The impact of aragonite saturation variability on shelled pteropods: An attribution study in the California Current System.

Observations from the California Current System (CalCS) indicate that the long-term trend in ocean acidification (OA) and the naturally occurring corrosive conditions for the CaCO3 mineral aragonite (saturation state Ω < 1) have a damaging effect on shelled pteropods, a keystone group of calcifying organisms in the CalCS. Concern is heightened by recent findings suggesting that shell formation and developmental progress are already impacted when Ω falls below 1.5. Here, we quantify the impact of low Ω conditions on pteropods using an individual-based model (IBM) with life-stage-specific mortality, growth, and behavior in a high-resolution regional hindcast simulation of the CalCS between 1984 and 2019. Special attention is paid to attributing this impact to different processes that lead to such low Ω conditions, namely natural variability, long-term trend, and extreme events. We find that much of the observed damage in the CalCS, and specifically >70% of the shell CaCO3 loss, is due to the pteropods' exposure to naturally occurring low Ω conditions as a result of their diel vertical migration (DVM). Over the hindcast period, their exposure to damaging waters (Ω < 1.5) increases from 9% to 49%, doubling their shell CaCO3 loss, and increasing their mortality by ~40%. Most of this increased exposure is due to the shoaling of low Ω waters driven by the long-term trend in OA. Extreme OA events amplify this increase by ~40%. Our approach can quantify the health of pteropod populations under shifting environmental conditions, and attribute changes in fitness or population structure to changes in the stressor landscape across hierarchical time scales.

The Neuropeptide HGAP Regulates Growth, Reproduction, Metamorphosis, Tissue Damage Repair, and Response against Starvation in Pacific Abalone.

INTRODUCTION: Neuropeptides regulate vital physiological processes in multicellular organisms, including growth, reproduction, metamorphosis, and feeding. Recent transcriptome analyses have revealed neuropeptide genes with potential roles in vertebrate and invertebrate growth and reproduction. Among these genes, haliotid growth-associated peptide (HGAP) was identified as a novel gene in abalone. METHODS: This study focused on HGAP in Pacific abalone (Haliotis discus hannai), where the complete cDNA sequence named Hdh-HGAP was identified and characterized. Samples from different experiments, such as metamorphosis, juvenile abalone growth, gonad development stages, muscle remodeling, and starvation, were collected for mRNA expression analysis. RESULTS: The sequence spans 552 bp, encoding 96 amino acids with a molecular weight of 10.96 kDa. Expression analysis revealed that Hdh-HGAP exhibited higher levels in muscle tissue. Notably, during metamorphosis, Hdh-HGAP exhibited greater expression in the trochophore, veliger, and juvenile stages than in the cell division stages. Regarding growth patterns, Hdh-HGAP was highly expressed during rapid growth compared to stunted, minimal, and normal growth. In gonadal development, Hdh-HGAP mRNA reached its highest expression level during the ripening stage, indicating a potential role in gonadal cell proliferation and maturation. The in vivo effects of GnRH on gonad development and the expression of the Hdh-HGAP neuropeptide indicate its involvement in regulating reproduction in Pacific abalone. While tissue remodeling is primarily governed by immune genes, Hdh-HGAP was also upregulated during muscle tissue remodeling. Conversely, Hdh-HGAP was downregulated during prolonged starvation. CONCLUSION: This study marks the first comprehensive exploration of the Hdh-HGAP neuropeptide gene in Pacific abalone, shedding light on its involvement in growth, reproduction, metamorphosis, tissue remodeling, and response to starvation, although regulatory mechanisms are mostly unknown.

Characterization of myoinhibitory peptide signaling system and its implication in larval metamorphosis and spawning behavior in Pacific abalone.

