Quantification of biogenic 3-D structures in marine sediments

Burrow and tube structures of marine infauna were quantified in sediments from cores obtained at 3 depths (75, 95 and 118 m) at 3 occasions over a 1-year period. The benthic communities at the two deeper stations were re-establishing subsequent to re-oxygenation after hypoxic conditions; the shallow station was a reference station unaffected by hypoxia. The benthic macrofauna was simultaneously quantified from sieved grab samples. The sediment cores were frozen and later sliced with a plane in horizontal layers. Biogenic structures in these sections were digitally analysed for numbers, area and volume. Number of tubes and burrows showed a general decline with depth in the sediment. The biogenic structures in the sediment at the reference station increased the sediment-water interface about 1.5 times compared to the surface area. The surface areas of burrows and tubes were 2 to 3 times larger at the reference station than at the deeper stations. The benthic fauna at the 95-m and 118-m stations showed a temporal increase in density and number of species. In contrast, development of biogenic surface areas in the sediment at these stations showed no significant temporal trend during the benthic faunal succession. Relic tubes of Melinna cristata were found throughout the sampling period at the 118-m station. The ecological importance of bioirrigation for biogeochemical processes is discussed.     

Seasonal Biotransformation of Naphthalene, Phenanthrene, and Benzo[a]pyrene in Surficial Estuarine Sediments

Transformation rates of naphthalene, phenanthrene, and benzo[a]pyrene in oxidized surficial sediments of a polluted urban estuary, Boston Harbor, Mass., were determined over a period of 15 months. Three sites characterized by muddy sediments were selected to represent a >300-fold range of ambient polycyclic aromatic hydrocarbon (PAH) concentration. Transformation rates were determined by a trace-level radiolabel PAH assay which accounted for PAH mineralization, the formation of polar metabolites, residue, and recovered parental PAHs in sediment slurries. Transformation rates of the model PAHs increased with increasing ambient PAH concentrations. However, turnover times for a given PAH were similar at all sites. The turnover times were as follows: naphthalene, 13.2 to 20.1 days; phenanthrene, 7.9 to 19.8 days, and benzo[a]pyrene, 53.7 to 82.3 days. At specific sites, rates were significantly affected by salinity, occasionally affected by temperature, but not affected by pH over the course of the study. Seasonal patterns of mineralization were observed for each of the PAHs at all sites. The timing of seasonal maxima of PAH mineralization varied from site to site. Seasonal potential heterotrophic activities as measured by acetate and glutamate mineralization rates did not always coincide with PAH mineralization maxima and minima, suggesting that the two processes are uncoupled in estuarine sediments.     

Effects of co-occurring aromatic hydrocarbons on degradation of individual polycyclic aromatic hydrocarbons in marine sediment slurries.

Rates of polycyclic aromatic hydrocarbon (PAH) degradation and mineralization were influenced by preexposure to alternate PAHs and a monoaromatic hydrocarbon at relatively high (100 ppm) concentrations in organic-rich aerobic marine sediments. Prior exposure to three PAHs and benzene resulted in enhanced [14C]naphthalene mineralization, while [14C]anthracene mineralization was stimulated only by benzene and anthracene preexposure. Preexposure of sediment slurries to phenanthrene stimulated the initial degradation of anthracene. Prior exposure to naphthalene stimulated the initial degradation of phenanthrene but had no effect on either the initial degradation or mineralization of anthracene. For those compounds which stimulated [14C]anthracene or [14C]naphthalene mineralization, longer preexposures (2 weeks) to alternative aromatic hydrocarbons resulted in an even greater stimulation response. Enrichment with individual PAHs followed by subsequent incubation with one or two PAHs showed no alteration in degradation patterns due to the simultaneous presence of PAHs. The evidence suggests that exposure of marine sediments to a particular PAH or benzene results in the enhanced ability of these sediments to subsequently degrade that PAH as well as certain other PAHs. The enhanced degradation of a particular PAH after sediments have been exposed to it may result from the selection and proliferation of specific microbial populations capable of degrading it. The enhanced degradation of other PAHs after exposure to a single PAH suggests that the populations selected have either broad specificity for PAHs, common pathways of PAH degradation, or both.     

