Freshwater systems are a significant component of the global carbon cycle (Tranvik et al., 2018) and they emit substantial amounts of methane (CH₄) to the atmosphere (Bastviken et al., 2011). Within freshwater systems, shallow lakes are considered to be biogeochemical hot spots and are distributed worldwide (Downing 2010; Holgerson and Raymond 2016). In some regions, shallow lakes can present two contrasting states: a clear water state dominated by submerged macrophytes, with low turbidity and low phytoplankton biomass, and a turbid water state dominated by phytoplankton, with high turbidity and no submerged vegetation (Scheffer et al., 1993; Sánchez et al., 2015). Submerged vegetation plays a key role in maintaining clear vegetated states by preventing sediment resuspension, by taking up nutrients from the water column, and by providing refuge for zooplankton, among others (Scheffer 2001; Hilt 2015). Their presence can also affect other processes, such as primary production, carbon burial rates and greenhouse gas (GHG) emissions (Hilt et al., 2017). In particular, the effect of submerged vegetation or phytoplankton on CH₄ dynamics is not well understood (Hilt et al., 2017). Submerged vegetation can enhance methanogenesis by providing macrophyte-derived carbon to the sediments (Emilson et al., 2018; Grasset et al., 2019), but phytoplankton-derived carbon has also been reported to enhance methanogenesis in sediments (Schwarz et al., 2008; West et al., 2012). Similarly, alternative states could have a differential effect on methane oxidation (MOX). The activity of methane oxidizing bacteria (MOB) depends mainly on, O₂ concentration and light penetration, where lower O₂ concentrations in combination with a reduction of light favors methanotrophs (Thottathil et al., 2018). Both submerged vegetation and phytoplankton can generate high oxygen concentrations and they can also diminish light penetration in the water column (Torremorell et al., 2009; Andersen et al., 2017). At the same time, it has been shown that submerged vegetation diminishes wind induced turbulence in the water column (Herb and Stefan 2005; Andersen et al., 2017), which could have a physical effect on gas exchange with the atmosphere, and this effect is not present in turbid phytoplanktonic lakes. Thus, it is not straightforward to predict potential biological—methanogenesis and methanotrophy—and physical - gas exchange with the atmosphere and also vertical and horizontal transport - differences in CH₄ dynamics between clear vegetated and turbid algal shallow lakes.

In a previous study we reported that clear vegetated shallow lakes from the Pampean Plain of Argentina had higher mean annual surface water CH₄ partial pressure (pCH₄) in comparison with turbid algal lakes (Baliña et al. under revision). However, we also reported that clear and turbid lakes presented similar mean annual CH₄ diffusive fluxes. The average CH₄ concentration in the water is the net balance between the rates of input to the water column, oxidation within the water column, and outflux to the atmosphere (Vachon et al., 2019; Noyce and Megonigal, 2021). Given that the average fluxes to the atmosphere were similar between states (Baliña et al. under revision), the differences in average pCH₄ could be the result of a physical effect due to differences in gas exchange, potentially combined with a biological effect, due to differences in the net balance between input and oxidation in the water column between the two states. CH₄ can be emitted from surface waters by diffusive flux, a strictly fickian process which depends on the concentration gradient at the water-air interface and the gas exchange velocity (k) and, in some cases, CH₄ can also be emitted in the form of microbubbles (Bastviken et al., 2004; Beaulieu et al., 2012). If CH₄ microbubbles are present, they can generate an additional flux of CH₄ to the atmosphere which will lead to increased measured k that is difficult to distinguish from that of purely diffusive k, and to a decoupling between CH₄ and CO₂ exchange velocities (Beaulieu et al., 2012; Prairie and del Giorgio 2013; McGinnis et al., 2015). This is a CH₄ emission pathway that is thought to relate positively to both water column turbulence and surface water CH₄ concentration (Prairie and del Giorgio 2013; McGinnis et al., 2015; Tang et al., 2016), yet it is currently difficult to predict whether these two contrasting lake states may be associated with an increased incidence of microbubbles.

In this study we assess both biological and physical factors influencing ambient surface pCH₄ in these shallow lakes: as biological factors, we explored potential differences in overall CH₄ oxidation between clear vegetated and turbid algal lakes, and by combining the patterns of oxidation and diffusive flux we infer potential differences in CH₄ input between states. As physical factors, we explored potential differences in kCH₄ between clear vegetated and turbid algal shallow lakes and also possible differences in the relationship between kCH₄ and wind, and between kCH₄ and pCH₄. In addition, we compared the patterns of kCO₂ and kCH₄ to infer differences in CH₄ microbubble dynamics between the two states.

