Lakes and reservoirs globally produce large quantities of methane and carbon dioxide in their sediments, which accumulate in the hypolimnia (bottom waters) during thermally stratified conditions. A key parameter controlling hypolimnetic greenhouse gas concentrations is dissolved oxygen. Land use and climate change have increased hypolimnetic anoxia worldwide in lakes and reservoirs, which is expected to affect their methane and carbon dioxide concentrations. We conducted whole-ecosystem oxygenation experiments to assess the effects of oxygen concentrations on dissolved hypolimnetic greenhouse gas concentrations in comparison to a reference reservoir and calculated the maximum hypolimnetic global warming potential in both reservoirs over three summers. We observed significantly greater hypolimnetic methane under anoxic conditions but similar carbon dioxide concentrations, leading to greater hypolimnetic global warming potential of anoxic hypolimnia. Our study indicates that the global warming potential of hypolimnetic greenhouse gas concentrations may increase as the prevalence of hypolimnetic anoxia increases due to global change.

Near-term, ecological forecasting with iterative model refitting and uncertainty partitioning has great promise for improving our understanding of ecological processes and the predictive skill of ecological models, but to date has been infrequently applied to predict biogeochemical fluxes. Bubble fluxes of methane (CH4) from aquatic sediments to the atmosphere (ebullition) dominate freshwater greenhouse gas emissions, but it remains unknown how best to make robust near-term CH4 ebullition predictions using models. Near-term forecasting workflows have the potential to address several current challenges in predicting CH4 ebullition rates, including: development of models that can be applied across time horizons and ecosystems, identification of the timescales for which predictions can provide useful information, and quantification of uncertainty in predictions. To assess the capacity of near-term, iterative forecasting workflows to improve ebullition rate predictions, we developed and tested a near-term, iterative forecasting workflow of CH4 ebullition rates in a small eutrophic reservoir throughout one open-water period. The workflow included the repeated updating of a CH4 ebullition forecast model over time with newly-collected data via iterative model refitting. We compared the CH4 forecasts from our workflow to both alternative forecasts generated without iterative model refitting and a persistence null model. Our forecasts with iterative model refitting estimated CH4 ebullition rates up to 2 weeks into the future [RMSE at 1-week ahead = 0.53 and 0.48 log(e)(mg CH4 m(-2) d(-1)) at 2-week ahead horizons]. Forecasts with iterative model refitting outperformed forecasts without refitting and the persistence null model at both 1- and 2-week forecast horizons. Driver uncertainty and model process uncertainty contributed the most to total forecast uncertainty, suggesting that future workflow improvements should focus on improved mechanistic understanding of CH4 models and drivers. Altogether, our study suggests that iterative forecasting improves week-to-week CH4 ebullition predictions, provides insight into predictability of ebullition rates into the future, and identifies which sources of uncertainty are the most important contributors to the total uncertainty in CH4 ebullition predictions.

Near-term iterative forecasting is a powerful tool for ecological decision support and has the potential to transform our understanding of ecological predictability. However, to this point, there has been no cross-ecosystem analysis of near-term ecological forecasts, making it difficult to synthesize diverse research efforts and prioritize future developments for this emerging field. In this study, we analyzed 178 near-term (<= 10-yr forecast horizon) ecological forecasting papers to understand the development and current state of near-term ecological forecasting literature and to compare forecast accuracy across scales and variables. Our results indicated that near-term ecological forecasting is widespread and growing: forecasts have been produced for sites on all seven continents and the rate of forecast publication is increasing over time. As forecast production has accelerated, some best practices have been proposed and application of these best practices is increasing. In particular, data publication, forecast archiving, and workflow automation have all increased significantly over time. However, adoption of proposed best practices remains low overall: for example, despite the fact that uncertainty is often cited as an essential component of an ecological forecast, only 45% of papers included uncertainty in their forecast outputs. As the use of these proposed best practices increases, near-term ecological forecasting has the potential to make significant contributions to our understanding of forecastability across scales and variables. In this study, we found that forecastability (defined here as realized forecast accuracy) decreased in predictable patterns over 1-7 d forecast horizons. Variables that were closely related (i.e., chlorophyll and phytoplankton) displayed very similar trends in forecastability, while more distantly related variables (i.e., pollen and evapotranspiration) exhibited significantly different patterns. Increasing use of proposed best practices in ecological forecasting will allow us to examine the forecastability of additional variables and timescales in the future, providing a robust analysis of the fundamental predictability of ecological variables.

