Body size is a “master trait” that affects all vital rates, including feeding, reproduction, excretion and metabolism (Kleiber, 1932, 1961; Schmidt-Nielsen, 1984; Hirst et al., 2014). Understanding what drives the body mass dependence of such a wide diversity of rates is of fundamental biological importance, indeed, this has been a much-debated topic over the last century. Recent work has explored body mass scaling exponents of metabolic rates within planktonic species (Hirst et al., 2014; Glazier et al., 2015) in order to better appreciate what controls these terms, and ultimately to better predict these rates for species and communities. These authors tested two groups of theories that predict the body-mass dependence of metabolism, those built upon internal transport networks (including the Metabolic Theory of Ecology; West et al., 1999; Savage et al., 2008; Banavar et al., 2010), and those based on a Surface Area model [a reapplication of Rubner’s surface dependent model of heat exchange in endotherms (Rubner, 1883), but more broadly applied to the influx and efflux of materials and energy]. Importantly, many zooplankton change body shape as they grow, while also using significant proportions of their body surface for the exchange of materials. While the major geometric scaling theories produce rather similar predictions when shape does not change over ontogeny (i.e., they are isomorphic), the predictions from these two groups of theory diverge starkly when organisms increasingly flatten or elongate in shape over ontogeny. These shape changes result in a reduction in the predicted scaling exponents of many resource transport model, but increase the predicted scaling exponent for the Surface Area dependent model. While the mass-scaling of respiration has been shown to correlate with body surface enlargement in many pelagic invertebrates (Hirst et al., 2014; Glazier et al., 2015), Hirst et al. (2016) predicted that body-mass scaling exponents for rates of soluble N excretion (b_N) should also then relate to the degree of body-shape change during growth. They tested this hypothesis using literature data on b_N for pelagic invertebrates across five different phyla, and found that b_N is significantly positively correlated with predicted surface area enlargement, whilst also co-varying with the mass-scaling of respiration rate (b_R). Indeed, intraspecific differences between b_N and b_R values have revealed there are shifts in the ratio of O₂-consumed to N-excreted over ontogeny. This suggests that changes in the relative anabolism and catabolism of proteins and lipids over development, may cause these consumption-excretion ratios to change too. In conclusion, diverse pelagic invertebrates, that dominate vast open water ecosystems, therefore appear to falsify the predictions of general metabolic scaling theories built upon resource-transport networks, while supporting predictions of surface-area dependent theory. Furthermore, ontogenetic variation in ratios of O₂ consumed to N excreted of these species, may not only provide insight into the developmental metabolism, but also the stoichiometry of ecological systems, including, for example, seasonal changes in N-budgets that are linked to pelagic animal life cycles.

Aquatic ecosystems can be sources of greenhouse gases (GHG), a process that is strongly controlled by the availability of C, N, and P, which can stimulate emission of nitrous oxide (N₂O), methane (CH₄), and carbon dioxide (CO₂) (Cao et al., 1996; Burgin et al., 2013; Nisbet et al., 2014; Deemer et al., 2016). However, mounting evidence suggests that benthic aquatic invertebrates such as midge larvae (Diptera: Chironomidae), snails (Gastropoda), and aquatic worms (Oligochaeta and Polychaeta) can enhance the emissions of GHG through high N excretion rates, by creating anoxic microenvironments within their guts, and through bioturbation and bioirrigation of surrounding sediments (Kristensen et al., 1991; Nielsen et al., 2004; Figueiredo-Barros et al., 2009; Stief et al., 2009; Heisterkamp et al., 2010; Nogaro and Burgin, 2014; Poulsen et al., 2014; Hölker et al., 2015; Mehring et al., 2017).

