Within- and transgenerational stress legacy effects of ocean acidification on red abalone (Haliotis rufescens) growth and survival.

Understanding the mechanisms by which individual organisms respond and populations adapt to global climate change is a critical challenge. The role of plasticity and acclimation, within and across generations, may be essential given the pace of change. We investigated plasticity across generations and life stages in response to ocean acidification (OA), which poses a growing threat to both wild populations and the sustainable aquaculture of shellfish. Most studies of OA on shellfish focus on acute effects, and less is known regarding the longer term carryover effects that may manifest within or across generations. We assessed these longer term effects in red abalone (Haliotis rufescens) using a multi-generational split-brood experiment. We spawned adults raised in ambient conditions to create offspring that we then exposed to high pCO2 (1180 µatm; simulating OA) or low pCO2 (450 µatm; control or ambient conditions) during the first 3 months of life. We then allowed these animals to reach maturity in ambient common garden conditions for 4 years before returning the adults into high or low pCO2 treatments for 11 months and measuring growth and reproductive potential. Early-life exposure to OA in the F1 generation decreased adult growth rate even after 5 years especially when abalone were re-exposed to OA as adults. Adult but not early-life exposure to OA negatively impacted fecundity. We then exposed the F2 offspring to high or low pCO2 treatments for the first 3 months of life in a fully factorial, split-brood design. We found negative transgenerational effects of parental OA exposure on survival and growth of F2 offspring, in addition to significant direct effects of OA on F2 survival. These results show that the negative impacts of OA can last within and across generations, but that buffering against OA conditions at critical life-history windows can mitigate these effects.

The effect of total alkalinity on growth performance and calcification in juvenile Pacific abalone Haliotis discus hannai.

A 45-day trial was conducted to study the effect of seawater total alkalinity (TA) level up- and downregulation on the growth performance and calcification of Haliotis discus hannai Ino, while seawater pH was maintained at pHNBS = 8.1. Although seawater was not acidified, the results showed that TA downregulation caused a significant reduction (P < 0.05) in the somatic tissue growth of juvenile abalone, while TA upregulation significantly increased growth performance (P < 0.05). Similar to the impacts of pH reduction, TA downregulation also induces a decline in CO2 buffering capacity, which may be the reason why somatic tissue growth was reduced, as lowered CO2 buffering capacity was reported to shift the acid-base balancing of abalone. Parts of the periostracum layer weremissing and exposed the inner shell layers of the individuals from the TA-downregulated group. Scanning electron microscopy (SEM) results showed calcium carbonate densely deposited onto the inner shell in the control and TA-upregulated groups, while sparsely deposited calcium carbonate was observed in the TA-downregulated group. The C: N ratio in the shell of individuals from the TA-downregulated group was significantly lower than that of the other two groups, indicating that less inorganic carbon was added to the shell. As a result, abalone grew lighter and thinner shells in TA-downregulated seawater. Although seawater was not acidified, TA downregulation also caused a reduction in the calcium carbonate saturation state (Ω), which induced the erosion of the surface shell and the interruption of calcium carbonate generation. In conclusion, although seawater pH remained at ambient levels, the lowered CO2 buffering capacity and Ω induced by seawater TA downregulation also showed a detrimental effect on the growth and calcification of Pacific abalone. The impact of ocean acidification on the growth of abalone should not be assessed using only seawater pH and/or pCO2 but rather taking into account all of carbonate chemistry, particularly the CO2 buffering capacity. Abalone cultivation is suggested to be carried out in seawater with a higher level of CO2 buffering capacity and Ω, which can be achieved through integrated culture with seaweed or increasing the seawater TA level.

Effects of heating rate and shell colour on the cardiac thermal performance in a polymorphic gastropod Batillaria attramentaria.

