Increasing comparability among coral bleaching experiments.

Coral bleaching is the single largest global threat to coral reefs worldwide. Integrating the diverse body of work on coral bleaching is critical to understanding and combating this global problem. Yet investigating the drivers, patterns, and processes of coral bleaching poses a major challenge. A recent review of published experiments revealed a wide range of experimental variables used across studies. Such a wide range of approaches enhances discovery, but without full transparency in the experimental and analytical methods used, can also make comparisons among studies challenging. To increase comparability but not stifle innovation, we propose a common framework for coral bleaching experiments that includes consideration of coral provenance, experimental conditions, and husbandry. For example, reporting the number of genets used, collection site conditions, the experimental temperature offset(s) from the maximum monthly mean (MMM) of the collection site, experimental light conditions, flow, and the feeding regime will greatly facilitate comparability across studies. Similarly, quantifying common response variables of endosymbiont (Symbiodiniaceae) and holobiont phenotypes (i.e., color, chlorophyll, endosymbiont cell density, mortality, and skeletal growth) could further facilitate cross-study comparisons. While no single bleaching experiment can provide the data necessary to determine global coral responses of all corals to current and future ocean warming, linking studies through a common framework as outlined here, would help increase comparability among experiments, facilitate synthetic insights into the causes and underlying mechanisms of coral bleaching, and reveal unique bleaching responses among genets, species, and regions. Such a collaborative framework that fosters transparency in methods used would strengthen comparisons among studies that can help inform coral reef management and facilitate conservation strategies to mitigate coral bleaching worldwide.

Host-symbiont coevolution, cryptic structure, and bleaching susceptibility, in a coral species complex (Scleractinia; Poritidae).

The 'species' is a key concept for conservation and evolutionary biology, yet the lines between population and species-level variation are often blurred, especially for corals. The 'Porites lobata species complex' consists of branching and mounding corals that form reefs across the Pacific. We used reduced representation meta-genomic sequencing to examine genetic relationships within this species complex and to identify candidate loci associated with colony morphology, cryptic genetic structure, and apparent bleaching susceptibility. We compared existing Porites data with bleached and unbleached colonies of the branching coral P. compressa collected in Kane'ohe Bay Hawai'i during the 2015 coral bleaching event. Loci that mapped to coral, symbiont, and microbial references revealed genetic structure consistent with recent host-symbiont co-evolution. Cryptic genetic clades were resolved that previous work has associated with distance from shore, but no genetic structure was associated with bleaching. We identified many candidate loci associated with morphospecies, including candidate host and symbiont loci with fixed differences between branching and mounding corals. We also found many loci associated with cryptic genetic structure, yet relatively few loci associated with bleaching. Recent host-symbiont co-evolution and rapid diversification suggests that variation and therefore the capacity of these corals to adapt may be underappreciated.

Coral hybridization or phenotypic variation? Genomic data reveal gene flow between Porites lobata and P. Compressa.

Major gaps remain in our understanding of the ecology, evolution, biodiversity, biogeography, extinction risk, and adaptive potential of reef building corals. One of the central challenges remains that there are few informative genetic markers for studying boundaries between species, and variation within species. Reduced representation sequencing approaches, such as RADseq (Restriction site Associated DNA sequencing) have great potential for resolving such relationships. However, it is necessary to identify loci in order to make inferences for endosymbiotic organisms such as corals. Here, we examined twenty-one coral holobiont ezRAD libraries from Hawai'i, focusing on P. lobata and P. compressa, two species with contrasting morphology and habitat preference that previous studies have not resolved. We used a combination of de novo assembly and reference mapping approaches to identify and compare loci: we used reference mapping to extract and compare nearly complete mitochondrial genomes, ribosomal arrays, and histone genes. We used de novo clustering and phylogenomic methods to compare the complete holobiont data set with coral and symbiont subsets that map to transcriptomic data. In addition, we used reference assemblies to examine genetic structure from SNPs (Single Nucleotide Polymorphisms). All approaches resolved outgroup taxa but failed to resolve P. lobata and P. compressa as distinct, with mito-nuclear discordance and shared mitochondrial haplotypes within the species complex. The holobiont and 'coral transcriptomic' datasets were highly concordant, revealing stronger genetic structure between sites than between coral morphospecies. These results suggest that either branching morphology is a polymorphic trait, or that these species frequently hybridize. This study provides examples of several approaches to acquire, identify, and compare loci across metagenomic samples such as the coral holobiont while providing insights into the nature of coral variability.

