Pacific Abalone Research Articles

Effects of different feeding patterns on growth, enzyme activity, and intestinal microbiome of the juvenile Pacific abalone Haliotis discus hannai

In northern China, the predominant food sources for abalone farming include fresh kelp, salted kelp, and formulated diets, and abalone can also endure prolonged starvation when food is scarce. While there is a dearth of comprehensive investigation into the effects of these feeding patterns on the temporal dynamics of enzyme activity, and intestinal microbiome of abalone. In this study, a total of 3600 juvenile Pacific abalones were allocated to 12 tanks, randomly divided into four groups based on the feeding patterns: fresh kelp (labeled as FK), salted kelp (SK), formulated feed (FF), and starved group (S). After a 60-day feeding trial, the results showed that abalone in the FK, SK, and FF groups exhibited significant positive effects on both weight and shell growth compared to the S group (P<0.05). Notably, the FF group displayed the highest values of the growth performance. The activity of digestive enzymes and antioxidant enzymes in the FF group had an up-regulated trend over the course of feeding trial, whereas amylase and lipase activity in abalone declined along with starvation. Moreover, the microbial Shannon index showed an overall decreasing trend over time in the FF, FK, and SK groups except for the S group. The ANOSIM analysis revealed that both feeding pattern and cultivating time significantly influenced the structure of the microbial communities. Across the feeding trial, Proteobacteria, Fusobacteriota, and Firmicutes were identified as the dominant phyla in the intestinal microbiome of abalone under different feeding patterns. The core genera were Psychrilyobacter, Mycoplasma, and Vibrio. Additionally, the neutral model analysis revealed that stochastic processes both play a key role in shaping the assembly of microbial communities in different feeding patterns. This study could offer valuable insights into the impact of different feeding patterns on growth and physiology of abalone, thereby contributing to the enhancement of management strategies in the northern Chinese abalone aquaculture industry.

Structure and function of vasa gene in gonadal gametogenesis of Pacific abalone

Pacific abalone (Haliotis discus hannai) is a marine gastropod mollusc with significant economic importance in both global fisheries and aquaculture. However, studies exploring the gonadal development and regulatory mechanisms of Haliotis discus hannai are limited. This study aimed to explore whether the vasa gene acted as a molecular marker for germ cells. Initially, the vasa gene was successfully cloned using the cDNA-end rapid amplification technique. The cloned gene had a 2478-bp-long open reading frame and encoded 825 amino acids. Then, a recombinant expression vector was constructed based on the Vasa protein, and an 87-kDa recombinant protein was prepared. Subsequently, a polyclonal antibody was prepared using the purified recombinant protein. The enzyme-linked immunosorbent assay (ELISA) confirmed the titer of the antibody to be ≥512 K. The immunohistochemical analysis revealed that Vasa was widely expressed in oogonia, Stage I oocytes, spermatogonia, and primary spermatocytes. The specific expression of Vasa in the hermaphroditic gonads of abalone was assessed using western blotting to investigate the effects of different photoperiods (12 L:12D, 24 L:0D, 18 L:6D, and 6 L:18D) on the gonadal development of abalone (P < 0.05), with higher expression levels observed in the ovarian proliferative and spermary maturing stages compared with other developmental stages (P < 0.05). Additionally, Vasa exhibited the highest expression in the spermary and ovary under a photoperiod of 18 L:6D (P < 0.05). These data demonstrated the key role of Vasa in developing germ cells in abalone. They shed light upon the molecular mechanism through which the photoperiod influenced Vasa expression and regulated gonadal development in abalone. The findings might provide theoretical references for analyzing the differentiation pattern of abalone germ cells and the genetic improvement and conservation of germplasm resources.

