Burrowing animals can profoundly affect the biological structure and ecosystem functions of their environments. For instance, burrowing crabs in soft-sediment coastal ecosystems, like salt marshes, can increase sediment deposition and facilitate sediment homogenization and turnover, with potential impacts to sediment biogeochemistry. However, the relative importance and overall impacts of burrowing crabs on sediment dynamics can vary considerably between, and within, salt marsh habitats. Past studies have suggested that sediment properties can influence how burrowing crabs will affect edaphic conditions in salt marshes, but these studies often assume homogenous sediment conditions and fail to consider how marsh sediment properties change with depth. Here, we conducted a series of field surveys in three tidal salt marshes with variable sediment properties to understand if salt marsh vertical sediment profiles can help predict the nature of burrowing crab-sediment relationships. We found that burrowing crabs homogenize sediments in all marshes, but their effects on sediment homogenization and edaphic conditions were greater in marshes with highly stratified vertical sediment profiles. Our study suggests that understanding the vertical sediment profile of a salt marsh may provide critical insights into how crab burrowing may influence the edaphic conditions and physical characteristics of marsh surface sediments — especially in restored, created, and managed salt marshes.

Site Descriptions

            To evaluate if marsh vertical sediment profile stratification informs burrowing crab effects on sediment edaphic conditions, we used a complex of two, 34-year-old constructed marshes and one natural marsh (~2,140 ± 15 - 2,290 ± 20 years old; Smyth 2020) along the West Fowl River in southern Alabama, USA (CON-1: 30°22’02.3” N, 88°09’06.8” W; CON-2: 30°22’03.5” N, 88°09’02.6” W; NAT: 30°22’02.5” N, 88°09’37.2” W). The natural marsh (hereafter, NAT) is approximately 1km from the two constructed sites (hereafter, CON-1 and CON-2), which are hydrologically connected to NAT by a dredged tidal channel. All three sites experience diurnal microtides with an amplitude of approximately 0.26 m (Smyth 2020) and range in elevation from 0.26-0.36 NAVD88 (Ledford and others 2021).

            The plant community at each marsh is dominated by Juncus roemerianus (hereafter, Juncus), with smaller patches of Spartina alterniflora and Distichlis spicata along creek banks and in the high marsh, respectively. The aboveground biomass at all three sites is comparable; however, CON-1 and CON-2 have lower belowground biomass, porewater and extractable nutrient concentrations, and organic matter content than NAT (Ledford and others 2020, Tatariw and others 2021). These differences likely result from the mitigation methods employed to create CON-1 and CON-2, which were converted to tidal marshes by harvesting pine savanna habitat and excavating topsoil down to a clay layer that was parallel with the water table (Vittor and others  1987). Today, this clay layer is still a prominent feature of CON-1 and CON-2 that can be found ~10-15 cm below the surface at both sites, under a layer of organic material, and results in a distinctly stratified vertical sediment profile (Figs 1a-b, Griffin Wood 2020).

Burrowing crabs are present at all three sites in similar densities [CON-1: 14 ± 3 burrows m^-2 (mean ± 1SE); CON-2: 16.5 ± 4 burrows m^-2; NAT: 16 ± 3 burrows m^-2), with their communities being primarily comprised of fiddler crabs (Minuca longisignalis, M. minax, Uca panacea, and U. spinicarpa). However, we have observed five Sesaema reticulatum individuals and several large burrows (2–3 cm diameter) — indicative of S. reticulatum activity— in the Juncus and D. spicata zones at NAT (Rinehart and Dybiec personal observations).

Characteristics of excavated sediment.

            We collected excavated sediments from areas within 2 cm of crab burrow openings at CON-1 (n = 4), CON-2 (n = 9) and NAT (n = 8). These sediments were returned to the laboratory where we dried them in an oven at 60˚ C to a constant weight to obtain bulk density. We ground the dried sediments with a mortar and pestle then ashed them in a muffle furnace (six hours at 550˚ C) to quantify the sediment organic matter (SOM) content via loss-on-ignition.

Following our initial evaluation of excavated sediment characteristics, we wanted to test if burrowing crabs were excavating clays from the dominant clay layer (~10–15 cm deep) at CON-1 and CON-2 (Griffin Wood 2020).  To do this, we collected additional excavated sediments from areas adjacent to crab burrow openings at all three marshes (n = 3 marsh^-1). The excavated sediments were then incubated for 12 hours in 10 ml of deionized (DI) water at ambient temperature. We also incubated three samples of only 10 ml of DI water (no sediment added) at ambient temperature to serve as a control. Following incubation, we used a Thermo Orion Star pH meter (model A211) to measure the pH of the sediment-water slurries and our control DI water samples. We then calculated the difference between the pH of our sediment-water slurries and the pH of our DI water controls. We implemented this approach because suspended clay particles can acidify water. Thus, slurries made from clay-rich sediments should acidify the DI water more than other sediment types (Keller and Matlack 1990).

