A DGGE-cloning method to characterize arbuscular mycorrhizal community structure in soil

Although arbuscular mycorrhizal fungi (AMF) are crucial for ecosystem functioning, characterizing AMF community structure in soil is challenging. In this study, nested polymerase chain reaction (PCR) and denaturing gradient gel electrophoresis (DGGE) were combined with cloning of fungal 18S ribosomal gene fragments for the rapid comparison of AMF community structure in soil. Reference AMF isolates, representing four major genera of AMF, were used to develop the method. Sequential amplification of 18S rDNA fragments by nested PCR using primer pairs AM1-NS31 and Glo1-NS31GC followed by DGGE analysis yielded a high-resolution band profile. In parallel, 18S rDNA fragment clone libraries were constructed and clones screened by DGGE. Sequence identity was inferred by matching the electrophoretic mobility of the sample fingerprint bands to that of bands from individual clones. The effectiveness of this approach was tested on soil samples from different ecosystems, yielding reproducible, complex DGGE band patterns specific to each site. The coupling of PCR–DGGE with clone library analysis provides a robust, reliable, and precise means to characterize AMF community structure in soils.     

Community structure of arbuscular mycorrhizal fungi associated with Robinia pseudoacacia in uncontaminated and heavy metal contaminated soils

The significance of arbuscular mycorrhizal fungi (AMF) in soil remediation has been widely recognized because of their ability to promote plant growth and increase phytoremediation efficiency in heavy metal (HM) polluted soils by improving plant nutrient absorption and by influencing the fate of the metals in the plant and soil. However, the symbiotic functions of AMF in remediation of polluted soils depend on plant–fungus–soil combinations and are greatly influenced by environmental conditions. To better understand the adaptation of plants and the related mycorrhizae to extreme environmental conditions, AMF colonization, spore density and community structure were analyzed in roots or rhizosphere soils of Robinia pseudoacacia. Mycorrhization was compared between uncontaminated soil and heavy metal contaminated soil from a lead–zinc mining region of northwest China. Samples were analyzed by restriction fragment length polymorphism (RFLP) screening with AMF-specific primers (NS31 and AM1), and sequencing of rRNA small subunit (SSU). The phylogenetic analysis revealed 28 AMF group types, including six AMF families: Glomeraceae, Claroideoglomeraceae, Diversisporaceae, Acaulosporaceae, Pacisporaceae, and Gigasporaceae. Of all AMF group types, six (21%) were detected based on spore samples alone, four (14%) based on root samples alone, and five (18%) based on samples from root, soil and spore. Glo9 (Rhizophagus intraradices), Glo17 (Funneliformis mosseae) and Acau3 (Acaulospora sp.) were the three most abundant AMF group types in the current study. Soil Pb and Zn concentrations, pH, organic matter content, and phosphorus levels all showed significant correlations with the AMF species compositions in root and soil samples. Overall, the uncontaminated sites had higher species diversity than sites with heavy metal contamination. The study highlights the effects of different soil chemical parameters on AMF colonization, spore density and community structure in contaminated and uncontaminated sites. The tolerant AMF species isolated and identified from this study have potential for application in phytoremediation of heavy metal contaminated areas.     

Species composition of arbuscular mycorrhizal fungi in two mountain meadows with differing management types and levels of plant biodiversity

Species composition of arbuscular mycorrhizal fungi (AMF) was analysed in two differently managed mountain grasslands in Thuringia (Germany). Arbuscular mycorrhizal fungi were studied in the roots of 18 dominant plant species from a total of 56 (32%). Additionally, spores of AMF were isolated from soil samples. Arbuscular mycorrhizal fungi species composition was analysed based on 96 sequences of the internal transcribed spacer of the nuclear ribosomal DNA, 72 originated from mycorrhizal roots, and 24 originated from AMF spores. Phylogenetic analyses revealed a total of 19 AMF species representing all genera of the Glomeromycota except Scutellospora and Pacispora. Despite a different farming intensity, resulting in remarkable differences concerning their plant species diversity (27 against 43 plant species), the diversity of AMF was found to be similar with 11 species on the intensively farmed meadow and ten species on the extensively farmed one. Nevertheless, species composition between both sites was clearly different. It thus seems likely that the AMF species composition, but not necessarily the species number, is related to above ground plant biodiversity in the system under study.     

