Seedling growth responses to experimental soil warming in a tropical forest, Panama, 2016-2022

Data were collected over the period 2016-2019, with seedling census each 3-months and soil parameters determined monthly.

Seedling height was determined by measuring from the soil surface to the shoot apex (apical meristem), using a tape measure. Light was determined by PPFD (Photosynthetic Photon Flux Density; µmol m-2 s-1), measured using a LI-250A light meter (LI-COR, Inc., Lincoln, NE, USA). PPFD was determined monthly above each individual seedling, both on an overcast day and a clear day for each month, providing two metrics of PPFD representing the average conditions on both an overcast and clear day. Light saturated photosynthesis (Asat) was determined using a portable photosynthetic system (LI-6400, LI-COR, Inc., Lincoln, NE, USA) fitted with a standard 2 × 3 cm leaf cuvette with a red-blue LED light source (6400-02B)) and a CO2 mixer (6400-01, LI-COR, Inc., Lincoln, NE, USA).

To assess whether soil warming affected nutrient availability to plants, we determined soil nitrogen (N) and phosphorus (P) mineralisation using in situ mixed-bed ion exchange resin membranes (Dowex Marathon Mr-3, Sigma-Aldrich). Resin was prepared by rinsing in 0.5 M HCl for 2 hours and then rinsed with DI H2O. Resin was placed inside nylon bags (5 g resin in each) and installed at 5 cm soil depth in systematically distributed locations in each plot (n = 5 per plot). Resin bags were deployed monthly for three years, and following collection were stored at 4oC until extraction. Extraction was performed using 0.5 M HCl, and PO4, NH4 and NO3, were determined using a Lachat Quickchem 8500 flow injection analyzer (Hach Co., Loveland, CO). Extractable PO4 (total mineralised P) and NH4 and NO3 (total mineralized N) were calculated from the difference between extracted N from resin deployed in the field and resin not deployed (blanks) and expressed as extractable PO4-P, NH4‐N and NO3‐N per g resin per day.

We determined relative growth rates (RGR) using a first order power-law model, which is especially suitable for the early stages of seedling growth when growth is rapid and non-asymptotic, but slowing over time:

dh / dt = tβ

where dh/dt is height increment over time (or RGR). To represent time in our models we used ‘relative month’ where the warming treatment began at relative month = 0. To calculate the power of the relationship (β), we used log-log plots. We determined RGR for each individual seedling (n = 30, 6 individuals per 5 plots; and for 6 species), by calculating the slope of log [seedling height]/log [time in months]. We used the power rate increase β to assess treatment effects on RGR.

All data were subject to quality control review prior to statistical analyses in R, by assessing outliers, to ensure correct identification of seedlings and correct use of blanks/standards for determination of final values.