High germline mutation rates, but not extreme population outbreaks, influence genetic diversity in a keystone coral predator.

Lewontin's paradox, the observation that levels of genetic diversity (π) do not scale linearly with census population size (Nc) variation, is an evolutionary conundrum. The most extreme mismatches between π and Nc are found for highly abundant marine invertebrates. Yet, the influences of new mutations on π relative to extrinsic processes such as Nc fluctuations are unknown. Here, we provide the first germline mutation rate (µ) estimate for a marine invertebrate in corallivorous crown-of-thorns sea stars (Acanthaster cf. solaris). We use high-coverage whole-genome sequencing of 14 parent-offspring trios alongside empirical estimates of Nc in Australia's Great Barrier Reef to jointly examine the determinants of π in populations undergoing extreme Nc fluctuations. The A. cf. solaris mean µ was 9.13 x 10-09 mutations per-site per-generation (95% CI: 6.51 x 10-09 to 1.18 x 10-08), exceeding estimates for other invertebrates and showing greater concordance with vertebrate mutation rates. Lower-than-expected Ne (~70,000-180,000) and low Ne/Nc values (0.0047-0.048) indicated weak influences of population outbreaks on long-term π. Our findings are consistent with elevated µ evolving in response to reduced Ne and generation time length, with important implications for explaining high mutational loads and the determinants of genetic diversity in marine invertebrate taxa.

Life-stage specificity and cross-generational climate effects on the microbiome of a tropical sea urchin (Echinodermata: Echinoidea).

Microbes play a critical role in the development and health of marine invertebrates, though microbial dynamics across life stages and host generations remain poorly understood in most reef species, especially in the context of climate change. Here, we use a 4-year multigenerational experiment to explore microbe-host interactions under the Intergovernmental Panel on Climate Change (IPCC)-forecast climate scenarios in the rock-boring tropical urchin Echinometra sp. A. Adult urchins (F0 ) were exposed for 18 months to increased temperature and pCO2 levels predicted for years 2050 and 2100 under RCP 8.5, a period which encompassed spawning. After rearing F1 offspring for a further 2 years, spawning was induced, and F2 larvae were raised under current day and 2100 conditions. Cross-generational climate effects were also explored in the microbiome of F1 offspring through a transplant experiment. Using 16S rRNA gene sequence analysis, we determined that each life stage and generation was associated with a distinct microbiome, with higher microbial diversity observed in juveniles compared to larval stages. Although life-stage specificity was conserved under climate conditions projected for 2050 and 2100, we observed changes in the urchin microbial community structure within life stages. Furthermore, we detected a climate-mediated parental effect when juveniles were transplanted among climate treatments, with the parental climate treatment influencing the offspring microbiome. Our findings reveal a potential for cross-generational impacts of climate change on the microbiome of a tropical invertebrate species.

Settlement cue selectivity by larvae of the destructive crown-of-thorns starfish.

Population irruptions of crown-of-thorns starfish (COTS) cause extensive degradation of coral reefs, threatening the structure and function of these important ecosystems. For population irruptions to initiate and spread, large numbers of planktonic larvae have to successfully transition into their benthic life-history stage (i.e. settlement), whereby larval behaviour and the presence of settlement cues may shape spatial patterns of recruitment and adult densities. Our results demonstrate that a wide range of coralline algae species induce COTS larvae to settle; however, the capacity to promote settlement success varied manyfold among algal species, ranging from greater than 90% in Melyvonnea cf. madagascariensis to less than 2% in Lithophyllum cf. kotschyanum and two Porolithon species at 24 h. Because many coralline algae species that promote high settlement success are prevalent in shallow reef habitats, our findings challenge the hypothesis that COTS larvae predominantly settle in deep water. Considering both larval behaviour and algal ecology, this study highlights the ecological significance of coralline algae communities in driving recruitment patterns of COTS. More specifically, the local abundance of highly inductive coralline algae (especially, Melyvonnea cf. madagascariensis) may explain some of the marked spatial heterogeneity of COTS populations and the incidence of population irruptions.

