Shaun P. Wilkinson

Intra-genomic variation in symbiotic dinoflagellates: recent divergence or recombination between lineages?

BackgroundThe symbiosis between corals and the dinoflagellate alga Symbiodinium is essential for the development and survival of coral reefs. Yet this fragile association is highly vulnerable to environmental disturbance. A coral’s ability to tolerate temperature stress depends on the fitness of its resident symbionts, whose thermal optima vary extensively between lineages. However, the in hospite population genetic structure of Symbiodinium is poorly understood and mostly based on analysis of bulk DNA extracted from thousands to millions of cells. Using quantitative single-cell PCR, we enumerated DNA polymorphisms in the symbionts of the reef-building coral Pocillopora damicornis, and applied a model selection approach to explore the potential for recombination between coexisting Symbiodinium populations.ResultsTwo distinct Symbiodinium ITS2 sequences (denoted C100 and C109) were retrieved from all P. damicornis colonies analysed. However, the symbiont assemblage consisted of three distinct Symbiodinium populations: cells featuring pure arrays of ITS2 type C109, near-homogeneous cells of type C100 (with trace ITS2 copies of type C109), and those with co-dominant C100 and C109 ITS2 repeats. The symbiont consortia of some colonies consisted almost entirely of these putative C100 × C109 recombinants.ConclusionsOur results are consistent with the occurrence of sexual recombination between Symbiodinium types C100 and C109. While the multiple-copy nature of the ITS2 dictates that the observed pattern of intra-genomic co-dominance may be a result of incomplete concerted evolution of intra-genomic polymorphisms, this is a less likely explanation given the occurrence of homogeneous cells of the C109 type. Conclusive evidence for inter-lineage recombination and introgression in this genus will require either direct observational evidence or a single-cell genotyping approach targeting multiple, single-copy loci.

Isolation by resistance across a complex coral reef seascape.

A detailed understanding of the genetic structure of populations and an accurate interpretation of processes driving contemporary patterns of gene flow are fundamental to successful spatial conservation management. The field of seascape genetics seeks to incorporate environmental variables and processes into analyses of population genetic data to improve our understanding of forces driving genetic divergence in the marine environment. Information about barriers to gene flow (such as ocean currents) is used to define a resistance surface to predict the spatial genetic structure of populations and explain deviations from the widely applied isolation-by-distance model. The majority of seascape approaches to date have been applied to linear coastal systems or at large spatial scales (more than 250 km), with very few applied to complex systems at regional spatial scales (less than 100 km). Here, we apply a seascape genetics approach to a peripheral population of the broadcast-spawning coral Acropora spicifera across the Houtman Abrolhos Islands, a high-latitude complex coral reef system off the central coast of Western Australia. We coupled population genetic data from a panel of microsatellite DNA markers with a biophysical dispersal model to test whether oceanographic processes could explain patterns of genetic divergence. We identified significant variation in allele frequencies over distances of less than 10 km, with significant differentiation occurring between adjacent sites but not between the most geographically distant ones. Recruitment probabilities between sites based on simulated larval dispersal were projected into a measure of resistance to connectivity that was significantly correlated with patterns of genetic divergence, demonstrating that patterns of spatial genetic structure are a function of restrictions to gene flow imposed by oceanographic currents. This study advances our understanding of the role of larval dispersal on the fine-scale genetic structure of coral populations across a complex island system and applies a methodological framework that can be tailored to suit a variety of marine organisms with a range of life-history characteristics.

Thermal Shock Induces Host Proteostasis Disruption and Endoplasmic Reticulum Stress in the Model Symbiotic Cnidarian Aiptasia

Coral bleaching has devastating effects on coral survival and reef ecosystem function, but many of the fundamental cellular effects of thermal stress on cnidarian physiology are unclear. We used label-free liquid chromatography-tandem mass spectrometry to compare the effects of rapidly (33.5 °C, 24 h) and gradually (30 and 33.5 °C, 12 days) elevated temperatures on the proteome of the model symbiotic anemone Aiptasia. We identified 2133 proteins in Aiptasia, 136 of which were differentially abundant between treatments. Thermal shock, but not acclimation, resulted in significant abundance changes in 104 proteins, including those involved in protein folding and synthesis, redox homeostasis, and central metabolism. Nineteen abundant structural proteins showed particularly reduced abundance, demonstrating proteostasis disruption and potential protein synthesis inhibition. Heat shock induced antioxidant mechanisms and proteins involved in stabilizing nascent proteins, preventing protein aggregation and degrading damaged proteins, which is indicative of endoplasmic reticulum stress. Host proteostasis disruption occurred before either bleaching or symbiont photoinhibition was detected, suggesting host-derived reactive oxygen species production as the proximate cause of thermal damage. The pronounced abundance changes in endoplasmic reticulum proteins associated with proteostasis and protein turnover indicate that these processes are essential in the cellular response of symbiotic cnidarians to severe thermal stress.

