Diazotrophy modulates cyanobacteria stoichiometry through functional traits that determine bloom magnitude and toxin production.

Harmful cyanobacterial blooms are an increasing threat to water quality. The interactions between two eco-physiological functional traits of cyanobacteria, diazotrophy (nitrogen (N)-fixation) and N-rich cyanotoxin synthesis, have never been examined in a stoichiometric explicit manner. We explored how a gradient of resource N:phosphorus (P) affects the biomass, N, P stoichiometry, light-harvesting pigments, and cylindrospermopsin production in a N-fixing cyanobacterium, Aphanizomenon. Low N:P Aphanizomenon cultures produced the same biomass as populations grown in high N:P cultures. The biomass accumulation determined by carbon, indicated low N:P Aphanizomenon cultures did not have a N-fixation growth tradeoff, in contrast to some other diazotrophs that maintain stoichiometric N homeostasis at the expense of growth. However, N-fixing Aphanizomenon populations produced less particulate cylindrospermopsin and had undetectable dissolved cylindrospermopsin compared to non-N-fixing populations. The pattern of low to high cyanotoxin cell quotas across an N:P gradient in the diazotrophic cylindrospermopsin producer is similar to the cyanotoxin cell quota response in non-diazotrophic cyanobacteria. We suggest that diazotrophic cyanobacteria may be characterized into two broad functional groups, the N-storage-strategists and the growth-strategists, which use N-fixation differently and may determine patterns of bloom magnitude and toxin production in nature.

Incorporating parameter variability into Monod models of nutrient-limited growth of non-diazotrophic and diazotrophic cyanobacteria.

Models are widely used tools in aquatic science to understand the mechanism of phytoplankton growth and anticipate the occurrence of harmful algal blooms. However, model parameterization remains challenging and issues that may introduce prediction uncertainty exist. Many models use the Monod equation to predict cyanobacteria growth rate based on ambient nutrient concentrations. The half-saturation concentrations in the Monod equation varies greatly among different studies and depends on environmental conditions. In this study, we estimated the growth rate due to nutrient limitations for two cyanobacteria species (Microcystis aeruginosa and Dolichospermum flos-aquae) using a modified Monod model which allows the half-saturation concentration to vary according to initial nitrogen (N) conditions. The model is calibrated against observations from laboratory experiment where cyanobacteria growth and ambient nutrient concentrations were measured simultaneously, which is rarely done in the literature. Our results show this modified model produce better predictions on growth rate and biomass, indicating many commonly used mechanistic models may need improvement regarding phytoplankton growth representation. Furthermore, our study quantifies the flexibility in cyanobacteria growth parameter across a wide range of environmental N in eutrophic lakes thus provides important information for large-scale modelling applications.

Stoichiometric imbalances complicate prediction of phytoplankton biomass in U.S. lakes: implications for nutrient criteria.

Using National Lakes Assessment data, we evaluated the influence of total N (TN), total P (TP), and other variables on lake chlorophyll-a concentrations. With simple linear regressions, high TN/TP samples biased predictions based on TN, and low TN/TP samples biased predictions based on TP. The bias problem was corrected, and correlation was improved, by splitting the dataset at the TN/TP ratio we estimated to be indicative of a balanced supply and developing separate regressions that predict chlorophyll-a based on TP, TN, dissolved inorganic N (DIN), dissolved organic carbon (DOC), non-algal light attenuation, depth, area, latitude, elevation, and conductivity. Both nutrients were excellent predictors, and non-algal light attenuation was the next most influential predictor. The regression analysis suggested that a potential for P only limitation (high TN/TP, 17% of samples) or N only limitation (low TN/TP, 14% of samples) can be inferred at the extremes of the TN/TP range. However, 69% of samples had an intermediate TN/TP ratio where it is difficult to infer anything about potential nutrient limitations (biomass could be N limited, P limited, N and P co-limited, or not limited by nutrients at all). Our results show that when developing phytoplankton response relationships using cross-lake datasets that span a wide range of trophic states, it is important to consider whether and how biomass is influenced by confounding factors - such as differences in the relative supply of N and P - so that biomass is not underestimated or overestimated, and nutrient criteria are not under-protective or over-protective.

