈黄色,且在KOH试剂中呈深褐色。根据采集的标本材料提供了它们的详细描述和显微结构图。     

When the ribosomal DNA does not tell the truth: The case of the taxonomic position of Kurtia argillacea, an ericoid mycorrhizal fungus residing among Hymenochaetales

The nuclear ribosomal DNA (nuc-rDNA) is widely used for the identification and phylogenetic reconstruction of Agaricomycetes. However, nuc-rDNA-based phylogenies may sometimes be in conflict with phylogenetic relationships derived from protein coding genes. In this study, the taxonomic position of the basidiomycetous mycobiont that forms the recently discovered sheathed ericoid mycorrhiza was investigated, because its nuc-rDNA is highly dissimilar to any other available fungal sequences in terms of nucleotide composition and length, and its nuc-rDNA-based phylogeny is inconclusive and significantly disagrees with protein coding sequences and morphological data. In the present work, this mycobiont was identified as Kurtia argillacea (= Hyphoderma argillaceum) residing in the order Hymenochaetales (Basidiomycota). Bioinformatic screening of the Kurtia ribosomal DNA sequence indicates that it represents a gene with a non-standard substitution rate or nucleotide composition heterogeneity rather than a deep paralogue or a pseudogene. Such a phenomenon probably also occurs in other lineages of the Fungi and should be taken into consideration when nuc-rDNA (especially that with unusual nucleotide composition) is used as a sole marker for phylogenetic reconstructions. Kurtia argillacea so far represents the only confirmed non-sebacinoid ericoid mycorrhizal fungus in the Basidiomycota and its intriguing placement among mostly saprobic and parasitic Hymenochaetales begs further investigation of its eco-physiology.     

Revealing the non-overlapping characteristics between original centers and genetic diversity of Purpureocillium lilacinum

Purpureocillium lilacinum is a free-living, multitrophic, cosmopolitan fungus. This study estimated the genetic diversity of P. lilacinum and its geographic distribution pattern based on a global ITS dataset. At the intercontinental levels, the highest genetic diversity was in Asia. Divergence time estimation indicated that Hap 5, Hap 9, Hap 31, and Hap 32 from the Hainan, Guangdong, Jiangxi, and Fujian regions of China in Asia were the earliest divergence haplotypes. The neutrality test indicated that P. lilacinum is undergoing population expansion. These results support that the southeastern coastal region of China is the original center of P. lilacinum, while the East Asian region adjacent to this origin is the center of the genetic diversity of P. lilacinum. The geographical distribution pattern of P. lilacinum showed that only one haplotype (Hap 1) was globally distributed and that most haplotypes were distributed in Asia, while the few remaining haplotypes were scattered on other continents. The results provide valuable information to elucidate the origin, genetic diversity, and evolution of fungi.     

Improving the recovery of qPCR-grade DNA from sludge and sediment

DNA extraction is often considered as the limiting step of most molecular approaches in ecology and environmental microbiology. Ten existing DNA extraction protocols were compared for recovery of DNA from sludge and a modified version of the protocol described by Porteous et al. (Mol Ecol 6:787–791, 1997) was determined to be the best method for recovery of DNA suitable for PCR. In this respect, it appeared that the commonly used guanidine isothiocyanate could impair the quality of the extracted DNA unless its concentration is lowered. Second, conditioning the samples as liquors as opposed to pellets critically impacts the outcome of the extraction. The suitability of the modified Porteous protocol for quantitative PCR applications is demonstrated in a series of experiments showing the absence of interfering coextracted inhibitors and the linear correspondence between the concentrations of input target DNA and PCR product. Interestingly, it is also shown that the nature of the environmental matrices affects the recovery yield of both circular plasmids and chromosomal DNA, resulting in an apparent fluctuation of the plasmid copy number per cell. This means that quantitative data obtained by PCR remain comparable as long as they apply to an identical target sequence extracted from a similar environment and amplified under the same conditions.     

