ABSTRACT
Freshwater ecosystems are under threat from rising pharmaceutical pollution. While such pollutants are known to elicit biological effects on organisms, we have limited knowledge on how these effects might cascade through food-webs, disrupt ecological processes, and shape freshwater communities. In this study, we used a mesocosm experiment to explore how the community impacts of a top-order predator, the eastern mosquitofish (Gambusia holbrooki), are mediated by exposure to environmentally relevant low (measured concentration: â¼10 ng/L) and high concentrations (â¼110 ng/L) of the pervasive pharmaceutical pollutant fluoxetine. We found no evidence that exposure to fluoxetine altered the consumptive effects of mosquitofish on zooplankton. However, once mosquitofish were removed from the mesocosms, zooplankton abundance recovered to a greater extent in control mesocosms compared to both low and high fluoxetine-exposed mesocosms. By the end of the experiment, this resulted in fundamental differences in community structure between the control and fluoxetine-treated mesocosms. Specifically, the control mesocosms were characterized by higher zooplankton abundances and lower algal biomass, whereas mesocosms exposed to either low or high concentrations of fluoxetine had lower zooplankton abundances and higher algal biomass. Our results suggest that fluoxetine, even at very low concentrations, can alter aquatic communities and hinder their recovery from disturbances.
Subject(s)
Fresh Water , Water Pollutants, Chemical , Zooplankton , Animals , Fresh Water/chemistry , Zooplankton/drug effects , Food Chain , Ecosystem , Fluoxetine , Fishes , CyprinodontiformesABSTRACT
We use a phase-sensitive measurement to perform a binary hypothesis testing, i.e., distinguish between one on-axis and two symmetrically displaced Gaussian point spread functions. In the sub-Rayleigh regime, we measure a total error rate lower than allowed by direct imaging. Our results experimentally demonstrate that linear-optical spatial mode transformations can provide useful advantages for object detection compared with conventional measurements, even in the presence of realistic experimental cross talk, paving the way for meaningful improvements in identifying, detecting, and monitoring real-world, diffraction-limited scenes.
ABSTRACT
Two novel, discrete lanthanide-macrocycle binary complexes for the detection of hydrogen sulfide are reported. The hydrogen sulfide sensing mechanism utilises the copper sequestration at a secondary binding site, with resulting bimetallic lanthanide(III)/copper(II) complexes (Ln = Eu3+ and Tb3+) exhibiting high selectivity, good sensitivity and excellent reversibility for aqueous hydrogen sulfide. The inclusion of the DO2A macrocycle and 4-(2-pyridyl)-1,2,3-triazole dipicolinic acid ligand, results in a complex with good solubility and stability. The europium(III) complex also displayed a low limit of detection (665 ppb) with a response time of 30 seconds with gaseous hydrogen sulfide. The improved water solubility and stability over a previous complex results in these sensors having the potential for use in environmental monitoring and biological studies for various functional settings.
ABSTRACT
Single-molecule localization microscopy enables three-dimensional fluorescence imaging at tens-of-nanometer resolution, but requires many camera frames to reconstruct a super-resolved image. This limits the typical throughput to tens of cells per day. While frame rates can now be increased by over an order of magnitude, the large data volumes become limiting in existing workflows. Here we present an integrated acquisition and analysis platform leveraging microscopy-specific data compression, distributed storage and distributed analysis to enable an acquisition and analysis throughput of 10,000 cells per day. The platform facilitates graphically reconfigurable analyses to be automatically initiated from the microscope during acquisition and remotely executed, and can even feed back and queue new acquisition tasks on the microscope. We demonstrate the utility of this framework by imaging hundreds of cells per well in multi-well sample formats. Our platform, implemented within the PYthon-Microscopy Environment (PYME), is easily configurable to control custom microscopes, and includes a plugin framework for user-defined extensions.
Subject(s)
Imaging, Three-Dimensional , Software , Microscopy, Fluorescence/methods , Single Molecule Imaging/methodsABSTRACT
We consider passive imaging tasks involving discrimination between known candidate objects and investigate the best possible accuracy with which the correct object can be identified. We analytically compute quantum-limited error bounds for hypothesis tests on any library of incoherent, quasimonochromatic objects when the imaging system is dominated by optical diffraction. We further show that object-independent linear-optical spatial processing of the collected light exactly achieves these ultimate error rates, exhibiting scaling superior to spatially resolved direct imaging as the scene becomes more severely diffraction limited. We apply our results to example imaging scenarios and find conditions under which superresolution object discrimination can be physically realized.
