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Tebuconazole (TBZ) is a widely used triazole fungicide at EU level on cereals and vines. It is relatively persistent in soil where it is transformed to various transformation products (TPs) which might be environmentally relevant. We assessed the dissipation of TBZ in soil under contrasting incubation conditions (standard vs winter simulated) that are relevant to its application scheme, determined its transformation pathway using advanced analytical tools and 14 C-labeled TBZ and assessed its soil microbial toxicity. Mineralization of 14 C-triazole-ring-labeled TBZ was negligible but up to 11% of 14 C-penyl-ring-labeled TBZ evolved as 14 CO 2 within 150 days of incubation. TBZ persistence increased at higher dose rates (×10 compared to the recommended agronomical dose ×1) and under winter simulated conditions compared to standard incubation conditions (at ×1 dose rate DT 50 of 202 and 88 days, respectively). Non-target suspect screening enabled the detection of 22 TPs of TBZ, among which 17 were unknown. Mass spectrometry analysis led to the identification of 1-(4-chlorophenyl) ethanone, a novel TP of TBZ, the formation of which and decay in soil was determined by gas chromatography mass spectrometry. Three hypothetical transformation pathways of TBZ, all converging to 1H-1,2,4-triazole are proposed based on suspect screening. The ecotoxicological effect of TBZ and of its TPs was assessed by measuring by qPCR the abundance of the total bacteria and the relative abundance of 11 prokaryotic taxa and 4 functional groups. A transient impact of TBZ on the relative abundance of all prokaryotic taxa (except α-proteobacteria and Bacteroidetes) and one functional microbial group (pcaH-carrying microorganisms) was observed. However the direction of the effect (positive or negative) varied, and in certain cases, depended on the incubation conditions. Proteobacteria was the most responsive phylum to TBZ with recovery observed 20 days after treatment. The ecotoxicological effects on the soil microorganisms were not correlated with 1-(4-chlorophenyl) ethanone., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Silver-induced selective pressure is becoming increasingly important due to the growing use of silver (Ag) as an antimicrobial agent in biomedical and commercial products. With demonstrated links between environmental resistomes and clinical pathogens, it is important to identify microbial profiles related to silver tolerance/resistance. We investigated the effects of ionic Ag stress on soil bacterial communities and identified resistant/persistent bacterial populations. Silver treatments of 50-400 mg Ag kg(-1) soil were established in five soils. Chemical lability measurements using diffusive gradients in thin-film devices confirmed that significant (albeit decreasing) labile Ag concentrations were present throughout the 9-month incubation period. Synchrotron X-ray absorption near edge structure spectroscopy demonstrated that this decreasing lability was due to changes in the Ag speciation to less soluble forms such as Ag(0) and Ag2S. Real-time PCR and Illumina MiSeq screening of 16S rRNA bacterial genes showed β-diversity changes, increasing α-diversity in response to Ag pressure, and immediate and significant reductions in 16S rRNA gene counts with varying degrees of recovery. These effects were more strongly influenced by exposure time than by Ag dose at these rates. Ag-selected dominant OTUs principally resided in known persister taxa (mainly Gram positive), including metal-tolerant bacteria and slow-growing Mycobacteria., (© FEMS 2015. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

The Po river plain lowland springs represent unique paradigms of managed environments. Their current locations used to be swamps that were drained 6-7 centuries ago, and they have been in constant use ever since. Our aims were to identify the effects of land use on the microbial communities of these soils, look for associated diversity drivers, and assess the applicability of ecology theories with respect to identified patterns. We screened the microbial diversity across a land use transect via high-throughput sequencing of partial 16S rrRNA gene amplicons. Land use had a major effect on soil properties and microbial community structures. Total organic carbon and pH were major diversity drivers for Bacteria, and pH was important for Archaea. We identified the potential contribution of soil amendments to the indigenous microbial communities, and also gained insights into potential roles of taxa in the organic carbon turnover. Verrucomicrobia coincided with the higher values of the recalcitrant organic carbon. Actinobacteria and Acidobacteria correlated with the more labile organic carbon. Finally, the higher diversity found in the soils less enzymatically active and relatively poorer in nutrients, may be explained to an extent by niche-based theories such as the resource heterogeneity hypothesis and Connell's intermediate disturbance hypothesis., (© 2013 Federation of European Microbiological Societies. Published by John Wiley & Sons Ltd. All rights reserved.)