Myoinhibitory peptides (MIPs) affect various physiological functions, including juvenile hormone signaling, muscle contraction, larval development, and reproduction in invertebrates. Although MIPs are ligands for MIP and/or sex peptide receptors (MIP/SPRs) in diverse arthropods and model organisms belonging to Lophotrochozoa, the MIP signaling system has not yet been fully investigated in mollusks. In this study, we identified the MIP signaling system in the Pacific abalone Haliotis discus hannai (Hdh). Similar to the invertebrate MIPs, a total of eight paracopies of MIPs (named Hdh-MIP1 to Hdh-MIP8), harboring a WX5-7Wamide motif, except for Hdh-MIP2, were found in the Hdh-MIP precursor. Furthermore, we characterized a functional Hdh-MIPR, which responded to the Hdh-MIPs, except for Hdh-MIP2, possibly linked with the PKC/Ca2+ and PKA/cAMP signaling pathways. Hdh-MIPs delayed larval metamorphosis but increased the spawning behavior. These results suggest that the Hdh-MIP signaling system provides insights into the unique function of MIP in invertebrates.

Dorsoventral decoupling of Hox gene expression underpins the diversification of molluscs.

In contrast to the Hox genes in arthropods and vertebrates, those in molluscs show diverse expression patterns with differences reported among lineages. Here, we investigate 2 phylogenetically distant molluscs, a gastropod and a polyplacophoran, and show that the Hox expression in both species can be divided into 2 categories. The Hox expression in the ventral ectoderm generally shows a canonical staggered pattern comparable to the patterns of other bilaterians and likely contributes to ventral patterning, such as neurogenesis. The other category of Hox expression on the dorsal side is strongly correlated with shell formation and exhibits lineage-specific characteristics in each class of mollusc. This generalized model of decoupled dorsoventral Hox expression is compatible with known Hox expression data from other molluscan lineages and may represent a key characteristic of molluscan Hox expression. These results support the concept of widespread staggered Hox expression in Mollusca and reveal aspects that may be related to the evolutionary diversification of molluscs. We propose that dorsoventral decoupling of Hox expression allowed lineage-specific dorsal and ventral patterning, which may have facilitated the evolution of diverse body plans in different molluscan lineages.

Evolved differences in energy metabolism and growth dictate the impacts of ocean acidification on abalone aquaculture.

Ocean acidification (OA) poses a major threat to marine ecosystems and shellfish aquaculture. A promising mitigation strategy is the identification and breeding of shellfish varieties exhibiting resilience to acidification stress. We experimentally compared the effects of OA on two populations of red abalone (Haliotis rufescens), a marine mollusc important to fisheries and global aquaculture. Results from our experiments simulating captive aquaculture conditions demonstrated that abalone sourced from a strong upwelling region were tolerant of ongoing OA, whereas a captive-raised population sourced from a region of weaker upwelling exhibited significant mortality and vulnerability to OA. This difference was linked to population-specific variation in the maternal provisioning of lipids to offspring, with a positive correlation between lipid concentrations and survival under OA. This relationship also persisted in experiments on second-generation animals, and larval lipid consumption rates varied among paternal crosses, which is consistent with the presence of genetic variation for physiological traits relevant for OA survival. Across experimental trials, growth rates differed among family lineages, and the highest mortality under OA occurred in the fastest growing crosses. Identifying traits that convey resilience to OA is critical to the continued success of abalone and other shellfish production, and these mitigation efforts should be incorporated into breeding programs for commercial and restoration aquaculture.

Evolution and biomineralization of pteropod shells.

Shelled pteropods, known as sea butterflies, are a group of small gastropods that spend their entire lives swimming and drifting in the open ocean. They build thin shells of aragonite, a metastable polymorph of calcium carbonate. Pteropod shells have been shown to experience dissolution and reduced thickness with a decrease in pH and therefore represent valuable bioindicators to monitor the impacts of ocean acidification. Over the past decades, several studies have highlighted the striking diversity of shell microstructures in pteropods, with exceptional mechanical properties, but their evolution and future in acidified waters remains uncertain. Here, we revisit the body-of-work on pteropod biomineralization, focusing on shell microstructures and their evolution. The evolutionary history of pteropods was recently resolved, and thus it is timely to examine their shell microstructures in such context. We analyse new images of shells from fossils and recent species providing a comprehensive overview of their structural diversity. Pteropod shells are made of the crossed lamellar and prismatic microstructures common in molluscs, but also of curved nanofibers which are proposed to form a helical three-dimensional structure. Our analyses suggest that the curved fibres emerged before the split between coiled and uncoiled pteropods and that they form incomplete to multiple helical turns. The curved fibres are seen as an important trait in the adaptation to a planktonic lifestyle, giving maximum strength and flexibility to the pteropod thin and lightweight shells. Finally, we also elucidate on the candidate biomineralization genes underpinning the shell diversity in these important indicators of ocean health.

