Esperanza Romero

Inter-laboratory evaluation of the ISO standard 11063 “Soil quality — Method to directly extract DNA from soil samples”

Extracting DNA directly from micro-organisms living in soil is a crucial step for the molecular analysis of soil microbial communities. However, the use of a plethora of different soil DNA extraction protocols, each with its own bias, makes accurate data comparison difficult. To overcome this problem, a method for soil DNA extraction was proposed to the International Organization for Standardization (ISO) in 2006. This method was evaluated by 13 independent European laboratories actively participating in national and international ring tests. The reproducibility of the standardized method for molecular analyses was evaluated by comparing the amount of DNA extracted, as well as the abundance and genetic structure of the total bacterial community in the DNA extracted from 12 different soils by the 13 laboratories. High quality DNA was successfully extracted from all 12 soils, despite different physical and chemical characteristics and a range of origins from arable soils, through forests to industrial sites. Quantification of the 16S rRNA gene abundances by real time PCR and analysis of the total bacterial community structure by automated ribosomal intergenic spacer analysis (A-RISA) showed acceptable to good levels of reproducibility. Based on the results of both ring-tests, the method was unanimously approved by the ISO as an international standard method and the normative protocol will now be disseminated within the scientific community. Standardization of a soil DNA extraction method will improve data comparison, facilitating our understanding of soil microbial diversity and soil quality monitoring.

Continuous-feeding vermicomposting as a recycling management method to revalue tomato-fruit wastes from greenhouse crops

Huge quantities of discarded fruits generated from greenhouse crops represent a worldwide environmental problem. The aim of this work was to assess the efficiency of vermicomposting as a recycling management option for biotransforming tomato-fruit wastes from greenhouses into an organic nutrient-rich product available for agricultural purposes. A pilot vermireactor was constructed. It was provided with a manure layer, where an initial population of Eisenia fetida was introduced and fed continuously at a high organic loading rate (13.6 kg TOC m(-3)wk(-1)) for 150 days. Vermicompost chemical and enzymatic parameters as well as the bacterial and fungal community structure were determined for 210 days (vermicomposting plus a maturation period). Earthworm biomass increased after 90 days, and then declined due to increasing pH, electrical conductivity and ammonium concentration. The temporal patterns of dehydrogenase, β-glucosidase, protease and urease were related to earthworm growth and the stabilization of organic matter. Bacterial DGGE profiles differed between the period of degradation of labile substrates and the maturation step. Fungal communities at the stage of maximum earthworm biomass differed most, suggesting a gut passage effect. The end product was chemically stable and enriched in nutrients, demonstrating that tomato-fruit wastes can be successfully vermicomposted into a valuable soil amendment. We suggest continuous-feeding vermicomposting as an environmentally sound management option for greenhouse wastes.

Dynamics of microbial communities related to biochemical parameters during vermicomposting and maturation of agroindustrial lignocellulose wastes.

Scarce information is available on the changes in abundance of microbial taxa during vermicomposting. Quantitative PCR and DGGE analysis were used to monitor variations in the microbial structure, relative abundance of four bacterial classes and fungi over the vermicomposting and maturation period of wet olive cake (O) and vine shoots (W). Multivariate correlation analysis between microbial structure and abundance, earthworm biomass and enzyme activities revealed similar and divergent interactions in both processes. Although Eisenia fetida development was different, significant correlations were found with β-glucosidase activity and with bacterial and fungal structure. In the vermicomposting period of O and W, a decline was found in bacteria (94% and 77%), fungi (93% and 94%), and Gammaproteobacteria (56% and 71%) but an increase in Betaproteobacteria and Actinobacteria (62-79%). Alphaproteobacteria increased only in O (26%). Despite the different initial lignocellulose wastes, the mature vermicomposts were similar in microbial and biochemical properties.

Evaluation of the sorption process for imidacloprid and diuron in eight agricultural soils from southern Europe using various kinetic models.

