A novel mechanism by which silica defends grasses against herbivory.

BACKGROUND AND AIMS: Previous studies have shown that silica in grass leaves defends them against small herbivores, which avoid high-silica grasses and digest them less efficiently. This study tested the idea that silica can reduce digestibility by preventing the mechanical breakdown of chlorenchyma cells. METHODS: Both the percentage of total chlorophyll liberated from high- and low-silica grass leaves by mechanical grinding and the chlorophyll content of locust faeces were measured. KEY RESULTS: High-silica grasses released less chlorophyll after grinding and retained more after passing through the gut of locusts, showing that silica levels correlated with increased mechanical protection. CONCLUSIONS: These results suggest that silica may defend grasses at least in part by reducing mechanical breakdown of the leaf, and that mechanical protection of resources in chlorenchyma cells is a novel and potentially important mechanism by which silica protects grasses.

The source and distribution of polychlorinated dibenzo-p-dioxin and polychlorinated dibenzofurans in sediments of Port Jackson, Australia.

Concentrations of polychlorinated dibenzo-p-dioxins (dioxins)(PCDDs) and polychlorinated dibenzofurans (PCDFs)(furans) in sediments from Port Jackson ranged from 32 to 4352 pg WHO-TEQ DF g(-1)dw with a mean of 712 pg g(-1)dw. Maximum total dioxins plus furans concentrations in this sediment was 6,290,000 pg g(-1)dw, which is the highest recorded in Australia and among the highest reported for sediment globally. Permanent bans were placed on fin fishing and prawn trawling in February, 2006 for the whole estuary, based on dioxin tissue burdens. A distinct congener profile corresponded to chemicals known to have been produced by industry on the shores of Homebush Bay in the upper estuary. These chemicals are being dispersed extensively and exceed pre-anthropogenic concentrations in even distant, less developed parts of the harbour. Remediation of sediments is currently being undertaken, with the intention of containing dispersion of dioxins and reducing body burdens in fin fish and prawn populations.

Identifying primary stressors impacting macroinvertebrates in the Salinas River (California, USA): relative effects of pesticides and suspended particles.

Laboratory dose-response experiments with organophosphate and pyrethroid pesticides, and dose-response experiments with increasing particle loads were used to determine which of these stressors were likely responsible for the toxicity and macroinvertebrate impacts previously observed in the Salinas River. Experiments were conducted with the amphipod Hyalella azteca, the baetid mayfly Procloeon sp., and the midge Chironomus dilutus (Shobanov, formerly Chironomus tentans). The results indicate the primary stressor impacting H. azteca was pesticides, including chlorpyrifos and permethrin. The mayfly Procloeon sp. was sensitive to chlorpyrifos and permethrin within the range of concentrations of these pesticides measured in the river. Chironomus dilutus were sensitive to chlorpyrifos within the ranges of concentrations measured in the river. None of the species tested were affected by turbidity as high as 1000 NTUs. The current study shows that pesticides are more important acute stressors of macroinvertebrates than suspended sediments in the Salinas River.

Monitoring structural changes in cells with high-frequency ultrasound signal statistics.

We investigate the use of signal envelope statistics to monitor and quantify structural changes during cell death using an in vitro cell model. Using a f/2.35 transducer (center frequency 20 MHz), ultrasound backscatter data were obtained from pellets of acute myeloid leukemia cells treated with a DNA-intercolating chemotherapy drug, as well as from pellets formed with mixtures of treated and untreated cells. Simulations of signals from pellets of mixtures of cells were generated as a summation of point scatterers. The signal envelope statistics were examined by fitting the Rayleigh and generalized gamma distributions. The fit parameters of the generalized gamma distribution showed sensitivity to structural changes in the cells. The scale parameter showed a 200% increase (p<0.05) between untreated and cells treated for 24 h. The shape parameter showed a 50% increase (p<0.05) over 24 h. Experimental results showed reasonable agreement with simulations. The results indicate that high-frequency ultrasound signal statistics can be used to monitor structural changes within a very low percentage of treated cells in a population, raising the possibility of using this technique in vivo.

High-frequency ultrasound scattering from microspheres and single cells.