Temporal and spatial trends in prey composition of wahoo Acanthocybium solandri: a diet analysis from the central North Pacific Ocean using visual and DNA bar-coding techniques.

A diet analysis was conducted on 444 wahoo Acanthocybium solandri caught in the central North Pacific Ocean longline fishery and a nearshore troll fishery surrounding the Hawaiian Islands from June to December 2014. In addition to traditional observational methods of stomach contents, a DNA bar-coding approach was integrated into the analysis by sequencing the cytochrome c oxidase subunit 1 (COI) region of the mtDNA genome to taxonomically identify individual prey items that could not be classified visually to species. For nearshore-caught A. solandri, juvenile pre-settlement reef fish species from various families dominated the prey composition during the summer months, followed primarily by Carangidae in autumn months. Gempylidae, Echeneidae and Scombridae were dominant prey taxa from the offshore fishery. Molidae was a common prey family found in stomachs collected north-east of the Hawaiian Archipelago while tetraodontiform reef fishes, known to have extended pelagic stages, were prominent prey items south-west of the Hawaiian Islands. The diet composition of A. solandri was indicative of an adaptive feeder and thus revealed dominant geographic and seasonal abundances of certain taxa from various ecosystems in the marine environment. The addition of molecular bar-coding to the traditional visual method of prey identifications allowed for a more comprehensive range of the prey field of A. solandri to be identified and should be used as a standard component in future diet studies.

Common misconceptions in molecular ecology: echoes of the modern synthesis.

The field of molecular ecology has burgeoned into a large discipline spurred on by technical innovations that facilitate the rapid acquisition of large amounts of genotypic data, by the continuing development of theory to interpret results, and by the availability of computer programs to analyse data sets. As the discipline grows, however, misconceptions have become enshrined in the literature and are perpetuated by routine citations to other articles in molecular ecology. These misconceptions hamper a better understanding of the processes that influence genetic variation in natural populations and sometimes lead to erroneous conclusions. Here, we consider eight misconceptions commonly appearing in the literature: (i) some molecular markers are inherently better than other markers; (ii) mtDNA produces higher F(ST) values than nDNA; (iii) estimated population coalescences are real; (iv) more data are always better; (v) one needs to do a Bayesian analysis; (vi) selective sweeps influence mtDNA data; (vii) equilibrium conditions are critical for estimating population parameters; and (viii) having better technology makes us smarter than our predecessors. This is clearly not an exhaustive list and many others can be added. It is, however, sufficient to illustrate why we all need to be more critical of our own understanding of molecular ecology and to be suspicious of self-evident truths.

Protein expression and genetic structure of the coral Porites lobata in an environmentally extreme Samoan back reef: does host genotype limit phenotypic plasticity?

The degree to which coral reef ecosystems will be impacted by global climate change depends on regional and local differences in corals' susceptibility and resilience to environmental stressors. Here, we present data from a reciprocal transplant experiment using the common reef building coral Porites lobata between a highly fluctuating back reef environment that reaches stressful daily extremes, and a more stable, neighbouring forereef. Protein biomarker analyses assessing physiological contributions to stress resistance showed evidence for both fixed and environmental influence on biomarker response. Fixed influences were strongest for ubiquitin-conjugated proteins with consistently higher levels found in back reef source colonies both pre and post-transplant when compared with their forereef conspecifics. Additionally, genetic comparisons of back reef and forereef populations revealed significant population structure of both the nuclear ribosomal and mitochondrial genomes of the coral host (F(ST) = 0.146 P < 0.0001, F(ST) = 0.335 P < 0.0001 for rDNA and mtDNA, respectively), whereas algal endosymbiont populations were genetically indistinguishable between the two sites. We propose that the genotype of the coral host may drive limitations to the physiological responses of these corals when faced with new environmental conditions. This result is important in understanding genotypic and environmental interactions in the coral algal symbiosis and how corals may respond to future environmental changes.

Conservation implications of complex population structure: lessons from the loggerhead turtle (Caretta caretta).