Characteristics of deposited sediment.

            We deployed 10 cm long, 2.5 cm internal diameter PVC crab burrow mimics along large (> 2m wide) tidal creeks at CON-1 and NAT in September 2021. Crab burrow mimics were sealed at one end with PVC socket caps. At each marsh, we deployed seven burrow mimics at 25 m intervals starting at the mouth of the tidal creek and running 150 m along the edge of the creek into the marsh. The crab burrow mimics were inserted vertically into the marsh sediments (sealed end down) until the open end of the mimic was flush with the sediment surface. We placed all crab burrow mimics ~0.5 m in from the marsh-tidal creek edge (horizontal distance) in stands of Juncus and S. alterniflora. All inserted mimics were filled with deionized water to prevent sediment deposition from rapidly flushing water (Wang and others 2010).

We collected the contents of each crab burrow mimic in October 2021 after one month. The collected sediments were transported in plastic bottles to the University of Alabama where the deposited sediments were oven dried at 60˚ C to a constant weight to obtain the dry weight burrow^-1. We then ashed the dried sediment in a muffle furnace (6 hours at 550˚ C) to estimate SOM via loss-on-ignition. We standardized total sediment deposition and SOM deposition by calculating the rate of total sediment deposition and SOM deposition crab burrow mimic^-1 day^-1.

Crab burrow density and habitat characteristic survey.

            In October 2021, we conducted an observational survey at all three marsh sites in the high Juncus zone to evaluate the relationships between crab burrow density, Juncus traits, porewater biogeochemistry, and sediment properties. Specifically, at each site we identified four, 0.25 m² plots with high crab burrow densities (i.e., ≥ 16 burrows m^-2; hereafter High Crab plots) and four, 0.25 m² plots with low crab burrow densities (i.e., ≤ 12 burrows m^-2; hereafter Low Crab plots). All plots were at least 0.5 m apart, but we did cluster all plots in the same habitat to minimize natural variability in marsh structure. We documented the total number of crab burrows and the diameter of every crab burrow in each plot at the time of sampling.

Juncus traits. We estimated percent cover, stem density, and mean stem height of Juncus in every plot. Juncus stem density was estimated by counting the number of individual stems within a 0.1 m² quadrat that was placed in the center of each plot. We then extrapolated these counts to the entire 0.25 m² plot. We estimated the mean Juncus stem height by measuring five, randomly-selected Juncus stems in each plot.

Biogeochemistry. We collected pore water samples from the center of each plot. Pore water was collected by inserting a 30 cm-long acrylic tube (4.7 mm diameter) with 1.5 mm holes drilled along the bottom 3 cm vertically into the sediment to a depth of 5 cm (McKee and others 1988). From each plot, we pulled two separate samples that were filtered in the field (0.45 μm, VWR) into vials and stored on ice. The first sample was pulled to determine nutrient concentrations and was frozen in the laboratory until analyses. We used colorimetric analyses for phosphate (PO₄^3-) and NOx (NO₃^- + NO₂^-) using a UV-vis spectrophotometer (Thermo Scientific GENESYS 50; Grasshof and others 1983, Schnetger and Lehners 2014). Additionally, we used fluorometric analysis for ammonium (NH₄⁺) using a fluorometer (Turner Designs 7200-02) equipped with a color-dissolved organic matter (CDOM)/ NH₄ UV module (Holmes and others 1999). The second porewater sample was transported to the laboratory where we used a Thermo Orion Star pH meter (model A211) and a YSI conductivity and salinity instrument (model 3100) to measure pH and salinity, respectively.

Sediment properties. To understand potential relationships between crab burrows and sediment conditions, we collected three, 10 cm deep sediment cores using a 5 cm diameter Russian peat corer. We standardized the locations of each core within every plot to minimize bias. Specifically, cores were all separated by 10 cm and formed a triangle in the center of the plot. All collected cores were then sub-sectioned in the field at 2.5 cm intervals and bagged. Sub-sections were weighed to obtain a wet mass (used in calculations of water content) and then oven dried at 60˚ C to a constant weight to obtain bulk density. Once dried, samples were ground with a mortar and pestle and then ashed in a muffle furnace (six hours at 550˚ C) to estimate SOM via loss-on-ignition. Additionally, we pooled sediment from replicate cores within each plot (n = 3 cores plot-1) and sent material from the 0–2.5 cm and 2.5–5 cm core subsections for analysis of %C and %N at the Alabama Stable Isotope Laboratory (ASIL). We then used %C and %N to calculate the mean C and N stocks in the top 0–2.5 and 2.5–5 cm of every plot.

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