Sensing of gene expression in live cells using electrical impedance spectroscopy and DNA-functionalized gold nanoparticles.

A novel electrical impedance spectroscopy-based method for non-destructive sensing of gene expression in living cells is presented. The approach used takes advantage of the robustness and responsiveness of electrical impedance spectroscopy and the highly specific and selective nature of DNA hybridization. The technique uses electrical impedance spectroscopy and gold nanoparticles functionalized with single-stranded DNA complementary to an mRNA of interest to provide reliable, real-time, and quantifiable data on gene expression in live cells. The system was validated by demonstrating specific detection of the uidA mRNA, which codes for the ß-glucuronidase (GUS) enzyme, in Solanum lycopersicum MsK8 cells. Gold nanoparticles were functionalized with single-stranded DNA oligonucleotides consisting of either a sequence complementary to uidA mRNA or an arbitrary sequence. The DNA-functionalized gold nanoparticles were mixed with cell suspensions, allowing the gold nanoparticles to penetrate into the cells. The impedance spectra of suspensions of cells with gold nanoparticles inserted within them were then studied. In suspensions of uidA-expressing cells and gold nanoparticles functionalized with the complementary single-stranded DNA oligonucleotide, the impedance magnitude in the frequency range of interest was significantly higher (146 %) in comparison to all other controls. Due to its highly selective nature, the methodology has the potential to be used as a precision agricultural sensing system for accurate and real-time detection of markers of stress, viral infection, disease, and normal physiological activities.

Measuring the effects of diethyl phthalate microplastics on marine algae growth using dielectric spectroscopy.

This paper presents the development of a dielectric spectroscopy-based method using a customized, transmission line probe, fabricated on a printed circuit board (PCB), for monitoring the effect of diethyl phthalate (DEP) microplastics on marine algae growth. Experiments were performed by exposing marine algae (Chlorella pyrenoidosa) to DEP (0-50 mg) for up to 6 days. In order to amplify the electrophysiological effects and improve the sensing, a glutaraldehyde crosslinking agent was used and encapsulated on the surface of the probe. The reflection coefficient (S11) and the complex permittivity (ɛ' & ɛ″) of the Medium Under Test (MUT) were investigated in the frequency range of 30 kHz-800 MHz. Without the presence of DEP, the number of algae (104 cells/mL) and chlorophyll content (mg/L) increased at the rates of 207.73 × 104 cells/mL and 148.1 mg/L per day, respectively. After 6 days of exposing Chlorella pyrenoidosa (C. pyrenoidosa) algae to different DEP concentrations, the growth rate decreased down to -11.92 × 104 cells/mL and -19.19 mg/L (50 mg DEP), respectively. Additionally, the linearity of the relationship kept decreasing as the DEP content increased from R2 = 0.9716 to R2 = 0.1050 and from R2 = 0.9293 to R2 = 0.4961, respectively. Dielectric spectroscopy using the custom, transmission line probe, at 740 MHz, showed linear relationship (-1.22 dB/day) between the reflection coefficient (S11) and hence complex permittivity (ɛ' & ɛ″) without the presence of DEP. However, as the DEP content increased, algae growth was prohibited more intensely, shown both from the number of algae and the chlorophyll content. This trend was reflected on S11 and subsequently on the complex permittivity. This relationship confirms the capability of this method to monitor the growth of marine algae in almost real-time. This dielectric spectroscopy method could be a potential, low-cost tool to examine the impact of microplastic pollutants on marine microorganisms.

The ElectroUteroGraph: A Novel Tool for Assessing Uterine Contractions of Non-Pregnant Women.

