bluuebunny/crossref_metadata_2025 huggingface dataset:
Controls on effective settling velocities of mud flocs in the Changjiang Estuary, China
Authors: Shi Z. , Zhou H. J.
Published: October 2004

Theoretical and experimental studies were undertaken to gain insight into physical parameters controlling the flocculation and settling properties of mud flocs in the Changjiang Estuary, China. The Rouse equation is applied to vertical profiles of suspended sediment concentration to determine the bulk mean settling velocity (ws) of sediment suspended in the Changjiang Estuary. Both in situ point‐sampled and acoustically measured profiles of suspended mud concentrations were fit selectively. The calculated settling velocities ws mainly ranged from 0·4 to 4·1 mm s−1 for the point‐sampled data set, and from 1·0 to 3·0 mm s−1 for the acoustically measured data set. Furthermore, the settling velocities of mud flocs increased with mean concentration (C̄) of mud flocs in suspension and were proportional to increasing bottom shear stress (τb) of tidal flow. The best equation for the field settling velocity of mud flocs in the Changjiang Estuary can be expressed by the power law: ws = mC̄n (m, 1·14–2·37; n, 0·84–1·03). It is suggested that C̄ and τb were the dominant physical parameters controlling the flocculation and ws of mud flocs in suspension. Copyright © 2004 John Wiley & Sons, Ltd.
Long‐term (18‐year) changes in sulphate concentrations in two Ontario headwater lakes and their inflows in response to decreasing deposition and climate variations
Authors: Eimers M. C. , Dillon P. J. , Watmough S. A.
Published: October 2004

Sulphate concentrations in two headwater lakes and their major inflows were evaluated over an 18 year period (1980–81 to 1997–98) during which time sulphate bulk deposition declined by approximately 40%. The two lake catchments represent either end of the spectrum of acid sensitivity in the Muskoka–Haliburton region of Ontario. Between 1980 and 1998, sulphate concentrations in Harp and Plastic Lakes decreased, but the decrease was much less than expected (28% and 21% respectively) given the magnitude of change in deposition. Sulphate export in streams draining into the lakes greatly exceeded sulphate input to catchments in most years, which quantitatively explains the response of lake‐sulphate concentration. Furthermore, temporal patterns in mean annual sulphate concentrations in streams were similar, and appeared to be related to climate factors. Specifically, catchment export of sulphate was greater and stream‐sulphate concentrations were higher in years that had warm, dry summers, i.e. when streamflow in many catchments ceased for up to several weeks. Increased sulphate export from catchments resulted in higher sulphate concentrations in lakes, but the response of lake sulphate was not as immediate or dramatic as the response of stream sulphate to changes in catchment dryness. Factors that affect sulphate retention or export in catchments exert a strong influence on sulphate concentrations in lakes and streams and need to be considered when evaluating the response of surface water chemistry to changes in sulphate deposition. Copyright © 2004 John Wiley & Sons, Ltd.
The application of an integrated biogeochemical model (PnET‐BGC) to five forested watersheds in the Adirondack and Catskill regions of New York
Authors: Chen Limin , Driscoll Charles T. , Gbondo‐Tugbawa Solomon , Mitchell Myron J. , Murdoch Peter S.
Published: October 2004

PnET‐BGC is an integrated biogeochemical model formulated to simulate the response of soil and surface waters in northern forest ecosystems to changes in atmospheric deposition and land disturbances. In this study, the model was applied to five intensive study sites in the Adirondack and Catskill regions of New York. Four were in the Adirondacks: Constable Pond, an acid‐sensitive watershed; Arbutus Pond, a relatively insensitive watershed; West Pond, an acid‐sensitive watershed with extensive wetland coverage; and Willy's Pond, an acid‐sensitive watershed with a mature forest. The fifth was Catskills: Biscuit Brook, an acid‐sensitive watershed. Results indicated model‐simulated surface water chemistry generally agreed with the measured data at all five sites. Model‐simulated internal fluxes of major elements at the Arbutus watershed compared well with previously published measured values. In addition, based on the simulated fluxes, element and acid neutralizing capacity (ANC) budgets were developed for each site. Sulphur budgets at each site indicated little retention of inputs of sulphur. The sites also showed considerable variability in retention of NO3−. Land‐disturbance history and in‐lake processes were found to be important in regulating the output of NO3− via surface waters. Deposition inputs of base cations were generally similar at these sites. Various rates of base cation outputs reflected differences in rates of base cation supply at these sites. Atmospheric deposition was found to be the largest source of acidity, and cation exchange, mineral weathering and in‐lake processes served as sources of ANC. Copyright © 2004 John Wiley & Sons, Ltd.
Export mechanisms for dissolved organic carbon and nitrate during summer storm events in a glaciated forested catchment in New York, USA
Authors: Inamdar Shreeram P. , Christopher Sheila F. , Mitchell Myron J.
Published: October 2004

