Multiplex PCR for Assessment of Tetracycline Resistance (tet) Genes in Antibiotic-Resistant Soil Bacteria and the Ecological Implications

Antibiotics are becoming increasingly prevalent in bacterial communities due to clinical and agricultural misuse and overuse in their environment. As exposure increases, so does the incidence of microbial resistance. Such is the case with bacterial resistance to tetracyclines, a phenotype often acquired through the horizontal gene transfer of tet genes between bacteria. The objective of this project was to analyze the bacterial diversity of tet resistance genes in soil from Miami-Dade County. Bacterial isolates were Gram-stained and the Kirby-Bauer antibiotic disk diffusion test was performed to determine each bacterium’s degree of resistance. The 16S rRNA gene from antibiotic-resistant isolates was amplified by PCR and sequenced to identify the isolates. All isolates’ tet genes were amplified by multiplex PCR, sequenced, and compared. Among eight isolates, three distinct species were positively identified based on their 16S rRNA sequences and four distinct tet genes were identified, though all tested susceptible to tetracycline via the Kirby-Bauer test. This project clarifies some aspects of the ecology of antibiotic resistance genes, their natural ecological function and the potential for the expansion of intrinsic multi-antibiotic resistance into new ecosystems and/or hosts.

Centrifuge modeling of oblique pipe-soil interaction in dense and loose sand

Due to relative ground movement, buried pipelines experience geotechnical loads. The imposed geotechnical loads may initiate pipeline deformations that affect system serviceability and integrity. Engineering guidelines (e.g., ALA, 2005; Honegger and Nyman, 2001) provide the technical framework to develop idealized structural models to analyze pipe‒soil interaction events and assess pipe mechanical response. The soil behavior is modeled using discrete springs that represent the geotechnical loads per unit pipe length developed during the interaction event. Soil forces are defined along three orthogonal directions (i.e., axial, lateral and vertical) to analyze the response of pipelines. Nonlinear load-displacement relationships of soil defined by a spring, is independent of neighboring spring elements. However, recent experimental and numerical studies demonstrate significant coupling effects during oblique (i.e., not along one of the orthogonal axes) pipe‒soil interaction events. In the present study, physical modeling using a geotechnical centrifuge was conducted to improve the current understanding of soil load coupling effects of buried pipes in loose and dense sand. A section of pipeline, at shallow burial depth, was translated through the soil at different oblique angles in the axial-lateral plane. The force exerted by the soil on pipe is critically examined to assess the significance of load coupling effects and establish a yield envelope. The displacements required to soil yield force are also examined to assess potential coupling in mobilization distance. A set of laboratory tests were conducted on the sand used for centrifuge modeling to find the stress-strain behavior of sand, which was used to examine the possible mechanisms of centrifuge model test. The yield envelope, deformation patterns, and interpreted failure mechanisms obtained from centrifuge modeling are compared with other physical modeling and numerical simulations available in the literature.

Mobility and toxicity of heavy metal(loid)s arising from contaminated wood ash application to a pasture grassland soil

The authors acknowledge the financial support the Scottish Government’s Rural and Environmental Sciences and Analytical Services (RESAS) in order to complete some of the soil and pore water sample analysis as well as the Czech Ministry of Education, Youth and Sports (COST CZ LD13068), the Czech Science Foundation (GAČR 14-02183P) and EU COST actionFP1407 (‘ModWoodLife’) short term scientific mission grant in order to complete the column leaching test.

Table 1. Typical soil sections, Centrum Sø

During the period May to August 1960, an Air Force scientific field party conducted earth science studies and tested a raised sand terrace, located about 224 km south of Station Nord, Northeast Greenland. The operation staged from Thule Air Force Base was climaxed by successful test Iandings on the terrace by C-119 and C-130 aircraft. Significant data were obtained from related investigations on a typical arctic lake, ice-free soils, meteorology, engineering geology, geomorphology, and electrical resistivity of soils.

