Characterization of glomalin as a hyphal wall component of arbuscular mycorrhizal fungi

Arbuscular mycorrhizal fungi (AMF) produce a protein, glomalin, quantified operationally in soils as glomalin-related soil protein (GRSP). GRSP concentrations in soil can range as high as several mg g⁻¹ soil, and GRSP is highly positively correlated with aggregate water stability. Given that AMF are obligate biotrophs (i.e. depending on host cells for their C supply), it is difficult to explain why apparently large amounts of glomalin would be produced and secreted actively into the soil, since the carbon could not be directly recaptured by the mycelium (and benefits to the AMF via increased soil structure would be diffuse and indirect). This apparent contradiction could be resolved by learning more about the pathway of delivery of glomalin into soil; namely, does this occur via secretion, or is glomalin tightly bound in the fungal walls and only released after hyphae are being degraded by the soil microbial community? In order to address this question, we grew the AMF Glomus intraradices in in vitro cultures and studied the release of glomalin from the mycelium and the accumulation of glomalin in the culture medium. Numerous protein-solubilizing treatments to release glomalin from the fungal mycelium were unsuccessful (including detergents, acid, base, solvents, and chaotropic agents), and the degree of harshness required to release the compound (autoclaving, enzymatic digestion) is consistent with the hypothesis that glomalin is tightly bound in hyphal and spore walls. Further, about 80% of glomalin (by weight) produced by the fungus was contained in hyphae and spores compared to that released into the culture medium, strongly suggesting that glomalin arrives mainly in soil via release from hyphae, and not primarily through secretion. These results point research on functions of glomalin and GRSP in a new direction, focusing on the contributions this protein makes to the living mycelium, rather than its role once it is released into the soil.     

Microsite differences in fungal hyphal length, glomalin, and soil aggregate stability in semiarid Mediterranean steppes

Glomalin is a recently discovered glypoproteinaceous substance produced by arbuscular mycorrhizal fungi (AMF) that plays an important role in structuring soil. We quantified soil fungal hyphal length and glomalin content at vegetated and open microsites in Stipa tenacissima steppes of SE Spain. Soils underneath the canopy of S. tenacissima had higher glomalin pools compared to open microsites. We also found significant differences between sites, suggesting the presence of landscape level heterogeneity in glomalin concentration. Soil fungal hyphal length also differed significantly among the sites, but there was no significant effect of microsite. Water-stable aggregates (1-2 mm diameter; WSA_1-2 mm), however, while differing among sites, did not vary as a function of microsite. Furthermore, WSA_1-2 mm was negatively correlated with glomalin fractions, as well as soil organic C. Carbonates were likely the major binding agents in these carbonate-rich (average carbonate content: 71%) soils, and not organic C (including glomalin). AMF-mediated stabilization of soil aggregates did not contribute to the formation and maintenance of fertile islands underneath the canopy of S. tenacissima.     

土地利用方式对球囊霉素土层分布的影响

球囊霉素对维持土壤有机碳平衡和土壤团聚体稳定性具有明显作用,但不同土地利用方式下土壤球囊霉素的土层分布模式及其影响因素尚不清楚.本研究选取农田、人工草地、果园和撂荒地4种不同土地利用方式,分别采集0～10 cm、10～20 cm、20～30 cm和30～40 cm 4个土层土样,通过测定土壤球囊霉素.pH、速效磷、有机碳和蛋白酶活性,研究不同土地利用方式下土壤球囊霉素土层分布的模式及影响因素.结果表明,土壤球囊霉素平均含量为1.41～3.18 mg·g-1,占土壤有机碳的6.98%～31.34%,是土壤中的一个重要碳库.土壤球囊霉素在不同土地利用方式和土层剖面之间表现出显著差异(P<0.01),具有明显的垂直分布特征,除撂荒地外,其随土层深度的增加表现出降低趋势.土壤球囊霉素分别与土壤速效磷、蛋白酶呈显著正相关(P<0.01).土壤速效磷在很大程度上决定土壤球囊霉素的含量和分布.土壤球囊霉素含量和土壤蛋白酶活性之间为间接相关关系,该关系有待于进一步研究.建议把球囊霉素作为研究丛枝菌根真菌生长状况和土壤生态系统波动的一个重要指标.     

