Influence of sampling effort on metrics of fish-based indices for the assessment of estuarine ecological quality

Multimetric fish-based indices have been increasingly gaining importance in Europe, as the Water Framework Directive (WFD) requires fish fauna, and particularly its composition and abundance, to be taken into account in the assessment of the ecological quality of continental surface waters, including transitional waters. These indices are composed of several metrics, mostly related with structural and functional characteristics of fish communities, such as species richness, the role of nursery areas, or trophic web structure. Therefore, ecological quality assessments should ensure that these structural and functional characteristics of fish communities were covered by the sampling methods used. In the present work, the influence of sampling effort on several metrics of the Estuarine Fish Assessment Index (EFAI) was studied. Pseudo-random samples were generated from data of four Portuguese estuaries and bootstrap cycles were performed, in order to obtain metrics’ means and standard deviations per number of hauls analysed. The number of hauls necessary for the means to level off differed with the metrics considered. Generally, for metrics on percentages (percentage of marine migrants, percentage of estuarine residents and percentage of piscivores) the curve levelled off with less than 20 hauls, both for the estuary as a whole and for different estuarine salinity zones. On the other hand, metrics on species richness required much larger samples. In order to decrease to −5% the current estimated bias of metrics, the WFD sampling costs would have to be more than 3 times higher than they currently are. The findings in the present study are of great importance for an effective assessment of estuarine ecological quality and particularly in the context of the WFD, as the metrics studied are common to other Member State indices.     

Estimating species richness using environmental DNA

The foundation for any ecological study and for the effective management of biodiversity in natural systems requires knowing what species are present in an ecosystem. We assessed fish communities in a stream using two methods, depletion‐based electrofishing and environmental DNA metabarcoding (eDNA) from water samples, to test the hypothesis that eDNA provides an alternative means of determining species richness and species identities for a natural ecosystem. In a northern Indiana stream, electrofishing yielded a direct estimate of 12 species and a mean estimated richness (Chao II estimator) of 16.6 species with a 95% confidence interval from 12.8 to 42.2. eDNA sampling detected an additional four species, congruent with the mean Chao II estimate from electrofishing. This increased detection rate for fish species between methods suggests that eDNA sampling can enhance estimation of fish fauna in flowing waters while having minimal sampling impacts on fish and their habitat. Modern genetic approaches therefore have the potential to transform our ability to build a more complete list of species for ecological investigations and inform management of aquatic ecosystems.     

Improved Methods for Capture,Extraction, and Quantitative Assay of Environmental DNA from Asian Bigheaded Carp (Hypophthalmichthys spp.)

Indirect, non-invasive detection of rare aquatic macrofauna using aqueous environmental DNA (eDNA) is a relatively new approach to population and biodiversity monitoring. As such, the sensitivity of monitoring results to different methods of eDNA capture, extraction, and detection is being investigated in many ecosystems and species. One of the first and largest conservation programs with eDNA-based monitoring as a central instrument focuses on Asian bigheaded carp (Hypophthalmichthys spp.), an invasive fish spreading toward the Laurentian Great Lakes. However, the standard eDNA methods of this program have not advanced since their development in 2010. We developed new, quantitative, and more cost-effective methods and tested them against the standard protocols. In laboratory testing, our new quantitative PCR (qPCR) assay for bigheaded carp eDNA was one to two orders of magnitude more sensitive than the existing endpoint PCR assays. When applied to eDNA samples from an experimental pond containing bigheaded carp, the qPCR assay produced a detection probability of 94.8% compared to 4.2% for the endpoint PCR assays. Also, the eDNA capture and extraction method we adapted from aquatic microbiology yielded five times more bigheaded carp eDNA from the experimental pond than the standard method, at a per sample cost over forty times lower. Our new, more sensitive assay provides a quantitative tool for eDNA-based monitoring of bigheaded carp, and the higher-yielding eDNA capture and extraction method we describe can be used for eDNA-based monitoring of any aquatic species.     