Ammonia oxidation has been intensively studied for its sensitivity to environmental shifts and stresses. However, acute stress effects on the occurrence and composition of ammonia oxidizing bacteria (AOB) and archaea (AOA) based on expression of related molecular markers in complex soil environments have been to an extent overlooked, particularly concerning transient but commonly occurring environmental changes like soil moisture shifts. The present study investigates the responses of AOB and AOA to moisture shifts and high Zn soil content. AmoA gene copies and transcripts of AOB and AOA along with potential nitrification activity were measured in a soil microcosm approach for investigating the referred environmental shifts. Moisture change from 87 to 50 % of the water holding capacity caused a ~99 % reduction of AOB but not of AOA amoA transcripts that did not change significantly. Increasing applied zinc concentrations resulted in a reduction of potential nitrification rates and negatively affected studied gene expressions of both AOB and AOA, with AOB being more responsive. Both 16 S rRNA and amoA transcripts of AOB had an inverse relation to the applied zinc, indicating a gradual loss in total cell activity. Our results suggest the existence of pronounced differences between AOB and AOA concerning ammonia oxidation activity.

Litter soil cover constitutes an important micro-ecosystem in sustainable viticulture having a key role in nutrient cycling and serving as a habitat of complex microbial communities. Ammonia-oxidizing bacteria (AOB) and archaea (AOA) are known to regulate nitrification in soil while little is known regarding their function and diversity in litter. We investigated the effects of two fungicides, penconazole and cyprodinil, commonly used in vineyards, on the function and diversity of total and active AOB and AOA in a microcosm study. Functional changes measured via potential nitrification and structural changes assessed via denaturating gradient gel electrophoresis (DGGE) at the DNA and RNA levels were contrasted with pesticide dissipation in the litter layer. The latter was inversely correlated with potential nitrification, which was temporarily inhibited at the initial sampling dates (0 to 21 days) when nearly 100 % of the applied pesticide amounts was still present in the litter. Fungicides induced changes in AOB and AOA communities with RNA-DGGE analysis showing a higher sensitivity. AOA were more responsive to pesticide application compared to AOB. Potential nitrification was less sensitive to the fungicides and was restored faster than structural changes, which persisted. These results support the theory of microbial redundancy for nitrification in a stressed litter environment.

The novel multi-million read generating sequencing technologies are very promising for resolving the immense soil 16S rRNA gene bacterial diversity. Yet they have a limited maximum sequence length screening ability, restricting studies in screening DNA stretches of single 16S rRNA gene hypervariable (V) regions. The aim of the present study was to assess the effects of properties of four consecutive V regions (V3-6) on commonly applied analytical methodologies in bacterial ecology studies. Using an in silico approach, the performance of each V region was compared with the complete 16S rRNA gene stretch. We assessed related properties of the soil derived bacterial sequence collection of the Ribosomal Database Project (RDP) database and concomitantly performed simulations based on published datasets. Results indicate that overall the most prominent V region for soil bacterial diversity studies was V3, even though it was outperformed in some of the tests. Despite its high performance during most tests, V4 was less conserved along flanking sites, thus reducing its ability for bacterial diversity coverage. V5 performed well in the non-redundant RDP database based analysis. However V5 did not resemble the full-length 16S rRNA gene sequence results as well as V3 and V4 did when the natural sequence frequency and occurrence approximation was considered in the virtual experiment. Although, the highly conserved flanking sequence regions of V6 provide the ability to amplify partial 16S rRNA gene sequences from very diverse owners, it was demonstrated that V6 was the least informative compared to the rest examined V regions. Our results indicate that environment specific database exploration and theoretical assessment of the experimental approach are strongly suggested in 16S rRNA gene based bacterial diversity studies.

The effects of a humic acid (HA) and its size-fractions on plants carbon deposition and the structure of microbial communities in the rhizosphere soil of maize (Zea mays L.) plants were studied. Experiments were conducted in rhizobox systems that separate an upper soil-plant compartment from a lower compartment, where roots are excluded from the rhizosphere soil by a nylon membrane. The upper rhizobox compartment received the humic additions, whereas, after roots development, the rhizosphere soil in the lower compartment was sampled and sliced into thin layers. The lux-marked biosensor Pseudomonas fluorescens 10586 pUCD607 biosensor showed a significant increase in the deposition of bioavailable sources of carbon in the rhizosphere of soils when treated with bulk HA, but no response was found for treatments with the separated size-fractions. PCR-DGGE molecular fingerprintings revealed that the structure of rhizosphere microbial communities was changed by all humic treatments and that the smaller and more bioavailable size-fractions were more easily degraded by microbial activity than the bulk HA. On the other hand, highly hydrophobic and strongly associated humic molecules in the bulk HA required additional plant rhizodeposition before their bio-transformation could occur. This work highlights the importance of applying advanced biological and biotechnological methods to notice changes occurring in plant rhizodeposition and rhizosphere microbial activity. Moreover, it suggests correlations between the molecular properties of humic matter and their effects on microbial communities in the rhizosphere as mediated by root exudation.

