Soil eDNA biomonitoring reveals changes in multitrophic biodiversity and ecological health of agroecosystems.

Soil health is integral to sustainable agroecosystem management. Current monitoring and assessment practices primarily focus on soil physicochemical properties, yet the perspective of multitrophic biodiversity remains underexplored. Here we used environmental DNA (eDNA) technology to monitor multitrophic biodiversity in four typical agroecosystems, and analyzed the species composition and diversity changes in fungi, bacteria and metazoan, and combined with the traditional physicochemical variables to establish a soil health assessment framework centered on biodiversity data. First, eDNA technology detected rich multitrophic biodiversity in four agroecosystems, including 100 phyla, 273 classes, 611 orders, 1026 families, 1668 genera and 1146 species with annotated classification, and the relative sequence abundance of dominant taxa fluctuates tens of times across agroecosystems. Second, significant differences in soil physicochemical variables such as organic matter (OM), total nitrogen (TN) and available phosphorus (AP) were observed among different agroecosystems, nutrients were higher in cropland and rice paddies, while heavy metals were higher in fish ponds and lotus ponds. Third, biodiversity metrics, including α and ß diversity, also showed significant changes across agroecosystems, the soil biota was generally more sensitive to nutrients (e.g., OM, TN or AP), while the fungal communities were mainly affected by heavy metals in October (e.g., Cu and Cr). Finally, we screened 48 sensitive organismal indicators and found significant positive consistency between the developed eDNA indices and the traditional soil quality index (SQI, reaching up to R2 = 0.58). In general, this study demonstrated the potential of eDNA technology in soil health assessment and underscored the importance of a multitrophic perspective for efficient monitoring and managing agroecosystems.

The sediment of a river having ''good ecological status" turned out to be toxic to midge Chironomus riparius larvae: Implication for environmental monitoring?

Globally, monitoring of the surface waters is largely limited to the physico-chemical analysis of water in rivers and lakes. Sediment state in the aquatic systems including sediment chemical content or the structure and diversity of benthic communities or ecotoxicological studies with natural sediments remains largely overlooked by the monitoring programs. Hence we assessed the potential toxicity of three riverine sediments on the life-cycle traits (emergence and reproduction) of midge Chironomus riparius via an ecotoxicological testing method over two generations (according to OECD test 233 guidelines). In addition, the riverine sediments were spiked with polyamide (nylon) microplastic particles (1 g kg-1) to analyze an additive effect of microplastic on the sediment toxicity. As model river systems, three rivers (Karchaghbyur, Gavaraget, Argichi) in the Lake Sevan basin (Armenia) were selected. Results of ecotoxicity testing were compared with the indices of water quality (derived from the physico-chemical analysis) and the indices of the ecological status of the rivers (derived from the analysis of benthic communities). The results of testing demonstrated an unexpectedly low emergence of midges after the first generation exposed to the sediment of the river having ''good ecological status'' - the Argichi. Sediments of the Karchaghbyur and Gavaraget rivers impeded the emergence and reproduction of midges after the second generation. An addition of polyamide particles to the sediments did not significantly affect the life-cycle traits of C. riparius indicating the primary effect of the sediments' condition. The discrepancy of biotesting result with that of the other two methods (which indicated ''average water quality'' and "good ecological status") underlies the importance of designing more comprehensive monitoring programs for better assessment and protection of aquatic systems and resources.

Evidence of interregional similarity in crayfish metabolomes at reference sites: Progress towards the metabolome as a biomonitoring tool.

It has been proposed that biomonitoring may benefit from the use of metabolomics (the study of all small molecules in an organism) to detect sub-lethal organism stress through changes in the metabolite profile (i.e., the metabolome). However, to integrate the metabolome into biomonitoring programs the amount of natural variability among and within populations of indicator taxa must be established prior to generating a reference condition. This study determined variation in the metabolome among ecoregion and stream of origin in the northern crayfish (Faxonius virilis) and if that variation inhibited detection of stressor effects at sites exposed to human activities. We collected crayfish from seven minimally disturbed streams (i.e., reference streams), distributed across three level II ecoregions in central Canada and compared their metabolomes. We found ecoregion and stream origin were poor predictors of crayfish metabolomes. This result suggests crayfish metabolomes were similar, despite differing environmental conditions. Metabolomes of crayfish collected from three stream sites exposed to agricultural activity and municipal wastewater (i.e., test sites) were then compared to the crayfish metabolomes from the seven reference streams. Findings showed that crayfish metabolomes from test sites were strongly differentiated from those at all reference sites. The consistency in the northern crayfish metabolome at the studied reference streams indicates that a single reference condition may effectively detect impacts of human activities across the sampled ecoregions.

