Using aquatic macrophyte community indices to define the ecological status of European lakes

Defining the overall ecological status of lakes according to the Water Framework Directive (WFD) is to be partially based on the species composition of the aquatic macrophyte community. We tested three assessment methods to define the ecological status of the macrophyte community in response to a eutrophication pressure as reflected by total phosphorus concentrations in lake water. An absolute species richness, a trophic index (TI) and a lake trophic ranking (LTR) method were tested at Europe-wide, regional and national scales as well as by alkalinity category, using data from 1,147 lakes from 12 European states. Total phosphorus data were used to represent the trophic status of individual samples and were plotted against the calculated TI and LTR values. Additionally, the LTR method was tested in some individual lakes with a relatively long time series of monitoring data. The TI correlated well with total P in the Northern European lake types, whereas the relationship in the Central European lake types was less clear. The relationship between total P and light extinction is often very good in the Northern European lake types compared to the Central European lake types. This can be one of the reasons for a better agreement between the indices and eutrophication pressure in the Northern European lake types. The response of individual lakes to changes in the abiotic environment was sometimes represented incorrectly by the indices used, which is a cause of concern for the use of single indices in status assessments in practice.     

Potential conflicts between environmental legislation and conservation exemplified by aquatic macrophytes

It is important that legislation on water quality issues of freshwaters is not in conflict with nature conservation purposes. So far, it is however unknown how the assessment of ecological status according to for example the Water Framework Directive (WFD) of the European Community relates to the status of lakes according to the Habitat Directive (HD) or to national environmental objectives including, e.g., the protection of important wetland areas and red-listed species. We used lake macrophyte classification schemes of Norway, Sweden, and Finland and a total of 1,014 lakes to evaluate the possible conflict between these directives and national legislation. The classification schemes represent mainly trophic indices penalizing lakes with elevated phosphorous concentrations. In general, high ecological status according to the WFD did not mean high number of red-listed species or high status according to the HD or other national environmental objectives. In Sweden 78%, in Norway 47%, and in Finland 29% of lakes with red-listed species were classified as lakes of moderate or worse ecological status based on the macrophyte classification scheme. These lakes thus did not fulfill the demands of the WFD. Restoration of surface water toward fulfilling the demands requires in practice a reduction of the trophic status. This might potentially result in for example the loss of red-listed species. To avoid such potential conflicts, we primarily suggest revising the national quality assessment systems toward implicitly incorporating nature conservation aspects, e.g., the number of red-listed species in a multi-metric assessment system.     

Development of a phytoplankton indicator system for the ecological assessment of brackish coastal waters (German Baltic Sea coast)

The implementation of the European Water Framework Directive (WFD) requires the development of ecologically-based classification systems for anthropogenically-induced eutrophication in all types of water bodies. Due to the inherent high temporal and spatial variability of hydrological and geochemical parameters of the coastal waters of the southern Baltic Sea, discrimination between anthropogenic impact and natural variability is necessary. The development of statistical methods for this discrimination was the main aim of this study. These methods were used to derive indicative phytoplankton parameters for different stages of eutrophication for the investigation area. For this purpose, a long-term phytoplankton data series was analysed, which covered a broad salinity and eutrophication gradient. In order to detect eutrophication effects, the analysis was restricted to phytoplankton spring bloom events and to the salinity range between 5 and 10 psu, i.e. superimposing seasonal and hydrodynamic effects were eliminated. An artificial abiotic degradation vector was developed based on four typical water quality parameters. A total of 11 potentially indicative phytoplankton parameters on different taxonomical levels arose from a correlation analysis with this degradation vector. These indicators were then tested for their ability to discriminate between three eutrophication levels. Finally, seven phytoplankton indices could be proposed: total phytoplankton biovolume, the percentage of diatoms and the biovolume of different size ranges of diatoms and one indicative species (Woronichinia compacta). Guest editors: A. Razinkovas, Z. R. Gasiūnaitė, J. M. Zaldivar & P. Viaroli European Lagoons and their Watersheds: Function and Biodiversity     

Performance of a new phytoplankton composition metric along a eutrophication gradient in Nordic lakes

A new phytoplankton metric is presented, which is developed from a large dataset of Norwegian lakes (>2,000 samples from >400 lakes). In contrast to previous metrics, this index is not built on selected ‘indicative’ taxa, but uses all available taxonomic information at genus and species level. Taxa optima with respect to lake trophic status (derived from total phosphorus concentrations) are used to calculate a phytoplankton trophic index (TI) for each sample. Analysis of the TI shows that phytoplankton communities exhibit highly non-linear responses to eutrophication in Norwegian lakes. Reference lakes are characterized by very similar TIs despite having considerable variation in total phosphorus and chlorophyll a concentrations. TI exhibits a non-linear distribution along the eutrophication gradient which separates unimpacted from impacted sites in the study area. We further show that TI exhibits smaller seasonal variations than chlorophyll a, making it a more reliable indicator for lake monitoring. Implications for its applicability within the WFD are discussed.     