Interspecific homeostatic regulation and growth across aquatic invertebrate detritivores: a test of ecological stoichiometry theory.

Across resource quality gradients, primary consumers must regulate homeostasis and release of nutrients to optimize growth and fitness. Based primarily on internal body composition, the ecological stoichiometry theory (EST) offers a framework to generalize interspecific patterns of these responses, yet the predictions and underlying assumptions of EST remain poorly tested across many species. We used controlled laboratory feeding experiments to measure homeostasis, nutrient release, and growth across seven field-collected aquatic invertebrate detritivore taxa fed wide resource carbon:nitrogen (C:N) and carbon:phosphorus (C:P) gradients. We found that most invertebrates exhibited strict stoichiometric homeostasis (average 1/H = - 0.018 and 0.026 for C:N and C:P, respectively), supporting assumptions of EST. However, the stoichiometry of new tissue production during growth intervals (growth stoichiometry) deviated - 30 to + 54% and - 145 to + 74% from initial body C:N and C:P, respectively, and across species, growth stoichiometry was not correlated with initial body stoichiometry. Notably, smaller non- and hemimetabolous invertebrates exhibited low, decreasing growth C:N and C:P, whereas larger holometabolous invertebrates exhibited high, often increasing growth C:N and C:P. Despite predictions of EST, interspecific sensitivity of egestion stoichiometry and growth rates to the resource gradient were weakly related to internal body composition across species. While the sensitivity of these patterns differed across taxa, such differences carried a weak phylogenetic signal and were not well predicted by EST. Our findings suggest that traits beyond internal body composition, such as feeding behavior, selective assimilation, and ontogeny, are needed to generalize interspecific patterns in consumer growth and nutrient release across resource quality gradients.

Substituting values for censored data from Texas, USA, reservoirs inflated and obscured trends in analyses commonly used for water quality target development.

We calculated four median datasets (chlorophyll a, Chl a; total phosphorus, TP; and transparency) using multiple approaches to handling censored observations, including substituting fractions of the quantification limit (QL; dataset 1 = 1QL, dataset 2 = 0.5QL) and statistical methods for censored datasets (datasets 3-4) for approximately 100 Texas, USA reservoirs. Trend analyses of differences between dataset 1 and 3 medians indicated percent difference increased linearly above thresholds in percent censored data (%Cen). This relationship was extrapolated to estimate medians for site-parameter combinations with %Cen > 80%, which were combined with dataset 3 as dataset 4. Changepoint analysis of Chl a- and transparency-TP relationships indicated threshold differences up to 50% between datasets. Recursive analysis identified secondary thresholds in dataset 4. Threshold differences show that information introduced via substitution or missing due to limitations of statistical methods biased values, underestimated error, and inflated the strength of TP thresholds identified in datasets 1-3. Analysis of covariance identified differences in linear regression models relating transparency-TP between datasets 1, 2, and the more statistically robust datasets 3-4. Study findings identify high-risk scenarios for biased analytical outcomes when using substitution. These include high probability of median overestimation when %Cen > 50-60% for a single QL, or when %Cen is as low 16% for multiple QL's. Changepoint analysis was uniquely vulnerable to substitution effects when using medians from sites with %Cen > 50%. Linear regression analysis was less sensitive to substitution and missing data effects, but differences in model parameters for transparency cannot be discounted and could be magnified by log-transformation of the variables.

Seasonal Differences in Relationships between Nitrate Concentration and Denitrification Rates in Ditch Sediments Vegetated with Rice Cutgrass.