Soil type and land use intensity determine the composition of arbuscular mycorrhizal fungal communities

The objective of this study was to test whether soil types can be characterized by their arbuscular mycorrhizal fungal (AMF) communities. To answer this question, a well-defined study area in the temperate climatic zone of Central Europe was chosen with a large spectrum of soils and parent materials. Representative soil samples were taken from three soil types (Cambisol, Fluvisol and Leptosol) at in total 16 sites differing in agricultural land use intensity (9 grasslands and 7 arable lands). AMF spores were isolated and morphologically identified directly from field soils and after reproduction in trap cultures. AMF diversity and community composition strongly depended on soil type and land use intensity, and several AMF species were characteristic for a specific soil type or a specific land use type and hence had a specific niche. In contrast, other AM fungi could be considered as ‘generalists’ as they were present in each soil type investigated, irrespective of land use intensity. An estimated 53% of the 61 observed AMF species could be classified as ‘specialists’ as (almost) exclusively found in specific soil types and/or under specific land use intensities; 28% appeared to be ‘generalists’ and 19% could not be classified. Plant species compositions (either natural or planted) had only a subordinate influence on the AMF communities. In conclusion, land use intensity and soil type strongly affected AMF community composition as well as the presence and prevalence of many AM fungi. Future work should examine how the differences in AMF species compositions affect important ecosystem processes in different soils and to which extent the loss of specific groups of AM fungi affect soil quality.     

丛枝菌根真菌对金叶六道木镉吸收及根际真菌群落结构的影响

为明确丛枝菌根(AM)真菌对促进绿化苗木镉(Cd)吸收的影响,通过盆栽试验比较接种不同AM真菌对12种绿化苗木Cd吸收的差异,并进一步分析接种对金叶六道木根际微生物数量和AM真菌群落结构的影响.结果表明,12种绿化苗木,加Cd处理6个月后,其叶片浓度的变化范围为0.25～2.59 mg·kg–1.接种AM真菌处理组的叶...     

长期保护性耕作对丛枝菌根真菌多样性的影响

为了明确我国北方干旱地区长期保护性耕作以及深松对丛枝菌根真菌(AMF)多样性的影响,笔者于2014年在山西省临汾市连续22年实施保护性耕作的长期定位试验基地,针对免耕覆盖(NTS)、深松免耕覆盖(SNTS)及传统耕作(TT)3种处理方式,进行了不同耕作条件下土壤AMF物种丰度、孢子密度、Shannon多样性指数以及AMF侵染率等因素的比较研究。结果显示,长期保护性耕作(NTS和SNTS)共分离鉴定出AMF 7属9种,其中根孢囊霉属(Rhizophagus)和斗管囊霉属(Funneliformis)各2种,球囊霉属(Glomus)、近明球囊霉属(Claroideoglomus)、无梗囊霉属(Acaulospora)、硬囊霉属(Sclerocystis)和隔球囊霉属(Septoglomus)各1种;而传统耕作(TT)共分离鉴定出AMF 6属8种,没有检测到无梗囊霉属。NTS、SNTS和TT处理在不同土层的AMF优势种基本一致,0~40 cm土层为摩西斗管囊霉(Fu.mosseae)和变形球囊霉(G.versiforme),40~80 cm土层为摩西斗管囊霉、变形球囊霉和聚丛根孢囊霉(Rh.aggregatum),80~120 cm土层为聚丛根孢囊霉,120 cm土层以下只有NTS和SNTS处理中存在聚丛根孢囊霉,说明保护性耕作措施促进了AMF向土壤深层发展。NTS和SNTS处理在同一土层的AMF物种丰度、孢子密度和Shannon多样性指数均高于TT处理,SNTS处理高于NTS处理。同一耕作措施不同土层的AMF物种丰度、孢子密度和Shannon多样性指数均随土层加深而逐渐降低;NTS和SNTS处理在小麦各生育期的丛枝侵染率和孢子密度均高于TT处理;各处理在小麦拔节期的AMF侵染率最高,分别为14.9%、16.1%和10.6%,而在收获期的土壤孢子密度最高,分别为111.7个·(100g)~(-1)、125.0个·(100g)~(-1)和90.3个·(100g)~(-1)。研究认为,长期免耕覆盖、尤其深松免耕覆盖,提高了AMF多样性。该研究结果可为中国北方旱作农田生态系统中AMF自然潜力的充分发挥,以及保护性耕作技术的合理应用提供科学依据。     