南黄海沉积物中的丝孢菌初报

从南黄海采集沉积物样品19份,共分离到74个丝孢菌分离物。除22个青霉属菌株未鉴定至种外,其余经鉴定属于18属20种。其中包括1个中国新记录属Devriesia;2个中国新记录种Devriesia pseudoamericana、Scedosporium dehoogii;其余18种为中国已知种。对中国新记录属种进行形态学描述和基于ITS序列的分子生物学分析,对18个国内已报道种则只作分布和生境的引证。所有菌种均保存在中国海洋大学海洋生物标本室(OUCMB)。     

Characterization of a formaldehyde degrading fungus Penicillium chrysogenum DY-F2 isolated from deep sea sediment

A formaldehyde-degrading fungus was isolated from deep sea sediment of East Pacific by enrichment culture technique and was identified as Penicillium chrysogenum DY-F2 based on microscopic spore morphology and 18S rRNA gene sequence analysis. The fungus showed high formaldehyde resistance and was able to grow in the presence of formaldehyde up to 3000 mg l⁻¹. The optimal temperature and pH for the growth of fungus in the presence of 1000 mg l⁻¹ of formaldehyde was 25 °C and 6.0, respectively. The fungus was able to degrade formaldehyde as the sole source of carbon and energy with the formation of formic acid as the intermediate. Degradation of formaldehyde by the fungus conformed to a first-order kinetic model. This study showed that the deep sea sediment fungi are the potential microbial resources for bioremediation of formaldehyde pollution in marine environment.     

Marine fungal communities in water and surface sediment of a sea cucumber farming system: habitat-differentiated distribution and nutrients driving succession

The planktonic and benthic fungi in a sea cucumber farming system were simultaneously investigated on three sampling dates. Analyses of SSU rRNA gene libraries of four samples revealed 131 fungal operational taxonomic units, of which 58 % were potentially novel. Chytridiomycota, Blastocladiomycota and Monoblepharidomycota were detected only from sediment, whereas ascomycetes and basidiomycetes dominated in sediment and water, respectively. Cryptomycota phylotypes were detected in both water and sediment samples. Based on terminal-restriction fragment length polymorphisms, distinct succession and contrasting community structure were found between planktonic and benthic habitats. Redundancy analysis indicated that the concentration of dissolved silicate in surface water and N:P in porewater were the most significant abiotic variables shaping the planktonic and benthic communities, respectively. This study indicates that plankton and benthos are distinct habitats for fungal distribution even in shallow coastal systems, and nutrients and stoichiometric ratios play important roles in driving succession of fungal communities.     

Phenylacetic acid-producing Rhizoctonia solani represses the biosynthesis of nematicidal compounds in vitro and influences biocontrol of Meloidogyne incognita in tomato by Pseudomonas fluorescens strain CHA0 and its GM derivatives

AIMS: The aim of the present investigation was to determine the influence of Rhizoctonia solani and its pathogenicity factor on the production of nematicidal agent(s) by Pseudomonas fluorescens strain CHA0 and its GM derivatives in vitro and nematode biocontrol potential by bacterial inoculants in tomato. METHODS AND RESULTS: One (Rs7) of the nine R. solani isolates from infected tomato roots inhibited seedling emergence and caused root rot in tomato. Thin layer chromatography revealed that culture filtrates of two isolates (Rs3 and Rs7) produced brown spots at Rf-values closely similar to synthetic phenylacetic acid (PAA), a phytotoxic factor. Filtrates from isolate Rs7, amended with the growth medium of P. fluorescens, markedly repressed nematicidal activity and PhlA'-'LacZ reporter gene expression of the bacteria in vitro. On the contrary, isolate Rs4 enhanced nematicidal potential of a 2,4-diacetylphloroglucinol overproducing mutant, CHA0/pME3424, of P. fluorescens strain CHA0 in vitro. Therefore, R. solani isolates Rs4 and Rs7 were tested more rigorously for their potential to influence biocontrol effectiveness of the bacterial agents. Methanol extract of the culture filtrates of PAA-producing isolate Rs7 resulting from medium amended with phenylalanine enhanced fungal repression of the production of nematicidal agents by bacteria, while amendments with zinc or molybdenum eliminated such fungal repression, thereby restoring bacterial potential to cause nematode mortality in vitro. A pot experiment was carried out, 3-week-old tomato seedlings were infested with R. solani isolates Rs4 or Rs7 and/or inoculated with Meloidogyne incognita, the root-knot nematode. The infested soil was treated with aqueous cell suspensions (10(8) CFU) of P. fluorescens strain CHA0 or its GM derivatives or left untreated (as a control). Observations taken 45 days after nematode inoculation revealed that, irrespective of the bacterial treatments, galling intensity per gram of fresh tomato roots was markedly higher in soil amended with isolate Rs4 than in Rs7-amended soils. Soil amendments with R. solani and the bacterial antagonists resulted in substantial reductions of the number of galls per gram of root. These results are contradictory to those obtained under in vitro conditions where culture filtrates of PAA-positive Rs7 repressed the production of nematicidal compounds. Plants grown in Rs7-amended soils, with or without bacterial inoculants, had lesser shoot and root weights than plants grown in nonamended or Rs4-amended soils. Moreover, amendments with Rs7 substantially retarded root growth and produced necrotic lesions that reduced the number of entry sites for invasion and subsequent infection by nematodes. Populations of P. fluorescens in the tomato rhizosphere were markedly higher in Rs7-amended soils. CONCLUSIONS: PAA-producing virulent R. solani drastically affects the potential of P. fluorescens to cause death of M. incognita juveniles in vitro and influences bacterial effectiveness to suppress nematodes in tomato roots. SIGNIFICANCE AND IMPACT OF THE STUDY: As most agricultural soils are infested with root-infecting fungi, including R. solani, it is likely that some PAA-producing isolates of the fungus may also be isolated from such soils. The inhibitory effect of PAA-producing R. solani on the biosynthesis of nematicidal agent(s) critical in biocontrol may reduce or even eliminate the effectiveness of fluorescent pseudomonads against root-knot nematodes, both in nursery beds and in field conditions. Introduction of bacterial inoculants, for the control of any plant pathogen, should be avoided in soils infested with PAA-producing R. solani. Alternatively, the agents could be applied together with an appropriate quantity of fungicide or chemicals such as zinc to create an environment more favourable for bacterial biocontrol action.     