Seasonal dynamics of riverine fish communities using eDNA

As fish communities are a major concern in rivers ecosystems, we investigated if their environmental (e)DNA signals vary according to the sampling period or hydromorphological conditions. Three rivers were studied over a year using eDNA metabarcoding approach. The majority of the species (c. 80%) were detected all year round in two rivers having similar hydromorphological conditions, whereas in the river affected by an upstream lake waterflow, more species were detected sporadically (42%). For all the rivers, in more than 98% of the occasional detections, the reads abundance represented <0.4% of the total reads per site and per sampling session. Even if the majority of the fish communities remained similar over the year for each of the three rivers, specific seasonal patterns were observed. We studied if the waterflow or the reproduction period had an effect on the observed dynamics. Waterflow, which influences eDNA downstream transportation, had a global influence in taxonomic richness, while the fishes' reproductive period had only an influence on certain species. Our results may help selecting the best sampling strategy according to research objectives. To study fish communities at local scale, seasons of low waterflow periods are recommended. This particularly helps to restraint effects of external eDNA coming from connections with other aquatic environment (tributaries, lakes, wetlands, sewage effluents, etc.). To obtain a more integrative overview of the fish community living in a river basin, high waterflow or breeding seasons are preferable for enhancing species detection probability, especially for rare species.     

Environmental DNA phylogeography: Successful reconstruction of phylogeographic patterns of multiple fish species from cups of water

Phylogeography is an integrative field of science linking micro- and macro-evolutionary processes, contributing to the inference of vicariance, dispersal, speciation, and other population-level processes. Phylogeographic surveys usually require considerable effort and time to obtain numerous samples from many geographical sites covering the distribution range of target species; this associated high cost limits their application. Recently, environmental DNA (eDNA) analysis has been useful not only for detecting species but also for assessing genetic diversity; hence, there has been growing interest in its application to phylogeography. As the first step of eDNA-based phylogeography, we examined (1) data screening procedures suitable for phylogeography and (2) whether the results obtained from eDNA analysis accurately reflect known phylogeographic patterns. For these purposes, we performed quantitative eDNA metabarcoding using group-specific primer sets in five freshwater fish species belonging to two taxonomic groups from a total of 94 water samples collected from western Japan. As a result, three-step data screening based on the DNA copy number of each haplotype detected successfully eliminated suspected false positive haplotypes. Furthermore, eDNA analysis could almost perfectly reconstruct the phylogenetic and phylogeographic patterns obtained for all target species with the conventional method. Despite existing limitations and future challenges, eDNA-based phylogeography can significantly reduce survey time and effort and is applicable for simultaneous analysis of multiple species in single water samples. eDNA-based phylogeography has the potential to revolutionize phylogeography.     

The role of phytoplankton diversity metrics in shallow lake and river quality assessment

Ecological water quality problems are frequently connected to increment of phytoplankton productivity and overdominance of some phytoplankton species. Metrics that show monotonously increasing or decreasing tendencies along stressor gradients is recommended for ecological state assessment. Diversity metrics are influenced by various physical disturbances and show high within-year variability; thus, there is no agreement on the usefulness of these metrics as state indicators.To test the usefulness of phytoplankton diversity in ecological state assessment we investigated the productivity–diversity relationships for lakes and rivers in the Carpathian Basin (Hungary). We demonstrated that the shape of productivity–diversity relationship depends on the investigated water body type. Regarding lakes, hump-shaped relationship was found for all computed metrics. Parallel with the increase in phytoplankton productivity values, diversity metrics showed monotonously increasing tendencies in rhithral and decreasing tendencies in large potamal rivers. We found no systematic relationship in the case of small lowland rivers.Changes of diversity metrics calculated for species and functional groups showed similar tendencies within the types, only the slopes of regression lines differ each other.The use of diversity metrics as ecological state indicators should be restricted to water body types where diversity decreases or increases monotonously with phytoplankton biomass. Regarding the lakes the use of diversity metrics is not recommended for ecological state assessment. In rhithral and large potamal river assessment, application of diversity metrics should be strongly considered. We demonstrated that diversity metrics can be useful components of multimetric indices proposed to use by the Water Framework Directive.     