Nitrate and dissolved organic carbon (DOC) concentrations during a summer storm for a forested catchment in the Adirondack Mountains displayed a clear separation in trajectories and timing of maximum values. Nitrate concentrations peaked early on the rising limb of the hydrograph, whereas DOC concentrations gradually increased through the rising limb with maximum concentrations following the discharge peak. Solute data from precipitation, throughfall, soilwater, and ground/till water indicated till water and near‐surface soil waters as the controlling end members for stream NO3− and DOC concentrations respectively. Streamflow concentrations of major base cations (Ca2+ and Mg2+), which were assumed to represent water originating from deep flow paths, matched the NO3− trajectory. These data suggest that streamflow NO3− concentrations are derived from till groundwater and that DOC is derived from near‐surface soil waters. We attributed the early expression of NO3− to the displacement of till waters by infiltrating precipitation. In contrast, we hypothesized that the delayed DOC concentrations occurred with surface and near‐surface runoff from near‐stream wetlands/peatlands and isolated saturated areas that became connected only under conditions of maximum water content in the catchment. Copyright © 2004 John Wiley & Sons, Ltd.
The episodic acidification of a stream with elevated concentrations of dissolved organic carbon
Authors: Wellington Brian I. , Driscoll Charles T.
Published: October 2004

Organic acids are generally thought to play a minor role in the episodic acidification of streams in the USA. In this study, we investigated the episodic acidification of a stream at the Hubbard Brook Experimental Forest in New Hampshire with high concentrations of dissolved organic carbon and naturally occurring organic acids. We studied three events in 2001: spring snowmelt, which occurred from 6 April to 14 May and resulted in two distinct melt events; and two rain events, one on 17 June and the other on 17 July. During snowmelt events organic acids were a minor contributor to the short‐term acidification of stream water, with increases in NO3− and dilution of base cations being the dominant mechanisms. During summer rainfall events, however, increases in inputs of organic acids were the dominant mechanism of episodic acidification when soil water was the dominant contributor to stream discharge (59 to 66% of peak stream discharge). We also found that precipitation events occurring after relatively wet antecedent conditions (17 July event) resulted in more severe acid episodes than events that followed drier antecedent conditions (17 June event). The minimum acid neutralizing capacity (ANC) was only −19 µeq l−1 for the 17 June event, whereas the minimum ANC for the 17 July event was much lower (−62 µeq l−1) although the total rainfall amount was similar for the two events. Copyright © 2004 John Wiley & Sons, Ltd.
Comparison of nitrogen solute concentrations within alder (Alnus incana ssp. rugosa) and non‐alder dominated wetlands
Authors: Hurd Todd M. , Raynal Dudley J.
Published: October 2004

This study examined differences in nitrogen solutes and groundwater flow patterns between a riparian wetland dominated by the N2‐fixing shrub, Alnus incana ssp. rugosa, and an upstream coniferous forested riparian wetland along a stream of the Adirondack Mountains, where some surface waters are susceptible to nitrogen excess. Channel water NO3− was up to 16 µmol l−1 greater in the alder reach, with peaks following maxima in groundwater dissolved inorganic nitrogen (DIN). NO3− at 25 cm depth was 30 µmol greater in the alder than in the conifer reach in April, and 24 µmol l−1 greater than channel water and 30 µmol l−1 greater than that of 125 cm groundwater in June. Dissolved organic nitrogen and NH4+ concentrations increased between 25 and 75 cm depths in both wetlands during the growing season. Inorganic nitrogen increased between the hillslope and stream in both wetlands, with the greatest increases in the alder reach during the dormant season. Greatest subsurface DIN (120 µmol l−1) occurred at 75 cm in the alder reach, within 1 m of the stream, between November (120 µmol l−1 NH4+) and a January thaw (60 µmol l−1 each of NH4+ and NO3−). Concentrations of deeper groundwater at 125 cm during this period were lower (10–30 µmol l−1). Lateral flow from the stream channel occurred in the alder reach during the dormant season, and channel water contribution to groundwater was correlated strongly to NO3− at 25 cm. These results indicate that nitrification is stimulated in the presence of alders and oxidized exchange flow, producing NO3− that may contribute to elevated channel water NO3− during periods of peak flow. Copyright © 2004 John Wiley & Sons, Ltd.
Evaluating the source of streamwater nitrate using δ15N and δ18O in nitrate in two watersheds in New Hampshire, USA
Authors: Pardo Linda H. , Kendall Carol , Pett‐Ridge Jennifer , Chang Cecily C. Y.
Published: October 2004

The natural abundance of nitrogen and oxygen isotopes in nitrate can be a powerful tool for identifying the source of nitrate in streamwater in forested watersheds, because the two main sources of nitrate, atmospheric deposition and microbial nitrification, have distinct δ18O values. Using a simple mixing model, we estimated the relative fractions in streamwater derived from these sources for two forested watersheds with markedly different streamwater nitrate outputs. In this study, we monitored δ15N and δ18O of nitrate biweekly in atmospheric deposition and in streamwater for 20 months at the Hubbard Brook Experimental Forest, New Hampshire, USA (moderate nitrogen export), and monthly in streamwater at the Bowl Research Natural Area, New Hampshire, USA (high nitrogen export). For rain, δ18O values ranged from +47 to +77‰ (mean: +58‰) and δ15N from −5 to +1‰ (mean: −3‰); for snow, δ18O values ranged from +52 to +75‰ (mean: +67‰) and δ15N from −3 to +2‰ (mean: −1‰). Streamwater nitrate, in contrast to deposition, had δ18O values between +12 and +33‰ (mean: +18‰) and δ15N between −3 and +6‰ (mean: 0‰). Since nitrate produced by nitrification typically has δ18O values ranging from −5 to +15‰, our field data suggest that most of the nitrate lost from the watersheds in streamflow was nitrified within the catchment. Our results confirm the importance of microbial nitrogen transformations in regulating nitrogen losses from forested ecosystems and suggest that hydrologic storage may be a factor in controlling catchment nitrate losses. Copyright © 2004 John Wiley & Sons, Ltd.
Regionalization of disturbance‐induced nitrogen leakage from mid‐Appalachian forests using a linear systems model
Authors: Eshleman Keith N. , Fiscus Daniel A. , Castro Nancy M. , Webb James R. , Herlihy Alan T.
Published: October 2004