Carryover effects of embryonic hypoxia exposure on adult fitness of the Pacific abalone

The widespread and severe drop in dissolved oxygen concentration in the open ocean and coastal waters has attracted much attention, but assessments of the impacts of environmental hypoxia on aquatic organisms have focused primarily on responses to current exposure. Past stress exposure might also affect the performance of aquatic organisms through carryover effects, and whether these effects scale from positive to negative based on exposure degree is unknown. We investigated the carryover effects of varying embryonic hypoxia levels (mediate hypoxia: 3.0−3.1 mg O2/L; severe hypoxia: 2.0−2.1 mg O2/L) on the fitness traits of adult Pacific abalone (Haliotis discus hannai), including growth, hypoxia tolerance, oxygen consumption, ammonia excretion rate, and biochemical responses to acute hypoxia. Moderate embryonic hypoxia exposure significantly improved the hypoxia tolerance of adult Pacific abalone without sacrificing growth and survival. Adult abalone exposed to embryonic hypoxia exhibited physiological plasticity, including decreased oxygen consumption rates under environmental stress, increased basal methylation levels, and a more active response to acute hypoxia, which might support their higher hypoxia tolerance. Thus, moderate oxygen declines in early life have persistent effects on the fitness of abalone even two years later, further affecting population dynamics. The results suggested that incorporating the carryover effects of embryonic hypoxia exposure into genetic breeding programs would be an important step toward rapidly improving the hypoxia tolerance of aquatic animals. The study also inspires the protection of endangered wild animals and other vulnerable species under global climate change.

Shellfish CO2 excretion is modulated by seawater carbonate chemistry but largely independent of pCO2.

Four species of shellfish, blue mussel (Mytilus galloprovincialis), Pacific abalone (Haliotis discus hannai), zhikong scallops (Chlamys farreri), and Pacific oyster (Crassostrea gigas), were exposed to decoupled carbonate system variables to investigate the impacts of different seawater carbonate parameters on the CO2 excretion process of mariculture shellfish. Six experimental groups with two levels of seawater pH (pH 8.1 and pH 7.7) and three levels of total alkalinity (TA = 1000, 2300, and 3600μmol/kg, respectively) were established, while pH 8.1 and TA = 2300μmol/kg was taken as control. Results showed that the CO2 excretion rates of these tested shellfish were significantly affected by the change in carbonate chemistry (P < 0.05). At the same TA level, animals incubated in the acidified group (pH 7.7) had a lower CO2 excretion rate than those in the control group (pH 8.1). In comparison, at the same pH level, the CO2 excretion rate increased when seawater TA level was elevated. No significant correlation between the CO2 excretion rate and seawater pCO2 levels (P > 0.05) was found; however, a significant correlation (P < 0.05) between CO2 excretion rate and TA-DIC (the difference between total alkalinity and dissolved inorganic carbon) was observed. Blue mussel has a significantly higher CO2 excretion rate than the other three species in the CO2 excretions per unit mass of soft parts, with no significant difference observed among these three species. However, in terms of CO2 excretion rate per unit mass of gills, abalone has the highest CO2 excretion rate, while significant differences were found between each species. Our studies indicate that the CO2 buffering capacity impacts the CO2 excretion rate of four shellfish species largely independent of pCO2. Since CO2 excretion is related to acid-base balancing, the results imply that the effects of other carbonate parameters, particularly the CO2 buffering capacity, should be studied to fully understand the mechanism of how acidification affects shellfish. Besides, the species difference in gill to soft parts proportion may contribute to the species difference in responding to ocean acidification.