Comparison of commonly used primer sets for evaluating arbuscular mycorrhizal fungal communities: Is there a universal solution?

Different primer systems have been developed to characterize arbuscular mycorrhizal fungal (AMF) communities; however, a direct comparison of their specificity, potential to describe diversity and representation of different phylogenetic lineages is lacking. Using seven root samples, we compared four routinely used AMF-specific primer systems for nuclear ribosomal DNA covering i) the partial small subunit (SSU), ii) the partial large subunit (LSU), iii) the partial SSU and internal transcribed spacer (ITS; “Redecker”) and iv) the partial SSU–ITS–partial LSU region (“Krüger”). In addition, a new primer combination v) covering the ITS2 region (ITS2) was included in the comparison. The “Krüger” primers tended to yield the highest AMF diversity and showed a significantly higher Shannon diversity index than the SSU primers. We found a strong bias towards the Glomeraceae in the LSU and SSU primer systems and differences in the composition of AMF communities based on the “Redecker” primer system. Our results confirm the crucial role of the choice of target rRNA marker region for analysing AMF communities. We also provide evidence that nested-PCR based data can be interpreted semi-quantitatively and that the extent of observed AMF community overdominance largely depends on the choice of primer.     

Design and evaluation of nematode 18S rDNA primers for PCR and denaturing gradient gel electrophoresis (DGGE) of soil community DNA

Consensus nematode 18S ribosomal DNA primers were designed by aligning available 18S sequences and identifying a variable region flanked by highly conserved regions. These primers were then used to amplify nematode 18S rDNA from whole soil community DNA extracted from a range of European grassland types. Cloning of the PCR amplicons (778 bp) followed by restriction digest analysis (RFLP) resulted in the recovery of 34 unique nematode sequences from the four grasslands studied. Comparison of these data with the limited number of 18S rDNA nematode sequences currently held in on-line databases revealed that all of the sequences could be assigned to known nematode taxa albeit tentatively in some cases. Two of the sequences recovered from the site in the Netherlands (wet, hay-grassland) were recovered in a clade that included a sequence of the genus Trichodorus whilst other sequences from this site showed similarity with 18S rDNA sequences of the genus Prismatolaimus (five sequences), Xiphinema (one sequence) and Enoplus (one sequence). Of the remaining sequences, two showed some affinity with Mylonchulus (UK, upland peat), four with Steinernema (UK) and one sequence with Mesorhabditis (Hungary, east European Steppe). Three sequences from the Netherlands and one from Hungary were recovered in a clade that included a sequence of the genus Pratylenchoides whilst three further sequences from the Netherlands and two from Hungary were recovered in a clade encompassing the genus Globodera. Of the remaining nine sequences, two (NL6, NL62) formed a distinct lineage within the Adenophorea with 90% bootstrap recovery in a paraphyletic clade that included sequences of Prismatolaimus and Trichodorus. Seven sequences (three from the Netherlands, three from the UK and one from Greece) were left unassigned though the tree topology suggested some relationship (58% bootstrap recovery) with the genus Cephalobus. To assess whether primers used to amplify 18S rDNA might be used to fingerprint genetic diversity in nematode communities in soil, the environmental sequence data were used to design a second set of primers carrying a GC-clamp. These primers amplified a 469 bp fragment internal to the region flanked by the primer set used to derive the nematode trees and were used to amplify 18S rDNA for subsequent analysis using denaturing gradient gel electrophoresis (DGGE). DGGE analysis of six major European grassland types revealed considerable genetic diversity between sites. However, the relationships seen with the DGGE data were inconsistent with previous studies where the same soils had been characterized with respect to functional and morphological diversity. To confirm that this second set of primers was amplifying nematode sequences, selected bands on the DGGE gels were extracted, PCR amplified and sequenced. The final alignment was 337 bases. These analyses revealed the presence of sequence signatures from the genera Paratrichodorus, Plectus, Steinernema, Globodera, Cephalobus and Pratylenchoides.     