Effects of co-occurring aromatic hydrocarbons on degradation of individual polycyclic aromatic hydrocarbons in marine sediment slurries

Rates of polycyclic aromatic hydrocarbon (PAH) degradation and mineralization were influenced by preexposure to alternate PAHs and a monoaromatic hydrocarbon at relatively high (100 ppm) concentrations in organic-rich aerobic marine sediments. Prior exposure to three PAHs and benzene resulted in enhanced [14C]naphthalene mineralization, while [14C]anthracene mineralization was stimulated only by benzene and anthracene preexposure. Preexposure of sediment slurries to phenanthrene stimulated the initial degradation of anthracene. Prior exposure to naphthalene stimulated the initial degradation of phenanthrene but had no effect on either the initial degradation or mineralization of anthracene. For those compounds which stimulated [14C]anthracene or [14C]naphthalene mineralization, longer preexposures (2 weeks) to alternative aromatic hydrocarbons resulted in an even greater stimulation response. Enrichment with individual PAHs followed by subsequent incubation with one or two PAHs showed no alteration in degradation patterns due to the simultaneous presence of PAHs. The evidence suggests that exposure of marine sediments to a particular PAH or benzene results in the enhanced ability of these sediments to subsequently degrade that PAH as well as certain other PAHs. The enhanced degradation of a particular PAH after sediments have been exposed to it may result from the selection and proliferation of specific microbial populations capable of degrading it. The enhanced degradation of other PAHs after exposure to a single PAH suggests that the populations selected have either broad specificity for PAHs, common pathways of PAH degradation, or both.     

Variation of a benthic heterotrophic bacteria community with different respiratory metabolisms in Coyuca de Benítez coastal lagoon (Guerrero, Mexico)

The fluctuations of the number, biomass and composition of the heterotrophic community were studied daily for two days, according to depth, pH, Eh, O2 and organic carbon concentration within a zone of the canal between the Coyuca de Benitez lagoon (Guerrero, Mexico) and the coastal waters. At the three moments of the day studied (6 am, 2 pm and 10 pm), the oxygen concentrations in the overlying water and in the superficial sediment layer were near air-saturation in the diurnal samplings (582 microM at 6 am and 665 microM at 2 pm), and sub-satured during the night (158 microM). In the sediments, the models of vertical distribution of Eh and organic carbon distributions were very irregular due to the bio-perturbation of the benthic, meio- and macrofauna, whose activity allows the superficial organic carbon to migrate towards sediment deeper layers. Vertical distribution of the different viable bacteria populations seems to be related to the hydrodynamic patterns of the communicating canal and sediments heterogeneity. In the sediment column, the heterotrophic bacteria total number varied from 6.8 to 20.3 x 108 cells cm(-3). The highest heterotrophic bacterial biomass values were encountered during the diurnal samplings (39.2 microgC.l(-1) at 6 am and 34.4 microgC.(l(-1) at 2 pm) and the lowest during the night (9.7 microgC.l(-1). The fluctuations of viable heterotrophic bacteria populations with different respiratory metabolisms (aerobic, microaerophilic and anaerobic) can be explained by the existence of suboxic microniches that appear when particles of sediment are resuspended due to the water circulation and the benthic infauna excavating activity, that allows the supernatant water oxygen to penetrate through its galleries towards deeper sediment zones. The statistical analysis (Multiple lineal regression model r2 > or = 0.5) showed that the on the whole, the hydrological parameters are not influence over the bacterial number and bacterial biomass distribution (r2 < or = 0.5), Nevertheless, the variations of the heterotrophic bacteria community observed in the two days sampling, seem to be governed (with F-values of 0.6 to 0.9) by the irregular flows of bio-available organic material and the sediment porosity.     

An Integrated Case Study for Evaluating the Impacts of an Oil Refinery Effluent on Aquatic Biota in the Delaware River: Integration and Analysis of Study Components