Genetic diversity and population structure of Pacific abalone (Haliotis discus hannai) using SNP genotyping data

The Pacific abalone Haliotis discus hannai is one of the most valuable mollusks in China. It also supports an important fishery in the northern coast of China, which has shown a declining tendency. The conservation of this species is attracting attentions under the potential threats of alien stocks and hybrid varieties disseminated in aquaculture. However, population genetics information on whole-genome level is lacking. The present study analyzed the genetic diversity, population structure, and inbreeding coefficient of nine populations collected from China and Japan using single nucleotide polymorphism (SNP) genotyping data from 214 samples. The results showed higher observed heterozygosity (Ho) than expected heterozygosity (He), and negative inbreeding coefficient (Fis) values indicating an excess of heterozygotes, and highlighting medium to high genetic diversity within the population. The genetic structures of the nine populations were categorized into three distinct subgroups. Pairwise Fst values based on all SNP loci and those for neutral markers were consistently low, indicating an overall genetic homogeneity between the populations. The distribution of runs of homozygosity (ROH) lengths suggests the potential occurrence of kinship relationships in various historical periods, including both distant and recent periods. The TreeMix and shared haplotypes suggested strong gene flow from Japanese populations of Pacific abalone to Chinese populations. These findings establish the stock status of the Pacific abalone in China, providing a basis for conservation, fishery management and utilization of the stocks in aquaculture.

First insight on the genetic base of the heritable variation of Pacific abalone (Haliotis discus hannai) resistance to Polydora hoplura infestation

Polydora hoplura is an invasive species and a harmful pest in commercial shellfish aquaculture, particularly for farmed abalone species. Different lines of treatment for Polydora affecting abalone have been explored, but with limited success. Using selective breeding to increase abalone resistance to and reduce shell damage from Polydora infestation could be a promising alternative. However, in abalone there is currently no information on the genetic variability for resistance to Polydora infestation. In the present study, heritability (h2) of traits associated with Pacific abalone resistance to P. hoplura were estimated after challenging abalone juveniles with trojan abalones heavily infested, using a nested experimental design, with 1 male crossed with 2–4 females. The heritability values were low but significant for some of those traits: survival h2=0.09; Infestation Intensity at Q50h2 = 0.16 ± 0.11; and the proportion of the shell area occupied by blisters caused by the mud-worms h2 = 0.29 ± 0.14. These results suggest a significant potential for improving resistance or tolerance to P. hoplura infestation through selective breeding in Pacific abalone.

Morphologic, cytometric, quantitative transcriptomic and functional characterisation provide insights into the haemocyte immune responses of Pacific abalone (Haliotis discus hannai).

In recent years, the abalone aquaculture industry has been threatened by the bacterial pathogens. The immune responses mechanisms underlying the phagocytosis of haemocytes remain unclear in Haliotis discus hannai. It is necessary to investigate the immune mechanism in response to these bacterial pathogens challenges. In this study, the phagocytic activities of haemocytes in H. discus hannai were examined by flow cytometry combined with electron microscopy and transcriptomic analyses. The results of Vibrio parahaemolyticus, Vibrio alginolyticus and Staphylococcus aureu challenge using electron microscopy showed a process during phagosome formation in haemocytes. The phagocytic rate (PP) of S. aureus was higher than the other five foreign particles, which was about 63%. The PP of Vibrio harveyi was about 43%, the PP peak of V. alginolyticus in haemocyte was 63.7% at 1.5h. After V. parahaemolyticus and V. alginolyticus challenge, acid phosphatase, alkaline phosphatase, total superoxide dismutase, lysozyme, total antioxidant capacity, catalase, nitric oxide synthase and glutathione peroxidase activities in haemocytes were measured at different times, differentially expressed genes (DEGs) were identified by quantitative transcriptomic analysis. The identified DEGs after V. parahaemolyticus challenge included haemagglutinin/amebocyte aggregation factor-like, supervillin-like isoform X4, calmodulin-like and kyphoscoliosis peptidase-like; the identified DEGs after V. alginolyticus challenge included interleukin-6 receptor subunit beta-like, protein turtle homolog B-like, rho GTPase-activating protein 6-like isoform X2, leukocyte surface antigen CD53-like, calponin-1-like, calmodulin-like, troponin C, troponin I-like isoform X4, troponin T-like isoform X18, tumor necrosis factor ligand superfamily member 10-like, rho-related protein racA-like and haemagglutinin/amebocyte aggregation factor-like. Some immune-related KEGG pathways were significantly up-regulated or down-regulated after challenge, including thyroid hormone synthesis, Th17 cell differentiation signalling pathway, focal adhesion, melanogenesis, leukocyte transendothelial migration, inflammatory mediator regulation of TRP channels, ras signalling pathway, rap1 signalling pathway. This study is the first step towards understanding the H. discus hannai immune system by adapting several tools to gastropods and providing a first detailed morpho-functional study of their haemocytes.