ABSTRACT
Passive imaging receivers that demultiplex an incoherent optical field into a set of orthogonal spatial modes prior to detection can surpass canonical diffraction limits on spatial resolution. However, these mode-sorting receivers exhibit sensitivity to contextual nuisance parameters (e.g., the centroid of a clustered or extended object), raising questions on their viability in realistic scenarios where prior information about the scene is limited. We propose a multistage detection strategy that segments the total recording time between different physical measurements to build up the required prior information for near quantum-optimal imaging performance at sub-Rayleigh length scales. We show, via Monte Carlo simulations, that an adaptive two-stage scheme that dynamically allocates recording time between a conventional direct detection measurement and a binary mode sorter outperforms idealized direct detection alone when no prior knowledge of the object centroid is available, achieving one to two orders of magnitude improvement in mean squared error for simple estimation tasks. Our scheme can be generalized for more sophisticated tasks involving multiple parameters and/or minimal prior information.
ABSTRACT
The first discrete bimetallic europium(iii)/copper(ii) complex for the fast, sensitive and selective luminescent detection of both aqueous and gaseous hydrogen sulfide has been developed. The chemosensor displayed an impressive response time of 30 seconds and a low theoretical limit of detection (100 ppb) for gaseous hydrogen sulfide.
Subject(s)
Coordination Complexes/chemistry , Hydrogen Sulfide/analysis , Luminescent Agents/chemistry , Luminescent Measurements/methods , Paper , Copper/chemistry , Europium/chemistry , Hydrogen Sulfide/chemistry , Limit of Detection , Luminescent Measurements/instrumentationABSTRACT
Because of their unique photochemical and photophysical properties, luminescent lanthanide-based complexes have long captivated chemists. In recent years, the number of reports of luminescent lanthanide complex-based probes for monitoring of biological and environmental processes has dramatically increased, namely, because of their selectivity for particular analytes, lower limits of detection, and the fact that they allow monitoring of analytes in real time. Lanthanide-based probes need to be paired with an appropriate antenna/sensitizer to allow maximum energy transfer, with the antenna typically covalently attached to the stable lanthanide chelate. We have recently investigated "dark" lanthanide-based probes where the sensitizer is not covalently linked to the lanthanide chelate. Herein we report the use of a luminescent lanthanide-based probe system for the detection of Zn2+ ions based on the formation of a ternary complex between a "dark" terbium complex, lumazine, and Zn2+. The terbium(III)-based probe incorporates a 1,4,7,10-tetraazacyclododecane-1,4,7,10-triacetic acid macrocyclic chelator covalently attached to a cyclen moiety, which is the Zn2+ ion binding group. In the presence of Zn2+ ions and lumazine (a strongly UV-absorbing sensitizer), a 1:1:1 ternary complex forms. The resulting complex is highly luminescent and selective for Zn2+ ions over other cations of environmental significance. Furthermore, with a limit of detection of 1.2 µM, this probe can detect the level of chronic zinc(II) concentrations denoted by the U.S. Environmental Protection Agency.