Response of a pan-European fish index (EFI+) to multiple pressures in rivers across Spain.

Rivers are ecosystems highly threatened by human activities and fish are an invaluable tool to measure and communicate environmental degradation and restoration. Fish bioassessment is crucial but notoriously difficult in Mediterranean-climate streams for a number of reasons, including low local species richness, faunas with high spatial turnover and generalist species, and scarcity of reference sites. In this study, we conducted the most comprehensive test of the pan-European fish index (EFI+) in the Iberian Peninsula, analysing its response to multiple anthropogenic pressures. We compiled a database, which we provide online, with 2970 electrofishing samples across Spain, involving 100,732 fish of 69 species. Principal component analyses of many quantitative variables were used to create new synthetic anthropogenic pressure indices. Correlation and multiple linear regression analyses were used to test the relationship between these pressures and the fish index (EFI+) and its four individual metrics scores (i.e., density of species intolerant to oxygen depletion, density of fish ≤150 mm of species intolerant to habitat degradation, richness of species of rheophilic reproduction habitat, and density of species of lithophilic reproduction habitat). We also obtained the same models but including the river basin district to test for spatial or methodological differences. Our results indicate that both the EFI+ index and its individual metrics respond to various anthropogenic pressures. These pressures explained about 36% of the variance of EFI+ values. Notably, downstream and mainstream reaches with higher agricultural or urban land uses, increased hydrologic alteration, and water and habitat quality impairment exhibited lower EFI+ values. Although less variance was explained for the individual metrics than for the fish index, they responded as expected to the different pressures. For instance, the richness of rheophilic species and the number of lithophilic fish decreased with hydrologic alteration, while the number of fish intolerant to oxygen depletion decreased with water quality impairment. Similar correlations were observed when river basin district was included in the model, but with higher explained variation and greater significance of the pressures. While it is possible to develop regional indices with more metrics and a stronger correlation with anthropogenic pressures, EFI+ is the only fish index that has been validated throughout the Spanish peninsular territory. Our results support the use of EFI+ in intercalibration exercises across Spain until better regional indices are developed.

The Roles of Microbes in Stream Restorations.

The goods and services provided by riverine systems are critical to humanity, and our reliance increases with our growing population and demands. As our activities expand, these systems continue to degrade throughout the world even as we try to restore them, and many efforts have not met expectations. One way to increase restoration effectiveness could be to explicitly design restorations to promote microbial communities, which are responsible for much of the organic matter breakdown, nutrient removal or transformation, pollutant removal, and biomass production in river ecosystems. In this paper, we discuss several design concepts that purposefully create conditions for these various microbial goods and services, and allow microbes to act as ecological restoration engineers. Focusing on microbial diversity and function could improve restoration effectiveness and overall ecosystem resilience to the stressors that caused the need for the restoration. Advances in next-generation sequencing now allow the use of microbial 'omics techniques (e.g., metagenomics, metatranscriptomics) to assess stream ecological conditions in similar fashion to fish and benthic macroinvertebrates. Using representative microbial communities from stream sediments, biofilms, and the water column may greatly advance assessment capabilities. Microbes can assess restorations and ecosystem function where animals may not currently be present, and thus may serve as diagnostics for the suitability of animal reintroductions. Emerging applications such as ecological metatranscriptomics may further advance our understanding of the roles of specific restoration designs towards ecological services as well as assess restoration effectiveness.

A versatile and economical method for collecting macroinvertebrates using Hester-Dendy multiplate samplers.