Heating rate has gained extensive attention in mechanistic understanding of physiological responses to changing thermal conditions in the context of climate change. In polymorphic gastropods, differences in the absorption of solar energy between dark- and light-coloured individuals lead to supposable differences in their heating rates and body temperatures in sunshine. In the present study, we examined the effect of heating rate on heart rate (HR) in a polymorphic gastropod Batillaria attramentaria. By using biomimetic models, we found that daily maximum temperature of snails with a dark unbanded shell (D-type morph) was higher than snails with a white line on the upper side of each whorl (UL-type morph) by 0.6 °C when exposed to sunlight, but there was no apparent difference in heating rates between D- and UL-type models. We measured HR of snails at various heating rates from 3.0 to 9.0 °C h-1. Faster heating rates significantly increased maximum thermal tolerance in both D- and UL-type snails, highlighting the importance to have thorough knowledge on the heating rate in the field to obtain accurate maximum thermal limit of gastropods. Critical temperature at which HR precipitously declines was higher in D-type snails than UL-type snails. Our results suggested that the impacts of heating rate as well as the shell colour should be considered to gain a mechanistic understanding of the population dynamics of polymorphic gastropods.

Seawater carbonate parameters function differently in affecting embryonic development and calcification in Pacific abalone (Haliotis discus hannai).

pH or pCO2 are usually taken to study the impact of ocean acidification on molluscs. Here we studied the different impact of seawater carbonate parameters on embryonic development and calcification of the Pacific abalone (Haliotis discus hannai). Early embryonic development was susceptible to elevated pCO2 level. Larvae hatching duration was positively and hatching rate was negatively correlated with the pCO2 level, respectively. Calcium carbonate (CaCO3) deposition of larval shell was found to be susceptible to calcium carbonate saturation state (Ω) rather than pCO2 or pH. Most larvae incubated in seawater with Ωarag = 1.5 succeeded in shell formation, even when seawater pCO2 level was higher than 3700 µatm and pHT was close to 7.4. Nevertheless, larvae failed to generate CaCO3 in seawater with Ωarag ≤ 0.52 and control level of pCO2, while seawater DIC level was lowered (≤ 852 µmol/kg). Surprisingly, some larvae completed CaCO3 deposition in seawater with Ωarag = 0.6 and slightly elevated DIC (2266 µmol/kg), while seawater pCO2 level was higher than 2700 µatm and pHT was lower than 7.3. This indicates that abalone may be capable of regulating carbonate chemistry to support shell formation, however, the capability was limited as surging pCO2 level lowered growth rate and jeopardized the integrity of larval shells. Larvae generated thicker shell in seawater with Ωarag = 5.6, while adult abalone could not deposit CaCO3 in seawater with Ωarag = 0.29 and DIC = 321 µmol/kg. This indicates that abalone may lack the ability to directly remove or add inorganic carbon at the calcifying sites. In conclusion, different seawater carbonate parameters play different roles in affecting early embryonic development and shell formation of the Pacific abalone, which may exhibit limited capacity to regulate carbonate chemistry.

Metabolic and transcriptional responses demonstrating enhanced thermal tolerance in domesticated abalone.

Global warming threatens aquatic systems and organisms. Many studies have focused on the vulnerability and stress responses of aquaculture organisms to future thermal conditions. However, it may be of more practical significance to reveal their acclimation potential and mechanisms. In this study, the physiological, metabolic, and transcriptional responses to long-term temperature acclimation of northern and southern populations of Pacific abalone Haliotis discus hannai, a commercially important gastropod sensitive to environmental changes, were compared. This study conducted two common-garden experiments, including a thermostatic experiment in the lab and an aquaculture experiment on the farm. The abalone population cultured in warmer southern waters was tolerant of ongoing high temperatures, whereas the abalone population originally cultured in cooler northern waters exhibited vulnerability to high temperatures but could enhance its thermal tolerance through the process of natural selection in warmer southern waters. This difference was linked to divergence in the metabolic and transcriptional processes of the two populations. The tolerant population exhibited a greater capacity for carbohydrate and amino acid metabolism regulation and energy redistribution to cope with heat stress. This capacity may have been selected for, and accumulated, over many generations because the tolerant population originated from the intolerant population over two decades ago. This work provides insight into the vulnerability and acclimation potential of abalone to heat stress and discloses the molecular and metabolic traits underlying this phenomenon. Future research on the ability of abalone and other commercial shellfish species to acclimate to global warming should take this potential into account.

Differential effects of ocean acidification and warming on biological functioning of a predator and prey species may alter future trophic interactions.

