While soil incubation experiments are important methods for exploring mechanisms, their short-term nature, exclusion of roots, and the large disturbance to natural soil structure make it difficult to use them in isolation to ask questions about real-world soil responses to new temperature regimes. In turn, elevational gradient studies cannot capture the responses of tropical biota to continued warming because nowhere on Earth has an analog 'future' climate for lowland tropical forests9. Cross-site comparisons are able to take into consideration the long-term adaptation of tropical forests to temperature11; however, current changes are happening at a rapid rate and may shift forests to new regimes at a pace at which they may be unable to adjust10. Further, while in situ field warming experiments that warm only the soil provide invaluable insights, they are limited in their ability to interpret how changing plant ecophysiology may interact with changes to belowground processes. In response to these limitations, we developed an in situ field warming experiment - Tropical Responses to Altered Climate Experiment [TRACE] - in a tropical forested ecosystem to evaluate soil respiration responses to chronic warming20. We experimentally warmed understory plants and soils 4 °C above ambient temperatures to a depth of at least 50 cm using a hexagonal array of infrared heaters. Three warmed plots were paired with three ambient (control) plots of similar topographic position (Lower slope, Mid slope, and Upper slope). For a year, half-hourly soil respiration rates were measured in each plot using automated soil respiration chambers, resulting in 57,450 rate measurements for our study period of September 28, 2016, to September 5, 2017. In addition to the high temporal resolution soil respiration fluxes measured within the temperature manipulation plots, we performed an additional 152 flux measurements between November 4-19, 2020, across the larger field site to assess spatial heterogeneity in soil respiration rates.
Experimental warming resulted in substantial, larger than expected increases in soil respiration in plots that were experimentally warmed relative to the unwarmed control plots (Figs. 1 and 2). Warming increased CO flux by 42% and 59% in Mid and Lower slope positions, respectively (paired T-test, df = 24,521 pairs [Lower slope]; df = 15,635 pairs [Mid slope]). In addition to warming-induced increases in soil respiration means, warming increased the variability of the rates, which may represent an increased variability in environmental conditions or a synergy between warming and soil moisture controls. The positive response of CO flux to in situ warming was substantially higher in the Upper slope position relative to the lower topographic positions (204%; df = 12,910 pairs). For two of the paired plots (Lower and Mid slope), the warming-induced increases in respiration rates were in the upper range of what has been reported for higher latitude forests; however, the relative responses of the Upper slope were greater than rates observed in any field warming experiment, regardless of ecosystem type or methodology. Given the already high soil respiration fluxes characteristic of this and other tropical forests, warming resulted in a large amount of additional CO released to the atmosphere, with 6.5, 9.7, and 81.7 Mg CO-C ha yr more CO being released in the warmed plots compared to the control plots in the Lower, Mid, and Upper slope positions, respectively. Specifically, the additional soil CO released from the Mid slope was equivalent to the total annual net primary productivity (NPP) of a temperate grassland, and the Lower slope additional CO released was equivalent to the total annual NPP of a temperate deciduous forest. For the Upper slope, there is nothing on record that compares to the additional CO flux other than the conversion of a tropical peatland forest to an oil palm plantation.
The inclusion of warmed understory plant communities combined with warmer soils in this in situ warming experiment allows for an integrated exploration of above versus belowground controls on soil respiration responses. This inclusion is important because plants grown under warmer temperatures could alter carbon allocation to root biomass and/or root exudates that, in turn, may influence the contribution of root respiration to total soil respiration, as well as the available carbon and fuel for microbial processes. Acclimation of root-specific respiration under warming conditions could also shift the contribution of roots to total soil respiration. In addition, changes to the microbial community, microbial activity, changes to abiotic or geochemical conditions, or some combination may drive observed increases in soil respiration rates in response to warming. Understanding how plant and soil responses to warming individually and interactively affect soil respiration is critical for accurately modeling and forecasting feedbacks to carbon cycling and future climate. While spatially variable, live fine root biomass did not differ among warmed and control plots prior to starting the warming treatment; however, after just six months of warming, live fine root biomass was 32% lower in warmed relative to control plots (Supplementary Fig. S1; two-way ANOVA P = 0.04, F = 5.83; followed by Tukey's test P < 0.05). Further, live fine root biomass of the Upper slope plots did not differ significantly from live fine root biomass in the Lower or Mid slope topographic positions (two-way ANOVA P > 0.05), suggesting cross-slope patterns were not due to topographic differences in root biomass. We observed no significant acclimation of root-specific respiration in response to warming, but, when combined with the reduced root biomass, the contribution of roots to total respiration declined. Although total and extractable carbon pools weren't altered by warming, soil microbial biomass carbon did increase significantly, growing by over 50% in the warmed plots relative to the unwarmed control plots.