Increased application of nitrogen (N) fertilizers in agricultural systems contributes to significant environmental impacts, including eutrophication of fresh and coastal waters. Rice cutgrass [ (L.) Sw.] can significantly enhance denitrification potential in agricultural ditch sediments and potentially reduce N export from agricultural watersheds, but relationships with known drivers are not well understood. To address this, we examined effects of nitrate (NO) availability on dinitrogen gas (N) and NO fluxes seasonally. Net denitrification rates were measured as positive N fluxes from vegetated intact sediment cores using membrane inlet mass spectrometry (MIMS). We developed Michaelis-Menten models for N fluxes across NO gradients in the spring, summer, and fall seasons. Summer N models exhibited the highest (maximum amount of net N flux) and (concentration of NO in the overlying water at which the net N flux is half of ), with a maximum production of N of ∼20 mg N m h. Maximum percentage NO retention occurred at 1 mg NO L in the overlying water in all seasons, except summer where maximum retention persisted from 1 to 5 mg NO L. Denitrification rates were strongly correlated with NO uptake by vegetated sediments in spring ( = 0.94, < 0.0001) and summer ( = 0.97, < 0.0001), but low NO uptake in fall and winter resulted in virtually no net denitrification during these seasons. Our results indicate that vegetated ditch sediments may act as effective NO sinks during the growing season. Ditch sediments vegetated with cutgrass not only immobilized a significant fraction of NO entering them but also permanently removed as much as 30 to 40% of the immobilized NO through microbial denitrification.

Comparing two periphyton collection methods commonly used for stream bioassessment and the development of numeric nutrient standards.

Periphyton is an important component of stream bioassessment, yet methods for quantifying periphyton biomass can differ substantially. A case study within the Arkansas Ozarks is presented to demonstrate the potential for linking chlorophyll-a (chl-a) and ash-free dry mass (AFDM) data sets amassed using two frequently used periphyton sampling protocols. Method A involved collecting periphyton from a known area on the top surface of variably sized rocks gathered from relatively swift-velocity riffles without discerning canopy cover. Method B involved collecting periphyton from the entire top surface of cobbles systematically gathered from riffle-run habitat where canopy cover was intentionally avoided. Chl-a and AFDM measurements were not different between methods (p = 0.123 and p = 0.550, respectively), and there was no interaction between method and time in the repeated measures structure of the study. However, significantly different seasonal distinctions were observed for chl-a and AFDM from all streams when data from the methods were combined (p < 0.001 and p = 0.012, respectively), with greater mean biomass in the cooler sampling months. Seasonal trends were likely the indirect results of varying temperatures. Although the size and range of this study were small, results suggest data sets collected using different methods may effectively be used together with some minor considerations due to potential confounding factors. This study provides motivation for the continued investigation of combining data sets derived from multiple methods of data collection, which could be useful in stream bioassessment and particularly important for the development of regional stream nutrient criteria for the southern Ozarks.

It Takes Two to Tango: When and Where Dual Nutrient (N & P) Reductions Are Needed to Protect Lakes and Downstream Ecosystems.

Preventing harmful algal blooms (HABs) is needed to protect lakes and downstream ecosystems. Traditionally, reducing phosphorus (P) inputs was the prescribed solution for lakes, based on the assumption that P universally limits HAB formation. Reduction of P inputs has decreased HABs in many lakes, but was not successful in others. Thus, the "P-only" paradigm is overgeneralized. Whole-lake experiments indicate that HABs are often stimulated more by combined P and nitrogen (N) enrichment rather than N or P alone, indicating that the dynamics of both nutrients are important for HAB control. The changing paradigm from P-only to consideration of dual nutrient control is supported by studies indicating that (1) biological N fixation cannot always meet lake ecosystem N needs, and (2) that anthropogenic N and P loading has increased dramatically in recent decades. Sediment P accumulation supports long-term internal loading, while N may escape via denitrification, leading to perpetual N deficits. Hence, controlling both N and P inputs will help control HABs in some lakes and also reduce N export to downstream N-sensitive ecosystems. Managers should consider whether balanced control of N and P will most effectively reduce HABs along the freshwater-marine continuum.

Dietary and taxonomic controls on incorporation of microbial carbon and phosphorus by detritivorous caddisflies.