Tab. 1+2+3: Selected principal soil properties in the oil exploitation region of KomiArcticOil (Usinsk) in the Russian tundra

Oil polluted and not oil polluted soils (crude oil hydrocarbons contents: 20-92500 mg/kg dry soil mass) under natural grass and forest vegetation and in a bog in the Russian tundra were compared in their principal soil ecological parameters, the oil content and the microbial indicators. CFE biomass-C, dehydrogenase and arylsulfatase activity were enhanced with the occurrence of crude oil. Using these parameters for purposes of controlling remediation and recultivation success it is not possible to distinguish bctween promotion of microbial activity by oil carbon or soil organic carbon (SOC). For this reason we think that these parameters are not appropriate to indicate a soil damage by an oil impact. In contrast the metabolie quotient (qC02), calculated as the ratio between soil basal respiration and the SIR biomass-C was adequate to indicate a high crude oil contamination in soil. Also, the ß-glucosidase activity (parameter ß-GL/SOC) was correlated negatively with oil in soil. The indication of a soil damage by using the stress parameter qCO, or the specific enzyme activities (activity/SOC) minimizes the promotion effect of the recent SOC content on microbial parameters. Both biomass methods (SIR, CFE) have technical problems in application for crude oil-contaminated and subarctic soils. CFE does not reflect the low C_mic level of the cold tundra soils. We recommend to test every method for its suitability before any data collection in series as well as application for cold soils and the application of ecophysiological ratios as R_mic/C_mic, C_mic/SOC or enzymatic activity/SOC instead of absolute data.

Undergrowth removal in Salgesch: Soil volumetric water content and temperature time series, hourly time resolution, 2010-2015

In the Salgesch forest in the Canton of Valais in Switzerland, the understory has been removed to test whether effects on pine tree vitality. The data set published here compromises 120 time series of 60 soil temperature and 60 volumetric water content (VWC) sensors (EC-TM and 5-TM) (Decagon Devices, WA, USA) at three soil depth levels (5, 30, 60 cm) employed in the direct vicinity of six control trees and six trees with the undergrowth removed. At the levels 5 and 60 cm, three replications were made whereas 4 replications were made at level 30 cm. Six loggers recorded hourly data since 2010 with 18% gaps or 11% when not considering winter months December, January and February. The figure attached to this repository shows the average VWC and temperature of all measurements within the same depth and treatment specific setting aggregated in a defined time interval and period. In addition to that, the standard deviations are plotted as transparent polygons. In case of insufficient values for calculating standard deviations, the setting specific mean standard deviation of the considered time period are inserted.

The impact of organic amendments on soil properties under mediterranean climatic conditions

Soil erosion and unsustainable land use produce adverse effects on SOC content. Soil management techniques and corrections can be applied for soil recovery, especially, with afforestaion purposes. This study presents the short term effects on the application of different treatments on soil properties for soil included in several sets of closed plots located in the experimental area of Pinarillo (Nerja, Spain). The analysed soil properties were: PH, EC, organic carbon, total nitrogen and total carbon. In order to verify possible differences, we applied the test of Mann-Whitney U in corroboration with the previous homogeneity test of variance.

Effect of particle loss on soil behaviour.

Soil particle loss can result in strength and volume reductions which are difficult to predict. This paper investigates the influence of the removal of fractions of selected particle sizes under different confining pressures. The mass loss process was reproduced by the dissolution of selected salt particle sizes and fractions from uniform Leighton Buzzard sand. The dissolution tests were performed in a triaxial cell customised to allow circulation of pore-fluid thereby allowing the dissolution/removal of the salt fraction. Test results from previously conducted oedometric dissolution tests and subsequent triaxial dissolution tests all show increases in void ratio. From the triaxial tests, a reduction in shear strength with increasing ductility was observed. Volumetric and strength behaviour were found to be related to the particle size and fraction material removed while shear-wave measurements obtained pre- and post-particle removal indicate significant changes in small-strain stiffness.

