Improved rock phosphate dissolution from organic acids is driven by nitrate assimilation of bacteria isolated from nitrate and CaCO3-rich soil.

Until now, the solubilization capacities of insoluble mineral P by soil microorganisms have been screened in vitro with media containing NH4+ as a nitrogen source. This presence of NH4+ will lead to an acidification of the medium responsible for the solubilization of the insoluble P. However, besides proton release, the production of organic acids can play a very important role in the release of free P. This physiological mechanism can largely depend on the source of nitrogen (NH4+vs NO3-) assimilated by the bacteria but the influence of the N source on the production of organic acids has yet to be studied. Our aim was to investigate if the N source assimilated by bacteria and the soil characteristics such as the dominant N source (NH4+vs NO3-) and CaCO3 contents might influence the bacterial capacities to solubilize rock phosphate. To fill this objective, we screened the capacity of bacteria isolated from 3 soils to solubilize rock phosphate in vitro in presence of NH4+or NO3-. Then, we selected the most efficient bacterial strains to identify and quantify the release of organic anions into the medium. Among the two hundred and forty-three bacterial strains isolated from the 3 soils, nine and seven isolates were identified with the highest % rock phosphate-solubilization values with NH4+ or NO3- as the sole N-source. Only one strain was able to release free Pi with NH4+ or NO3- as the sole N-source. The most predominant organic acids released by almost all isolates were gluconic acid, lactic acid, glycolic acid, acetic acid, formic acid and pyruvic acid regardless the N-source. However, with NO3- as source of N, the highest concentrations on those acids were found together with the highest release of free Pi into the medium. Molecular analysis of 16S rRNA indicated that almost all strains belonged to Bacillus and Paenibacillus genera. The PCA analysis between soil properties and bacterial capacities to release organic acids and free Pi also revealed that soil factors such as CaCO3 and soil NO3- content positively influenced the release of organic acids by bacteria grown in vitro. Our results concluded that the bacterial rock phosphate-solubilization was intimately related to organic acids production which in turn seemed to be driven by the assimilation of NO3- by bacteria. Therefore, the N-source might be considered a key factor to take into consideration during the screening and selection of suitable strains involved in the P-solubilization.

Application of dry olive residue-based biochar in combination with arbuscular mycorrhizal fungi enhances the microbial status of metal contaminated soils.

Biochar made-up of dry olive residue (DOR), a biomass resulting from the olive oil extraction industry, has been proposed to be used as a reclamation agent for the recovery of metal contaminated soils. The aim of the present study was to investigate whether the soil application of DOR-based biochar alone or in combination with arbuscular mycorrhizal fungi (AMF) leads to an enhancement in the functionality and abundance of microbial communities inhabiting metal contaminated soils. To study that, a greenhouse microcosm experiment was carried out, where the effect of the factors (i) soil application of DOR-based biochar, (ii) biochar pyrolysis temperature (considering the variants 350 and 500 °C), (iii) soil application dose of biochar (2 and 5%), (iv) soil contamination level (slightly, moderately and highly polluted), (v) soil treatment time (30, 60 and 90 days) and (vi) soil inoculation with Funneliformis mosseae (AM fungus) on ß-glucosidase and dehydrogenase activities, FA (fatty acid)-based abundance of soil microbial communities, soil glomalin content and AMF root colonization rates of the wheat plants growing in each microcosm were evaluated. Biochar soil amendment did not stimulate enzyme activities but increased microbial abundances. Dehydrogenase activity and microbial abundances were found to be higher in less contaminated soils and at shorter treatment times. Biochar pyrolysis temperature and application dose differently affected enzyme activities, but while the first factor did not have a significant effect on glucosidase and dehydrogenase, a higher biochar dose resulted in boosted microbial abundances. Soil inoculation with F. mosseae favored the proliferation of soil AMF community and increased soil glomalin content as well as rates of AMF root colonization. This factor also interacted with many of the others evaluated to significantly affect soil enzyme activities, microbial abundances and AMF community. Our results indicate that the application of DOR-based biochar along with AMF fungi is an appropriate approach to improve the status of microbial communities in soils with a moderate metal contamination at short-term.

Special Issue "Microorganisms in Recycling and Valorization of Organic Waste for Sustainable Soil Health and Management".