Clear vegetated lakes had three-fold higher mean annual pCH₄ than turbid algal lakes (1,181.7 ± 1,375.8 ppmv and 358.9 ± 390.4 ppmv, respectively) (p = 0.002, df = 1, F = 10.6; Figure 2A; data from Baliña et al. (under revision)). Nonetheless, clear vegetated and turbid algal lakes had similar mean annual CH₄ diffusive fluxes (14.4 ± 24.2 and 19.9 ± 33.5 mmol m⁻² d⁻¹, respectively) (Figure 2B; data from Baliña et al. (under revision)). Mean annual δ¹³C-CH₄ was similar between clear vegetated and turbid algal shallow lakes and in both states and the isotopic signatures corresponded to enriched values related to fresh sediment CH₄ (Figure 2C and Supplementary Table S2). Our estimates of mean annual CH₄ fraction of oxidation (F_ox), obtained using the non-steady state closed model and assuming an average fractionation (α) of 1.021, were also very similar between clear vegetated and turbid algal lakes (average 0.57 ± 0.09 and 0.58 ± 0.11, respectively; Figure 2C). The steady state open model was not appropriate for these systems, since in a significant number of cases it yielded F_ox values higher than one (Supplementary Figure S2), as has been observed in other studies (Bastviken et al., 2002; Barbosa et al., 2018; Thottathil et al., 2018). Using α = 1.005 we obtained nonsensical F_ox values for both closed and open models, whereas using α = 1.031 we obtained logical F_ox values for both closed and open models that were in the range of the results obtained with α of 1.021 (Supplementary Figure S3).

There was a large range in measured k ₆₀₀ values based on either CO₂ or CH₄ across lakes (from 0.3 to 59.8 m d⁻¹), and although both agreed well for approximately 32% of observations, there was a high proportion of points that deviated significantly from the expected 1:1 relationship, and most of these points corresponded to turbid shallow lakes (Figure 3). The overall mean annual k ₆₀₀ CH₄ (12.7 ± 15.1 m d⁻¹) was significantly (6.4 times) higher than mean annual k ₆₀₀ CO₂ (1.98 ± 1.43 m d⁻¹), (p < 0.0001, df = 1, F = 19.7; Figure 4A). There were also differences between states in the patterns of gas exchange: mean annual k ₆₀₀ CH₄ was 3 times higher in turbid lakes (19.3 ± 18.9 m d⁻¹) than in clear lakes (6.7 ± 7.0 m d⁻¹) (p = 0.0092, df = 1, F = 7.5, Figure 4B), whereas mean annual k ₆₀₀ CO₂ was similar between clear and turbid lakes (2.0 ± 1.5 m d⁻¹ and 2.0 ± 1.4 m d⁻¹, respectively, Figure 4C).

We explored the relationships between K₆₀₀ and wind speed and also between K₆₀₀ and CH₄ and CO₂ partial pressure in the water, separately for clear vegetated and turbid algal lakes. Mean annual wind speed was similar between clear vegetated and turbid algal lakes (Supplementary Figure S4). There was a significant positive relationship between k ₆₀₀ CO₂ and wind speed (Figure 5A) and also between k ₆₀₀ CH₄ and wind speed (Figure 5B), but in both cases these relationships were present only for turbid algal lakes and were not significant for clear vegetated lakes. These two relationships with wind in turbid lakes, however, are strikingly different: the slope of the k ₆₀₀ CH₄ vs. wind relationship (4.9 ± 1.9) is one order of magnitude higher than that of k ₆₀₀ CO₂ (0.4 ± 0.1), and the intercepts are significantly different as well (6 ± 7.7 vs. 1 ± 0.5, respectively). Regarding dissolved GHG, pCO₂ was weakly and negatively related to k ₆₀₀ CO₂ in both clear vegetated and turbid algal shallow lakes, and the relationship was similar for both states (Figure 5C). Likewise, there was a weak (but not significant) negative relationship between ambient pCH₄ and k ₆₀₀ CH₄ in clear vegetated lakes yet, interestingly, there was a strong significant positive relationship between pCH₄ and k ₆₀₀ CH₄ in turbid algal lakes (Figure 5D).