Heterotrophic microbes on detritus play critical roles in the nutrition of detritivorous animals, yet few studies have examined factors controlling the acquisition of microbial nutrients toward detritivore growth, which is termed "incorporation". Here, we assessed effects of detrital substrate identity (leaf type), background nutrients, and detritivore species identity on detritivore incorporation of microbial carbon (C) and phosphorus (P) in leaf litter diets. We fed oak and maple litter conditioned under two nutrient concentrations (50 or 500 µg P L(-1)) to the detritivorous caddisfly larvae Ironoquia spp., Lepidostoma spp., and Pycnopsyche lepida and used the radioisotopes 14C as glucose and 33P as phosphate to dually trace incorporation of microbial C and P by caddisflies. Incorporation efficiencies of microbial C (mean ± SE = 12.3 ± 1.3%) were one order of magnitude higher than gross growth efficiencies for bulk detrital C from recent studies (1.05 ± 0.08%). Litter type did not affect incorporation of microbial nutrients; however, caddisflies incorporated microbial P 11 % less efficiently when fed litter from the higher P concentration. Two lower body C:P species (Pycnopsyche and Ironoquia) exhibited 9.9 and 7.1% greater microbial C and 19.0 and 17.7% greater microbial P incorporation efficiencies, respectively, than the higher body C:P species (Lepidostoma). Our findings support ecological stoichiometry theory on post-ingestive regulation that animals fed lower C:P diets should reduce P incorporation efficiency due to excess diet P or alleviation of P-limited growth, and that lower C:P species must incorporate dietary C and P more efficiently to support fast growth of P-rich tissues.

Implementing Effects-Based Water Quality Criteria for Eutrophication in Beaver Lake, Arkansas: Linking Standard Development and Assessment Methodology.

To address water quality standards needed to prevent accelerated eutrophication, many states in the United States have developed effects-based standards related to nutrients. In many cases, this has resulted in specific standards for Secchi transparency (ST) and phytoplankton biomass measured as sestonic chlorophyll (chl-). The state of Arkansas recently adopted its first effects-based water quality criteria for Beaver Lake in northwestern Arkansas, which was a growing-season geometric mean chl- <8 µg L and an annual average ST >1.1 m. However, the adopted standard did not have a predefined assessment methodology that outlined the frequency and duration of potential exceedances. This study used hydrologic frequency analysis to estimate the risk of exceeding these water quality standards using measured and modeled data from Beaver Lake from 2001 to 2014. Beaver Lake conformed to common models in reservoir limnology in that ST was least and chl- was greatest in the river-reservoir transition zone and decreased in the downstream direction toward the dam. Greater chl- and lesser ST was clearly related to total phosphorus concentrations along this gradient. Thus, the risk of exceeding the water quality criteria decreased in a downstream direction. There were substantial differences in the probability of exceeding the adopted water quality criteria based on both spatial and temporal variation in the potential assessment periods. Based on the way the standard was developed and the risk of exceeding these standards derived from data collected before the standards were in place, we recommend that a minimum of half of the years assessed be necessary to result in a water quality violation. A number of other assessment considerations are presented that could provide flexibility to regulatory agencies in assessing water quality standards.

Contrasting Nutrient Mitigation and Denitrification Potential of Agricultural Drainage Environments with Different Emergent Aquatic Macrophytes.

Remediation of excess nitrogen (N) in agricultural runoff can be enhanced by establishing wetland vegetation, but the role of denitrification in N removal is not well understood in drainage ditches. We quantified differences in N retention during experimental runoff events followed by stagnant periods in mesocosms planted in three different vegetation treatments: unvegetated, cutgrass [ (L.) Sw.], and common cattail ( L.). We also quantified denitrification rates using membrane inlet mass spectrometry from intact cores extracted from each mesocosm treatment. All treatments retained 60% or more of NO-N loads during the 6-h experimental runoff event, but mesocosms planted with cutgrass had significantly higher (68%) retention than the cattail (60%) or unvegetated (61%) treatments. After the runoff event, mesocosms planted in cattail reduced NO-N concentrations by >95% within 24 h and cutgrass achieved similar reductions within 48 h, whereas reductions in the unvegetated mesocosms were significantly less (65%). Cores from cutgrass mesocosms had significantly higher average denitrification rates (5.93 mg m h), accounting for as much as 56% of the immobilized NO-N within 48 h, whereas denitrification rates were minimal in cores from the unvegetated (-0.19 mg m h) and cattail (0.2 mg m h) mesocosms. Our findings have implications for mitigating excess NO-N in agricultural runoff. While vegetated treatments removed excess NO-N from the water column at similar and significantly higher rates than unvegetated treatments, the high denitrification rates observed for cutgrass highlight the potential for permanent removal of excess N from agricultural runoff in vegetated ditches and wetlands.