Taken together, these results suggest that the large increase in soil respiration rates in response to warming was due to an increase in microbial-derived CO efflux, while simultaneously experiencing a concomitant decline in the contribution of root-derived CO. Similarly, a belowground-only in situ soil warming experiment in Panama found a large increase in soil respiration in response to warming, with increases in CO efflux that were primarily derived from microbial sources with no change in root contribution. It is possible that the reduced allocation to roots by plants observed at our site was driven by warming of the aboveground vegetation, and not by warmer soils. These differential results highlight the potential for above- and belowground interactions to play an important role when considering the key drivers of the response of soil respiration fluxes to warmer temperatures in tropical ecosystems.
Given the especially high rates of soil CO efflux, we used several approaches to confirm that our observed emissions increases were indeed primarily driven by warming and not by stochastic spatial variability or measurement error (Supplementary Materials and "Methods" section). Whereas the Upper slope is clearly a "hotspot" for higher soil respiration rates, throughout the study, all plots exhibited extreme "hot moments" of soil respiration, where observed flux values were several times higher than the mean value (Fig. 3), which is characteristic of trace gas fluxes in this system. While there was high temporal variation throughout the year, we found no significant diurnal variation in soil respiration. We further conducted a spatial survey of 30 locations randomly selected outside of the plot locations across the 40 m × 60 m TRACE area to quantify the spatial variability of soil CO fluxes across the TRACE landscape (Supplementary Fig. S2). We unsurprisingly found variability across space but no evidence that the warmed plots were systematically located on landscape-level hot spots. Additionally, pre-treatment data showed no statistical differences in soil respiration rates between paired warmed and unwarmed plots prior to the initiation of warming (Supplementary Fig. S3), and running a generalized least squares model with and then without the Upper slope (i.e., the control-warming paired plots with particularly large differences in CO flux rates) did not change the finding that warmed plots had significantly higher soil respiration rates (Supplementary Table S1, Supplementary Materials and "Methods" section). Field sensitivity analyses failed to detect evidence that invertebrate or animal presence in the chamber could have produced such high respiration rates (Supplementary Materials and "Methods" section).
Despite sustained high rates of soil CO flux, we found no significant difference in total or available (i.e., extractable) carbon in the surface soils (0-10 cm) of the plots either before or after 6 months of warming. It remains unclear whether or for how long soil carbon stocks can be sustained at current levels despite such high CO flux rates, and it will be critical to determine the source of additional carbon that is fueling elevated soil respiration rates, particularly in the Upper Slope. For example, there could be greater carbon loss from deeper soils, which could be especially relevant for the Upper slope, where the soils are considerably deeper. Inputs from the declining fine root pool, altered litter layer decomposition, or a concentration of preferential flow paths in the more aerated upper slopes could also explain the high fluxes. There could also be abiotic factors influencing soil carbon availability via an increase in desorption reactions or a shift in the physicochemical environment. Ascertaining which of these potential sources of carbon are driving our responses requires further study. That said, regardless of the carbon source, our observed increases in soil respiration rates in response to warming are conspicuously greater than the 95% range of that observed for warming experiments in northern hemisphere forests (12-31% compared to our observed 42-204%), suggesting large temperature sensitivity of the tropical forest carbon cycle. Further, there were notable increases in the variability of soil respiration at higher temperatures (Fig. 1). Increased variability in soil CO flux under chronically warmer conditions has substantial implications for the equilibrium soil C stock size if highly variable emissions are not counteracted by similarly variable and large soil C inputs. These responses in both the mean and variation provide critical insight into quantifying tropical forest feedbacks to climate change in a range of conceptual and numerical models, and also contradict the supposition that tropical forests may be relatively insensitive to warming. In addition, our observed increase in microbial biomass carbon (~50%) was greater than the increase observed in the soil warming experiment in Panama or any northern hemisphere ecosystem, and is contrary to observations from tropical elevation studies, highlighting the exceptional value of additional data that elucidate in situ tropical forest responses to warming in global assessments.