Effective accumulative temperature affects gonadal maturation by controlling expression of GnRH, GnRH receptor, serotonin receptor and APGWamide gene in Pacific abalone, Haliotis discus hannai during broodstock conditioning in hatcheries.

Water temperature is a crucial environmental factor that influences reproductive function of abalone. Broodstock conditioning exposed to effective accumulative temperature (EAT) is a common practice in abalone hatcheries. To understand the molecular mechanism underlying the regulation of gonadal maturation and reproduction of Haliotis discus hannai exposed to EAT and induced spawning period, changes in expression of neuroendocrine genes encoding two gonadotropin releasing hormone (Hdh-GnRH, GnRH-like peptide), GnRH receptor (HdhGnRH-R), serotonin receptor (5-HTHdh) and Hdh-APGWamide in neural ganglia and gonadal tissues were examined. Gonadosomatic index (GSI) was significantly increased with increasing EAT °C-days. Expression levels of Hdh-GnRH, GnRH-like peptide, HdhGnRH-R, 5-HTHdh and Hdh-APGWamide mRNA were significantly increased with increasing EAT °C-days in ganglion (where the gene synthesized) and gonadal tissues. The significant increase in mRNA expression of each examined gene started from EAT 500 to 750°C-days, reached an initial peak at 1000°C-days, suggesting gonadal maturation started from the onset of EAT and slowly continued until 750°C-days, then at 1000°C-days reached to initial peak developmental period. The maturation reached to spawning state at 1000°C-days and peaked at 1500°C-days. Hdh-GnRH showed significantly higher mRNA expression in pleuropedal ganglion and branchial ganglion, whereas GnRH like peptide showed higher expression in cerebral ganglion, and HdhGnRH-R, 5-HTHdh and Hdh-APGWamide showed higher expression in pleuropedal ganglion. All genes were expressed higher at higher EAT °C-days. During induced spawning period, higher mRNA expression of examined genes was observed at the time of spawning; however, a sharp decrease occurred after spawning, suggesting that these genes are involved in spawning activities. Taken together, these results indicate that an increase of EAT °C-days can increase expression of neuroendocrine genes and enhance gonadal maturation. Besides all these genes are involved in the process of spawning induction, and increase of GSI has a positive correlation with the increase of gene expression.

Effects of Antifreeze Protein III on Sperm Cryopreservation of Pacific Abalone, Haliotis discus hannai.

Pacific abalone (Haliotis discus hannai) is a highly commercial seafood in Southeast Asia. The aim of the present study was to improve the sperm cryopreservation technique for this valuable species using an antifreeze protein III (AFPIII). Post-thaw sperm quality parameters including motility, acrosome integrity (AI), plasma membrane integrity (PMI), mitochondrial membrane potential (MMP), DNA integrity, fertility, hatchability, and mRNA abundance level of heat shock protein 90 (HSP90) were determined to ensure improvement of the cryopreservation technique. Post-thaw motility of sperm cryopreserved with AFPIII at 10 µg/mL combined with 8% dimethyl sulfoxide (DMSO) (61.3 ± 2.7%), 8% ethylene glycol (EG) (54.3 ± 3.3%), 6% propylene glycol (PG) (36.6 ± 2.6%), or 2% glycerol (GLY) (51.7 ± 3.0%) was significantly improved than that of sperm cryopreserved without AFPIII. Post-thaw motility of sperm cryopreserved with 2% MeOH and 1 µg/mL of AFPIII was also improved than that of sperm cryopreserved without AFPIII. A combination of 10 µg/mL AFPIII with 8% DMSO resulted in the highest post-thaw motility, showing AI of 60.1 ± 3.9%, PMI of 67.2 ± 4.0%, and MMP of 59.1 ± 4.3%. DNA integrity of sperm cryopreserved using 10 µg/mL AFPIII combined with 8% DMSO was not significantly (p > 0.05) different from that of fresh sperm. Cryopreservation using a combination of AFPIII with 8% DMSO improved fertilization and hatching rates of sperm compared to that of cryopreservation without supplementation of 10 µg/mL AFPIII. Sperm cryopreserved using AFPIII showed higher mRNA abundance levels of HSP90 than those cryopreserved without AFPIII. Results of the present study suggest that 10 µg/mL AFPIII combined with 8% DMSO can be used for large scale cryopreservation of Pacific abalone sperm and for hatchery production.