Palliative effect of abalone insulin-related peptide 2 on emersion stress-induced hyperglycemia in the Pacific abalone (Haliotis discus hannai)

The Pacific abalone (Haliotis discus hannai), an economically important marine mollusk in East Asia, is often exposed to extreme environments, such as high water temperatures during summer and air exposure during handling or transportation. These stressful conditions lead to hyperglycemia and the generation of reactive oxygen species (ROS), resulting in oxidative stress and subsequent cell damage in the organism. To understand the role of insulin-related peptides in the regulation of elevated hemolymph glucose, hyperglycemia was confirmed in abalone under high water temperature and air (emersion) exposure (p < 0.05). To prove the existence of a glucose regulatory system in abalone, changes in hemolymph glucose were measured after bovine insulin injection, and hemolymph glucose was lowered by bovine insulin injection in hyperglycemia caused by both stressors (p < 0.05), supporting the existence of a glucose regulatory system functionally similar to that of vertebrates. Among the three abalone insulin-related peptides (AIPs), which were previously reported to be upregulated along with hemolymph glucose levels upon feeding, the relative mRNA expression of AIP2 in the cerebral ganglion showed a close correlation with high water temperature and emersion stress, which peaked 12 h after treatment (p < 0.05). Based on these results, we focused on the function of AIP2 in glucose homeostasis and characterized the localization of AIP2 mRNA-expressing cells in the cerebral ganglion using in situ hybridization. Positive signals from RNA probes were detected in neural cortex cells in the cerebral ganglion. Furthermore, suppressing AIP2 mRNA expression using RNA interference with double-stranded RNA (dsRNA) revealed that dsRNA-AIP2 injected abalone maintained high hemolymph glucose levels after 24 h of recovery following 1 h of emersion stress (p < 0.01). In contrast, abalones that received dsRNA-GFP and molluscan saline showed relatively rapid clearance of hemolymph glucose. This study demonstrates that AIP2 gene expression affects the recovery rate from emersion stress-induced hyperglycemia by regulating hemolymph glucose.

Structural and functional characterization of Hdh-HSBP1 and its involvement in heat stress and early development in Pacific abalone, Haliotis discus hannai

Heat shock factor binding protein 1 (HSBP1) is known to regulate the activity of heat shock factor 1 (HSF1) and the early development of organisms. To understand the involvement of HSBP1 in the heat shock response and embryonic and larval development of Pacific abalone (Haliotis discus hannai), the Hdh-HSBP1 gene was sequenced from the digestive gland (DG) tissue. The full-length sequence of Hdh-HSBP1 encompassed 738 nucleotides, encoding an 8.42 kDa protein consisting of 75 deduced amino acids. The protein contains an HSBP1 domain and a coiled-coil domain, which are conserved features in the HSBP1 protein family. Protein-protein molecular docking revealed that the coiled-coil region of Hdh-HSBP1 binds to the coiled-coil region of Hdh-HSF1. Tissue expression analysis demonstrated that the highest Hdh-HSBP1 expression occurred in the DG, whereas seasonal expression analysis revealed that this gene was most highly expressed in summer. In heat-stressed abalone, the highest expression of Hdh-HSBP1 occurred at 30 °C. Moreover, time-series analysis revealed that the expression of this gene began to increase significantly at 6 h post-heat stress, with higher expression observed at 12 h and 24 h post-heat stress. Furthermore, Hdh-HSBP1 mRNA expression showed a link to ROS production. Additionally, the expression of Hdh-HSBP1 showed significantly higher expression in the early stages of embryonic development in Pacific abalone. These results suggest that Hdh-HSBP1 plays a crucial role in the stress physiology of Pacific abalone by interacting with Hdh-HSF1, as well as its embryonic development.