The ‘leakage’ of nitrate‐nitrogen (nitrate‐N) to surface waters is a common (but not universal) response of forest ecosystems to both human‐induced and natural disturbances. There are several reported examples of the transient leakage of nitrate‐N to surface waters from eastern US forests that have sustained outbreaks of defoliating insects, such as the introduced gypsy moth (Lymantria dispar) larva. Previous research has suggested that annual nitrate‐N leakage from disturbed forests can be modelled using an empirically derived unit nitrogen export response function (UNERF) model. The model represents annual nitrate‐N export as a linear deterministic process in both space and time and is analogous to a unit hydrograph. The goal of the present study was to verify and apply a regionalized, lithology‐based UNERF model that references the geographic distribution of bedrock class and the timing and extent of gypsy moth defoliation of forests in the non‐glaciated highlands of the Chesapeake Bay watershed. Despite an inability to verify the model for most individual watersheds within the study area, the model was able to reproduce the statistical distribution of annual nitrate‐N export to streams that comprised our target population. During water year 1991 (the year following peak defoliation) the model results indicated that regional annual nitrate‐N export had transiently increased by nearly 1500% from a baseline rate of about 0·1 kg ha−1to a peak value approaching 1·5 kg ha−1. We thus conclude that natural vegetation disturbance is an important mechanism by which dissolved nitrogen is leaked from forested lands to small streams, rivers, and Chesapeake Bay. The present study also illustrates how simple, empirically derived linear systems approaches like the UNERF model can be successfully applied to problems where regionalization is a primary goal. Copyright © 2004 John Wiley & Sons, Ltd.
Changes in biogeochemical cycling following forest defoliation by pine wilt disease in Kiryu experimental catchment in Japan
Authors: Tokuchi Naoko , Ohte Nobuhito , Hobara Satoru , Kim Su‐Jin , Masanori Katsuyama
Published: October 2004

Changes in nutrient budgets and hydrological processes due to the natural disturbance of pine wilt disease (PWD) were monitored in a small, forested watershed in Japan. The disturbance caused changes in soil nitrogen transformations. Pre‐disturbance, mineralized nitrogen remained in the form of NH4+, whereas in disturbed areas most mineralized nitrogen was nitrified. Stream NO3− concentrations increased following PWD. There was a delay between time of disturbance and the increase of NO3− in ground and stream waters. Stream concentrations of NO3− and cations (Ca2+ + Mg2+) were significantly correlated from 1994 to 1996, whereas the correlation among NO3−, H+, and SO42− was significant only in 1995. Although both cation exchange and SO42− adsorption buffered protons, cation exchange was the dominant and continuous mechanism for acid buffering. SO42− adsorption was variable and highly pH dependent. The disturbance also resulted in slight delayed changes of input–output nutrient balances. The nitrogen contribution to PWD litter inputs was 7·39 kmol ha−1, and nitrogen loss from streamwater was less than 0·5 kmol ha−1 year−1 throughout the observation period. This large discrepancy suggested substantial nitrogen immobilization. Copyright © 2004 John Wiley & Sons, Ltd.
Matrix hydrodynamic properties of carbonate rocks from the Betic Cordillera (Spain)
Authors: Pulido‐Bosch A. , Motyka J. , Pulido‐Leboeuf P. , Borczak S.
Published: October 2004

Laboratory tests performed on 181 rock samples from boreholes drilled in different areas of the Betic Cordillera allow us to calculate their hydraulic conductivity and open porosity values. Higher values are generally associated with Miocene calcareous sandstones, although hydraulic conductivity reaches its highest values in some isolated limestone and dolostone samples. Lower values were found in marly limestones and marbles. Specific yield ranged from 0 to 0·0798, with a mean value of 0·00579. A total of 79 samples did not release water during the specific yield test, while another 11 samples released water for more than 30 min. Such wide ranges of variation show the great diversity of behaviour that the matrix of the carbonate rocks can have, referring to water storage and transfer, and its influence on pollutants spread, for example. A weak relation between interconnected porosity and hydraulic conductivity was found. The relation between interconnected porosity and specific yield is slightly stronger, except in the case of the dolomites, where a high correlation was found. No dependence on depth was found for hydraulic conductivity and interconnected porosity. Copyright © 2004 John Wiley & Sons, Ltd.
Estimates of flow resistance and eddy viscosity coefficients for 2D modelling on vegetated floodplains
Authors: Vionnet C. A. , Tassi P. A. , Martín Vide J. P.
Published: October 2004

