Impact of DOM from composted "alperujo" on soil structure, AM fungi, microbial activity and growth of Medicago sativa

Water-soluble extracts from compost may represent an alternative nutrient and organic matter source for crop production under drip irrigation. Dissolved organic matter (DOM), extracted from composted "alperujo", the main by-product from the Spanish olive oil industry, was applied to soil alone or in combination with either Glomus intraradices Schenck and Smith or a mixture of G. intraradices, Glomus deserticola (Trappe, Bloss. and Menge) and Glomus mosseae (Nicol and Gerd.) Gerd. and Trappe. Response measurements included mycorrhizal colonisation, nutrient uptake and growth of Medicago sativa and microbiological and physical properties in the rhizosphere. Dissolved organic matter was added to soil at concentrations of 0, 50, 100 or 300mgCkg(-1) substrate. During the four months of the experiment, the plants were harvested three times. Both mycorrhizal inoculation treatments significantly increased soil aggregate stability. Only the mycorrhizal inoculations increased microbial biomass C and protease and phosphatase activities and decreased water-soluble C, particularly the mixture of arbuscular mycorrhizal fungi. At the third harvest, the greatest increase in growth of M. sativa was observed in the inoculated plants with shoot biomass being 38% greater than for plants grown in the soil amended with the highest dose of DOM and 57% greater than for control plants. The addition of DOM was not sufficient to restore soil structure and microbial activity and did not affect the mycorrhizal development of introduced populations of arbuscular mycorrhizal fungi, but, depending on the dose, its fertiliser efficiency for improving plant growth was apparent.     

Landfill Leachate Ecotoxicity Experiments using Lemna minor

The effects of varying concentrations of landfill leachate on the growth, frond area, chlorophyll content and fluorescence of four strains of Lemna minor were assessed. Growth fluorescence and frond chlorophyll content decreased after seven days exposure to leachate, although responses differed between the strains and end parameters. A L. minor bioassay was used to assess leachate toxicity and the effectiveness of a constructed wetland treatment system and pre-treatment aeration and settlement in reducing toxicity. Pre-treatments were found to significantly reduce toxicity, so their incorporation in any treatment system may increase pollutant stripping.     

A Sensitive Electrochemical Impedance Spectroscopy Method for Detection of Polyimide Degradation by Microorganisms

An electrochemical impedance spectroscopy (EIS) technique was evaluated for monitoring microbial degradation of electronic packaging polyimides. The microbial inoculum was a mixed culture of fungi isolated previously from deteriorated polyimides. The active fungal consortium comprised Aspergillus versicolor, Cladosporium cladosporioides, and a Chaetomium species. After inoculation, fungal growth on the polyimides resulted in distinctive EIS spectra indicative of polymer insulation failure, which directly related to polymer integrity. Degradation appeared to occur in a number of steps and two distinctive stages in the decline of film resistance were observed in the inoculated EIS cells within the 2 and 10 weeks after inoculation. The early stage of resistance decrease may be related to the ingress of water molecules and ionic species into the polymeric materials, whereas the second stage probably resulted from partial degradation of the polymers by fungal growth on the polymer film. The relationship between changes of impedance spectra and microbial degradation of the polymer was further supported by scanning electron microscopy (SEM) observations of fungi growing on the surface of the inoculated polyimides. Our data indicate that the EIS can be used in detection of early degradation of resistant polymers and polyimides that are susceptible to biodeterioration.     

Accumulation of Heavy Metals by in vitro cultures of plants

The aim of the project is to study heavy metals accumulation by the selected plants in both laboratory and field conditions. Within the experiments the aspen (Populus tremula × tremuloides), sunflower (Helianthus annuus) and corn (Zea mays) plants were studied. The reasons for this selection were: a fast growth of these plants, an accumulation capacity and an ability to survive in different types of soils. The study was carried out on the aspen plantlets grown in vitro. The plants were exposed to the aqueous solutions having concentrations 0.1 mM, 0.5 mM of Pb²⁺ or Ni²⁺, respectively. The accumulation capacityfor aspen, was about 70% of Pb²⁺ originally present in the solution. The starting concentration of Pb²⁺ (0.5 mM) exhibited no negative impact on the growth. Besides in vitro expositions, a pilot-scale phytoremediation experiment was carried out at the polluted industrial area (Zn – 75000 mg/kg), (Pb – 16000 mg/kg), (Cr – 590 mg/kg), (Cd – 90 mg/kg) and (Cu – 1700 mg/kg).     