Independently, ocean warming (OW) and acidification (OA) from increased anthropogenic atmospheric carbon dioxide are argued to be two of the greatest threats to marine organisms. Increasingly, their interaction (ocean acidification and warming, OAW) is shown to have wide-ranging consequences to biological functioning, population and community structure, species interactions and ecosystem service provision. Here, using a multi-trophic experiment, we tested the effects of future OAW scenarios on two widespread intertidal species, the blue mussel Mytilus edulis and its predator Nucella lapillus. Results indicate negative consequences of OAW on the growth, feeding and metabolic rate of M. edulis and heightened predation risk. In contrast, Nucella growth and metabolism was unaffected and feeding increased under OAW but declined under OW suggesting OA may offset warming consequences. Should this differential response between the two species to OAW, and specifically greater physiological costs to the prey than its predator come to fruition in the nature, fundamental change in ecosystem structure and functioning could be expected as trophic interactions become disrupted.

Playing it safe; risk-induced trait responses increase survival in the face of predation.

Predation risk effects are impacts on prey caused by predators that do not include consumption. These can include changes in prey behaviour, physiology, and morphology (i.e. risk-induced trait responses), which can have consequences to individual fitness and population dynamics (i.e. non-consumptive effects). While these risk-induced trait responses (RITRs) can lower individual fitness as compared to prey not exposed to risk, they are assumed to increase fitness in the presence of predators. While much work has been built upon this assumption, most evidence occurs in consumptive experiments where the trait values of consumed prey are unknown. We have little evidence showing individuals with a greater magnitude of RITR have greater survival. Here, we tested the hypothesis that RITRs increase survival in the presence of predators, but come at a cost to growth. We tested this hypothesis using Nucella lapillus as prey and Carcinus maenas as a predator and including mussels as a basal resource in a two-phase mesocosm experimental set-up. In phase 1, Nucella were placed into either a control or risk treatment (exposure to non-lethal Carcinus) for 28 days and their behaviour and growth measured. In phase 2, a lethal Carcinus was added to all mesocosms (non-lethal crabs were removed), and survival was recorded for 15 days. At the treatment (group) level, we found that Nucella exposed to predation risk in phase 1 had significantly greater risk aversion behaviour (summed score of risky vs. safe behaviour) and significantly lower growth. In phase 2, we found that Nucella exposed to predation risk had greater survival. At the individual level (regardless of treatment), we found that Nucella with greater risk aversion scores in phase 1 had significantly higher survival in phase 2 when exposed to a lethal predator, but this came at a cost to their growth. This study provides some of the first empirical evidence, at both the group and individual level, testing a long-held assumption that predation risk-induced behavioural responses increase survival in the face of direct predation, but that these responses come at a cost to the prey. These results add to our growing understanding of the benefits of RITRs to individual fitness and non-consumptive effects generally.

The paralytic shellfish toxin effect on bioenergetic constituents of the fishery resource Chorus giganteus (Gastropoda: Muricidae).

Alexandrium catenella, one of the most common harmful microalgae observed in southern Chile, produces paralytic shellfish toxins, which can affect many organisms throughout the trophic chain. This research evaluated how paralytic shellfish toxins affected the principal bioenergetic constituents and fatty acids composition of the carnivorous snail Chorus giganteus. Snails were separated into a "toxic" group that was fed the toxic clam Mulinia edulis (which was previously fed A. catenella), and a "non-toxic" group, fed non-toxic clams. Both groups were kept under these conditions for 63 days. Our results indicated no difference in the ingestion rate of toxic versus non-toxic snails; however, a higher protein level was identified in toxic snails. The total lipid content proved to be no different in toxic versus non-toxic snails; although, an effect of the toxic diet on the fatty acid profile of C. giganteus was observed. High levels of essential polyunsaturated fatty acids, especially docosahexaenoic acid (DHA) and eicosapentaenoic acid (EPA) in toxic snails, were identified. Our results suggest that exposure to paralytic shellfish toxins, through diet, may cause changes in the biochemical composition of C. giganteus, which may have a subsequent impact on its energetic physiology.

Responses of early life stages of European abalone (Haliotis tuberculata) to ocean acidification after parental conditioning: Insights from a transgenerational experiment.