Assessing the proportion of biological cells in a volume of interest undergoing structural changes, such as cell death, using high-frequency ultrasound (20-100 MHz), requires the development of a theoretical model of scattering by any arbitrary cell ensemble. A prerequisite to building such a model is to know the scattering by a single cell in different states. In this paper, a simple model for the high-frequency acoustic scattering by one cell is proposed. A method for deducing the backscatter transfer function from a single, subresolution scatterer is also devised. Using this method, experimental measurements of backscatter from homogeneous, subresolution polystyrene microspheres and single, viable eukaryotic cells, acquired across a broad, continuous range of frequencies were compared with elastic scattering theory and the proposed cell scattering model, respectively. The resonant features observed in the backscatter transfer function of microspheres were found to correspond accurately to theoretical predictions. Using the spacing of the major spectral peaks in the transfer functions obtained experimentally, it is possible to predict microsphere diameters with less than 4% error. Such good agreement was not seen between the cell model and the measured backscatter from cells. Possible reasons for this discrepancy are discussed.

A comparison of in situ and laboratory toxicity tests with the estuarine amphipod Eohaustorius estuarius.

Amphipod survival in laboratory and in situ exposures was assessed using the eastern Pacific Haustoriid species Eohaustorius estuarius. Toxicity test results were compared using intact (unhomogenized) and homogenized sediment samples in both field and laboratory exposures. Experiments were conducted in Moss Landing Harbor, California, an impaired waterbody under 303 (d) of the US Clean Water Act. Synoptic laboratory and in situ sediment toxicity tests were conducted at two stations: Sandholdt Bridge (SB), the most contaminated station in the harbor, and at the South Jetty (SJ), a more marine station near the mouth of the harbor. We found that Eohaustorius is amenable to in situ testing. Despite highly variable field salinity regimes (6-32 PSU at SB and 21-34 PSU at SJ), in situ control survival was 87% and 84% at SB and SJ, respectively. Amphipod survival was lower in the in situ exposures relative to the laboratory exposures at both sites. Survival at SB was 30% and 76% in the homogenized in situ and laboratory samples, respectively, and 40% and 64% in the intact (unhomogenized) in situ and laboratory samples, respectively. Neither the homogenized or intact samples from the SJ station were toxic in laboratory experiments, but amphipod survival was only 40% in the intact in situ exposure at this station, possibly due to predation. These experiments suggest that the interaction of contaminants and variable physical parameters such as salinity and temperature may have resulted in lower survival in the in situ exposures. Sediment homogenization prior to in situ deployment may have reduced effects of predators in some samples.

Integrated assessment of the impacts of agricultural drainwater in the Salinas River (California, USA).

The Salinas River is the largest of the three rivers that drain into the Monterey Bay National Marine Sanctuary in central California. Large areas of this watershed are cultivated year-round in row crops and previous laboratory studies have demonstrated that acute toxicity of agricultural drainwater to Ceriodaphnia dubia is caused by the organophosphate (OP) pesticides chlorpyrifos and diazinon. In the current study, we used a combination of ecotoxicologic tools to investigate incidence of chemical contamination and toxicity in waters and sediments in the river downstream of a previously uncharacterized agricultural drainage creek system. Water column toxicity was investigated using a cladoceran C. dubia while sediment toxicity was investigated using an amphipod Hyalella azteca. Ecological impacts of drainwater were investigated using bioassessments of macroinvertebrate community structure. The results indicated that Salinas River water downstream of the agricultural drain is acutely toxic to Ceriodaphnia, and toxicity to this species was highly correlated with combined toxic units (TUs) of chlorpyrifos and diazinon. Laboratory tests were used to demonstrate that sediments in this system were acutely toxic to H. azteca, which is a resident genus. Macroinvertebrate community structure was moderately impacted downstream of the agricultural drain input. While the lowest macroinvertebrate abundances were measured at the station demonstrating the greatest water column and sediment toxicity and the highest concentrations of pesticides, macroinvertebrate metrics were more significantly correlated with bank vegetation cover than any other variable. Results of this study suggest that pesticide pollution is the likely cause of laboratory-measured toxicity in the Salinas River samples and that this factor may interact with other factors to impact the macroinvertebrate community in the system.

Causes of sediment toxicity to Mytilus galloprovincialis in San Francisco Bay, California.

Since the San Francisco Regional Monitoring Program (RMP) sampling began, elutriate samples prepared with sediment from the Grizzly Bay monitoring station have been consistently toxic to bivalve larvae (Mytilus galloprovincialis). An investigation into the cause of toxicity was initiated with a Phase I Toxicity Identification Evaluation (TIE) using bivalve embryos. TIE results and chemical analyses of elutriate samples suggested that divalent metals were responsible for the observed toxicity. Following the initial characterization of trace metals as toxicants, additional TIEs were performed on elutriates prepared from three additional Grizzly Bay samples collected between 1997 and 2001. Additional TIEs included ethylenediamine tetraacetic acid (EDTA) treatments in a sediment-water interface (SWI) exposure system, and the use of a cation exchange column with serial elution of sample fractions with hydrochloric acid of increasing normality. EDTA significantly reduced toxicity in overlying water in the SWI system. The cation exchange column reduced both toxicity and concentrations of trace metals, and serial elution of the column added back both toxicity and specific metals contained in individual acid fractions. Chemical analyses of three elutriate samples demonstrated copper concentrations were within the range toxic to bivalves. Results of Phase I TIEs, additional Phase II treatments, SWI exposures, and metals analyses indicate the potential for metal toxicity in sediments from this estuarine site. When combined with the results of standard TIE methods, a solid-phase cation extraction and elution approach identified copper as the most probable cause of toxicity.