Spatial distribution of arbuscular mycorrhizal fungi, glomalin and soil enzymes under the canopy of Astragalus adsurgens Pall. in the Mu Us sandland, China

To measure and manage plant growth in arid and semi-arid sandlands, improved understanding of the spatial patterns of desert soil resources and the role of arbuscular mycorrhizal (AM) fungi is needed. Spatial patterns of AM fungi, glomalin and soil enzyme activities were investigated in five plots located in the Mu Us sandland, northwestern China. Soils to 50 cm depth in the rhizosphere of Astragalus adsurgens Pall. were sampled. The study demonstrated that A. adsurgens Pall. could form strong symbiotic relationships with AM fungi. Arbuscular mycorrhizal fungal status and distributions were significantly different among the five studied plots. Correlation coefficient analysis demonstrated that spore density was significantly and positively correlated with soil organic carbon (SOC), soil acid phosphatase and to two Bradford-reactive soil protein (BRSP) fractions (P < 0.01). Colonization of arbuscules and vesicles were positively correlated with protease activity. The BRSP fractions were also significantly and positively correlated to edaphic factors (e.g. SOC, available nitrogen, and Olsen phosphorus) and soil enzymes (e.g. soil urease and acid phosphatase). The means of total BRSP and easily extractable BRSP were 0.95 mg g⁻¹ and 0.5 mg g⁻¹ in all data, respectively. The levels of BRSP in the desert soil were little lower than those in native and arable soils, but the ratios of BRSP to SOC were much higher than farmland soils. The results of this study support the conclusion that glomalin could be an appropriate index related to the level of soil fertility, especially in desert soil. Moreover, AM fungal colonizations and glomalin might be useful to monitor desertification and soil degradation.     

Behavior of Bradford-reactive substances is consistent with predictions for glomalin

There is considerable controversy concerning detection in soils of the protein, glomalin, which is produced by arbuscular mycorrhizal fungi. Glomalin was originally defined as a substance that cross reacts with a monoclonal antibody formed against a substance in the cell walls of an arbuscular mycorrhizal fungus. Thus, one can use an immunological approach to detect glomalin. However, it was recently shown that other proteins cross react with the antibody. The other, more common, approach involves assay of soil protein using the Bradford reaction. This approach assumes that the Bradford assay is specific to protein, and that the assayed protein is largely glomalin, either because other proteins are in low concentration, or because the extraction process eliminates the possibility of their detection. These assumptions, however, have been called into question recently. One way to test whether the Bradford assay can be useful in quantifying glomalin is to determine whether the concentrations of Bradford-reactive substances are consistent with predictions for glomalin. For example, if recently produced glomalin is more labile than older glomalin, the concentrations of the two fractions should not be highly correlated. Moreover, when a contrast is established between mycorrhizal and nonmycorrhizal vegetation, recently produced glomalin should soon occur in higher concentration in soils supporting mycorrhizal vegetation. Older glomalin should not be found in higher concentrations in the soils of mycorrhizal vegetation until some time later. We tested these predictions by employing the Bradford assay during the course of a three-year field experiment in which canola (nonmycorrhizal) and soy (mycorrhizal) were grown in separate plots in year 1, both of which were followed by maize (mycorrhizal) in years 2 and 3. The correlation between the concentrations of fraction 1 Bradford-reactive substances (also known as easily extractable glomalin and frequently assumed to be recently produced) and fraction 2 (the more difficult-to-extract fraction and frequently assumed to be older glomalin), was very poor. In year 1, the concentration of fraction 1 was significantly greater in soy plots than in canola plots. Finally, fraction 2 was only significantly higher in the former soy plots than in former canola plots in years 2 and 3. These data support the hypothesis that the Bradford assay was useful in detecting glomalin in this case.     