Cross-generational response of a tropical sea urchin to global change and a selection event in a 43-month mesocosm study.

Long-term experimental investigations of transgenerational plasticity (TGP) and transgenerational acclimatization to global change are sparse in marine invertebrates. Here, we test the effect of ocean warming and acidification over a 25-month period of Echinometra sp. A sea urchins whose parents were acclimatized at ambient or one of two near-future (projected mid and end of the 21st century) climate scenarios for 18 months. Several parameters linked to performance exhibited strong effects of future ocean conditions at 9 months of age. The Ambient-Ambient group (A-A, both F0 and F1 at ambient conditions) was significantly larger (21%) and faster in righting response (31%) compared to other groups. A second set of contrasts revealed near-future scenarios caused significant negative parental carryover effects. Respiration at 9 months was depressed by 59% when parents were from near-future climate conditions, and righting response was slowed by 28%. At 10 months, a selective pathogenic mortality event led to significantly higher survival rates of A-A urchins. Differences in size and respiration measured prior to the mortality were absent after the event, while a negative parental effect on righting (29% reduction) remained. The capacity to spawn at the end of the experiment was higher in individuals with ambient parents (50%) compared to other groups (21%) suggesting persistent parental effects. Obtaining different results at different points in time illustrates the importance of longer term and multigeneration studies to investigate effects of climate change. Given some animals in all groups survived the pathogenic event and that effects on physiology (but not behavior) among groups were eliminated after the mortality, we suggest that similar events could constitute selective sweeps, allowing genetic adaptation. However, given the observed negative parental effects and reduced potential for population replenishment, it remains to be determined if selection would be sufficiently rapid to rescue this species from climate change effects.

Cross-generational effects of climate change on the microbiome of a photosynthetic sponge.

Coral reefs are facing increasing pressure from rising seawater temperatures and ocean acidification. Sponges have been proposed as possible winners in the face of climate change; however, little is known about the mechanisms underpinning their predicted tolerance. Here we assessed whether microbiome-mediated cross-generational acclimatization could enable the photosynthetic sponge Carteriospongia foliascens to survive under future climate scenarios. To achieve this, we first established the potential for vertical (cross-generational) transmission of symbionts. Sixty-four amplicon sequence variants accounting for >90% of the total C. foliascens microbial community were present across adult, larval and juvenile life stages, showing that a large proportion of the microbiome is vertically acquired and maintained. When C. foliascens were exposed to climate scenarios projected for 2050 and 2100, the host remained visibly unaffected (i.e. no necrosis/bleaching) and the overall microbiome was not significantly different amongst treatments in adult tissue, the respective larvae or recruits transplanted amongst climate treatments. However, indicator species analysis revealed that parental exposure to future climate scenarios altered the presence and abundance of a small suite of microbial taxa in the recruits, thereby revealing the potential for microbiome-mediated cross-generational acclimatization through both symbiont shuffling and symbiont switching within a vertically acquired microbiome.

Adjusting Tropical Marine Water Quality Guideline Values for Elevated Ocean Temperatures.

Increased frequency of summer heatwaves and poor water quality are two of the most prevalent and severe pressures faced by coral reefs. While these pressures often co-occur, their potential risks to tropical marine species are usually considered independently. Here, we extended the application of multisubstance-Potentially Affected Fraction (ms-PAF) to a nonchemical stressor, elevated sea surface temperature. We then applied this method to calculate climate-adjusted water quality guideline values (GVs) for two reference toxicants, copper and the herbicide diuron, for tropical marine species. First, we developed a species sensitivity distribution (SSD) for thermal stress based on published experimental data for 41 tropical benthic marine species using methods adapted from water quality GV derivation. This enabled quantitative predictions of community effects as temperatures exceeded acclimation values. The resulting protective temperature values (PTx) were similar to temperatures known to initiate coral bleaching and are therefore relevant for application in multistressor risk assessments. The extended ms-PAF method enabled the adjustment of current water quality GVs to account for thermal stress events. This approach could be applied to other ecosystems and other non-contaminant stressors (e.g., sediment, low salinity, anoxia, and ocean acidification), offering an alternative approach for deriving environmental GVs, reporting and assessing the risk posed by multiple stressors.