A series of statistical and graphical techniques incorporating a “weight of evidence” approach were used to interpret results from an integrated Triad case study designed to determine potential environmental impacts to aquatic biota in the Delaware River that may be linked to PAHs found in Motiva's oil refinery effluent. Sediment concentrations of various metals, PCBs and LMW PAHs exceeding both ERL and ERM sediment quality guidelines (SQGs) were reported in the study area. However, most chemical contaminants did not exceed their respective SQGs. Results from a long-term sediment coring study indicated that there was no evidence of significant historical PAH contamination of sediments related to Motiva's exceedences. PAHs comprising the Motiva “fingerprint” were found in the surficial sediments at four near-field sites but non-Motiva PAH concentrations (background) were shown to be significantly higher at other far-field sites (non-Motiva influence). Chronic sediment toxicity appears to have significant relationships to the patterns of most PAH isomers, certain PCB isomers, and certain metals. However, sediment toxicity does not appear to be related to the PAH isomers that are characteristic of Motiva's effluent nor to the near-field sites. Impacted benthic communities were reported in the study area, primarily at one near-field and two far-field sites. However, there were no apparent relationships between benthic community health and sediment contaminants. The status of benthic communities does not appear to be related to PAHs derived from the Motiva effluent. The “weight of evidence” analysis developed from a systematic and comprehensive series of statistical and graphical assessments indicates that, although the study area displayed some degree of sediment contamination, chronic sediment toxicity, and benthic health impacts, these environmental effects generally could not be related to Motiva's exceedences.     

Biogenic Sediment Compounds in Relation to Marine Meiofaunal Abundances

Correlations between a series of biogenic sediment compounds, commonly used in ecological studies, and a major component of the benthic infauna, the meiofauna, were studied on the continental margin off Southwest Africa (Angola) and in a central oceanic region of the Atlantic Ocean (Mid Oceanic Ridge). Biogenic sediment compounds chosen for this investigation (electron-transport-system activity, total adenylates and energy charge, particulate proteins, chloroplastic pigments) are obviously not suitable for a quick and rough estimation of meiofaunal abundances. Nevertheless, biogenic sediment compounds might reflect quite well the activity and biomass of the total benthic infauna, including all size classes (from bacteria to macrofaunal organisms) and/or the total particulate organic matter within the sediments. Furthermore, analyzing biogenic sediment compounds leads to a better understanding of environmental conditions and biological activities of benthic organisms. Consequently, despite their limitations, biochemical sediment parameters may be very useful in benthic ecological studies to obtain rapid information on the eco-status of the benthic system.     

Dissolved Oxygen Saturation Controls PAH Biodegradation in Freshwater Estuary Sediments

Polycyclic aromatic hydrocarbons (PAHs) are common contaminants in terrestrial and aquatic environments and can represent a significant constituent of the carbon pool in coastal sediments. We report here the results of an 18-month seasonal study of PAH biodegradation and heterotrophic bacterial production and their controlling biogeochemical factors from 186 sediment samples taken in a tidally influenced freshwater estuary. For each sampling event, measurements were averaged from 25–45 stations covering ∼250 km². There was a clear relationship between bacterial production and ambient temperature, but none between production and bottom water dissolved oxygen (DO) % saturation or PAH concentrations. In contrast with other studies, we found no effect of temperature on the biodegradation of naphthalene, phenanthrene, or fluoranthene. PAH mineralization correlated with bottom water DO saturation above 70% (r² > 0.99). These results suggest that the proportional utilization of PAH carbon to natural organic carbon is as much as three orders of magnitude higher during cooler months, when water temperatures are lower and DO % saturation is higher. Infusion of cooler, well-oxygenated water to the water column overlying contaminated sediments during the summer months may stimulate PAH metabolism preferentially over non-PAH organic matter.     

Polycyclic Aromatic Hydrocarbons in Sediments: An Overview of Risk-Related Issues

This series of articles address site-specific issues associated with evaluating exposure and toxicity of polycyclic aromatic hydrocarbons (PAHs) in sediments. These include factors that influence the opportunity to come into contact with PAHs in sediments (bioaccessibility) and those that affect the potential for transfer from sediment to ecological and human receptors (bioavailability). Although organic carbon is viewed as an important matrix for sorbing PAHs, studies have shown that there are various forms of carbon, some of which are highly sorptive. These latter forms vary on a site-specific basis and including this form of carbon in the assessment can reduce the uncertainty associated with estimating exposure. Other site-specific factors associated with water clarity, depth, and light penetration can result in enhanced toxicity of PAHs as a result of photoactivation. Chemical analyses of sediments increasingly include the alkylated PAH compounds. Although analyses of this suite of PAH compounds is being conducted to support ecological assessments and forensic analyses, there is little guidance on how to interpret the alkylated PAHs with respect to human health risks. An approach for accomplishing this is suggested.     