Goal: Uterine contractility is known to play significant role in women's health. Ultrasonography and magnetic resonance imaging have been used for assessing uterine peristalsis, however they lack practicality, objectivity, and cost-effectiveness. In this paper, the ElectroUteroGraph (EUG) and novel electrodes are introduced, to cover the unmet need of practical intrauterine contractility assessment. The EUG measures biopotentials produced by uterine muscle contraction, similar to the basis of electrocardiography. Methods: The EUG was used to fifteen healthy, non-pregnant women of reproductive age. Amplitude and frequency-related features were derived from our recordings. Results: The EUG and novel electrodes did not cause any pain or discomfort to the patients, over their multiple recording sessions. The collected data showed difference between the proliferative and luteal phase of menstrual cycle (p < 0.05). Conclusions: The EUG can accurately measure uterine electrical activity, in a simple, standardized, safe and pain-free approach, leading to objective evaluation of uterine peristalsis.

Towards optimization of plant cell detection in suspensions using impedance-based analyses and the unified equivalent circuit model.

An improved approach for comparative study of plant cells for long term and continuous monitoring using electrical impedance spectroscopy is demonstrated for tomato and tobacco plant cells (MSK8 and BY2) in suspensions. This approach is based on the locations and magnitudes of defining features in the impedance spectra of the recently reported unified equivalent circuit model. The ultra-wide range (4 Hz to 20 GHz) impedance spectra of the cell lines were measured using custom probes, and were analyzed using the unified equivalent circuit model, highlighting significant negative phase peaks in the ~ 1 kHz to ~ 10 MHz range. These peaks differ between the tomato and tobacco cells, and since they can be easily defined, they can potentially be used as the signal for differentiating between different cell cultures or monitoring them over time. These findings were further analysed, showing that ratios relating the resistances of the media and the resistance of the cells define the sensitivity of the method, thus affecting its selectivity. It was further shown that cell agglomeration is also an important factor in the impedance modeling in addition to the overall cell concentration. These results can be used for optimizing and calibrating electrical impedance spectroscopy-based sensors for long term monitoring of cell lines in suspension for a given specific cell and media types.

Electrical impedance spectroscopy of plant cells in aqueous buffer media over a wide frequency range of 4 Hz to 20 GHz.

Electrical impedance spectroscopy was performed on suspensions of plant cells in aqueous buffer media over a wide frequency range of 4 Hz to 20 GHz. Custom probes were designed, manufactured, and used for these investigations. Experiments were performed with a custom-made parallel plate probe and impedance analysers in the low-frequency range (4 Hz to 5 MHz), with a custom-made coaxial airline probe and a vector network analyser in the mid-frequency range (100 kHz to 3 GHz), and with a commercial open-ended probe and a vector network analyser in the high-frequency range (200 MHz to 20 GHz). The impedance data acquired were processed in order to eliminate the effects of parasitics and compensate for geometrical differences between the three probes. Following this, the data were fitted to a unified model consisting of the Randles and Debye models. The data were also normalized to a reference measurement, in order to accentuate the effects of cell concentration on the impedance of the suspensions.•The methodology allows for impedance spectroscopy of cell suspensions over a wide frequency range spanning 10 orders of magnitude.•It allows for compensation of parasitics and of geometrical variations between probes, using mathematical techniques.

Electrical Impedance Spectroscopy of plant cells in aqueous biological buffer solutions and their modelling using a unified electrical equivalent circuit over a wide frequency range: 4Hz to 20 GHz.

A simple, ultra-wide frequency range, equivalent circuit for plant cell suspensions is presented. The model incorporates both the interfacial interactions of the suspension with the electrode, dominant at low frequencies, and the molecule and cell polarization mechanisms dominant at higher frequencies. Such model is useful for plant cell characterization allowing a single set of parameters over >9 orders of magnitude, whilst allows electronic simulations over the whole frequency range using a single model, simplifying the design of electronic systems of integrated plant cell sensors. The model has been experimentally validated in the frequency range of 4 Hz-20 GHz with each component in the circuit representing a physical phenomenon. Various cell concentrations (MSK8 tomato cells in Murashige and Skoog media) have been investigated, showing clear correlations of the cell capacitance increasing within the range of 200-600 pF, whilst cell resistance (R) decreasing within the range of approximately 0.8-3 kΩ within the cell concentration X-Y cells/mL range. This is the first model ever reported that covers such a wide frequency range and includes both interfacial and polarization effects in this simple form.

