Physiology, fast and slow: bacterial response to variable resource stoichiometry and dilution rate.

Microorganisms grow despite imbalances in the availability of nutrients and energy. The biochemical and elemental adjustments that bacteria employ to sustain growth when these resources are suboptimal are not well understood. We assessed how Pseudomonas putida KT2440 adjusts its physiology at differing dilution rates (to approximate growth rates) in response to carbon (C), nitrogen (N), and phosphorus (P) stress using chemostats. Cellular elemental and biomolecular pools were variable in response to different limiting resources at a slow dilution rate of 0.12 h-1, but these pools were more similar across treatments at a faster rate of 0.48 h-1. At slow dilution rates, limitation by P and C appeared to alter cell growth efficiencies as reflected by changes in cellular C quotas and rates of oxygen consumption, both of which were highest under P- and lowest under C- stress. Underlying these phenotypic changes was differential gene expression of terminal oxidases used for ATP generation that allows for increased energy generation efficiency. In all treatments under fast dilution rates, KT2440 formed aggregates and biofilms, a physiological response that hindered an accurate assessment of growth rate, but which could serve as a mechanism that allows cells to remain in conditions where growth is favorable. Our findings highlight the ways that microorganisms dynamically adjust their physiology under different resource supply conditions, with distinct mechanisms depending on the limiting resource at slow growth and convergence toward an aggregative phenotype with similar compositions under conditions that attempt to force fast growth. IMPORTANCE: All organisms experience suboptimal growth conditions due to low nutrient and energy availability. Their ability to survive and reproduce under such conditions determines their evolutionary fitness. By imposing suboptimal resource ratios under different dilution rates on the model organism Pseudomonas putida KT2440, we show that this bacterium dynamically adjusts its elemental composition, morphology, pools of biomolecules, and levels of gene expression. By examining the ability of bacteria to respond to C:N:P imbalance, we can begin to understand how stoichiometric flexibility manifests at the cellular level and impacts the flow of energy and elements through ecosystems.

Global-to-Local Dependencies in Phosphorus Mass Flows and Markets: Pathways to Improving System Resiliency in Response to Exogenous Shocks.

Uneven global distribution of phosphate rock deposits and the supply chains to transport phosphorus (P) make P fertilizers vulnerable to exogenous shocks, including commodity market shocks; extreme weather events or natural disasters; and geopolitical instability, such as trade disputes, disruption of shipping routes, and war. Understanding bidirectional risk transmission (global-to-local and local-to-global) in P supply and consumption chains is thus essential. Ignoring P system interdependencies and associated risks could have major impacts on critical infrastructure operations and increase the vulnerability of global food systems. We highlight recent unanticipated events and cascading effects that have impacted P markets globally. We discuss the need to account for exogenous shocks in local assessments of P flows, policies, and infrastructure design choices. We also provide examples of how accounting for undervalued global risks to the P industry can hasten the transition to a sustainable P future. For example, leveraging internal P recycling loops, improving plant P use efficiency, and utilizing legacy soil P all enhance system resiliency in the face of exogenous shocks and long-term anticipated threats. Strategies applied at the local level, which are embedded within national and global policy systems, can have global-scale impacts in derisking the P supply chain.

Understanding stoichiometric constraints on growth using resource use efficiency imbalances.

Growth is a function of the net accrual of resources by an organism. Energy and elemental contents of organisms are dynamically linked through their uptake and allocation to biomass production, yet we lack a full understanding of how these dynamics regulate growth rate. Here, we develop a multivariate imbalance framework, the growth efficiency hypothesis, linking organismal resource contents to growth and metabolic use efficiencies, and demonstrate its effectiveness in predicting consumer growth rates under elemental and food quantity limitation. The relative proportions of carbon (%C), nitrogen (%N), phosphorus (%P), and adenosine triphosphate (%ATP) in consumers differed markedly across resource limitation treatments. Differences in their resource composition were linked to systematic changes in stoichiometric use efficiencies, which served to maintain relatively consistent relationships between elemental and ATP content in consumer tissues and optimize biomass production. Overall, these adjustments were quantitatively linked to growth, enabling highly accurate predictions of consumer growth rates.