The problem of quantifying the effects of flexible plants on flow resistance and eddy viscosity by vegetated floodplains is first addressed with a one‐dimensional (1D) approximation based upon the so‐called lateral distribution method. The estimates so obtained are then tested with two‐dimensional (2D) numerical simulations based on the full shallow water equations through the use of the computational code Telemac‐2D. Data obtained on a physical model of the Besòs River (Spain), whose floodplains were covered with plastic ornamental plants to mimic the effect of flexible vegetation, is used for the validation of the numerical results. Additionally, the values of flow resistance estimated numerically with the 1D and 2D simulations are compared with values obtained in a rectangular flume under flow conditions (slope, water depth and artificial lining) similar to those used on the reduced model. It is then established that as more physical mechanisms are included in the mathematical model used to study the problem, the ratio between the floodplain and the main channel flow resistance coefficient increases. The approach demonstrates that whenever enough flow data is available, the lateral distribution method delivers values of flow resistance and eddy viscosity which are highly consistent with 2D numerical modelling. This finding could mean considerable savings in the burdensome task of specifying flow resistance and turbulence dissipation values for 2D modelling of large compound channel systems. Copyright © 2004 John Wiley & Sons, Ltd.
Protein‐like fluorescence intensity as a possible tool for determining river water quality
Authors: Baker Andy , Inverarity Roger
Published: October 2004

The results of a comparison between chemical water quality determinants and river water fluorescence on the River Tyne, NE England, demonstrate that tryptophan‐like fluorescence intensity shows statistically significant relationships between nitrate, phosphate, ammonia, biochemical oxygen demand (BOD) and dissolved oxygen. Tryptophan‐like fluorescence intensity at the 280 nm excitation/350 nm emission wavelength fluorescence centre correlates with both phosphate (r = 0·80) and nitrate (r = 0·87), whereas tryptophan‐like fluorescence intensity at the 220 nm excitation/350 nm emission wavelength centre correlates with BOD (r = 0·85), ammonia (r = 0·70) and dissolved oxygen (r = −0·65). The strongest correlations are between tryptophan‐like fluorescence intensity and nitrate and phosphate, which in the Tyne catchment derive predominantly from point and diffuse source sewage inputs. The correlation between BOD and the tryptophan‐like fluorescence intensity suggests that this fluorescence centre is related to the bioavailable or labile dissolved organic matter pool. The weakest correlations are observed between tryptophan‐like fluorescence intensity and ammonia concentration and dissolved oxygen. The weaker correlation with ammonia is due to removal of the ammonia signal by wastewater treatment, and that with dissolved oxygen due to the natural aeration of the river such that this is not a good indicator of water quality. The observed correlations only hold true when treated sewage, sewerage overflows or cross connections, or agricultural organic pollutants dominate the water quality—this is not true for two sites where airport deicer (propylene glycol, which is non‐fluorescent) or landfill leachate (which contains high concentrations of humic and fulvic‐like fluorescent DOM) dominate the dissolved organic matter in the river. Mean annual tryptophan‐like fluorescence intensity agrees well with the General Water Quality Assessment as determined by the England and Wales environmental regulators, the Environment Agency. Copyright © 2004 John Wiley & Sons, Ltd.
Sensitivity evaluation of the Daphtoxkit and Thamnotoxkit microbiotests on blind samples
Authors: Törökné Andrea
Published: October 2004

At the Oregon State University Workshop we utilized two ‘culture/maintenance‐free’ microbiotests, the Thamnotoxkit F™ with the anostracan crustacean Thamnocephalus platyurus and the Daphtoxkit F™ magna with the cladoceran crustacean Daphnia magna, to determine the toxicity of water samples in a blind format. The Thamnotoxkit was applied to 7 samples and the Daphtoxkit to 12 samples. The chemical composition of the water samples to which the Toxkit microbiotests had been applied was disclosed a posteriori by the organizers and appeared to contain chlordimeform, colchicine, phosdrin, mercuric chloride, sodium arsenite, metham sodium, sodium cyanide, trimethylol propane phosphate, p‐chlorophenol and a natural sediment sample containing mercury. Three of the water samples were blanks that had not been spiked with chemicals. No false positives were obtained with the two Toxkits and all the toxic waters were earmarked as such by the microbiotests, except trimethylol propane phosphate, which was not found to induce acute effects in Daphnia magna in the non‐diluted water sample containing 100 mg l−1 of this compound. Copyright © 2004 John Wiley & Sons, Ltd.
EILATox‐Oregon Workshop: blind study evaluation of vitotox test with genotoxic and cytotoxic sample library
Authors: Meriläinen Jari , Lampinen Jorma
Published: October 2004

In order to assess the robustness, sensitivity and speciﬁcity of a recently developed Vitotox™ test, 17 blind coded chemicals and three environmental water samples were tested at the EILATox‐Oregon Workshop using the Thermo Electron Vitotox kit. The Vitotox test is a rapid geno‐ and cytotoxicity test using standard 96‐ or 384‐well microtitre plates. The genotoxicity test is based on two genetically modiﬁed Salmonella typhimurium strains containing bacterial luciferase operon from Vibrio ﬁsheri under the SOS inducible promoter. The SOS system is an inducible network in Escherichia coli that responds to DNA damage and activates DNA repair. The Vitotox genotoxicity test bacteria strain carries bacterial luciferase genes under the control of SOS inducible promoter and therefore any DNA damage inside the cells induces the production of bacterial luciferase. The luciferase expression is then followed with a microtitre plate luminometer for 3 h after mixing different dilutions of sample with the test bacteria. The genotoxicity index is calculated for each dilution and the genotoxicity of the sample is interpreted based on kinetic time curves and genotoxicity vs concentration/dilution curves. Cytotoxicity of the sample is determined simultaneously with another test strain containing the same luciferase operon controlled by the constitutive promoter. This bacterium produces constant bioluminescence and any decrease of the bioluminescence production is used as a marker for cytotoxicity.