DNA Fingerprinting of Paecilomyces Strains of Potential use for the Biological Control of Pests

Summary Telomeric fingerprinting was found to be highly differentiating for Paecilomyces fumosoroseus and Paecilomyces lilacinus isolates in comparison to intron splice site PCR and is therefore a good method for quality control of future products based on these fungi. Although the telomeric restriction length polymorphisms correctly divided the isolates into their appropriate species, further correlation with host range or geographical origin of the isolates was not found. In this respect, intron splice site PCR was more informative taxonomically. The chromosome numbers inferred from telomeric fingerprints were seven chromosomes for P. lilacinus and between six and nine chromosomes for P. fumosoroseus.     

黑曲霉htmA基因的分离鉴定及其功能分析

本研究通过分析比较黑曲霉基因组与人、哺乳动物和酿酒酵母基因组序列同源性,首次分离鉴定了黑曲霉htmA基因,该基因长3459bp,编码1083个氨基酸。已知真核生物的htmA基因编码一种类α-甘露糖苷酶I的非必须蛋白HTMA,在内质网中参与降解非正确折叠的糖蛋白,htmA基因的破坏会延迟非正确折叠糖蛋白的降解。为分析htmA基因在黑曲霉中的功能,运用同源重组技术敲除黑曲霉基因组中的htmA基因,获得htmA基因缺失突变菌株,并进行了缺失株外源漆酶分泌能力的检测。结果表明黑曲霉htmA基因的破坏延缓了外源漆酶的降解,由此推测黑曲霉htmA基因编码蛋白HTMA具有与酵母、哺乳动物HTM1P/EDEM类似的功能作用。     

A PCR-based method for detecting the mycelia of stipitate hydnoid fungi in soil

To reduce the reliance on sporocarp records for conservation efforts, information on the below-ground distribution of specific fungal species, such as stipitate hydnoid fungi, is required. Species-specific primers were developed within the internal transcribed spacer (ITS1 and ITS2) regions for 12 hydnoid fungal species including Bankera fuligineoalba, Hydnellum aurantiacum, H. caeruleum, H. concrescens, H. ferrugineum, H. peckii, Phellodon confluens, P. melaleucus, P. niger, P. tomentosus, Sarcodon glaucopus and S. squamosus. The specificity of the primer pairs was tested using BLAST searches and PCR amplifications. All primers amplified DNA only of the target species with the exception of those designed for P. melaleucus. In order to assess the ability of the primers to detect DNA from mycelium in soil, DNA extracted from soil samples taken from around solitary H. peckii sporocarps was amplified with the H. peckii primer 1peck and ITS2. H. peckii DNA was detected in 70% of all soil samples and up to 40 cm away from the base of individual sporocarps. The development of these species-specific primers provides a below-ground alternative for monitoring the distribution of these rare fungi.     