A large portion of the CH₄ produced in freshwater and marine sediments that is not released by ebullition is oxidized to CO₂ or assimilated by methanotrophic bacteria (Bastviken et al., 2008). Some species of midge larvae and zooplankton have been shown to assimilate methane-derived C through consumption of methanotrophic bacteria (Deines et al., 2007), as evidenced by exceptionally low stable isotopic ratios (δ¹³C as low -64‰ for midge larvae; Jones et al., 2008). It is still unclear if differences in faunal isotopic ratios among aquatic ecosystems can be consistently linked to differences in ecosystem function, or if the effects of methanotroph consumption by invertebrates are substantial enough to influence emissions across the air–water interface of lakes and wetlands. For example, Kajan and Frenzel (1999) observed that both production and oxidation of CH₄ were enhanced in chironomid burrows in rice paddies, but there was no net effect on benthic CH₄ flux. The feeding activity of bacterivorous zooplankton such as Cladocera has been shown to suppress methanotrophic activity in laboratory mesocosms (Kankaala et al., 2007), but this has not yet been demonstrated to affect CH₄ fluxes at large scales. Conversely, bioturbation is a non-consumptive mechanism by which benthic fauna may influence CH₄ flux, which has been demonstrated in manipulative laboratory studies (Figueiredo-Barros et al., 2009) but has yet to be linked to differences in faunal stoichiometry.

While much work is needed to further elucidate the enhancement of microbial metabolic pathways and GHG flux by aquatic invertebrates, previous studies have demonstrated enhancement of GHG flux by invertebrates under highly controlled conditions in laboratories. An assessment of the effects of mixed assemblages (and likely resulting in a wide range of nutrient stoichiometry) under variable conditions is important to our understanding of faunal influence on GHG fluxes in aquatic ecosystems. Since taxa such as Tubificinae have been shown to enhance GHG flux (Nogaro and Burgin, 2014; Mehring et al., 2017) and also to reach high densities in eutrophic aquatic environments (Devine and Vanni, 2002), invertebrate enhancement of GHG emissions from aquatic ecosystems may be linked both to anthropogenically induced nutrient loading and resulting shifts in aquatic community structures. Given the variable environmental conditions in mixed biotic assemblages outside of controlled laboratory conditions, the degree to which the effects of invertebrates and their corresponding C:N:P can be detected relative to other drivers of GHG flux in field settings requires further investigation.

ES is an important framework for examining paired biogeochemical processes; however, ES studies in both terrestrial and aquatic systems are biased toward C, N, and P while trace elements are often neglected (Sterner and Elser, 2002). Recently, Kaspari and Powers (2016) argued the importance of expanding traditional models of co-limitation to include all 25 of life’s building elements. Including non-essential trace elements is also crucial to a holistic stoichiometric approach (MacNeill et al., 2016). Arsenic (As), mercury (Hg), selenium (Se) and other non-essential trace elements have been well studied individually (Boening, 2000; Farag et al., 2003; Schaller et al., 2010; Walters et al., 2015), but their pairings with other, more common elements have less frequently been evaluated (but see Wang et al., 2013). Integrating trace elements, their interactions with each other and their interactions with C, N, and P into studies of ES will provide a more complete picture of elemental cycling in ecosystems (Wang et al., 2013). The toxic trace element As can alter both ecosystem structure and function: In terms of ecosystem structure, As contamination decreases stream invertebrate abundance and diversity (Chaffin et al., 2005). Functionally, As affects cycling of common (N and P) stream nutrients (Lottig et al., 2007; Rodriguez Castro et al., 2015; MacNeill et al., 2016). In freshwaters, P is usually in the form of phosphate (PO₄^3-), which shares the same chemical structure as arsenate (AsO₄^3-), the most common form of As in oxygenated freshwaters (Button et al., 1973; Schaller et al., 2010). Consequently, As can be taken into bacterial, algal, and animal cells in place of P and decouple oxidative- and photo-phosphorylation, hindering energy production (Finnegan and Chen, 2012). Cells are less able to distinguish between As and P when P is low relative to As (Rodriguez Castro et al., 2015) and in particular when total P is less than ∼50 μg/L, as is the case in a majority of freshwaters (Villanueva et al., 2000; Binkley et al., 2004; Hall et al., 2013). Recently published research shows that As metabolism by the algae Chlorella vulgaris depends on the relative amount of P, which determines both uptake of P and the dominant metabolite excreted by cells (Baker and Wallschläger, 2016).