Proteome changes in larval zebrafish (Danio rerio) and fathead minnow (Pimephales promelas) exposed to (±) anatoxin-a.

Anatoxin-a and its analogues are potent neurotoxins produced by several genera of cyanobacteria. Due in part to its high toxicity and potential presence in drinking water, these toxins pose threats to public health, companion animals and the environment. It primarily exerts toxicity as a cholinergic agonist, with high affinity at neuromuscular junctions, but molecular mechanisms by which it elicits toxicological responses are not fully understood. To advance understanding of this cyanobacteria, proteomic characterization (DIA shotgun proteomics) of two common fish models (zebrafish and fathead minnow) was performed following  (±) anatoxin-a exposure. Specifically, proteome changes were identified and quantified in larval fish exposed for 96 h (0.01-3 mg/L (±) anatoxin-a and caffeine (a methodological positive control) with environmentally relevant treatment levels examined based on environmental exposure distributions of surface water data. Proteomic concentration - response relationships revealed 48 and 29 proteins with concentration - response relationships curves for zebrafish and fathead minnow, respectively. In contrast, the highest number of differentially expressed proteins (DEPs) varied between zebrafish (n = 145) and fathead minnow (n = 300), with only fatheads displaying DEPs at all treatment levels. For both species, genes associated with reproduction were significantly downregulated, with pathways analysis that broadly clustered genes into groups associated with DNA repair mechanisms. Importantly, significant differences in proteome response between the species was also observed, consistent with prior observations of differences in response using both behavioral assays and gene expression, adding further support to model specific differences in organismal sensitivity and/or response. When DEPs were read across from humans to zebrafish, disease ontology enrichment identified diseases associated with cognition and muscle weakness consistent with the prior literature. Our observations highlight limited knowledge of how (±) anatoxin-a, a commonly used synthetic racemate surrogate, elicits responses at a molecular level and advances its toxicological understanding.

Sestonic Chlorophyll- Shows Hierarchical Structure and Thresholds with Nutrients across the Red River Basin, USA.

The Red River is a transboundary, multijurisdictional basin where water-quality standards are often different across state lines. The state agencies with USEPA Region VI focused resources to organize water-quality data from within this basin and have it statistically analyzed to evaluate the relationships between nutrients and sestonic chlorophyll- (chl-a). There were 152 sites within the Red River basin that had nutrient and sestonic chl-a measurements; these sites were narrowed down to 132 when a minimum number of observations was required. Sestonic chl-a levels increased with increasing nutrient concentrations; these regressions were used to predict nutrient concentrations at 10 µg chl-a L. Total nitrogen (TN) and phosphorus (TP) concentrations (at 10 µg chl-a L) varied across the Red River basin and its ecoregions from 0.10 to 0.22 mg TP L and 0.75 to 2.11 mg TN L. Nutrient thresholds were also observed with sestonic chl-a at 0.14 mg TP L and 0.74 mg TN L using categorical and regression tree analysis (CART). The CART analysis also revealed that hierarchical structure was important when attempting to predict sestonic chl-a from TN, TP, and conductivity. The ranges of TN and TP concentrations that resulted in chl-a concentrations that exceeded 10 µg chl-a L were similar in magnitude to the threshold in TN and TP that resulted in increased sestonic chl-a. This corroborating evidence provides useful guidance to the states with jurisdiction within the Red River basin for establishing nutrient criteria, which may be similar when the Red River and its tributaries cross political boundaries.

Phosphorus mitigation to control river eutrophication: murky waters, inconvenient truths, and "postnormal" science.