Effectiveness of the garlic (Allium sativum) juice on removing the Pacific abalone postlarvae (Haliotis discus hannai) and subsequent growth evaluation

The Pacific abalone (Haliotis discus hannai) is widely cultivated in East Asian countries. Aquaculture of this species has become a thriving industry in China, with adequate seed production being a key contributing factor. However, maintaining sufficient diatom abundance during the nursery phase remains challenging. In this study, we assessed a transfer nursery culture method that involves prolonged cultivation with a sufficient diatom supply. This method involved treating postlarvae on diatom-depleted plates with a garlic juice (GJ) solution to facilitate detachment, followed by transfer to newly diatom-precoated plates for further growth in the nursery. We investigated the effects of garlic juice concentration (GJC) and animal size on detachment and mortality rates using a two-factor design. Our negative binomial regression analysis uncovered significant impacts of both factors and their interactions on the target traits. We noticed that the optimal GJC for smaller groups of abalone postlarvae (2.2 ± 1.0 mm) was 0.7 %, while for larger groups (3.8 ± 2.2 mm), it increased to 1.0 %. Notably, suitable GJC concentrations during post-treatment recovery resulted in no apparent mortality in Pacific abalone postlarvae. Our study demonstrated the effectiveness of the transfer nursery method, showing an improvement of over ∼1.1 mm in shell length and a 43.1 % increase in survival during a 30-day experiment. Our study presents a significant advancement, as it likely introduces the first reported method of large-scale postlarval removal of H. discus hannai for prolonged nursery periods with sufficient diatom provision. The integration of GJ treatment and the transfer nursery method exhibited promising results, highlighting the benefits of its application for enhancing husbandry procedures, nursery management, and the overall industrial productivity of Pacific abalone.

Application of ultrasonography in abalone gonadal evaluation

The precise assessment of the gonad status and gamete development level is an essential step in abalone breeding. However, traditional methods are damaging to the animals, causing the waste of a quality parent. Ultrasound imaging as a noninvasive technique is a candidate substitute. This study aimed to verify the application of ultrasonography to the assessment of abalone gonad condition. Pacific abalones (n = 20) were randomly selected for ultrasonography, another six abalones in different gonad maturation stages (mature, between mature and immature, and immature) were also subjected to the same detection and histological analysis. Abalones (n = 10) were induced to spawn, and the changes in the gonad index and the types of germ cells were determined before and after spawning. Thirty-six female abalones were selected and divided into a high-gonad index group (HGI) and a low-gonad index group (LGI) according to the gonad index determined by ultrasonography, and their reproductive performance was compared during the entire breeding process. There was no significant difference in gonad index or gonad relative average thickness (GRAT) obtained from ultrasonography and histological methods. The gonad index and GRAT measured by ultrasound images significantly decreased after spawning. After spawning, a minimal quantity of mature gametes remained in gonad tissue as reflected in histology images. In a reproductive trial, the HGI group showed comprehensive and overwhelming reproductive advantages compared to the LGI group in total fecundity, relative fecundity, fertilization rate, hatching rate, abnormality rate, and attachment rate. Ultrasonography was demonstrated to be useful as a noninvasive method to evaluate abalone gonad condition. The ultimate goal is to achieve tracing and monitoring the abalone reproductive cycle through the ultrasound technique.