ABSTRACT
Denitrification (DNF) and dissimilatory nitrate reduction to ammonium (DNRA) are two competing nitrate reduction pathways that remove or recycle nitrogen, respectively. However, factors controlling the partitioning between these two pathways are manifold and our understanding of these factors is critical for the management of N loads in constructed wetlands. An important factor that controls DNRA in an aquatic ecosystem is the electron donor, commonly organic carbon (OC) or alternatively ferrous iron and sulfide. In this study, we investigated the role of natural organic carbon (NOC) and acetate at different OC/NO3- ratios and ferrous iron on the partitioning between DNF and DNRA using the 15N-tracer method in slurries from four constructed stormwater urban wetlands in Melbourne, Australia. The carbon and nitrate experiments revealed that DNF dominated at all OC/NO3- ratios. The higher DNF and DNRA rates observed after the addition of NOC indicates that nitrate reduction was enhanced more by NOC than acetate. Moreover, addition of NOC in slurries stimulated DNRA more than DNF. Interestingly, slurries amended with Fe2+ showed that Fe2+ had significant control on the balance between DNF and DNRA. From two out of four wetlands, a significant increase in DNRA rates (pâ¯<â¯.05) at the cost of DNF in the presence of available Fe2+ suggests DNRA is coupled to Fe2+ oxidation. Rates of DNRA increased 1.5-3.5 times in the Fe2+ treatment compared to the control. Overall, our study provides direct evidence that DNRA is linked to Fe2+ oxidation in some wetland sediments and highlights the role of Fe2+ in controlling the partitioning between removal (DNF) and recycling (DNRA) of bioavailable N in stormwater urban constructed wetlands. In our study we also measured anammox and found that it was always <0.05% of total nitrate reduction in these sediments.
ABSTRACT
An in-depth study of the interaction of a trinuclear terbium(III)-dizinc(II) complex with an array of nucleotides differing in the type of nucleobase and number of phosphate groups, as well as cyclic versus acyclic variants, is presented. The study examined the nature of the interaction and the efficiency at which guanine was able to sensitize terbium(III) luminescence. Competitive binding and titration studies were performed to help establish the nature/mode of the interactions. These established that (1) interaction occurs by the coordination of phosphate groups to zinc(II) (in addition to uridine in the case of uridine monophosphate), (2) acyclic nucleotides bind more strongly than cyclic counterparts because of their higher negative charge, (3) guanine-containing nucleotides are able to sensitize terbium(III) luminescence with the efficiency of sensitization following the order guanosine monophosphate (GMP) > guanosine diphosphate > guanosine triphosphate because of the mode of binding, and (4) nucleoside monophosphates bind to a single zinc(II) ion, whereas di- and triphosphates appear to bind in a bridging mode between two host molecules. Furthermore, it has been shown that guanine is a sensitizer of terbium(III) luminescence. On the basis of the ability of GMP to effectively sensitize terbium(III)-based luminescence while cyclic GMP (cGMP) does not, the complex has been utilized to monitor the catalytic conversion of cGMP to GMP by a phosphodiesterase enzyme in real time using time-gated luminescence on a benchtop fluorimeter. The complex has the potential to find broad application in monitoring the activity of enzymes that process nucleotides (co)substrates, including high-throughput drug-screening programs.
Subject(s)
Coordination Complexes/chemistry , Guanosine Monophosphate/chemistry , Phosphoric Diester Hydrolases/analysis , Terbium/chemistry , Zinc/chemistry , Coordination Complexes/chemical synthesis , Coordination Complexes/radiation effects , Cyclic GMP/chemistry , Enzyme Assays , Light , Luminescence , Spectrophotometry , Water/chemistryABSTRACT
A multitude of biologically active pharmaceuticals contaminate surface waters globally, yet their presence in aquatic food webs remain largely unknown. Here, we show that over 60 pharmaceutical compounds can be detected in aquatic invertebrates and riparian spiders in six streams near Melbourne, Australia. Similar concentrations in aquatic invertebrate larvae and riparian predators suggest direct trophic transfer via emerging adult insects to riparian predators that consume them. As representative vertebrate predators feeding on aquatic invertebrates, platypus and brown trout could consume some drug classes such as antidepressants at as much as one-half of a recommended therapeutic dose for humans based on their estimated prey consumption rates, yet the consequences for fish and wildlife of this chronic exposure are unknown. Overall, this work highlights the potential exposure of aquatic and riparian biota to a diverse array of pharmaceuticals, resulting in exposures to some drugs that are comparable to human dosages.
