Dataset: Recruit qPCR data

This section includes methodology for several related treatments and experiments conducted in the course of research presented in Williamson et al. (2021).   Each BCO-DMO page provides access to one of these datasets, see the "Related Datasets" section for access to all related datasets.

Parents of recruits used in this study were located on Horseshoe Reef in Key Largo, FL at 5 - 8 meters depth.

Symbiont uptake experiment   (see related dataset "Recruit survivorship and symbiont acquisition" https://www.bco-dmo.org/dataset/920853)

Throughout symbiont uptake period, a dissecting microscope was used to count the number of: (1) surviving recruits and (2) recruits visibly infected with symbionts. When infection was first observed in all aquaria (day 21) and again after 60 days in their treatments, three to five recruits from each aquarium were sacrificed using a razor blade. To standardize sampling, only solitary polyps not clumped with others were sacrificed. Sacrificed recruits were placed in individual 1.5-mL microcentrifuge tubes with 200 μL of 1% SDS + DNAB and incubated at 65 °C for one hour. Genomic DNA was extracted from SDS archives following modified organic extraction methods (Rowan and Powers 1991; Baker and Cunning 2016).

Growth and symbiont density (see related dataset "Recruit area measurements" https://www.bco-dmo.org/dataset/920846)

Five months (150 days) after settlement, a random sample of recruits from each of the four original treatments was photographed under a dissecting microscope with QCapture Suite Plus. Only solitary polyps were photographed to maximize accuracy of area measurements. ImageJ was used to calculate recruit skeletal area in mm2. A subset of the photographed recruits was then sacrificed to measure symbiont identity and density at the time of growth measurements.

Heat stress experiment (see related dataset "Heat stress experiment data" https://www.bco-dmo.org/dataset/920837)

A subset of plugs with each symbiont type were placed into new aquaria, where temperature was increased from 22 to 28 °C over six days, and then to 32 °C over 48 h. At this point, all recruits (n = 66) were infected with symbionts. About half the recruits (n = 32) were pre-exposed to mild heat stress (reared at 31 °C during Experiment 1), while the other half (n = 34) were naïve to heat stress (reared at 29 °C). Temperature was maintained at 32 °C for ten days, then raised to 33 °C for ten days, then raised to 34 °C for 40 days. Every two to five days, recruits were scored as “healthy,” “pale,” “bleached,” or “dead” using a fluorescence microscope. After 60 days, all remaining living recruits were sacrificed for symbiont community analysis. 

Recruit PCR (qPCR) (data included in this dataset)

Quantitative PCR (qPCR) assays were used to identify Symbiodiniaceae to genus level and quantify symbiont-to-host (S:H) cell ratios for each recruit sampled. Since O. faveolata commonly hosts members of Symbiodinium, Breviolum, Cladocopium, and Durusdinium (Kemp et al. 2015), assays targeting specific actin loci for each genus were performed using a QuantStudio 3 Real-Time PCR Instrument (Applied Biosystems, USA). Assays for O. faveolata, Symbiodinium, and Breviolum followed reactions described in Cunning and Baker (2013), whereas Cladocopium and Durusdinium assays were multiplexed as described in Cunning et al. (2015).   
Instruments: For qPCR data, assays targeting specific actin loci for each genus were performed using a QuantStudio 3 Real-Time PCR Instrument (Applied Biosystems, USA).