Behavioral and physiological responses of nocturnal marine shellfish to predation pressure

Predator-prey interactions are critical in ecological communities. As the wild resources of Pacific abalone are on the verge of exhaustion, efforts to reveal the effects of predation by hostile organisms on the efficiency of its bottom sowing and proliferation are crucial for the success of resource restoration. In this study, abalone and its predator, starfish Asterias amurensis, were placed in a customized monitoring unit. Through continuous infrared video monitoring, predation pressure was found to cause a significant decrease in the distance moved, duration of movement, cumulative feeding duration and orexin levels in hemolymph (P < 0.01). It also caused a significant increase in the average velocity of movement, group cohesion, coordination and cholecystokinin levels in hemolymph (P < 0.01). Surprisingly, when a shelter was set up in the environment, the cumulative feeding duration of abalone was significantly extended in the presence of the predator (P < 0.001), whereas when the predator was removed, the distance moved, duration of movement, and cumulative feeding duration of abalone were found to significantly increase during continuous 24 h monitoring (P < 0.01). These results first identified the anti-predation strategy by nocturnal marine organisms when exposed to predation risk, and supported the pivotal role of shelter in helping abalone make a trade-off between predation pressure and survival needs, which not only provides a new insight to optimizing the strategy for prevention and control of marine ecological disasters and strengthening the protection of marine endangered species, but also results in a productive attempt to restore the marine benthic ecosystem.

Ascorbic acid biosynthesis in Pacific abalone Haliotis discus hannai Ino and L-gulonolactone oxidase gene loss as an independent event

Up to now, it has been believed that invertebrates are unable to synthesize ascorbic acid (AA) in vivo. However, in the present study, the full-length CDs (Coding sequence) of L-gulonolactone oxidase (GLO) from Pacific abalone (Haliotis discus hannai Ino) were obtained through molecular cloning. The Pacific abalone GLO contained a FAD-binding domain in the N-termination, and ALO domain and conserved HWAK motif in the C-termination. The GLO gene possesses 12 exons and 11 introns. The Pacific abalone GLO was expressed in various tissues, including the kidney, digestive gland, gill, intestine, muscle and mantle. The GLO activity assay revealed that GLO activity was only detected in the kidney of Pacific abalone. After a 100-day feeding trial, dietary AA levels did not significantly affect the survival, weight gain, daily increment in shell length, and feed conversion ratio of Pacific abalone. The expression of GLO in the kidney was downregulated by dietary AA. These results implied that the ability to synthesize AA in abalone had not been lost. From the evolutionary perspective, the loss of GLO occurred independently as an independent event by matching with the genomes of various species. The positive selection analysis revealed that the GLO gene underwent purifying selective pressure during its evolution. In conclusion, the present study provided direct evidence to prove that the GLO activity and the ability to synthesize AA exist in abalone. The AA synthesis ability in vertebrates might have originated from invertebrates dating back 930.31 million years.

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.

Effects of reproductive condition on haemocyte parameters in the Pacific abalone Haliotis discus hannai during the spawning period

ABSTRACT The assessment of morphology and immunology of specimens at different maturity stages is essential for the determination of spawning strategy and reducing the risk of mortality. In this paper, female and male Pacific abalones (Haliotis discus hannai) at different statuses (pre-spawning, spawning and post-spawning) were collected to estimate the effects of spawning on the immune response. The haemocyte parameters, including total haemocyte counts (THC), haemocyte mortality, and reactive oxygen species production (ROS) were analysed using flow cytometry. THC decreased after spawning, which was lowest 3 days post-spawning. Haemocyte mortality was higher during spawning, and was highest 3 days post-spawning. Abalone also reduced their metabolic activity after spawning, reflected by an increase in ROS in both females and males. Moreover, our results show that female abalone reach the spawning period with a higher degree of activities of haemocytes and immune responses than male. The results confirmed that variability in abalone haemocytes during the spawning period might related to individual metabolic conditions.