Surface water δ ¹³C-CH₄ corresponded to enriched methane in both clear vegetated and turbid algal shallow lakes (between −50‰ and −25‰), followed by also similar percentages of CH₄ oxidation (mean of 57%) and, therefore, implying a high methanotrophic activity in both states. A comparable range of δ¹³C-CH₄ was reported for a subtropical shallow wetland from Australia (−53 and −39‰, Jeffrey et al., 2019) and a shallow Boreal lakes (−60 and −35‰, Desrosiers et al., 2021). A wider range of F_ox, also obtained using a non-steady state model, was reported for different habitat types within a shallow boreal lake (Desrosiers et al., 2021), with values ranging from 34% to 56% in Brasenia and Typha dominated habitats and down to 31% in open water areas. A wide range has also been reported for subtropical (15%–36%, Jeffrey et al., 2019), tropical (34%–100%; Barbosa et al., 2018), boreal (57%–75%; Bastviken et al., 2002) and temperate lakes (2%–97%; Thottathil et al., 2018) of varying size. This broad range of F_ox both within and across aquatic systems, highlights the complexity in the regulation of CH₄ oxidation. In this regard, the convergence in annual average F_ox between clear vegetated and turbid algal lakes despite their contrasting environmental conditions is remarkable.

Although there was significant seasonal variability in pCH₄ in both clear vegetated and turbid algal lakes (Baliña et al. (under revision)), this variation was centered around very different mean annual pCH₄ values for each state. Assuming that these annual means reflect an average steady state partial pressure of each lake state and are not varying greatly, then the amount of CH₄ exchanged at the water-air interface on an annual basis reflects the total CH₄ input to the water column (which includes CH₄ production in the sediments and water column, as well as lateral input) minus the CH₄ oxidized. Considering that annually the fraction of oxidation and the rates of diffusive flux were similar between states, we can infer that on an annual basis CH₄ input would be rather comparable between clear vegetated and turbid algal shallow lakes. CH₄ production in the sediments depends mainly on temperature, oxygen concentration and on the amount and type of organic matter (Megonigal et al., 2003; Duc et al., 2010). Both macrophyte and phytoplankton derived organic matter are known to favor methanogenesis (Emilson et al., 2018; Yan et al., 2019), but the extent to which the dominance of these different sources of organic matter may condition carbon cycling and methanogenesis at the ecosystem level is not well understood, with few studies having assessed this impact (Brothers et al., 2013). Although in this study we do not explicitly address CH₄ production, these results imply a comparable mean annual CH₄ input between clear vegetated and turbid algal states, which could be plausible since the characteristics of both states seem to favor sediment methanogenesis. Independent of sediment production, several authors have demonstrated that there is also a significant potential for CH₄ production in the water column (Grossart et al., 2011; Bogard et al., 2014; Günthel et al., 2019). Whereas there are studies that have explored possible isotopic signatures for this CH₄ (Günthel et al., 2020; Hartmann et al., 2020), there is to date no clear information of what the isotopic signature of this fresh pelagic CH₄ could be. Therefore, we could not include this source of CH₄ in the isotopic mass balance. Nonetheless, we consider that for the purpose of the present study it is sufficient to include a sedimentary methane source to derive a first order estimate of oxidation extent.

In our study, k ₆₀₀ CH₄ was 6.4 times higher than k ₆₀₀ CO₂. Previous studies have reported k ₆₀₀ CH₄ to be on average 1.8-fold higher than k ₆₀₀ CO₂ in two boreal lakes (Rantakari et al., 2015), 2.3-fold higher in a Canadian hydroelectric reservoir and boreal lakes (Prairie and del Giorgio 2013), 2.5-fold times higher in oligotrophic Lake Stechlin (McGinnis et al., 2015), and 2.5-fold higher in a tropical reservoir from Brazil (Paranaíba et al., 2018). In all cases, the differences between CH₄ and CO₂ exchange velocities were explained by the presence of CH₄ microbubbles, which result in a k ₆₀₀ CH₄ that is higher than that from diffusive exchange alone (Beaulieu et al., 2012; Prairie and del Giorgio 2013) and, therefore, generates a decoupling between k ₆₀₀ CH₄ and k ₆₀₀ CO₂ (Prairie and del Giorgio 2013; McGinnis et al., 2015). Moreover, we observed that turbid lakes had higher k ₆₀₀ CH₄ than clear lakes, which would suggest a differential CH₄ microbubble formation between states. CH₄ microbubbles are thought to be produced as a combination of CH₄ supersaturation and turbulence (Vagle et al., 2010; Prairie and del Giorgio 2013; McGinnis et al., 2015). Although clear vegetated lakes had a higher mean annual pCH₄ than turbid algal lakes, overall pCH₄ was high in both states, and even higher in comparison with other studies that reported CH₄ microbubble formation (Prairie and del Giorgio 2013; McGinnis et al., 2015; Tang et al., 2016). Therefore, both clear vegetated and turbid algal lakes could potentially harbor the production of CH₄ microbubbles in terms of the amount of CH₄ present in the water column. On the other hand, water column turbulence is substantially different between states, as evidenced by the different relationship that exists between k ₆₀₀ CO₂ and wind speed and as has been reported in previous studies (Herb and Stefan 2005; Andersen et al., 2017): in clear vegetated lakes submerged vegetation tends to suppress wind induced turbulence in the water column, whereas in turbid algal lakes the absence of submerged vegetation allows a higher wind induced turbulence. This might explain the observed apparent higher CH₄ microbubble formation in turbid algal lakes, leading to higher k ₆₀₀ CH₄ in comparison with clear vegetated lakes, in spite of average lower pCH₄.