Elemental and macromolecular plasticity of Chlamydomonas reinhardtii (Chlorophyta) in response to resource limitation and growth rate.

With the ongoing differential disruption of the biogeochemical cycles of major elements that are essential for all life (carbon, nitrogen, and phosphorus), organisms are increasingly faced with a heterogenous supply of these elements in nature. Given that photosynthetic primary producers form the base of aquatic food webs, impacts of changed elemental supply on these organisms are particularly important. One way that phytoplankton cope with the differential availability of nutrients is through physiological changes, resulting in plasticity in macromolecular and elemental biomass composition. Here, we assessed how the green alga Chlamydomonas reinhardtii adjusts its macromolecular (e.g., carbohydrates, lipids, and proteins) and elemental (C, N, and P) biomass pools in response to changes in growth rate and the modification of resources (nutrients and light). We observed that Chlamydomonas exhibits considerable plasticity in elemental composition (e.g., molar ratios ranging from 124 to 971 for C:P, 4.5 to 25.9 for C:N, and 15.1 to 61.2 for N:P) under all tested conditions, pointing to the adaptive potential of Chlamydomonas in a changing environment. Exposure to low light modified the elemental and macromolecular composition of cells differently than limitation by nutrients. These observed differences, with potential consequences for higher trophic levels, included smaller cells, shifts in C:N and C:P ratios (due to proportionally greater N and P contents), and differential allocation of C among macromolecular pools (proportionally more lipids than carbohydrates) with different energetic value. However, substantial pools of N and P remained unaccounted for, especially at fast growth, indicating accumulation of N and P in forms we did not measure.

Uncovering the spatio-temporal dynamics of crop-specific nutrient budgets in China.

Nitrogen (N) and phosphorus (P) are two critical nutrients for agroecosystems. In meeting food demands, human use of both nutrients has crossed planetary boundaries for sustainability. Further, there has been a dramatic shift in their relative inputs and outputs, which may generate strong N:P imbalances. Despite enormous efforts on agronomic N and P budgets, the spatio-temporal characteristics of different crop types in using nutrients are unknown as are patterns in the stoichiometric coupling of these nutrients. Thus, we analyzed the annual crop-specific N and P budgets and their stoichiometric relations for producing ten major crops at the provincial level of China during 2004-2018. Results show that, China has generally witnessed excessive N and P input over the past 15 years, with the N balance remaining stable while the P balance increasing by more than 170%, thus resulting in a decline in the N:P mass ratios from 10.9 in 2004 to 3.8 in 2018. Crop-aggregated nutrient use efficiency (NUE) of N has increased by 10% in these years while most crops have shown a decreasing trend of this indicator for P, which reduced NUE of P from 75% to 61% during this period. At the provincial level, the nutrient fluxes of Beijing and Shanghai have obviously declined, while the nutrient fluxes of provinces such as Xinjiang and Inner Mongolia have increased significantly. Although N management has made progress, P management should be further explored in the future due to eutrophication concerns. More importantly, N and P management strategies for sustainable agriculture in China should take account of not only the absolute nutrient use, but also their stoichiometric balance for different crops in different locations.

Revisiting the growth rate hypothesis: Towards a holistic stoichiometric understanding of growth.

The growth rate hypothesis (GRH) posits that variation in organismal stoichiometry (C:P and N:P ratios) is driven by growth-dependent allocation of P to ribosomal RNA. The GRH has found broad but not uniform support in studies across diverse biota and habitats. We synthesise information on how and why the tripartite growth-RNA-P relationship predicted by the GRH may be uncoupled and outline paths for both theoretical and empirical work needed to broaden the working domain of the GRH. We found strong support for growth to RNA (r2  = 0.59) and RNA-P to P (r2  = 0.63) relationships across taxa, but growth to P relationships were relatively weaker (r2  = 0.09). Together, the GRH was supported in ~50% of studies. Mechanisms behind GRH uncoupling were diverse but could generally be attributed to physiological (P accumulation in non-RNA pools, inactive ribosomes, translation elongation rates and protein turnover rates), ecological (limitation by resources other than P), and evolutionary (adaptation to different nutrient supply regimes) causes. These factors should be accounted for in empirical tests of the GRH and formalised mathematically to facilitate a predictive understanding of growth.