Crown-of-Thorns Sea Star Acanthaster cf. solaris Has Tissue-Characteristic Microbiomes with Potential Roles in Health and Reproduction.

Outbreaks of coral-eating crown-of-thorns sea stars (CoTS; Acanthaster species complex) cause substantial coral loss; hence, there is considerable interest in developing prevention and control strategies. We characterized the microbiome of captive CoTS and assessed whether dysbiosis was evident in sea stars during a disease event. Most tissue types had a distinct microbiome. The exception was female gonads, in which the microbiomes were highly variable among individuals. Male gonads were dominated (>97% of reads) by a single Mollicutes-related operational taxonomic unit (OTU). Detailed phylogenetic and microscopy analysis demonstrated the presence of a novel Spiroplasma-related bacterium in the spermatogenic layer. Body wall samples had high relative abundance (43 to 64% of reads) of spirochetes, likely corresponding to subcuticular symbionts reported from many echinoderms. Tube feet were characterized by Hyphomonadaceae (24 to 55% of reads). Pyloric cecal microbiomes had high alpha diversity, comprising many taxa commonly found in gastrointestinal systems. The order Oceanospirillales (genera Endozoicomonas and Kistimonas) was detected in all tissues. A microbiome shift occurred in diseased individuals although differences between tissue types were retained. The relative abundance of spirochetes was significantly reduced in diseased individuals. Kistimonas was present in all diseased individuals and significantly associated with diseased tube feet, but its role in disease causation is unknown. While Arcobacter was significantly associated with diseased tissues and Vibrionaceae increased in diversity, no single OTU was detected in all diseased individuals, suggesting opportunistic proliferation of these taxa in this case. This study shows that CoTS have tissue-characteristic bacterial communities and identifies taxa that could play a role in reproduction and host health.IMPORTANCE Coral-eating crown-of-thorns sea stars (CoTS; Acanthaster species complex) are native to the Indo-Pacific, but during periodic population outbreaks they can reach extreme densities (>1,000 starfish per hectare) and function as a pest species. On the Great Barrier Reef, Australia, CoTS have long been considered one of the major contributors to coral loss. There has been significant investment in a targeted control program using lethal injection, and there is interest in developing additional and complementary technologies that can increase culling efficiencies. The biology of CoTS has been studied extensively, but little is known about their associated microbiome. This cultivation-independent analysis of the CoTS microbiome provides a baseline for future analyses targeting the functional role of symbionts, the identification of pathogens, or the development of reproduction manipulators.

Losing a winner: thermal stress and local pressures outweigh the positive effects of ocean acidification for tropical seagrasses.

Seagrasses are globally important coastal habitat-forming species, yet it is unknown how seagrasses respond to the combined pressures of ocean acidification and warming of sea surface temperature. We exposed three tropical species of seagrass (Cymodocea serrulata, Halodule uninervis, and Zostera muelleri) to increasing temperature (21, 25, 30, and 35°C) and pCO2 (401, 1014, and 1949 µatm) for 7 wk in mesocosms using a controlled factorial design. Shoot density and leaf extension rates were recorded, and plant productivity and respiration were measured at increasing light levels (photosynthesis-irradiance curves) using oxygen optodes. Shoot density, growth, photosynthetic rates, and plant-scale net productivity occurred at 25°C or 30°C under saturating light levels. High pCO2 enhanced maximum net productivity for Z. muelleri, but not in other species. Z. muelleri was the most thermally tolerant as it maintained positive net production to 35°C, yet for the other species there was a sharp decline in productivity, growth, and shoot density at 35°C, which was exacerbated by pCO2 . These results suggest that thermal stress will not be offset by ocean acidification during future extreme heat events and challenges the current hypothesis that tropical seagrass will be a 'winner' under future climate change conditions.