Anthropogenic activity generates huge amounts of solid organic wastes [...].

Studying Microbial Communities through Co-Occurrence Network Analyses during Processes of Waste Treatment and in Organically Amended Soils: A Review.

Organic wastes have the potential to be used as soil organic amendments after undergoing a process of stabilization such as composting or as a resource of renewable energy by anaerobic digestion (AD). Both composting and AD are well-known, eco-friendly approaches to eliminate and recycle massive amounts of wastes. Likewise, the application of compost amendments and digestate (the by-product resulting from AD) has been proposed as an effective way of improving soil fertility. The study of microbial communities involved in these waste treatment processes, as well as in organically amended soils, is key in promoting waste resource efficiency and deciphering the features that characterize microbial communities under improved soil fertility conditions. To move beyond the classical analyses of metataxonomic data, the application of co-occurrence network approaches has shown to be useful to gain insights into the interactions among the members of a microbial community, to identify its keystone members and modelling the environmental factors that drive microbial network patterns. Here, we provide an overview of essential concepts for the interpretation and construction of co-occurrence networks and review the features of microbial co-occurrence networks during the processes of composting and AD and following the application of the respective end products (compost and digestate) into soil.

The grazing activity of Acrobeloides sp. drives phytate mineralisation within its trophic relationship with bacteria.

The microbial loop has been suggested as an alternative route for better utilization of phytate, a poorly available P source to plants. We hypothesized that bacterial grazer activity might dramatically enhance bacterial migration and proliferation, increasing the probability of phytate hydrolysis by bacterial phytases and, thus, phytate mineralization and release of free phosphate. We tested this hypothesis in a two-compartment system with a solid medium containing phytate or free phosphate as the source of P. Two bacterial species, B. subtilis 168 or Bradyrhizobium sp., with or without bacterial grazing nematodes belonging to Acrobeloides sp. previously fed on each of the bacterial species, were inoculated at a single point in the medium. Whatever the P source, nematode migration within both zones allowed the proliferation of bacteria. However, B. subtilis 168 was more efficient in using phytate than Bradyrhizobium sp. since the highest bacterial cell density and free phosphate concentrations were reached by Acrobeloides sp. fed on B. subtilis 168. The grazer activity seemed to be crucial to enhance phytate mineralization, despite Acrobeloides sp. showing a higher preference to feed on Bradyrhizobium sp. This study provides new insights into the effects of bacterial grazer activity on phytate mineralization.

Implications of mycoremediated dry olive residue application and arbuscular mycorrhizal fungi inoculation on the microbial community composition and functionality in a metal-polluted soil.

Metal-polluted soils represent hostile environments affecting the composition and functions of soil microbial communities. This study evaluated the implication of combining the mycoremediated dry olive residue (MDOR) amendment application with the inoculation of the arbuscular mycorrhizal fungi (AMF) Funneliformis mosseae in restoring the quality, composition, and functionality of soil microbial communities. To achieve this aim, a mesocosms experiment was set up that included three variations: i) with and without application of Penicillium chrysogenum-10-transformed MDOR (MDOR_Pc), and Chondrosterum purpureum-transformed MDOR (MDOR_Cp) amendments; ii) with and without F. mosseae inoculation; and iii) 30-day and 60-day soil treatment time. As a result of this combined treatment, changes in the soil labile organic C and N fractions were observed throughout the experiment. Increases in the abundance of phospholipid fatty acids (PLFAs) for bacteria, actinobacteria, and Gram- and Gram+ bacteria were also recorded at the end of the experiment. The addition of MDOR amendments boosted fungal and AM fungi communities. AM fungi root and soil colonization was also enhanced as the result of improvement nutrient turnover and spatial conditions caused by adding MDOR in combination with an inoculation of F. mosseae. The composition and functionality of microbial communities seemed to be an important ecological attribute indicating an apparently fully functional restoration of this metal-polluted soil and therefore suggesting the suitability of the combined MDOR and AM fungus treatment as a reclamation practice.

A comparative study to evaluate natural attenuation, mycoaugmentation, phytoremediation, and microbial-assisted phytoremediation strategies for the bioremediation of an aged PAH-polluted soil.

