A web-based tool for assessing the condition of benthic diatom assemblages in streams and rivers of the conterminous United States.

Benthic diatom assemblages are known to be indicative of water quality but have yet to be widely adopted in biological assessments in the United States due to several limitations. Our goal was to address some of these limitations by developing regional multi-metric indices (MMIs) that are robust to inter-laboratory taxonomic inconsistency, adjusted for natural covariates, and sensitive to a wide range of anthropogenic stressors. We aggregated bioassessment data from two national-scale federal programs and used a data-driven analysis in which all-possible combinations of 2-7 metrics were compared for three measures of performance. After ranking the best-performing MMIs, we selected the final MMIs by evaluating stress-response relations in independent regional datasets of diatom samples paired with measures of several water-quality stressors, including herbicides and streamflow flashiness. Each regional MMI performed well at calibration sites and represented diverse aspects of the structure and function of diatom communities. Most metrics included in the best MMIs were modeled to account for natural variation including climate, topography, soil characteristics, lithology, and groundwater influence on streamflow. MMI performance improved with higher numbers of component metrics, but this effect diminished beyond six metrics. Component metrics of MMIs were associated with a broad suite of measured stressors in every region, including salinity, nutrients, herbicides, and streamflow flashiness. We provide a web-based software application that allows users in the conterminous United States to apply our MMIs to their own datasets and compare MMI scores from their sites to a broader regional context.

Physical habitat in conterminous US streams and rivers, Part 1: Geoclimatic controls and anthropogenic alteration.

Anthropogenic alteration of physical habitat structure in streams and rivers is increasingly recognized as a major cause of impairment worldwide. As part of their assessment of the status and trends in the condition of rivers and streams in the U.S., the U.S. Environmental Protection Agency's (USEPA) National Aquatic Resource Surveys (NARS) quantify and monitor channel size and slope, substrate size and stability, instream habitat complexity and cover, riparian vegetation cover and structure, anthropogenic disturbance activities, and channel-riparian interaction. Like biological assemblages and water chemistry, physical habitat is strongly controlled by natural geoclimatic factors that can obscure or amplify the influence of human activities. We developed a systematic approach to estimate the deviation of observed river and stream physical habitat from that expected in least-disturbed reference conditions. We applied this approach to calculate indices of anthropogenic alteration of three aspects of physical habitat condition in the conterminous U.S. (CONUS): streambed sediment size and stability, riparian vegetation cover, and instream habitat complexity. The precision and responsiveness of these indices led the USEPA to use them to evaluate physical habitat condition in CONUS rivers and streams. The scores of these indices systematically decreased with greater anthropogenic disturbance at river and stream sites in the CONUS and within ecoregions, which we interpret as a response of these physical habitat indices to anthropogenic influences. Although anthropogenic activities negatively influenced all three physical habitat indices in the least-disturbed sites within most ecoregions, natural geoclimatic and geomorphic factors were the dominant influences. For sites over the full range of anthropogenic disturbance, analyses of observed/expected sediment characteristics showed augmented flood flows and basin and riparian agriculture to be the leading predictors of streambed instability and excess fine sediments. Similarly, basin and riparian agriculture and non-agricultural riparian land uses were the leading predictors of reduced riparian vegetation cover complexity in the CONUS and within ecoregions. In turn, these reductions in riparian vegetation cover and complexity, combined with reduced summer low flows, were the leading predictors of instream habitat simplification. We conclude that quantitative measures of physical habitat structure are useful and important indicators of the impacts of human activities on stream and river condition.

Physical habitat in conterminous US streams and Rivers, part 2: A quantitative assessment of habitat condition.