We present an inexpensive, versatile, and robust mounting system for Hester-Dendy (HD) multiplate samplers that are useful in aquatic biological studies and freshwater biomonitoring programs. Detailed instructions are provided outlining the construction and deployment of a concrete block system featuring threaded anchors for screwing in HD columns in a vertical position. Additionally, eye bolts provide a central attachment point for cabling the block securely to the stream or river bank, and for attachment of a buoy or physiochemical data logger if desired. All the components of the block system are inexpensive, readily available, and assembled with no special skills required. The system offers superior ease-of-use and a more standardized sampling device compared to other methods.

No lukewarm diatom communities-the response of freshwater benthic diatoms to phosphorus in streams as basis for a new phosphorus diatom index (PDISE).

In the present study, we developed a new Swedish phosphorus diatom index (PDISE) to improve the poor fit of existing indices to match the needs of water managers to detect and mitigate eutrophication. We took advantage of a large amount of data (820 Swedish stream sites) collected in recent years. During our work, we found an unexpected bimodal response of the diatom assemblages to phosphorus. The taxa clustered either into an assemblage with a low or with a high site-specific averaged TP optimum (a calculated value comprised of the diatom taxa-specific optima). We could not find a characteristic diatom assemblage for sites with intermediate site-specific averaged TP optima. To our knowledge, this bimodal community response has not been shown earlier. The PDISE correlated more strongly than the currently used TDI to changes in TP concentrations. Thus, the PDISE should replace the TDI in the Swedish standard method. The modeled TP optima (expressed as categories) were different compared to the TDI for most of the taxa included in the index, indicating that the realized niche for these morphotaxa was different between Sweden and the UK where the TDI was developed originally. With a r2 of 0.68, the correlation of the PDISE to TP is among the highest reported for other diatom nutrient indices globally; thus, we believe that it might be worth to test it for other bioregions with similar geography and climate.

Ecological integrity assessment of streams in the light of natural ecoregions and anthropic land use.

Reference conditions for river bioassessment should be established inside ecoregions. Our objectives were (1) to implement a bioassessment methodology for Uruguayan prairie streams regarding ecoregions and land use and (2) to assess the ecological integrity of streams of the Río Negro basin in Uruguay. Due to logistical constraints, sampling was divided into two collection trips: one including the upper basin in fall 2015 and the other including the lower basin in spring 2016. Basins were analyzed separately due to seasonal and geographical differences. In the streams sampled in fall 2015, conductivity, total nitrogen (TN), and total phosphorus (TP) were higher in sedimentary ecoregions than in crystalline ones, independent on land use. In those sampled in spring 2016, these variables showed the highest values in the ecoregions dominated by agriculture. Eighty percent of the sampled streams presented the impact of cattle in their riparian zone. Discriminant analysis showed a similar composition of macroinvertebrates among ecoregions in 2015, but different composition between land uses. Conversely, in 2016, there were differences among some ecoregions, but not between land uses. Agriculture was correlated with tolerant invertebrates, while natural land use and afforestation were correlated with sensitive ones. The BMWP-Colombia showed the impact of livestock on streams, but in general good water quality, while an index of genera for Uruguay, indicated that all streams are eutrophic, thereby confirming the importance of using different types of metrics. Due to its geographical homogeneity and small size, a smaller number of ecoregions could be defined for stream assessments in the Río Negro basin.

Development of the Karun macroinvertebrate tolerance index (KMTI) for semi-arid mountainous streams in Iran.

The most robust approach to ecological monitoring and assessment is the use of regionally calibrated indicators. These should be calculated based on collocated biological (response) and physicochemical (stressor) variables and an objective rating and scoring system. In developing countries, a frequent lack of financial and technical resources for monitoring has led to many environmental problems being overlooked, such as the degradation of streams, rivers, and watersheds. In this paper, we propose the Karun Macroinvertebrate Tolerance Index (KMTI) for application to rivers in the Karun River basin, which is the largest watershed in Iran, draining semi-arid mountainous regions. The KMTI is the first biological index specifically developed and calibrated for Iranian water resources. Benthic macroinvertebrates, physical habitat, hydromorphic, and water quality data were collected and measured at 54 sites across four seasons in 2018 and 2019. A total of 101 families of benthic macroinvertebrates belonging to eight classes and 21 orders were identified, and tolerance values were determined for 95 families. The KMTI was found to be most efficient in identifying ecological degradation when data were used from winter samples with a discrimination efficiency (DE) 90% and a four-season mean of 84.3%. Also, the best DE of the water quality classification table based on the KMTI index was equal to 86.9%.

