Pesticide transformation by a variant of CYPBM3 with improved peroxygenase activity

The activity of the mutant CYPBM3 “21B3”, which is able to use hydrogen peroxide as the final electron acceptor, was evaluated against two major environmental pollutants; organochlorine and organophosphorus pesticides. This evolved CYP from Bacillus megaterium is able to transform a variety of structurally different pesticides. The catalytic parameters for two organochlorine; dichlorophen (k_cat = 9.2 min⁻¹, K_M = 64.1 μM) and linuron (k_cat = 226.5 min⁻¹, K_M = 468.2 μM), and two organophosphorus compounds; parathion (k_cat = 10.9 min⁻¹, K_M = 59.3 μM) and chlorpyrifos (k_cat = 9.2 min⁻¹, K_M = 226.5 μM) were determined giving catalytic efficiencies between 0.143 and 1.107 min⁻¹ μM⁻¹. CYPBM3 “21B3” has the ability to both activate and detoxify organophosphorus pesticides, as demonstrated by the chemical nature of the reaction products. The capacity to transform structurally diverse compounds together with the great stability, easy production and relatively inexpensive cofactors needed, makes CYPBM3 “21B3” an enzyme with a potential use on the environmental field.     

Influence of different plant species on water repellency in Mediterranean heathland soils

It is established that soil hydrophobicity reduces soil infiltration rates, and enhances runoff flow and soil erosion. Water repellency has been studied with special interest in coniferous and eucalyptus forests, particularly after burning, but the number of studies concerning Mediterranean heathlands is still very low. In this paper, we study the occurrence and persistence of water repellency in soil samples collected under different plant species susceptible to induce soil hydrophobicity (Erica arborea, Erica australis, Calluna vulgaris, Quercus lusitanica and Rhododendron ponticum) in a natural protected area in southern Spain. Great attention has been paid to the relationships between soil water repellency and environmental factors as organic matter content and soil acidity. The largest hydrophobicity was measured in soil samples collected under E. australis, E. arborea and C. vulgaris. For these species, the organic matter content and pH showed positive and negative correlations with the persistence of water repellency, respectively. The hydrophobicity originated by humic substances in the soil seems to be the only explanation for slight soil water repellency under Q. lusitanica or R. ponticum. The patchy patterns of occurrence and persistence of soil water repellency is governed by the spatial distribution of the studied species and modulated by other factors. Soil surface water repellent layers reduce the infiltration rates and limit the water storage capacity. However, the macropore flow can be enhanced on non-repellent layers, cracks or roots such us the wetting's front shown. The vegetation effects on soil hydrology should be considered for afforestation works and flooding control.     

Analysis of genetic variability in <Emphasis Type="Italic">Couepia</Emphasis> accessions using AFLP markers

Couepia is a genus distributed in tropical regions of America. The nut of some Couepia species is used as fresh fruit and oil source for local communities. Despite its critical situation of conservation and its economic potential, there is a lack of information on the genetic variability of Couepia species. This study examines AFLP variation among 40 accessions of Couepia collected in the Colombian Amazonian region, representing two species: Couepia dolichopoda and Couepia subcordata. The individuals were examined for 96 markers generated from four EcoRI/MseI primer pairs, with 80% polymorphism across all accessions studied. According to cluster analysis, 40 accessions were grouped into two major clusters, corresponding to the two species analyzed, except one case whose situation is discussed. In C. dolichopoda accessions, significant correlation between the clustering pattern and the geographical origin was detected; the extent of variation within and among its collect sites was examined by AMOVA. The knowledge about the genetic variability of the accessions examined contributes to development of Couepia conservational efforts.     