Aggregate stability and glomalin in alternative crop rotations for the central Great Plains

 Land productivity, along with improvement or maintenance of soil health, must be evaluated together to achieve sustainable agricultural practices. Winter wheat-fallow (W-F) has been the prevalent cropping system in the central Great Plains for 60 years where moisture is a limitation to crop production. Alternative cropping systems show that producers can crop more frequently if residue management and minimum tillage are used. The impact of different crops, crop rotations and tillage management practices on soil quality was assessed by measuring aggregate stability and glomalin production by arbuscular mycorrhizal (AM) fungi in soil from cropping trials established in 1990. Crops were wheat (W), corn (C), proso millet (M), and sunflower (S). Rotations sampled were W-F, W-C-M, W-C-M-F, W-C-F, and W-S-F. In the same area as the cropping trials, soils were taken from a perennial grass (crested wheatgrass) and from a buffer area that had been planted to Triticale for the past 2 years but prior to that had been extensively plowed for weed control. We found that aggregate stability and glomalin were linearly correlated (r=0.73, n=54, P<0.001) across all treatments sampled. Highest and lowest aggregate stability and glomalin values were seen in perennial grass and Triticale soils, respectively. Aggregate stability in W-S-F was significantly lower than in the other crop rotations (P≤0.03), while W-C-M had significantly higher glomalin than the other rotations (P<0.05). Differences between crop rotations and the perennial grass indicate that selected comparisons should be studied in greater detail to determine ways to manage AM fungi to increase glomalin and aggregate stability in these soils. Received: 16 March 1999     

接种丛枝菌根真菌的种植紫花苜蓿土壤中球囊霉素含量与去除的关系

以菲和芘为多环芳烃（PAHs）代表物,以紫花苜蓿（Medicago sativa L.）为宿主植物,选用幼套球囊霉（Glomus etunicatum, Ge）、摩西球囊霉（Glomus mosseae,Gm）和层状球囊霉（Glomus lamellosum,Gla）3种丛枝菌根真菌（AMF）,研究了接种AMF下土壤中AMF菌丝密度、球囊霉素含量与PAHs去除率的关系。35~75 d,接种Ge、Gm、Gla处理的土壤中菌丝密度、总球囊霉素含量、易提取球囊霉素含量均随时间延长而显著增大,与不接种对照相比,75 d时接种Ge、Gm、Gla处理的土壤中易提取球囊霉素含量提高了48.58%、55.99%和50.23%,总球囊霉素含量则提高了38.75%、50.95%和46.12%。接种AMF促进了土壤中菲和芘的去除,随着时间（35~75 d）延长,接种Ge、Gm、Gla处理的土壤中菲去除率分别高达83.4%~92.7%、82.1%~93.8%、86.9%~93.4%,芘去除率达42.2%~63.5%、43.7%~69.2%、44.6%~66.4%。接种Ge、Gm和Gla处理土壤中AMF菌丝密度、总球囊霉素含量均与土壤中菲和芘的去除率之间存在极显著正相关关系,表明接种AMF提高了土壤中AMF菌丝密度和总球囊霉素含量,并促进了土壤中PAHs的去除。研究结果为阐明丛枝菌根修复PAHs污染土壤的规律及机理提供了依据。     

Carbon and nitrogen in operationally defined soil organic matter pools

Humic substances [humic acid (HA), fulvic acid (FA), and insoluble humin], particulate organic matter (POM), and glomalin comprise the majority (ca 75%) of operationally defined extractable soil organic matter (SOM). The purpose of this work was to compare amounts of carbon (C) and nitrogen (N) in HA, FA, POM, and glomalin pools in six undisturbed soils. POM, glomalin, HA, and FA in POM, and glomalin, HA, and FA in POM-free soil were extracted in the following sequence: (1) POM fraction separation from the soil, (2) glomalin extraction from the POM fraction and POM-free soil, and (3) co-extraction of HA and FA from the POM fraction and POM-free soil. Only trace amounts of HA and FA were present in the POM fraction, while POM-associated glomalin (POM-glomalin) and POM alone contributed 2 and 12%, respectively, of the total C in the soil. Mean combined weights for chemically extracted pools from POM and from POM-free soil were 9.92 g glomalin, 1.12 g HA, and 0.88 g FA kg⁻¹ soil. Total protein and C, N, and H concentrations showed that glomalin and HA were, for the most part, separate pools, although protein was detected in HA extracts. Even though percentage carbon was higher in HA than in glomalin, glomalin was a larger (almost nine times) operationally defined pool of soil organic C. Glomalin was also the largest pool of soil N of all the pools isolated, but all pools combined only contained 31% of the total N in the soil.     