As a miniaturized microtitre plate assay the Vitotox test requires a very small quantity of the sample material. The samples used in the workshop were diluted 1 : 10 or 1 : 100 before testing. Genotoxicity and cytotoxicity data were collected at dilutions of 1 : 10–1 : 2000. When the samples of the EILATox‐Oregon Workshop were tested using the Vitotox test, four coded chemicals out of 17 were determined to be genotoxic. Seven chemicals and one environmental sample were found to be cytotoxic. Three chemical samples were found to be both geno‐ and cytotoxic. Copyright © 2004 John Wiley & Sons, Ltd.
Detection of bioavailable heavy metals in EILATox‐Oregon samples using whole‐cell luminescent bacterial sensors in suspension or immobilized onto fibre‐optic tips
Authors: Hakkila Kaisa , Green Tal , Leskinen Piia , Ivask Angela , Marks Robert , Virta Marko
Published: October 2004

At the EILATox‐Oregon Workshop, nine luminescent whole‐cell bacterial sensors were used for the determination of bioavailable metals in blind samples (17 synthetic and 3 environmental). A non‐inducible luminescent control strain was used to determine sample matrix effects and bacterial toxicity. Whole‐cell bacterial sensors capable of determining arsenic, inorganic mercury and its organic derivatives, cadmium, lead or copper were used in suspensions and a bacterial sensor for the detection of inorganic mercury was immobilized onto ﬁbre‐optic tips using calcium alginate. Bioavailable amounts of metals were estimated using calibration plots, that were constructed to determine the range of metals giving rise to a linear relationship between luminescence and the amount of metals present in the standard solutions. EILATox‐Oregon sample 5, which contained 74 mg l−1 of Hg, gave a signiﬁcant response with both formats of the mercury sensor. The bioavailable amounts of mercury according to the measurement of bacterial sensor in suspension and immobilized onto a ﬁbre‐optic tip were 76 and 93 mg l−1, respectively. The bacterial sensor for arsenic and copper showed a response with sample 6 (58 mg l−1 of As) and sample 8 (400 mg l−1 of metham sodium), respectively. This study showed that the bacterial sensors in suspension or immobilized onto optical ﬁbres are capable of quantifying bioavailable metals from unknown samples. The measurement protocol of bacterial sensors is simple and possible to perform in the ﬁeld. Moreover, the samples do not need any pretreatment before analysis. Construction and characterization of the strain for the detection of bioavailable copper are described. Copyright © 2004 John Wiley & Sons, Ltd.
Toxicity detection from EILATox‐Oregon Workshop samples by using kinetic photobacteria measurement: the flash method
Authors: Hakkila Kaisa , Lappalainen Juha , Virta Marko
Published: October 2004

The Flash test, which can be used to measure the toxicity from clear, turbid and coloured samples with identical protocol, was used in the EILATox‐Oregon Workshop. During the workshop, 17 synthetic samples and three environmental samples were tested. In the Flash method the photobacteria Vibrio ﬁscheri are initially dispensed on top of the sample and the light output of the bioluminescence reaction is recorded at a rate of several readings per second. The change in the light production was measured at two points: 30 s and 30 min after exposure at room temperature. Toxicity calculations were done by using either the peak height of the sample or the peak height of the control sample (maximum signal at 0–2 s). When the peak height of the sample is used as a reference, the colour and turbidity effect of solid particles are taken into account throughout the whole measurement period. In the EILATox‐Oregon Workshop toxicity was seen in samples 1 (chlordimeform), 4 (phosdrin), 5 (HgCl2), 6 (sodium arsenite), 8 (metham sodium), 12 (p‐chlorophenol), 14 (sodium selenite) and 17 (MNNG). Only samples 1, 4 and 12 showed different ec50 values between different calculations of toxicity. The ec50 values were calculated and compared with the ec50 values of Microtox derived from the literature. The Flash test will not replace the standardized photobacteria test, but is the advanced version of it and is needed when the sample matrix is complex and when speed and low costs are required. Copyright © 2004 John Wiley & Sons, Ltd.
Blind and naïve classiﬁcation of toxicity by fish chromatophores
Authors: Mojovic Ljiljana , Dierksen Karen P. , Upson Rosalyn H. , Caldwell Bruce A. , Lawrence Jeannine R. , Trempy Janine E. , McFadden Philip N.
Published: October 2004

Cellular and molecular pathways involved in the ability of animals to change color have been studied previously as biosensors and cytosensors of active and toxic agents, but such studies generally have been limited to just a few standardized agents. Here we describe the performance of cultured chromatophore pigment cells from the ﬁn tissue of Siamese ﬁghting ﬁsh as sensors of toxic agents under blind sampling conditions at the September 2002 EILATox‐Oregon Workshop. Detection was accomplished by monitoring motor protein‐mediated movements of cellular pigment in chromatophores at both the gross population level as well as in singly imaged cells. Pigment responses were recorded both during the exposure of chromatophores to each blind sample as well as afterwards when the cells were examined for after‐effects by challenging them with clonidine, an adrenergic drug that induces standardized pigment movements. After recording all results and upon breaking the key to reveal the identities of the toxic agents, it was found that all of the toxic samples in the study had been distinguished accurately from non‐toxic controls that were included among the blind samples. Furthermore, it was revealed that most of the toxic agents detected had never before been tested or calibrated against chromatophores, demonstrating that detection can be achieved under naïve conditions that have not been optimized for the analysis of any particular toxic agent. Finally, by organizing the results into categories of pigment responses, a binary classiﬁcation tree was generated that distinguished each toxic agent as having a distinct response pattern from the others. Thus, chromatophore‐based cytosensors can discover toxicity in the absence of prior knowledge of the agent in question, and the categories of responses of the cells can be used to distinguish one toxic agent from another. Copyright © 2004 John Wiley & Sons, Ltd.
Water toxicity detection by a panel of stress‐responsive luminescent bacteria
Authors: Pedahzur R. , Polyak B. , Marks R.S. , Belkin S.
Published: October 2004