Causes of ambient toxicity in the Calleguas Creek watershed of southern California.

A combination of toxicity tests, chemical analyses, and Toxicity Identification Evaluations (TIEs) were used to investigate receiving water toxicity in the Calleguas Creek watershed of southern California. Studies were conducted from 1995 through 1999 at various sites to investigate causes of temporal variability of toxicity throughout this system. Causes of receiving water toxicity varied by site and species tested. Investigations in the lower watershed (Revolon Slough, Santa Clara Drain, Beardsley Wash) indicated that toxicity of samples to the cladoceran Ceriodaphnia dubia was due to elevated concentrations of the organophosphate pesticide chlorpyrifos, while causes of intermittent toxicity to fathead minnows (Pimephales promelas) and the alga Selanastrum capricornutum were less clear. Investigations at sites in the middle and upper reaches of the watershed (Arroyo Simi and Conejo Creek) indicated that the pesticide diazinon was the probable cause of receiving water toxicity to Ceriodaphnia. Elevated ammonia was the cause of toxicity to fathead minnows in the upper watershed sites. Results of these and previous studies suggest that biota are impacted by degraded stream quality from a variety of point and non-point pollution sources in the Calleguas Creek watershed. Water quality resource manager's efforts to identify contaminant inputs and implement source control will be improved with the findings of this study.

Ultrasonic spectral parameter characterization of apoptosis.

Ultrasound (US) spectral analysis methods are used to analyze the radiofrequency (RF) data collected from cell pellets exposed to chemotherapeutics that induce apoptosis and other chemicals that induce nuclear transformations. Calibrated backscatter spectra from regions-of-interest (ROI) were analyzed using linear regression techniques to calculate the spectral slope and midband fit. Two f/2 transducers, with operating frequencies of 30 and 34 MHz (relative bandwidths of 93% and 78%, respectively) were used with a custom-made imaging system that enabled the collection of the raw RF data. For apoptotic cells, the spectral slope increased from 0.37 dB/MHz before drug exposure to 0.57 dB/MHz 24 h after, corresponding to a change in effective scatterer radius from 8.7 to 3.2 microm. The midband fit increased in a time-dependent fashion, peaking at 13dB 24 h after exposure. The statistical deviation of the spectral parameters was in close agreement with theoretical predictions. The results provide a framework for using spectral parameter methods to monitor apoptosis in in vitro and in in vivo systems and are being used to guide the design of system and signal analysis parameters.

An evaluation of methods for calculating mean sediment quality guideline quotients as indicators of contamination and acute toxicity to amphipods by chemical mixtures.

Mean sediment quality guideline quotients (mean SQGQs) were developed to represent the presence of chemical mixtures in sediments and are derived by normalizing a suite of chemicals to their respective numerical sediment quality guidelines (SQGs). Mean SQGQs incorporate the number of SQGs exceeded and the degree to which they are exceeded and are used for comparison with observed biological effects in the laboratory or field. The current research makes it clear, however, that the number and type of SQGs used in the derivation of these mean quotients can influence the ability of mean SQGQ values to correctly predict acute toxicity to marine amphipods in laboratory toxicity tests. To determine the optimal predictive ability of mean SQGQs, a total of 18 different chemical combinations were developed and compared. The ability of each set of mean SQGQs to correctly predict the presence and absence of acute toxicity to amphipods was determined using three independent databases (n = 605, 2753, 226). Calculated mean SQGQ values for all chemical combinations ranged from 0.002 to 100. The mean SQGQ that was most predictive of acute toxicity to amphipods is calculated as SQGQ1 = ((sigma ([cadmium]/4.21 )([copper]/270)([lead]/ 12.18)([silver]/1.77)([zinc]/ 410)([total chlordane]/6)([dieldrin]/8)([total PAHoc]/1,800)([total PCB]/400))/9). Both the incidence and magnitude of acute toxicity to amphipods increased with increasing SQGQI values. To provide better comparability between regions and national surveys, SQGQ1 is recommended to serve as the standard method for combination of chemicals and respective SQGs when calculating mean SQGQs.