Ocean acidification has little effect on developmental thermal windows of echinoderms from Antarctica to the tropics.

As the ocean warms, thermal tolerance of developmental stages may be a key driver of changes in the geographical distributions and abundance of marine invertebrates. Additional stressors such as ocean acidification may influence developmental thermal windows and are therefore important considerations for predicting distributions of species under climate change scenarios. The effects of reduced seawater pH on the thermal windows of fertilization, embryology and larval morphology were examined using five echinoderm species: two polar (Sterechinus neumayeri and Odontaster validus), two temperate (Fellaster zelandiae and Patiriella regularis) and one tropical (Arachnoides placenta). Responses were examined across 12-13 temperatures ranging from -1.1 °C to 5.7 °C (S. neumayeri), -0.5 °C to 10.7 °C (O. validus), 5.8 °C to 27 °C (F. zelandiae), 6.0 °C to 27.1 °C (P. regularis) and 13.9 °C to 34.8 °C (A. placenta) under present-day and near-future (2100+) ocean acidification conditions (-0.3 pH units) and for three important early developmental stages 1) fertilization, 2) embryo (prehatching) and 3) larval development. Thermal windows for fertilization were broad and were not influenced by a pH decrease. Embryological development was less thermotolerant. For O. validus, P. regularis and A. placenta, low pH reduced normal development, albeit with no effect on thermal windows. Larval development in all five species was affected by both temperature and pH; however, thermal tolerance was not reduced by pH. Results of this study suggest that in terms of fertilization and development, temperature will remain as the most important factor influencing species' latitudinal distributions as the ocean continues to warm and decrease in pH, and that there is little evidence of a synergistic effect of temperature and ocean acidification on the thermal control of species ranges.

In situ developmental responses of tropical sea urchin larvae to ocean acidification conditions at naturally elevated pCO2 vent sites.

Laboratory experiments suggest that calcifying developmental stages of marine invertebrates may be the most ocean acidification (OA)-sensitive life-history stage and represent a life-history bottleneck. To better extrapolate laboratory findings to future OA conditions, developmental responses in sea urchin embryos/larvae were compared under ecologically relevant in situ exposures on vent-elevated pCO2 and ambient pCO2 coral reefs in Papua New Guinea. Echinometra embryos/larvae were reared in meshed chambers moored in arrays on either venting reefs or adjacent non-vent reefs. After 24 and 48 h, larval development and morphology were quantified. Compared with controls (mean pH(T) = 7.89-7.92), larvae developing in elevated pCO2 vent conditions (pH(T) = 7.50-7.72) displayed a significant reduction in size and increased abnormality, with a significant correlation of seawater pH with both larval size and larval asymmetry across all experiments. Reciprocal transplants (embryos from vent adults transplanted to control conditions, and vice versa) were also undertaken to identify if adult acclimatization can translate resilience to offspring (i.e. transgenerational processes). Embryos originating from vent adults were, however, no more tolerant to reduced pH. Sea temperature and chlorophyll-a concentrations (i.e. larval nutrition) did not contribute to difference in larval size, but abnormality was correlated with chlorophyll levels. This study is the first to examine the response of marine larvae to OA scenarios in the natural environment where, importantly, we found that stunted and abnormal development observed in situ are consistent with laboratory observations reported in sea urchins, in both the direction and magnitude of the response.

Echinometra sea urchins acclimatized to elevated pCO2 at volcanic vents outperform those under present-day pCO2 conditions.

