Aim

The mutualistic relationship between anemones and anemonefishes is one of the most iconic examples of symbiosis. However, while anemonefishes have been extensively studied in terms of genetic connectivity, such information is lacking entirely for host sea anemones. Here, we provide the first information on the broad-scale population structure and phylogeographic patterns of three species of host sea anemone, Heteractis magnifica, Stichodactyla mertensii, and Entacmaea quadricolor. We evaluate if there is concordance in genetic structure across several distinct biogeographic areas within the Indo-Pacific region and to what extent the observed patterns may concur with those found for anemonefishes. 

Location

Indo-Pacific, including the Red Sea.

Taxon

Heteractis magnifica, Stichodactyla mertensii, and Entacmaea quadricolor

Methods

Microsatellite markers and a combination of statistical methods includingBayesian clustering, Isolation by Distance (IBD), Analysis of Molecular Variance (AMOVA), and Principal Components Analysis (PCA) were used to determine population structure. The congruence among distance matrices method (CADM) was used to assess similarity in spatial genetic patterns among species.

Results

Significant population structure was identified in the three host anemone species. Each species is likely composed of at least two genetic clusters corresponding to two biogeographic regions, the Red Sea and the rest of the Indo-Pacific. Two of the three anemone species seem to be experiencing admixture where the two main clusters overlap (the Maldives). IBD analyses in the Red Sea revealed differences in gene flow among species, suggesting more limited dispersal potential for E. quadricolorthan forS. mertensiiand H. magnifica. Clonality is documented in S. mertensii for the first time.

Main conclusions

This research documents the genetic population structure for three ecologically important host sea anemones across the Indo-Pacific and provides valuable insights regarding their biogeography and evolution. Specifically, we found high levels of genetic divergence between populations across different biogeographic regions, suggesting different evolutionary lineages within species. At the same time, common geographic overlap of population structures suggests similar evolutionary histories among all three species. Interestingly, the observed patterns are congruent to some extent with structure reported for several anemonefish species, reflecting their close ecological association.

The final datasets consisted of multilocus genotypes for 205 H. magnifica individuals, 122 S. mertensiiindividuals, and 249 E. quadricolor individuals. For each species two sheets are provided. One that includes all specimens including clones (WITH CLONES) and one for which only one representative of each clone was included (NO CLONES). The genotypes for each microsatellite are given in base pairs. 

A tentacle of each specimens was collected using dissecting scissors and forceps whilst SCUBA diving at 42 sites across the Indo-Pacific and Red Sea. Specimens were placed in 2 ml vials and stored in 96% ethanol. 

All forward sequences were labelled with a fluorescent dye (6-FAM, NED, PET, VIC). PCR conditions followed the Qiagen PCR Multiplex kit protocol with modifications as in Gatins et al. (2018); a total of 10 µL was used for each individual reaction mix, including 5 µL of Multiplex PCR MasterMix (Qiagen), 1 µL of primers (2 µM; see Table S1, Supporting information), 3.3 µL of water and 0.7 µL DNA (50-150 ng/µL). The thermocycler conditions for PCR amplifications were: 95 ºC for 15 min, then 25 cycles of 94 ºC for 30 s, annealing at a locus-specific temperature (57/60 ºC, see Table S1, Supporting information) for 90 s, and an extension at 72 °C for 60 s, with a final extension set at 60 °C for 30 min. Further details regarding microsatellite and PCR protocols can be found in Gatins et al. (2018). Final PCR products of 10 µL were diluted with 130 µL MilliQ water before being sent for fragment size analysis using a GeneScan 500-LIZ size standard and an ABI 3730xl genetic analyser (Applied Biosystems, USA) in the Biosciences CORE laboratory at King Abdullah University of Science and Technology, Saudi Arabia. Genotyping was completed using Geneious v. 8.1.6 

Gatins, R., Saenz-Agudelo, P., Scott, A., & Berumen, M. L. (2018). Development and characterization of new polymorphic microsatellite markers in four sea anemones: Entacmaea quadricolor, Heteractis magnifica, Stichodactyla gigantea, and Stichodactyla mertensii. Marine Biodiversity, 48, 1283–1290. https://doi.org/10.1007/s12526-016-0576-0