CO2 absorption is leading to ocean acidification (OA), which is a matter of major concern for marine calcifying species. This study investigated the effects of simulated OA on the reproduction of European abalone Haliotis tuberculata and the survival of its offspring. Four-year-old abalone were exposed during reproductive season to two relevant OA scenarios, ambient pH (8.0) and low pH (7.7). After five months of exposure, abalone were induced to spawn. The gametes, larvae and juveniles were then exposed for five months to the same pH conditions as their parents. Several biological parameters involved in adult reproduction as well as in larval, post-larval and juvenile fitness were measured. No effects on gametes, fertilisation or larval oxidative stress response were detected. However, developmental abnormalities and significant decreases in shell length and calcification were observed at veliger stages. The expression profile of a GABA A receptor-like gene appeared to be regulated by pH, depending on larval stage. Larval and post-larval survival was not affected by low pH. However, a lower survival and a reduction of growth were recorded in juveniles at pH 7.7. Our results confirm that OA negatively impacts larval and juvenile fitness and suggest the absence of carry-over effects on abalone offspring. This may compromise the survival of abalone populations in the near future.

Effects of one-year exposure to ocean acidification on two species of abalone.

Ocean acidification (OA) resulting from the absorption of excess atmospheric CO2 by the ocean threatens the survival of marine calcareous organisms, including mollusks. This study investigated the effects of OA on adults of two abalone species (Haliotis diversicolor, a subtropical species, and Haliotis discus hannai, a temperate species). Abalone were exposed to three pCO2 conditions for 1 year (ambient, ~ 880, and ~ 1600 µatm), and parameters, including mortality, physiology, immune system, biochemistry, and carry-over effects, were measured. Survival decreased significantly at ~ 800 µatm pCO2 for H. diversicolor, while H. discus hannai survival was negatively affected only at a higher OA level (~ 1600 µatm pCO2). H. diversicolor exhibited depressed metabolic and excretion rates and a higher O:N ratio under OA, indicating a shift to lipids as a metabolism substrate, while these physiological parameters in H. discus hannai were robust to OA. Both abalone failed to compensate for the pH decrease of their internal fluids because of the lowered hemolymph pH under OA. However, the reduced hemolymph pH did not affect total hemocyte counts or tested biomarkers. Additionally, H. discus hannai increased its hemolymph protein content under OA, which could indicate enhanced immunity. Larvae produced by adults exposed to the three pCO2 levels were cultured in the same pCO2 conditions and larval deformation and shell length were measured to observe carry-over effects. Enhanced OA tolerance was observed for H. discus hannai exposed under both of the OA treatments, while that was only observed following parental pCO2 ~ 880 µatm exposure for H. diversicolor. Following pCO2 ~ 1600 µatm parental exposure, H. diversicolor offspring exhibited higher deformation and lower shell growth in all pCO2 treatments. In general, H. diversicolor were more susceptible to OA compared with H. discus hannai, suggesting that H. diversicolor could be unable to adapt to acidified oceans in the future.

Functional analysis of LFRFamide signaling in Pacific abalone, Haliotis discus hannai.

The invertebrate LFRFamide (LFRFa) and short neuropeptide F (sNPF), consisting of 6 to 10 amino acids, are orthologs for bilaterian NPF/Y, which consist of 36 to 40 amino acids. Recently, a molluscan G protein-coupled receptor (GPCR) for NPF was characterized in Pacific abalone (Haliotis discus hannai). To address the functional evolutionary route of the invertebrate LFRFa and NPF signaling system, in this study, we identified cDNAs encoding LFRFa precursors and the sNPF receptor (Hdh-sNPFR) in Pacific abalone. Four LFRFa mature peptides with 6 or 7 amino acids were predicted: GSLFRFa, GGLFRFa, GTLFRFa, and GSTLFRFa. Hdh-sNPFR was identified as a classical rhodopsin-like GPCR and classified into a molluscan sNPFR group. In HEK293 cells, Hdh-sNPFR was mainly localized in the cell membranes and internalized in the cytoplasm following treatment with LFRFa peptides. Reporter assays demonstrated that LFRFa peptides inhibit forskolin-stimulated cAMP accumulation in Hdh-sNPFR-expressing HEK293 cells. LFRFa precursor and Hdh-sNPFR transcripts were more strongly expressed in the cerebral and pleural-pedal ganglia of Pacific abalone than in the peripheral tissues such as the ovary, gills, intestine, and hepatopancreas. The levels of LFRFa transcripts in the ovary, intestine, and hepatopancreas were significantly higher in mature female abalone than in immature females. Injection of LFRFa induced the egg release and spawning behavior of mature abalone, but suppressed food intake. These results suggest that LFRFa peptides are endogenous ligands for Hdh-sNPFR involved in food intake and reproduction through a Gαi-protein dependent signaling pathway.

