Adult culture and spawning

Adult Nematostella vectensis sea anemones were collected from a salt marsh in Brigantine, New Jersey in the fall of 2020, and spawning was induced. Larvae (n = 1 cohort with mixed parentage) were then cultured to the planula stage (3 d post-fertilization) for experimentation. An aquarium population of adult Galaxea fascicularis colonies (n = 9 females, 10 males) spawned during August 2023 at Carnegie Science (Baltimore, MD, USA), yielding a cohort of planulae (n = 1 cohort with mixed parentage) that were used in experiments within 48 h. Adult Porites astreoides colones (n = 20) were collected in Bermuda (32°22′13″N, 64°44′27″W) during July 2023. Brooded planulae (n = 4 cohorts) were collected and maintained in culture for ~ 48 h.

At the time of use in experimentation, larvae from all 3 species were in the planula stage and would become competent to settle within 72–96 h (Hand and Uhlinger 1992; Goodbody-Gringley et al. 2018; Wei et al. 2023). Artificial seawater was used for culturing and experimentation for N. vectensis and G. fascicularis, while flow-through, natural seawater facilities at the Bermuda Institute for Ocean Science were used for P. astreoides.

Dissoved Oxygen Treatments

Stage-matched, swimming planulae of N. vectensis, G. fascicularis, and P. astreoides (N = 1,200–2,400 larvae species-1) were divided into six replicate groups (three normoxia/control and three hypoxia) and exposed to 6 h of normoxia (dissolved oxygen (DO) = 6.8–8.69 mg L-1) or severe hypoxia (DO = 1.58–1.8 mg L-1; seawater deoxygenated using N2 gas) inside sealed glass jars (500 mL) overnight from 21:00 h to 03:00 h the following day. The jars were placed at the ambient culture temperature for each species (N. vectensis: 18°C; G. fascicularis: 27°C; P. astreoides: 28°C). At the end of the treatment period, the jars were uncapped and groups of 20–30 larvae (N = 60–90 larvae treatment-1 species-1) were transferred to 1.5 mL tubes without seawater for storage at -80°C until processing

Measurements

Detailed methods can be found in the associated publication (Glass and Barott 2025).

Ash-free dry weight (AFDW; i.e., organic biomass) of larvae and juveniles was quantified from burned, dried tissue.
Chlorophyll content within algal endosymbionts of larvae and juveniles was quantified via spectroscopy.
Heat tolerance of P. astreoides larval symbionts was quantified via changes in photochemical yield (collected with a pulse-amplitude-modulation (PAM) fluorometer) over time at 36°C.
Heat tolerance of larvae was quantified via visual observation of survival over time at 36°C. Photochemical yield of P. astreoides larvae and juveniles as quantified using a pulse-amplitude-modulation (PAM) fluorometer.
Metabolic rates of larvae and juveniles was quantified via respirometry.
Protein content larvae and juveniles was quantified via Bradford assays.
Larval settlement rates were quantified via brightfield microscopy.
Larval and juvenile sizes were quantified via brightfield microscopy.
Larval swimming behavior immediately following the oxygen treatments was quantified via imaging using iPhone 14 (Apple).
Larval and juvenile symbiont density was quantified via flow cytometry.