A common temperature dependence of nutritional demands in ectotherms.

In light of ongoing climate change, it is increasingly important to know how nutritional requirements of ectotherms are affected by changing temperatures. Here, we analyse the wide thermal response of phosphorus (P) requirements via elemental gross growth efficiencies of Carbon (C) and P, and the Threshold Elemental Ratios in different aquatic invertebrate ectotherms: the freshwater model species Daphnia magna, the marine copepod Acartia tonsa, the marine heterotrophic dinoflagellate Oxyrrhis marina, and larvae of two populations of the marine crab Carcinus maenas. We show that they all share a non-linear cubic thermal response of nutrient requirements. Phosphorus requirements decrease from low to intermediate temperatures, increase at higher temperatures and decrease again when temperature is excessive. This common thermal response of nutrient requirements is of great importance if we aim to understand or even predict how ectotherm communities will react to global warming and nutrient-driven eutrophication.

Sustained stoichiometric imbalance and its ecological consequences in a large oligotrophic lake.

Considerable attention is given to absolute nutrient levels in lakes, rivers, and oceans, but less is paid to their relative concentrations, their nitrogen:phosphorus (N:P) stoichiometry, and the consequences of imbalanced stoichiometry. Here, we report 38 y of nutrient dynamics in Flathead Lake, a large oligotrophic lake in Montana, and its inflows. While nutrient levels were low, the lake had sustained high total N: total P ratios (TN:TP: 60 to 90:1 molar) throughout the observation period. N and P loading to the lake as well as loading N:P ratios varied considerably among years but showed no systematic long-term trend. Surprisingly, TN:TP ratios in river inflows were consistently lower than in the lake, suggesting that forms of P in riverine loading are removed preferentially to N. In-lake processes, such as differential sedimentation of P relative to N or accumulation of fixed N in excess of denitrification, likely also operate to maintain the lake's high TN:TP ratios. Regardless of causes, the lake's stoichiometric imbalance is manifested in P limitation of phytoplankton growth during early and midsummer, resulting in high C:P and N:P ratios in suspended particulate matter that propagate P limitation to zooplankton. Finally, the lake's imbalanced N:P stoichiometry appears to raise the potential for aerobic methane production via metabolism of phosphonate compounds by P-limited microbes. These data highlight the importance of not only absolute N and P levels in aquatic ecosystems, but also their stoichiometric balance, and they call attention to potential management implications of high N:P ratios.

Microplastics in Flathead Lake, a large oligotrophic mountain lake in the USA.

Microplastics are contaminants that are closely associated with human activity and are often abundant even in remote areas. As the largest natural freshwater lake in the western USA, Flathead Lake is a suitable site to study microplastics in lakes in less-populated areas of North America. Our assessment of microplastics in lake surface water samples showed that microplastic densities and concentrations in Flathead Lake were similar to those in other lakes located in less-populated areas around the world, with densities ranging from 8.00 × 104 to 4.22 × 105 particles/km2 with a mean concentration of 1.89 × 105 particles/km2. Dry deposition rates for microplastics ranged from 4 to 140 particles/m2/day with an average of 69 particles/m2/day and were significantly higher in the fall. Microplastic concentrations in wet deposition ranged from 0.006 particles/mL to 0.050 particles/mL with highest concentrations in winter and lowest in summer. Fibrous microplastics were predominant in both lake water and atmospheric deposition. The high densities of microplastics in the sample sites located near the Flathead River inlet suggests that the river is an important source of microplastics to Flathead Lake. The high densities of microplastics and high proportions of non-fibrous microplastics near populated areas of the lake imply that local human activities also affect microplastics in Flathead Lake. Although the annual flux of microplastics in dry deposition was higher than that in wet deposition, the relatively modest difference suggests that precipitation might enhance the deposition of microplastics. The results of this study indicate that instituting increased control measures that target both reducing the microfibers generated by laundry and improving the overall level of plastic waste management in the watershed may help in controlling microplastic levels in Flathead Lake.