Mollusc shell shape as pollution biomarkers: Which is the best biological model?

Alterations in mollusc shells have been proposed contamination biomarkers. We used geometric morphometrics analyses associated with analytical determinations of contaminants to select suitable biological models among species widely distributed on coastal zones. The study was carried out using Lottia subrugosa (herbivore limpet), Crassostrea brasiliana (filter-feeder bivalve), and Stramonita brasiliensis (carnivore gastropod) obtained along a marked contamination gradient at Santos Estuarine System (Brazil). L. subrugosa and S. brasiliensis presented distinct shapes along the gradient, while no significant differences in shell form were seen for C. brasiliana. Indeed, limpets and snails presented morphometric parameters consistent with measured contamination levels hazardous substances. Based on cross-validation models, the reliability of morphometric responses was over 75% for the herbivore and carnivore species. In addition, for S. brasiliensis, a 95.2% confidence was detected in most contaminated sites. Therefore, shell alterations on carnivorous gastropods should be further investigated, seeking to be effectively employed as pollution biomarkers.

Bursting emerges from the complementary roles of neurons in a four-cell network.

Reciprocally inhibitory modules that form half-center oscillators require mechanisms for escaping or being released from inhibition. The central pattern generator underlying swimming by the nudibranch mollusc, Dendronotus iris, is composed of only four neurons that are organized into two competing modules of a half-center oscillator. In this system, bursting activity in left-right alternation is an emergent property of the network as a whole; none of the neurons produces bursts on its own. We found that the unique synaptic actions and membrane properties of the two neurons in each module (Si2 and the contralateral Si3) play complementary roles in generating stable bursting in this network oscillator. Although Si2 and Si3 each inhibits its contralateral counterpart, Si2 plays a dominant role in evoking fast and strong inhibition of the other module, the termination of which initiates postinhibitory rebound in the Si3 of that module by activating a hyperpolarization-activated inward current. Within each module, the synaptic actions and membrane properties of the two neurons complement each other: Si3 excites Si2, which then feeds back slow inhibition to Si3, terminating the burst. Using dynamic clamp, we showed that the magnitude of the slow inhibition sets the period of the oscillator. Thus, the synaptic actions of Si2 provide the hyperpolarization needed for the other module to rebound stably, whereas the membrane properties of Si3 in each module cause it to rebound first and excite Si2 to maintain the burst until terminated by the slow inhibition from Si2, which releases the other module to become active.NEW & NOTEWORTHY Half-center oscillators composed of reciprocally inhibitory neurons have been posited for over a century to underlie the production of rhythmic movements. The Dendronotus swim central pattern generator may be the simplest such circuit with only two pairs of bilaterally represented neurons. This study completes the description of the mechanism by which this network oscillator functions, showing how stable rhythmic activity arises from the complementary membrane and synaptic properties of the two neurons in the competing modules.

Multiple-stressor effects of ocean acidification, warming and predation risk cues on the early ontogeny of a rocky-shore keystone gastropod.

To understand how climate change stressors might affect marine organisms and support adequate projections it is important to know how multiple stressors may be modulated by the presence of other species. We evaluated the direct effects of ocean warming (OW) and ocean acidification (OA) together with non-consumptive effects (NCEs) of the predatory crab Acanthocyclus hassleri on early ontogeny fitness-related traits of the commercially important rocky-shore keystone gastropod Concholepas concholepas. We measured the response of nine traits to these stressors at either the organismal level (survival, growth, feeding rates, tenacity, metabolic rate, calcification rate) or sub-organismal level (nutritional status, ATP-supplying capacity, stress condition). C. concholepas survival was not affected by any of the stressors. Feeding rates were not affected by OW or OA; however, they were reduced in the presence of crab NCEs compared with control conditions. Horizontal tenacity was affected by the OA × NCEs interaction; in the presence of NCEs, OA reduced tenacity. The routine metabolic rate, measured by oxygen consumption, increased significantly with OW. Nutritional status assessment determined that carbohydrate content was not affected by any of the stressors. However, protein content was affected by the OA × NCEs interaction; in the absence of NCEs, OA reduced protein levels. ATP-supplying capacity, measured by citrate synthase (CS) activity, and cellular stress condition (HSP70 expression) were reduced by OA, with reduction in CS activity found particularly at the high temperature. Our results indicate C. concholepas traits are affected by OA and OW and the effects are modulated by predator risk (NCEs). We conclude that some C. concholepas traits are resilient to climate stressors (survival, growth, horizontal tenacity and nutritional status) but others are affected by OW (metabolic rate), OA (ATP-supplying capacity, stress condition), and NCEs (feeding rate). The results suggest that these negative effects can adversely affect the associated community.