Assessment of the possible reuse of MSW coming from landfill mining of old open dumpsites

The present study addresses the theme of recycling potential of old open dumpsites by using landfill mining. Attention is focused on the possible reuse of the residual finer fraction (<4 mm), which constitutes more than 60% of the total mined material, sampled in the old open dumpsite of Lavello (Southern Italy). We propose a protocol of analysis of the landfill material that links chemical analyses and environmental bioassays. This protocol is used to evaluate the compatibility of the residual matrix for the disposal in temporary storages and the formation of “bio-soils” to be used in geo-environmental applications, such as the construction of barrier layers of landfills, or in environmental remediation activities. Attention is mainly focused on the presence of heavy metals and on the possible interaction with test organisms. Chemical analyses of the residual matrix and leaching tests showed that the concentration of heavy metals is always below the legislation limits. Biological acute tests (with Lepidum sativum, Vicia faba and Lactuca sativa) do not emphasize adverse effects to the growth of the plant species, except the bioassay with V. faba, which showed a dose–response effect. The new developed chronic bioassay test with Spartium junceum showed a good adaptation to stress conditions induced by the presence of the mined landfill material. In conclusion, the conducted experimental activities demonstrated the suitability of the material to be used for different purposes.     

The Effect of Interaction Between White-rot Fungi and Indigenous Microorganisms on Degradation of Polycyclic Aromatic Hydrocarbons in Soil

White-rot fungi applied for soil bioremediation have to compete with indigenous soil microorganisms. The effect of competition on both indigenous soil microflora and white-rot fungi was evaluated with regard to degradation of polycyclic aromatic hydrocarbons (PAH) with different persistence in soil. Sterile and non-sterile soil was artificially contaminated with ¹⁴C-labeled PAH consisting of three (anthracene), four (pyrene, benz[a]anthracene) and five fused aromatic rings (benzo[a]pyrene, dibenz[a,h]anthracene). The two fungi tested,Dichomitus squalens and Pleurotus ostreatus, produced similar amounts of ligninolytic enzymes in soil, but PAH mineralization by P. ostreatus was significantly higher. Compared to the indigenous soil microflora, P.ostreatus mineralized 5-ring PAH to a larger extent, while the indigenous microflora was superior in mineralizing 3-ring and 4-ring PAH. In coculture the special capabilities of both soil microflora and P. ostreatus were partly restricted due to antagonistic interactions, but essentially preserved. Thus, soil inoculation with P. ostreatus significantly increased the mineralization of high-molecular-weight PAH, and at the same time reduced the mineralization of anthracene and pyrene. Regarding the mineralization of low-molecular-weight PAH, the stimulation of indigenous soil microorganisms by straw amendment was more efficient than application of white-rot fungi.     

Effects of Liming and Ash Application on Below Ground Ectomycorrhizal Community Structure in Two Norway Spruce Forests

The effect of lime and ash additions upon the below ground ectomycorrhizal communities was studied at two spruce forest sites, Horröd and Hasslöv, in Southern Sweden. At Hasslöv, fifteen years after the addition of 8.75 ton ha^-1 of dolomite, the species richness of the mycorrhizal community was unchanged. However, the liming resulted in an almost total replacement of species normally found under the site conditions, with only three morphotypes from a total of 24 distinguished being common to both the control and limed plots. At Horröd, four years after the application of 3.25 ton ha^-1 crushed lime, there was a significant reduction in the number of species recorded. This was not a result of a reduction in root tips sampled, since there was no significant treatment effect upon root tip abundance. Instead, it appeared that the less common species present in the control plots had been lost following liming. At the same site, the addition of 4.28 ton ha^-1 of wood ash had little effect upon the mycorrhizal community as a whole, with only small changes recorded in the abundance of a small number of species. This study highlights the urgent need for additional data on the temporal and load response of ectomycorrhizal fungi to the addition of lime. This information is vital in view of the proposed widespread use of lime as a counter measure to soil acidification.     