Rigorous assessments of the ecological condition of water resources and the effect of human activities on those waters require quantitative physical, chemical, and biological data. The U.S. Environmental Protection Agency's river and stream surveys quantify river and stream bed particle size and stability, instream habitat complexity and cover, riparian vegetation cover and structure, and anthropogenic disturbance activities. Physical habitat is strongly controlled by natural geoclimatic factors that co-vary with human activities. We expressed the anthropogenic alteration of physical habitat as O/E ratios of observed habitat metric values divided by values expected under least-disturbed reference conditions, where site-specific expected values vary given their geoclimatic and geomorphic context. We set criteria for good, fair, and poor condition based on the distribution of O/E values in regional least-disturbed reference sites. Poor conditions existed in 22-24% of the 1.2 million km of streams and rivers in the conterminous U.S. for riparian human disturbance, streambed sediment and riparian vegetation cover, versus 14% for instream habitat complexity. Based on the same four indicators, the percentage of stream length in poor condition within 9 separate U.S. ecoregions ranged from 4% to 42%. Associations of our physical habitat indices with anthropogenic pressures demonstrate the scope of anthropogenic habitat alteration; habitat condition was negatively related to the level of anthropogenic disturbance nationally and in nearly all ecoregions. Relative risk estimates showed that streams and rivers with poor sediment, riparian cover complexity, or instream habitat cover conditions were 1.4 to 2.6 times as likely to also have fish or macroinvertebrate assemblages in poor condition. Our physical habitat condition indicators help explain deviations in biological conditions from those observed among least-disturbed sites and inform management actions for rehabilitating impaired waters and mitigating further ecological degradation.

The Relation of Lotic Fish and Benthic Macroinvertebrate Condition Indices to Environmental Factors Across the Conterminous USA.

National and regional ecological assessments are essential for making rational decisions concerning water body conservation and management at those spatial extents. We analyzed data from 4597 samples collected from 3420 different sites across the conterminous USA during the U.S. Environmental Protection Agency's 2008-2009 and 2013-2014 National Rivers and Streams Assessment. We evaluated the relationship between both fish and macroinvertebrate multimetric index (MMI) condition scores and 38 environmental factors to assess the relative importance of natural versus anthropogenic predictors, contrast site-scale versus watershed-scale predictors, and examine ecoregional and assemblage differences. We found that most of the environmental factors we examined were related to either fish and/or macroinvertebrate MMI scores in some fashion and that the factors involved, and strength of the relationship, varied by ecoregion and between assemblages. Factors more associated with natural conditions were usually less important in explaining MMI scores than factors more directly associated with anthropogenic disturbances. Local site-scale factors explained more variation than watershed-scale factors. Random forest and multiple regression models performed similarly, and the fish MMI-environment relationships were stronger than macroinvertebrate MMI-environment relationships. Among ecoregions, the strongest environmental relationships were observed in the Northern Appalachians and the weakest in the Southern Plains. The fish and macroinvertebrate MMIs were only weakly correlated with each other, and they generally responded more strongly to different groups of variables. These results support the use of multiple assemblages and the sampling of multiple environmental indicators in ecological assessments across large spatial extents.

Taxonomic harmonization may reveal a stronger association between diatom assemblages and total phosphorus in large datasets.

Diatom data have been collected in large-scale biological assessments in the United States, such as the U.S. Environmental Protection Agency's National Rivers and Streams Assessment (NRSA). However, the effectiveness of diatoms as indicators may suffer if inconsistent taxon identifications across different analysts obscure the relationships between assemblage composition and environmental variables. To reduce these inconsistencies, we harmonized the 2008-2009 NRSA data from nine analysts by updating names to current synonyms and by statistically identifying taxa with high analyst signal (taxa with more variation in relative abundance explained by the analyst factor, relative to environmental variables). We then screened a subset of samples with QA/QC data and combined taxa with mismatching identifications by the primary and secondary analysts. When these combined "slash groups" did not reduce analyst signal, we elevated taxa to the genus level or omitted taxa in difficult species complexes. We examined the variation explained by analyst in the original and revised datasets. Further, we examined how revising the datasets to reduce analyst signal can reduce inconsistency, thereby uncovering the variation in assemblage composition explained by total phosphorus (TP), an environmental variable of high priority for water managers. To produce a revised dataset with the greatest taxonomic consistency, we ultimately made 124 slash groups, omitted 7 taxa in the small naviculoid (e.g., Sellaphora atomoides) species complex, and elevated Nitzschia, Diploneis, and Tryblionella taxa to the genus level. Relative to the original dataset, the revised dataset had more overlap among samples grouped by analyst in ordination space, less variation explained by the analyst factor, and more than double the variation in assemblage composition explained by TP. Elevating all taxa to the genus level did not eliminate analyst signal completely, and analyst remained the most important predictor for the genera Sellaphora, Mayamaea, and Psammodictyon, indicating that these taxa present the greatest obstacle to consistent identification in this dataset. Although our process did not completely remove analyst signal, this work provides a method to minimize analyst signal and improve detection of diatom association with TP in large datasets involving multiple analysts. Examination of variation in assemblage data explained by analyst and taxonomic harmonization may be necessary steps for improving data quality and the utility of diatoms as indicators of environmental variables.