A panel of Escherichia coli strains harbouring different stress‐responsive promoters fused to a lux reporter system was used to assess the potential toxicity of 17 unknown model water samples. Using liquid cultures, nine out of 14 toxic samples were properly identiﬁed as toxic, whereas ﬁve were false negatives. All three non‐toxic controls were identiﬁed correctly (no false positives). Two strains containing promoter–lux fusions were also tested when immobilized onto ﬁbre‐optic tips. One genotoxic sample and six toxic samples were correctly identiﬁed in this manner. The potential advantages and limitations in the use of genetically engineered bacteria as biosensors for water toxicity are discussed in view of these results. Copyright © 2004 John Wiley & Sons, Ltd.
Detection and classiﬁcation of threat agents via high‐content assays of mammalian cells
Authors: Tencza Sarah B. , Sipe Michael A.
Published: October 2004

One property common to all chemical or biological threat agents is that they damage mammalian cells. A threat detection and classiﬁcation method based on the effects of compounds on cells has been developed. This method employs high‐content screening (HCS), a concept in drug discovery that enables those who practice cell‐based assays to generate deeper biological information about the compounds they are testing. A commercial image‐based cell screening platform comprising ﬂuorescent reagents, automated image acquisition hardware, image analysis algorithms, data management and informatics was used to develop assays and detection/classiﬁcation methods for threat agents. These assays measure a cell's response to a compound, which may include activation or inhibition of signal transduction pathways, morphological changes or cytotoxic effects. Data on cell responses to a library of compounds was collected and used as a training set. At the EILATox‐Oregon Workshop, cellular responses following exposure to unknown samples were measured by conducting assays of p38 MAP kinase, NF‐κB, extracellular‐signal related kinase (ERK) MAP kinase, cyclic AMPresponse element binding protein (CREB), cell permeability, lysosomal mass and nuclear morphology. Although the assays appeared to perform well, only four of the nine toxic samples were detected. However the system was speciﬁc, because no false positives were detected. Opportunities for improvement to the system were identiﬁed during the course of this enlightening workshop. Some of these improvements were applied in subsequent tests in the Cellomics laboratories, resulting in a higher level of detection. Thus, an HCS approach was shown to have potential in detecting threat agents, but additional work is necessary to make this a comprehensive detection and classiﬁcation system. Copyright © 2004 John Wiley & Sons, Ltd.
Cultured neuronal networks as environmental biosensors
Authors: O'Shaughnessy Thomas J. , Gray Samuel A. , Pancrazio Joseph J.
Published: October 2004

Contamination of water by toxins, either intentionally or unintentionally, is a growing concern for both military and civilian agencies and thus there is a need for systems capable of monitoring a wide range of natural and industrial toxicants. The EILATox‐Oregon Workshop held in September 2002 provided an opportunity to test the capabilities of a prototype neuronal network‐based biosensor with unknown contaminants in water samples. The biosensor is a portable device capable of recording the action potential activity from a network of mammalian neurons grown on glass microelectrode arrays. Changes in the action potential ﬁring rate across the network are monitored to determine exposure to toxicants. A series of three neuronal networks derived from mice was used to test seven unknown samples. Two of these unknowns later were revealed to be blanks, to which the neuronal networks did not respond. Of the ﬁve remaining unknowns, a signiﬁcant change in network activity was detected for four of the compounds at concentrations below a lethal level for humans: mercuric chloride, sodium arsenite, phosdrin and chlordimeform. These compounds — two heavy metals, an organophosphate and an insecticide — demonstrate the breadth of detection possible with neuronal networks. The results generated at the workshop show the promise of the neuronal network biosensor as an environmental detector but there is still considerable effort needed to produce a device suitable for routine environmental threat monitoring. Copyright © 2004 John Wiley & Sons, Ltd.
Use of a complex approach for assessment of metamizole sodium and acetylsalicylic acid toxicity, genotoxicity and cytotoxicity
Authors: Arkhipchuk V. V. , Goncharuk V. V. , Chernykh V. P. , Maloshtan L. N. , Gritsenko I. S.
Published: October 2004