Changes in Extractability of Cr and Pb in a Polycontaminated Soil After Bioaugmentation With Microbial Producers of Biosurfactants, Organic Acids and Siderophores

Partly because of the low bioavailability of metals, the soil cleaning-up using phytoremediation is usually time-consuming. In order to enhance the amount of metals at the plant's disposal, the soil bioaugmentation coupled together with phytoextraction is an emerging technology. In this preliminary work, two agricultural soils which mainly differed in their Cr, Hg and Pb contents (LC, low-contaminated soil; HC, high-contaminated soil) were bioaugmented in laboratory conditions by either bacterial (Bacillus subtilis, Pseudomonas aeruginosa, Pseudomonas fluorescens or Ralstonia metallidurans) or fungal inocula (Aspergillus niger or Penicillium simplicissimum) and incubated during three weeks. The LC soil pots bioaugmented with A. niger and P. aeruginosa contained higher concentrations of Cr (0.08 and 0.25 mg.kg⁻¹ dw soil) and Pb (0.25 and 0.3 mg.kg⁻¹ dw soil) in the exchangeable fraction F1 (extraction with MgCl₂) by comparison with the non-bioaugmented soil where neither Cr nor Pb was detected. Conversely, immobilization of Cr and Pb in the soil were observed with the other microorganisms. The soil bioaugmentation not only modified the metal speciation for the most easily extractable fractions but also modified the distribution of metals in the other fractions, to a lesser extent nevertheless. The difference in microbial concentrations between the bioaugmented or not HC soils reached up to 1.8 log units. Thus the microorganisms that we chose for the soil bioaugmentation were competitive towards the indigenous microflora. The PCA analysis showed close positive relationships between the microorganisms which potentially produced siderophores in the soil and the amount of Cr and Pb in the fraction F1.     

Effects of Acidification and its Mitigation with Lime and Wood Ash on Forest Soil Processes: A Review

Anthropogenic acid deposition causes forest soil acidification and perturbation of the soil forming processes. The impact of soil acidification on tree growth is discussed in view of the role of mycorrhizal fungi in weathering and nutrient uptake. A review has been carried out of experiments involving treatments of forest soil by lime and wood ash, where soil properties and soil solution composition have been investigated. Results from these experiments in Europe and North America are summarized. In general, the content of C in the mor layer decreased as a result of treatment due to higher microbial activity and soil respiration as well as increased leakage of DOC. In addition, the content of N in the mor layer, in general, decreased after treatment and there are occasional peaks of high NO₃concentrations in soil solution. In nearly all reviewed investigations the pH of the deep mineral soil solution decreased and Al, SO₄and NO₃concentrations increased after treatment. These effects are probably due to the high ionic strength and increased microbial activity as a consequence of the treatments. In the soil, pH, CEC and base saturation increased in the upper horizons, but decreases in the upper mineral soil are also reported. In general, there was no increase in tree growth as a result of these treatments. The positive effects of the treatments on soil processes and tree growth are therefore questionable. In view of these conclusions, an investigation was carried out on the soil and soil solution chemistry and the role of mycorrhizal fungi in a spruce stand treated with two doses of lime and another treated with lime/ash in southern Sweden. The results of this investigation is reported in this volume.     

Fungal and enzymatic treatment of mature municipal landfill leachate

The aim of our study was to evaluate biotreatability of mature municipal landfill leachate by using white rot fungus and its extracellular enzymes. Leachates were collected in one active and one closed regional municipal landfill. Both chosen landfills were operating for many years and the leachates generated there were polluted by organic and inorganic compounds. The white rot fungus Dichomitus squalens was able to grow in the mature leachate from the closed landfill and as it utilizes present organic matter as a source of carbon, the results were showing 60% of DOC and COD removal and decreased toxicity to the bacterium Aliivibrio fischeri. On the other hand, growth of the fungus was inhibited in the presence of the leachate from the active landfill. However, when the leachate was introduced to a crude enzyme filtrate containing extracellular ligninolytic enzymes, removal levels of COD and DOC reached 61% and 44%, respectively. Furthermore, the treatment led to detoxification of the leachate to the bacterium Aliivibrio fischeri and to reduction of toxicity (42%) to the plant Sinapis alba. Fungal and enzymatic treatment seems to be a promising biological approach for treatment of mature landfill leachates and their application should be further investigated.     