High-frequency, dielectric spectroscopy for the detection of electrophysiological/biophysical differences in different bacteria types and concentrations.

This paper describes a novel technique to quantify and identify bacterial cultures of Bacillus Subtilis (2.10-1.30 × 109 CFU mL-1) and Escherichia Coli (1.60-1.00 × 109 CFU mL-1), in corn oil using dielectric spectroscopy at elevated frequencies of 0.0100-20.0 GHz. This technique is using the electrophysiological/biophysical differences (e.g. gram positive and gram negative) between various bacteria types, as a basis to distinguish between bacteria concentrations and bacteria types. A close-ended, coaxial probe of 20.0 mm long sample-holder was developed and used to calculate the dielectric constant from the measured S parameters of the bacterial cultures, using the Nicolson-Ross-Weir method. This technique shows a linear relationship (r2 ≥ 0.999) between the dielectric constant and the cell concentration, at 16.0 GHz. The sensitivity of the technique is 0.177 × 109 (CFU mL-1)-1 for B. Subtilis (with a size of 10.0 × 1.00 µm), 0.322 × 109 (CFU mL-1)-1 for E. Coli (with a size of 2.00 × 0.500 µm) and 0.913 × 109 (CFU mL-1) -1 for their 1:1 mixture, while the response time is 60.0s. The dependency of dielectric constant on the bacterial cell concentration at a given frequency can be potentially exploited for measuring bacterial concentrations and biophysical differences.

Conserving and extending the useful life of the largest aquifer in North America: the future of the High Plains/Ogallala aquifer.

The water-level decline of the High Plains/Ogallala aquifer is one of the largest water management concerns in the United States. The economy and livelihood of people living in that vast region depend almost exclusively on water extracted from that aquifer. A debate about its future is ongoing, and questions remain as to how best to conserve the groundwater resource. Maintaining the aquifer will require reductions in pumping and irrigated hectarage and adopting additional conservation measures. Eventually, the agricultural system will have to be based dominantly on the renewable water resources of the region. In effect, this means a limited-irrigation and/or dry-farming regime. What Kansas is currently doing to further extend the life of the aquifer is presented here together with additional measures that could be taken. A key management approach to help sustain the aquifer in western Kansas is to divide the aquifer into subunits on which to base localized management decisions. Another recently adopted measure is the establishment of local enhanced management areas, which would allow locally agreed upon specific corrective controls in those areas. History has shown that incentive and voluntary plans alone have not been successful in halting water-level declines. Thus, limits and timelines need to be set and checks must be in place to enforce strict administration of conservation measures. It is advocated that water laws be reformed and modernized so that "water rights" are constrained by the current availability of water and the preservation of the resource base for future generations.

Towards sustainable groundwater use: setting long-term goals, backcasting, and managing adaptively.

The sustainability of crucial earth resources, such as groundwater, is a critical issue. We consider groundwater sustainability a value-driven process of intra- and intergenerational equity that balances the environment, society, and economy. Synthesizing hydrogeological science and current sustainability concepts, we emphasize three sustainability approaches: setting multigenerational sustainability goals, backcasting, and managing adaptively. As most aquifer problems are long-term problems, we propose that multigenerational goals (50 to 100 years) for water quantity and quality that acknowledge the connections between groundwater, surface water, and ecosystems be set for many aquifers. The goals should be set by a watershed- or aquifer-based community in an inclusive and participatory manner. Policies for shorter time horizons should be developed by backcasting, and measures implemented through adaptive management to achieve the long-term goals. Two case histories illustrate the importance and complexity of a multigenerational perspective and adaptive management. These approaches could transform aquifer depletion and contamination to more sustainable groundwater use, providing groundwater for current and future generations while protecting ecological integrity and resilience.