A complex approach based on the use of test organisms belonging to different systematic groups (plants, invertebrates and vertebrates), as well as the nucleolar biomarker and the micronucleus test on their cells, was applied to assess the toxicity, cytotoxicity and genotoxicity of two pharmaceutical substances (metamizole sodium and acetylsalicylic acid) applied at ic50 concentrations for mammalian cells. The compound acetylsalicylic acid was evaluated at a concentration (1.6 × 103 mg l−1) that was non‐toxic for bioassays based on ﬁsh (Carassius auratus gibelio) and hydra (Hydra attenuata) and acutely toxic for bioassays with ceriodaphnia (Ceriodaphnia afﬁnis) and onion (Allium cepa). The metamizole sodium solution (6.25%) demonstrated acute toxicity for the whole set of test organisms. Both drugs, after their 30–360 min inﬂuence on the test organisms, ﬁrst changed the nucleolar size in plant and animal cells (i.e. the transcriptional activity of ribosomal genes was affected most signiﬁcantly). Moreover, the metamizole sodium solution caused nucleolar structural damage in 90% of hydra cells as early as after 30 min of exposure. The acetylsalicylic acid solution inhibited essentially the rate of cell division in the meristem of onion roots (the mitotic index decreased to 9.6‰, as compared 51.7‰ for the control). The carp incubation and the onion germination in the acetylsalicylic acid solution showed a reproducible increase in the frequency of cells with micronuclei (2 and 5.5 times, respectively) and double nuclei (3 and 1.5 times, respectively). The approach described herein may be applied for obtaining rapid, cost‐efﬁcient and useful supplementary data on different types of toxicity for marketed drugs as well as for drugs under development. Copyright © 2004 John Wiley & Sons, Ltd.
Response characteristics of an aquatic biomonitor used for rapid toxicity detection
Authors: van der Schalie W. H. , Shedd T. R. , Widder M. W. , Brennan L. M.
Published: October 2004

The response characteristics of an aquatic biomonitor that detects toxicity by monitoring changes in bluegill (Lepomis macrochirus Raﬁnesque) ventilatory and movement patterns were evaluated in single chemical laboratory studies at concentrations near the 96‐h LC50 concentration and at the EILATox‐Oregon Workshop in sequential tests of multiple unknown samples. Baseline data collected prior to exposure allows each ﬁsh to serve as its own control. When at least 70% of exposed ﬁsh exhibit ventilatory or movement parameters signiﬁcantly different from baseline observations, a group alarm is declared. In the laboratory studies, the aquatic biomonitor responded to the majority of chemicals at the 96‐h lc50 within an hour or less, although substantially higher response times were found for malathion and pentachlorophenol. Workshop tests of single chemical concentrations presented as blind samples were consistent with the laboratory test results. There were no alarms under control conditions in any test. Although data are limited, the aquatic biomonitor appears to respond more rapidly to chemicals causing membrane irritation, narcosis or polar narcosis than to acetylcholinesterase inhibitors or oxidative phosphorylation uncouplers. All four monitored parameters (ventilatory rate, cough rate, ventilatory depth and movement) contributed to identiﬁcation of ﬁrst alarms at acutely toxic levels. Understanding these response patterns can be useful in data interpretation for biomonitor applications such as surface water monitoring for watershed protection, wastewater treatment plant efﬂuent monitoring or source water monitoring for drinking water protection. Copyright © 2004 John Wiley & Sons, Ltd.
Monitoring genotoxicity during the photocatalytic degradation of p‐nitrophenol
Authors: Sekler M. Shani , Levi Y. , Polyak B. , Novoa A. , Dunlop P. S. M. , Byrne J. A. , Marks R. S.
Published: October 2004

p‐Nitrophenol is a common structural unit of many pesticides and was chosen as a model compound to monitor genotoxicity during photocatalytic degradation. The genotoxicity of p‐nitrophenol (PNP) and its breakdown products was measured using a bioluminescent bacterial bioassay, Vitotox™. The genotoxic potential decreased with the concomitant photocatalytic degradation of the parent PNP concentration. The rate of genotoxicity reduction was slower than the rate of removal of the parent PNP, due to the formation of genotoxic by‐products. After 6 h of photocatalytic treatment the total genotoxicity was removed. These results indicate that bioassays can be used as a simple and highly sensitive method for monitoring the general toxicity of chemical pollutants before, during and after photocatalytic treatment or other destructive processes. Copyright © 2004 John Wiley & Sons, Ltd.
Responses of fish chromatophore‐based cytosensor to a broad range of biological agents
Authors: Dierksen Karen P. , Mojovic Ljiljana , Caldwell Bruce A. , Preston R. Ryan , Upson Rosalyn , Lawrence Jeannine , McFadden Philip N. , Trempy Janine E.
Published: October 2004

A cytosensor based on living chromatophores from Betta splendens Siamese ﬁghting ﬁsh was used to test several classes of biologically active agents. Tested agents include neurotransmitters, adenyl cyclase activators, cytoskeleton effectors, cell membrane effectors and protein synthesis inhibitors. Characteristic cell responses were analyzed, and potential cytosensor applications were considered. Streptococcus pyogenes toxins streptolysin S and streptolysin O, Clostridium tetani tetanolysin, Staphylococcus aureus alpha‐toxin and Vibrio parahemolyticus hemolysin, all bacterial toxins that act on cell membranes, elicited a strong response from chromatophores. A comparison of puriﬁed toxin to actual bacterial culture from Vibrio parahemolyticus demonstrated a nearly identical chromatophore cell response pattern. This suggests that the cytosensor response is reﬂective of bacterial toxin production. Copyright © 2004 John Wiley & Sons, Ltd.
3D real time methodology monitoring cement failures in THA
Authors: Qi Gang , Li Jihui , Mann Kenneth A. , Mouchon W. Paul , Hamstad Marvin A. , Salehi Abraham , Whitten Stephen A.
Published: October 2004