Effects of Soil Organic Matter Properties and Microbial Community Composition on Enzyme Activities in Cryoturbated Arctic Soils

Enzyme-mediated decomposition of soil organic matter (SOM) is controlled, amongst other factors, by organic matter properties and by the microbial decomposer community present. Since microbial community composition and SOM properties are often interrelated and both change with soil depth, the drivers of enzymatic decomposition are hard to dissect. We investigated soils from three regions in the Siberian Arctic, where carbon rich topsoil material has been incorporated into the subsoil (cryoturbation). We took advantage of this subduction to test if SOM properties shape microbial community composition, and to identify controls of both on enzyme activities. We found that microbial community composition (estimated by phospholipid fatty acid analysis), was similar in cryoturbated material and in surrounding subsoil, although carbon and nitrogen contents were similar in cryoturbated material and topsoils. This suggests that the microbial community in cryoturbated material was not well adapted to SOM properties. We also measured three potential enzyme activities (cellobiohydrolase, leucine-amino-peptidase and phenoloxidase) and used structural equation models (SEMs) to identify direct and indirect drivers of the three enzyme activities. The models included microbial community composition, carbon and nitrogen contents, clay content, water content, and pH. Models for regular horizons, excluding cryoturbated material, showed that all enzyme activities were mainly controlled by carbon or nitrogen. Microbial community composition had no effect. In contrast, models for cryoturbated material showed that enzyme activities were also related to microbial community composition. The additional control of microbial community composition could have restrained enzyme activities and furthermore decomposition in general. The functional decoupling of SOM properties and microbial community composition might thus be one of the reasons for low decomposition rates and the persistence of 400 Gt carbon stored in cryoturbated material.

Small strain elastic behaviour of unsaturated soil investigated by bender/extender element testing

The aim of this project was to investigate very small strain elastic behaviour of soils under unsaturated conditions, using bender/extender element (BEE) testing. The behaviour of soils at very small strains has been widely studied under saturated conditions, whereas much less work has been performed on very small strain behaviour under unsaturated conditions. A suction-controlled double wall triaxial apparatus for unsaturated soil testing was modified to incorporate three pairs of BEEs transmitting both shear and compression waves with vertical and horizontal directions of wave transmission and wave polarisation. Various different techniques for measuring wave travel time were investigated in both the time domain and the frequency domain and it was concluded that, at least for the current experimental testing programme, peak-to-first-peak in the time domain was the most reliable technique for determining wave travel time. An experimental test programme was performed on samples of compacted speswhite kaolin clay. Two different forms of compaction were employed (i.e. isotropic and anisotropic). Compacted kaolin soil samples were subjected to constant suction loading and unloading stages at three different values of suction, covering both unsaturated conditions (s= 50kPa and s= 300kPa) and saturated conditions (s=0). Loading and unloading stages were performed at three different values of stress ratio (η=0, η=1 and η=-1 ). In some tests a wetting-drying cycle was performed before or within the loading stage, with the wetting-drying cycles including both wetting-induced swelling and wetting-induced collapse compression. BEE tests were performed at regular intervals throughout all test stages, to measure shear wave velocity Vs and compression wave velocity Vp and hence to determine values of shear modulus G and constrained modulus M. The experimental test programme was designed to investigate how very small strain shear modulus G and constrained modulus M varied with unsaturated state variables, including how anisotropy of these parameters developed either with stress state (stress-induced anisotropy) or with previous straining (strain-induced anisotropy). A new expression has been proposed for the very small strain shear modulus G of an isotropic soil under saturated and unsaturated conditions. This expression relates the variation of G to only mean Bishop’s stress p* and specific volume v, and it converges to a well-established expression for saturated soils as degree of saturation approaches 1. The proposed expression for G is able to predict the variation of G under saturated and unsaturated conditions at least as well as existing expressions from the literature and it is considerably simpler (employing fewer state variables and fewer soil constants). In addition, unlike existing expressions from the literature, the values of soil constants in the proposed new expression can be determined from a saturated test. It appeared that, in the current project at least, any strain-induced anisotropy of very small strain elastic behaviour was relatively modest, with the possible exception of loading in triaxial extension. It was therefore difficult to draw any firm conclusion about evolution of strain-induced anisotropy and whether it depended upon the same aspects of soil fabric as evolution of anisotropy of large strain plastic behaviour. Stress-induced anisotropy of very small strain elastic behaviour was apparent in the experimental test programme. An attempt was made to extend the proposed expression for G to include the effect of stress-induced anisotropy. Interpretation of the experimental results indicated that the value of shear modulus was affected by the values of all three principal Bishop’s stresses (in the direction of wave transmission, the direction of wave polarisation and the third mutually perpendicular direction). However, prediction of stress-induced anisotropy was only partially successful, and it was concluded that the effect of Lode angle was also significant.