Polycyclic Aromatic Hydrocarbons in Coastal Sediment of Klang Strait,Malaysia: Distribution Pattern,Risk Assessment and Sources

Concentration, source, and ecological risk of polycyclic aromatic hydrocarbons (PAHs) were investigated in 22 stations from surface sediments in the areas of anthropogenic pollution in the Klang Strait (Malaysia). The total PAH level in the Klang Strait sediment was 994.02±918.1 µg/kg dw. The highest concentration was observed in stations near the coastline and mouth of the Klang River. These locations were dominated by high molecular weight PAHs. The results showed both pyrogenic and petrogenic sources are main sources of PAHs. Further analyses indicated that PAHs primarily originated from pyrogenic sources (coal combustion and vehicular emissions), with significant contribution from petroleum inputs. Regarding ecological risk estimation, only station 13 was moderately polluted, the rest of the stations suffered rare or slight adverse biological effects with PAH exposure in surface sediment, suggesting that PAHs are not considered as contaminants of concern in the Klang Strait.     

Risk assessment of polycyclic aromatic hydrocarbons (PAHs) in sediments from a mixed-use reservoir

Although reservoirs in China are of great significance, very few studies on risk assessment have been reported for reservoirs. This study investigated distribution characteristics, cancer and ecological risks, and source diagnosis of 16 priority polycyclic aromatic hydrocarbons (PAHs) in sediments from Shitou Koumen Reservoir in Jilin Province, China. A total of 12 sediment samples were collected from the reservoir in August (wet season) 2014. Total PAH concentrations in sediment samples ranged from 1294.51 ng/g to 2755.35 ng/g with a mean concentration of 1757.54 ng/g. For individual PAHs, average concentration of Nap was the highest, 800.56 ng/g, while Acy, Fla, BkF, and DahA were undetected in sediment samples. Light PAHs (2–3 rings) accounting for 74.21% was a dominant PAH compositional pattern. Pearson correlation analysis was carried out; results showed that total PAHs was strongly correlated with the highly enriched sedimentary PAHs, and pH was a major factor in controlling PAH distribution. Lifetime cancer risk was employed to assess cancer risk; results indicated that the fish-culturing area was exposed to cancer risk. The molecular diagnostic ratios of isomeric PAHs were applied to identify possible PAH sources; primary PAH sources were identified as oil-related activities, burning agricultural wastes, vehicular emissions, and industrial discharges.     

Distribution characteristics,sources, and ecological risk assessment of polycyclic aromatic hydrocarbons in sediments from Haizhou Bay,China

Sixteen USEPA priority polycyclic aromatic hydrocarbons (PAHs) were analyzed by gas chromatography–mass spectrometry. Twenty samples were collected from the surface sediments of Haizhou Bay in this survey. This research aimed to identify the PAHs' contamination level, composition pattern, pollution sources, and assess the ecological risk of PAHs. The results showed that the sum of PAH concentrations ranged from 116.6 ng g^?1 to 2414.9 ng g^?1 (mean: 662.42 ng g^?1), which is higher than the reported values for different wetlands worldwide. Three- and four-ring PAHs (accounts for more than 70% of the total PAH content) were predominant in the wetland sediment. The PAHs source distribution in the surface sediments were determined using diagnostic ratio and PCA/MLR. Consequently, multiple PAHs sources were found. Of the total PAH, 79.25% was derived from vehicular emission, 20.75% from coal combustion. The effect range low/effect range median (ERL/ERM) values indicated a low toxicity risk level. However, the fluoranthene concentrations exceeded the ERL level, and even the ERM level, in some stations. The mean effects range–median quotient (M-ERM-Q) suggests a low ecological risk for the PAHs in the sediments.     

Vertical Profile of PAHs and Dredging Depth of River Sediments

Sediment dredging is an effective engineering measure to reduce the negative effects of PAHs pollution on water environment. The dredging depth is a key parameter in environmental dredging engineering. A guidance of environmental dredging depth needed to reduce the toxicity risk was developed which was specifically designed for the removal of river sediments contaminated by PAHs based on the mean effect range median quotients (mean-ERM-q). The dredging depths for removing river sediments were calculated based on the vertical profile of PAHs content and the quality guidelines of sediment. Pinghu water system was carried out to determine the dredging depths of sediments from nine rivers by the proposed method. Each sediment core was collected from different river. The results showed the profiles of PAHs in sediment cores were irregular and diverse due to river dredging events and human activities in different periods. The risk assessment of PAHs toxicity showed one of nine cores with moderate-high probability, 4 of nine cores with low-moderate probability and the others with low toxicity risk. The achievement can offer a reference to the dredging engineering in other similar river systems.     