Rising atmospheric CO2 concentrations will significantly reduce ocean pH during the 21st century (ocean acidification, OA). This may hamper calcification in marine organisms such as corals and echinoderms, as shown in many laboratory-based experiments. Sea urchins are considered highly vulnerable to OA. We studied an Echinometra species on natural volcanic CO2 vents in Papua New Guinea, where they are CO2 -acclimatized and also subjected to secondary ecological changes from elevated CO2 . Near the vent site, the urchins experienced large daily variations in pH (>1 unit) and pCO2 (>2000 ppm) and average pH values (pHT 7.73) much below those expected under the most pessimistic future emission scenarios. Growth was measured over a 17-month period using tetracycline tagging of the calcareous feeding lanterns. Average-sized urchins grew more than twice as fast at the vent compared with those at an adjacent control site and assumed larger sizes at the vent compared to the control site and two other sites at another reef near-by. A small reduction in gonad weight was detected at the vents, but no differences in mortality, respiration, or degree of test calcification were detected between urchins from vent and control populations. Thus, urchins did not only persist but actually 'thrived' under extreme CO2 conditions. We suggest an ecological basis for this response: Increased algal productivity under increased pCO2 provided more food at the vent, resulting in higher growth rates. The wider implication of our observation is that laboratory studies on non-acclimatized specimens, which typically do not consider ecological changes, can lead to erroneous conclusions on responses to global change.

Changes in microbial communities in coastal sediments along natural CO2 gradients at a volcanic vent in Papua New Guinea.

Natural CO2 venting systems can mimic conditions that resemble intermediate to high pCO2 levels as predicted for our future oceans. They represent ideal sites to investigate potential long-term effects of ocean acidification on marine life. To test whether microbes are affected by prolonged exposure to pCO2 levels, we examined the composition and diversity of microbial communities in oxic sandy sediments along a natural CO2 gradient. Increasing pCO2 was accompanied by higher bacterial richness and by a strong increase in rare members in both bacterial and archaeal communities. Microbial communities from sites with CO2 concentrations close to today's conditions had different structures than those of sites with elevated CO2 levels. We also observed increasing sequence abundance of several organic matter degrading types of Flavobacteriaceae and Rhodobacteraceae, which paralleled concurrent shifts in benthic cover and enhanced primary productivity. With increasing pCO2 , sequences related to bacterial nitrifying organisms such as Nitrosococcus and Nitrospirales decreased, and sequences affiliated to the archaeal ammonia-oxidizing Thaumarchaeota Nitrosopumilus maritimus increased. Our study suggests that microbial community structure and diversity, and likely key ecosystem functions, may be altered in coastal sediments by long-term CO2 exposure to levels predicted for the end of the century.

Ocean acidification induces biochemical and morphological changes in the calcification process of large benthic foraminifera.

Large benthic foraminifera are significant contributors to sediment formation on coral reefs, yet they are vulnerable to ocean acidification. Here, we assessed the biochemical and morphological impacts of acidification on the calcification of Amphistegina lessonii and Marginopora vertebralis exposed to different pH conditions. We measured growth rates (surface area and buoyant weight) and Ca-ATPase and Mg-ATPase activities and calculated shell density using micro-computer tomography images. In A. lessonii, we detected a significant decrease in buoyant weight, a reduction in the density of inner skeletal chambers, and an increase of Ca-ATPase and Mg-ATPase activities at pH 7.6 when compared with ambient conditions of pH 8.1. By contrast, M. vertebralis showed an inhibition in Mg-ATPase activity under lowered pH, with growth rate and skeletal density remaining constant. While M. vertebralis is considered to be more sensitive than A. lessonii owing to its high-Mg-calcite skeleton, it appears to be less affected by changes in pH, based on the parameters assessed in this study. We suggest difference in biochemical pathways of calcification as the main factor influencing response to changes in pH levels, and that A. lessonii and M. vertebralis have the ability to regulate biochemical functions to cope with short-term increases in acidity.

Larvae of the coral eating crown-of-thorns starfish, Acanthaster planci in a warmer-high CO2 ocean.