One species, two developmental modes: a case of geographic poecilogony in marine gastropods.

BACKGROUND: Poecilogony, the presence of two developmental modes in the same animal species, is a rare phenomenon. Few cases of poecilogony have been suggested for marine invertebrates including molluscs and even less stood extensive testing, mostly revealing a species pair with differing developmental modes. We studied a textbook example of poecilogony in the viviparous snail Planaxis sulcatus (Gastropoda: Planaxidae), for the first time throughout its entire distribution range. RESULTS: In the Western Indian Ocean and Red Sea this intertidal species is observed to have large, shelled juveniles, whereas in the Indo-West Pacific planktotrophic veliger larvae are released from a subhaemocoelic brood pouch. We uncovered a shift in developmental modes across its range: from west to east successively earlier developmental stages are released. Furthermore, genetic data based on mitochondrial DNA suggests to recognize P. sulcatus as a single species rather than a group of cryptic species. A reconstruction of the ancestral area of P. sulcatus based on molecular data outlines the Western Indian Ocean and the Indo-West Pacific as area of origin. CONCLUSION: The findings supporting Planaxis sulcatus as a single widespread species and the geographical shift from one reproductive mode to another suggest for this species to truly represent a case of geographic poecilogony, i.e. differing developmental modes between populations of the same species. Furthermore, the results of our ancestral range estimation imply the release of planktotrophic larvae as the ancestral developmental mode.

Over high or low dietary protein levels depressed the growth, TOR signaling, apoptosis, immune and anti-stress of abalone Haliotis discus hannai.

A 120-day feeding trial was conducted to investigate the effects of relative higher and lower dietary protein levels on the growth, immunity and anti-stress of abalone Haliotis discus hannai fed diets with 17.64% (low), 30.49% (normal) and 43.27% (high) of proteins, respectively. The results showed that compared with 30.49% of dietary protein, 17.64% and 43.27% of dietary protein levels significantly decreased the weight gain rate and the activities of α-amylase, trypsin, alanine aminotransferase and aspartate aminotransferase in the hepatopancreas and serum of abalone (P < 0.05). Abalone fed 30.49% of dietary protein had the highest activity of superoxidase, acid phosphatase, alkaline phosphatase, lysozyme and the total anti-oxidative capacity, and the lowest content of malondialdehyde in the serum and hepatopancreas (P < 0.05). The gene expressions of TOR, S6k, Bcl-2, IκB, NfκB, TNF-α and Nrf2 were significantly up-regulated in the group with 30.49% of dietary protein (P < 0.05). Pathological abnormalities in hepatocyte cells of abalone were found in the groups with 17.64% and 43.27% of dietary protein. Meanwhile, accumulative mortalities of abalone after the Vibrio parahaemolyticus challenge test and heat stress test were significantly increased within these two groups (P < 0.05). In conclusion, the excessive (43.27) or deficient (17.64) dietary protein levels depressed the growth and immunity of abalone. Combined with the stress tests results, 17.63% or 43.27% of dietary protein contents are not recommended to the abalone facing the stress of vibriosis or high-water temperature (≥28 °C).

Effects of Bacillus lincheniformis feeding frequency on the growth, digestion and immunity of Haliotis discus hannai.