Subject(s)
Food Chain , Pharmaceutical Preparations/analysis , Rivers/chemistry , Water Pollutants, Chemical/analysis , Animals , Australia , Diet , Invertebrates/metabolism , Larva/metabolism , Predatory Behavior , Risk Factors , Spiders/metabolismABSTRACT
A simple sample preparation method based on a modified liquid-phase extraction approach to extract selected pharmaceuticals and personal care products from freshwater organisms is described. Extracted samples were analysed using liquid chromatography with Q-Exactive plus hybrid quadrupole Orbitrap mass spectrometry, using 2.6 µm C18 media. A 0.1% v/v acetic acid/acetonitrile mobile phase was applied over a 20 min gradient. Method detection limits in full scan mode were ca. 0.04-2.38 ng of analyte per g of sample. Linearity ranged from 0.9750 to 0.9996 over the calibration range of 0.01-100 µg/L; MS mass accuracy was <2 ppm for most analytes. This method was applied to quantify six pharmaceuticals and personal care products in seven invertebrate samples. For tandem mass spectrometry analysis, selection of precursor ions was performed for each pharmaceutical, with Mass Frontier software illustrating the fragmentation mechanism. Effects of collision energy on intensities of ions was further investigated. The tandem mass spectrometry condition resulting in the highest signal of respective selected product ion was selected to confirm each pharmaceutical, which was initially observed in the full scan mode. Results indicate that pharmaceuticals and personal care products found to be present in water-ways, may be incorporated into organisms that live in the environment of affected water streams.
Subject(s)
Fresh Water/chemistry , Water Pollutants, Chemical/analysis , Animals , Chromatography, High Pressure Liquid , Invertebrates , Tandem Mass SpectrometryABSTRACT
This study developed an analytical approach for sub-ppb level detection and confirmation of 13 pharmaceuticals and personal care products (PPCPs) in water samples using ultra high pressure liquid chromatography hyphenated with a quadrupole Orbitrap mass spectrometer (UHPLC- Q-Orbitrap-MS). Sample preparation was performed by using solid phase extraction (SPE) employing hydrophilic-lipophilic balance cartridges, with elution of sorbed analytes using methanol. Acceptable automatic gain control (AGC) target and maximum injection time (IT) were 1×106 and 200ms, respectively, resulting in a mass accuracy <2ppm. High response signals with sufficient data points per peaks (20-30) were obtained whilst maintaining high resolution of approximately 70,000 full width at half maximum. Extracted ion chromatograms provided quantitative analysis with linearity (R2) ranging from 0.9875 to 0.9993 and method detection limits ranging from 0.01-0.61ngmL-1. Compounds were further analysed by MS/MS analysis, with the MS operated in parallel reaction monitoring (PRM) mode under precursor ion analysis intervals and collision energies chosen for the different PPCPs. The developed method was applied to analyse water samples obtained from sources in Victoria, Australia.
Subject(s)
Chromatography, High Pressure Liquid/methods , Pharmaceutical Preparations/analysis , Pharmaceutical Preparations/isolation & purification , Tandem Mass Spectrometry/methods , Water Pollutants, Chemical/analysis , Water Pollutants, Chemical/isolation & purification , Australia , Limit of Detection , Solid Phase Extraction/methodsABSTRACT
Synthesized ferrihydrite (Fh) with the dosages of 0.3, 0.6 and 0.9 cm thickness (labeled as Fh, 2Fh and 3Fh respectively, equivalent to 248-774 g/m2) were deployed to serve as the reactive capping layer covering the Ornamental Lake sediments, the Royal Botanic Garden of Melbourne. The sediments were exposed to an alternating regime of oxic/anoxic conditions using laboratory reactors for 45 days. Dynamics of dissolved oxygen (DO), pH, filterable reactive phosphorus (FRP), filterable ammonium (NH4+), nitrate and nitrite (NOx), total dissolved nitrogen (TDN) and dissolved iron (Fe) of overlying water were examined. After incubation, O2 and H2S profiles across the water-sediment interface were observed with microelectrodes. The element distributions in the upper sediments were tested as well. Results showed that DO and pH kept relatively stable during oxic period, while decreased significantly during anoxic period. Fh cappings decreased both DO and pH, and inhibited the release of FRP. No significant increments of FRP in overlying waters were observedduring anoxic period. Fh cappings prompted the releases of NH4+ and TDN, while inhibited that of NOx.NH4+increased while NOx decreased during anoxic period. Fe(II) and TFe increased only in 3Fh, especially during anoxic conditions. Fh cappings increased O2 and H2S concentrations across the water-sediment interfaces. TP and TN in the sediments decreased after capping, while TFe increased significantly. We concluded that 0.6 cm thickness of (496 g/m2) Fh capping could sequestrate P, even experiencing redox conditions.