Complex population structure can result from either sex-biased gene flow or population overlap during migrations. Loggerhead turtles (Caretta caretta) have both traits, providing an instructive case history for wildlife management. Based on surveys of maternally inherited mtDNA, pelagic post-hatchlings show no population structure across the northern Atlantic (phi(ST) < 0.001, P = 0.919), subadults in coastal habitat show low structure among locations (phi(ST) = 0.01, P < 0.005), and nesting colonies along the southeastern coast of the United States have strong structure (phi(ST) = 0.42, P < 0.001). Thus the level of population structure increases through progressive life history stages. In contrast, a survey of biparentally inherited microsatellite DNA shows no significant population structure: R(ST) < 0.001; F(ST) = 0.002 (P > 0.05) across the same nesting colonies. These results indicate that loggerhead females home faithfully to their natal nesting colony, but males provide an avenue of gene flow between regional nesting colonies, probably via opportunistic mating in migratory corridors. As a result, all breeding populations in the southeastern United States have similar levels of microsatellite diversity (H(E) = 0.70-0.89), whereas mtDNA haplotype diversity varies dramatically (h = 0.00-0.66). Under a conventional interpretation of the nuclear DNA data, the entire southeastern United States would be regarded as a single management unit, yet the mtDNA data indicate multiple isolated populations. This complex population structure mandates a different management strategy at each life stage. Perturbations to pelagic juveniles will have a diffuse impact on Atlantic nesting colonies, mortality of subadults will have a more focused impact on nearby breeding populations, and disturbances to adults will have pinpoint impact on corresponding breeding populations. These findings demonstrate that surveys of multiple life stages are desirable to resolve management units in migratory marine species.

Foundations of gregariousness: a dispersal polymorphism among the planktonic larvae of a marine invertebrate.

Theory predicts that selection should favor genotypes that can vary their tendency to disperse in habitats that are spatially or temporally variable or those that remain near their carrying capacity. Although many marine habitats appear to fit these criteria, confirmed examples of dispersal polymorphism among marine invertebrates are exceedingly rare. Competent larvae of the gregarious tubeworm, Hydroides dianthus, settle specifically in response to living conspecific worms, but a small proportion of each spawn settle nonspecifically on uninhabited substrata concurrently with their gregarious siblings. Here, using a parental half-sib analysis, we show that the proportion of a spawn settling in response to uninhabited biofilm is highly heritable. When estimated as a continuous trait based on a one-way ANOVA, heritability is estimated to be 0.83 +/- 0.31. When founder production was analyzed as a threshold trait, heritability was estimated to be 0.68 +/- 0.10 based on the breeding design experiment and 0.65 +/- 0.09 based on the artificial selection experiments. Realized heritability based on the selection experiments was considerably lower, however (0.17 per generation and 0.02 cumulative). Artificial selection was ineffectual at sequentially increasing the proportion of founder larvae among inbred family lines, but after three generations of selection, the proportion of larvae settling in response to biofilm was significantly higher among inbred lines than among the field-collected parents. The obligate planktonic larval stage common among so many marine invertebrates is thought to preclude the evolution of dispersal polymorphisms in these animals. Theoretical expectations of variable dispersal may instead be realized through individual behavioral differences resulting in differential transport or settlement preference, but this possibility remains largely unexplored among marine invertebrates.

Increased throughput for fragment analysis on an ABI PRISM 377 automated sequencer using a membrane comb and STRand software.

This manuscript outlines our protocol for using a freely downloadable fragment analysis software package (STRand) together with a 96+4 RapidLoad membrane comb to increase throughput of samples for fragment analysis on ABI sequencers without costly upgrades from the manufacturer. We outline how using these products allows one to score 90 lanes of sample per gel on an ABI PRISM 377XL (64-lane sequencer), saving both time and money in the processing of samples. This protocol is a major modification to those suggested by the manufacturer. This protocol gives more consistent results that are easier to score than standard protocols, and it reduces reagent costs. Interest in fragment analysis (primarily microsatellites and AFLPs) is steadily increasing among both population ecologists and geneticists, and methods that simultaneously increase sample throughput while reducing costs associated with these analyses by over 50% per gel should prove useful to anyone using an ABI, MJ Basestation, or LI-COR automated sequencer for fragment analysis.