Ecological quality scales based on phytoplankton for the implementation of Water Framework Directive in the Eastern Mediterranean

Structural changes of phytoplankton communities, often expressed through ecological indices, constitute one of the metrics for the implementation of the European Water Framework Directive (WFD). In the current study a thorough analysis of the efficiency of 22 ecological indices was performed and a small number was selected for the development of five-level water quality scales (High, Good, Moderate, Poor, and Bad). The analysis was performed on simulated communities free of the noise of field communities due to uncontrolled factors or stochastic processes. Two criteria were set for the sensitivity of indices, namely their monotonicity and linearity across the studied eutrophication spectrum. The whole procedure was based on the development of a five-level quality assessment scheme based on phytoplankton abundance. Among the indices tested, the Menhinick diversity index and three indices of evenness were the most efficient, showing consistency (monotonic behavior) and linearity and were therefore used for the development of quality scales for the WFD. An Integrated Phytoplankton Index (IPI) based on three phytoplankton metrics, chlorophyll a, abundance, and diversity is also proposed. The efficiency of these indices was evaluated for a number of sites in the Aegean, already classified in the past by various methods based on nutrient concentrations or phytoplankton data. The results indicate that the various phytoplankton metrics (chlorophyll a, abundance, and diversity) assessed or proposed in the current study, carry their own information showing differences in the final classification of areas. Therefore the establishment of synthetic indices as the IPI seems to be advantageous for the integrated assessment of coastal water quality in the framework of European policies as the WFD.     

Exploring the robustness of macrophyte-based classification methods to assess the ecological status of coastal and transitional ecosystems under the Water Framework Directive

Identifying and quantifying the factors that contribute to the potential misclassification of the ecological status of water bodies is a major challenge of the Water Framework Directive (WFD). The present study compiles extensive biomonitoring data from a range of macrophyte-based classification methods developed by several European countries. The data reflect spatial and temporal variation as well as inter-observer variation. Uncertainty analysis identified that factors related to the spatial scale of sampling generally contributed most to the uncertainty in classifying water bodies to their ecological status, reflecting the high horizontal and depth-related heterogeneity displayed by macrophyte communities. In contrast, the uncertainty associated with temporal variation was low. In addition, inter-observer variation, where assessed, did not contribute much to overall uncertainty, indicating that these methods are easily transferable and insensitive to observer error. The study, therefore, suggests that macrophyte-based sampling schemes should prioritize large spatial replication over temporal replication to maximize the effectiveness and reliability of water body classification within the WFD. We encourage conducting similar uncertainty analyses for new/additional ecological indicators to optimize sampling schemes and improve the reliability of classification of ecological status.     

Ecological threshold responses in European lakes and their applicability for the Water Framework Directive (WFD) implementation: synthesis of lakes results from the REBECCA project

The objective of this synthesis is to present the key messages and draw the main conclusions from the work on lakes in the REBECCA project, pointing out their links to theoretical ecology and their applicability for the WFD implementation. Type-specific results were obtained from analyses of large pan-European datasets for phytoplankton, macrophytes, macroinvertebrates and fish, and indicators and relationships showing the impact of eutrophication or acidification on these biological elements were constructed. The thresholds identified in many of the response curves are well suited for setting ecological status class boundaries and can be applied in the intercalibration of classification systems. Good indicators for phytoplankton (chrysophytes, cyanobacteria) and macrophytes (isoetids and charaphytes) responses to eutrophication were identified, and the level of eutrophication pressure needed to reach the thresholds for these indicators was quantified. Several existing metrics developed for macrophytes had low comparability and need further harmonisation to be useful for intercalibration of classification systems. For macroinvertebrates, a number of metrics developed for rivers turned out to be less useful to describe lake responses to eutrophication and acidification, whereas other species based indicators were more promising. All the biological elements showed different responses in different lake types according to alkalinity and humic substances, and also partly according to depth. Better harmonisation of monitoring methods is needed to achieve better precision in the dose–response curves. Future research should include impacts of hydromorphological pressures and climate change, as well as predictions of timelags involved in responses to reduction of pressures.     