Spatial and Temporal Variations in the Sediment Habitat of Ranunculus spp. in Lowland Chalk Streams – Implications for Ecological Status?

Ranunculus spp. are the dominant plants of lowland chalk stream habitats in England. The spatial variability of sediment characteristics (silt–clay, organic matter, total phosphorus and total nitrogen content) within stands of Ranunculus spp. was investigated in 12 rivers in lowland England. Variability was found to be high and there were no discernible differences between samples taken from within Ranunculus and a limited number of samples from bare substrate. For two of these rivers, comparisons were also made between reaches upstream and downstream of waste water treatment works outfalls in terms of the characteristics of the sediments within Ranunculus stands. In one river a clear increase in sediment nutrient, fine and organic material content was observed downstream but in the other there was no consistent difference. Temporal variability was considered for two rivers in the Frome catchment, Dorset, by analysing the monthly variability in sediment organic matter and silt–clay content beneath Ranunculus stands over an annual cycle of growth and die-back. Whilst a clear pattern of fine and organic material retention consistent with seasonal plant growth patterns was evident at one site, the three sites displayed different temporal patterns. This inconsistency is believed to reflect differences in sediment supply at the three sites.     

Biodegradation of chlorinated organic pesticides endosulfan and chlorpyrifos in soil extract broth using fungi

The present study was designed to screen 20 fungi for their potential to degrade the chlorinated organic pesticides endosulfan and chlorpyrifos. Fungi were first screened for their tolerance to various concentrations of target pesticides using soil extract agar and subsequent degradation studies were performed in soil extract broth containing 25 mg/L of the individual pesticide. Pesticide degradation was evaluated using gas chromatography. Other parameters, such as pH and mycelial weight, were also determined. Based on percent growth inhibition of test fungi and subsequent analysis of EC₅₀ values, the overall results revealed that chlorpyrifos showed significantly more growth inhibition in all tested fungi compared with endosulfan. Trametes hirsuta showed complete degradation of both α‐ and β‐endosulfan isomers and Cladosporium cladosporioides displayed maximum degradation of chlorpyrifos. All test fungi degraded endosulfan more efficiently than chlorpyrifos, except Phanerochaete chrysosporium, Trichoderma harzianum, and Trichoderma virens which showed higher degradation of chlorpyrifos than endosulfan. It was also found that all tested fungi degraded α‐endosulfan more efficiently than β‐endosulfan. Endosulfan sulfate was found to be the major degradation product with all tested fungi. Fungi which showed more endosulfan degradation also produced more endosulfan sulfate. However, less endosulfan sulfate was detected with T. hirsuta and Trametes versicolor, although they degraded endosulfan more efficiently.     

Degradation of L- and DL-lactic acid oligomers in the presence ofFusarium moniliforme andPseudomonas putida

Poly(-alkanoates) derived from lactic acid enantiomers are known to degrade easily hydrolytically in aqueous media. The ability of two microorganisms, a filamentous fungus,Fusarium moniliforme, and a bacterium,Pseudomonas putida, to assimilate the degradation by-products of poly(lactic acid) (PLA), namely, lactic acid, lactyllactic acid dimers, and higher oligomers, was investigated in liquid culture. To distinguish the influence of chirality on bioassimilation, two series of substrates were considered which derived from the racemic and the L-form of lactic acid, respectively. The fate of these compounds was monitored by HPLC. Under the selected conditions,DL- andL-lactic acids were totally used by the two microorganisms regardless of the enantiomeric composition. Both microorganisms degraded the LL-dimer rather rapidly. However,F. moniliforme acted more rapidly thanP. putida. It is likely that the DD-dimer also biodegraded but at a slower rate, especially in the case of the fungi. Higher racemic oligomers were slowly assimilated by the two microorganisms, whereas higher L-oligomers appeared biostable probably because of their crystallinity. A synergistic effect was observed when both microorganisms were present in the same culture medium containing racemic oligomers.Presented at the 4th International Workshop on Biodegradable Plastics and Polymers, October 11–14, 1995. Durham, New Hampshire.     