Oxygen availability is decreasing in many lakes and reservoirs worldwide, raising the urgency for understanding how anoxia (low oxygen) affects coupled biogeochemical cycling, which has major implications for water quality, food webs, and ecosystem functioning. Although the increasing magnitude and prevalence of anoxia has been documented in freshwaters globally, the challenges of disentangling oxygen and temperature responses have hindered assessment of the effects of anoxia on carbon, nitrogen, and phosphorus concentrations, stoichiometry (chemical ratios), and retention in freshwaters. The consequences of anoxia are likely severe and may be irreversible, necessitating ecosystem-scale experimental investigation of decreasing freshwater oxygen availability. To address this gap, we devised and conducted REDOX (the Reservoir Ecosystem Dynamic Oxygenation eXperiment), an unprecedented, 7-year experiment in which we manipulated and modeled bottom-water (hypolimnetic) oxygen availability at the whole-ecosystem scale in a eutrophic reservoir. Seven years of data reveal that anoxia significantly increased hypolimnetic carbon, nitrogen, and phosphorus concentrations and altered elemental stoichiometry by factors of 2-5x relative to oxic periods. Importantly, prolonged summer anoxia increased nitrogen export from the reservoir by six-fold and changed the reservoir from a net sink to a net source of phosphorus and organic carbon downstream. While low oxygen in freshwaters is thought of as a response to land use and climate change, results from REDOX demonstrate that low oxygen can also be a driver of major changes to freshwater biogeochemical cycling, which may serve as an intensifying feedback that increases anoxia in downstream waterbodies. Consequently, as climate and land use change continue to increase the prevalence of anoxia in lakes and reservoirs globally, it is likely that anoxia will have major effects on freshwater carbon, nitrogen, and phosphorus budgets as well as water quality and ecosystem functioning.

Near-term, iterative ecological forecasts with quantified uncertainty have great potential for improving lake and reservoir management. For example, if managers received a forecast indicating a high likelihood of impending impairment, they could make decisions today to prevent or mitigate poor water quality in the future. Increasing the number of automated, real-time freshwater forecasts used for management requires integrating interdisciplinary expertise to develop a framework that seamlessly links data, models, and cyberinfrastructure, as well as collaborations with managers to ensure that forecasts are embedded into decision-making workflows. The goal of this study is to advance the implementation of near-term, iterative ecological forecasts for freshwater management. We first provide an overview of FLARE (Forecasting Lake And Reservoir Ecosystems), a forecasting framework we developed and applied to a drinking water reservoir to assist water quality management, as a potential open-source option for interested users. We used FLARE to develop scenario forecasts simulating different water quality interventions to inform manager decision-making. Second, we share lessons learned from our experience developing and running FLARE over 2 years to inform other forecasting projects. We specifically focus on how to develop, implement, and maintain a forecasting system used for active management. Our goal is to break down the barriers to forecasting for freshwater researchers, with the aim of improving lake and reservoir management globally.

As climate and land use increase the variability of many ecosystems, forecasts of ecological variables are needed to inform management and use of ecosystem services. In particular, forecasts of phytoplankton would be especially useful for drinking water management, as phytoplankton populations are exhibiting greater fluctuations due to human activities. While phytoplankton forecasts are increasing in number, many questions remain regarding the optimal model time step (the temporal frequency of the forecast model output), time horizon (the length of time into the future a prediction is made) for maximizing forecast performance, as well as what factors contribute to uncertainty in forecasts and their scalability among sites. To answer these questions, we developed near-term, iterative forecasts of phytoplankton 1-14 days into the future using forecast models with three different time steps (daily, weekly, fortnightly), that included a full uncertainty partitioning analysis at two drinking water reservoirs. We found that forecast accuracy varies with model time step and forecast horizon, and that forecast models can outperform null estimates under most conditions. Weekly and fortnightly forecasts consistently outperformed daily forecasts at 7-day and 14-day horizons, a trend that increased up to the 14-day forecast horizon. Importantly, our work suggests that forecast accuracy can be increased by matching the forecast model time step to the forecast horizon for which predictions are needed. We found that model process uncertainty was the primary source of uncertainty in our phytoplankton forecasts over the forecast period, but parameter uncertainty increased during phytoplankton blooms and when scaling the forecast model to a new site. Overall, our scalability analysis shows promising results that simple models can be transferred to produce forecasts at additional sites. Altogether, our study advances our understanding of how forecast model time step and forecast horizon influence the forecastability of phytoplankton dynamics in aquatic systems and adds to the growing body of work regarding the predictability of ecological systems broadly.