Localisation of phosphomonoesterase activity in ectomycorrhizal fungi grown on different phosphorus sources

Phosphorus (P) is a major limiting nutrient for plants in boreal forest ecosystems where a substantial part of the total P is sequestered in organic compounds. Some ectomycorrhizal (ECM) fungi are known to produce phosphomonoesterases, enzymes that degrade organic P sources. Here, we test 16 ECM species for this enzymatic activity by growing them on media containing orthophosphate, phytic acid or apatite. A method with an overlay gel that determined both phosphomonoesterase activity and its spatial distribution was developed. The phosphomonoesterase activity was not significantly higher when growing on organic P; conversely some isolates only produced measurable enzyme activity when grown on apatite. Species-specific variations with respect to phosphomonoesterase activity as well as growth responses to different substrates were found. The production of phosphomonoesterases was found to be widespread in ECM fungi and the enzyme activity did not need induction by organic P. The enzyme activity was highest in the central parts of the mycelia, potentially reflecting breakdown and recycling of phospholipids from old hyphae or potentially higher mycelial density.     

Phosphorus speciation in Myall Lake sediment, NSW, Australia

The amount of phosphorus and its fractions in the sediment of Lake Myall, NSW, Australia, was assessed using a sequential extraction technique. Five sedimentary phosphorus reservoirs were measured, namely loosely sorbed phosphorus (NH₄Cl–P), iron associated phosphorus (BD–P), calcium bound phosphorus (HCl–P), metal oxide bound phosphorus (NaOH–P) and residual phosphorus (organic and refractory P, Res-P). Samples were taken from the deep and shallow sites of the lake. During the analysis, the average concentrations of each fraction of phosphorus was calculated. The results depicted that the total phosphorus (TP) content and chemically extractable phosphorus in both fine and coarse sediment fractions from the deep sites of the lake were significantly higher than those of the shallow sites, except for HCl–P extracted from the fine sediment fraction. Sediment TP was also strongly and positively correlated to sediment Fe. The phosphorus in the sediment mainly consisted of BD–P and Res-P, while NH₄Cl–P and HCl–P only contributed a minor part. The rank order of the different phosphorus extracts was similar for the two sites, namely Residual-P > BD–P > NaOH–P > HCl–P > NH₄Cl–P.     

The quality of organic matter mediates the response of heterotrophic biofilms to phosphorus enrichment of the water column and substratum

1. Heterotrophic biofilms are important drivers of community respiration, nutrient cycling and decomposition of organic matter in stream ecosystems. Both organic matter quality and nutrient levels have been shown to affect biofilm biomass and activity individually, but both factors have rarely been manipulated simultaneously. 2. To experimentally manipulate the organic matter quality and phosphorus (P) levels of both the substratum and water column, we first used cellulose cloth as a low‐quality organic material and enhanced its quality and P‐content by amending the underlying agar with maltose and P, respectively (Experiment I). To manipulate water column P, artificial substrata were incubated in low‐ and high‐P sites of a whole‐stream P‐enrichment in lowland Costa Rica. 3. Results from Experiment I suggest that heterotrophic biofilm respiration on cellulose cloth is co‐limited by carbon (C) and P. Biofilm respiration responded in an additive manner to combined effects of maltose and P‐enrichment of water column and synergistically to maltose and high‐P in substrata. 4. As decomposing organic matter that supports heterotrophic biofilms varies naturally in its labile C content along with other physical and chemical properties, we conducted a second experiment (Experiment II) in which we amended leaf discs from two species (Trema integerrima, a labile C source and Zygia longifolia, a recalcitrant C source) with maltose. We incubated the substrata in low‐ and high‐P sites of the P‐enrichment stream. 5. Results from Experiment II indicate that biofilm respiration on a labile C source (Trema) was not C‐limited, while biofilm respiration on a recalcitrant C source (Zygia) was C‐limited. Phosphorus stimulated the biofilm respiration and breakdown rate on Trema, but not on Zygia, supporting the hypothesis that the stimulatory effect of P‐enrichment is dependent on the availability of labile C in decomposing leaves. 6. Our results suggest that the interactive effects of organic matter quality and nutrient loading of streams can significantly increase microbial biofilm activity, potentially altering the trophic base of stream food webs. Researchers should consider both the organic matter quality and the enrichment of both water column and substrata to better predict the effects of anthropogenic nutrient loading to stream the ecosystems.     

