The difference in soil carbon and nitrogen content between ant nests and control sites varied with elevation. The extent to which control soil was enriched with nitrogen was significantly positively correlated with elevation (and the climatic factors that covary with elevation (Table S2)) across both mountain ranges, and the extent to which control soil was enriched with carbon was significantly positively correlated with elevation in the Sierra Nevada (Table S3). At higher elevations, nest soil had higher amounts of carbon and nitrogen (Fig. 1; Table 1) than adjacent control soil samples. This pattern was consistent with our expectations. However, at lower elevations, nest soil tended to have reduced carbon and nitrogen relative to controls (Table 1), which seemed counter-intuitive. The unexpected pattern of nutrient depletion in low elevation ant nests led us to consider possible limitations of studies investigating soil properties in ant nests. While moisture of control soil significantly increased with elevation in the Sierra Nevada, there was no significant pattern along the elevational gradient of the San Jacinto Mountains (Table S3). We observed no significant pattern of soil moisture in ant nests compared with controls (Table 1). Here, we describe and interpret the patterns that we found in our study, and then we consider potential sources of noise in our sampling approach and suggest steps that could be taken to reduce variance not associated with the biological research questions in future studies.
Table 1 Linear mixed-effects model results Intriguingly, data from both ranges showed a trend of low elevation nests containing soil depleted in carbon and nitrogen compared to control soils, whereas high elevation nest soils were enriched in carbon and nitrogen compared to control soils. While these consistent trends emerged, the magnitude of ant nest effects on soil varied between the mountain ranges and transects within range. The relative increase or decrease in carbon and nitrogen levels in ant nest soil compared to control soil was up to an order of magnitude greater in the F. sibylla nests in the Sierra Nevada compared to the F. francoeuri nests in the San Jacinto mountains. Thus, our results highlight the complexity of generalizing the effects of ant nests on soil properties across elevations and landscapes that may vary in water drainage, soil type, and climate.
We were unsurprised to find higher soil carbon and nitrogen in nest soil compared with control soil in many sites. This result is consistent with studies that have found that Formica nests contain higher amounts of nutrients and organic matter (Kristiansen and Amelung 2001; Drager et al. 2016), which suggests a large community of soil microbes in nest soils that would encourage greater rates of decomposition and nitrogen mineralization (as has been found in harvester ants (Wagner and Jones 2006)). We also expected differences in the effects of nests across elevation gradients, as landscape-level processes affect the distribution of soil nutrients. Our results align with studies that have found a differential effect of ants on soil processes and function across various climates and land-use regimes (Folgarait 1998).
The result that nest soils showed nutrient depletion compared to control soil in many lower-elevation sites was more surprising. We identify three alternative, biologically relevant hypotheses that may explain this counter-intuitive finding. The pattern could result from soil texture and permeability; generally, more sandy soil is nutrient-poor because nutrients tend to leach through (e.g., Ge et al. 2019). Observationally, higher-elevation sites appeared to have more compact soil, which may limit the depth of nests and resulting nutrient accumulation. Conversely, ants may excavate deeper in less-compact soil, bringing vegetation and prey items well below the soil surface and bringing nutrient-poor soil to the surface. Our complementary research revealed that ant nest depth depends on surface temperature in laboratory-based groups of Formica podzolica workers (Sankovitz and Purcell 2021). Hence, nest architecture may vary consistently with elevation, causing standard soil sampling approaches, like the one used here, to yield samples from different portions of nests at different points along elevation gradients. To investigate the possibility that our standardized sampling approach systematically collected soil from different portions of nests across sites, we revisited one low and one high elevation site in the San Jacinto Mountains and took soil cores 35 cm deep. However, we found no correlation of carbon or nitrogen with soil depth in these preliminary samples (Table S4).
Several issues with our sampling protocol could yield significant differences in soil metrics that are not mediated by biologically interesting effects of ant nests on soil and should be adjusted in future studies. First, even if thoroughly homogenized, the quantity of soil used by the elemental analyzer is minuscule—mere grams—making obtaining a representative sample difficult. In this study, we took measurements from a single soil sample within the nest and a second one near the nest (nest and control sample, respectively). Comparing multiple nest and control samples for each nest could provide a more representative measure. Second, ant nests complicate this process further because the bioturbation and addition of organic matter by ants, in addition to variation in nest architecture, create a plot of soil that is highly variable in all three dimensions. We took samples from a standard 10 cm below the soil surface, following established protocols (Nkem et al. 2000; Wagner et al. 2004), but our concurrent study indicated that nest depth could strongly covary with elevation and temperature (Sankovitz and Purcell 2021). Hence, we may have breached the nest in some samples and not others, with a bias associated with the average temperature of each locality. This sampling approach might be accurate for ants that excavate superficial nests, like Argentine ants, but both sampling soil cores and investigating the depth of the highest nest chambers may be necessary to ensure that equivalent samples are compared in ant species that dig deeper nests. Due to these potential limitations, the results presented here should be considered intriguing patterns that hint at ecologically significant processes at play but should be investigated further with sampling encompassing a broader spectrum of possible nest architecture.
Much research has focused on the effects of ants on ecosystem processes within single populations (De Bruyn and Conacher 1990; MacMahon et al. 2000; Decaëns et al. 2002; Folgarait et al. 2002), but few studies have examined how these effects vary across environmental gradients. Both our original samples and the follow-up samples taken using a soil core contained significant variability, demonstrating the difficulty of taking a representative soil sample inside ant nests. Despite our efforts to homogenize soil samples, analyzing mere grams of soil from an entire nest probably does not capture a representative profile of nest soil. We recommend that future research on ant soil interactions sample soil from multiple locations within an ant nest, both parallel and perpendicular to the soil surface. Overall, our results suggest that the magnitude of ant nest effects on nutrients is likely influenced by factors that vary with elevation, like climate and soil type. Future research on ant ecosystem engineering should work to generalize how ants affect soil properties in distinct geological and climatic zones.