This commentary examines an "inconvenient truth" that phosphorus (P)-based nutrient mitigation, long regarded as the key tool in eutrophication management, in many cases has not yet yielded the desired reductions in water quality and nuisance algal growth in rivers and their associated downstream ecosystems. We examine why the water quality and aquatic ecology have not recovered, in some case after two decades or more of reduced P inputs, including (i) legacies of past land-use management, (ii) decoupling of algal growth responses to river P loading in eutrophically impaired rivers; and (iii) recovery trajectories, which may be nonlinear and characterized by thresholds and alternative stable states. It is possible that baselines have shifted and that some disturbed river environments may never return to predisturbance conditions or may require P reductions below those that originally triggered ecological degradation. We discuss the practical implications of setting P-based nutrient criteria to protect and improve river water quality and ecology, drawing on a case study from the Red River Basin in the United States. We conclude that the challenges facing nutrient management and eutrophication control bear the hallmarks of "postnormal" science, where uncertainties are large, management intervention is urgently required, and decision stakes are high. We argue a case for a more holistic approach to eutrophication management that includes more sophisticated regime-based nutrient criteria and considers other nutrient and pollutant controls and river restoration (e.g., physical habitat and functional food web interactions) to promote more resilient water quality and ecosystem functioning along the land-freshwater continuum.

The effects of salinity and N:P on N-rich toxins by both an N-fixing and non-N-fixing cyanobacteria.

Freshwater ecosystems are experiencing increased salinization. Adaptive management of harmful algal blooms (HABs) contribute to eutrophication/salinization interactions through the hydrologic transport of blooms to coastal environments. We examined how nutrients and salinity interact to affect growth, elemental composition, and cyanotoxin production/release in two common HAB genera. Microcystis aeruginosa (non-nitrogen (N)-fixer and microcystin-LR producer; MC-LR) and Aphanizomenon flos-aquae (N-fixer and cylindrospermopsin producer; CYN) were grown in N:phosphorus (N:P) 4 and 50 (by atom) for 21 and 33 days, respectively, then dosed with a salinity gradient (0 - 10.5 g L-1). Both total MC-LR and CYN were correlated with particulate N. We found Microcystis MC-LR production and release was affected by salinity only in the N:P 50 treatment. However, Aphanizomenon CYN production and release was affected by salinity regardless of N availability. Our results highlight how cyanotoxin production and release across the freshwater - marine continuum are controlled by eco-physiological differences between N-acquisition traits.

Within-river phosphorus retention: accounting for a missing piece in the watershed phosphorus puzzle.

The prevailing "puzzle" in watershed phosphorus (P) management is how to account for the nonconservative behavior (retention and remobilization) of P along the land-freshwater continuum. This often hinders our attempts to directly link watershed P sources with their water quality impacts. Here, we examine aspects of within-river retention of wastewater effluent P and its remobilization under high flows. Most source apportionment methods attribute P loads mobilized under high flows (including retained and remobilized effluent P) as nonpoint agricultural sources. We present a new simple empirical method which uses chloride as a conservative tracer of wastewater effluent, to quantify within-river retention of effluent P, and its contribution to river P loads, when remobilized under high flows. We demonstrate that within-river P retention can effectively mask the presence of effluent P inputs in the water quality record. Moreover, we highlight that by not accounting for the contributions of retained and remobilized effluent P to river storm-flow P loads, existing source apportionment methods may significantly overestimate the nonpoint agricultural sources and underestimate wastewater sources in mixed land-use watersheds. This has important implications for developing effective watershed remediation strategies, where remediation needs to be equitably and accurately apportioned among point and nonpoint P contributors.

Nitrogen fixation: a poorly understood process along the freshwater-marine continuum.

Although N2 fixation is a major component of the global N cycle and has been extensively studied in open-ocean and terrestrial ecosystems, rates and ecological dynamics remain virtually unknown for the inland and coastal aquatic ecosystems (lakes, wetlands, rivers, streams, estuaries) that connect terrestrial and marine biomes. This is due to the diversity of these habitats, as well as the traditional paradigm that N2 fixation rates were low to nonexistent, and therefore not important, in these ecosystems. We identify three major research themes to advance understanding of aquatic N2 fixation: 1) the biological diversity of diazotrophs and variability of N2 fixation rates, 2) the ecological stoichiometry of N2 fixation, and 3) the upscaling of N2 fixation rates from genes to ecosystems. Coordinating research across these areas will advance limnology and oceanography by fully integrating N2 fixation into ecological dynamics of aquatic ecosystems from local to global scales.