Pesticides are intentionally applied to agricultural fields for crop protection. They can harm non-target organisms such as soil microorganisms involved in important ecosystem functions with impacts at the global scale. Within the frame of the pesticide registration process, the ecotoxicological impact of pesticides on soil microorganisms is still based on carbon and nitrogen mineralization tests, despite the availability of more extensive approaches analyzing the abundance, activity or diversity of soil microorganisms. In this study, we used a high-density DNA microarray (PhyloChip) and 16S rDNA amplicon next-generation sequencing (NGS) to analyze the impact of the organophosphate insecticide chlorpyrifos (CHL), the phenyl-urea herbicide isoproturon (IPU), or the triazole fungicide tebuconazole (TCZ) on the diversity and composition of the soil bacterial community. To our knowledge, it is the first time that the combination of these approaches are applied to assess the impact of these three pesticides in a lab-to-field experimental design. The PhyloChip analysis revealed that although no significant changes in the composition of the bacterial community were observed in soil microcosms exposed to the pesticides, significant differences in detected operational taxonomic units (OTUs) were observed in the field experiment between pesticide treatments and control for all three tested pesticides after 70 days of exposure. NGS revealed that the bacterial diversity and composition varied over time. This trend was more marked in the microcosm than in the field study. Only slight but significant transient effects of CHL or TCZ were observed in the microcosm and the field study, respectively. IPU was not found to significantly modify the soil bacterial diversity or composition. Our results are in accordance with conclusions of the Environmental Food Safety Authority (EFSA), which concluded that these three pesticides may have a low risk toward soil microorganisms. [ABSTRACT FROM AUTHOR]

The authors provide an updated and integrated view of the adaptation of soil microorganisms to elevated concentrations of trace elements. Starting with a summary of the occurrence of trace elements in soils and their effects on soil microorganisms, the scientific evidence underlying adaptation of microorganisms to trace elements from species to community level is discussed. Insights are given regarding the main physiological processes involved in the resistance of bacteria to toxic elements including the potential importance of horizontal gene transfer in the adaptation process. The review continues with a discussion of how new molecular and biotechnological techniques can enrich this field of study. Scientific evidence is utilized in constructing an illustration of microbial community responses with reference to ecological indicators during various adaptation stages, while the related effects on community biological functionality and resilience are discussed. The authors conclude with an evaluation of the importance of considering adaptation in risk assessment and possible remediation of trace element–contaminated sites. [ABSTRACT FROM PUBLISHER]

Assessment of dissipation constitutes an integral part of pesticides risk assessment since it provides an estimate of the level and the duration of exposure of the terrestrial ecosystem to pesticides. Within the frame of an overall assessment of the soil microbial toxicity of pesticides, we investigated the dissipation of a range of dose rates of three model pesticides, isoproturon (IPU), tebuconazole (TCZ), and chlorpyrifos (CHL), and the formation and dissipation of their main transformation products following a tiered lab-to-field approach. The adsorption of pesticides and their transformation products was also determined. IPU was the least persistent pesticide showing a dose-dependent increase in its persistence in both laboratory and field studies. CHL dissipation showed a dose-dependent increase under laboratory conditions and an exact opposite trend in the field. TCZ was the most persistent pesticide under lab conditions showing a dose-dependent decrease in its dissipation, whereas in the field TCZ exhibited a biphasic dissipation pattern with extrapolated DT 90s ranging from 198 to 603.4 days in the × 1 and × 2 dose rates, respectively. IPU was demethylated to mono- (MD-IPU) and di-desmethyl-isoproturon (DD-IPU) which dissipated following a similar pattern with the parent compound. CHL was hydrolyzed to 3,5,6-trichloro-2-pyridinol (TCP) which dissipated showing a reverse dose-dependent pattern compared to CHL. Pesticides adsorption affinity increased in the order IPU < TCZ < CHL. IPU transformation products showed low affinity for soil adsorption, whereas TCP was weakly adsorbed compared to its parent compound. The temporal dissipation patterns of the pesticides and their transformation products will be used as exposure inputs for assessment of their soil microbial toxicity. [ABSTRACT FROM AUTHOR]