Subject(s)
Environmental Restoration and Remediation/methods , Ferric Compounds/chemistry , Phosphorus/analysis , Water Pollutants, Chemical/analysis , Wetlands , Eutrophication , Geologic Sediments , Iron , Lakes , Nitrogen/analysis , Oxidation-Reduction , Oxygen , Plants , WaterABSTRACT
A bimetallic terbium(iii)/copper(ii) complex (Tb-1·Cu2+) for the time-gated luminescent detection of hydrogen sulfide in aqueous samples is reported. The probe shows excellent selectivity towards HS- over various anions and cations, including the ions common to natural waterways and waste water samples, displaying a 73-fold increase in luminescence in the presence of sulfide. The probe exhibits extremely fast reaction times and a low limit of detection (130 nM). The probe was used to quantify sulfide in an industrial "sour water" sample, with the result in excellent agreement with those from two independent assay methods (methylene blue and AzMC).
ABSTRACT
A new bifunctional macrocyclic chelator featuring a conjugatable alkynyl-naphthalimide fluorophore pendant group has been prepared and its Gd(III) complex coupled to a cell-penetrating lipidated azido-Tat peptide derivative using Cu(I)-catalysed "click" chemistry. The resulting fluorescent conjugate is able to enter CAL-33 tongue squamous carcinoma cells, as revealed by confocal microscopy, producing a very modest anti-proliferative effect (IC50 = 93 µM). Due to the photo-reactivity of the naphthalimide moiety, however, the conjugate's cytotoxicity is significantly enhanced (IC50 = 16 µM) upon brief low-power UV-A irradiation.
Subject(s)
Antineoplastic Agents/metabolism , Coordination Complexes/metabolism , Naphthalimides/metabolism , Photosensitizing Agents/metabolism , Antineoplastic Agents/pharmacology , Cell Line, Tumor , Cell Proliferation/drug effects , Chelating Agents/metabolism , Chelating Agents/pharmacology , Click Chemistry , Coordination Complexes/pharmacology , Gadolinium/chemistry , Heterocyclic Compounds, 1-Ring/chemistry , Humans , Naphthalimides/pharmacology , Photosensitizing Agents/pharmacology , tat Gene Products, Human Immunodeficiency Virus/chemistryABSTRACT
Two new bifunctional macrocyclic chelate ligands that form luminescent terbium(III) complexes featuring an alkyne group for conjugation to (bio)molecules via the Cu(I)-catalyzed "click" reaction were synthesized. Upon ligation, the complexes exhibit a significant luminescent enhancement when excited at the λ(max) of the "clicked" products. To demonstrate the utility of the complexes for luminescent labeling, they were conjugated in vitro to E. coli aspartate/glutamate-binding protein incorporating a genetically encoded p-azido-L-phenylalanine or p-(azidomethyl)-L-phenylalanine residue. The complexes may prove useful for time-gated assay applications.
Subject(s)
Alkynes/chemistry , Proteins/chemistry , Terbium/chemistry , LuminescenceABSTRACT
The long-term efficacy of stormwater treatment systems requires continuous pollutant removal without substantial re-release. Hence, the division of incoming pollutants between temporary and permanent removal pathways is fundamental. This is pertinent to nitrogen, a critical water body pollutant, which on a broad level may be assimilated by plants or microbes and temporarily stored, or transformed by bacteria to gaseous forms and permanently lost via denitrification. Biofiltration systems have demonstrated effective removal of nitrogen from urban stormwater runoff, but to date studies have been limited to a 'black-box' approach. The lack of understanding on internal nitrogen processes constrains future design and threatens the reliability of long-term system performance. While nitrogen processes have been thoroughly studied in other environments, including wastewater treatment wetlands, biofiltration systems differ fundamentally in design and the composition and hydrology of stormwater inflows, with intermittent inundation and prolonged dry periods. Two mesocosm experiments were conducted to investigate biofilter nitrogen processes using the stable isotope tracer 15NO3(-) (nitrate) over the course of one inflow event. The immediate partitioning of 15NO3(-) between biotic assimilation and denitrification were investigated for a range of different inflow concentrations and plant species. Assimilation was the primary fate for NO3(-) under typical stormwater concentrations (â¼1-2 mg N/L), contributing an average 89-99% of 15NO3(-) processing in biofilter columns containing the most effective plant species, while only 0-3% was denitrified and 0-8% remained in the pore water. Denitrification played a greater role for columns containing less effective species, processing up to 8% of 15NO3(-), and increased further with nitrate loading. This study uniquely applied isotope tracing to biofiltration systems and revealed the dominance of assimilation in stormwater biofilters. The findings raise important questions about nitrogen release upon plant senescence, seasonally and in the long term, which have implications on the management and design of biofiltration systems.