To study the effects of Bacillus lincheniformis feeding frequency on the survival and growth of Haliotis discus hannai abalone, we measured the expression levels of nonspecific immune genes and monitored the anti-Vibrio parahaemolyticus immune reaction. H. discus hannai (shell length: 32.75 ± 2.63 mm, body weight: 4.91 ± 0.34 g) was selected to perform a 70 d laboratory culture experiment including a 14 d V. parahaemolyticus artificial infection experiment. The control group (C) was fed normal commercial feed every day. The M1 experimental group was given experimental feed and basal feed on alternating days until the end of the experiment. The M2 experimental group was given experimental feed for 4 d and basal feed for 3 d, and this cycle was repeated every 7 d until the end of the experiment. The M3 experimental group was given experimental feed for 2 d and basal feed for 5 d, and this cycle was repeated every 7 d until the end of the experiment. The M4 group was continuously given experimental feed for the duration of the experiment. The concentration of added B. lincheniformis in each experimental group was 105 cfu/g (according to the quantity of viable bacteria). The specific growth rate (as measured by body weight) and the feed conversion efficiency of the abalone in M1 and M2 were significantly higher than those in M4 and C (P < 0.05). The cellulose and lipase activities of abalone in M1, M2 or M4 were significantly higher than those in M3 or C (P < 0.05). The acid phosphatase, superoxide dismutase, total haemocyte counts, O2- levels generated by respiratory bursts, and the expression levels of Mn-SOD, TPx, GSTs and GSTm in abalone in the M2 group were significantly higher than those in any other feeding frequency group (P < 0.05). At the end of the V. parahaemolyticus infection, the cumulative mortality of the abalone in M2 was significantly lower than that in any other group (P < 0.05). Consequently, given the growth advantages and the enhancement of immune function, the feeding plan in which B. lincheniformis was applied for 4 d per week, and basal feed was then applied for 3 d, did not lead to a high level of immune reaction, immune fatigue or waste of resources, but increased the growth rate of individuals and their resistance to V. parahaemolyticus infection.

Eobania vermiculata as a potential indicator of nitrate contamination in soil.

The effects of nitrates were analysed on the land snail Eobania vermiculata, a good bioindicator to assess the effects of certain pollutants in soil. It is known that the molluscs are very sensitive to contamination substances and can be used as sentinel organism for environmental pollution assessment. The nitrates are present in fertilizers and in food additives and their excess can not only be harmful to the environment but also dangerous for the humans. Indeed, in the mammals the nitrates are converted into nitrites and can cause a series of complications as the formation of methaemoglobin and cancers. In this study, adult organisms of E. vermiculata were exposed to soil containing 2000 mg/L of nitrates for 30 days to evaluate the stool microbiome and the histological changes at the level of the foot. Eggs of these snails were similarly treated to observe their hatching, survival and development. Histological changes were observed at level of the foot of adult snails exposed to nitrate and in their stools was evident an increase of bacteria, especially those that have a high ability to exploit nitrates and nitrogen as nutrients. Instead, the treated eggs showed changes in hatching, hypopigmentation of newborn snails and a decrease of their survival in time. The overall information obtained from these endpoints can provide important information regarding the quality of the environment. In addition, they also showed that the invertebrate organism E. vermiculata despite being a simple organism is very useful and efficient for ecotoxicological studies.

Imposex Incidence in Gastropod Species from Santa Marta Coastal Zone, Colombian Caribbean Sea.

Imposex is a phenomenon widely associated with environmental exposure to organotin compounds which were quite common components of antifouling paints applied on boats and ship hulls. Here we study the incidence of imposex in neogastropods and its relation with water quality and maritime traffic in the coastal strip of Santa Marta, Colombia. Imposex was determined via specialized indexes and related to the organisms' size, somatic conditions, variables of water quality and maritime traffic, in a space-time assessment. There was evidence of imposex in five species Plicopurpura patula, Vasula deltoidea, Stramonita haemastoma, S. floridana, and Gemophos auritulus. Purpura patula and Vasula deltoidea species were found in all sampling sites. The results have proved that imposex is highly influenced by the maritime traffic variable, with greater prevalence during the dry season, and with P. patula being more sensitive than V. deltoidea.

Transcriptome analysis of air-breathing land slug, Incilaria fruhstorferi reveals functional insights into growth, immunity, and reproduction.