Biological treatments are considered an environmentally option to clean-up polluted soil with polycyclic aromatic hydrocarbons (PAHs). A pot experiment was conducted to comparatively evaluate four different strategies, including natural attenuation (NA), mycoaugmentation (M) by using Crucibulum leave, phytoremediation (P) using maize plants, and microbial-assisted phytoremediation (MAP) for the bioremediation of an aged PAH-polluted soil at 180 days. The P treatment had higher affinity degrading 2-3 and 4 ring compounds than NA and M treatments, respectively. However, M and P treatments were more efficient in regards to naphthalene, indeno[l,2,3-c,d]pyrene and benzo[g,h,i]perylene degradation respect to NA. However, 4, 5-6 rings undergo a strong decline during the microbe-assisted phytoremediation, being the treatment which determined the highest rates of PAHs degradation. Sixteen PAH compounds, except fluorene and dibenzo[a,h]anthracene, were found in maize roots, whereas the naphthalene, phenanthrene, anthracene, fluoranthene, and pyrene were accumulated in the shoots, in both P and MAP treatments. However, higher PAH content in maize biomass was achieved during the MAP treatment respect to P treatment. The bioconversion and translocation factors were less than 1, indicating that phystabilization/phytodegradation processes occurred rather than phytoextraction. The microbial biomass, activity and ergosterol content were significantly boosted in the MAP treatment respect to the other treatments at 180 days. Ours results demonstrated that maize-C. laeve association was the most profitable technique for the treatment of an aged PAH-polluted soil when compared to other bioremediation approaches.

Risk element immobilization/stabilization potential of fungal-transformed dry olive residue and arbuscular mycorrhizal fungi application in contaminated soils.

The use of biotransformed dry olive residue (DOR) as organic soil amendment has recently been proposed due to its high contents of stabilized organic matter and nutrients. The potential of biotransformed DOR to immobilize risk elements in contaminated soils might qualify DOR as a potential risk element stabilization agent for in situ soil reclamation practices. In this experiment, the mobility of risk elements in response to Penicillium chrysogenum-10-transformed DOR, Funalia floccosa-transformed DOR, Bjerkandera adusta-transformed DOR, and Chondrostereum purpureum-transformed DOR as well as arbuscular mycorrhizal fungi (AMF), Funneliformis mosseae, inoculation was investigated. We evaluated the effect of these treatments on risk element uptake by wheat (Triticum aestivum L.) plants in a pot experiment with Cd, Pb, and Zn contaminated soil. The results showed a significant impact of the combined treatment (biotransformed DOR and AMF inoculation) on wheat plant growth and element mobility. The mobile proportions of elements in the treated soils were related to soil pH; with increasing pH levels, Cd, Cu, Fe, Mn, P, Pb, and Zn mobility decreased significantly (r values between -0.36 and -0.46), while Ca and Mg mobility increased (r = 0.63, and r = 0.51, respectively). The application of biotransformed DOR decreased risk element levels (Cd, Zn), and nutrient concentrations (Ca, Cu, Fe, Mg, Mn) in the aboveground biomass, where the elements were retained in the roots. Thus, biotransformed DOR in combination with AMF resulted in a higher capacity of wheat plants to grow under detrimental conditions, being able to accumulate high amounts of risk elements in the roots. However, risk element reduction was insufficient for safe crop production in the extremely contaminated soil.

Stabilization of different starting materials through vermicomposting in a continuous-feeding system: Changes in chemical and biological parameters.

