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Rouzé H, Lecellier GJ, Saulnier D, Planes S, Gueguen Y, Wirshing HH, Berteaux-Lecellier V. (2016) An updated assessment of Symbiodinium that associate with common scleractinian corals from Moorea (French Polynesia) reveals high diversity among background symbionts and a novel finding of clade B.PeerJ Preprints4:e2607v1https://doi.org/10.7287/peerj.preprints.2607v1
The adaptative bleaching hypothesis (ABH) states that depending on the symbiotic flexibility of coral hosts (i.e., the ability of corals to “switch” or “shuffle” their algal symbionts), coral bleaching can lead to a change in the composition of their associated Symbiodinium community, and, thus, contribute to the coral’s overall survival. In order to determine the flexibility of corals, molecular tools are required to provide accurate species delineations, and to detect low levels of coral-associated Symbiodinium. Here, we used highly sensitive quantitative (real-time) PCR (qPCR) technology to analyse five common coral species from Moorea (French Polynesia), previously screened using only traditional conventional molecular methods, to assess the presence of low-abundance (background) Symbiodinium. Similar to other studies, each coral species exhibited a strong specificity to a particular clade, irrespective of the environment. In addition, however, each of the five species harboured at least one additional Symbiodinium clade, among clades A-D, at background levels. Unexpectedly, and for the first time in French Polynesia, clade B was detected as a coral symbiont. These results increase the number of known coral-Symbiodinium associations from corals found in French Polynesia, and likely indicate an underestimation of the ability of the corals in this region to associate with and/or “shuffle” different Symbiodinium clades. Altogether our data suggest that corals from French Polynesia may manage a trade-off between optimizing symbioses with a specific Symbiodinium clade(s), and maintaining associations with particular background clades that may play a role in the ability of corals to respond to environmental change.
Standard curves for different primers, corresponding to Ct values versus logarithmic 10 fold dilution of purified 28S PCR products. Each plot corresponds to individual Ct values obtained from three technical replicates. Primer efficacities were deduced by linear regression: a) mix of purified 28S PCR products from Symbiodinium A-F each concentrated at equal concentration, 1•1011 x e(-0.681x) (clade A, R2= 0.999), 1•1011 x e(-0.676x) (clade B, R2= 0.999), 1•1012 x e(-0.687x) (clade C, R2= 0.983), 1•1012 x e(-0.694x) (clade D, R2= 0.998), 6•1010 x e(-0.663x) (clade E, R2= 0.998) and 8•1010 x e(-0.694x) (clade F, R2= 0.997); and b) mix of coral DNA each concentrated at equal concentration, universal coral primer set, 1.25•104 x e(-0.694x) (mix of 10 coral species: R2=0.999), 9.02•105 x e(-0.705x) (mix of A.cytherea: R2=1.000), 6.58•105 x e(-0.692x) (mix of P.rus: R2=0.999) and 1.0•106 x e(-0.629x) (mix of P.damicronis: R2=0.996).
Efficacities of specific-clade primer sets on Symbiodinium cells isolated from corals
Standard curves for clade-specific primer sets for clades A, C and D corresponding to cell densities versus logarithmic 10 fold-dilution of isolated coral-symbiotic Symbiodinium. Each plot corresponds to individual Ct values obtained from three technical replicates. Primer efficacities were deduced by linear regression: 1•1010 x e(-0.628x) (clade A, R2=0.85), 1•108 x e(-0.45x) (clade C, R2=0.91) and 4•106 x e(-0.363x) (clade D, R2=0.92).
Phylogenetic tree of Symbiodinium using sequences of 28S rDNA
Phylogenetic tree of Symbiodinium clades A-F derived from bayesian analyses using sequences of 28S rDNA from Moorea (in bold black), BURR collection strains described in Table S2 (in bold grey), and published genetic sequence from GenBank (regular font). Bayesian posterior probabilities (first values) and Maximum Likelihood bootstrap support values (second value) are presented following the substitution model of Kimura 2-parameter with a proportion of invariable sites.
Symbiodinium clades C and F in seawater samples of Moorea
Presence of Symbiodinium clade C, E and F in seawater samples of Moorea. PCR amplification (primer sets clade-specific of Yamashita et al. 2011) on DNA extracts from saturated 0.2μm filters (filtration volume: 6-9 L) of 4 seawater samples (M1, M2, M3 and M4)., T-: negative control (no DNA); TD: positive control (clade D DNA). A 100bp amplicon characterizes a positive amplification. Uppercase letters indicate the corresponding clade.
Values of Ct obtained by qPCR assays of the corresponding clade-specific primer sets on different DNAs issued from cultured Symbiodinium strains (clades A to F; BURR collection). Each DNA was tested at an equal amount of 10ng per reaction. (-) means no amplification or unspecific amplifications based on dissociation curves analysis, and (*) symbiotic strains used in mix at same concentration and tested with each primer sets.
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