In addition to the interchangeability of As and P, the cycles of N and P are intimately linked (Cross et al., 2005; Schade et al., 2011). Because the cycles of N and P are so intertwined, it is likely that the As cycle is linked to the N cycle through P. Toxic effects of As tend to be greater in P limited environments (Rodriguez Castro et al., 2015) and P limitation depends on relative N availability (Tessier and Raynal, 2003; Schade et al., 2011; Rodriguez Castro et al., 2015). Therefore, linkages with N may explain why previous studies have not satisfactorily resolved how As affects P uptake (Pringle, 1991; Lottig et al., 2007; Hoellein et al., 2012). MacNeill et al. (2016) found evidence that ambient dissolved N:P, rather than P concentration alone or relative As:P, influences the amount of As removed from the water column by biofilm (assemblages of bacteria, algae, and fungi growing on rocks) uptake. The relative N:P dissolved in water as a driver of As uptake by biofilms has implications for the amount of As, metabolized by, retained in, and transferred through food webs. Therefore, expanding the framework of ES to include trace elements is important to understand their relationships with common elements and their effects on ecosystem functioning.

ES, together with biogeochemistry has been applied to understanding invasive species and changes to aquatic food webs in the San Francisco Bay Delta (Glibert et al., 2011; Glibert, 2012). In this ecosystem, the food web has changed significantly over the past decades, from phytoplankton to fish. Using 30 years of records of nutrient loads and concentrations and abundances of phytoplankton, zooplankton, macroinvertebrates, and fish it was shown that changes in ratios of N and P, together with changes in N form, have been significant drivers of changes in the food web (Figure 2). Members of different trophic levels were found to have different correlations with N and P, as did taxa within trophic levels. These patterns were consistent with the premise that the fish community shifted to species that were proportionately more P-rich over time as N and P ratios increased due to substantial increases in N loading and reductions in P. The patterns were also consistent with increased importance of a benthic food web following reductions in P loading. Changes in external nutrient loads also drove changes in biogeochemical fluxes at the sediment water interface, leading to increasing abundance of macrophytes, clams, and of the toxic algae Microcystis, along with more omnivorous fish fueled by a benthic food web. The picture that has emerged of this ecosystem is one where changes in the food web are now understood to follow the conceptual model of stoichiometry, and not purely stochastic events. Previously considered one of the most heavily invaded estuaries in the world, it is now clear that environmental changes, including nutrient ratios and concentrations, interact with vectors of invasion to enhance their success.

Invasive quagga mussels (Dreissena rostriformis bugensis) have caused unprecedented ecological and environmental changes in Lake Michigan. Declines in primary production, fish biomass, and turbidity as well as significant changes to food web structure, phytoplankton composition, and nutrient cycling pathways have all occurred as a result of the introduction of quagga mussels (Bunnell et al., 2006; Cuhel and Aguilar, 2013; Lin and Guo, 2016). As efficient ecosystem engineers, quagga mussels voraciously filter pelagic particulate matter and excrete/egest nutrients in the benthos resulting in significant alterations to water column and benthic chemistry (Schindler and Scheuerell, 2002; Madenjian et al., 2015). Specifically, nutrients and organic matter that served as an energy source for forage fish have been intercepted by quagga mussels and sequestered in the benthos. Therefore, quantifying the specific mechanisms and pathways by which invasive quagga mussels have altered organic C and nutrient cycling are needed to understand the response of the Lake Michigan ecosystem to these non-indigenous bivalves. In the absence of particulate organic matter, which has become scarce in the water column of Lake Michigan, quagga mussels have been shown to efficiently remove materials in the dissolved and colloidal phase (DeVilbiss and Guo, 2017). For example, laboratory incubations have demonstrated the ability of quagga mussels to efficiently remove material as small as 0.5 μm, indicating their potential to directly uptake DOM in the water column. Quagga mussels also directly excrete DOM, with smaller mussels excreting at a significantly higher rate than larger mussels. Excreted DOM had unique chromophoric and fluorescent properties characteristic of protein-like materials, a colloidal size spectrum centered at 1–5 kDa, low TOC/TDN ratios (1.1 ± 0.1) but higher TDN/TDP ratios (33 ± 4) and was predominately composed of structural (refractory) polysaccharides. These results indicated that excreted DOM was chemically altered not only in regards to C molecules, but in N:P ratios as well. Based on initial estimations, only around 11% of consumed organic C was excreted/egested by quagga mussels, indicating that quagga mussels may be a potential sink for organic matter as well as a CO₂ source via metabolism.