Assessing water-quality changes in US rivers at multiple geographic scales using results from probabilistic and targeted monitoring.

Two commonly used approaches for water quality monitoring are probabilistic and targeted. In a probabilistic approach like the US Environmental Protection Agency's National Rivers and Streams Assessment, monitoring sites are selected using a statistically representative approach. In a targeted approach like that used by many monitoring organizations, monitoring sites are chosen individually to answer specific questions. One important goal of both approaches is documenting long-term changes in water quality. Here, we compare chloride change results in US rivers and streams between the early 2000s and early 2010s from both approaches. The probabilistic approach provided an unbiased representation of change in all US rivers and streams, but was designed to measure low-streamflow conditions within a spring/summer index period during periodic survey years. The targeted approach was focused on larger, more developed watersheds but samples were collected frequently throughout the assessment period in different seasons and streamflows. The probabilistic results showed a small decrease in chloride concentrations in rivers and streams with the lowest concentrations, but no consistent increase or decrease in the remainder. The increased granularity of the targeted results showed that there was, in fact, a mix of changes occurring, with increases at 132 sites, decreases at 112 sites, and relatively stable conditions at 55 sites. The combined results suggest that chloride is not responding to a widespread, common driver across the USA and that management of chloride would be most effective when targeted regionally or locally.

A synoptic survey of microbial respiration, organic matter decomposition, and carbon efflux in U.S. streams and rivers.

We analyzed ecoenzyme activities related to organic matter processing in 1879 streams and rivers across the continental U.S. as part of the USEPA's National Rivers and Streams Assessment. Ecoenzymatic stoichiometry was used to construct models for carbon use efficiency (CUE) and decomposition (-k). Microbial respiration (Rm) was estimated from sediment organic carbon stocks, CUE and -k. The streams and rivers were classified by size (headwaters: 1st-order; streams: 2nd-3rd order; small rivers: 4th-5th order; big rivers 6th-7th order; and great rivers ≥ 8th order) and condition class (least, intermediate and most disturbed), and grouped into nine ecoregions. There were ecoregion, stream size, and condition class effects for CUE, -k, and Rm, with Rm increasing from eastern ecoregions through the plains to the western ecoregions. CUE, -k, and Rm decreased with increasing streams size and increased with increasing disturbance. Rm, CUE, and -k were correlated with water and sediment chemistry; CUE and -k were also correlated with stream bed fine sediments; and CUE was further correlated with catchment land cover. Rm was extrapolated to ecoregional and national scales, and the results suggest that microbial assemblages account for 12% of the total CO2 outgassing, and nearly 50% of the aquatic metabolism C losses, from U.S. streams and rivers. Cumulative respiratory C losses increased from headwaters to small streams, then decreased with increasing stream size. This U-shaped respiration curve was not evident when streams were viewed by disturbance classes, suggesting that anthropogenic disturbances mask the expected organic matter processing signature of the river continuum.

Using DNA metabarcoding to characterize national scale diatom-environment relationships and to develop indicators in streams and rivers of the United States.