Anammox: an option for ammonium removal in bioreactor landfills

Experiments carried out in bioreactor landfill simulators demonstrated that more than 40% of the total N was transferred into the liquid and gas phases during the incubation period of 380 days. Ammonium, an end product of protein degradation and important parameter to consider during landfill closure, tends to accumulate up to inhibitory levels in the leachate of landfills especially in landfills with leachate recirculation. Most efforts to remove ammonium from leachate have been focused on ex situ and partial in situ methods such as nitrification, denitrification and chemical precipitation. Besides minimal contributions from other N-removal processes, Anammox (Anaerobic Ammonium Oxidation) bacteria were found to be active within the simulators. Anammox is considered to be an important contributor to remove N from the solid matrix. However, it was unclear how the necessary nitrite for Anammox metabolism was produced. Moreover, little is known about the nature of residual nitrogen in the waste mass and possible mechanisms to remove it. Intrusion of small quantities of O₂ is not only beneficial for the degradation process of municipal solid waste (MSW) in bioreactor landfills but also significant for the development of the Anammox bacteria that contributed to the removal of ammonium. Volatilisation and Anammox activity were the main N removal mechanisms in these pilot-scale simulators. The results of these experiments bring new insights on the behaviour, evolution and fate of nitrogen from solid waste and present the first evidence of the existence of Anammox activity in bioreactor landfill simulators.     

Factors affecting polycyclic aromatic hydrocarbon biodegradation by Aspergillus flavus

This study investigated the ability of fungi isolated from highly contaminated soil to biodegrade polycyclic aromatic hydrocarbon (PAH) compounds, as well as the effect of several parameters on the biodegradation ability of these fungi. The isolated fungi were identified using ITS rDNA sequencing and tested using 2,6‐dichlorophinolendophenol to determine their preliminary ability to degrade crude oil. The top‐performing fungi, Aspergillus flavus and Aspergillus fumigatus, were selected to test their ability to biodegrade PAH compounds as single isolates. After 15 days of incubation, A. flavus degraded 82.7% of the total PAH compounds, with the complete degradation of six compounds, whereas A. fumigatus degraded 68.9% of the total PAHs, with four aromatic compounds completely degraded. We also tested whether different temperatures, pH, and nitrogen sources influenced the growth of A. flavus and the degradation rate. The degradation process was optimal at a temperature of 30°C, pH of 5.5, and with nitrogen in the form of yeast extract. Finally, the ability of the fungal candidate, A. flavus, to degrade PAH compounds under these optimum conditions was studied. The results showed that 95.87% of the total PAHs, including 11 aromatic compounds, were completely degraded after 15 days of incubation. This suggests that A. flavus is a potential microorganism for the degradation of PAH compounds in aqueous cultures.     

Correlation between physicochemical and ecotoxicological approaches to estimate landfill leachates toxicity

Leachates from municipal solid waste (MSW) landfills may contain a huge diversity of contaminants; these wastewaters should be considered as potentially hazardous complex mixtures, representing a potential environmental risk for surface and groundwater. Current MSW landfill wastes regulatory approaches deem exclusively on the physicochemical characterization and does not contemplate the ecotoxicological assessment of landfill leachates. However, the presence of highly toxic substances in consumer products requires reconsideration on the need of more specific ecotoxicological assessments. The main aim of this study was to evaluate the toxicity of different MSW landfill leachates using a battery of toxicity tests including acute toxicity tests with Daphnia magna and the anuran Xenopus laevis and the in vitro toxicity test with the fish cell line RTG-2. The additional objective was to study the possible correlation between physicochemical properties and the toxicity results obtained for untreated landfill leachates. The results showed that the proposed test battery was effective for the ecotoxicological characterization of MSW landfill leachates. A moderate to strong correlation between the measured physicochemical parameters and the calculated toxicity units was detected for all toxicity assays. Correlation factors of 0.85, 0.86 and 0.55 for Daphnia, Xenopus and RTG-2 tests, respectively, were found. The discriminant analysis showed that certain physicochemical parameters could be used for an initial categorization of the potential aquatic acute toxicity of leachates; this finding may facilitate leachates management as the physicochemical characterization is currently the most common or even only monitoring method employed in a large majority of landfills. Ammonia, alkalinity and chemical oxygen demand (COD), together with chloride, allowed a proper categorization of leachates toxicity for up to 75% of tested samples, with a small percentage of false negatives.     