The influence of tillage on the structure of rhizosphere and root-associated arbuscular mycorrhizal fungal communities

Soil environmental factors affect the structure of arbuscular mycorrhizal (AM) fungal communities present in soil. However, it is not understood to which degree management practices such as tillage lead to dissimilarities between intra- and extraradical AM fungal communities. This study aims to assess the influence of two different soil management practices (conventional tillage and no-till) on the diversity of AMF communities, both in rhizosphere soil and inside corn roots. We hypothesized that under no-till, roots are colonized as they grow through the undisturbed fungal mycelia left from the previous crop whereas under conventional tillage they are colonized by those propagules that survived disturbance and can re-establish in their new relocated and mixed environment. We predicted that the degree of similarity of AM fungal communities inside versus outside the roots would be greater under no-till than under tillage. Using terminal restriction fragment length polymorphism (T-RFLP) and denaturing gradient gel electrophoresis we observed a different AM fungal community present in roots under no-till than under conventional tillage. Moreover, the communities present in the rhizosphere soil were different than in the roots of the corn plants. These results suggest that soil management does alter the diversity of AM fungal communities associated with corn roots and that plants influence the structure of the AMF community colonizing their roots. Sequencing results indicated that the majority of AMF species found in this agricultural soil was Glomus spp. However, further work is required to determine the extent to which AM fungal genotypic alterations by soil management influences competitive relationships.     

Comparisons of different hypervariable regions of rrs genes for fingerprinting of microbial communities in paddy soils

The use of molecular approaches based on 16S rDNA-PCR in microbial ecology has revealed a tremendous prokaryotic diversity in environmental samples. However, there is little or no systematic evaluation of the impacts of hypervariable (V) regions of rrs genes choice on microbial community analysis in soil samples, especially the detailed information about the dominant groups preferentially amplified by different primer pairs. In the present study, eight primer pairs were detected to compare the different V regions for fingerprinting microbial communities in a paddy soil irrigated with petroleum-wastewater, using denaturing gradient gel electrophoresis (DGGE) and amplified ribosomal DNA restriction analysis (ARDRA) techniques. Results reveal the obvious PCR bias produced by different V regions. Both ARDRA analysis of 16S rDNA clone library and DGGE suggest that V₁-V₃ region amplified with primer pair 8f-519r produced the most informative fingerprinting profiles. Additionally, V₃-V₅ region amplified with 341f-907r was another preferable choice for microbial diversity in petroleum-contaminated soil. The V₄-V₅ region and single V region (V₁, V₃, and V₈) were not recommended for the future study of microbial diversity in soil samples. Phylogenetic analysis of 123 sequences from libraries constructed by amplicons generated from six different V regions suggests that different dominant groups were amplified with distinct primer sets. In detail, V₁-V₃ library (amplified with 8f-519r) and V₃-V₅ library were dominated by Actinobacteria (20.4%) (particularly in genus Arthrobacter), V₁-V₃ library (amplified with 63f-518r) was dominated by γ-Proteobacteria (25.0%) and α-Proteobacteria (22.0%) (particularly in genus Brevundimonas), V₃ library was dominated by β-Proteobacteria (22.3%) (particularly in genus Gallionella) and α-Proteobacteria (20.0%), V₆-V₈ library was dominated by Chlamydiae (20.4%) and β-Proteobacteria (20.4%), V₈ library was dominated by γ-Proteobacteria (27.2%) (particularly in genus Acinetobacter) and β-Proteobacteria (14.0%). The present work strongly recommends that primer pairs should be chosen cautiously in community diversity analysis based on PCR amplification of 16S rDNA, and involving at least two different 16S rDNA universal primer pairs would perform better.     