Bacterial communities in surface and basal ice of a glacier terminus in the headwaters of Yangtze River on the Qinghai-Tibet Plateau.

BACKGROUND: On the front lines of climate change, glacier termini play crucial roles in linking glaciers and downstream ecosystems during glacier retreat. However, we lack a clear understanding of biological processes that occur in surface and basal ice at glacier termini. METHODS: Here, we studied the bacterial communities in surface ice and basal ice (the bottom layer) of a glacier terminus in the Yangtze River Source, Qinghai-Tibet Plateau. RESULTS: Surface and basal ice harbored distinct bacterial communities but shared some core taxa. Surface ice communities had a higher α-diversity than those in basal ice and were dominated by Proteobacteria, Firmicutes, Bacteroidetes, Actinobacteria, and Cyanobacteria while basal ice was dominated by Firmicutes and Proteobacteria. The bacterial communities were also substantially different in functional potential. Genes associated with functional categories of cellular processes and metabolism were significantly enriched in surface ice, while genes connected to environmental information processing were enriched in basal ice. In terms of biogeochemical cycles of carbon, nitrogen, phosphorus, and sulfur, bacterial communities in surface ice were enriched for genes connected to aerobic carbon fixation, aerobic respiration, denitrification, nitrogen assimilation, nitrogen mineralization, sulfur mineralization, alkaline phosphatase, and polyphosphate kinase. In contrast, bacterial communities in basal ice were enriched for genes involved in anaerobic carbon fixation, fermentation, nitrate reduction, 2-aminoethylphosphonic acid pathway, G3P transporter, glycerophosphodiester phosphodiesterase, and exopolyphosphatase. Structural equation modeling showed that total nitrogen and environmental carbon:phosphorus were positively while environmental nitrogen:phosphorus was negatively associated with taxonomic ß-diversity which itself was strongly associated with functional ß-diversity of bacterial communities. CONCLUSIONS: This study furthers our understanding of biogeochemical cycling of the mountain cryosphere by revealing the genetic potential of the bacterial communities in surface and basal ice at the glacier terminus, providing new insights into glacial ecology as well as the influences of glacier retreat on downstream systems.

Species invasion progressively disrupts the trophic structure of native food webs.

Species invasions can have substantial impacts on native species and ecosystems, with important consequences for biodiversity. How these disturbances drive changes in the trophic structure of native food webs through time is poorly understood. Here, we quantify trophic disruption in freshwater food webs to invasion by an apex fish predator, lake trout, using an extensive stable isotope dataset across a natural gradient of uninvaded and invaded lakes in the northern Rocky Mountains, USA. Lake trout invasion increased fish diet variability (trophic dispersion), displaced native fishes from their reference diets (trophic displacement), and reorganized macroinvertebrate communities, indicating strong food web disruption. Trophic dispersion was greatest 25 to 50 y after colonization and dissipated as food webs stabilized in later stages of invasion (>50 y). For the native apex predator, bull trout, trophic dispersion preceded trophic displacement, leading to their functional loss in late-invasion food webs. Our results demonstrate how invasive species progressively disrupt native food webs via trophic dispersion and displacement, ultimately yielding biological communities strongly divergent from those in uninvaded ecosystems.

Key rules of life and the fading cryosphere: Impacts in alpine lakes and streams.