Because temperature and moisture often co-vary, we also explored how the application of the warming treatment affected both soil temperature and moisture at the site. Soil temperatures increased similarly across the warmed plots at all topographic positions in response to experimental warming and were on average 3.99 °C warmer than control plots (Fig. 3a-c). In contrast, while soil volumetric water content (VWC) did not differ among the plots prior to starting the treatment, soil moisture responded very differently to warming depending on topographic position (Fig. 3d-f). Soils in the Lower and Mid slope plots were drier under the warming treatment (-0.03 g HO g soil [6.8% lower] for Lower slope and -0.06 g HO g soil [16% lower] for Mid slope between warmed and control plots, p < 0.001), while the soils of the Upper slope were surprisingly wetter in the warmed plots compared to control (+0.06 g HO g soil [18% higher], p < 0.001, Fig. 3). There is a range of explanations for the occurrence of greater soil moisture in the warmed Upper slope plot when compared with the control, including lower litter quality combined with extensive drying of the litter layer, which could slow decomposition and create a buffer against soil moisture loss. More work is needed to isolate the cause. Regardless, these data suggest that warming will, in part, regulate soil respiration through interactions with soil moisture, and that in situ experiments could further elucidate interactive controls with longer-term assessments of the connections among soil temperature, moisture, and biological activity.
Within the context of a warming climate, it is worth noting that lowland tropical forests experience a very narrow temperature range compared to all other terrestrial environments on Earth. In this system, a mean experimental increase of 4 °C nearly doubled the total ecosystem temperature range. Specifically, the mean diurnal temperatures of the control plots (~20-26 °C) overlap less than half of the range of the warmed plots (~22-32 °C, Figs. 2 and 4). Therefore, while the magnitude of warming that these soils experienced was in line with other ecosystem warming studies, the temperature increase relative to the temperature range was well beyond that imposed in other aboveground plus belowground field warming studies and pushed the warmed plots into a new climate space. The future climate warming expected in tropical forests will also have these large proportional increases in temperature.
Using diurnal averaged soil respiration rates, temperature, water content, and topographic position, we evaluated soil temperature and soil moisture effects on soil respiration using a generalized least squares modeling approach (Supplementary Materials and "Methods" section). Model results revealed that, regardless of warming treatment, the relationship between normalized respiration response and soil moisture on Lower and Mid slope positions was significantly negative (p < 0.001, Supplementary Table S1, Supplementary Fig. S4), while Upper slope positions, while also having a significantly negative relationship between soil respiration rate and moisture, exhibited a significantly positive interaction term between normalized respiration response and soil moisture (p < 0.05, Supplementary Table S1, Supplementary Fig. S4). In other words, soil respiration in the lower topographic positions declined during times when soils were wetter (i.e., after rainfall), consistent with decreased soil aeration, but this relationship was much weaker in Upper slope positions. The change in this statistical relationship was driven by occasional extreme respiration rates, which were observed in Upper slope plots during periods of high soil moisture values, regardless of warming status (Figs. 3 and 4). This points to more complex moisture and temperature interactions than expected for this system.