Changes in water temperature: The effect of triploid performance in Pacific abalone (Haliotis discus hannai)

The triploid Pacific abalone (Haliotis discus hannai) has significant economic value and promising application prospects. However, the optimal cultural conditions that maximize the advantages of triploidy in the abalone have yet to be determined. This study examined the properties of triploid Pacific abalone after 3 months of culture under three constant temperatures (18 °C, 22 °C, and 26 °C) to fill up this information gap. The results showed that the survival rate of triploids in the 26 °C group was significantly higher than that of diploids (P < 0.05), while the survival rate of triploids in the other two groups was similar to that of diploids (P > 0.05). Except for the shell width of the 22 °C group and the shell length of the 26 °C group, triploid Pacific abalone exhibited significantly superior growth traits compared to diploids at all temperatures (P < 0.05). The most notable improvement was observed in the foot muscle weight trait, where there were triploid advantage rates of 81.44% (18 °C), 76.21% (22 °C), and 54.96% (26 °C). The specific growth rate and the ratio of shell length and body weight indicated that triploids grew the fastest in the 22 °C group. Furthermore, the thermal resistance of diploids and triploids was evaluated through the Arrhenius break temperature (ABT) of cardiac performance. The results demonstrated that the ABT of triploids was significantly higher than that of diploids (P < 0.05, ABT2N = 30.81 ± 0.43 °C, ABT3N = 31.38 ± 0.72 °C). Both diploid and triploid foot adhesion forces were measured, and the results showed that triploid foot adhesion force was stronger than that of diploids (P < 0.05). Additionally, there was a significant positive correlation between foot adhesion force and meat yield traits (foot muscle weight, fullness index (FI) and foot muscle-soft tissue index (FMSI)) (P < 0.05). In summary, this study confirmed the triploid advantage in meat yield of Pacific abalone and identified a temperature range that maximizes their growth rate. Moreover, triploids exhibited improved thermal resistance and adhesion capacity compared to diploids, with an increase in meat yield. The results of this study could serve as a reference for identifying a culture model suitable for triploid Pacific abalone and thus could promote the development of the triploid aquaculture industry.

Molecular and structural analysis of Hdh-MIRP3 and its impact on reproductive regulation in female Pacific abalone, Haliotis discus hannai

Molluscan insulin-related peptides (MIRP) play a crucial role in various biological processes, including reproduction and larval development in mollusk species. To investigate the involvement of MIRP in the ovarian development of Pacific abalone (Haliotis discus hannai), the Hdh-MIRP3 was cloned from cerebral ganglion (CG). Hdh-MIRP3 cDNA was 993 bp long, encoded a 13.22 kDa peptide, comprising 118 amino acids. Fluorescence in situ hybridization confirmed the localization of Hdh-MIRP3 in the CG and ovary. Molecular docking revealed that Hdh-MIRP3 binds to the N-terminal region of Hdh-IRP-R. Tissue expression analysis showed the highest Hdh-MIRP3 expression in the CG, followed by ovarian tissue. Hdh-MIRP3 expression was significantly upregulated in the CG and ovary during the ripe stage of seasonal ovarian development and in effective accumulative temperature conditioned abalone. Furthermore, siRNA silencing of Hdh-MIRP3 significantly downregulated the expression of four reproduction-related genes, including Hdh-GnRH, Hdh-GnRH-R, Hdh-IRP-R, and Hdh-VTG in both the CG and ovary, and Hdh-MIRP3 as well. These results indicate that Hdh-MIRP3 acts as a regulator of ovarian development in Pacific abalone. Additionally, expression analysis indicated that Hdh-MIRP3 plays a role in embryonic and larval development. Overall, the present findings elucidate the role of Hdh-MIRP3 in reproductive development in female Pacific abalone.