Assessing fish metrics and biotic indices in a Mediterranean stream: effects of uncertain native status of fish

Implementation of the Water Framework Directive requires tools for measuring and monitoring the ecological status of aquatic ecosystems. Several indices are in use in the Iberian Peninsula, although there has been little comparison among them. We sampled the fish assemblage and limnological features of the Tordera stream (NE Spain) quarterly from September 2001 to May 2003 to evaluate the usefulness of several fish metrics and to compare habitat quality and biotic indices currently in use. Data for eight biotic and abiotic indices for this and three other Catalan river basins were also compiled in order to analyse the relationships among indices. In the Tordera stream, fish abundance and richness increased with stream order except in the last sampling site that had the lowest fish abundance owing to the effects of drought and water abstraction. Although most indices were positively correlated, some displayed low or null correlations particularly for the Tordera basin which is more affected by water abstraction and less by pollution; a commonly used physico-chemical index (ISQA) was the least correlated. In a regional fish index (IBICAT) under development, the brown trout (Salmo trutta) has been previously considered as introduced in the Tordera basin. Here, we report an old published record that demonstrates that trout was present before 1845 and we argue that its status should be considered as uncertain given the current information available. Whether brown trout is treated as native or introduced to this river basin has profound effects on the results of fish metrics because of its dominance in the upper reaches. We briefly discuss the role of introduced species, particularly in headwater streams, in the development of fish indices. Our study exemplifies the need for careful, basin-specific assessment of native/introduced status in the development of fish metrics. Handling editor: K. Martens     

Environmental DNA metabarcoding provides enhanced detection of the European eel Anguilla anguilla and fish community structure in pumped river catchments

The European eel Anguilla anguilla (eel hereafter) is critically endangered and has a catadromous life cycle, which means adult eels that live in pumped catchments must pass through pumps during their downstream spawning migration. Policy makers are currently lacking detailed site-by-site eel distribution information to estimate the overall impact of individual pumping stations on eel escapement, and as such lack the data to enable informed prioritisation of pumping station management and targeted mitigation. This study investigated whether environmental DNA (eDNA) metabarcoding can provide increased detection sensitivity for eel and fish community structure in highly regulated pumped catchments, when compared directly to current standard practice fish survey protocols (seine netting/electric fishing). Eels were detected in 14 of 17 sites (82.4%) using eDNA metabarcoding in contrast to 3 of 17 sites (17.6%) using traditional catch methods. In addition, when using eDNA monitoring, species richness was higher in 16 of 17 sites (94.1%), and site occupancy was greater than or equal to traditional methods for 23 of 26 of the fish species detected (88.5%). Although eDNA methods presented significantly higher average species richness and species site occupancy overall, eDNA and catch methods were positively correlated in terms of species richness and site occupancy. It was therefore found that eDNA metabarcoding was a high-sensitivity method for detecting eels in pumped catchments while also increasing the detection of overall fish community structure compared to traditional catch methods. In addition, this study highlights how eDNA monitoring is especially suited to increase the detection of particular species, with traditional methods sufficient for others. This high sensitivity, coupled with the ability to sample multiple sites in a short time frame, suggests that eDNA metabarcoding workflows could be invaluable tools when prioritising pumping station management.     

Performance of eDNA assays to detect and quantify an elusive benthic fish in upland streams

The sensitivity and specificity of eDNA-based monitoring, coupled with its potential utility to estimate population density or biomass, makes it a useful tool in invasive species management. In this study, we investigated the potential of the eDNA method to improve the detection of the elusive invasive fish, oriental weatherloach (Misgurnus anguillicaudatus), in a river system where a density gradient of the species occurs. We compared detection rates between eDNA and conventional monitoring methods and examined the relationship between eDNA and abundance in a flowing environment. The eDNA method had a higher site detection rate than conventional methods (63 vs. 38%). Weatherloach eDNA was detected at all sites where the fish has been previously caught and none of the sites where the species has not been caught for the past 7 years. There was an increasing density trend going downstream based on long-term conventional monitoring, but the eDNA concentration in water samples reflected this trend only in a continuous section of the river where impoundments were absent. We did not find a positive relationship between eDNA concentration and contemporary abundance estimates in our study area. A high eDNA concentration was recorded at a site (DVC) which was designated a low density site based on long-term catch data. This discrepancy was a likely result of physical habitat characteristics which influenced the efficiency of the conventional methods used. This study highlighted the challenges of inferring density from eDNA data in flowing water because habitat features may confound results, necessitating careful consideration for results to be useful to management.     