Recent advancements in DNA techniques, metabarcoding, and bioinformatics could help expand the use of benthic diatoms in monitoring and assessment programs by providing relatively quick and increasingly cost-effective ways to quantify diatom diversity in environmental samples. However, such applications of DNA-based approaches are relatively new, and in the United States, unknowns regarding their applications at large scales exist because only a few small-scale studies have been done. Here, we present results from the first nationwide survey to use DNA metabarcoding (rbcL) of benthic diatoms, which were collected from 1788 streams and rivers across nine ecoregions spanning the conterminous USA. At the national scale, we found that diatom assemblage structure (1) was strongly associated with total phosphorus and total nitrogen concentrations, conductivity, and pH and (2) had clear patterns that corresponded with differences in these variables among the nine ecoregions. These four variables were strong predictors of diatom assemblage structure in ecoregion-specific analyses, but our results also showed that diatom-environment relationships, the importance of environmental variables, and the ranges of these variables within which assemblage changes occurred differed among ecoregions. To further examine how assemblage data could be used for biomonitoring purposes, we used indicator species analysis to identify ecoregion-specific taxa that decreased or increased along each environmental gradient, and we used their relative abundances of gene reads in samples as metrics. These metrics were strongly correlated with their corresponding variable of interest (e.g., low phosphorus diatoms with total phosphorus concentrations), and generalized additive models showed how their relationships compared among ecoregions. These large-scale national patterns and nine sets of ecoregional results demonstrated that diatom DNA metabarcoding is a robust approach that could be useful to monitoring and assessment programs spanning the variety of conditions that exist throughout the conterminous United States.

A one health approach for monitoring antimicrobial resistance: developing a national freshwater pilot effort.

Antimicrobial resistance (AMR) is a world-wide public health threat that is projected to lead to 10 million annual deaths globally by 2050. The AMR public health issue has led to the development of action plans to combat AMR, including improved antimicrobial stewardship, development of new antimicrobials, and advanced monitoring. The National Antimicrobial Resistance Monitoring System (NARMS) led by the United States (U.S) Food and Drug Administration along with the U.S. Centers for Disease Control and U.S. Department of Agriculture has monitored antimicrobial resistant bacteria in retail meats, humans, and food animals since the mid 1990's. NARMS is currently exploring an integrated One Health monitoring model recognizing that human, animal, plant, and environmental systems are linked to public health. Since 2020, the U.S. Environmental Protection Agency has led an interagency NARMS environmental working group (EWG) to implement a surface water AMR monitoring program (SWAM) at watershed and national scales. The NARMS EWG divided the development of the environmental monitoring effort into five areas: (i) defining objectives and questions, (ii) designing study/sampling design, (iii) selecting AMR indicators, (iv) establishing analytical methods, and (v) developing data management/analytics/metadata plans. For each of these areas, the consensus among the scientific community and literature was reviewed and carefully considered prior to the development of this environmental monitoring program. The data produced from the SWAM effort will help develop robust surface water monitoring programs with the goal of assessing public health risks associated with AMR pathogens in surface water (e.g., recreational water exposures), provide a comprehensive picture of how resistant strains are related spatially and temporally within a watershed, and help assess how anthropogenic drivers and intervention strategies impact the transmission of AMR within human, animal, and environmental systems.

Understanding the effects of phosphorus on diatom richness in rivers and streams using taxon-environment relationships.