Outbreaks of crown-of-thorns starfish (COTS), Acanthaster planci, contribute to major declines of coral reef ecosystems throughout the Indo-Pacific. As the oceans warm and decrease in pH due to increased anthropogenic CO2 production, coral reefs are also susceptible to bleaching, disease and reduced calcification. The impacts of ocean acidification and warming may be exacerbated by COTS predation, but it is not known how this major predator will fare in a changing ocean. Because larval success is a key driver of population outbreaks, we investigated the sensitivities of larval A. planci to increased temperature (2-4 °C above ambient) and acidification (0.3-0.5 pH units below ambient) in flow-through cross-factorial experiments (3 temperature × 3 pH/pCO2 levels). There was no effect of increased temperature or acidification on fertilization or very early development. Larvae reared in the optimal temperature (28 °C) were the largest across all pH treatments. Development to advanced larva was negatively affected by the high temperature treatment (30 °C) and by both experimental pH levels (pH 7.6, 7.8). Thus, planktonic life stages of A. planci may be negatively impacted by near-future global change. Increased temperature and reduced pH had an additive negative effect on reducing larval size. The 30 °C treatment exceeded larval tolerance regardless of pH. As 30 °C sea surface temperatures may become the norm in low latitude tropical regions, poleward migration of A. planci may be expected as they follow optimal isotherms. In the absence of acclimation or adaptation, declines in low latitude populations may occur. Poleward migration will be facilitated by strong western boundary currents, with possible negative flow-on effects on high latitude coral reefs. The contrasting responses of the larvae of A. planci and those of its coral prey to ocean acidification and warming are considered in context with potential future change in tropical reef ecosystems.

Impacts of ocean warming on echinoderms: A meta-analysis.

Rising ocean temperatures are threatening marine species and populations worldwide, and ectothermic taxa are particularly vulnerable. Echinoderms are an ecologically important phylum of marine ectotherms and shifts in their population dynamics can have profound impacts on the marine environment. The effects of warming on echinoderms are highly variable across controlled laboratory-based studies. Accordingly, synthesis of these studies will facilitate the better understanding of broad patterns in responses of echinoderms to ocean warming. Herein, a meta-analysis incorporating the results of 85 studies (710 individual responses) is presented, exploring the effects of warming on various performance predictors. The mean responses of echinoderms to all magnitudes of warming were compared across multiple biological responses, ontogenetic life stages, taxonomic classes, and regions, facilitated by multivariate linear mixed effects models. Further models were conducted, which only incorporated responses to warming greater than the projected end-of-century mean annual temperatures at the collection sites. This meta-analysis provides evidence that ocean warming will generally accelerate metabolic rate (+32%) and reduce survival (-35%) in echinoderms, and echinoderms from subtropical (-9%) and tropical (-8%) regions will be the most vulnerable. The relatively high vulnerability of echinoderm larvae to warming (-20%) indicates that this life stage may be a significant developmental bottleneck in the near-future, likely reducing successful recruitment into populations. Furthermore, asteroids appear to be the class of echinoderms that are most negatively affected by elevated temperature (-30%). When considering only responses to magnitudes of warming representative of end-of-century climate change projections, the negative impacts on asteroids, tropical species and juveniles were exacerbated (-51%, -34% and -40% respectively). The results of these analyses enable better predictions of how keystone and invasive echinoderm species may perform in a warmer ocean, and the possible consequences for populations, communities and ecosystems.

Induction of larval settlement in crown-of-thorns starfish is not mediated by conspecific cues.

Population irruptions of crown-of-thorns starfish (COTS; Acanthaster spp.) remain a major cause of coral reef degradation throughout the Pacific and Indian Oceans and are inherently modulated by larval settlement and recruitment success. Gregarious larval settlement, as exhibited by many other ecologically important marine invertebrates, can catalyse population growth and replenishment. However, whether conspecific cues induce or influence the settlement of COTS larvae remains a critical information gap. This experimental study examined the induction of COTS settlement in response to a range of conspecific cues associated with early- and late-stage herbivorous juveniles, corallivorous juveniles and adults. Competent COTS larvae were generally not induced to settle by the presence of conspecifics or cues associated with conspecifics, while the settlement success of COTS in the presence of coralline algae was not inhibited or enhanced by adding conspecific conditioned seawater. Rather than being reinforced by gregarious settlement, the recruitment of COTS populations appears dependent on associative settlement cues (i.e., coralline algae and/or associated microbial communities) signalling suitable benthic habitat.