Quantifying uncertainties in biologically-based water quality assessment: A pan-European analysis of lake phytoplankton community metrics

Lake phytoplankton are adopted world-wide as a sensitive indicator of water quality. European environmental legislation, the EU Water Framework Directive (WFD), formalises this, requiring the use of phytoplankton to assess the ecological status of lakes and coastal waters. Here we provide a rigorous assessment of a number of proposed phytoplankton metrics for assessing the ecological quality of European lakes, specifically in response to nutrient enrichment, or eutrophication, the most widespread pressure affecting lakes. To be useful indicators, metrics must have a small measurement error relative to the eutrophication signal we want them to represent among lakes of different nutrient status. An understanding of variability in metric scores among different locations around a lake, or due to sampling and analytical variability can also identify how best this measurement error is minimised.To quantify metric variability, we analyse data from a multi-scale field campaign of 32 European lakes, resolving the extent to which seven phytoplankton metrics (including chlorophyll a, the most widely used metric of lake quality) vary among lakes, among sampling locations within a lake and through sample replication and processing. We also relate these metrics to environmental variables, including total phosphorus concentration as an indicator of eutrophication.For all seven metrics, 65–96% of the variance in metric scores was among lakes, much higher than variability occurring due to sampling/sample processing. Using multi-model inference, there was strong support for relationships between among-lake variation in three metrics and differences in total phosphorus concentrations. Three of the metrics were also related to mean lake depth. Variability among locations within a lake was minimal (<4%), with sub-samples and analysts accounting for much of the within-lake metric variance. This indicates that a single sampling location is representative and suggests that sub-sample replication and standardisation of analyst procedures should result in increased precision of ecological assessments based upon these metrics.For three phytoplankton metrics being used in the WFD: chlorophyll a concentration, the Phytoplankton Trophic Index (PTI) and cyanobacterial biovolume, >85% of the variance in metric scores was among-lakes and total phosphorus concentration was well supported as a predictor of this variation. Based upon this study, we can recommend that these three proposed metrics can be considered sufficiently robust for the ecological status assessment of European lakes in WFD monitoring schemes.     

Comparing macroinvertebrate indices to detect organic pollution across Europe: a contribution to the EC Water Framework Directive intercalibration

With the EC Water Framework Directive (WFD) the ecological status of a water body is defined by comparing the observed biological community composition present with near-natural reference conditions. The ecological status is then classified into five quality classes (high, good, moderate, poor and bad). It is of great importance that `good ecological status' has the same meaning within the European Union, since water bodies not measuring up to these standards have to be improved. Therefore, the Ecological Quality Ratios (EQR) at high-good, and good-moderate quality class boundaries will be intercalibrated. Each country has to report physical, chemical, and biological data from two sites at each of these boundaries and since most data exist for benthic macroinvertebrates, this quality element will be of great importance in the intercalibration process. The aim of this study was therefore to compare the results of different benthic macroinvertebrate metrics used to assess the impact of organic pollution (including eutrophication) (one of the major human impacts on European streams). A selection of the data sampled in the AQEM project was evaluated, where benthic macroinvertebrate- and abiotic data from four countries (Austria, the Czech Republic, Portugal and Sweden) and seven `stream types' were included. An organic pollution (including eutrophication) gradient was defined using Principal Component Analysis and the boundaries for high-good and good-moderate ecological status set by the partners from each country were used to define arbitrary class boundaries. The Average Score Per Taxon (ASPT) was well correlated with the stress gradient in most stream types, whereas the Saprobic Index worked clearly better than ASPT in those countries (Austria and the Czech Republic) where macroinvertebrates are generally identified to lower (species) as opposed to a higher (genus or family) level of identification. Defining harmonised class boundaries is difficult; this process has to consider the natural differences between stream types (e.g. in the reference values of metrics) but has to eliminate different perceptions of ecological quality.     