Over time, there is potential for organisms to adapt or acclimate to their new environment. Thus, we derived the Q of soil respiration for each of the plots (defined as the multiplicative change in soil respiration for every 10 °C increase in temperature, Fig. 4b, Supplementary Materials, and "Methods" section) to explore if organisms in the warmed plots had an altered relationship between warming and respiration rates. Q was significantly reduced with warming from a Q of 2.51 ± 1.23 (controls) to a Q of 0.71 ± 1.30 (warmed; Fig. 4b, p < 0.001). The Q values of the control plots fell within the range of the global mean of 1.6-3, while the mean Q of the warmed plots falls well below that range. These Q values indicate that soil respiration rate decreased per unit of increased temperature in the warmed plots (i.e., Q < 1), despite the fact that respiration rates themselves were higher in the warmed plots. In summary, while the sensitivity to temperature was reduced in response to warming, the higher respiration rates suggest a shift toward overall higher basal metabolic rates. Crucially, this finding suggests that the microbial community is exhibiting a highly plastic response to chronic +4 °C warming, with acclimatization or adaptation responses that both compensate for (reduced Q) and enhance (higher respiration rates) the response to temperature. This has important implications not only for the relationship between temperature and CO efflux for such carbon-rich forests, but also for the use of a single Q in Earth System Model forecasts of future climate.
Overall, the higher respiration rates in the warmed relative to control plots, independent of temperature sensitivity, coincided with a significantly higher soil microbial biomass in the warmed plots, which has been shown to correlate with higher soil respiration rates. While soil respiration in the warmed plots was higher across all temperatures (Figs. 2 and 3), the sensitivity (i.e., slope) of soil respiration in the warmed plots was flat to negative (Fig. 4b), suggesting that warming induced a systemic shift in function. Given the decline in root biomass observed in the warmed plots, we attribute this shift in function to heterotrophic rather than autotrophic responses. These functional shifts could include a change in microbial carbon use efficiency, a shift in the microbial community composition, and/or a change in the distribution of biotic activity vertically through the soil profile (e.g., driven by warming-induced changes to soil hydrology).
Contrary to the long-held paradigm that tropical forest responses to increased temperature will be relatively muted in their role in climate change feedbacks, we found large increases in soil respiration rates in response to in situ experimental warming that interacted in complex ways with soil moisture. Due to the amount of carbon released, these increased rates have substantial implications for forecasts of future climate at the global scale. While soil respiration showed signs of acclimation with respect to Q, the respiration rates were substantially higher in warmed relative to control plots (+42-205%) across all three topographic positions. This study contributes the second field-based observation from an in situ experiment that a warming climate may lead to large increases in CO fluxes from carbon-rich tropical soils, adding observations from a a-seasonal wet tropical forest on highly weathered, deep soils to the previous results from a seasonally dry tropical forest with relatively shallow soils. Taken together, the work demonstrates a potential for large carbon losses from tropical forest ecosystems in a warmer world, and highlights the immense value in evaluating the response of soil respiration to warmer temperatures across a range of tropical forested ecosystems, which include 30 Holdridge Life Zones spanning lowland dry deciduous to montane wet evergreen. Each of these forest types reflects an incredible level of diversity in forest structure, community composition, and soil conditions that make it unlikely these systems will exhibit a single response trajectory to a changing climate. While experimental warming resulted in a substantial increase in soil respiration at both this forest site and the site in Panama, the underlying controls, as well as the magnitude of the response, appear to be different. For example, our data suggested some acclimation/adaptation of respiration rates combined with a decrease in root respiration contributions to CO flux, whereas the seasonally dry forest in Panama found no significant effect of warming on root-derived CO flux and no signs of adaptation/acclimation. At both sites, large amounts of additional carbon continued to be released to the atmosphere from warmed plots with no signs of declining in the first 1-2 years after initiating warming, though how this will moderate with time remains to be seen. Nevertheless, understanding the underlying mechanisms driving the response of soil respiration to warmer temperatures is critical for accurate representation of tropical ecosystems in global models and assessing the magnitude and duration of feedback to future climate over the long term.