In this study the feasibility of Eisenia andrei to digest great amount of wastes including horse manure (HM), apple pomace (AP), grape pomace (GP), and digestate (DG) was monitored through a continuous-feeding system. New layers of fresh material were gradually added to form an aged-profile of layers in order to understand the interaction between earthworms and microorganisms during vermicomposting. Thus, changes in chemical and biological parameters were evaluated for 240days. The earthworm population reached maximum values in 120 d-old-layer, which was related to an increase in overall microbial biomass, assayed as dehydrogenase activity, in all of the processed materials. The pH was generally alkaline or neutral in all of the materials. The electrical conductivity did not modify significantly during vermicomposting, except in the case of the processed GP, and DG. The stabilization, in all of the processed materials, was detected after 240 d of vermicomposting, as indicated the decline in the content of dissolved organic carbon (DOC). The N-NO3- content exhibited an enhanced in the processed HM and AP, while a generalized decreased was found in the GP, and DG materials in 240 d-old-layer. The decline in microbial biomass activity, in all processed substrates, was related to a decrease in the earthworm activity after 240 d of vermicomposting, indicating a high degree of stabilization. However, the ß-glucosidase, phosphatase, protease, and o-diphenol oxidase activities were different according to the age of layers and type of processed material. The phytotoxicity test indicated that the end products of the processed AP and DG were chemically stable and enriched with nutrients in comparison with the HM and GP vermicompost. This fact indicates to stabilization (maturation) in the end product, which is important for its safe disposal as an organic nutrient-rich product.

Risk element sorption/desorption characteristics of dry olive residue: a technique for the potential immobilization of risk elements in contaminated soils.

Olive oil production is one of the most relevant agroindustrial activities in the Mediterranean region and generates a huge amount of both solid and semi-solid wastes, the uncontrolled disposal of which might lead to serious environmental problems. Due to its organic matter and mineral nutrient content, the waste material can be applied to agricultural soil as a fertilizer. However, due to its high organic matter content, dry olive residue (DOR), commonly called "alperujo," has the potential to immobilize risk elements in contaminated soils. The main objective of this study was to assess the possible effect of DOR on sorption of risk elements such as cadmium (Cd), lead (Pb), and zinc (Zn) in the soil. A set of batch sorption experiments were carried out to assess the ability of DOR to adsorb Cd, Pb, and Zn where the effect of the preceding biotransformation of DOR by four species of fungi: Penicillium chrysogenum, Coriolopsis floccosa, Bjerkhandera adusta, and Chondrostereum purpureum was compared. The Freundlich and Langmuir sorption isotherms were calculated to assess the sorption characteristics of both transformed and non-transformed DOR. The results showed good potential sorption capacity of DOR, especially for Pb and to a lesser extent for Cd and Zn. Better sorption characteristics were reported for the biotransformed DOR samples, which are expected to show higher humification of the organic matter. However, the desorption experiments showed weakness and instability of the DOR-bound elements, especially in the case of Zn. Thus, future research should aim to verify the DOR sorption pattern in contaminated soil as well as the potential stabilization of the DOR element bounds where the increase of the pH levels of the DOR samples needs to be taken into account.

Organic and inorganic amendment application on mercury-polluted soils: effects on soil chemical and biochemical properties.

On the basis of a previous study performed in our laboratory, the use of organic and inorganic amendments can significantly modify the Hg mobility in soil. We have compared the effectiveness of organic and inorganic amendments such as digestate and fly ash, respectively, reducing the Hg mobility in Chernozem and Luvisol soils differing in their physicochemical properties. Hence, the aim of this work was to compare the impact of digestate and fly ash application on the chemical and biochemical parameters in these two mercury-contaminated soils in a model batch experiment. Chernozem and Luvisol soils were artificially contaminated with Hg and then incubated under controlled conditions for 21 days. Digestate and fly ash were applied to both soils in a dose of 10 and 1.5 %, respectively, and soil samples were collected after 1, 7, 14, and 21 days of incubation. The presence of Hg in both soils negatively affected to processes such as nitrification, provoked a decline in the soil microbial biomass C (soil microbial biomass C (MBC)), and the microbial activities (arylsulfatase, and ß-glucosaminidase) in both soils. Meanwhile, the digestate addition to Chernozem and Luvisol soils contaminated with Hg improved the soil chemical properties (pH, dissolved organic carbon (DOC), N (Ntot), inorganic-N forms (N-NH4 (+) and N-NO3 (-))), as consequence of high content in C and N contained in digestate. Likewise, the soil MBC and soil microbial activities (dehydrogenase, arylsulfatase, and ß-glucosaminidase) were greatly enhanced by the digestate application in both soils. In contrast, fly ash application did not have a remarkable positive effect when compared to digestate in Chernozem and Luvisol soil contaminated with mercury. These results may indicate that the use of organic amendments such as digestate considerably improved the soil health in Chernozem and Luvisol compared with fly ash, alleviating the detrimental impact of Hg. Probably, the chemical properties present in digestate may determine its use as a suitable amendment for the assisted-natural attenuation of mercury-polluted soils.