Freshwater phytoplankton communities are currently experiencing multiple global change stressors, including increasing frequency and intensity of storms. An important mechanism by which storms affect lake and reservoir phytoplankton is by altering the water column's thermal structure (e.g., changes to thermocline depth). However, little is known about the effects of intermittent thermocline deepening on phytoplankton community vertical distribution and composition or the consistency of phytoplankton responses to varying frequency of these disturbances over multiple years. We conducted whole-ecosystem thermocline deepening manipulations in a small reservoir. We used an epilimnetic mixing system to experimentally deepen the thermocline via five short (24-72 hr) mixing events across two summers, inducing potential responses to storms. For comparison, we did not manipulate thermocline depth in two succeeding summers. We collected weekly depth profiles of water temperature, light, nutrients, and phytoplankton biomass as well as bottle samples to assess phytoplankton community composition. We then used time-series analysis and multivariate ordination to assess the effects of intermittent thermocline deepening due to both our experimental manipulations and naturally occurring storms on phytoplankton community structure. We observed inter-annual and intra-annual variability in phytoplankton community response to thermocline deepening. We found that peak phytoplankton biomass was significantly deeper in years with a higher frequency of thermocline deepening events (i.e., years with both manipulations and natural storms) due to altered thermal stratification and more variable depth distributions of soluble reactive phosphorus. Furthermore, we found that the depth of peak phytoplankton biomass was linked to phytoplankton community composition, with certain taxa being associated with deep or shallow biomass peaks, often according to functional traits such as optimal growth temperature, mixotrophy, and low-light tolerance. For example, Cryptomonas taxa, which are low-light tolerant and mixotrophic, were associated with deep peaks, while the cyanobacterial taxon Dolichospermum was associated with shallow peaks. Our results demonstrate that abrupt thermocline deepening due to water column mixing affects both phytoplankton depth distribution and community structure via alteration of physical and chemical gradients. In addition, our work supports previous research that phytoplankton depth distributions are related to phytoplankton community composition at inter-annual and intra-annual timescales. Variability in the inter-annual and intra-annual responses of phytoplankton to abrupt thermocline deepening indicates that antecedent conditions and the seasonal timing of surface water mixing may mediate these responses. Our findings emphasise that phytoplankton depth distributions are sensitive to global change stressors and effects on depth distributions should be taken into account when predicting phytoplankton responses to increased storms under global change.

Associations between animals and microbes affect not only the immediate tissues where they occur, but also the entire host. Metabolomics, the study of small biomolecules generated during metabolic processes, provides a window into how mutualistic interactions shape host biochemistry. The Hawaiian bobtail squid, Euprymna scolopes, is amenable to metabolomic studies of symbiosis because the host can be reared with or without its species-specific symbiont, Vibrio fischeri. In addition, unlike many invertebrates, the host squid has a closed circulatory system. This feature allows a direct sampling of the refined collection of metabolites circulating through the body, a focused approach that has been highly successful with mammals. Here, we show that rearing E. scolopes without its natural symbiont significantly affected one-quarter of the more than 100 hemolymph metabolites defined by gas chromatography mass spectrometry analysis. Furthermore, as in mammals, which harbor complex consortia of bacterial symbionts, the metabolite signature oscillated on symbiont-driven daily rhythms and was dependent on the sex of the host. Thus, our results provide evidence that the population of even a single symbiont species can influence host hemolymph biochemistry as a function of symbiotic state, host sex and circadian rhythm.

Sheet 1: Relative expression values of ssrA from bacteria cells fraction or OMV fractions. Cells grown in three different media: a tryptone-based medium (LBS) or LBS with the addition of either glycerol (32.6 mM) or GlcNAc (10 mM). S1A Fig. Sheet 2: OD600 values over 24 h of bacteria growth in tryptone-based medium (LBS). S1A Fig. Sheet 3: OD600 values over 24 h of bacteria growth in minimum medium. S1A Fig. Sheet 4: Motility in soft agar of WT or DeltassrA cells measured as the diameter of the outer migration ring at 3 and 7 h post inoculation. S2B Fig. Sheet 5: Respiration rates of WT, DeltassrA, DeltassrA + ssrA, and a nonluminescent lux-deletion mutant (Deltalux) normalized to OD600. Fig 2C. Sheet 6: RCI between WT and DeltassrA in co-inoculated light organs after 24, 48, and 72 h. S2D Fig. Sheet 7: Relative expression values of ssrA and smpB. S2E Fig. GlcNAc, N-acetyl-glucosamine; LBS, Luria-Bertani salt medium; OD600, optical density at 600 nm; OMV, outer membrane vesicle; RCI, relative competitive index; WT, wild type. (XLSX) Copyright: CC BY 4.0

Sheet 1: Survival proportions of juvenile squid colonized by either WT, DeltassrA, DeltassrA + ssrA, or DeltasmpB. Fig 4B, S6B Fig. Sheet 2: Dry weight of juvenile squid immediately after hatching ("Hatch") or at 4 d post hatching when kept APO or colonized with WT, DeltassrA, or a nonluminescent mutant (Deltalux) strain. Fig 4C. Sheet 3: Quantification of internal yolk-sac area of juvenile squid immediately after hatching ("Hatch") or at 2 d post hatching colonized with WT or DeltassrA. Fig 4D. APO, aposymbiotic; WT, wild type. (XLSX) Copyright: CC BY 4.0