Differential effect of arbuscular mycorrhizal fungal communities from ecosystems along management gradient on the growth of forest understorey plant species

Arbuscular mycorrhizal fungi (AMF) are important functional components of ecosystems. Although there is accumulating knowledge about AMF diversity in different ecosystems, the effect of forest management on diversity and functional characteristics of AMF communities has not been addressed. Here, we used soil inoculum representing three different AM fungal communities (from a young forest stand, an old forest stand and an arable field) in a greenhouse experiment to investigate their effect on the growth of three plant species with contrasting local distributions - Geum rivale, Trifolium pratense and Hypericum maculatum. AM fungal communities in plant roots were analysed using the terminal restriction fragment length polymorphism (T-RFLP) method. The effect of natural AMF communities from the old and young forest on the growth of studied plant species was similar. However, the AMF community from the contrasting arable ecosystems increased H. maculatum root and shoot biomass compared with forest inocula and T. pratense root biomass compared to sterile control. According to ordination analysis AMF inocula from old and young forest resulted in similar root AMF communities whilst plants grown with AM fungi from arable field hosted a different AMF community from those grown with old forest inocula. AMF richness in plant roots was not related to the origin of AMF inoculum. G. rivale hosted a significantly different AM fungal community to that of T. pratense and H. maculatum. We conclude that although the composition of AM fungal communities in intensively managed stands differed from that of old stands, the ecosystem can still offer the ‘symbiotic service’ necessary for the restoration of a characteristic old growth understorey plant community.     

广西沿海地区红树林根系土壤中放线菌的分离与鉴定

本研究通过分离纯培养,从广西北海及防城港红树林根系土壤中分离出放线菌并提取其总DNA,用放线菌通用引物对获得菌株的16SrDNA进行PCR扩增,对获得的扩增产物进行DNA序列测定及菌株鉴定。研究结果表明,从红树林根系土壤样品中分离出15株典型放线菌菌株。16SrDNA测序比对鉴定结果显示,15株典型放线菌菌株中有12株属于链霉菌属（Streptomyces）,是常见菌属;3株属于拟诺卡氏菌属（Nocardiopsis）,为稀有放线菌。本研究分离纯化获得15株典型放线菌,初步揭示了广西沿海地区红树林土壤中放线菌的多样性。     

磷肥对酸性红壤上玉米不同部位丛枝菌根真菌群落的影响

为了研究不同磷肥水平下酸性红壤上玉米不同部位丛枝菌根真菌(Arbuscular mycorrhizal fungi, AMF)群落多样性和组成结构,明确玉米不同部位AMF群落的分布特征及对磷肥的响应差别,为提高酸性红壤磷素利用提供理论依据。本实验设置三个磷肥水平：不施磷、低磷(25 mg P /kg)和高磷(100 mg P /kg),玉米培养4周后,测定玉米生物量和土壤理化性质,利用高通量测序技术检测玉米根部、根际和非根际土壤AMF群落结构和多样性。结果显示,随着磷肥水平增加,玉米生物量显著提高,高磷处理下玉米地上部磷含量显著高于不施磷和低磷处理。取样部位(根部、根际和非根际)显著影响了AMF群落优势属球囊霉属(Glomus)、巨孢囊霉属(Paraglomus)和近明球囊霉属(Claroideoglomus)相对丰度,但是磷肥影响不显著。类似的,取样部位而不是磷肥显著影响了AMF群落香农指数和物种丰富度。非度量多维标度(NMDS)结果显示,根部样品与非根际和根际土壤样品群落距离更远,而相同取样部位中不同磷肥水平间群落组成更为相似；置换多元方差分析(PERMANOVA)进一步表明,取样部位而不是磷肥显著影响了AMF群落组成结构,主要表现在根部样品与根际和非根际土壤不同。因此,酸性红壤上玉米不同部位对AMF群落的影响明显高于磷肥作用,表明AMF应用于酸性红壤时应重点考虑作物部位的特性。     