Organic matter addition, N, and residue burning effects on infiltration, biological, and physical properties of an intensively tilled silt-loam soil

Seventy years of different management treatments have produced significant differences in runoff, erosion, and ponded infiltration rate in a winter wheat (Triticum aestivum L.)–summer fallow experiment in OR, USA. We tested the hypothesis that differences in infiltration are due to changes in soil structure related to treatment-induced biological changes. All plots received the same tillage (plow and summer rod-weeding). Manure (containing 111 kg N ha⁻¹), pea (Pisum sativum L.), vine (containing 34 kg N ha⁻¹), or N additions of 0, 45 and 90 kg ha⁻¹ were treatment variables with burning of residue as an additional factor within N-treatments. We measured soil organic C and N, water stability of whole soil, water stable aggregates, percolation through soil columns, glomalin, soil-aggregating basidiomycetes, earthworm populations, and dry sieve aggregate fractions. Infiltration was correlated (r = 0.67–0.95) to C, N, stability of whole soil, percolation, and glomalin. Basidiomycete extracellular carbohydrate assay values and earthworm populations did not follow soil C concentration, but appeared to be more sensitive to residue burning and to the addition of pea vine residue and manure. Dry sieve fractions were not well correlated to the other variables. Burning reduced (p < 0.05) water stability of whole soil, total glomalin, basidiomycetes, and earthworm counts. It also reduced dry aggregates of 0.5–2.0 mm size, but neither burning nor N fertilizer affected total C or total N or ponded infiltration rate. Water stability of whole soil and of 1–2-mm aggregates was greater at 45 kg N ha⁻¹ than in the 0 and 90 kg N ha⁻¹ treatments. Zero N fertilizer produced significantly greater 0.5–2.0 mm dry aggregate fractions. We conclude that differences in infiltration measured in the field are related to relatively small differences in aggregate stability, but not closely related to N or residue burning treatments. The lack of an effect of N fertilizer or residue burning on total C and N, along with the excellent correlation between glomalin and total C (r = 0.99) and total N (r = 0.98), indicates that the major pool of soil carbon may be dependent on arbuscular mycorrhizal fungi.     

Dynamics of arbuscular mycorrhizal fungi and glomalin in the rhizosphere of Artemisia ordosica Krasch. in Mu Us sandland, China

To understand the ecological significance of arbuscular mycorrhizal (AM) associations in semi-arid and arid lands, the temporal and spatial dynamics of AM fungi and glomalin were surveyed in Mu Us sandland, northwest China. Soil samples in the rhizosphere of Artemisia ordosica Krasch. were collected in May, July and October 2007, respectively. Arbuscular, hyphal and total root infection and spore density of AM fungi peaked in summer. The mean contents of total Bradford-reactive soil proteins (T-BRSPs, TG) and easily extractable Bradford-reactive soil proteins (EE-BRSPs, EEG) reached maximal values in spring. Spore density and two BRSPs fractions were the highest in the 0-10 cm soil layer, but the ratios of two BRSPs fractions to soil organic carbon (SOC) were the highest in the 30-50 cm soil layer. Hyphal infection was negatively correlated with soil enzymatic activity (soil urease and acid phosphatase) (P < 0.05). Arbuscular infection was negatively correlated with soil acid phosphatase (P < 0.01). Spore density was positively correlated with edaphic factors (soil available N, Olsen P, and SOC) and soil enzymatic activity (soil acid and alkaline phosphatase) (P < 0.01). Two BRSPs fractions were positively correlated with edaphic factors (soil available N and SOC) and soil enzymatic activity (soil urease, acid and alkaline phosphatase) (P < 0.01). TG was positively correlated with soil Olsen P (P < 0.05). We concluded that the dynamics of AM fungi and glomalin have highly temporal and depth patterns, and influenced by nutrient availability and enzymatic activity in Mu Us sandland, and suggest that glomalin are useful indicators for evaluating soil quality and function of desert ecosystem on the basis of its relationship to AM fungal community, soil nutrient dynamics and carbon cycle.     