Refining benthic macroinvertebrate kick sampling protocol for wadeable rivers and streams in Ethiopia.

Streams and rivers cover a larger proportion of the Earth's surface but are highly affected by human pressures. Conversely, bioassessment methods are in their infancy in developing countries such as Ethiopia. In this study, we compared 2- and 3-min macroinvertebrate kick samples at multiple locations for both riffle habitat (RH) and multihabitat (MH) approaches. The performance of each method was evaluated statistically using benthic macroinvertebrate metrics and diversity indices. Results of the Kruskal-Wallis analysis in this study showed no significant differences among methods tested in minimally impacted streams in Ethiopia and generally performed equally irrespective of the methods employed except for total abundances and Ephemeroptera abundances. Furthermore, multivariate analysis of the relative abundances of macroinvertebrate communities using analysis of similarity (ANOSIM), RELATE, non-metric multidimensional scaling (MDS), and classification strength-sampling method comparability (CS-SMC) indicated a high similarity in the macroinvertebrate communities recorded among all methods employed in this study area. However, the index of multivariate dispersion (IMD) test showed variations in relative abundances of macroinvertebrate communities among the methods. In summary, if the focus is not on rare taxa and the required information is not dependent on additional evidence provided by the use of lower taxonomic levels of identification (genus and species), the results of the present study support the use of the shorter 2-min RH kick sampling method for the bioassessment of wadeable rivers and streams in Ethiopia.

Metals Alter Membership but Not Diversity of a Headwater Stream Microbiome.

Metal contamination from mining or natural weathering is a common feature of surface waters in the American west. Advances in microbial analyses have created the potential for routine sampling of aquatic microbiomes as a tool to assess the quality of stream habitat. We sought to determine if microbiome diversity and membership were affected by metal contamination and identify candidate microbial taxa to be used to indicate metal stress in stream ecosystems. We evaluated microbiome membership from sediments at multiple sites within the principal drainage of an EPA superfund site near the headwaters of the Upper Arkansas River, Leadville, CO. From each sample, we extracted DNA and sequenced the 16S rRNA gene amplicon on the Illumina MiSeq platform. We used the remaining sediments to simultaneously evaluate environmental metal concentrations. We also conducted an artificial stream mesocosm experiment using sediments collected from two of the observational study sites. The mesocosm experiment had a two-by-two factorial design: (i) location (upstream or downstream of contaminating tributary), and (ii) treatment (metal exposure or control). We found no difference in diversity between upstream and downstream sites in the field. Similarly, diversity changed very little following experimental metal exposure. However, microbiome membership differed between upstream and downstream locations and experimental metal exposure changed microbiome membership in a manner that depended on origin of the sediments used in each mesocosm.IMPORTANCE Our results suggest that microbiomes can be reliable indicators of ecosystem metal stress even when surface water chemistry and other metrics used to assess ecosystem health do not indicate ecosystem stress. Results presented in this study, in combination with previously published work on this same ecosystem, are consistent with the idea that a microbial response to metals at the base of the food web may be affecting primary consumers. If effects of metals are mediated through shifts in the microbiome, then microbial metrics, as presented here, may aid in the assessment of stream ecosystem health, which currently does not include assessments of the microbiome.

Morphological vs. DNA metabarcoding approaches for the evaluation of stream ecological status with benthic invertebrates: Testing different combinations of markers and strategies of data filtering.