Arbuscular mycorrhizal fungi in degraded typical steppe of inner Mongolia

Arbuscular mycorrhizal (AM) fungi may have some potential use in the restoration of degraded grassland through beneficial effects on plant growth and soil quality. A field investigation was conducted in three grassland sites of typical steppe in Inner Mongolia. The three plant communities, one of which was undegraded, one moderately degraded and the third severely degraded, were studied by collecting soil samples and samples of four plant species that occurred in all three sites. The percentage of root length colonized by AM fungi was estimated and the species composition and diversity of AM fungus spores recovered from the soil were determined using spore morphological characteristics. Although differences between the sites may have been due partly to other factors, it is likely that the degree of degradation was an important factor. No decline was found in the AM colonization of the roots of the indicator plant species in the moderately or severely degraded plant communities, and two plant species showed higher colonization status in the two degraded areas. Glomus geosporum and Scutellospora calospora were the dominant AM fungi in the undegraded steppe, while G. geosporum and Glomus aggregatum dominated the two degraded sites which also had low spore densities, species richness and diversity indices. However, different AM species showed different distributions among the three plant communities and the results indicate that both biotic and abiotic factors were important in determining the AMF communities, with biotic factors possibly the more important. Copyright © 2008 John Wiley & Sons, Ltd.     

我国南方红壤酸化问题及改良修复技术研究进展

  目的  研究间作对作物根际土壤理化性质、球囊霉素相关土壤蛋白含量和AMF群落的影响,为我国南方红壤坡耕地种植模式的选择提供理论依据。  方法  以红壤坡耕地为研究对象,设置玉米单作、马铃薯单作、玉米与马铃薯间作3个处理,分析各作物在不同种植模式下其根际土壤理化性质、球囊霉素相关土壤蛋白含量、AMF群落的组成和多样性的变化,以及各指标之间的相互关系。  结果  玉米、马铃薯间作与单作相比均显著增加了两种作物根际土壤总球囊霉素和易提取球囊霉素含量,增加幅度为10.5% ~ 30.1%。玉米间作与玉米单作相比根际土壤有机质含量显著降低了10.6%,马铃薯间作与马铃薯单作相比根际土壤有机质和速效磷含量分别显著增加了13.5%和46.5%。玉米间作相比于玉米单作其根际土壤AMF群落特有的操作分类单元数量有增加趋势,而马铃薯间作相比马铃薯单作有降低趋势。非度量多维尺度法分析结果表明,种植模式较明显的改变了玉米根际土壤中AMF的群落组成,但没有明显改变马铃薯根际土壤中AMF的群落组成。玉米间作与玉米单作相比根际土壤AMF群落的Chao1指数显著增加了9.3%,香农指数有增加趋势,其值为1.3%,辛普森指数有降低趋势,其值为7.6%。马铃薯间作与马铃薯单作相比根际土壤AMF群落的辛普森指数有增加趋势,其值为53.3%,Chao1指数显著降低了10.8%,香农指数有降低趋势,其值为10.6%。在玉米、马铃薯的各处理中,根际土壤AMF群落优势属为巨孢囊霉属、无梗囊霉属和根孢霉属。冗余分析得出,pH是影响作物根际土壤AMF群落丰度、α多样性指数和球囊霉素相关土壤蛋白含量的主要因素。  结论  玉米与马铃薯间作可显著增加作物根际土壤的球囊霉素相关土壤蛋白含量,提高玉米根际土壤AMF群落组成的多样性,这一研究结果可为云南省红壤坡耕地种植模式的选择提供理论依据。     

Arbuscular mycorrhizal fungal diversity along a Tibetan elevation gradient

Rhizosphere samples were taken from herbaceous plants along an altitudinal gradient on Segrila Mountain slope. Root colonization and spore biodiversity of arbuscular mycorrhizal fungi (AMF) from different altitudes were analyzed. Of the 146 plant species representing 45 families investigated, 72.2% of plant species were colonized by AMF and formed typical AM structures. A broad range of AM fungal taxa, 62 taxa representing all 4 orders of AMF were isolated from the soil. The composition of the AMF spore community was quite different at different positions along the elevation gradient. Some AM fungi, such as Scutellospora, preferred some specific elevations, or a range of elevation. Intensity of root colonization (M%) and spore density were negatively correlated with the altitude of the study sites. Species richness showed a decreasing trend with increasing elevation but the Shannon–Weiner index was unaffected by elevation. Isolate frequency and relative abundance of AMF also showed quite different distribution patterns among taxonomic families. The drivers of these changes in the AM fungal assemblages is not known and cannot be determined conclusively using such a comparative study along an environmental gradient.     