Direct cloning of a xylanase gene from the mixed genomic DNA of rumen fungi and its expression in intestinal Lactobacillus reuteri

A relatively newly defined xylanase gene, xynR8, was obtained directly from a mixed DNA sample prepared from unpurified rumen fungal cultures by PCR amplification. The DNA sequence of xynR8 revealed that the gene was 884 bp in size and encoded amino acid sequences with a molecular weight of 27.9 kDa. XynR8 belonged to glycosyl hydrolase family 11, and the catalytic site residues were also found in its amino acid sequence. The main hydrolysis products of XynR8 were xylobiose, xylotriose and xylotetrose, which indicated that it belonged to the endoxylanases. The xynR8 gene was constructed so as to express and secrete under the control of the Lactococcus lactis lac A promoter and its secretion signal, and was transformed into L. reuteri Pg4, a strain isolated from the gastrointestinal tract of broiler chickens. The L. reuteri transformants harboring xynR8 not only acquired the capacity to break down xylan, but also maintained their high adhesion efficiency to mucin and mucus and their resistance to bile salts and acid.     

中国木层孔菌属三个新记录种

报道了木层孔菌属Phellinus 3个中国新记录种。赤杨木层孔菌Phellinus alni典型特征为菌盖具有较宽的同心环带和清晰的密集环纹菌核;黑木层孔菌P. nigricans具有较大的担孢子;东方木层孔菌P. orienticus具有平伏的子实体,担孢子相对较小。对这3个种进行了详细的描述和显微结构绘图。     

Degradation rates of organic phosphorus in lake sediment

Phosphorus (P) binding groups were identified in phytoplankton, settling particles, and sediment profiles by ³¹P NMR spectroscopy from the Swedish mesotrophic Lake Erken. The ³¹P NMR analysis revealed that polyphosphates and pyrophosphates were abundant in the water column, but rapidly mineralized in the sediment. Orthophosphate monoesters and teichoic acids degraded more slowly than DNA-P, polyphosphates, and P lipids. Humic acids and organic acids from phytoplankton were precipitated from the NaOH extract by acidification and identified by ³¹P NMR spectroscopy. The precipitated P was significantly more recalcitrant than the P compound groups remaining in solution, but does not constitute a major sink of P as it did not reach a stable concentration with depth, which indicates that it may eventually be degraded. Since P also precipitated from phytoplankton, the origin of humic-P can not be related solely to allochthonous P.     

一种适用于PCR反应的酵母菌、无绿藻及丝状真菌DNA提取方法

目的 介绍一种从酵母、无绿藻及丝状真菌中提取DNA以用于PCR反应的方法。方法 所用菌种包括临床分离的未知菌株和保藏菌株共23株：未知酵母菌（5株）、真皮毛孢子菌（1株）、糠秕马拉色菌（1株）、季也蒙念珠菌（5株）、未知丝状真菌（6株）、无绿藻（1株）、烟曲霉（2株）、拟青霉菌（1株）、茎点霉（1株）。用溶细胞酶（lyticase）结合Biospin真菌基因组DNA提取试剂盒提取基因组DNA,A260／A280检测纯度并计算质量浓度,用真菌通用引物ITS1／ITS4扩增真菌核糖体基因（rDNA）内转录间区ITS基因,经PCR扩增检验所提取的DNA质量。结果 成功提取所有23株真菌基因组DNA,其纯度及质量浓度能满足PCR反应的要求。结论 用溶细胞酶结合Biospin真菌基因组DNA提取试剂盒从酵母菌、无绿藻及丝状真菌提取的DNA可用于PCR反应。     

Measurement of mercury methylation in lake water and sediment samples

The production of various eremophilane-type sesquiterpenes by Penicillium roqueforti strains has allowed us to propose a biochemical pathway for PR toxin synthesis. A time-course study of P. roqueforti metabolite production by high-performance liquid chromatography was performed to check this hypothetical pathway. The results obtained suggested that eremofortin C was the direct precursor of PR toxin in the P. roqueforti cell. Attempts to determine the amount of PR toxin in the mycelium failed. It was shown that the absence of PR toxin in mycelium was due to its instability during the extraction procedure.