Dynamic Phycobilin Pigment Variations in Diazotrophic and Non-diazotrophic Cyanobacteria Batch Cultures Under Different Initial Nitrogen Concentrations.

Increased anthropogenic nutrient loading has led to eutrophication of aquatic ecosystems, which is the major cause of harmful cyanobacteria blooms. Element stoichiometry of cyanobacteria bloom is subject to nutrient availabilities and may significantly contribute to primary production and biogeochemical cycling. Phycobilisome is the antenna of the photosynthetic pigment apparatus in cyanobacteria, which contains phycobilin pigments (PBPs) and linker proteins. This nitrogen (N)-rich protein complex has the potential to support growth as a N-storage site and may play a major role in the variability of cyanobacteria N stoichiometry. However, the regulation of PBPs during bloom formation remains unclear. We investigated the temporal variation of N allocation into PBPs and element stoichiometry for two ubiquitous cyanobacteria species, Microcystis aeruginosa and Dolichospermum flos-aquae, in a batch culture experiment with different initial N availabilities. Our results indicated that the N allocation into PBPs is species-dependent and tightly regulated by the availability of nutrients fueling population expansion. During the batch culture experiment, different nutrient uptake rates led to distinct stoichiometric imbalances of N and phosphorus (P), which substantially altered cyanobacteria C: N and C: P stoichiometry. Microcystis invested cellular N into PBPs and exhibited greater flexibility in C: N and C: P stoichiometry than D. flos-aquae. The dynamics of such N-rich macromolecules may help explain the N stoichiometry variation during a bloom and the interspecific difference between M. aeruginosa and D. flos-aquae. Our study provides a quantitative understanding of the elemental stoichiometry and the regulation of PBPs for non-diazotrophic and diazotrophic cyanobacteria blooms.

Comment on "Models predict planned phosphorus load reduction will make Lake Erie more toxic".

Hellweger et al. (Reports, 27 May 2022, pp. 1001) predict that phosphorus limitation will increase concentrations of cyanobacterial toxins in lakes. However, several molecular, physiological, and ecological mechanisms assumed in their models are poorly supported or contradicted by other studies. We conclude that their take-home message that phosphorus load reduction will make Lake Erie more toxic is seriously flawed.

Change point analysis of phosphorus trends in the Illinois River (Oklahoma) demonstrates the effects of watershed management.

Detecting water quality improvements following watershed management changes is complicated by flow-dependent concentrations and nonlinear or threshold responses that are difficult to detect with traditional statistical techniques. In this study, we evaluated the long-term trends (1997-2009) in total P (TP) concentrations in the Illinois River of Oklahoma, and some of its major tributaries, using flow-adjusted TP concentrations and regression tree analysis to identify specific calendar dates in which change points in P trends may have occurred. Phosphorus concentrations at all locations were strongly correlated with stream flow. Flow-adjusted TP concentrations increased at all study locations in the late 1990s, but this trend was related to a change in monitoring practices where storm flow samples were specifically targeted after 1998. Flow-adjusted TP concentrations decreased in the two Illinois River sites after 2003. This change coincided with a significant decrease in effluent TP concentrations originating with one of the largest municipal wastewater treatment facilities in the basin. Conversely, flow-adjusted TP concentrations in one tributary increased, but this stream received treated effluent from a wastewater facility where effluent TP did not decrease significantly over the study period. Results of this study demonstrate how long-term trends in stream TP concentrations are difficult to quantify without consistent long-term monitoring strategies and how flow adjustment is likely mandatory for examining these trends. Furthermore, the study demonstrates how detecting changes in long-term water quality data sets requires statistical methods capable of identifying change point and nonlinear responses.