Long-term effects of contrasting pCO2 levels on the scope for growth in the carnivorous gastropod Concholepas concholepas.

We evaluated the effect of contrasting pCO2 levels: lower (390 µatm), moderate (700 µatm) and extreme (1000 µatm), on the scope for growth of the keystone snail Concholepas concholepas over an exposure period of 6 months. Juvenile snails were collected from rocky intertidal habitats and acclimated for 5 months to those pCO2 levels. Subsequently, three groups of snails were randomly taken (n = 7 for each treatment) and reared for an additional 1 month for each of the three pCO2 levels. Physiological traits related with energy gain and energy expenditure were quantified. The scope for growth index decreased significantly with increases in pCO2, yielding negative values throughout the experimental period for the snails exposed to 1000 µatm pCO2, probably due to the extra energy required to maintain their metabolic functions in balance. This suggests that future climate change scenarios with elevated pCO2 levels could threaten the growth and other basic functions of juvenile snails of this species.

Onset of Late Cretaceous diversification in Europe's freshwater gastropod fauna links to global climatic and biotic events.

The Mesozoic rise of the European freshwater gastropod fauna is still poorly understood. Compared to the well documented Cenozoic history, little is known about the patterns and processes underlying the early diversification preceding their extinction crisis at the K-Pg boundary. We assess what is probably a first pulse of diversification of the Cenozoic-type fauna in the Late Cretaceous along with the potential abiotic and biotic controls for shifts in species diversification. We find strong support that the increase in the speciation rate in the Santonian (~ 85 Myr ago) is linked to a global sea level rise, which caused extensive flooding of continental areas and the formation of vast brackish-water ecosystems. The following decline of the speciation rate coincides with a rise in diversity and reflects increasing interspecific competition. The peak in the speciation rate postdates the Cenomanian-Turonian Thermal Maximum, which probably limited the potential for diversification among freshwater gastropods due to ecological constraints. The peak coincides moreover with the end phase of the Cretaceous Terrestrial Revolution, which sparked the radiation of angiosperms. The expansion and diversification of flowering plants, being an important food source for freshwater gastropods today, could have formed a necessary basis for gastropod diversification.

Weapons or deterrents? Nudibranch molluscs use distinct ecological modes of chemical defence against predators.

Defensive chemicals are used by plants and animals to reduce the risk of predation through different mechanisms, including toxins that cause injury and harm (weapons) and unpalatable or odiferous compounds that prevent attacks (deterrents). However, whether effective defences are both toxins and deterrents, or work in just one modality is often unclear. In this study, our primary aim was to determine whether defensive compounds stored by nudibranch molluscs acted as weapons (in terms of being toxic), deterrents (in terms of being distasteful) or both. Our secondary aim was to investigate the response of different taxa to these defensive compounds. To do this, we identified secondary metabolites in 30 species of nudibranch molluscs and investigated their deterrent properties using antifeedant assays with three taxa: rock pool shrimp, Palaemon serenus, and two fish species: triggerfish Rhinecanthus aculeatus and toadfish Tetractenos hamiltoni. We compared these results to toxicity assays using brine shrimp Artemia sp. and previously published toxicity data with a damselfish Chromis viridis. Overall, we found no clear relationship between palatability and toxicity, but instead classified defensive compounds into the following categories: Class I & II-highly unpalatable and highly toxic; Class I-weakly unpalatable and highly toxic; Class II-highly unpalatable but weakly toxic; WR (weak response)-weakly unpalatable and weakly toxic. We also found eight extracts from six species that did not display activity in any assays indicating they may have very limited chemical defensive mechanisms (NR, no response). We found that the different classes of secondary metabolites were similarly unpalatable to fish and shrimp, except extracts from Phyllidiidae nudibranchs (isonitriles) that were highly unpalatable to shrimp but weakly unpalatable to fish. Our results pave the way towards better understanding how animal chemical defences work against a variety of predators. We highlight the need to disentangle weapons and deterrents in future work on anti-predator defences to better understand the foraging decisions faced by predators, the resultant selection pressures imposed on prey and the evolution of different anti-predator strategies.