The effectiveness of various treatments in changing the nutrient status and bioavailability of risk elements in multi-element contaminated soil.

Potential changes in the mobility and bioavailability of risk and essential macro- and micro-elements achieved by adding various ameliorative materials were evaluated in a model pot experiment. Spring wheat (Triticum aestivum L.) was cultivated under controlled condition for 60 days in two soils, uncontaminated Chernozem and multi-element contaminated Fluvisol containing 4900 ± 200 mg/kg Zn, 35.4 ± 3.6 mg/kg Cd, and 3035 ± 26 mg/kg Pb. The treatments were all contained the same amount of sulfur and were as follows: (i) digestate from the anaerobic fermentation of biowaste, (ii) fly ash from wood chip combustion, and (iii) ammonium sulfate. Macro- and micro-nutrients Ca, Mg, K, Fe, Mn, Cu, P, and S, and risk elements Cd, Cr, Pb, and Zn were assayed in soil extracts with 0.11 mol/l solution of CH3COOH and in roots, shoots, and grain of wheat after 30 and 60 days of cultivation. Both digestate and fly ash increased levels of macro- and micro-nutrients as well as risk elements (especially Cd and Zn; the mobility of Pb decreased after 30 days of cultivation). The changes in element mobility in ammonium sulfate-treated soils appear to be due to both changes in soil pH level and inter-element interactions. Ammonium sulfate tended to be the most effective measure for increasing nutrient uptake by plants in Chernozem but with opposite pattern in Fluvisol. Changes in plant yield and element uptake in treated plants may have been associated with the higher proline content of wheat shoots cultivated in both soils compared to control. None of the treatments decreased uptake of risk elements by wheat plants in the extremely contaminated Fluvisol, and their accumulation in wheat grains significantly exceeded maximum permissible levels; these treatments cannot be used to enable cereal and other crop production in such soils. However, the combination of increased plant growth alongside unchanged element content in plant biomass in pots treated with digestate and fly ash suggests that these treatments have a beneficial impact on yield and may be effective treatments in crops grown for phytoremediation.

Applications of organic and inorganic amendments induce changes in the mobility of mercury and macro- and micronutrients of soils.

Both soil organic matter and sulfur (S) can reduce or even suppress mercury (Hg) mobility and bioavailability in soil. A batch incubation experiment was conducted with a Chernozem and a Luvisol artificially contaminated by 440 mg · kg(-1) Hg showing wide differences in their physicochemical properties and available nutrients. The individual treatments were (i) digestate from the anaerobic fermentation of biowaste; (ii) fly ash from wood chip combustion; and (iii) ammonium sulfate, and every treatment was added with the same amount of S. The mobile Hg portion in Chernozem was highly reduced by adding digestate, even after 1 day of incubation, compared to control. Meanwhile, the outcome of these treatments was a decrease of mobile Hg forms as a function of incubation time whereas the contents of magnesium (Mg), potassium (K), iron (Fe), manganese (Mn), copper (Cu), zinc (Zn), and phosphorus (P) were stimulated by the addition of digestate in both soils. The available calcium (Ca) contents were not affected by the digestate addition. The experiment proved digestate application as the efficient measure for fast reduction of mobile Hg at extremely contaminated soils. Moreover, the decrease of the mobile mercury portion was followed by improvement of the nutrient status of the soils.

Arbuscular mycorrhizal fungi alleviate oxidative stress induced by ADOR and enhance antioxidant responses of tomato plants.