The present work proposed a methodology to monitor cement microcrack formation in the cemented femoral stem construct using the acoustic emission technique. This technique provides a unique means to automatically tally the number of microcracks, to visualize microcrack distribution, and to animate the progress of crack formation in a given time window of a fatigue test. In this work, the formulation of microcrack source location was derived and a computer program was developed specifically for the proposed application. The program was validated using computer simulation and standard pencil lead break tests. It was found that the mathematical errors complied with the acceptable minimal error. Based on the pencil lead break tests, the average technical error used to estimate the resolution of this technique was 4.7 mm at the present stage. The program was then used to monitor the fatigue damage in precoated cemented femoral hip constructs loaded for a total of more than five million cycles. Two types of microcrack activities were observed in the experiments: Type I and Type II microcracks. A Type I microcrack was a crack that was captured by four or more sensors, and therefore its location was defined uniquely by a set of coordinates. A Type II microcrack was a crack that was captured by three or less sensors, therefore it was unlocatable. Both counts of Type I and Type II microcrack were tallied with respect to the day of fatigue tests. Acoustic emission microcrack graphs were used to visualize the distribution of Type I microcracks in the construct. It was found that the Type I microcracks distributed mainly over the proximal third of the stem. The amount of microcrack events decreased significantly as the number of loading cycles increased. © 2004 Wiley Periodicals, Inc. J Biomed Mater Res 71A: 391–402, 2004
Preparation and characterization of a highly macroporous biodegradable composite tissue engineering scaffold
Authors: Guan Limin , Davies John E.
Published: October 2004

A unique composite scaffold for bone‐tissue engineering applications has been prepared by combining biodegradable poly(lactide‐co‐glycolide) (PLGA) with bioresorbable calcium phosphate (CaP) cement particles through the process of particle fusion and phase separation/particle leaching. The scaffold is characterized by a highly interconnected macroporosity, with macropores of 0.8–1.8 mm and porosities ranging from 81% to 91%, and improved mechanical properties with respect to the polymer alone, producing excellent dimensional stability. The scaffold properties were controlled by adjusting the processing parameters, including PLGA molar mass and concentration, CaP/PLGA ratio, and porogen size. The differences in mechanical properties between dry, wet/room temperature, and wet/37°C testing conditions, of which the latter are more relevant for materials to be employed in a biological milieu, were investigated. Thus, a scaffold made from PLGA IV 1.13, PLGA concentration 12.5%, and CaP/PLGA ratio 2:1 exhibited significantly different compressive strengths of 0.16 MPa and 0.04 MPa when tested under dry and wet/37°C conditions, respectively. © 2004 Wiley Periodicals, Inc. J Biomed Mater Res 71A: 480–487, 2004
Bone morphogenetic protein‐2 decorated silk fibroin films induce osteogenic differentiation of human bone marrow stromal cells
Authors: Karageorgiou Vassilis , Meinel Lorenz , Hofmann Sandra , Malhotra Ajay , Volloch Vladimir , Kaplan David
Published: October 2004

Bone morphogenetic protein (BMP)‐2 has a critical role in bone formation and regeneration. Therefore, the ability to immobilize this molecule in certain matrices may be crucial in bone tissue engineering. Using carbodiimide chemistry, BMP‐2 was directly immobilized on silk fibroin films. Whereas human bone marrow stromal cells cultured on unmodified silk fibroin films in the presence of osteogenic stimulants exhibited little if any osteogenesis, the same cells cultured on BMP‐2 decorated films in the presence of osteogenic stimulants differentiated into an osteoblastic lineage as assessed by their significantly elevated alkaline phosphatase activity, calcium deposition, and higher transcript levels of collagen type I, bone sialoprotein, osteopontin, osteocalcin, BMP‐2, and cbfa1. Using cell culture inserts, it was demonstrated that differentiation was induced by the immobilized protein and not by protein released into the culture medium. Comparison with a similar amount of medium‐supplemented BMP‐2, where no additional protein was added with medium changes, showed that delivery of BMP‐2 immobilized on the biomaterial surface was more efficient than soluble delivery. The results illustrate that BMP‐2 covalently coupled on silk biomaterial matrices retains biological function in vitro based on the induction of osteogenic markers in seeded bone marrow stromal cells. © 2004 Wiley Periodicals, Inc. J Biomed Mater Res 71A: 528–537, 2004
Carboxy‐terminal fragment of osteogenic growth peptide regulates myeloid differentiation through RhoA
Authors: Mattii Letizia , Fazzi Rita , Moscato Stefania , Segnani Cristina , Pacini Simone , Galimberti Sara , D'Alessandro Delfo , Bernardini Nunzia , Petrini Mario
Published: October 2004

The carboxy‐terminal fragment of osteogenic growth peptide, OGP(10–14), is a pentapeptide with bone anabolic effects and hematopoietic activity. The latter activity appears to be largely enhanced by specific growth factors. To study the direct activity of OGP(10–14) on myeloid cells, we tested the pentapeptide proliferating/differentiating effects in HL60 cell line. In this cell line, OGP(10–14) significantly inhibited cell proliferation, and enhanced myeloperoxidase (MPO) activity and nitroblue tetrazolium reducing ability. Moreover, it induced cytoskeleton remodeling and small GTP‐binding protein RhoA activation. RhoA, which is known to be involved in HL60 differentiation, mediated these effects as shown by using its specific inhibitor, C3. Treatment with GM‐CSF had a comparable OGP(10–14) activity on proliferation, MPO expression, and RhoA activation. Further studies on cell proliferation and RhoA activation proved enhanced activity by association of the two factors. These results strongly suggest that OGP(10–14) acts directly on HL60 cells by activating RhoA signaling although other possibilities cannot be ruled out. © 2004 Wiley‐Liss, Inc.