Marine water environmental DNA metabarcoding provides a comprehensive fish diversity assessment and reveals spatial patterns in a large oceanic area

Current methods for monitoring marine fish (including bony fishes and elasmobranchs) diversity mostly rely on trawling surveys, which are invasive, costly, and time‐consuming. Moreover, these methods are selective, targeting a subset of species at the time, and can be inaccessible to certain areas. Here, we used environmental DNA (eDNA), the DNA present in the water column as part of shed cells, tissues, or mucus, to provide comprehensive information about fish diversity in a large marine area. Further, eDNA results were compared to the fish diversity obtained in pelagic trawls. A total of 44 5 L‐water samples were collected onboard a wide‐scale oceanographic survey covering about 120,000 square kilometers in Northeast Atlantic Ocean. A short region of the 12S rRNA gene was amplified and sequenced through metabarcoding generating almost 3.5 million quality‐filtered reads. Trawl and eDNA samples resulted in the same most abundant species (European anchovy, European pilchard, Atlantic mackerel, and blue whiting), but eDNA metabarcoding resulted in more detected bony fish and elasmobranch species (116) than trawling (16). Although an overall correlation between fishes biomass and number of reads was observed, some species deviated from the common trend, which could be explained by inherent biases of each of the methods. Species distribution patterns inferred from eDNA metabarcoding data coincided with current ecological knowledge of the species, suggesting that eDNA has the potential to draw sound ecological conclusions that can contribute to fish surveillance programs. Our results support eDNA metabarcoding for broad‐scale marine fish diversity monitoring in the context of Directives such as the Common Fisheries Policy or the Marine Strategy Framework Directive.     

A conceptual guide to measuring species diversity

Three metrics of species diversity – species richness, the Shannon index and the Simpson index – are still widely used in ecology, despite decades of valid critiques leveled against them. Developing a robust diversity metric has been challenging because, unlike many variables ecologists measure, the diversity of a community often cannot be estimated in an unbiased way based on a random sample from that community. Over the past decade, ecologists have begun to incorporate two important tools for estimating diversity: coverage and Hill diversity. Coverage is a method for equalizing samples that is, on theoretical grounds, preferable to other commonly used methods such as equal-effort sampling, or rarefying datasets to equal sample size. Hill diversity comprises a spectrum of diversity metrics and is based on three key insights. First, species richness and variants of the Shannon and Simpson indices are all special cases of one general equation. Second, richness, Shannon and Simpson can be expressed on the same scale and in units of species. Third, there is no way to eliminate the effect of relative abundance from estimates of any of these diversity metrics, including species richness. Rather, a researcher must choose the relative sensitivity of the metric towards rare and common species, a concept which we describe as ‘leverage.' In this paper we explain coverage and Hill diversity, provide guidelines for how to use them together to measure species diversity, and demonstrate their use with examples from our own data. We show why researchers will obtain more robust results when they estimate the Hill diversity of equal-coverage samples, rather than using other methods such as equal-effort sampling or traditional sample rarefaction.     

基于环境DNA宏条码和底拖网的珠江河口鱼类多样性

文章采用环境DNA宏条码和底拖网对珠江河口鱼类多样性进行了研究, 并对两种方法进行了比较。利用环境DNA宏条码检测到了175种鱼类, 而利用底拖网采集到了47种鱼类, 结合两种方法共检测出179种鱼类, 隶属于15 目63科128属。其中两种方法共同识别了鱼类43种, 占总检测物种的24.02%, 基于底拖网的调查未能收集到基于环境DNA宏条码检测到的大多数物种。根据Shannon指数和Simpson指数显示, DNA宏条码所检测珠江河口鱼类群落α多样性显著高于底拖网方法(P<0.05)。两种方法的PCoA结果均显示珠江河口鱼类群落存在空间结构, 基于环境DNA宏条码的分析显示空间重叠更多。两种方法基于冗余分析均显示溶解氧和盐度是影响鱼类群落结构的主要环境因子。研究表明, 环境DNA 宏条形码是一种环保且可靠的评估方法, 将其搭载到现有调查可以更好地了解河口鱼类多样性。     