Most models of diffusive gas exchange in lakes have positively linked k ₆₀₀ to wind speed (Sebacher et al., 1983; Raymond and Cole 2001; Guérin et al., 2007) and, if CH₄ microbubbles are present, k ₆₀₀ CH₄ is also expected to have a positive correlation with pCH₄ (Prairie and del Giorgio 2013). Our results suggest a fundamentally different response of CH₄ and CO₂ to wind forcing in turbid algal lakes, evidenced by the slope of the regressions, which also point towards CH₄ microbubble formation. In clear vegetated lakes, in contrast, neither k ₆₀₀ CH₄ nor k ₆₀₀ CO₂ were significantly related to wind, yet k ₆₀₀ CH₄ was nevertheless consistently higher than k ₆₀₀ CO₂, also implying CH₄ microbubble formation but with a different behavior towards wind turbulence. The positive and expected relationship between k ₆₀₀ CH₄ and pCH₄ was found only for turbid algal lakes and is consistent with a pattern of wind induced microbubble formation that is enhanced by increasing supersaturation of CH₄. This was not the outcome for clear vegetated lakes, implying that this wind vs. pCH₄ interaction is largely suppressed in vegetated habitats. On the other hand, ambient pCO₂ was weakly and negatively related to k ₆₀₀ CO₂ in both states in a very similar manner, which is coherent with the expected role of gas exchange as a modulator of ambient gas concentrations in surface waters, as has been previously reported for lakes (Lapierre et al., 2013) and rivers (Rocher-Ros et al., 2019). Therefore, in turbid algal lakes the combination of wind and pCH₄ determine k ₆₀₀ CH₄, yet in clear vegetated lakes, gas exchange is largely decoupled from wind, and under this circumstance a reciprocal relationship is established between pCH₄ and gas exchange: the wind-independent and relatively constant k in clear vegetated lakes leads to high pCH₄ because it acts as a lid, yet pCH₄ also appears to influence k ₆₀₀ CH₄, because the high pCH₄ leads to higher k ₆₀₀ CH₄ relative to CO₂ through microbubble formation. Submerged vegetation therefore influences gas dynamics two-fold in these clear vegetated lakes: directly by modulating the effect of wind on water column turbulence and therefore on gas exchange velocity, as is the case on CO₂ exchange, but also indirectly, by altering the dynamics of microbubble formation and therefore of CH₄ exchange.

Previous studies have also reported a strong impact of aquatic vegetation on gas exchange in shallow lakes. Kosten et al. (2016) reported a lower CH₄ exchange velocity in the presence of free-floating vegetation in comparison with open water sites, where the higher pCH₄ detected below the floating vegetation was partly explained by the lower k. Likewise, Barbosa et al. (2020) reported that vegetated and open water habitats from a tropical floodplain lake had similar CH₄ diffusive fluxes but that vegetated habitats had higher pCH₄, which was linked to a higher k in open water sites. Martinsen et al. (2020) also reported a lower CO₂ exchange velocity in a small shallow lake when submerged macrophytes were more abundant and related this observation with a negative effect of vegetation on the mixing of the water column. In our case, k ₆₀₀ CO₂ did not differ in average magnitude between lake states, but as pointed out above, the relationship between k ₆₀₀ CO₂ and wind differed markedly between states. An almost complete decoupling between exchange velocity and wind has been reported in previous studies carried out in small lakes (Heiskanen et al., 2014; Tedford et al., 2014; Tan et al., 2021), where wind-based models (Cole and Caraco 1998; Crusius and Wanninkhof 2003; MacIntyre et al., 2010) did not adequately explain the observed patterns in gas exchange, and where other factors, such as convection, had a stronger influence on gas exchange velocities. In our study, wind speed was a good predictor for k ₆₀₀ CO₂ and k ₆₀₀ CH₄ but only in turbid algal lakes. In clear vegetated lakes, submerged vegetation seems to decouple this relationship, therefore the use of wind speed would not be a good predictor for exchange velocities in these systems.