Macroinvertebrate assemblages are the most common bioindicators used for stream biomonitoring, yet the standard approach exhibits several time-consuming steps, including the sorting and identification of organisms based on morphological criteria. In this study, we examined if DNA metabarcoding could be used as an efficient molecular-based alternative to the morphology-based monitoring of streams using macroinvertebrates. We compared results achieved with the standard morphological identification of organisms sampled in 18 sites located on 15 French wadeable streams to results obtained with the DNA metabarcoding identification of sorted bulk material of the same macroinvertebrate samples, using read numbers (expressed as relative frequencies) as a proxy for abundances. In particular, we evaluated how combining and filtering metabarcoding data obtained from three different markers (COI: BF1-BR2, 18S: Euka02 and 16S: Inse01) could improve the efficiency of bioassessment. In total, 140 taxa were identified based on morphological criteria, and 127 were identified based on DNA metabarcoding using the three markers, with an overlap of 99 taxa. The threshold values used for sequence filtering based on the "best identity" criterion and the number of reads had an effect on the assessment efficiency of data obtained with each marker. Compared to single marker results, combining data from different markers allowed us to improve the match between biotic index values obtained with the bulk DNA versus morphology-based approaches. Both approaches assigned the same ecological quality class to a majority (86%) of the site sampling events, highlighting both the efficiency of metabarcoding as a biomonitoring tool but also the need for further research to improve this efficiency.

Multiple co-occurrent alien invaders constrain aquatic biodiversity in rivers.

A greater understanding and effective management of biological invasions is a priority for biodiversity conservation globally. Many freshwater ecosystems are experiencing the colonization and spread of multiple co-occurrent alien species. Here the implications of both the relative abundance and richness of alien invaders on aquatic macroinvertebrate taxonomic and functional richness, ecosystem quality, and functional redundancy are assessed using long-term data from rivers in England. Based on the most common aquatic invaders, results indicated that their richness, rather than abundance, was the most important factor negatively affecting aquatic macroinvertebrate biodiversity. However, the response of functional redundancy was negatively affected by invader abundance at the river basin scale. The response of communities varied as the number of invading taxa increased, with the most marked reductions following the colonization of the first few invaders. Results indicate that different facets of multiple biological invasions influence distinct aspects of aquatic biodiversity. Preventing the establishment of new invaders and limiting invader taxa richness within a community should therefore be a conservation priority. These findings will assist river scientists in understanding mechanisms driving changes in biodiversity and facilitate the testing of ecological theories while also ensuring environmental managers and regulators can prioritize conservation / management opportunities.

Assessing mining impacts: The influence of background geochemical conditions on diatom and macroinvertebrate communities in subarctic streams.

Mining has changed landscapes locally in northern Fennoscandia and there is an increasing pressure for exploitation of the remaining mineral deposits of the region. Mineral deposits, even if unmined, can strongly influence stream water chemistry, stream biological communities and the ability of organisms to tolerate stressors. Using data sampled from six mining areas with three active (gold and chrome), two closed (gold) and one planned mine (phosphate), we examined how mineral deposits and mining influence water chemistry and diatom and macroinvertebrate communities in subarctic streams in Finnish Lapland. We supplemented the data by additional samples compiled from databases and further assessed how variation in background geological conditions influences bioassessments of the impacts arising from mining. We found that water specific conductivity was elevated in our study streams draining through catchments with a high mineral potential. Mining effects were mainly seen as increased concentration of nitrogen. Influence of mineral deposits was detected in composition of diatom and macroinvertebrate communities, but communities in streams in areas with a high mineral potential were as diverse as those in streams in areas with a low mineral potential. Mining impacts were better detected for diatoms using a reference condition based on sites with a high than low mineral potential, while for macroinvertebrates, the responses were generally less evident, likely because of only minor effects of mining on water chemistry. Community composition and frequencies of occurrence of macroinvertebrate taxa were, however, highly similar between mine-influenced streams and reference streams with a high potential for minerals indicating that the communities are strongly structured by the natural influence of mineral deposits. Incorporating geochemistry into the reference condition would likely improve bioassessments of both taxonomic groups. Replicated monitoring in potentially impacted sites and reference sites would be the most efficient framework for detecting environmental impacts in streams draining through mineral-rich catchments.

Land-use impacts on benthic macroinvertebrate assemblages in pampean streams (Argentina).

