Effects of Crumb Rubber Leachate across Ecosystems
Description
Created one leachate for planaria and worms. Using the same rubber crumbs, made another leachate for marine dinoflagellates. Measured a number of biological outcomes
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Planaria Swimming Speed Apparatus Opaque plastic chamber (200 × 140 × 60 mm) covered with one-way tint film. A 1,000-lumen flashlight projected a 31.65 mm light circle onto a 90 mm Petri dish centered in the arena, positioned 152 mm below the light. Graph paper (2.5 mm grid) calibrated distance. Individual planaria were placed in the dish center; time to exit the light ring was recorded to calculate swimming speed. S3. Planaria Experiments without Preconditioning Duration: 8 days. Sample size: 52 (26 control, 26 leachate). Baseline: body length, eyespot distance, swimming speed. Procedure: worms fed 0.2 g earthworm, bisected, heads discarded, tails placed in jars with pond water vs. leachate (300 mL). Endpoints: survivorship, regenerated length, eyespot distance, swimming speed. S4. Planaria Experiments with Preconditioning Duration: 14 days. Sample: 60 (30 control, 30 leachate). Preconditioning: 7 days in pond water vs. leachate (300 mL). Bisection into head/tail segments reseeded into same treatment. Endpoints: survivorship, body length, regeneration of both segments after 7 days. S5. Earthworm Enclosures and Soil Adults of E. fetida with clitellum, 0.19–0.22 g, in OMRI-certified compost (350 g + 75 mL water/leachate). Enclosures (26.5 × 6.9 × 24.3 cm; viewing width 2.6 cm) housed one worm each; eight 7-day trials yielded 24 worms per treatment. S6. Earthworm Camera Setup Soil displacement recorded using Arducam OV5647 sensors on Raspberry Pi 3b+, positioned 70 cm away, programmed in Python to capture images every 10 min, stored on external drives. S7. Earthworm Image Processing Images cropped to enclosure face; pixel differences converted to area (1 pixel = 0.06522 cm²). Each trial produced 675–1,084 frames (mean 989.8 ± 54.7). Data from 39 worms (21 control, 18 leachate). To avoid pseudoreplication, soil displacement was averaged per worm and compared with one-way ANOVA. S8. Earthworm Body Mass Twelve kg soil distributed into 120 jars (90 g/jar). Controls (N=30) received 12 mL water; treatments (N=30) 12 mL leachate. Seventy worms rested overnight, weighed, and placed one per jar. After 14 days, worms reweighed to calculate mass change. S9. Earthworm Stress Test Following Pochron et al., worms placed individually in Petri dishes (35°C, 250 μmol m⁻² s⁻¹ light). Dishes inspected at 5-min intervals; unresponsive worms scored as dead. Survival time recorded in minutes. S10. Dinoflagellates P. fusiformis cultures (PyroFarms) maintained in Conviron chamber (12:12 light-dark, 273 µmol·m⁻²·s⁻¹, 21°C). Nutrients added biweekly. Thirty Petri dishes prepared with 5 mL culture. Treatments: control (0%), low (2.5%), high (9.75%) leachate. Dishes stirred, capped, and returned to chamber. S12. Dinoflagellate Population Counts taken nightly at 2000 h for 7 days. Dishes photographed with iPhone 15 Pro Max against black background. Images processed in ImageJ (8-bit, thresholded, “Analyze Particles”) to quantify individuals.
Institutions
- Stony Brook University
Categories
Funders
- U.S. National Science FoundationUnited States