Terrestrial runoff controls the bacterial community composition of biofilms along a water quality gradient in the Great Barrier Reef.

16S rRNA gene molecular analysis elucidated the spatiotemporal distribution of bacterial biofilm communities along a water quality gradient. Multivariate statistics indicated that terrestrial runoff, in particular dissolved organic carbon and chlorophyll a concentrations, induced shifts of specific bacterial communities between locations and seasons, suggesting microbial biofilms could be suitable bioindicators for water quality.

Productivity gains do not compensate for reduced calcification under near-future ocean acidification in the photosynthetic benthic foraminifer species Marginopora vertebralis.

Changes in the seawater carbonate chemistry (ocean acidification) from increasing atmospheric carbon dioxide (CO2 ) concentrations negatively affect many marine calcifying organisms, but may benefit primary producers under dissolved inorganic carbon (DIC) limitation. To improve predictions of the ecological effects of ocean acidification, the net gains and losses between the processes of photosynthesis and calcification need to be studied jointly on physiological and population levels. We studied productivity, respiration, and abundances of the symbiont-bearing foraminifer species Marginopora vertebralis on natural CO2 seeps in Papua New Guinea and conducted additional studies on production and calcification on the Great Barrier Reef (GBR) using artificially enhanced pCO2 . Net oxygen production increased up to 90% with increasing pCO2 ; temperature, light, and pH together explaining 61% of the variance in production. Production increased with increasing light and increasing pCO2 and declined at higher temperatures. Respiration was also significantly elevated (~25%), whereas calcification was reduced (16-39%) at low pH/high pCO2 compared to present-day conditions. In the field, M. vertebralis was absent at three CO2 seep sites at pHTotal levels below ~7.9 (pCO2 ~700 µatm), but it was found in densities of over 1000 m(-2) at all three control sites. The study showed that endosymbiotic algae in foraminifera benefit from increased DIC availability and may be naturally carbon limited. The observed reduction in calcification may have been caused either by increased energy demands for proton pumping (measured as elevated rates of respiration) or by stronger competition for DIC from the more productive symbionts. The net outcome of these two competing processes is that M. vertebralis cannot maintain populations under pCO2 exceeding 700 µatm, thus are likely to be extinct in the next century.

Developing an effective marine eDNA monitoring: eDNA detection at pre-outbreak densities of corallivorous seastar (Acanthaster cf. solaris).

Outbreaks of the corallivorous Crown-of-Thorns Seastar (CoTS) Acanthaster cf. solaris contribute significantly to coral reef loss. Control of outbreaks is hampered because standard monitoring techniques do not detect outbreaks at early (low density) stages, thus preventing early intervention. We previously demonstrated that eDNA monitoring can detect CoTS at intermediate densities. Here, we test whether detection probability can be improved by (i) targeted site selection or collection at specific times and (ii) moving from an average eDNA copy number approach (based on the limit of quantification) to a presence/absence approach (based on the limit of detection). Using a dataset collected over three years and multiple reef sites, we demonstrated that adding water residence age, sea surface level and temperature into generalized linear models explained low amounts of variance of eDNA copy numbers. Site specific CoTS density, by contrast, was a significant predictor for eDNA copy numbers. Bayesian multi-scale occupancy modelling of the presence/absence data demonstrated that the probability of sample capture (θ) on most reefs with intermediate or high CoTS densities was >0.8. Thus, confirming CoTS presence on these reefs would only require 2-3 samples. Sample capture decreased with decreasing CoTS density. Collecting ten filters was sufficient to reliably (based on the lower 95 % Credibility Interval) detect CoTS below nominal outbreak levels (3 Ind. ha-1). Copy number-based estimates may be more relevant to quantify CoTS at higher densities. Although water residence age did contribute little to our models, sites with higher residence times may serve as sentinel sites accumulating eDNA. The approach based on presence or absence of eDNA facilitates eDNA monitoring to detect CoTS densities below outbreak thresholds and we continue to further develop this method for quantification.