Differential light responses of Mediterranean tree saplings: linking ecophysiology with regeneration niche in four co-occurring species

The ecophysiological mechanisms underlying plant-plant interactions and forest regeneration processes in Mediterranean ecosystems are poorly understood, and the experimental evidence for the role of light availability in these processes is particularly scant. We analyzed the effects of high and low irradiances on 31 ecological, morphological and physiological variables in saplings of four late-successional Mediterranean trees, two deciduous (Acer opalus subsp. granatense (Boiss.) Font Quer & Rothm. and Quercus pyrenaica Willd.) and two evergreen (Pinus nigra Arnold subsp. salzmannii (Dunal) Franco and Quercus ilex L.), which coexist in mature montane forests. Species differed in both their capacity to withstand high radiation and in their shade tolerance. The two deciduous species were the least tolerant to high radiation, exhibiting both dynamic and chronic photoinhibition in full sunlight, with severe implications for gas exchange and photosynthetic performance. Excess light severely limited the survival of A. opalus subsp. granatense, even minor reductions of excessive radiation (from full sunlight to 80% sunlight) being crucial for sapling survival. Among species, P. nigra was the most tolerant of high irradiances but showed the poorest shade acclimation. Interspecific differences in the mechanisms of response to light provide a partial explanation of the differential regeneration patterns previously reported for these species, with shade-tolerant plants (i.e., deciduous broadleaf species) benefiting the most from associations with nurse plants. We conclude that light availability is an important environmental factor defining the regeneration niche of Mediterranean woody species.     

Solid-phase microextraction in the analysis of virgin olive oil volatile fraction: characterization of virgin olive oils from two distinct geographical areas of northern Italy

SPME was employed to characterize the volatile profile of virgin olive oils produced in two geographical areas of northern Italy: the region of the Gulf of Trieste and the area near Lake Garda. There are as yet no data on the headspace composition of virgin olive oils from these regions, characterized by particular conditions of growth for Olea europaea. Using the SPME technique coupled to GC-MS and GC-FID, the volatile components of 42 industrially produced virgin olive oil samples were identified and the principal compounds quantitatively analyzed. Significant differences in the proportion of volatile constituents from oils of different varieties and geographical origins were detected. The results suggest that besides the genetic factor, environmental conditions influence the volatile formation.     

A simple analytical method for dhurrin content evaluation in cyanogenic plants for their utilization in fodder and biofumigation

Cyanogenic plants have some potential as biocidal green manure crops in limiting several soilborne pests and pathogens. Sorghum (Sorghum bicolor (L.) Moench) and Sudangrass (Sorghum bicolor subsp. sudanense (P.) Stapf), in fact, contain the cyanogenic glucoside p-hydroxy-(S)-mandelonitrile-β-D-glucoside (dhurrin) as a substrate of its secondary defensive system able to release hydrogen cyanide following tissue lesions due to biotic or abiotic factors. Given that dhurrin content is correlated with the biofumigant efficacy of the plants, a high dhurrin content could be a positive character for utilization of sorghum and Sudangrass as biocidal green manure plants. For chemical characterization of the available germplasm, a simple, safe, and accurate method is necessary. In this paper, a new method for dhurrin analysis, based on methanol extraction and high-performance liquid chromatography, is reported and discussed. The feasibility of this analytical procedure was tested by evaluating dhurrin level in roots and stems during cultivation of four different sorghum and Sudangrass varieties in agronomic trials performed in 2008 in the Po valley (Italy). The dhurrin content ranged from 0.16 ± 0.04 to 7.14 ± 0.32 mg g(-1) on dried matter (DM) in stems and from 1.38 ± 0.02 to 6.57 ± 0.09 mg g(-1) on DM in roots, showing statistical differences among the tested germplasms that could be linked to the efficacy of their utilization as biofumigant plants. The method also opens new perspectives for the characterization of sorgum plants as fodder, for which the presence of dhurrin is considered to be negative for its well-known toxicity.     

