Multiplexed ISSR genotyping by sequencing distinguishes two precious coral species (Anthozoa: Octocorallia: Coralliidae) that share a mitochondrial haplotype

Abstract

Background: Precious corals known as coralliid corals (Anthozoa: Octocorallia) play an important role in increasing the biodiversity of the deep sea. Currently, these corals are highly threatened because of overfishing that has been brought on by an increased demand and elevated prices for them.The deep sea precious corals Pleurocorallium elatius and P. konojoi are distributed in Japanese waters and have distinct morphological features: (1) the terminal branches of the colony form of P. elatius are very fine, while those of P. konojoi are blunt and rounded, (2) the autozooids of P. elatius are arranged in approximately four rows, while those of P. konojoi are clustered in groups. However, previous genetic analysis using mtDNA and nuclear DNA did not indicate monophyly. Therefore, it is important to clarify their species status to allow for their conservation.

Methodology: We collected a total of 87 samples (60 of Corallium japonicum and 27 of P. konojoi) from around the Ryukyu Islands and Shikoku Island, which are geographically separated by approximately 1,300 km. We used a multiplexed inter-simple sequence repeat (ISSR) genotyping by sequencing (MIG-seq) and obtained 223 SNPs with which to perform STRUCTURE analysis and principle coordinate analysis (PCoA). In addition, two relatively polymorphic mtDNA regions were sequenced and compared.

Results: P. elatius and P. konojoi share a same mtDNA haplotype, which has been previously reported. However, MIG-seq analysis clearly distinguished the two species based on PCoA and STRUCTURE analysis, including 5% of species-specific fixed SNPs.

Conclusion: This study indicated that P. elatius and P. konojoi are different species and therefore both species should be conserved separately. Our findings highlight the importance of the conservation of these two species, especially P. elatius, whose population has been dramatically depleted over the last 100 years. The study also demonstrated the effectiveness and robustness of MIG-seq for defining closely related octocoral species that were otherwise indistinguishable using traditional genetic markers (mtDNA and EF).

Keywords: Deep sea coral; Incomplete lineage sorting; MIG-seq; Octocoral; Species delimitation.

Figure 1. MtDNA haplotype network considering gap regions (A) and maximum likelihood phylogenetic tree excluding gap regions (B).

P. elatius (PE) is shown in pink and P. konojoi (PK) is shown in black/white. (A) The size of the circle represents the number of haplotypes found in the analysis. (B) Only the nodes with bootstrap values (>50) were indicated.

Figure 2. STRUCTURE (A) and PCoA result (B).

(A) K = 2 with Mean (LnProb) = − 5666.970 and Mean (similarity score) among 10 runs = 0.999 and PCoA results using 223 SNPs data obtained by MIG-seq analysis (B). X axis indicates 18.5% and Y axis indicates 3.52% of the data. PK and PE represent P. konojoi and, P. elatius respectively.