Glomalin extraction and measurement

We investigated extraction from soil of glomalin, a glycoprotein produced by arbuscular mycorrhizal fungi, and we examined its measurement. The most commonly used protocols for extracting glomalin require autoclaving of soil in citrate solution, followed by centrifugation to separate the supernatant, and then measurement by either Bradford protein assay or enzyme-linked immunosorbent assay (ELISA). We found that lengthening the time of autoclaving increased easily extractable glomalin extraction. Delay of centrifugation after autoclaving, however, diminished Bradford-reactive substances in the supernatant, suggesting that extracted substances might be reversibly immobilized on soil particles. Surprisingly, increasing the volume of extraction solution did not accelerate extraction of “total glomalin”, but instead, substantially increased the amount extracted. Multiple autoclave cycles nevertheless denature glomalin, which may not be as heat-resistant as thought. Repeated 1-h autoclaving of supernatant diminished both its Bradford-reactive substances (7.3% h^?1) and immunoreactive protein (22% during the first hour and 9.5% h^?1 of the remainder thereafter), although a large initial volume of extractant could reduce the loss of immunoreactive protein. Proteins and polyphenols that survive the extraction process are measured non-specifically by the Bradford assay. When we added other glycoproteins to dry soils, we recovered a maximum 34% bovine serum albumin and 22% bovine mucin, primarily in the first two, 1-h extraction cycles. These added proteins may adhere to soil organic matter and thereby be protected from denaturation. In addressing the endpoint of glomalin extraction, we found that the Michaelis–Menten equation closely fits cumulative glomalin per extraction cycle such that its asymptote provides an objective estimate of total extractable glomalin for a given set of extraction conditions. Additionally, the equation provides a curvature parameter that reflects the soil-specific efficiency of an extraction protocol. Although the soils that we investigated with 7.6% or more soil organic matter had the most asymptotic total glomalin, they were extracted the least efficiently.     

长期稻草还田对土壤球囊霉素和土壤C、N的影响

A long-term experiment was conducted to investigate how long-term fertilization and rice straw incorporation into soil affect soil glomalin, C and N. The combined application of chemical fertilizer and straw resulted in a significant increase in both soil easily extractable glomalin （EEG） and total glomalin （TG） concentrations, as compared with application of only chemical fertilizer or no fertilizer application. The EEG and TG concentrations of the NPKS （nitrogen, phosphorus, and potassium fertilizer application ＋ rice straw return） plot were 4.68% and 5.67% higher than those of the CK （unfertilized control） plot, and 9.87% and 6.23% higher than those of the NPK （nitrogen, phosphorus, and potassium fertilizer applied annually） plot, respectively. Application of only chemical fertilizer did not cause a statistically significant change of soil glomalin compared with no fertilizer application. The changes of soil organic C （SOC） and total N （TN） contents demonstrated a similar trend to soil glomalin in these plots. The SOC and TN contents of NPKS plot were 15.01% and 9.18% higher than those of the CK plot, and 8.85% and 14.76% higher than those of the NPK plot, respectively. Rice straw return also enhanced the contents of microbial biomass C （MBC） and microbial biomass N （MBN） in the NPKS plot by 7.76% for MBC and 31.42% for MBN compared with the CK plot, and 12.66% for MBC and 15.07% for MBN compared with the NPK plots, respectively. Application of only chemical fertilizer, however, increased MBN concentration, but decreased MBC concentration in soil.     