Repurposing environmental DNA samples—detecting the western pearlshell (Margaritifera falcata) as a proof of concept

Information on the distribution of multiple species in a common landscape is fundamental to effective conservation and management. However, distribution data are expensive to obtain and often limited to high‐profile species in a system. A recently developed technique, environmental DNA (eDNA) sampling, has been shown to be more sensitive than traditional detection methods for many aquatic species. A second and perhaps underappreciated benefit of eDNA sampling is that a sample originally collected to determine the presence of one species can be re‐analyzed to detect additional taxa without additional field effort. We developed an eDNA assay for the western pearlshell mussel (Margaritifera falcata) and evaluated its effectiveness by analyzing previously collected eDNA samples that were annotated with information including sample location and deposited in a central repository. The eDNA samples were initially collected to determine habitat occupancy by nonbenthic fish species at sites that were in the vicinity of locations recently occupied by western pearlshell. These repurposed eDNA samples produced results congruent with historical western pearlshell surveys and permitted a more precise delineation of the extent of local populations. That a sampling protocol designed to detect fish was also successful for detecting a freshwater mussel suggests that rapidly accumulating collections of eDNA samples can be repurposed to enhance the efficiency and cost‐effectiveness of aquatic biodiversity monitoring.     

Assessing the efficiency of connectivity measures with regard to the EU-Water Framework Directive in a Danube-tributary system

The effect of restoring connectivity for fish by the construction of 11 fish ladders in the Pielach and Melk rivers, both tributaries to the Danube in Austria, was monitored using electric fishing and fish traps between 1999 and 2004. In order to assess the efficiency of connectivity rehabilitation measures pre- and post-project data combining electric fishing and trap catch data were analyzed by means of three fish-based assessment methods: a MUlti-Level concept for a Fish-based, river-type-specific Assessment of ecological integrity (MULFA), the Fish Index Austria (FIA) and the European Fish Index (EFI). The effect of adding qualitative trap catch data to electric fishing data on metrics and indices was also tested and the magnitude of the effect was related to the distance of the sites from the river mouth. The results clearly demonstrated the significant contribution of connectivity rehabilitation measures to the ecological integrity of rivers like the River Pielach where morphological conditions are good, whereas remaining channelization still limits the success of connectivity measures in the River Melk. Trap catch data were found to represent an essential source of additional information to assess the efficiency of connectivity measures shortly after their implementation. The negative correlations of the magnitude of the effect of different indices and metrics with the distance of assessment sites from river mouths obviously underline the importance of the river Danube as a source for the re-colonization process. While the indices tested were found to have limited ability to reflect short-term response of fish assemblages to continuum rehabilitation, guild metrics were able to detect improvements of the ecological status shortly after the implementation of connectivity measures. Six metrics showed significant differences between pre and post-project data reflecting the expected increase of the ecological integrity: (1) Fish Region Index (FRI; FIA, MULFA), (2) number of subdominant species and (3) number of flow-guilds (FIA), (4) number of type specific species (MULFA), (5) number of benthic species and (6) number of potamodromous species (EFI); the FRI differences were only significant when trap catch data were added. The EFI indicated a decline of ecological integrity through increases in the density of omnivorous species and the relative number of tolerant species as well as a decrease in the relative number of intolerant species. Significantly decreasing responses with the distance from the river mouth were documented by the EFI and MULFA-index, the FRI (FIA, MULFA), total biomass and for the number of type specific species (MULFA). Guest editors: R. L. Welcomme & G. Marmulla Hydropower, Flood Control and Water Abstraction: Implications for Fish and Fisheries     

Assessment of fish communities using environmental DNA: Effect of spatial sampling design in lentic systems of different sizes