Kinetic studies are of great concern for understanding the processes and parameters involved in the sorption of pollutants by soils. Sorption kinetics of imidacloprid and diuron in eight soils of different characteristics, with very low organic carbon content were investigated. Pseudosecond-order kinetic reactions closely correlate with the experimental kinetic (R(2) > 0.98) in all soils. The sorbed amount of diuron was higher than that for imidacloprid. The low OC content of these soils correlated neither with the sorbed amount nor with the kinetic parameters for both pesticides. Imidacloprid sorption was correlated with silt and sand content and cation exchange capacity (CEC); meanwhile for diuron, no correlation was found. Thus, sorption kinetics take place throughout different mechanisms related mainly to the chemical character of the pesticides. Sorption kinetic parameters determined using three of the four models selected (pseudosecond-order kinetic reactions, Elovich equation, and Weber-Morris models) have been shown to be worthy to distinguish the process controlling the sorption kinetic of both pesticides.

Feasibility of vermicomposting for vegetable greenhouse waste recycling

This study was conducted in order to evaluate the feasibility of Eisenia andrei for vermicomposting heterogeneous-plant (HP), tomato-plant (P), and damaged tomato-fruit (T) greenhouse vegetable wastes. Earthworm growth and reproduction were monitored over a 12-week period, and variations in chemical parameters, enzyme activity, phytotoxicity test, and genetic fingerprinting of bacterial communities were evaluated. While high rates of salinity prevented earthworm survival in HP and P (>10 dS m(-1)), T was vermicomposted recording an adequate earthworm growth and cocoon production. The latter waste was successfully stabilized, as indicated by the significant decrease in its TOC content ( approximately 13-26%) and C:N ratio ( approximately 16-36%) and its high germination indices ( approximately 39-72%). The similar enzyme activities levels and bacterial community fingerprintings recorded in diverse vermicomposts obtained from T waste indicate that this type of waste favoured the existence of analogous bacterial communities responsible for the high degree of stabilization and maturity detected.

Biosolids and biosolids-ash as sources of heavy metals in a plant-soil system

Generally, the potential for biosolids (digested or composted)to contribute heavy metals to the soil-plant system has beencompared with commercial fertilizers and other organic wastesbut not with biosolids-ash. An column study was conducted in agreenhouse to determine the availability, extractability andleachability of metals in a degraded, non-calcareous soilamended with different biosolids (200 Mg ha-1). Thebiosolids investigated were dewatered, anaerobically digestedbiosolids, composted biosolids and biosolids-ash. The columns(26 cm) were planted with wheat (Triticum aestivum L. cvMexa). The addition of digested biosolids decreased the drymatter yield of wheat. Treatments including organic biosolidsincreased Cu and Zn concentrations in wheat roots, straw andgrain, whereas the addition of biosolids-ash did not affect theconcentrations of these metals in wheat. Concentrations of Ni,Co, Pb, Cr and Cd in wheat were below reliable detection limits(0.06, 0.05, 0.1, 0.06 and 0.02 mg kg-1, respectively).After harvesting, total and AB-DTPA extractable Cu, Zn and Pbincreased in the upper layer of the soil amended with thedifferent biosolids studied, whereas levels of AB-DTPAextractable Ni and Co were affected only when the soil wasamended with digested or composted biosolids. Total chromiumincreased only in treatments including organic biosolids. TheAB-DTPA extractable Cu, Zn and Pb in the lower layer of thesoil in treatments including biosolids evidenced downwardmovement of these metals. However, absence of these metals incolumn leachates indicates that this movement was gradual.