The taste characteristics and metabolite variations of two Pacific abalone strains with different glycogen contents

The flavor of shellfish has become one of the key selection goals in aquatic breeding since it influences consumer preferences. Glycogen has been proven to be a primary taste-enhancing component in most shellfish. But fewer studies were reported on Pacific abalone, and the contribution of glycogen to taste is unclear. In this study, the human sensory test, electronic tongue, ultra-high-performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS), and multivariate statistical methods were used to investigate differences in flavor between two groups (high and low groups, respectively) of Pacific abalone with different glycogen contents. The results of the two flavor evaluations consistently showed that the scores of umami and sweetness were significantly higher in the high glycogen content group. Pacific abalone with a higher glycogen content has a better flavor. A total of 295 substantially differential metabolites were identified between the two groups. Lysine, glutamic acid, glycine, serine, threonine, nucleotides, phosphatidylcholine (PC), and phosphatidylethanolamine (PE) were considered to be the main taste-enhancing components in the high group. This study proves the significance of glycogen content in Pacific abalone on flavor characteristics and provides new insights into the underlying metabolic causes of flavor differences. Additionally, the findings also provide guidance for selection breeding of quality-related traits in Pacific abalone.

Genome-wide identification and expression profiles of sex-related gene families in the Pacific abalone Haliotis discus hannai

In recent years, members of the Dmrt family, TGF-β superfamily and Sox family have been recognized as crucial genes for sex determination/differentiation across diverse animal species. Nevertheless, knowledge regarding the abundance and potential functions of these genes in abalone remains limited. In this study, a total of 5, 10, and 7 members of the Dmrt family, the TGF-β superfamily and the Sox family, respectively, were identified in the Pacific abalone Haliotis discus hannai. Sequence characteristics, phylogenetic relationships and spatiotemporal expression profiles of these genes were investigated. Notably, HdDmrt-04 (Dmrt1/1L-like) emerged as a potential mollusc-specific gene with a preponderance for expression in the testis. Interestingly, none of the TGF-β superfamily members exhibited specific or elevated expression in the gonads, highlighting the need for further investigation into their role in abalone sex differentiation. The Sox proteins in H. discus hannai were categorized into 7 subfamilies: B1, B2, C, D, E, F, and H. Among them, HdSox-07 (SoxH-like) was observed to play a crucial role in testis development, while HdSox-03 (SoxB1-like) and HdSox-04 (SoxC-like) probably cooperate in abalone ovary development. Taken together, the results of the present study suggested that HdDmrt-04 and HdSox-07 can be used as male-specific markers for gonad differentiation in H. discus hannai and imply conservation of their functions across invertebrates and vertebrates. Our findings provide new insights into the evolution and genetic structure of the Dmrt family, the TGF-β superfamily and the Sox family in abalone and pave the way for a deeper understanding of sex differentiation in gastropods.

Easy-to-Use CRISPR-Cas9 Genome Editing in the Cultured Pacific Abalone (Haliotis discus hannai).

The Pacific abalone is an important aquaculture shellfish and serves as an important model in basic biology study. However, the study of abalone is limited by lack of highly efficient and easy-to-use gene-editing tools. In this paper, we demonstrate efficient gene knockout in Pacific abalone using CRISPR-Cas9. We developed a highly effective microinjection method by nesting fertilized eggs in a low-concentration agarose gel. We identified the cilia developmental gene β-tubulin and light-sensitive transmembrane protein r-opsin as target genes and designed highly specific sgRNAs for modifying their genomic sequences. Sanger sequencing of the genomic regions of β-tubulin and r-opsin genes from injected larvae identified various genomic long-fragment deletions. In situ hybridization showed gene expression patterns of β-tubulin and r-opsin were significantly altered in the mosaic mutants. Knocking out β-tubulin in abalone embryos efficiently affected cilia development. Scanning electron microscopy and swimming behavior assay showed defecting cilia and decreased motility. Moreover, knocking out of r-opsin in abalone embryos effectively affected the expression and development of eyespots. Overall, this work developed an easy-to-use mosaic gene knockout protocol for abalone, which will allow researchers to utilize CRISPR-Cas9 approaches to study unexploited abalone biology and will lead to novel breeding methods for this aquaculture species.