Parametric study of MSW landfill settlement model

A newly developed and validated constitutive model that accounts for primary compression and time-dependent mechanical creep and biodegradation is used for parametric study to investigate the effects of model parameters on the predicted settlement of municipal solid waste (MSW) with time. The model enables the prediction of stress strain response and yield surfaces for three components of settlement: primary compression, mechanical creep, and biodegradation. The MSW parameters investigated include compression index, coefficient of earth pressure at-rest, overconsolidation ratio, and biodegradation parameters of MSW. A comparison of the predicted settlements for typical MSW landfill conditions showed significant differences in time-settlement response depending on the selected model input parameters. The effect of lift thickness of MSW on predicted settlement is also investigated. Overall, the study shows that the variation in the model parameters can lead to significantly different results; therefore, the model parameter values should be carefully selected to predict landfill settlements accurately. It is shown that the proposed model captures the time settlement response which is in general agreement with the results obtained from the other two reported models having similar features.     

Verification of exponential distribution of arrival and service times at municipal solid waste facilities

Vehicle queuing at municipal solid waste (MSW) facilities causes economic and environmental damage. In the United States, typical MSW receiving facilities (transfer, recycling, energy recovery, and landfill) have three queues: one at the entrance weighing station, one at the waste tipping location, and one at the exit weighing station. A common method of determining queue behavior relies on equations that assume exponentially distributed arrival and service times, but there has not been a comprehensive study to determine whether this assumption is valid for a variety of MSW facilities and conditions. In this study, data were gathered from two transfer stations, two energy recovery plants, and one transfer/landfill facility. Among the five facilities there was a variety of queuing styles. The data were plotted as an inverse exponential relationship and linearized. The linearized plots were regressed and R ² values were calculated. It was determined that the negative exponential relationship can be used to describe arrival times at all three queues and service times at weighing queues. The queuing equations are therefore theoretically supported for use at entering and exiting weighing queues and moderately theoretically supported for use at tipping queues.     

Waste settlement in bioreactor landfill models

Prediction of landfill settlement is one of the important parameters that affects the design and maintenance of bioreactor landfills. Due to the large number of variables involved in the settlement mechanism, accurate prediction of landfill settlement is a real challenge. The operational protocol of a landfill, the presence of municipal sludge from treatment plants, the addition of soybean peroxidase (SBP) enzymes, and the fraction of organic matter in the municipal solid waste (MSW) have to be reflected in the parameters of any model used to predict the settlement of MSW. In this work, a biodegradation-induced settlement model incorporating two parameters (A and B) was developed. The settlement data of two researchers were used to estimate the parameter values with two different approaches; the first considered the overall experiment and results, and the second separated the aerobic phase, if present, from the anaerobic phase. The rate of initial settlement occurring under aerobic conditions has been greater than that under anaerobic conditions. Parameters increased with the increase in the concentration of enzymes and with the presence of sludge in both aerobic and anaerobic stages. Increasing organic content of MSW has resulted in the enhancement of the biodegradation rate and settlement. This has been reflected on the higher values of the parameters compared to their values in the absence of organic waste.     

Use of fungal technology in soil remediation: A Case Study

Two white rot fungi Irpex lacteus and Pleurotus ostreatus and a PAH-degrading bacterial strain of Pseudomonas putida were used as inoculum for bioremediation of petroleum hydrocarbon-contaminated soil from a manufactured-gas-plant-area. Also two cocultures comprising a fungus with Pseudomonas putida were applied. After 10-week treatment out of 12 different PAHs, concentration of phenanthrene, anthracene, fluoranthene and pyrene decreased up to 66%. The ecotoxicity of the soil after bioremediation did not reveal any effect on the survival of Daphnia magna, a crustacian. However, the toxic effect on seed germination of plant Brassica alba and oxidoreductase activity of bacterium Bacillus cereus decreased after 5 and 10 weeks of treatment.