Testing the European stream typology of the Water Framework Directive for macroinvertebrates

In the EU Water Framework Directive (WFD) a typological framework is defined for assessing the ecological quality of water bodies in the future. The conditions in the Directive impose a strong demand for `new' assessment systems. During the AQEM project an assessment system was developed for European streams using macroinvertebrates. The aim of this study was to test if the typology suggested in the WFD is useful for developing an assessment system for macroinvertebrates in streams. In total 889 streams of 29 stream types were sampled in eight countries all over the major geographical gradients in Europe. These stream types fit the WFD typological demands and fit to the major European geographic regions (ecoregions). The sites included gradients from reference conditions (for the definition see Nijboer et al., 2004) to sites with bad ecological quality. Despite standardisation there were large differences between the participating countries concerning the number of taxa, the number of specimens and the taxonomic resolution. The data, including macroinvertebrates and environmental variables were analysed by using Canonical Correspondence Analysis (CCA). The observed macroinvertebrate distribution largely supported the WFD typological criteria. This means that the major macroinvertebrate distribution patterns in European streams follow climatological and geomorphological conditions and are well distinguished in terms of stream types. Furthermore, it was shown that large scale factors affected the macroinvertebrate distribution even on a very fine scale. Most explanatory variables seemed to be scale independent. Even at a fine scale major factors concerning geology, geomorphology and hydrology added to the species occurrences. Within stream types morphology together with physico-chemistry best explained the macroinvertebrates distribution. In conclusion, the WFD typology is useful for an assessment system for streams using macroinvertebrates. The large scale factors were indeed the variables that explained most of the variation in species composition. But as these factors even strongly act at the scale of stream types, a further refinement is most probably necessary to disentangle typological actors from water quality ones.     

Comparison of five methods for assessing impacts of nutrient enrichment using estuarine case studies

There are several approaches for assessing nutrient enrichment impacts in marine waters including the OSPAR* Comprehensive Procedure, the TRIX* ranking process, the WFD, ASSETS* and EPA NCA*. They differ in definitions and application, but all use key indicators for evaluating eutrophication status. Data from two estuaries in the United Kingdom (UK) were used to test the hypothesis that these five methods would result in the same outcome. The intent is to provide managers with information for selecting an appropriate method, insight about how each method works, how results from different methods compare and a basis for improvement of methods. Results from all approaches indicate that both estuaries required management intervention, but detailed results differed. Methods using more than one biological indicator show that secondary biological impacts were minimal, with the exception of moderate macroalgal problems in the Medway. Comparison of final results was difficult due to differences in: timeframes of data analysis (seasonal versus annual), characteristics included in indicator metrics (concentration, spatial coverage, frequency of occurrence), and methods for combining indicators to determine eutrophication status. This comparison suggests that use of annual data, inclusion frequency of occurrence, spatial coverage and, secondary biological indicators in the index, and a multi-category rating scale results in a more representative assessment.     

A synthesis of the biological quality elements for the implementation of the European Water Framework Directive in the Mediterranean ecoregion: The case of Saronikos Gulf

Benthic macroinvertebrates, macroalgae and phytoplankton constitute the biological quality elements proposed in the Water Framework Directive (WFD, 2000/60/EC) to be used for the classification of the ecological status of a water body. In the context of the preparation for the implementation of WFD, classification schemes for all three elements have been developed and tested for all the European ecoregions. In the present work, the classification schemes, with the corresponding metric tools and the interpretations of the class boundaries according to the normative definitions of WFD, are presented for each biological element in Saronikos Gulf, as case study in the Mediterranean ecoregion. The combination of the three biological elements into an integrated classification for coastal water bodies has been one of the major issues addressed in the context of WFD guidelines. Results are interpreted and validated, through an ecological viewpoint, on the basis of relevant environmental data. Moreover, this work presents a way to combine the EUNIS system to the typology of water bodies in the Mediterranean ecoregion.     