Distribution and efficiency of Rhizobium leguminosarum strains nodulating Phaseolus vulgaris in Northern Spanish soils: Selection of native strains that replace conventional N fertilization

Phaseolus vulgaris is a legume extensively cultivated in Spain, León province being the most important producer. This province produces selected varieties of common bean highly appreciated by their quality that warrants a Protected Geographic Indication (PGI). In this work we analysed the rhizobia present in nodules of the variety “Riñón” in several soils from León province in order to select native rhizobial strains to be used as biofertilizers. The analysis of rrs and housekeeping genes of these strains showed that they belong to two phylogenetic groups within Rhizobium leguminosarum (I and II). Although the group II strains were most abundant in nodules, very effective strains were also found in group I. Strains LCS0306 from group I and LBM1123 from group II were the best nitrogen fixers among all strains isolated and were selected for field experiments. The field research showed that the biofertilization of common bean with native and selected rhizobial strains can completely replace the fertilization with chemical N fertilizers. The biofertiliser designed in such way, was valid for the whole agroecological area, regardless the specific properties of each soil and microclimatic conditions. This conclusion can be generalised as a strategy for the development of biofertilisers in different agroecological conditions worldwide.     

Clinical pharmacokinetics of a dexmedetomidine–methadone combination in dogs undergoing routine anaesthesia after buccal or intramuscular administration

This study aimed to define the pharmacokinetic profiles of dexmedetomidine and methadone administered simultaneously in dogs by either an oral transmucosal route or intramuscular route and to determine the bioavailability of the oral transmucosal administration relative to the intramuscular one of both drugs, so as the applicability of this administration route in dogs. Twelve client‐owned dogs, scheduled for diagnostic procedures, were treated with a combination of dexmedetomidine hydrochloride (10 μg/kg) and methadone hydrochloride (0.4 mg/kg) through an oral transmucosal route or intramuscularly. Oral transmucosal administration caused ptyalism in most subjects, and intramuscular administration caused transient peripheral vasoconstriction. The results showed reduced and delayed absorption of both dexmedetomidine and methadone when administered through an oral transmucosal route, with median (range) C_max values of 0.82 (0.42–1.49) ng/ml and 13.22 (2.80–52.30) ng/ml, respectively. The relative bioavailability was low: 16.34% (dexmedetomidine) and 15.5% (methadone). Intramuscular administration resulted in a more efficient absorption profile, with AUC and C_max values for both drugs approximately 10 times higher. Dexmedetomidine and methadone administered simultaneously by an oral transmucosal route using injectable formulations were not well absorbed through the oral mucosa. Nevertheless, additional studies on these drugs combination using alternative administration routes are recommended.     

Meta‐analysis of the prevalence of thermotolerant Campylobacter in food‐producing animals worldwide

The objective of this meta‐analysis was to summarize available information on the prevalence of thermotolerant Campylobacter (TC) in different food‐producing animals worldwide. Databases (i.e., PubMed, ScienceDirect, Scopus) were searched from 1980 to 2017 unrestricted by language. The inclusion criteria were as follows: prevalence or incidence studies, published in peer‐reviewed journals, and they must have reported the total number of animal samples studied and the number of samples that were positive for the presence of TC. When the identification of Campylobacter species was available, this information was included in the analysis. Multilevel random‐effect meta‐analysis models were fitted to estimate mean occurrence rate of TC and to compare them among different factors potentially associated with the outcome. The mean occurrence rate of TC in food‐producing animals was 0.424 (95% CI: 0.394–0.455), and the mean occurrence rate of Campylobacter jejuni and Campylobacter coli were 0.214 and 0.133, respectively. Pigs and poultry showed the highest prevalence of TC; however, there were differences in the prevalence of each Campylobacter species. Campylobacter jejuni was observed in broilers (0.322; 95% CI: 0.273–0.377) and hens (0.395; 95% CI: 0.265–0.542), while C. coli was restricted essentially in pigs (0.553; 95% CI: 0.541–0.650). The prevalence of C. jejuni in intensively bred cattle was higher (0.302; 95% CI: 0.227–0.389) than the prevalence in extensively bred cattle (0.172; 95% CI: 0.119–0.242) while the prevalence of C. coli was similar (0.051; 95% CI: 0.028–0.091 vs. 0.050; 95% CI: 0.027–0.091) in both production systems. Agar with or without blood used for the isolation of TC did not affect the prevalence observed. The method of species identification did not seem to generate differences in the prevalence of Campylobacter species. The prevalence of Campylobacter in primary food production has a strong impact on the entire agri‐food chain. National authorities must monitor the situation with the aim to establish the appropriate risk management measures.