Diversity of arbuscular mycorrhizal fungi along an environmental gradient in the Brazilian semiarid

Plants host, environmental characteristics and dispersal limitation are the main factors affecting the diversity of arbuscular mycorrhizal fungi (AMF) in global scale, whereas at the local scale other factors might also influence the composition of the AMF community. Therefore, the objective of this study was to assess the diversity of AMF and to relate it with the richness of plant species, season (dry or rainy) and soil parameters along an environmental gradient to know which are the drivers of the spatial distribution of AMF in the Brazilian semiarid. The gradient was composed of a dry forest (DF), a transitional zone (TZ) and a moist forest (MF). Ten soil samples were collected during the rainy (August 2011) and dry (February 2012) seasons in each site. The soil properties were determined, as well as AMF diversity, the latter based on morphological spore identification. There were significant differences between the DF and the other two areas in most soil chemical parameters, whereas the majority of soil attributes in the MF and TZ were similar. Altogether, 50 AMF species were identified, and the genera Acaulospora and Glomus were predominant. The AMF community structure in DF was significantly different from the other two areas by ordination (NMDS) and statistical method (PERMANOVA). However, the highest species diversity, based on the Shannon index, occurred in the TZ. The AMF community structure differed between seasons, with greater spore abundance in the dry season. Spatial AMF distribution was influenced by plants host, season, but the soil was the main factor. Four edaphic attributes showed approximately 60% of correlation with AMF community composition (Zn, Mg, base saturation and clay) based on the BIO-ENV analysis. We conclude that vegetation, seasonal variations and soil type affect the AMF diversity, and that the latter is a key factor for the similarity/dissimilarity of AMF communities between areas in the Brazilian semiarid.     

Phosphorus fertilisation management modifies the biodiversity of AM fungi in a tropical savanna forage system

In the present study we investigated how the community of arbuscular mycorrhizal fungi (AMF) in roots of Centrosema macrocarpum responded to different doses and sources of phosphorus (40 kg ha⁻¹ of P as rock phosphate, 150 kg ha⁻¹ of P as rock phosphate and 75 kg ha⁻¹ of P as diammonium phosphate together with 75 kg ha⁻¹ of P as rock phosphate) in a Venezuelan savanna ecosystem. We also related AMF diversity to soil parameters (total N, total P, available P, extractable K, total Ca, total Mg, total Fe, total Cu, total Zn, total Mn, glomalin-related soil protein, microbial biomass C, dehydrogenase, urease and acid phosphatase activities, water-soluble carbon and carbohydrates and aggregate stability) at different doses of P. The AM fungal small-subunit (SSU) rRNA genes were subjected to PCR, cloning, SSCP, sequencing and phylogenetic analyses. Nine fungal types were identified: six belonged to the genus Glomus and three to Acaulospora. The majority of fungal types showed high similarity to sequences of known glomalean isolates: Aca 1 to Acaulospora mellea, Aca 2 to Acaulospora rugosa, Aca 3 to Acaulospora spinosa, Glo 1 to Glomus intraradices and Glo 3 to Glomus fasciculatum. The control treatment was dominated by species belonging to the genus Acaulospora. However, when the soil was fertilised with low doses of P, the colonisation of roots increased and there was a change in the AMF diversity, the genus Glomus dominating. The AM development and the abundance of AM fungal types in roots were decreased dramatically by the fertilisation with high doses of P, without differences between the sources of P used. The available P in soil was negatively correlated with the AMF diversity. In conclusion, the application of low doses of P as rock phosphate stimulated mycorrhization and enhanced the soil quality parameters except water-soluble carbohydrates, helping to offset a loss of fertility in P-poor tropical savanna soils.     