M-AMBI derived from taxonomic levels higher than species allows Ecological Status assessments of benthic habitats in new geographical areas

Under the Water Framework Directive (WFD) benthic Ecological Quality Ratios (EQRs) are important tools for assessing Ecological Status (ES) of coastal and transitional waters. Calculation of the Multivariate-AZTI Marine Biotic Index (M-AMBI) EQR is based on the proportions of sensitive and stress tolerant benthic invertebrate species, number of species and Shannon–Wiener diversity. The sensitivity of many tropical/sub-tropical taxa has not yet been determined, presenting a barrier to the direct transfer of WFD EQRs, and the ideas underpinning them, to the management of coastal waters beyond Europe. To overcome this we examine using higher taxonomic level data with M-AMBI.Before applying such approaches to assessing ES in new geographical regions it is essential to determine the effects of using higher taxonomic level data on M-AMBI in areas where the tool was developed. To this end, we use macrofaunal data from three well studied sites in north-western Europe to examine the effects of using taxonomic level data higher than species on M-AMBI. Using the European datasets M-AMBI ES classification was shown to be robust to changes in taxonomic level data. We test the suitability of family-level M-AMBI for assessing ES in subtropical Hong Kong waters. Family level M-AMBI was useful in detecting stress in Hong Kong, where it successfully detected temporal and spatial shifts in ES in response to seasonal hypoxia and salinity variability, and anthropogenic organic enrichment.     

Response of macroalgae and macroinvertebrates to anthropogenic disturbance gradients in rocky shores

The compliance of macroalgal and macroinvertebrate assemblages to anthropogenic disturbance gradients (e.g., nutrient enrichment) was investigated at intertidal rocky shores. Macroalgae and macroinvertebrates presented parallel behavior, both showing shifts in the communities’ structural variation along the gradients, in which an higher number of opportunistic species (and higher abundances) were found in more stressful sites (close to the disturbance source), in contrast to less disturbed sites (far from the disturbance source), which showed higher presence of more sensitive species (and higher abundance of several of them).The macroinvertebrate abundance and taxonomic composition, which are parameters required by the Water Framework Directive (WFD) to be included in tools for the ecological quality status assessment, responded to the disturbance gradient. Results suggest that the macroinvertebrate biological element might be considered an indicator of disturbance in intertidal rocky shores as good as the macroalgae, and therefore the development of a specific methodology based solely on benthic macroinvertebrates of rocky shores, presently a gap in the ecological quality status assessment for the WFD, seems feasible.     

Response of ecological indices to nutrient and chemical contaminant stress factors in Eastern Mediterranean coastal waters

Environmental data produced throughout monitoring activities in the framework of the implementation of Water Framework Directive 2000/60/EC (WFD) in Eastern Mediterranean (Greece) were used to assess the sensitivity and response of ecological indices against trace metals, eutrophication and multiple stress factors. The applied ecological indices include multi-metric eutrophication indices, a physicochemical status index applied for the first time in the Greek marine area, benthic indices, phytoplankton biomass index, and integrated status indices assessed through the application of the decision tree integration scheme. To investigate the exceedances in the eco-stoichiometric relationship between nutrients, considered a stressing factor, all physicochemical elements influenced directly or indirectly by eutrophication, such as nutrient concentrations, water transparency, oxygen saturation, particulates concentration, and sediment organic content, were related to ecological indices. Also, chemical contaminant stress factors represented by heavy metal concentrations in the water, as well as multiple stress factors represented by a pressure index, were related to ecological indices. A graphical visualization multivariate tool and statistical correlations were used to evaluate the sensitivity or explanatory power of the tested ecological indices against single and multiple stress factors. Results showed a strong response of all ecological indices to stress factors, although a diversification of sensitivity was evident. Primary production-related indices, i.e., macroalgae and chlorophyll-a indices, are more sensitive to particulates and nitrogen, while secondary production-related indices, i.e., benthic macroinvertebrates indices and eutrophication indices, including nutrients, are more sensitive to phosphates in the water column. The macroalgae index shows the strongest sensitivity to multiple stress factors. Among metals, mostly cadmium seems to match all indices⿿ performance. Nutrient relationships were shown as critical to eutrophication and ecological status.     

Phytoplankton communities and biomass size structure (fractionated chlorophyll “a”), along trophic gradients of the Basque coast (northern Spain)