Changes in phosphorus concentrations affect periphytic diatom composition in streams, yet we rarely observe strong relationships between diatom richness and phosphorus. In contrast, changes in conductivity are strongly associated with differences in both diatom composition and richness. We hypothesised that we could better understand the mechanisms that control the phosphorus-richness relationship by examining relationships between phosphorus and the occurrence of individual diatom taxa, comparing these with relationships between conductivity and taxon occurrence, and documenting how niche breadths of taxa affect richness patterns. We estimated relationships between phosphorus and taxon occurrence using DNA metabarcoding data of diatoms collected from 1,811 sites distributed across the conterminous U.S.A. and contrasted patterns in these relationships with those between conductivity and taxon occurrence. The distribution of taxon optima for phosphorus was bimodal, with most optima located at either the maximum or minimum observed phosphorus concentration. The distribution of taxon optima for conductivity was unimodal. Niche breadths of taxa for phosphorus and for conductivity both generally increased with optimum values. The distribution of conductivity optima gave rise to a prominent hump-shaped relationship between richness and conductivity. The relationship between richness and phosphorus was also slightly hump-shaped, but this relationship would not be expected from the bimodal distribution of optima. Instead, we determined that broad niche breadths caused the hump-shaped relationship between richness and phosphorus. Our results highlight the nuanced effects that increased P loadings exert on diatom assemblages in rivers and streams and identify reasons that weak relationships between taxon richness and increased phosphorus have been observed. These findings allow us to better describe how excess phosphorus and subsets of taxa and their niche breadths contribute to patterns of taxa richness in diatom assemblages, and to improve the tools used to manage phosphorus pollution.

Patterns in and predictors of stream and river macroinvertebrate genera and fish species richness across the conterminous USA.

Both native and non-native taxa richness patterns are useful for evaluating areas of greatest conservation concern. To determine those patterns, we analyzed fish and macroinvertebrate taxa richness data obtained at 3475 sites collected by the USEPA's National Rivers and Streams Assessment. We also determined which natural and anthropogenic variables best explained patterns in regional richness. Macroinvertebrate and fish richness increased with the number of sites sampled per region. Therefore, we determined residual taxa richness from the deviation of observed richness from predicted richness given the number of sites per region. Regional richness markedly exceeded average site richness for both macroinvertebrates and fish. Predictors of macroinvertebrate-genus and fish-species residual-regional richness differed. Air temperature was an important predictor in both cases but was positive for fish and negative for macroinvertebrates. Both natural and land use variables were significant predictors of regional richness. This study is the first to determine mean site and regional richness of both fish and aquatic macroinvertebrates across the conterminous USA, and the key anthropogenic drivers of regional richness. Thus, it offers important insights into regional USA biodiversity hotspots.

Biogeography and co-occurrence of 16 planktonic species of Keratella Bory de St. Vincent, 1822 (Rotifera, Ploima, Brachionidae) in lakes and reservoirs of the United States.

We report current distributions of 16 species of Keratella Bory de St. Vincent, 1822 within lakes and reservoirs of the U.S. Specimens were identified from 988 lakes and reservoirs during spring and summer 2012 as part of the U.S. Environmental Protection Agency's National Lakes Assessment (NLA). We performed a co-occurrence analysis to determine correlations between species-pair occurrences and a niche-centroid analysis to predict optimal water quality conditions for each species. While a high degree of overlap in geographic range was observed among the species, distribution maps showed that K. quadrata was largely confined to more northern latitudes and K. americana was confined primarily to the eastern U.S. Other common species were either ubiquitously or sparsely distributed across ecoregions, suggesting that their distribution may be more related to inter-species competition or local water quality parameters. This study expands the limited pool of knowledge on rotifer biogeography within the U.S.

Environmental factors influencing the quantitative distribution of microcystin and common potentially toxigenic cyanobacteria in U.S. lakes and reservoirs.