Freshwater fish biodiversity is quickly decreasing and requires effective monitoring and conservation. Environmental DNA (eDNA)‐based methods have been shown to be highly sensitive and cost‐efficient for aquatic biodiversity surveys, but few studies have systematically investigated how spatial sampling design affects eDNA‐detected fish communities across lentic systems of different sizes. We compared the spatial patterns of fish diversity determined using eDNA in three lakes of small (SL; 3 ha), medium (ML; 122 ha) and large (LL; 4,343 ha) size using a spatially explicit grid sampling method. A total of 100 water samples (including nine, 17 and 18 shoreline samples and six, 14 and 36 interior samples from SL, ML and LL, respectively) were collected, and fish communities were analysed using eDNA metabarcoding of the mitochondrial 12S region. Together, 30, 35 and 41 fish taxa were detected in samples from SL, ML, and LL, respectively. We observed that eDNA from shoreline samples effectively captured the majority of the fish diversity of entire waterbodies, and pooled samples recovered fewer species than individually processed samples. Significant spatial autocorrelations between fish communities within 250 m and 2 km of each other were detected in ML and LL, respectively. Additionally, the relative sequence abundances of many fish species exhibited spatial distribution patterns that correlated with their typical habitat occupation. Overall, our results support the validity of a shoreline sampling strategy for eDNA‐based fish community surveys in lentic systems but also suggest that a spatially comprehensive sampling design can reveal finer distribution patterns of individual species.     

基于eDNA metabarcoding探究北京市主要河流鱼类分布及影响因素

使用eDNA宏条形码(eDNA metabarcoding)和地笼法检测了北京市3条水系在夏季和秋季两个季节的鱼类多样性, 旨在研究北京市鱼类群落的空间格局特征, 探索适用于北京鱼类生物多样性监测及保护的新方法。结果表明: 在北京市的34个采样点中, 利用eDNA metabarcoding共检测出鱼类55种, 显著高于传统方法所捕获的鱼类种数(35种), 鱼类组成以鲤形目和鲈形目为主。山区河流清水鱼的多样性要显著高于城区河流, 城区河流(北运河水系)群落结构较为单一, 以鲫(Carassius auratus)、麦穗鱼(Pseudorasbora parva)、泥鳅(Misgurnus anguillicaudatus)等耐污种为优势种; 山区河流(潮白河水系及大清河水系)以宽鳍鱲(Zacco platypus)、拉氏鱥(Rhynchocypris lagowskii)、马口鱼(Opsariichthys uncirostris)等为优势种。不同季节影响清水鱼群落结构的环境因子不同, 夏季主要是总溶解固体和电导率, 秋季主要是海拔和温度。清水鱼丰富度与环境因子及人类活动的相关性表明, 清水鱼的丰富度随着总溶解固体及灯光指数增加而显著降低, 且均与海拔、温度等存在显著相关性。本研究证明了eDNA metabarcoding方法用于监测北京市鱼类多样性及其时空分布的可行性。     

Reliable eDNA detection and quantification of the European weather loach (Misgurnus fossilis)

The European weather loach (Misgurnus fossilis) is a cryptic and poorly known fish species of high conservation concern. The species is experiencing dramatic population collapses across its native range to the point of regional extinction. Although environmental DNA (eDNA)-based approaches offer clear advantages over conventional field methods for monitoring rare and endangered species, accurate detection and quantification remain difficult and quality assessment is often poorly incorporated. In this study, we developed and validated a novel digital droplet PCR (ddPCR) eDNA-based method for reliable detection and quantification, which allows accurate monitoring of M. fossilis across a number of habitat types. A dilution experiment under laboratory conditions allowed the definition of the limit of detection (LOD) and the limit of quantification (LOQ), which were set at concentrations of 0.07 and 0.14 copies μl^–1, respectively. A series of aquarium experiments revealed a significant and positive relationship between the number of individuals and the eDNA concentration measured. During a 3 year survey (2017–2019), we assessed 96 locations for the presence of M. fossilis in Flanders (Belgium). eDNA analyses on these samples highlighted 45% positive detections of the species. On the basis of the eDNA concentration per litre of water, only 12 sites appeared to harbour relatively dense populations. The other 31 sites gave a relatively weak positive signal that was typically situated below the LOQ. Combining sample-specific estimates of effective DNA quantity (Q_e) and conventional field sampling, we concluded that each of these weak positive sites still likely harboured the species and therefore they do not represent false positives. Further, only seven of the classified negative samples warrant additional sampling as our analyses identified a substantial risk of false-negative detections (i.e., type II errors) at these locations. Finally, we illustrated that ddPCR outcompetes conventional qPCR analyses, especially when target DNA concentrations are critically low, which could be attributed to a reduced sensitivity of ddPCR to inhibition effects, higher sample concentrations being accommodated and higher sensitivity obtained.     