Microbial nitrogen dynamics in organic and mineral soil horizons along a latitudinal transect in western Siberia

Soil N availability is constrained by the breakdown of N-containing polymers such as proteins to oligopeptides and amino acids that can be taken up by plants and microorganisms. Excess N is released from microbial cells as ammonium (N mineralization), which in turn can serve as substrate for nitrification. According to stoichiometric theory, N mineralization and nitrification are expected to increase in relation to protein depolymerization with decreasing N limitation, and thus from higher to lower latitudes and from topsoils to subsoils. To test these hypotheses, we compared gross rates of protein depolymerization, N mineralization and nitrification (determined using N-15 pool dilution assays) in organic topsoil, mineral topsoil, and mineral subsoil of seven ecosystems along a latitudinal transect in western Siberia, from tundra (67 degrees N) to steppe (54 degrees N). The investigated ecosystems differed strongly in N transformation rates, with highest protein depolymerization and N mineralization rates in middle and southern taiga. All N transformation rates decreased with soil depth following the decrease in organic matter content. Related to protein depolymerization, N mineralization and nitrification were significantly higher in mineral than in organic horizons, supporting a decrease in microbial N limitation with depth. In contrast, we did not find indications for a decrease in microbial N limitation from arctic to temperate ecosystems along the transect. Our findings thus challenge the perception of ubiquitous N limitation at high latitudes, but suggest a transition from N to C limitation of microorganisms with soil depth, even in high-latitude systems such as tundra and boreal forest.

Analyses of the community compositions of root rot pathogenic fungi in the soybean rhizosphere soil

Soybean ( Glycine max [L.] Merr.) root rot is an important disease of soybean under continuous cropping, and root rot is widely distributed throughout the world. This disease is extremely harmful, and it is difficult to prevent and control. The study aimed to elucidate the composition of root rot pathogenic fungal communities in the continuous cropping of soybean. In this study, we employed PCRDGGE technology to analyze the communities of root rot pathogenic fungi in soybean rhizosphere soil subjected to continuous cropping during a season with a high incidence of root rot in Heilongjiang province, China, the main soybean producing area in China. The results of 13 DGGE bands were analyzed by phylogenetic revealed that the predominant root rot pathogenic fungi in rhizosphere soil in the test area were Pythium ultimum and Fusarium species. The results of cluster analysis showed that the duration of continuous cropping, the soybean variety and the plant growth stage all had significant effects on the diversity of root rot pathogenic fungi in rhizosphere soil.

Isolation of a potential anticancer agent with protein phosphatase inhibitory activity from soil-derived Penicillium sp strain H9318

Purpose: To determine the effect of the secondary metabolites from Penicillium sp. H9318 on cytotoxicity and cell cycle progression. Methods: A yeast PP1 inhibitory screening system was carried out to confirm the presence of anti- PP1c activity in crude acetone extracts of strain H9318. The extracts were fractionated and identified as Fraction S1 and Citrinin 9318 (CTN9318). Various cancer cell lines were used to test for the toxicity of the crude acetone extracts, Fraction S1 and Citrinin 9318, using MTT viability assay. Results: It was found that a colorectal cancer cell line, HT-29, was susceptible to Fraction S1 and Citrinin 9318. A propidium iodide (PI)-incorporated DNA assay was used to show that there was G2/M arrest in HT-29 by Citrinin 9318. Conclusion: Citrinin 9318 inhibits the viability of HT-29 via mitotic block. The results suggest that Citrinin 9318 is capable of exerting cytotoxicity and mitotic arrest in a colon cancer cell line, HT29

Impact of HydroPolymers on the soil biological components in mediterranean drylands