BACKGROUND: Incilaria (= Meghimatium) fruhstorferi is an air-breathing land slug found in restricted habitats of Japan, Taiwan and selected provinces of South Korea (Jeju, Chuncheon, Busan, and Deokjeokdo). The species is on a decline due to depletion of forest cover, predation by natural enemies, and collection. To facilitate the conservation of the species, it is important to decide on a number of traits related to growth, immunity and reproduction addressing fitness advantage of the species. RESULTS: The visceral mass transcriptome of I. fruhstorferi was enabled using the Illumina HiSeq 4000 sequencing platform. According to BUSCO (Benchmarking Universal Single-Copy Orthologs) method, the transcriptome was considered complete with 91.8% of ortholog genes present (Single: 70.7%; Duplicated: 21.1%). A total of 96.79% of the raw read sequences were processed as clean reads. TransDecoder identified 197,271 contigs that contained candidate-coding regions. Of a total of 50,230 unigenes, 34,470 (68.62% of the total unigenes) annotated to homologous proteins in the Protostome database (PANM-DB). The GO term and KEGG pathway analysis indicated genes involved in metabolism, phosphatidylinositol signalling system, aminobenzoate degradation, and T-cell receptor signalling pathway. Many genes associated with molluscan innate immunity were categorized under pathogen recognition receptor, TLR signalling pathway, MyD88 dependent pathway, endogenous ligands, immune effectors, antimicrobial peptides, apoptosis, and adaptation-related. The reproduction-associated unigenes showed homology to protein fem-1, spermatogenesis-associated protein, sperm associated antigen, and testis expressed sequences, among others. In addition, we identified key growth-related genes categorized under somatotrophic axis, muscle growth, chitinases and collagens. A total of 4822 Simple Sequence Repeats (SSRs) were also identified from the unigene sequences of I. fruhstorferi. CONCLUSIONS: This is the first available genomic information for non-model land slug, I. fruhstorferi focusing on genes related to growth, immunity, and reproduction, with additional focus on microsatellites and repeating elements. The transcriptome provides access to greater number of traits of unknown relevance in the species that could be exploited for in-depth analyses of evolutionary plasticity and making informed choices during conservation planning. This would be appropriate for understanding the dynamics of the species on a priority basis considering the ecological, health, and social benefits.

Spermatogenesis and lobular cyst type of testes organization in marine gastropod Littorina saxatilis (Olivi 1792).

Although individual stages of spermatogenesis in Littorina saxatilis are well studied at the electron microscopic level, the gonad structure and the spatial localization of gametes at different stages of maturation remain unclear. Using differential-interference contrast (DIC) for observations of fresh tissue we show that the mature testis consists of numerous ovoid lobules forming larger lobes. The lobules of intact mature testes of L. saxatilis are filled with randomly arranged multicellular cysts containing gametes at different stages of maturation. Gametes within a cyst are highly synchronized in respect of the differentiation degree. At the same time, no spatial gradient in the arrangement of cysts according to the maturation degree of gametes in them was observed in any of the studied lobules. The male gonads contain cysts with early spermatids, mid, late spermatids, and spermatozoa. Using silver-staining, DAPI, and chromomycin A3 (CMA3) staining, we identify 20 main types of nucleus organization in differentiating sperm. Premature and mature male gonads contain cysts with a mosaic arrangement as well as rare solitary cyst cells, goniablast cysts, or separate spermatogonia in between them. Our data indicate that the testis structure in L. saxatilis cannot be attributed to the tubular type, as previously thought. It corresponds to the lobular cyst type but individual lobules contain cysts with gametes at the same stage of development. It is similar to the testis structure of several fishes, amphibians, and Drosophila melanogaster. This type of the gonad organization has never been described in gastropods before.

Effects of the probiotic Bacillus amyloliquefaciens on the growth, immunity, and disease resistance of Haliotis discus hannai.