间作小麦和接种AM真菌协同提高蚕豆抗枯萎病能力和根际微生物碳代谢活性

【目的】接种丛枝菌根 (arbuscular mycorrhizal,AM) 真菌和间作均是防治蚕豆枯萎病的有效方法,从土壤微生物学角度研究两者协同减轻蚕豆枯萎病的机理,对控制蚕豆枯萎病传播具有重要意义。【方法】利用盆栽试验方法,进行了间作和接种AM真菌摩西管柄囊霉 (Funneliformis mosseae,Fm) 和扭形球囊霉 (Glomus tortuosum,Gt) 试验。设蚕豆单作对照 (MF)、蚕豆小麦间作 (IF)、蚕豆单作接种Fm (MFFm)、蚕豆小麦间作接种Fm (IFFm)、蚕豆单作接种Gt (MFGt)、蚕豆小麦间作接种Gt (IFGt) 6个处理。于蚕豆开花期 (生长70天) 取土壤样品,测定蚕豆幼苗生长、枯萎病发生、根际镰刀菌数量和微生物碳代谢活性。【结果】间作显著增加蚕豆幼苗干重93.0%、降低蚕豆枯萎病病情指数71.4%,接菌显著增加蚕豆幼苗干重55.3%、降低病情指数76.6%,其中接种Fm真菌对蚕豆幼苗干重的影响更大,对病情指数的抑制效果更好。间作接菌显著增加蚕豆幼苗干重100%、降低病情指数89.8%。Biolog微平板测试结果显示,间作提高根际微生物碳代谢活性32.3%;接菌提高微生物活性85.4%;间作接菌提高微生物活性122%。主成分分析结果表明,间作和接菌均明显改变了根际微生物的群落结构,并主要改变了对碳水化合物类、氨基酸和羧酸类碳源的利用。相关性分析结果显示,枯萎病发病率和病情指数与根际镰刀菌数量呈极显著正相关关系,与AWCD值、Shannon多样性指数和丰富度指数均呈极显著负相关。【结论】蚕豆与小麦间作和接菌对抑制蚕豆枯萎病和促进蚕豆生长均具有积极效应,间作显著提高了AM真菌的定殖率,二者协同提高了根际微生物活性,改变了微生物群落结构,并抑制了病原菌增殖,进而控制蚕豆枯萎病发生。     

The application of an organic amendment modifies the arbuscular mycorrhizal fungal communities colonizing native seedlings grown in a heavy-metal-polluted soil

A mesocosm experiment was conducted to investigate whether communities of arbuscular mycorrhizal (AM) fungi associated with roots of native (Piptatherum miliaceum, Retama sphaerocarpa, Psoralea bituminosa, Coronilla juncea, and Anthyllis cytisoides) and for comparison (Lolium perenne) seedlings in a heavy-metal-contaminated, semiarid soil were affected by the application of composted sugar beet waste. We also investigated whether there were relation between AMF diversity and metal concentration (Al, Cd, Cu, Fe, Mn, Pb and Zn) and total P in shoot as well as some soil parameters (total organic carbon and total N) when the SB waste was added to the soil. We analyzed a portion of approximately 795 base pairs of the small-subunit (SSU) rRNA gene by nested PCR, cloning, sequencing, and phylogenetic analyses. Twelve different AMF sequence types were distinguished: seven of these belonged to Glomus group A, one to Glomus group B, one to Diversispora, one to Archaeospora, and two to Paraglomus. The AM fungal populations colonizing roots in a heavy-metal-polluted soil were quite dependent on the host plant, the highest diversity values being obtained in authochtonous plants recognized as metallophytes, such as P. bituminosa, and in an allochtonous, invasive species (L. perenne). No significant correlation was found between AMF diversity and plant metal concentration and soil parameters. Excepting P. bituminosa, when sugar beet waste was added to soil, the populations of AM fungi in roots increased and the shoot metal concentrations decreased in all host plant species studied. Therefore, the addition of sugar beet waste can be considered a good strategy for the remediation and/or phytostabilization of mine tailing sites.