Combined effects of ocean warming and acidification on the larval stages of the European abalone Haliotis tuberculata.

This study examined the physiological responses of the larval stages of Haliotis tuberculata, an economically important abalone, to combined temperature (17 °C and 19 °C) and pH (ambient pH and -0.3 units, i.e., +200% increase in seawater acidity) in a full factorial experiment. Tissue organogenesis, shell formation, and shell length significantly declined due to low pH. High temperature significantly increased the proportion of fully shelled larvae at 24 h post-fertilization (hpf), but increased the proportion of unshelled larvae at 72 hpf. Percentage of swimming larvae at 24 hpf, 72 hpf and 96 hpf significantly declined due to high temperature, but not because of low pH. Larval settlement increased under high temperature, but was not affected by low pH. Despite the fact that no interaction between temperature and pH was observed, the results provide additional evidence on the sensitivity of abalone larvae to both low pH and high temperature. This may have negative consequences for the persistence of abalone populations in natural and aquaculture environments in the near future.

Finite element analysis relating shape, material properties, and dimensions of taenioglossan radular teeth with trophic specialisations in Paludomidae (Gastropoda).

The radula, a chitinous membrane with embedded tooth rows, is the molluscan autapomorphy for feeding. The morphologies, arrangements and mechanical properties of teeth can vary between taxa, which is usually interpreted as adaptation to food. In previous studies, we proposed about trophic and other functional specialisations in taenioglossan radulae from species of African paludomid gastropods. These were based on the analysis of shape, material properties, force-resistance, and the mechanical behaviour of teeth, when interacting with an obstacle. The latter was previously simulated for one species (Spekia zonata) by the finite-element-analysis (FEA) and, for more species, observed in experiments. In the here presented work we test the previous hypotheses by applying the FEA on 3D modelled radulae, with incorporated material properties, from three additional paludomid species. These species forage either on algae attached to rocks (Lavigeria grandis), covering sand (Cleopatra johnstoni), or attached to plant surface and covering sand (Bridouxia grandidieriana). Since the analysed radulae vary greatly in their general size (e.g. width) and size of teeth between species, we additionally aimed at relating the simulated stress and strain distributions with the tooth sizes by altering the force/volume. For this purpose, we also included S. zonata again in the present study. Our FEA results show that smaller radulae are more affected by stress and strain than larger ones, when each tooth is loaded with the same force. However, the results are not fully in congruence with results from the previous breaking stress experiments, indicating that besides the parameter size, more mechanisms leading to reduced stress/strain must be present in radulae.

Origin and significance of two pairs of head tentacles in the radiation of euthyneuran sea slugs and land snails.

The gastropod infraclass Euthyneura comprises at least 30,000 species of snails and slugs, including nudibranch sea slugs, sea hares and garden snails, that flourish in various environments on earth. A unique morphological feature of Euthyneura is the presence of two pairs of sensory head tentacles with different shapes and functions: the anterior labial tentacles and the posterior rhinophores or eyestalks. Here we combine molecular phylogenetic and microanatomical evidence that suggests the two pairs of head tentacles have originated by splitting of the original single tentacle pair (with two parallel nerve cords in each tentacle) as seen in many other gastropods. Minute deep-sea snails of Tjaernoeia and Parvaplustrum, which in our phylogeny belonged to the euthyneurans' sister group (new infraclass Mesoneura), have tentacles that are split along much of their lengths but associated nerves and epidermal sense organs are not as specialized as in Euthyneura. We suggest that further elaboration of cephalic sense organs in Euthyneura closely coincided with their ecological radiation and drastic modification of body plans. The monotypic family Parvaplustridae nov., superfamily Tjaernoeioidea nov. (Tjaernoeiidae + Parvaplustridae), and new major clade Tetratentaculata nov. (Mesoneura nov. + Euthyneura) are also proposed based on their phylogenetic relationships and shared morphological traits.