The effects of a diet containing the probiotic Bacillus amyloliquefaciens on the survival and growth of Haliotis discus hannai were evaluated by measuring growth and hematological parameters and the expression levels of nonspecific immune genes. In addition, the abalone's response to Vibrio parahaemolyticus infection was assessed. H. discus hannai (shell length: 29.35 ±â€¯1.81 mm, body weight: 4.28 ±â€¯0.23 g) were exposed to an 8-week culture experiment in indoor aquariums and a 2-week V. parahaemolyticus artificial infection experiment. In each experiment, the control group (C) was fed daily with the basal feed; the experimental groups were fed daily with the experimental feed, prepared by spraying B. amyloliquefaciens onto the basal feed at final concentrations of 103 (group A1), 105 (A2), and 107 (A3) cfu/g. The survival rate, body weight specific growth rate, and food conversion efficiency in A2 and A3 were significantly higher than those in A1 and C (P < 0.05). The total number of blood lymphocytes, the O2- and NO levels produced from respiratory burst, the activities of acid phosphatase, superoxide dismutase, and catalase, and the expression levels of catalase and thiol peroxidase in A2 were not significantly different from those in A3, but these factors were significantly higher in A2 compared to A1 and C (P < 0.05). The total antioxidant capacity and expression levels of glutathione S-transferase in A1, A2 and A3 were significantly higher than those in C (P < 0.05). At day 9 after infection with V. parahaemolyticus, all abalone in C were dead; at the end of the experiment, the cumulative mortality of abalone in A2 was significantly lower than that in any other group (P < 0.05). Thus, the experimental feed containing 105 cfu/g B. amyloliquefaciens not only facilitated the food intake and growth of abalone, but also effectively enhanced their non-specific immunity and resistance to V. parahaemolyticus infection. In this regard, B. amyloliquefaciens may be a useful probiotic strain for abalone aquaculture.

Mannan oligosaccharide increases the growth performance, immunity and resistance capability against Vibro Parahemolyticus in juvenile abalone Haliotis discus hannai Ino.

This trial was conducted to investigate the effect of mannose oligosaccharides (MOS) on the growth performance, antioxidation, immunity and disease resistance of Vibro Parahemolyticus in juvenile abalone Haliotis discus hannai Ino. Four formulated diets were produced to contain 0.00 g/kg, 0.40 g/kg, 0.80 g/kg and 1.60 g/kg Actigen®, with functional ingredients of MOS, respectively. Accordingly, the experimental diets were named as A0, A4, A8 and A16. After 120-days feeding trial, the best growth performance was observed in A8 group (P < 0.05) and there was no significant difference in A0, A4 and A16 groups. With the increase of dietary MOS, the activity of the total antioxidant capacity in hepatopancreas is increasingly elevated (P < 0.05) while no significant difference was observed on activity of glutathione S-transferase (P > 0.05). The activities of superoxide dismutase and glutathione peroxidase were firstly increased and then decreased, with the highest values in A8 group (P < 0.05). Immune-related parameters were significantly affected by dietary MOS inclusion. Specifically, the activities of alkaline phosphatase and acid phosphatase in hepatopancreas and serum of abalone fed diets containing MOS were significantly higher than those of control A0 group (P < 0.05). Moreover, the highest values of both enzymes were observed in hepatopancreas of A8 group but in serum of A16 group, respectively. The lysozyme activities in hepatopancreas and serum of A4 group were significantly higher than those of other groups (P < 0.05) and there was no significant difference in A0, A8 and A16 groups (P > 0.05). The activities of cytophagy and respiratory burst in serum of abalone were not significantly affected by dietary MOS content (P > 0.05). The mRNA levels of focal adhesion kinase and integrin-linked kinase were gradually elevated with the increase of dietary MOS, with the highest value recorded in A16 group (P < 0.05). The gene expression of caspse-3 in A8 group was dramatically higher than those of other groups (P < 0.05) and there was no significant difference in A0, A4 and A16 groups (P > 0.05). The mRNA level of nuclear factor-κB was not significantly affected by dietary MOS (P > 0.05). During 56 h of V. Parahemolyticus challenge period, the accumulated mortality rate of abalone fed diets containing MOS were significantly lower than that of control A0 group in each time point (P < 0.05). Overall, the lowest rate was happened in A8 group (P < 0.05). In conclusion, MOS inclusion in diet has obviously positive effect on growth, immunity and disease resistance capability of abalone, with the optimal level of Actigen® at 0.80 g/kg in diet.

Development of Cerata in the Cladobranchian Sea Slug Pteraeolidia semperi (Mollusca: Gastropoda: Nudibranchia).