Understanding effects of nutrient addition on microbial to ecosystem-scale metabolic processes is essential to expanding theoretical predictions of elemental limitation among ecosystems (Elser et al., 2007). Ecosystems that are autotrophic are generally co-limited by N and P (Elser et al., 2007), whereas donor-controlled, detritus-based ecosystems are dominated by heterotrophic consumers whose responses to added nutrients depend on the stoichiometry of detrital resources (Manning et al., 2015). Added N and P both accelerate C loss in detritus-based streams through enhanced organic matter breakdown and export (Benstead et al., 2009; Rosemond et al., 2015; Manning et al., 2016), as well as through substrate-specific and whole-stream ER (Suberkropp et al., 2010; Kominoski et al., 2017). Litter breakdown rates are constrained by microbial nutrient limitation (both N and P) at low-to-moderate concentrations through changes in litter C:N and C:P stoichiometry (Kominoski et al., 2015; Manning et al., 2015). These collective findings emphasize the importance of microbial processes on ecosystem C loss and the potential for long-term vulnerability to sustained C losses with sustained or increased N and P availability (Alexander and Smith, 2006), which ultimately can be linked to nutrient stoichiometry.

Long-term studies of nutrient enrichment in forest streams show declines in ecosystem-scale C. Studies of added N and P in streams of the Coweeta Long Term Ecological Research Program in the southern Appalachians, United States, illustrate that nutrients increase C loss through enhanced microbial respiration rates and invertebrate feeding activities (Benstead et al., 2009; Suberkropp et al., 2010). Increasing N and P concentrations while maintaining N:P ratios can accelerate in-stream biological process that result in up to a 50% reduction in residence time of terrestrial C (Rosemond et al., 2015). Declines in organic matter standing stocks and increases in associated respiration rates with nutrient enrichment, appear to be driven more by N than P. Nutrient enrichment can alter the relationships between N and P supply ratio and ecosystem-level processes. For example, prior to nutrient enrichment whole-stream ER in Coweeta streams was higher at lower N:P, but during enrichment ER increased with increasing N:P (Kominoski et al., 2017). Increased heterotrophy from microbial to ecosystem-scales can occur at concentrations of N and P that are now common among pristine and human-impacted ecosystems (Alexander and Smith, 2006).

A central challenge for ecologists is to understand how climate warming will influence GPP and ER, due to the central role these processes play in structuring food web production and C and nutrient cycles (Peterson et al., 2001; Raymond et al., 2013; Hotchkiss et al., 2015). The combined frameworks of metabolic ecology and ES offer promise for developing a mechanistic understanding of how temperature influences freshwater metabolism (Sterner and Elser, 2002; Sibly et al., 2012). Yet, more explicit consideration of the coupling between metabolic theory and ES is required (Sterner, 2004; Cross et al., 2015). A growing literature suggests that temperature dependences of ecosystem processes may diverge strongly from predictions, particularly when temperature influences—or is associated with—changes in resource supply (Anderson-Teixeira et al., 2008; Valett et al., 2008; Yvon-Durocher et al., 2012; Huryn et al., 2014; Welter et al., 2015). A better mechanistic understanding of how temperature and nutrients interact to influence metabolism will likely improve these predictive models.

Model ecosystems, that are natural, can provide a powerful tool for quantifying these mechanisms at the ecosystem level. The Hengill geothermal area in Iceland represents one such natural laboratory for examining how temperature influences the structure and function of stream ecosystems (O’Gorman et al., 2012, 2014) by allowing a combination of field surveys, stream-side channel experiments, and whole-stream temperature manipulations. Recent experiments have discovered that temperature dependences (measured as apparent “activation energies”; Brown et al., 2004) for GPP and ER were 6.5- and 2.7-fold higher, respectively, than predicted by Metabolic Theory; interestingly, these relationships were similar to the temperature dependency of N₂-fixation (Welter et al., 2015), suggesting a strong interaction between temperature and nutrient supply. The stronger than expected temperature dependencies for GPP and ER likely resulted from N-limitation of production at low temperatures and release from N-limitation at warm temperatures by N₂-fixation and the addition of “new” N. In addition, these studies showed that N limitation was further alleviated by a temperature-induced increase in N use efficiency (Williamson et al., 2016). A similar increase in flux-based N use efficiency was found in a survey of natural geothermal streams, as well as a whole-stream warming experiment in this Icelandic catchment (Hood et al., unpublished data). Taken together, these results promise that a better understanding of the interactive effects of temperature and nutrients on organisms and elemental fluxes can be used to develop a strong mechanistic understanding of how climate warming will influence river metabolism.