A comparison of algal,macroinvertebrate, and fish assemblage indices for assessing low-level nutrient enrichment in wadeable Ozark streams

Biotic indices for algae, macroinvertebrates, and fish assemblages can be effective for monitoring stream enrichment, but little is known regarding the value of the three assemblages for detecting perturbance as a consequence of low-level nutrient enrichment. In the summer of 2006, we collected nutrient and biotic samples from 30 wadeable Ozark streams that spanned a nutrient-concentration gradient from reference to moderately enriched conditions. Seventy-three algal metrics, 62 macroinvertebrate metrics, and 60 fish metrics were evaluated for each of the three biotic indices. After a group of candidate metrics had been identified with multivariate analysis, correlation procedures and scatter plots were used to identify the four metrics having strongest relations to a nutrient index calculated from log transformed and normalized total nitrogen and total phosphorus concentrations. The four metrics selected for each of the three biotic indices were: algae—the relative abundance of most tolerant diatoms, the combined relative abundance of three species of Cymbella, mesosaprobic algae percent taxa richness, and the relative abundance of diatoms that are obligate nitrogen heterotrophs; macroinvertebrate—the relative abundance of intolerant organisms, Baetidae relative abundance, moderately tolerant taxa richness, and insect biomass; fish—herbivore and detritivore taxa richness, pool species relative abundance, fish catch per unit effort, and black bass (Micropterus spp.) relative abundance.All three biotic indices were negatively correlated to nutrient concentrations but the algal index had a higher correlation (rho = ?0.89) than did the macroinvertebrate and fish indices (rho = ?0.63 and ?0.58, respectively). Biotic index scores were lowest and nutrient concentrations were highest for streams with basins having the highest poultry and cattle production. Because of the availability of litter for fertilizer and associated increases in grass and hay production, cattle feeding capacity increases with poultry production. Studies are needed that address the synergistic effect of poultry and cattle production on Ozark streams in high production areas before ecological risks can be adequately addressed.     

Macroinvertebrate indicators of lake acidification: analysis of monitoring data from UK, Norway and Sweden

Although the acid sensitivity of many invertebrate species in lakes is well known, methods for assessment of lake acidification based on macroinvertebrate samples are less developed than for rivers. This article analyses a number of existing metrics developed for assessment of river acidification, and evaluates their performance for assessment of lake acidification. Moreover, new species-based indicators of lake acidification were developed and tested. The selected dataset contains 668 samples on littoral macroinvertebrates from 427 lakes with almost 60% of the samples from Sweden and the rest from UK and Norway. Flexible, non-parametric regression models were used for explorative analyses of the pressure–response relationships. The metrics have been assessed according to their response to pH, the degree of non-linearity of the response and the influence of humic compounds. Acid-sensitive metrics often showed a threshold in response to pH between 5.8 and 6.5. Highly acid-tolerant metrics were typically dominant across the whole pH range. Humic level had a positive effect for most acid-sensitive metrics. Generally, most metrics showed a more non-linear response pattern for the humic lakes than for clear lakes. The significant relationship between these macroinvertebrate metrics and acidification shows that there is a potential for developing further the assessment systems for ecological quality of lakes based on these metrics, although the metrics explained a low % of the variation (<30%). In order to improve the predictive power of the biotic metrics across the acidified part of Europe, further harmonization and standardisation of sampling effort and taxa identification are needed.