Cladobranchian sea slugs are characterized by a number of dorsal projections, called "cerata," which are presumably involved in such biological functions as kleptocnidal defense, gas exchange, and symbiotic photosynthesis. Here, we investigated the developmental pattern of ceras formation in a cladobranchian, Pteraeolidia semperi, using field-collected individuals at various postembryonic developmental stages. As the body length increased, the total number of cerata increased in a logistic manner, up to 280 per individual. On the dorsal side of the body, the cerata exhibited a conspicuous formation of repeated, laterally-paired clusters, or rows, along the antero-posterior axis of the animals. As the body length increased, the number of ceras rows increased in a logistic manner, reaching a plateau at around 15 rows per individual. Two types of ceras clusters were observed: well-developed ceras clusters forming a glove-like structure with a basal bulge, which tended to be found in larger animals and at the anterior body region, and less-developed ceras clusters without the bulge, which tended to be found in smaller animals and at the posterior body region. Statistical and simulation analyses suggested that bulge formation underlies increased ceras number, even after the plateaued formation of new ceras rows. These results indicate that, in the postembryonic development of P. semperi, the increase of dorsal cerata entails the following processes: (i) increase of the number of ceras rows, (ii) formation of the basal bulge in each ceras cluster, and (iii) increase of the number of cerata per ceras cluster.

Quantifying dispersal from hydrothermal vent fields in the western Pacific Ocean.

Hydrothermal vent fields in the western Pacific Ocean are mostly distributed along spreading centers in submarine basins behind convergent plate boundaries. Larval dispersal resulting from deep-ocean circulations is one of the major factors influencing gene flow, diversity, and distributions of vent animals. By combining a biophysical model and deep-profiling float experiments, we quantify potential larval dispersal of vent species via ocean circulation in the western Pacific Ocean. We demonstrate that vent fields within back-arc basins could be well connected without particular directionality, whereas basin-to-basin dispersal is expected to occur infrequently, once in tens to hundreds of thousands of years, with clear dispersal barriers and directionality associated with ocean currents. The southwest Pacific vent complex, spanning more than 4,000 km, may be connected by the South Equatorial Current for species with a longer-than-average larval development time. Depending on larval dispersal depth, a strong western boundary current, the Kuroshio Current, could bridge vent fields from the Okinawa Trough to the Izu-Bonin Arc, which are 1,200 km apart. Outcomes of this study should help marine ecologists estimate gene flow among vent populations and design optimal marine conservation plans to protect one of the most unusual ecosystems on Earth.

Aging and retinoid X receptor agonists on masculinization of female Pomacea canaliculata, with a critical appraisal of imposex evaluation in the Ampullariidae.

Ampullariidae are unique among gastropods in that females normally show a primordium of the copulatory apparatus (CApp). The aims of this study were (a) to quantitatively evaluate the development and growth of the female CApp with age; (b) to compare the effects of RXR and PPARγ agonists in adult females of known age and (c) to explore the effect of masculinizing RXR agonists on the expression of RXR in the CApp. It was found that the CApp grows and develops with age. A significant increase in penile sheath length (PsL) and also in a developmental index (DI) was observed in 7-8 months old females, as compared with 4-5 months old ones. A reported endogenous agonist of RXR, 9-cis retinoic acid (9cis-RA), as well as two organotin compounds, tributyltin (TBT) and triphenyltin (TPT) which have been also reported to bind to RXR, were injected and its masculinizing effects were measured. Also, the effect of a PPARγ agonist, rosiglitazone, was studied. All studied RXR agonists, but not the PPARγ agonist, were effective in increasing PsL, penile length (PL) and DI. Finally, the expression of the RXR in the CApp was studied (Western blot) in control, TBT, TPT, and 9cis-RA treated females. A significantly increased expression of RXR was only observed after 9cis-RA treatment. It is concluded that (a) development and growth of the CApp is significantly affected by female age; (b) reported RXR agonists, but not a PPARγ agonist, cause female masculinization of young females. An appraisal of previous studies of female masculinization in the Ampullariidae has also been made and it is emphasized that the masculinizing effect of aging should be considered, particularly when interpreting field data.