In recent decades agriculture has intensified in the Argentine Pampa, and pesticide application has also increased. Livestock fields, although being progressively replaced by crops, are still commonly interspersed with crop fields. The objective of the present work is to assess the effects of land use on the benthic invertebrate assemblages of streams in the main Argentine agricultural region. Two areas were sampled during the 2011/12 growing season (November-March): Arrecifes, a homogeneous intensively cultivated area, and La Plata, a heterogeneous area of mixed livestock pasture, cropland and biological reserve. Nutrient concentrations in water were significantly higher in the streams surrounded by cropland. Measured pesticides in stream sediments were those most commonly used in crop production: chlorpyrifos, cypermethrin, lambda-cyhalothrin, endosulfan and its degradation product endosulfan sulfate. Detection frequency and pesticide concentrations were generally higher in streams surrounded by cropland than in streams surrounded by pasture or reserve. Macroinvertebrate assemblages were significantly different in streams with different land uses. Palaemonidae (Decapoda) and Caenidae (Ephemeroptera) were the taxa best represented in the reserve. Hyalellidae (Amphipoda) and Hirudinea were dominant at the streams surrounded by livestock fields. Within the streams surrounded by croplands, Oligochaeta and Hirudinea were best represented in La Plata while Chironomidae, Gastropoda and Oligochaeta were dominant at Arrecifes. Present evidence suggests that agrochemical applications contribute, in combination with other environmental variables, to the observed differences in macroinvertebrate assemblages in streams of different land use.

Biotic indices of hydrological variability as tools to inform dynamic ecological status assessments in river ecosystems.

River biomonitoring uses biotic indices that assess human-induced degradation, including pollution, by comparison with type-specific static biological reference conditions. However, index scores that indicate pollution may reflect biological responses to natural hydrological variability associated with low flows and drying, leading biomonitoring schemes to misclassify sites as degraded. To address this, dynamic, site-specific adjustments of static biological reference conditions have been proposed, but current biomonitoring tools cannot facilitate implementation of these adjustments. We analyzed 329 samples from mediterranean-climate rivers in Greece, to evaluate the use of six stressor-specific macroinvertebrate-based indices of hydrological variability (CEFI, DEHLI, ELF, LIFE, LIFENZ, MIS-index) as tools to facilitate dynamic adjustments of static biological reference conditions. We examined macroinvertebrate assemblage responses to physicochemical and land use drivers in relation to each sample's hydrological conditions, as assessed by the six indices. We evaluated index performance beyond the region of development by exploring correlations among indices, including correlations with the region-specific Greek ELF index, for which 100% of taxa were represented. We also examined the influence of inorganic nutrient pollution on index performance by comparing index scores from samples with and without nutrient pollution. Season, water temperature, agricultural land use and nutrient pollution were major drivers of macroinvertebrate assemblage composition. Indices were positively correlated but correlation strength varied considerably, driven primarily by taxonomic representation (the proportion of sampled taxa included in each index's calculation), and potentially also by differences in river types, taxonomic resolution and sampling strategies. All indices identified site-specific hydrological conditions both in the presence and absence of nutrient pollution. We recommend the development of region-specific biotic indices of hydrological variability, or regional adaptation of existing indices, to represent 100% of the regional taxa pool and thus to enable acceptable performance beyond their region of development. Such indices could inform dynamic adaptation of static biological reference conditions by assessing site-specific hydrological conditions based on a macroinvertebrate assemblage, without the collection of additional, abiotic field data. Application of our proposed approach could prevent misclassification of ecological status, thus avoiding time-demanding and costly mismanagement of rivers and streams.

Integrating Regional Frameworks and Local Variability for Riverine Bioassessment.

Regional frameworks enable bioassessment methods to detect anthropogenic effects on ecosystems amid natural variability. Conventional approaches to regionalization have used coarse geographical frameworks to separate sites similar in their ecological (ecoregion) or faunal (basin) characteristics. Expectations for individual streams are then adjusted for within-region variability in local environmental characteristics. Integrating regional frameworks and local variability may improve the sensitivity and performance of bioassessments. In this study, we used a biologically-informed stream classification to develop an integrated regional framework for bioassessment considering the effects of ecoregion, basin, and local environmental variables on wadeable stream fish communities of South Carolina, USA. Our integrated framework was compared against conventional regional frameworks indexing ecoregions or basins alone. Frameworks were evaluated by their ability to (1) efficiently partition community variation and (2) allow for the detection of anthropogenic effects on fish communities. We found an integrated framework better described natural variability in stream fish communities. In addition, we found highly regional relationships between fish metrics and anthropogenic disturbance among frameworks, suggesting appropriate bioassessment metrics will differ across regions in our study area. Differences in community response to disturbance among frameworks emphasize the importance of testing metrics for their hypothesized sensitivity before using them in bioassessment. This study ultimately supports the integration of regional frameworks across spatial scales to classify streams for bioassessment, and provides an analytical framework from which to evaluate biotic variation and metric utility in the context of bioassessment.