Alpine regions are changing rapidly due to loss of snow and ice in response to ongoing climate change. While studies have documented ecological responses in alpine lakes and streams to these changes, our ability to predict such outcomes is limited. We propose that the application of fundamental rules of life can help develop necessary predictive frameworks. We focus on four key rules of life and their interactions: the temperature dependence of biotic processes from enzymes to evolution; the wavelength dependence of the effects of solar radiation on biological and ecological processes; the ramifications of the non-arbitrary elemental stoichiometry of life; and maximization of limiting resource use efficiency across scales. As the cryosphere melts and thaws, alpine lakes and streams will experience major changes in temperature regimes, absolute and relative inputs of solar radiation in ultraviolet and photosynthetically active radiation, and relative supplies of resources (e.g., carbon, nitrogen, and phosphorus), leading to nonlinear and interactive effects on particular biota, as well as on community and ecosystem properties. We propose that applying these key rules of life to cryosphere-influenced ecosystems will reduce uncertainties about the impacts of global change and help develop an integrated global view of rapidly changing alpine environments. However, doing so will require intensive interdisciplinary collaboration and international cooperation. More broadly, the alpine cryosphere is an example of a system where improving our understanding of mechanistic underpinnings of living systems might transform our ability to predict and mitigate the impacts of ongoing global change across the daunting scope of diversity in Earth's biota and environments.

Bacterial Communities in Stream Biofilms in a Degrading Grassland Watershed on the Qinghai-Tibet Plateau.

Grassland is among the largest terrestrial biomes and is experiencing serious degradation, especially on the Qinghai-Tibet Plateau (QTP). However, the influences of grassland degradation on microbial communities in stream biofilms are largely unknown. Using 16S rRNA gene sequencing, we investigated the bacterial communities in stream biofilms in sub-basins with different grassland status in the Qinghai Lake watershed. Grassland status in the sub-basins was quantified using the normalized difference vegetation index (NDVI). Proteobacteria, Bacteroidetes, Cyanobacteria, and Verrucomicrobia were the dominant bacterial phyla. OTUs, 7,050, were detected in total, within which 19 were abundant taxa, and 6,922 were rare taxa. Chao 1, the number of observed OTUs, and phylogenetic diversity had positive correlations with carbon (C), nitrogen (N), and/or phosphorus (P) in biofilms per se. The variation of bacterial communities in stream biofilms was closely associated with the rate of change in NDVI, pH, conductivity, as well as C, N, P, contents and C:N ratio of the biofilms. Abundant subcommunities were more influenced by environmental variables relative to the whole community and to rare subcommunities. These results suggest that the history of grassland degradation (indicated as the rate of change in NDVI) influences bacterial communities in stream biofilms. Moreover, the bacterial community network showed high modularity with five major modules (>50 nodes) that responded differently to environmental variables. According to the module structure, only one module connector and 12 module hubs were identified, suggesting high fragmentation of the network and considerable independence of the modules. Most of the keystone taxa were rare taxa, consistent with fragmentation of the network and with adverse consequences for bacterial community integrity and function in the biofilms. By documenting the properties of bacterial communities in stream biofilms in a degrading grassland watershed, our study adds to our knowledge of the potential influences of grassland degradation on aquatic ecosystems.

Improvement in municipal wastewater treatment alters lake nitrogen to phosphorus ratios in populated regions.

Large-scale and rapid improvement in wastewater treatment is common practice in developing countries, yet this influence on nutrient regimes in receiving waterbodies is rarely examined at broad spatial and temporal scales. Here, we present a study linking decadal nutrient monitoring data in lakes with the corresponding estimates of five major anthropogenic nutrient discharges in their surrounding watersheds over time. Within a continuous monitoring dataset covering the period 2008 to 2017, we find that due to different rates of change in TN and TP concentrations, 24 of 46 lakes, mostly located in China's populated regions, showed increasing TN/TP mass ratios; only 3 lakes showed a decrease. Quantitative relationships between in-lake nutrient concentrations (and their ratios) and anthropogenic nutrient discharges in the surrounding watersheds indicate that increase of lake TN/TP ratios is associated with the rapid improvement in municipal wastewater treatment. Due to the higher removal efficiency of TP compared with TN, TN/TP mass ratios in total municipal wastewater discharge have continued to increase from a median of 10.7 (95% confidence interval, 7.6 to 15.1) in 2008 to 17.7 (95% confidence interval, 13.2 to 27.2) in 2017. Improving municipal wastewater collection and treatment worldwide is an important target within the 17 sustainable development goals set by the United Nations. Given potential ecological impacts on biodiversity and ecosystem function of altered nutrient ratios in wastewater discharge, our results suggest that long-term strategies for domestic wastewater management should not merely focus on total reductions of nutrient discharges but also consider their stoichiometric balance.