Influence of sample manipulation on contaminant flux and toxicity at the sediment-water interface.

Toxicities of sediments from San Diego and San Francisco Bays were compared in laboratory experiments using sea urchin (Strongylocentrotus purpuratus) embryos exposed to pore water and at the sediment-water interface (SWI). Toxicity was consistently greater to embryos exposed at the SWI to intact (unhomogenized) sediment samples relative to homogenized samples. Measurement of selected trace metals indicated considerably greater fluxes of copper, zinc, and cadmium into overlying waters of intact sediment samples. Inhibition of sea urchin embryo development was generally greater in sediment pore waters relative to SWI exposures. Pore water toxicity may have been due to elevated unionized ammonia concentrations in some samples. The results indicate that invertebrate embryos are amenable to SWI exposures, a more ecologically relevant exposure system, and that sediment homogenization may create artifacts in laboratory toxicity experiments.

A large-scale categorization of sites in San Francisco Bay, USA, based on the sediment quality triad, toxicity identification evaluations, and gradient studies.

Sediment quality was assessed in San Francisco Bay, California, USA, using a two-tiered approach in which 111 sites were initially screened for sediment toxicity. Sites exhibiting toxicity were then resampled and analyzed for chemical contamination, recurrent toxicity, and, in some cases, benthic community impacts. Resulting data were compared with newly derived threshold values for each of the metrics in a triad-based weight-of-evidence evaluation. Sediment toxicity test results were compared with tolerance limits derived from reference site data, benthic community data were compared with threshold values for a relative benthic index based on the presence and abundance of pollution-tolerant and -sensitive taxa, and concentrations of chemicals and chemical mixtures were compared with sediment quality guideline-based thresholds. A total of 57 sites exceeded threshold values for at least one metric, and each site was categorized based on triad inferences. Nine sites were found to exhibit recurrent sediment toxicity associated with elevated contaminant concentrations, conditions that met program criteria for regulatory attention. Benthic community impacts were also observed at three of these sites, providing triad evidence of pollution-induced degradation. Multi- and univariate correlations indicated that chemical mixtures, heavy metals, chlordanes, and other organic compounds were associated with measured biological impacts in the Bay. Toxicity identification evaluations indicated that metals were responsible for pore-water toxicity to sea urchin larvae at two sites. Gradient studies indicated that the toxicity tests and benthic community metrics employed in the study predictably tracked concentrations of chemical mixtures in Bay sediments.

Evaluation and use of sediment toxicity reference sites for statistical comparisons in regional assessments.

Sediment reference sites were used to establish toxicity standards against which to compare results from sites investigated in San Francisco Bay (California, USA) monitoring programs. The reference sites were selected on the basis of low concentrations of anthropogenic chemicals, distance from active contaminant sources, location in representative hydrographic areas of the Bay, and physical features characteristic of depositional areas (e.g., fine grain size and medium total organic carbon [TOC]). Five field-replicated sites in San Francisco Bay were evaluated over three seasons. Samples from each site were tested with nine toxicity test protocols and were analyzed for sediment grain size and concentrations of trace metals, trace organics, ammonia, hydrogen sulfide, and TOC. The candidate sites were found to have relatively low concentrations of measured chemicals and generally exhibited low toxicity. Toxicity data from the reference sites were then used to calculate numerical tolerance limits to be used as threshold values to determine which test sites had significantly higher toxicity than reference sites. Tolerance limits are presented for four standard test protocols, including solid-phase sediment tests with the amphipods Ampelisca abdita and Eohaustorius estuarius and sea urchin Strongylocentrotus purpuratus embryo/larval development tests in pore water and at the sediment-water interface (SWI). Tolerance limits delineating the lowest 10th percentile (0.10 quantile) of the reference site data distribution were 71% of the control response for Ampelisca, 70% for Eohaustorius, 94% for sea urchin embryos in pore water, and 87% for sea urchins embryos exposed at the SWI. The tolerance limits are discussed in terms of the critical values governing their calculation and the management implications arising from their use in determining elevated toxicity relative to reference conditions.

Statistical significance of sediment toxicity test results: threshold values derived by the detectable significance approach.

A number of methods have been employed to determine the statistical significance of sediment toxicity test results. To allow consistency among comparisons, regardless of among-replicate variability, a protocol-specific approach has been used that considers protocol performance over a large number of comparisons. Ninetieth-percentile minimum significant difference (MSD) values were calculated to determine a critical threshold for statistically significant sample toxicity. Significant toxicity threshold values (as a percentage of laboratory control values) are presented for six species and nine endpoints based on data from as many as 720 stations. These threshold values are useful for interpreting sediment toxicity data from large studies and in eliminating cases where statistical significance is assigned in individual cases because among-replicate variability is small.