Abstract: The influence of humic substances (HS) on crops and soil microbial communities still lacks a complete understanding. The influence of HS on chemical composition of maize rhizodeposits and diversity of rhizosphere microbial communities was studied here in rhizobox systems. Eleven different HS were characterized by elemental analyses and 13C-CPMAS NMR, and applied to the upper compartment of rhizobox systems cultivated with maize. Once a full rhizoplane was formed, the lower compartment of the rhizoboxes was sampled with a slicing device, in order to obtain rhizopshere (0–2mm) and bulk (>10mm) soil samples, which were subjected to chemical analyses of organic acids and sugars (pentoses and hexoses), bioassays with a luminescent biosensor for the detection and quantification of bioavailable C forms, and analyzed for bacterial diversity by means of PCR-DGGE fingerprint. Multivariate techniques were applied to assess correlations between the parameters involved and to assess the most relevant chemical drivers of rhizosphere microbial diversity. Results showed that the hydrophilic and poorly aromatic HS changed the pattern of sugars and organic acids released in the rhizosphere by maize roots, and this, in turn, affected the composition of rhizosphere microbial communities. Differences in chemical and microbiological composition were also found, but to a lesser extent, among bulk soil samples of different treatments, thus indicating that rhizodeposits can influence soil composition at several mm distance from the root plane. Multiple regression analyses suggested a prominent role of succinic and malic acid in shaping the diversity of microbial communities. The approach and results described here proved to be useful to test bioactivity of HS towards an agricultural crop, and identify correlations between chemical and microbiological constituents at the tri-phase soil–roots–microbes system. [Copyright &y& Elsevier]

Abstract: This study investigated the responses to Zn of selected enzymatic activities, potential nitrification and microbial activity in a soil contaminated with Zn at concentrations ranging from 74 to 3490 mg kg−1. The main aims of the study were to evaluate different models for EC50 estimation and to compare the relative sensitivity to Zn of the different biological properties studied. The EC50 for each property was calculated using three different models, namely a logistic dose response model, and a full and a partial inhibition model based on Michaelis–Menten kinetics. Both total (aqua regia extractable) and labile (CaCl2 extractable) Zn content were assessed and employed for application of the models. A response to increasing Zn concentrations was observed for most of the biological properties except for fluorescein diacetate hydrolysis activity (FDA) and acid phosphatase activity. Among the three models tested, the logistic model had the widest applicability and generally gave the best results in terms of EC50 estimation and R 2 fit. Among the two kinetics models tested, the partial inhibition model was applicable only in a few cases, but in these cases results are generally better than for the full inhibition model. The utilization of either total or labile Zn data did not significantly affect the outcomes of the models. The sensitivity of the biological properties to Zn was ranked according to the models as follows: nitrate reductase > potential nitrification > β-galactosidase > dehydrogenase > phenol oxidase = urease = arylsulphatase. These ranking results were generally in accordance with evidence provided by other researchers, thus showing that the relative sensitivity of these properties, especially the most sensitive ones, is generally constant among different soil types. The relevance of this finding for soil ecological risk assessment is discussed. [Copyright &y& Elsevier]

Abstract: Phospholipid fatty acid (PLFA) analysis has gained great importance in the study of soil microbial community structure. This structure can give indication of the soil status. Purpose of the present paper is to analyse PLFA patterns in altered agricultural soils in order to develop a soil status alteration index. Soils subjected either to intensive agricultural exploitation, or to overflow by municipal and industrial wastes, or to irrigation with saline waters were analysed for PLFA content and compared to adjacent untreated soils by means of different statistical techniques. Principal component analysis separated PLFAs in three groups: unsaturated PLFAs (first axis, 48% of total variance), monounsaturated and cyclopropane PLFAs (second axis, 28% of total variance) and polyunsaturated PLFAs (third axis, 24% of total variance). By means of canonical discriminant analysis, a soil alteration index (SAI) was produced from 15 PLFAs using two data sets. A third data set was used to test the SAI general validity together with other data sets reported in literature. The index validity was confirmed in most cases: SAI gave higher scores for control soils and was generally able to classify soils according to their reported degree of alteration. [Copyright &y& Elsevier]