Comparison of benthic macroinvertebrate assessment methods along a salinity gradient in headwater streams.

Benthic macroinvertebrate community assessments are used commonly to characterize aquatic systems and increasingly for identifying their impairment caused by myriad stressors. Yet sampling and enumeration methods vary, and research is needed to compare their abilities to detect macroinvertebrate community responses to specific water quality variables. A common assessment method, rapid bioassessment, uses subsampling procedures to identify a fixed number of individual organisms regardless of total sample abundance. In contrast, full-enumeration assessments typically allow for expanded community characterization resulting from higher numbers of identified organisms within a collected sample. Here, we compared these two sampling and enumeration methods and their abilities to detect benthic macroinvertebrate response to freshwater salinization, a common stressor of streams worldwide. We applied both methods in headwater streams along a salinity gradient within the coal-mining region of central Appalachia USA. Metrics of taxonomic richness, community composition, and trophic function differed between the methods, yet most metrics exhibiting significant response to SC for full-enumeration samples also did for rapid bioassessment samples. However, full-enumeration yielded taxonomic-based metrics consistently more responsive to the salinization gradient. Full-enumeration assessments may potentially provide more complete characterization of macroinvertebrate communities and their response to increased salinization, whereas the more cost-effective and widely employed rapid bioassessment method can detect community alterations along the full salinity gradient. These findings can inform decisions regarding such tradeoffs for assessments of freshwater salinization in headwater streams and highlight the need for similar research of sampling and enumeration methodology in other aquatic systems and for other stressors.

Spatial correlation of macroinvertebrate assemblages in streams and the implications for bioassessment programs.

Stream bioassessment using benthic macroinvertebrate assemblages is widely implemented by regulatory agencies, yet a critical question regarding spatial autocorrelation and sample independence remains: How much of a stream network does a point sample represent? Macroinvertebrate assemblages vary along a longitudinal gradient, likely due to a combination of natural and anthropogenic factors that alter water physiochemistry and habitat. A better understanding of how these gradients affect macroinvertebrate assemblage variance could prevent spatial over- and under-sampling within bioassessment efforts. This project investigated longitudinal patterns (10 s of km) of macroinvertebrate assemblages in 14 Wisconsin streams. Spatial autocorrelation was assessed using Moran's I and other multivariate methods with an emphasis on estimating the distance at which assemblages no longer display spatial correlation. Within most streams, there was a linear, direct relationship between assemblage dissimilarity and longitudinal distance, with distance to independence (DTI) ranging from 1.7-13.5 km. DTI was most strongly affected by conductivity, which is often a surrogate for a suite of anthropogenic effects. With increasing conductivity, DTI increases, suggesting more homogenous assemblages in disturbed streams. Natural factors like watershed size, channel gradient, and riparian slope also affected DTI. Considering spatial correlation in monitoring designs could improve the efficiency and application of regulatory bioassessment programs.

Sampling Efforts for Estimating Fish Species Richness in Western USA River Sites.

Fish species richness is an indicator of river ecological condition but it is particularly difficult to estimate in large unwadeable rapidly flowing rivers. Intensive multi-gear sampling is time consuming, logistically complex and expensive. However, insufficient sampling effort underestimates species richness and yields inaccurate data about the ecological condition of river sites. We raft-electrofished 10 river sites in 10 different ecoregions and six western USA states for distances equal to 300 times their mean wetted channel widths (MCWs) to estimate the effort needed to approach asymptotes in fish species richness. To collect 90% of the observed fish species at the sites, we found that an average of 150 MCWs (ranging 80-210 MCWs) were needed, with the number of MCWs increasing in rivers with a higher proportion of spatially rare species. Frequently, the second or third additional 100 MCWs produced only one or two additional singletons or doubletons (species occurring only once or twice at a site). Before initiating sampling programs for adequately estimating species richness, we recommend assessing sampling effort, particularly if rare or uncommon species are expected or desired.