Effects of grassland degradation on ecological stoichiometry of soil ecosystems on the Qinghai-Tibet Plateau.

Grasslands across the world are being degraded due to the impacts of overgrazing and climate change. However, the influences of grassland degradation on carbon (C), nitrogen (N), and phosphorus (P) dynamics and stoichiometry in soil ecosystems are not well studied, especially at high elevations where ongoing climate change is most pronounced. Ecological stoichiometry facilitates understanding the biogeochemical cycles of multiple elements by studying their balance in ecological systems. This study sought to assess the responses of these soil elements to grassland degradation in the Qinghai Lake watershed on the Qinghai-Tibet Plateau (QTP), which has an average elevation of >4000 m and is experiencing serious grassland degradation due to its sensitivity and vulnerability to external disturbances. Substituting space for time, we quantified normalized difference vegetation index to gauge grassland degradation. C, N, and P concentrations and their molar ratios in soil and in soil microbial biomass were also measured. The results showed that grassland degradation decreased the concentrations of C and N, as well as the ratios of C:P and N:P in soil. The soil became relatively more P rich and thus N limitation is anticipated to be more apparent with grassland degradation. Moreover, C, N, and P concentrations in soil microbial biomass decreased with increased grassland degradation. C:N:P ratios of soil microbial biomass were highly constrained, suggesting that soil microorganisms exhibited a strong homeostatic behavior, while the variations of microbial biomass C:N:P ratios suggest changes in microbial activities and community structure. Overall, our study revealed that grassland degradation differentially affects soil C, N, and P, leading to decreased C:N and N:P in soil, as well as decreased C, N, and P concentrations in soil microbial biomass. This study provides insights from a stoichiometric perspective into microbial and biogeochemical responses of grassland ecosystems as they undergo degradation on the QTP.

Water Depth Underpins the Relative Roles and Fates of Nitrogen and Phosphorus in Lakes.

Eutrophication mitigation is an ongoing priority for aquatic ecosystems. However, the current eutrophication control strategies (phosphorus (P) and/or nitrogen (N)) are guided mainly by nutrient addition experiments in small waters without encompassing all in-lake biogeochemical processes that are associated largely with lake morphological characteristics. Here, we use a global lake data set (573 lakes) to show that the relative roles of N vs P in affecting eutrophication are underpinned by water depth. Mean depth and maximum depth relative to mixing depth were used to distinguish shallow (mixing depth > maximum depth), deep (mixing depth < mean depth), and transitional (mean depth ≤ mixing depth ≤ maximum depth) lakes in this study. TN/TP ratio (by mass) was used as an indicator of potential lake nutrient limitation, i.e., N only limitation if N/P < 9, N + P colimitation if 9 ≤ N/P < 22.6, and P only limitation if N/P ≥ 22.6. The results show that eutrophication is favored in shallow lakes, frequently (66.2%) with N limitation while P limitation predominated (94.4%) in most lakes but especially in deep ones. The importance of N limitation increases but P limitation decreases with lake trophic status while N and P colimitation occurs primarily (59.4%) in eutrophic lakes. These results demonstrate that phosphorus reduction can mitigate eutrophication in most large lakes but a dual N and P reduction may be needed in eutrophic lakes, especially in shallow ones (or bays). Our analysis helps clarify the long debate over whether N, P, or both control primary production. While these results imply that more resources be invested in nitrogen management, given the high costs of nitrogen pollution reduction, more comprehensive results from carefully designed experiments at different scales are needed to further verify this modification of the existing eutrophication mitigation paradigm.

Genomic adaptations in information processing underpin trophic strategy in a whole-ecosystem nutrient enrichment experiment.