Arbuscular mycorrhizal fungi in the Brazilian Atlantic forest: A gradient of environmental restoration

Arbuscular mycorrhizal fungi (AMF) are of great importance for the successful regeneration of degraded natural areas. The objective of this study was to examine how the time of environmental recuperation is affecting the occurrence and diversity of AMF species in riparian areas belonging to the Atlantic Forest biome in the State of São Paulo, Brazil. The study involved a native forest area (NT) and a gradient of environmental restoration: five (R05), ten (R10), and twenty (R20) years after reforestation. Soil samples were collected in the rainy (January) and dry season (June). Chemical, physical and microbiological analyses were performed including the amount of glomalin and quantification of AMF spores. The frequency of occurrence of genera and ecological indices, as richness (R), Shannon's diversity (H) and Simpson's dominance index (Is) were calculated. The largest spore number was found in R05 and the highest richness and diversity indices of AMF species in NT. Considering the two sampling periods and the four areas studied, we found 22 AMF species, and the genera Glomus and Acaulospora were the most frequent. A Canonical Discriminant Analysis showed that Glomus viscosum, Acaulospora scrobiculata, Acaulospora mellea and Scutellospora heterogama were the species that contributed the most to distinguishing the areas. Moisture, density and glomalin were positively correlated with the number of spores, however, soil nitrate showed a negative correlation. This work gives a better understanding of the interactions between AMF and forest soils and allows to know the distribution of AMF species according to environmental recovery time.     

Extraction of Glomalin and Associated Compounds with Two Chemical Solutions in Cultivated Tepetates of Mexico

Glomalin is a glycoprotein produced by the hyphae of arbuscular mycorrhizal fungi (AMF). The chemical methods usually employed to extract glomalin from the soil obtain something more than this pure glycoprotein, and therefore it would be better to call this fraction soil protein related to glomalin (SPRG) or glomalin associated with humic substances (GAHS). On this account, its isolation is controversial. The SPGR or GAHS has a significant influence on the physical, chemical, and biological properties of soils and could then be considered as an indicator of soil use change. In the present study, the storage of SPRG was evaluated, as well as carbon (C) associated with the latter (CG) and the content of soil organic C (SOC) in cultivated tepetates in the State of Mexico. Tepetates are hardened volcanic tuffs of the fragipan type, ameliorated for agricultural production. The specific objectives of the present study were (1) to evaluate the SPRG levels in tepetates, (2) measure the contribution of carbon (C) made by the SPRG to soil organic C (SOC), and (3) compare the extraction of SPRG with sodium pyrophosphate and sodium citrate. The samples used in this experiment came from 87 tepetate-cultivated plots (0–20 cm) located in the Texcoco River basin, State of Mexico. The levels of SPRG were observed among traces, 2.3 mg g^–1 for citrate and up to 5.6 mg g^–1 for pyrophosphate. The latter reactive allowed us to extract nearly three times more SPRG and two times more C-SPRG than sodium citrate (P?=?0.05) in the tepetates having about 4% of SOC; yet when the latter was less than 0.5%, the extraction levels with both solutions were similar. The SPRG and CG were closely correlated with SOC (r > 0.90). Tepetates have levels of SPRG similar to those observed in arid soils.     

Response of Arbuscular Mycorrhizal Fungi in Different Soil Tillage Systems to Long‐Term Swine Slurry Application

Swine slurry is a common agricultural fertilizer in many countries. However, its long‐term use in large amounts can cause excess nutrient accumulation, alter soil compounds, and potentially influence critical microbial populations such as arbuscular mycorrhizal fungi, which have important roles in plant nutrition and soil sustainability. This work determined if arbuscular mycorrhizal status, external mycelium, and glomalin‐related soil protein content were affected by long‐term swine slurry application to different soil tillage systems. The experiment was conducted on a clayey oxisol, in southern Brazil. Swine slurry (0, 30, 60, 90, and 120 m³ ha⁻¹ y⁻¹) was applied for 15 years to conventional tillage and no tillage soil prior to the summer (soybean or maize) and winter (wheat or oats) crop seasons. Swine slurry decreased mycorrhizal root colonization, spore number, and total external mycelium. Swine slurry increased active external mycelium and both easily extractable and total glomalin‐related soil protein. No‐tillage soil had more glomalin‐related soil protein than conventional tillage soil. The most significant response variables were root colonization, easily extractable glomalin‐related soil protein, and total external arbuscular mycorrhizal mycelia. Long‐term application of swine slurry in this environment, even at high rates, did not adversely affect crop yield but did influence arbuscular mycorrhizae fungi and their products in the soil environment. Benefits of swine slurry application for crop nutrition must be weighed against potential adverse consequences for the size, activity, and benefits of the mycorrhizal community to subsequent annual crops. Copyright © 2014 John Wiley & Sons, Ltd.     

