Surf smelt accelerate usage of endogenous energy reserves under climate change

elevated pCO2 on Y:E (Table 2). Embryo heart rates were not significantly affected by tempera- ture or pCO2 stressors (Table 3). Surf smelt embryo heart rates in ambient temperature treat- ments were similar between pCO2 treatments. As temperature increased, the average heartrate increased between 13˚C and 18˚C at elevated pCO2 and a decreased at ambient pCO2 but vari- ability was high, and these patterns were not significant (S1 Fig). Larvae yolk and oil globule usage Larval yolk size decreased over time in all treatments with day having a significant negative effect on larval yolk size (Table 3; Fig 2). Temperature also had a significant, negative effect on Fig 1. Embryo yolk usage. Proportion of yolk area to total egg area (Y:E) of surf smelt embryos (n = 216/day) that were submerged in treatment water for 13 days. Each point is an average measurement representative of the temperature treatment, pCO2 treatment (points offset for visualization), and day with standard error shown as whiskers. Day, pCO2 and Temperature were significant factors predicting embryo yolk usage. https://doi.org/10.1371/journal.pone.0270491.g001 Table 3. Linear mixed effect model results. Value Std. Error df t-value p-value Embryo heart rate Intercept 11.8181 0.9540 65 12.3879 <0.0000 Temperature -0.0252 0.0230 26 -1.0925 0.2846 pCO2 0.0008 0.0005 65 1.8712 0.0657 Larval yolk sac size Intercept 0.3389 0.0270 383 12.5504 <0.0000 Day -0.0283 0.0019 383 -14.3675 <0.0000 pCO2 0.000004 <0.0000 16 0.6234 0.5418 Temperature -0.0111 0.0017 383 -6.4106 <0.0000

Larval oil globule size Intercept 0.0823 0.0071 383 11.5417 <0.0000 Day -0.0075 0.0007 383 -10.8844 <0.0000 pCO2 -0.000001 <0.0000 16 -0.8062 0.4319 Temperature -0.0018 0.0005 383 -4.1379 <0.0000 Summary of linear mixed effect models examining the effect of day, temperature, and pCO2 treatment level on multiple parameter measurements. https://doi.org/10.1371/journal.pone.0270491.t003 PLOS ONE Surf smelt response to climate co-stressors PLOS ONE | https://doi.org/10.1371/journal.pone.0270491 June 27, 2022 7 / 14