Effect of vermicomposting process on pesticide sorption capability using agro-industrial wastes

The aim of this work was to study the use of organic wastes from olive oil (alperujo), winery, and alcohol industries (vine shoots, spent grape marc, lees cake, and biosolids vinasse) as sorbents for pesticide control. The pesticide sorption capability of these organic wastes and the effect of vermicomposting process was also studied. The insecticide imidacloprid and different herbicides (diuron, metsulfuron-methyl, sulfuron-methyl, and flazasulfuron) were applied. The vermicomposting process was more effective for the agro-industrial wastes with a low lignin content. The sorption capacity of these wastes, natural or previously vermicomposted, was low for the anionic herbicides with respect to hydrophobic pesticides. Adsorption isotherms by the different wastes of confidor (imidacloprid 20% w/v) and diurokey (diuron 80% w/w) fit the Freundlich model (R 2 > 0.933). The larger K f values (231–138 µg1/n−1mL−1g−1) correspond to the spent grape marc, untreated or vermicomposted, due to the high lignin content of this waste.

Photodegradation of mecoprop and dichlorprop on dry, moist and amended soil surfaces exposed to sunlight

The effects of environmental conditions on the photodecomposition of two phenoxy-alkanoic acid herbicides were investigated on different soil surfaces conditions under sunlight exposition. A technique has been developed to study this process on moist nonsterile soil surfaces. A slow rate of disappeareance occurs in the three soils exposed to sunlight in absence of water. Photolysis of these herbicides is controlled by soil texture and its adsorption capacity. The organic matter does not have a sensitizing effect on the photodecomposition of these herbicides on dry soil surfaces. On nonsterile moist soil surfaces exposed to sunlight, the photolytic process prevails in the two first days of exposure and the transformation kinetics fits the Hoerl function better than the first-order exponential equation.

Use of DGGE and COMPOCHIP for investigating bacterial communities of various vermicomposts produced from different wastes under dissimilar conditions

This study describes the use of denaturing gradient gel electrophoresis (DGGE) and COMPOCHIP (i.e. a microarray targeting typical bacteria of stabilized organic materials and pathogenic bacteria) for investigating the bacterial communities of four different vermicomposts. These included a commercial vermicompost produced from cattle manure (CM) and three vermicomposts produced at pilot-scale by recycling: damaged tomato fruits (DT); olive-mill waste mixed with biosolids (OB); and winery wastes (WW). DGGE provided distinctive fingerprints of each vermicompost, which were statistically related to their particular chemical features. The comparison of the various vermicompost fingerprints showed that they contained bacterial communities with an average similarity coefficient of close to 80%. COMPOCHIP detected the presence of Sphingobacterium, Streptomyces, Alpha-Proteobacteria, Delta-Proteobacteria, and Firmicutes in all the vermicomposts. COMPOCHIP showed differences in the abundance of particular bacterial taxa among the vermicomposts, giving an idea about the usefulness of each vermicompost in the search for bacteria valuable to biotechnology. The joint use of DGGE and COMPOCHIP is a useful tool to compare vermicompost bacterial communities and to assess the potential of different vermicomposts as bioactive organic materials.

Isolation of an endosulfan-degrading bacterium from a coffee farm soil: persistence and inhibitory effect on its biological functions.

Endosulfan is a lypophilic persistent organic pollutant (POP) that has caused widespread concern due to its persistence in the environment, toxicity and bioaccumulation in living organisms. The aim of this study is to isolate endosulfan-degrading bacteria taken from five coffee farms historically exposed to this insecticide which could be used in future remediation strategies. The biodegradation capability of the isolated strain as well as endosulfans impact on some of the strains biological functions was studied. Endosulfan and its metabolites were analyzed using TLC and GC-MS. The isolated strain, capable of growing in a liquid culture treated with this insecticide as the sole sulfur source rather than a carbon source, was selected for further study. The isolated bacterium is Gram-negative, having the morphological and biochemical characteristics of Azotobacter sp. The remaining concentrations after 6 days, using 2 and 10 mg l(-1) of endosulfan, were 57.6 and 72.3% respectively, and the degradation constants were 0.12 d(-1) and 0.26 d(-1). Four metabolites were detected, one of which was identified as endosulfan ether. Endosulfan reduced nitrogenase activity but had no impact on indole 3-acetic acid production. Thus, these results suggest that this strain has the potential to act as a biocatalyst in endosulfan degradation.