Elevated CO2 increases the effect of an arbuscular mycorrhizal fungus and a plant-growth-promoting rhizobacterium on structural stability of a semiarid agricultural soil under drought conditions

In arid and semiarid Mediterranean regions, an increase in the severity of drought events could be caused by rising atmospheric CO₂ concentrations. We studied the effects of the interaction of CO₂, water supply and inoculation with a plant-growth-promoting rhizobacterium (PGPR), Pseudomonas mendocina Palleroni, or inoculation with an arbuscular mycorrhizal (AM) fungus, Glomus intraradices (Schenk & Smith), on aggregate stabilisation of the rhizosphere soil of Lactuca sativa L. cv. Tafalla. The influence of such structural improvements on the growth of lettuce was evaluated. We hypothesised that elevated atmospheric CO₂ concentration would increase the beneficial effects of inoculation with a PGPR or an AM fungus on the aggregate stability of the rhizosphere soil of lettuce plants. Leaf hydration, shoot dry biomass and mycorrhizal colonisation were decreased significantly under water-stress conditions, but this decrease was more pronounced under ambient vs elevated CO₂. The root biomass decreased under elevated CO₂ but only in non-stressed plants. Under elevated CO₂, the microbial biomass C of the rhizosphere of the G. intraradices-colonised plants increased with water stress. Bacterial and mycorrhizal inoculation and CO₂ had no significant effect on the easily-extractable glomalin concentration. Plants grown under elevated CO₂ had a significantly higher percentage of stable aggregates under drought stress than under well-watered conditions, particularly the plants inoculated with either of the assayed microbial inocula (about 20% higher than the control soil). We conclude that the contribution of mycorrhizal fungi and PGPR to soil aggregate stability under elevated atmospheric CO₂ is largely enhanced by soil drying.     

Arbuscular mycorrhizal fungi and soil aggregation in a no‐tillage system with crop rotation

Crop rotation adoption in no‐tillage systems (NTS) has been recommended to increase the biological activity and soil aggregation, suppress soil and plant pathogens, and increase the productivity aiming at the sustainability of agricultural areas. In this context, this study aimed to assess the effect of crop rotation on the arbuscular mycorrhizal fungi (AMF) community and soil aggregation in a soil cultivated for nine years under NTS. Treatments consisted of combinations of three summer crop sequences and seven winter crops. Summer crop sequences consisted of corn (Zea mays L.) monoculture, soybean (Glycine max L. Merrill) monoculture, and soybean–corn rotation. Winter crops consisted of corn, sorghum (Sorghum bicolor (L.) Moench), sunflower (Helianthus annuus L.), sunn hemp (Crotalaria juncea L.), pigeon pea (Cajanus cajan (L.) Millsp.), oilseed radish (Raphanus sativus L.), and millet (Pennisetum americanum (L.) Leeke). Soil samples were collected at a depth of 0–0.10 m for analyses of soil chemical, physical, and biological attributes. Spore abundance, total glomalin, and soil aggregate stability index were higher in the soil under corn monoculture. The highest values of aggregate mean weight diameter were observed in the soybean–corn rotation (3.78 mm) and corn monoculture (3.70 mm), both differing from soybean monoculture (3.15 mm), while winter crops showed significant differences only between sorghum (3.96 mm) and pigeon pea (3.25 mm). Two processes were identified in the soil under summer crop sequences. The first process was observed in PC1 (spore abundance, total glomalin, easily extractable glomalin, pH, P, and Mg²⁺) and was related to AMF; the second process occurred in PC2 (aggregate mean weight diameter, soil aggregate stability index, K⁺, and organic matter) and was related to soil aggregation. The nine‐year no‐tillage system under the same crop rotation adoption influenced AMF abundance in the soil, especially with corn cultivation in the summer crop sequence, which promoted an increased total external mycelium length and number of spores of AMF. In addition, it favored an increased soil organic matter content, which is directly related to the formation and stability of soil aggregates in these managements.     