Soil degradation affects more than 52 million ha of land in counties of the European Union. This problem is particularly serious in Mediterranean areas, where the effects of anthropogenic activities (tillage on slopes, deforestation, and pasture production) add to problems caused by prolonged periods of drought and intense and irregular rainfall. Soil microbiota can be used as an indicator of the soil healthy in degraded areas. This is because soil microbiota participates in the cycle elements and in the organic matter decomposition. All this helps to the young plants establishment and in long term protect the soils against the erosion. During dry periods in the Mediterranean areas, the lack of water entering the soil matrix leads to a loss of soil microbiological activity and it turns into a lower soil production capabilities. Under these conditions, the aim of this study was to evaluate the positive effect on soil biological components produced by an hydro absorbent polymer (Terracottem). The aim of the experiment was to evaluate the impact assessment of an hydropolymer (Terracottem) on the soil biological components. An experimental flowerpot layout was established in June 2015 and 12 variants with different amount of Terracottem were applied as follow: i) 3.0 kg.m3 ; ii) 1.5 kg.m3 and; iii) 0 kg.m3. In all the variants were tested the further additives: a) 1% of glucose, b) 50 kg N.ha-1 of Mineral nitrogen, c) 1% of Glucose + 50 kg N.ha-1 of Mineral nitrogen d) control (no additive). According to natural conditions, humidity have been kept at 15% in all the variants. During four weeks, mineral nitrogen leaching and soil respiration have been measured in each flowerplot. Respiration has been quantified four times every time while moistening containers and alkaline soda lime has been used as a sorbent. The amount of CO 2 increase has been measured with the sorbent. Leaching of mineral nitrogen has been quantified by ion exchange resins (IER). IER pouches have been placed on the bottom of each container, and after completion of the experiment mineral nitrogen leaching has been evaluated by distillation and titration method. Results from respiration have shown statistically significant differences between the variants. According to control, soil with polymers have shown significant difference when comparing respiration with independence of the additive used. CO 2 production in the first week has exceeded the sum of the outputs of the following weeks. Mineral nitrogen leaching measurement has shown statistically significant differences. The lowest leaching has been occurred in control variant, while the highest in variant containing only the addition of mineral nitrogen. Research results may conclude that the biological part of the test soil is not limited by a lack of components, the only thing that suppresses its activity is the lack of moisture. After moistening it leads to a rapid growth of soil activity, without causing the nutrients loss. Besides, Terracottem has affected soil activity neither positively nor negatively, but it considers being a suitable tool for reducing the drought impact in arid and semi-arid areas.

SAMPLING BIAS IN EVALUATING THE PROBABILITY OF SEISMICALLY INDUCED SOIL LIQUEFACTION WITH SPT & CPT CASE HISTORIES

Several deterministic and probabilistic methods are used to evaluate the probability of seismically induced liquefaction of a soil. The probabilistic models usually possess some uncertainty in that model and uncertainties in the parameters used to develop that model. These model uncertainties vary from one statistical model to another. Most of the model uncertainties are epistemic, and can be addressed through appropriate knowledge of the statistical model. One such epistemic model uncertainty in evaluating liquefaction potential using a probabilistic model such as logistic regression is sampling bias. Sampling bias is the difference between the class distribution in the sample used for developing the statistical model and the true population distribution of liquefaction and non-liquefaction instances. Recent studies have shown that sampling bias can significantly affect the predicted probability using a statistical model. To address this epistemic uncertainty, a new approach was developed for evaluating the probability of seismically-induced soil liquefaction, in which a logistic regression model in combination with Hosmer-Lemeshow statistic was used. This approach was used to estimate the population (true) distribution of liquefaction to non-liquefaction instances of standard penetration test (SPT) and cone penetration test (CPT) based most updated case histories. Apart from this, other model uncertainties such as distribution of explanatory variables and significance of explanatory variables were also addressed using KS test and Wald statistic respectively. Moreover, based on estimated population distribution, logistic regression equations were proposed to calculate the probability of liquefaction for both SPT and CPT based case history. Additionally, the proposed probability curves were compared with existing probability curves based on SPT and CPT case histories.