Overall, our results imply a roughly comparable mean annual CH₄ input to the water column between lakes in turbid algal and clear vegetated states, the latter inferred from similar mean annual CH₄ diffusive fluxes and mean annual CH₄ fraction of oxidation. We also observed that mean annual pCH₄ in clear lakes was 3 times higher than in turbid lakes, while mean annual k ₆₀₀ CH₄ in turbid lakes was 3 times higher than in clear lakes. Furthermore, the higher k ₆₀₀ CH₄ in turbid lakes was associated with a positive relationship with wind and pCH₄, and these relationships were absent in clear vegetated shallow lakes. Therefore, the higher pCH₄ in clear vegetated lakes could be explained by their lower average k ₆₀₀ CH₄ and also by the absence of a relation between k ₆₀₀ CH₄ and wind, which further suggests a lower CH₄ microbubble formation. These patterns seem to be driven by a physical effect produced by the submerged vegetation over the mixing of the water column: a reduction of water column turbulence apparently leads to both a lower exchange velocity and also to a reduction of CH₄ microbubble formation, consequently leading to higher surface water pCH₄ in clear vegetated lakes (Figure 6). Furthermore, whereas pCO₂ is at least in part controlled by k ₆₀₀ CO₂, as expected, pCH₄ seems to be differentially controlled depending on the lake state: on one hand, in clear lakes pCH₄ is primarily controlled by gas exchange, yet k ₆₀₀ CH₄ is to some degree also influenced by pCH₄, since there is some degree of microbubble formation and k ₆₀₀ CH₄ is still systematically higher than k ₆₀₀ CO₂. In turbid algal lakes, on the other hand, pCH₄ appears to influence the apparent k ₆₀₀ CH₄ through wind-driven CH₄ microbubble formation, but there is also a control of k ₆₀₀ CH₄ over pCH₄ (Figure 6). Therefore, physical rather than biological processes - ie. methanogenesis and methanotrophy-seem to be controlling the differences observed in surface water mean annual pCH₄ between clear vegetated and turbid algal shallow lakes from the Pampean plain.

The observation that k ₆₀₀ CH₄ is systematically higher than k ₆₀₀ CO₂ is consistent with the formation and subsequent emission of CH₄ microbubbles, and similar observations have been reported for rivers (Beaulieu et al., 2012; Campeau et al., 2014), lakes (McGinnis et al., 2015; Rantakari et al., 2015; Jansen et al., 2020), and reservoirs (Prairie and del Giorgio 2013; Paranaíba et al., 2018), with enhancement values ranging from 1.8 to 2.5. Although this appears to be a widespread phenomenon, the data are still sparse because there are surprisingly few studies that have measured CH₄ air-water exchange in parallel to that of another gas that can be used as a reference. As a result, the CH₄ microbubble dynamics in inland waters remains poorly constrained (Jansen et al., 2020), and this adds a large degree of uncertainty to current models and budgets of freshwater CH₄ emissions that already include ebullitive, diffusive and plant mediated fluxes. Here we have shown that there is indeed an interaction between wind velocity and surface water CH₄ concentration in determining the relative enhancement of k ₆₀₀ CH₄ but that this combined effect is only present in turbid shallow lakes. In vegetated clear lakes, the presence of submerged macrophytes seems to greatly dampen wind-induced water column turbulence, and under this scenario, k ₆₀₀ CH₄ responds to neither wind speed nor to pCH₄. Had we sampled lakes in only one state, we may have perhaps concluded that pCH₄ regulates k, or that k regulates pCH₄, and yet both are occurring but under different habitat and climatic combinations. It is clear that the regulation of water-air CH₄ exchange is complex, and an improved understanding of this process will require parallel CH₄ and CO₂ - or another reference gas - measurements carried out in a wide range of habitat and climatic conditions.