Functioning of a Shallow-Water Sediment System during Experimental Warming and Nutrient Enrichment

Effects of warming and nutrient enrichment on intact unvegetated shallow-water sediment were investigated for 5 weeks in the autumn under simulated natural field conditions, with a main focus on trophic state and benthic nitrogen cycling. In a flow-through system, sediment was exposed to either seawater at ambient temperature or seawater heated 4°C above ambient, with either natural or nutrient enriched water. Sediment–water fluxes of oxygen and inorganic nutrients, nitrogen mineralization, and denitrification were measured. Warming resulted in an earlier shift to net heterotrophy due to increased community respiration; primary production was not affected by temperature but (slightly) by nutrient enrichment. The heterotrophic state was, however, not further strengthened by warming, but was rather weakened, probably because increased mineralization induced a shortage of labile organic matter. Climate-related warming of seawater during autumn could therefore, in contrast to previous predictions, induce shorter but more intensive heterotrophic periods in shallow-water sediments, followed by longer autotrophic periods. Increased nitrogen mineralization and subsequent effluxes of ammonium during warming suggested a preferential response of organisms driving nitrogen mineralization when compared to sinks of ammonium such as nitrification and algal assimilation. Warming and nutrient enrichment resulted in non-additive effects on nitrogen mineralization and denitrification (synergism), as well as on benthic fluxes of phosphate (antagonism). The mode of interaction appears to be related to the trophic level of the organisms that are the main drivers of the affected processes. Despite the weak response of benthic microalgae to both warming and nutrient enrichment, the assimilation of nitrogen by microalgae was similar in magnitude to rates of nitrogen mineralization. This implies a sustained filter function and retention capacity of nutrients by the sediment.     

PAH mineralization and bacterial organotolerance in surface sediments of the Charleston Harbor estuary

Semi-volatile organic compounds (SVOCs) in estuarine waters can adversely affect biota but watershed sources can be difficult to identify because these compounds are transient. Natural bacterial assemblages may respond to chronic, episodic exposure to SVOCs through selection of more organotolerant bacterial communities. We measured bacterial production, organotolerance and polycyclic aromatic hydrocarbon (PAH) mineralization in Charleston Harbor and compared surface sediment from stations near a known, permitted SVOC outfall (pulp mill effluent) to that from more pristine stations. Naphthalene additions inhibited an average of 77% of bacterial metabolism in sediments from the more pristine site (Wando River). Production in sediments nearest the outfall was only inhibited an average of 9% and in some cases, was actually stimulated. In general, the stations with the highest rates of bacterial production also were among those with the highest rates of PAH mineralization. This suggests that the capacity to mineralize PAH carbon is a common feature amongst the bacterial assemblage in these estuarine sediments and could account for an average of 5.6% of bacterial carbon demand (in terms of production) in the summer, 3.3% in the spring (April) and only 1.2% in winter (December).     

A Statistical Evaluation of Sediment Quality Triad Benthic Community Measurements in PAH-Impacted Sediments at a Manufactured Gas Plant Site along the Hudson River

The sediment quality triad (SQT) assumes that three measurements (sediment chemistry, laboratory bioassay, and benthic macroinvertebrate counts) comprise an independent assessment of impact, which when integrated using a weight-of-evidence approach provides a comprehensive assessment of risk. An SQT assessment was conducted on 41 sediment samples collected adjacent to a manufactured gas plant site on the freshwater reach of the Hudson River in New York State. The assessment shows that the benthic macroinvertebrate data did not correlate with either sediment or pore water polycyclic aromatic hydrocarbon (PAH) concentrations, nor did these data show consistent relationships to the results of laboratory bioassay testing (Hyalella azteca 28-day survival or biomass). The benthic community across the site and reference areas was comprised of few taxa, all of which were pollution-tolerant organisms with tolerance values greater than or equal to five. Only in significantly impacted sediment samples with PAH concentrations in the thousands of milligrams per kilogram, pore water concentrations above 100 toxic units, and visible non-aqueous phase liquid present in the sample did the benthic macroinvertebrate data show a response. In contrast, sediment and pore water PAH measurements and H. azteca toxicity testing provided consistent interpretation of impact. These results illustrate that benthic macroinvertebrate data may contain less information value and be a more challenging line of evidence to interpret in triad studies conducted in certain ecological settings; in this case, a large-order river with a relatively depauperate benthic community dominated by species tolerant of PAHs.     

