Replacing long-lived, rarely disturbed vegetation with short-lived, frequently disturbed vegetation is a widespread phenomenon in the Anthropocene that can influence ecosystem functioning and soil development by reducing the abundance of deep roots. We explore how sources and fate of soil CO2 vary with organic substrate source, abundance of respiring biota (i.e., roots and soil microbes), season, and soil depth. We quantified multiple isotopic signatures of CO2 (δ13C, Δ14C, δ18O) as well as concentrations and δ18O of free O2 in the upper 5 m of soil at sites where root abundances and soil organic C have been previously quantified: in late-successional forests, cultivated fields, and ~ 80 y old regenerating pine forests growing on previously cultivated land. We hypothesized that soil CO2sources would vary across soil depth and land cover, reflecting varying abundances of organic substrates, and seasonally as the dominance of root vs. microbial CO2 production changes through the year. δ13C–CO2 revealed respiration of C4-derived substrates in cultivated fields particularly during the growing season. This effect was not evident in soils of regenerating pine or older hardwood forests, suggesting that ~ 80 y of pine inputs to reforested soils have been sufficient to dominate microbial substrate selection over any remnant, historic agricultural C4 inputs. Δ14C–CO2 diverged by land use at 3 and 5 m, indicating that more recently-produced photosynthate is available for mineralization in forests compared to cultivated plots, and in late-successional forests compared to regenerating pine forests. At 1.5, 3, and 5 m in forested plots we observed evidence of respiratory demands on soil pore space O2. In these soils, we observed declines in [O2] compared to other depths and to the agricultural plots and concurrent increases in δ18O of free O2, consistent with the idea that roots and heterotrophic soil microbes are more active where photosynthate is more available. The δ18O–CO2 values, a likely proxy for δ18O of soil porewater, exhibited 18O enrichment during the winter, when many sampling wells were flooded, compared to growing season values. These data suggest an isotopically-distinct and laterally-flowing source of CO2-laden porewater during winter months. Combined, these datasets document how ~ 80 y of forest regeneration can provide sufficient C inputs to mask any microbial mineralization of decades-old organic inputs, but belowground C inputs still lag those of late successional forests. We also infer that lateral and vertical flows of water can serve as a sink for biotically-generated CO2, and that where deep soil [CO2] is lower due to lower root and microbial activities, production of carbonic acid is also diminished. Where reaction rates are weathering limited, a paucity of deep roots imposed by anthropogenic land cover change thus may limit the production of this agent of soil development and the C sink represented by the silicate weathering it can promote. The data suggest deep and persistent effects of the loss of deeply rooted long-lived vegetation on deep soil C storage and transformations that promote acid-dissolution weathering reactions that help form soil itself.
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