Tannic acid reduces recovery of water-soluble carbon and nitrogen from soil and affects the composition of Bradford-reactive soil protein

Tannins are plant-derived polyphenolic compounds that precipitate proteins, bind to metals and complex with other compounds. Solutions of tannic acid, or other phenolic compounds, were added to soil samples to determine if they would affect recovery of soluble soil carbon (WSC) or –nitrogen (WSN) or influence the extraction and composition of Bradford-reactive soil protein (BRSP), associated with glomalin. Tannic acid-C added with water was not completely recovered from samples and the amount of total net WSC and WSN recovered was reduced, suggesting formation of insoluble complexes. By comparison, non-tannin phenolics like gallic acid, or methyl gallate, had little effect on extraction of WSC or WSN while a simple gallotannin derived from tannic acid, 1,2,3,4,6-penta-O-galloyl-d-glucose (PGG), inhibited extraction most. The C and N concentrations in BRSP increased when soil samples were treated with tannic acid or PGG before extraction, a procedure that includes autoclaving. Increases were greatest in the 10–20 cm compared to 0–5 cm depth. Accompanying these were declines in the ratio of absorbance at 465 and 665 nm (E4/E6 ratio) of BRSP extracts suggesting formation of larger or heavier molecules. In contrast, C and N composition in lyophilized BRSP was unaffected or even slightly reduced when tannic acid or PGG were added to the BRSP extract solution after the extraction process. We conclude that some tannins can reduce the solubility of labile soil C and N, at least temporarily and given unpredictability of response associated with phenolic substances, the Bradford assay should not be relied on to quantify pools or composition of soil proteins like glomalin.     

Survival of inocula and native AM fungi species associated with shrubs in a degraded Mediterranean ecosystem

Reconstitution of the potential of soil mycorrhizal inoculum is a key step in revegetation programs for semiarid environments. We tested the effectiveness of inoculation with native arbuscular mycorrhizal (AM) fungi or with an allochthonous AM fungus, Glomus claroideum, with respect to the growth of four shrub species, the release of mycorrhizal propagules in soil, within and outside the canopy, and the improvement of soil structural stability. Two years after outplanting, the mixture of native endophytes was more effective than, for Olea europaea subsp. sylvestris, Retama sphaerocarpa and Rhamnus lycioides, or equally as effective as, for Pistacia lentiscus, the non-native AM fungus Glomus claroideum, with respect to increasing shoot biomass and foliar NPK contents. The increases in glomalin concentration and structural stability produced by inoculation treatments in the rhizosphere soil of the all shrub species, except R. lycioides, ranged from about 55 to 173% and 13 to 21%, respectively. The mixture of native AM fungi produced the highest levels of mycorrhizal propagules in soil from the center of the canopy of P. lentiscus and R. lycioides, while plants of O. europaea and R. sphaerocarpa inoculated with G. claroideum had more mycorrhizal propagules than did those inoculated with the mixture of native fungi. The number of mycorrhizal propagules in soil outside the canopy of the four shrub species was 5-35 times higher in inoculation treatments than in soil of the non-inoculated plants.     

Differential decomposition of arbuscular mycorrhizal fungal hyphae and glomalin

Arbuscular mycorrhizal fungi (AMF) are obligate symbionts of most higher plants. In addition to being a major component of soil microbial biomass, AMF hyphae produce glomalin, a recalcitrant glycoproteinaceous substance highly correlated with soil aggregate water stability. This study addresses the lack of knowledge concerning the decomposition of hyphae and glomalin. We used an experimental design that exploited the lack of saprobic capabilities of AMF hyphae by incubating field soil samples in the dark, and hence in the absence of plant or AMF hyphal growth. In 150 days, hyphal length decreased 60%, while glomalin, quantified by the Bradford protein assay, declined only 25%. Immuno-reactive glomalin decreased 46%. This study serves as a proof-of-concept for further examination of factors that influence decomposition of AMF hyphae using similar experimental designs.