On understanding and predicting groundwater response time.

An aquifer system, when perturbed, has a tendency to evolve to a new equilibrium, a process that can take from just a few seconds to possibly millions of years. The time scale on which a system adjusts to a new equilibrium is often referred to as "response time" or "lag time." Because groundwater response time affects the physical and economic viability of various management options in a basin, natural resource managers are increasingly interested in incorporating it into policy. However, the processes of how groundwater responds to land-use change are not well understood, making it difficult to predict the timing of groundwater response to such change. The difficulty in estimating groundwater response time is further compounded because the data needed to quantify this process are not usually readily available. This article synthesizes disparate pieces of information on aquifer response times into a relatively brief but hopefully comprehensive review that the community of water professionals can use to better assess the impact of aquifer response time in future groundwater management investigations. A brief exposition on dimensional/scaling analysis is presented first, followed by an overview of aquifer response time for simplified aquifer systems. The aquifer response time is considered first from a water-quantity viewpoint and later expanded to incorporate groundwater age and water-quality aspects. Monitoring programs today, as well as water policies and regulations, should address this issue of aquifer response time so that more realistic management expectations can be reached.

Soil nitrogen balance under wastewater management: field measurements and simulation results.

The use of treated wastewater for irrigation of crops could result in high nitrate-nitrogen (NO(3)-N) concentrations in the vadose zone and ground water. The goal of this 2-yr field-monitoring study in the deep silty clay loam soils south of Dodge City, Kansas, was to assess how and under what circumstances N from the secondary-treated, wastewater-irrigated corn reached the deep (20-45 m) water table of the underlying High Plains aquifer and what could be done to minimize this problem. We collected 15.2-m-deep soil cores for characterization of physical and chemical properties; installed neutron probe access tubes to measure soil-water content and suction lysimeters to sample soil water periodically; sampled monitoring, irrigation, and domestic wells in the area; and obtained climatic, crop, irrigation, and N application rate records for two wastewater-irrigated study sites. These data and additional information were used to run the Root Zone Water Quality Model to identify key parameters and processes that influence N losses in the study area. We demonstrated that NO(3)-N transport processes result in significant accumulations of N in the vadose zone and that NO(3)-N in the underlying ground water is increasing with time. Root Zone Water Quality Model simulations for two wastewater-irrigated study sites indicated that reducing levels of corn N fertilization by more than half to 170 kg ha(-1) substantially increases N-use efficiency and achieves near-maximum crop yield. Combining such measures with a crop rotation that includes alfalfa should further reduce the accumulation and downward movement of NO(3)-N in the soil profile.

The science and practice of environmental flows and the role of hydrogeologists.

Conflicts between ecosystems and human needs for fresh water are increasing. The purpose of this paper is to raise awareness in the hydrogeologic community of environmental flows (EFs) and to address the major challenges involved in their protection. Ground water is a key component of EFs, and therefore hydrogeologists are called upon to get involved in the ongoing debates about maintaining healthy riverine ecosystems. Promising opportunities for achieving EFs in both underallocated and overallocated basins as well as new methods for protecting fresh water ecosystems developed in different countries are outlined. EF protection measures include private water trusts, "upside-down instream flow water rights," the "public trust" doctrine, and water markets, among other measures. A number of knowledge gaps are identified, to which hydrogeologists could contribute, such as our rudimentary knowledge about ground water-dependent ecosystems, aspects of stream-aquifer interactions, and the impacts of land-use changes. The values that society places on the different uses of water ultimately determine where the water is allocated. EF requirements can be legitimately recognized and addressed by basing the environmental needs of hydrologic systems on robust science, focusing on increasing the productivity of water use, engaging society in understanding the benefits and costs of decisions that affect ecosystems, and taking advantage of various opportunities for achieving EF goals.