Several universal genomic traits affect trade-offs in the capacity, cost, and efficiency of the biochemical information processing that underpins metabolism and reproduction. We analyzed the role of these traits in mediating the responses of a planktonic microbial community to nutrient enrichment in an oligotrophic, phosphorus-deficient pond in Cuatro Ciénegas, Mexico. This is one of the first whole-ecosystem experiments to involve replicated metagenomic assessment. Mean bacterial genome size, GC content, total number of tRNA genes, total number of rRNA genes, and codon usage bias in ribosomal protein sequences were all higher in the fertilized treatment, as predicted on the basis of the assumption that oligotrophy favors lower information-processing costs whereas copiotrophy favors higher processing rates. Contrasting changes in trait variances also suggested differences between traits in mediating assembly under copiotrophic versus oligotrophic conditions. Trade-offs in information-processing traits are apparently sufficiently pronounced to play a role in community assembly because the major components of metabolism-information, energy, and nutrient requirements-are fine-tuned to an organism's growth and trophic strategy.

Cascading influences of grassland degradation on nutrient limitation in a high mountain lake and its inflow streams.

Nitrogen (N) and phosphorus (P) are key growth-limiting nutrients for organisms and their absolute and relative supplies regulate the structure and function of ecosystems. Landcover changes lead to modifications of terrestrial biogeochemistry, consequently influencing aquatic nutrient conditions. This study sought to evaluate the potential impacts of grassland degradation on nutrient availability and nutrient limitation in the Qinghai Lake (China) and its inflow streams. We sampled nutrient concentrations and tested stream nutrient limitation by conducting nutrient diffusing substrata (NDS) bioassays in streams flowing through subbasins with different grassland status. To test nutrient limitation and the responses of lake phytoplankton to stream inflows, bioassays were conducted by adding different nutrients (N, P, and joint NP) as well as water from different streams to lake water with phytoplankton, respectively. In general, N concentrations as well as N:P ratios decreased while P concentrations increased with decreased normalized difference vegetation index (NDVI, an index of vegetation status), especially in September, suggesting that grassland degradation (low NDVI) has the potential to differentially decrease N availability and increase P availability in streams. Consistent with this, relative responses (RR) of stream periphyton to P and combined NP enrichments in the NDS bioassays decreased with stream P concentrations while increased with stream water N:P ratios. Lake phytoplankton responded strongly to P and combined NP addition indicating strong P-limitation of lake phytoplankton. RR of lake phytoplankton to stream water decreased with nitrate concentration and N:P ratios in stream water and increased with the concentrations of ammonium, total phosphorus, and soluble reactive phosphorus, indicating that stream water with higher P but lower N and N:P from degraded subcatchments is associated with increased impact on P-limited Lake phytoplankton. Overall, this study suggests that grassland degradation has the potential to differentially influence the nutrients delivered to streams with substantial increases in P but decreases in N and N:P, alleviating P limitation of stream periphyton and, ultimately, stimulating P-limited phytoplankton growth in the lake.

The stoichiometric legacy of fire regime regulates the roles of micro-organisms and invertebrates in decomposition.

Decadal-scale increases in fire frequency have the potential to deplete ecosystems of essential nutrients and consequently impede nutrient-limited biological processes via stoichiometric imbalance. Decomposition, a fundamental ecosystem function and strong driver of future fire occurrence, is highly sensitive to nutrient availability and is, therefore, particularly important in this context. Here we show that 40 yr of quadrennial (4yB) and biennial (2yB) prescribed burning result in severely P- and N-depleted litter stoichiometry, respectively, relative to fire exclusion. These effects exacerbated the nutrient limitation of microbial activities, constraining litter decomposition by 42.1% (4yB) and 23.6% (2yB) relative to unburned areas. However, invertebrate-driven decomposition largely compensated for the diminished capacity of micro-organisms under 4yB, suggesting that invertebrates could have an important stabilizing influence in fire-affected ecosystems. This effect was strongly positively coupled with the strength of microbial P-limitation and was not obviously or directly driven by fire regime-induced changes in invertebrate community assemblage. Together, our results reveal that high-frequency fire regimes promote nutrient-poor, carbon-rich ecosystem stoichiometry and, in doing so, disrupt ecosystem processes and modify the relative functionality of micro-organisms and invertebrates.