Metabolite profiling of symbiont and host during thermal stress and bleaching in the coral Acropora aspera

Rising seawater temperatures pose a significant threat to the persistence of coral reefs. Despite the importance of these systems, major gaps remain in our understanding of how thermal stress and bleaching affect the metabolic networks that underpin holobiont function. We applied gas chromatography–mass spectrometry (GC–MS) metabolomics to detect changes in the intracellular free metabolite pools (polar and semi-polar compounds) of in hospite dinoflagellate symbionts and their coral hosts (and any associated microorganisms) during early- and late-stage thermal bleaching (a reduction of approximately 50 and 70% in symbiont density, respectively). We detected characteristic changes to the metabolite profiles of each symbiotic partner associated with individual cellular responses to thermal, oxidative and osmotic stress, which progressed with the severity of bleaching. Alterations were also indicative of changes to energy-generating and biosynthesis pathways in both partners, with a shift to the increased catabolism of lipid stores. Specifically, in symbiont intracellular metabolite pools, we observed accumulations of multiple free fatty acids, plus the chloroplast-associated antioxidant alpha-tocopherol. In the host, we detected a decline in the abundance of pools of multiple carbohydrates, amino acids and intermediates, in addition to the antioxidant ascorbate. These findings further our understanding of the metabolic changes that occur to symbiont and host (and its associated microorganisms) during thermal bleaching. These findings also provide further insight into the largely undescribed roles of free metabolite pools in cellular homeostasis, signalling and acclimation to thermal stress in the cnidarian–dinoflagellate symbiosis.

The distribution of intra-genomically variable dinoflagellate symbionts at Lord Howe Island, Australia

The symbiotic dinoflagellates of corals and other marine invertebrates (Symbiodinium) are essential to the development of shallow-water coral reefs. This genus contains considerable genetic diversity and a corresponding range of physiological and ecological traits. Most genetic variation arises through the accumulation of somatic mutations that arise during asexual reproduction. Yet growing evidence suggests that occasional sexual reproductive events also occur within, and perhaps between, Symbiodinium lineages, further contributing to the pool of genetic variation available for evolutionary adaptation. Intra-genomic variation can therefore arise from both sexual and asexual reproductive processes, making it difficult to discern its underlying causes and consequences. We used quantitative PCR targeting the ITS2 locus to estimate proportions of genetically homogeneous symbionts and intra-genomically variable Symbiodinium (IGV Symbiodinium) in the reef-building coral Pocillopora damicornis at Lord Howe Island, Australia. We then sampled colonies through time and at a variety of spatial scales to find out whether the distribution of these symbionts followed patterns consistent with niche partitioning. Estimated ratios of homogeneous to IGV Symbiodinium varied between colonies within sites (metres to tens of metres) and between sites separated by hundreds to thousands of metres, but remained stable within colonies through time. Symbiont ratios followed a temperature gradient, with the local thermal maximum emerging as a negative predictor for the estimated proportional abundance of IGV Symbiodinium. While this pattern may result from fine-scale spatial population structure, it is consistent with an increased susceptibility to thermal stress, suggesting that the evolutionary processes that generate IGV (such as inter-lineage recombination and the accumulation of somatic mutations at the ITS2 locus) may have important implications for the fitness of the symbiont and that of the coral host.

phylogram: an R package for phylogenetic analysis with nested lists

The R environment continues to gain popularity as a platform for bioinformatic analysis, due to the reproducible code-based workflow and the many powerful analytical tools available in a suite of open-source packages such as ape (Paradis, Claude, and Strimmer 2004), phangorn (Schliep 2011) and Phytools (Revell 2012). These packages typically employ a tree structure known as the “phylo” object, whose primary element is an integer matrix with one row for each edge in the graph, and two columns giving the indices of the connecting nodes. This is a versatile and memory-efficient structure suitable for most applications encountered by evolutionary biologists, and hence a comprehensive array of tools has been developed for editing, analyzing and visualizing trees in this format.