Subject(s)
Cyclonic Storms , Denitrification , Filtration/methods , Nitrogen/isolation & purification , Nitrogen/metabolism , Bacteria/metabolism , Biodegradation, Environmental , Carex Plant/metabolism , Filtration/instrumentation , Nitrates/isolation & purification , Nitrogen Isotopes , Oxygen/analysis , Porosity , WaterABSTRACT
Second derivative ultra-violet spectrophotometric methods are described for the measurement of nitrate and total nitrogen in freshwaters using flow analysis techniques. A simple flow system consisting of a peristaltic pump and a single-reflection flow-through cell was used for the measurement of nitrate. Quantification of total nitrogen using alkaline peroxodisulfate photo-digestion was achieved by incorporating an ultra-violet photo-reactor, a hollow-fibre filter and a debubbler into the flow system. The nitrate system featured a limit of detection of 0.04 mg N L(-1), 0.4%RSD (1 mg N L(-1) as nitrate, n=10), a coefficient of determination (R(2)) of 0.9995 over the calibration range 0.0-2.0 mg N L(-1), and a data acquisition time of 1.5s per spectrum. The total nitrogen system featured a limit of detection of 0.05 mg N L(-1), 1%RSD (1 mg N L(-1) as ammonium chloride, n=10), a coefficient of determination of 0.9989 over the calibration range 0.0-2.0 mg N L(-1), and a throughput of 5 sample h(-1) measured in triplicate. Digestions of five model nitrogen compounds returned recoveries of >88%. Determinations carried out using the developed systems show a high degree of agreement with data obtained using reference methods. These methods require no colorimetric reagents and eliminate the requirement for a toxic cadmium reduction column. The overlap of chloride and nitrate spectra in seawater is not eliminated entirely by the use of second derivative spectrophotometry, and consequently the methods are restricted to the analysis of freshwaters.
ABSTRACT
The export of nitrogen from urban catchments is a global problem, and denitrifying bacteria in stream ecosystems are critical for reducing in-stream N. However, the environmental factors that control the composition of denitrifying communities in streams are not well understood. We determined whether denitrifying community composition in sediments of nine streams on the eastern fringe of Melbourne, Australia was correlated with two measures of catchment urban impact: effective imperviousness (EI, the proportion of a catchment covered by impervious surfaces with direct connection to streams) or septic tank density (which affects stream water chemistry, particularly stream N concentrations). Denitrifying community structure was examined by comparing terminal restriction fragment length polymorphisms of nosZ genes in the sediments, as the nosZ gene codes for nitrous oxide reductase, the last step in the denitrification pathway. We also determined the chemical and physical characteristics of the streams that were best correlated with denitrifying community composition. EI was strongly correlated with community composition and sediment physical and chemical properties, while septic tank density was not. Sites with high EI were sandier, with less fine sediment and lower organic carbon content, higher sediment cations (calcium, sodium and magnesium) and water filterable reactive phosphorus concentrations. These were also the best small-scale environmental variables that explained denitrifying community composition. Among our study streams, which differed in the degree of urban stormwater impact, sediment grain size and carbon content are the most likely drivers of change in community composition. Denitrifying community composition is another in a long list of ecological indicators that suggest the profound degradation of streams is caused by urban stormwater runoff. While the relationships between denitrifying community composition and denitrification rates are yet to be unequivocally established, landscape-scale indices of environmental impact such as EI may prove to be useful indicators of change in microbial communities.