Ediacaran–Cambrian bioturbation did not extensively oxygenate sediments in shallow marine ecosystems

The radiation of bioturbation during the Ediacaran–Cambrian transition has long been hypothesized to have oxygenated sediments, triggering an expansion of the habitable benthic zone and promoting increased infaunal tiering in early Paleozoic benthic communities. However, the effects of bioturbation on sediment oxygen are underexplored with respect to the importance of biomixing and bioirrigation, two bioturbation processes which can have opposite effects on sediment redox chemistry. We categorized trace fossils from the Ediacaran and Terreneuvian as biomixing or bioirrigation fossils and integrated sedimentological proxies for bioturbation intensity with biogeochemical modeling to simulate oxygen penetration depths through the Ediacaran–Cambrian transition. Ultimately, we find that despite dramatic increases in ichnodiversity in the Terreneuvian, biomixing remains the dominant bioturbation behavior, and in contrast to traditional assumptions, Ediacaran–Cambrian bioturbation was unlikely to have resulted in extensive oxygenation of shallow marine sediments globally.     

Ecological Risk Assessment of Sediments in Sydney Harbour,Nova Scotia,Canada

Contamination within sediments of Sydney Harbour (once a major industrial port) were evaluated using a multiple lines-of-evidence (LOE) ecological risk assessment (ERA) approach prior to divestiture of the harbor. The multiple LOE approach included: (1) measurement of polycyclic aromatic hydrocarbons (PAHs), polychlorinated biphenyls (PCBs), metals, metalloids, petroleum hydrocarbons(PHCs), and total organic carbon (TOC) concentrations in surface sediments from multiple Sydney Harbour locations; (2) identification and application of sediment quality guidelines (SQGs) from various jurisdictions; (3) comparisons of harbor sediment chemistry against background/reference sediment chemistry; (4) determining number and frequency of exceedances over SQGs; (5) calculating mean probable effect level-quotients (PEL-Qs); (6) PAH forensic source evaluation; (7) review of previous sediment chemistry and biota tissue data; and (8) characterizing benthic habitat at harbor stations. The ERA determined that current sediments exhibited mostly low probability of adverse effects. Furthermore, contaminated sediments exhibiting a high probability of adverse effects were localized to only a few stations within the harbor. Ongoing natural recovery of harbor sediments is likely responsible for attenuating contaminants that historically were higher than those measured in this study and were previously distributed over much wider areas of the harbor. Results suggest that legacy industrial activities and current urban sewage effluents are the major sources of contamination in Sydney Harbour sediments.     

Evaluating Human Risk from Exposure to Alkylated PAHs in an Aquatic System

Polycyclic aromatic hydrocarbons (PAHs) are a large class of organic chemicals typically found as mixtures in the aquatic environment from natural, petrogenic, and pyrogenic sources. People can be exposed to PAHs through ingestion or dermal contact with contaminated sediments or through ingestion of finfish and shellfish exposed to contaminated sediments. Although more than 100 PAHs have been identified, human exposure and risk are commonly evaluated for 18 individual PAHs. Other PAHs, such as alkylated PAHs, likely contribute to biological activity of environmental PAH mixtures; however, insufficient toxicity data are available to quantify their potential risk. This article presents an initial evaluation of the potential for human health risk from exposure to alkylated PAHs in sediment and fish. Individual alkylated PAHs have been observed to have potentially mutagenic, tumor-promoting, or carcinogenic activity. However, except for 1-and 2-methylnaphthalene, insufficient toxicity data are available to quantify toxicity or cancer risk from exposure to individual alkylated PAHs or mixtures of alkylated PAHs. This article describes a proposed strategy to better understand the potential human health risk from exposure to alkylated PAHs. Implementation of this strategy will contribute to evaluations of human exposure to complex PAH mixtures in the environment.