Eutrophication is a major threat to coastal ecosystems. Within Europe, the Water Framework Directive (WFD) has established the need of developing methods of assessment. Bulk chlorophyll “a” is used world-wide as an indicator of eutrophication. However, the size structure of the phytoplankton communities has not been investigated, in detail, in relation to eutrophication pressures. This study investigates the suitability of fractionated chlorophyll “a” (<10 and >10 μm), as an indicator of eutrophication. Along the Basque coast (northern Spain) several water bodies, ranging from offshore waters to the heads of the estuaries, were surveyed during 2008. Physico-chemical conditions and chlorophyll “a” showed a distinct spatial gradient. Trophic richness increased towards the middle and inner parts of the estuaries, where nutrient inputs, from natural or anthropogenic sources, together with the residence time of the water are usually higher. In summer, phytoplankton biomass and abundance decreased, generally, in offshore and coastal waters; in estuaries, they tended to increase. The chlorophyll distribution within the two size fractions was coherent with the phytoplankton taxonomic composition. In summer, the relative abundance of small-sized cells (diatoms and non-siliceous taxa) increased. The relationships between size-fractionated chlorophyll and physico-chemical variables differed, when comparing the offshore and coastal waters, with the estuaries. In the offshore and coastal waters, a strong seasonality was observed; in estuaries, an important spatial component was found. Fractionated chlorophyll provided complementary useful information on anthropogenic pressures. However, more research is necessary to utilise this variable as a tool for ecological status assessment, within the WFD.     

Applying macrophyte community indicators to assess anthropogenic pressures on shallow soft bottoms

Vegetated soft bottoms are under pressure due to a number of anthropogenic stressors, such as coastal exploitation and eutrophication. The ecological value of these biotopes has gained recognition through international conventions and the EU directives, which request methods for assessment of the environmental status of coastal areas. However, currently there is no appropriate method for assessing the status of shallow vegetated soft bottoms in the northern Baltic Sea. Therefore, we developed a macrophyte community index and tested its response in relation to important pressures (eutrophication and boating activity) and natural gradients (topographic openness, depth and salinity) on shallow bays in the northern Baltic Sea. The macrophyte index, and hence the proportion of sensitive to tolerant species, decreased with increasing phosphorus concentration, turbidity and level of boating activity, while the cumulative cover of macrophytes only showed a negative trend in response to increasing turbidity. Juvenile fish abundance was positively related to the index, indicating importance of sensitive macrophyte species for ecosystem functioning. As the index was tested in a wide geographic area, and showed a uniform response across natural gradients, it is a promising tool for assessment of environmental status that may be applied also in other vegetated soft-bottom areas.     

Macroinvertebrate and diatom communities as indicators for the biological assessment of river Picentino (Campania,Italy)

Pollution of rivers is an increasing problem that affects biological diversity and structure of natural ecosystem. The present study reported the results of the preliminary analysis of diatom and macroinvertebrate communities of river Picentino (Italy, Campania) in respect of the WFD/60/2000/EC. Because of the sensitive to a variety of environmental factors the two categories of organism are used as excellent indicators of changes taking place in water ecosystems, especially eutrophication. Sampling of benthic diatoms and macroinvertebrates has been carried out in five stations along the river during April–May 2014 in order to apply the ICMi and the STAR_ICMi and assess river quality. The data showed a diversification in diatomic and macroinvertebrate communities in relation to environmental stresses and level of pollution, with the disappearance of species higher sensitive to eutrophication and organic load in the upper course of the river. River water quality was found to deteriorate from the upperstream to the downstream because of the increasing of human impact and the intensive agriculture activity along the river.We conclude that the monitoring of diatomic and macroinvertebrate communities could give detailed information about the ecological status of rivers. However it is necessary to increase the achieved data by monitoring other biological communities in order to define adequated strategies to save and preserve the rivers habitat.     

Evaluating the relative contributions of hydroperiod and soil fertility on growth of south Florida mangroves

The requirements of the European Water Framework Directive (WFD), aimed at an integrative assessment methodology for evaluating the ecological status of water bodies are frequently being achieved through multimetric techniques, i.e. by combining several indices, which address different stressors or different components of the biocoenosis. This document suggests a normative methodology for the development and application of Multimetric Indices as a tool with which to evaluate the ecological status of running waters. The methodology has been derived from and tested on a European scale within the framework of the AQEM and STAR research projects, and projects on the implementation of the WFD in Austria and Germany. We suggest a procedure for the development of Multimetric Indices, which is composed of the following steps: (1) selection of the most suitable form of a Multimetric Index; (2) metric selection, broken down into metric calculation, exclusion of numerically unsuitable metrics, definition of a stressor gradient, correlation of stressor gradients and metrics, selection of candidate metrics, selection of core metrics, distribution of metrics within the metric types, definition of upper and lower anchors and scaling; (3) generation of a Multimetric Index (general or stressor-specific approach); (4) setting class boundaries; (5) interpretation of results. Each step is described by examples.