Sediment quality in Los Angeles Harbor, USA: a triad assessment.

Sediment quality in the Los Angeles and Long Beach Harbor area of southern California, USA, was assessed from 1992 to 1997 as part of the California State Water Resources Control Board's Bay Protection and Toxic Cleanup Program and the National Oceanic and Atmospheric Administration's National Status and Trends Program. The assessment strategy relied on application of various components of the sediment quality triad, combined with bioaccumulation measures, in a weight-of-evidence approach to sediment quality investigations. Results of bulk-phase chemical measurements, solid-phase amphipod toxicity tests, pore-water toxicity tests with invertebrate embryos, benthic community analyses (presented as a relative benthic index), and bioaccumulation measures indicated that inner harbor areas of this system are polluted by high concentrations of a mixture of sediment-associated contaminants and that this pollution is highly correlated with toxicity in laboratory experiments and degradation of benthic community structure. While 29% of sediment samples from this system were toxic to amphipods (Rhepoxynius abronius or Eohaustorius estuarius), 79% were toxic to abalone embryos (Haliotis rufescens) exposed to 100% pore-water concentrations. Statistical analyses indicated that amphipod survival in laboratory toxicity tests was significantly correlated with the number of crustacean species and the total number of species measured in the benthos at these stations. Triad measures were incorporated into a decision matrix designed to classify stations based on degree of sediment pollution, toxicity, benthic community degradation, and, where applicable, tissue concentrations in laboratory-exposed bivalves and feral fish.

Ultrasound imaging of apoptosis: high-resolution non-invasive monitoring of programmed cell death in vitro, in situ and in vivo.

A new non-invasive method for monitoring apoptosis has been developed using high frequency (40 MHz) ultrasound imaging. Conventional ultrasound backscatter imaging techniques were used to observe apoptosis occurring in response to anticancer agents in cells in vitro, in tissues ex vivo and in live animals. The mechanism behind this ultrasonic detection was identified experimentally to be the subcellular nuclear changes, condensation followed by fragmentation, that cells undergo during apoptosis. These changes dramatically increase the high frequency ultrasound scattering efficiency of apoptotic cells over normal cells (25- to 50-fold change in intensity). The result is that areas of tissue undergoing apoptosis become much brighter in comparison to surrounding viable tissues. The results provide a framework for the possibility of using high frequency ultrasound imaging in the future to non-invasively monitor the effects of chemotherapeutic agents and other anticancer treatments in experimental animal systems and in patients.

An investigation of the flow dependence of temperature gradients near large vessels during steady state and transient tissue heating.

Temperature distributions measured during thermal therapy are a major prognostic factor of the efficacy and success of the procedure. Thermal models are used to predict the temperature elevation of tissues during heating. Theoretical work has shown that blood flow through large blood vessels plays an important role in determining temperature profiles of heated tissues. In this paper, an experimental investigation of the effects of large vessels on the temperature distribution of heated tissue is performed. The blood flow dependence of steady state and transient temperature profiles created by a cylindrical conductive heat source and an ultrasound transducer were examined using a fixed porcine kidney as a flow model. In the transient experiments, a 20 s pulse of hot water, 30 degrees C above ambient, heated the tissues. Temperatures were measured at selected locations in steps of 0.1 mm. It was observed that vessels could either heat or cool tissues depending on the orientation of the vascular geometry with respect to the heat source and that these effects are a function of flow rate through the vessels. Temperature gradients of 6 degrees C mm(-1) close to large vessels were routinely measured. Furthermore, it was observed that the temperature gradients caused by large vessels depended on whether the heating source was highly localized (i.e. a hot needle) or more distributed (i.e. external ultrasound). The gradients measured near large vessels during localized heating were between two and three times greater than the gradients measured during ultrasound heating at the same location, for comparable flows. Moreover, these gradients were more sensitive to flow variations for the localized needle heating. X-ray computed tomography data of the kidney vasculature were in good spatial agreement with the locations of all of the temperature variations measured. The three dimensional vessel path observed could account for the complex features of the temperature profiles. The flow dependences of the transient temperature profiles near large vessels during the pulsed experiments were consistent with the temperature distributions measured in the steady state experiments and provided unique insights into the process of convective heat transfer in tissues. Finally, it was shown that even for very short treatment times (3-20 s), large vessels had significant effects on the tissue temperature distributions.