The behaviour of tomato plants inoculated with arbuscular mycorrhizal (AM) fungi grown in the presence of aqueous extracts from dry olive residue (ADOR) was studied in order to understand how this symbiotic relationship helps plants to cope with oxidative stress caused by ADOR. The influence of AM symbiosis on plant growth and other physiological parameters was also studied. Tomato plants were inoculated with the AM fungus Funneliformis mosseae and were grown in the presence of ADOR bioremediated and non-bioremediated by Coriolopsis floccosa and Penicillium chrysogenum-10. The antioxidant response as well as parameters of oxidative damage were examined in roots and leaves. The data showed a significant increase in the biomass of AM plant growth in the presence of ADOR, regardless of whether it was bioremediated. The establishment and development of the symbiosis were negatively affected after plants were exposed to ADOR. No differences were observed in the relative water content (RWC) or PS II efficiency between non-AM and AM plants. The increase in the enzymatic activities of superoxide dismutase (SOD; EC 1.15.1.1), catalase (CAT; EC 1.11.1.6) and glutathione-S-transferase (GST; EC 2.5.1.18) were simultaneous to the reduction of MDA levels and H2O2 content in AM root growth in the presence of ADOR. Similar H2O2 levels were observed among non-AM and AM plants, although only AM plants showed reduced lipid peroxidation content, probably due to the involvement of antioxidant enzymes. The results highlight how the application of both bioremediated ADOR and AM fungi can alleviate the oxidative stress conditions, improving the growth and development of tomato plants.

Oxidative stress induced in sunflower seedling roots by aqueous dry olive-mill residues.

The contamination of soils with dry olive-mill residue can represent a serious problem as being an environmental stressor in plants. It has been demonstrated that inoculation of aqueous extract of olive oil-mill residue (ADOR) with saprobe fungi removes some phenolic compounds. In this paper we studied the effect of ADOR uninoculated or inoculated with saprobe fungi in sunflower seedling roots. The germination and root growth, O(2)·(-) generation, superoxide dismutase (SOD) and extracellular peroxidases (EC-POXs) activities, and the content of some metabolites involved in the tolerance of stress were tested. The roots germinated in ADOR uninoculated show a decrease in meristem size, resulting in a reduction of the root length and fresh weight, and in the number of layers forming the cortex, but did not alter the dry weight, protein and soluble amino acid content. ADOR caused the decreases in O(2)·(-) generation and EC-POX's activities and protein oxidation, but enhanced SOD activity, lipid peroxidation and proline content. Fluorescence imaging showed that ADOR induced O(2)·(-) and H(2)O(2) accumulation in the roots. The increase in SOD and the decrease in EC-POX's activities might be involved in the enhancement of H(2)O(2) content and lipid peroxidation. Control roots treated with ADOR for 10 min show an oxidative burst. Roots germinated in ADOR inoculated with saprobe fungi partially recovered normal levels of ROS, morphological characteristics and antioxidant activities. These results suggested that treatment with ADOR caused a phytotoxic effect during germination inducing an oxidative stress. The inoculation of ADOR with saprobe fungi limited the stress.

Defence response of tomato seedlings to oxidative stress induced by phenolic compounds from dry olive mill residue.

ADOR is an aqueous extract obtained from the dry olive mill residue (DOR) which contains the majority of its soluble phenolic compounds, which are responsible for its phytotoxic properties. Some studies have shown that ADOR negatively affects seed germination. However, to date, few studies have been carried out on the effect of ADOR on the oxidative stress of the plant. It is well known that saprobe fungi can detoxify these phenolic compounds and reduce the potential negative effects of ADOR on plants. To gain a better understanding of the phytotoxic effects and oxidative stress caused by this residue, tomato seeds were germinated in the presence of ADOR, treated and untreated with Coriolopsis rigida, Trametes versicolor, Pycnoporus cinnabarinus and Penicillium chrysogenum-10 saprobe fungi. ADOR sharply reduced tomato seed germination and also generated high levels of malondialdehyde (MDA), O(2)(-) and H(2)O(2). However, bioremediated ADOR did not negatively affect germination and reduced MDA, O(2)(-) and H(2)O(2) content in different ways depending on the fungus used. In addition, the induced defense response was studied by analyzing the activity of both antioxidant enzymes (superoxide dismutase (SOD), catalase, ascorbate peroxidasa, glutathione reductase (GR), peroxidases and coniferil alcohol peroxidasa) and detoxification enzymes (glutathione-S-transferase (GST)). Our findings suggest that, because ADOR is capable of inducing oxidative stress, tomato seedlings trigger a defense response through SOD, GR, and GST activity and through antioxidant and lignification processes. On the other hand, the bioremediation of ADOR plays an important role in counteracting the oxidative stress induced by the untreated residue.