Many species of cyanobacteria are capable of producing toxins and causing nuisance blooms, however response to environmental conditions is likely taxon-specific. Environmental factors influencing cyanobacterial composition and toxin production in lakes have been examined in many studies; yet are often confined to individual water bodies, or to a small number of systems within the same region. Here, data from the 2012 USEPA National Lakes Assessment are used to examine relationships between biovolume of common potentially-toxigenic cyanobacteria (Aphanizomenon spp., Cylindrospermopsis spp., Dolichospermum spp., Microcystis spp. and Planktothrix spp.) and environmental variables across the entire conterminous United States, and results are compared across nine distinct ecoregions. Total phosphorus and water clarity were identified as the most influential environmental factors correlated with phytoplankton community composition. The Northern, Southern and Temperate Plains ecoregions displayed the highest biovolumes of potentially toxigenic taxa on average, as well as highest mean concentrations of microcystin. In those three ecoregions, samples with microcystin concentrations greater than 1 ppb were primarily dominated by Planktothrix spp. while in all other ecoregions Dolichospermum spp. was the dominant genus. Canonical Correlation Analysis revealed a strong association between high microcystin concentrations and high nutrient concentrations (total nitrogen and total phosphorus), and between high microcystin concentrations and low percentage of watershed forest cover. Results from this study indicate that the likely occurrence of potentially toxigenic taxa in lakes and reservoirs is predictable on a biogeographical basis, depending on morphological and water quality characteristics. Data from this study may be useful to regional managers attempting to prevent or mitigate nuisance cyanobacterial blooms.

CD4+ and CD8+ T cell receptor repertoire perturbations with normal levels of T cell receptor excision circles in HIV-infected, therapy-naive adolescents.

Our objective was to determine whether treatment-naive HIV-infected adolescents manifest abnormalities in thymus function and peripheral T cell repertoire, and to assess relationships of these immunologic characteristics with each other, with plasma HIV virus load, and T cell surface markers. TCR Vbeta repertoire was determined by CDR3 length spectratyping in purified CD4(+) and CD8(+) T cells of high-risk, HIV-negative adolescents and of treatment-naive, HIV-infected adolescents. Thymus function was investigated by the simultaneous examination of T cell receptor excision circles (TRECs) in the CD4(+) and CD8(+) T cell subsets. HIV-infected adolescents exhibited significantly greater perturbations in their TCR Vbeta repertoire in comparison with HIV-negative subjects. Perturbations in the CD8(+) T cell compartment were more profound in comparison with CD4(+) T cells. The CD4(+) TCR Vbeta perturbations were negatively correlated with the total and phenotypically naive CD4(+) T cells, and with CD4(+) TRECs. CD8(+) TRECs, although not correlated with CD8(+) TCR Vbeta perturbations, showed negative correlation with memory and activated CD8(+) T cells. Interestingly, TRECs in CD4(+) and CD8(+) T cells were not significantly different between HIV-infected and uninfected adolescents. The TCR Vbeta repertoire in adolescents is profoundly perturbed even in early stages of HIV infection, when total CD4(+) cell counts in most subjects are within normal limits. The correlative analyses demonstrating negative association of CD4(+) cell TRECs with CD4(+) TCR Vbeta perturbations and of CD8(+) TRECs with CD8(+) cell activation markers provide evidence of the intense activation of the central and peripheral immune compartments in this study population.

Assessment of thymic activity in human immunodeficiency virus-negative and -positive adolescents by real-time PCR quantitation of T-cell receptor rearrangement excision circles.

Circular DNA molecules known as T-cell receptor rearrangement excision circles (TREC) arise during T-cell development and are present in cells that have recently emigrated from the thymus. In cross-sectional studies, the number of peripheral blood lymphocytes bearing TREC decreases with age, consistent with an anatomically demonstrated loss of thymic epithelial tissue. TREC numbers increase following hematopoietic stem cell transplantation and during therapy for human immunodeficiency virus (HIV) infection. Quantitation of TREC has therefore been proposed as a parameter of thymic activity. In this study, we used real-time PCR to quantify TREC in peripheral blood samples obtained longitudinally from HIV-seronegative adolescents. TREC values in peripheral blood T cells were very stable throughout adolescence, once thought to be a time of rapid involution of the thymus. In addition, in a cross-sectional analysis, we examined TREC values in a cohort of HIV-positive adolescents and found evidence of ongoing thymopoiesis in perinatally infected individuals, despite lifelong infection. These data demonstrate the utility of TREC assessment in adolescents and that HIV infection does not uniformly result in accelerated thymic involution in childhood.