Dose‐dependent protective effects of dietary selenium on abalone Haliotis discus hannai Ino against the toxicity of waterborne copper

This study was designed to assess the protective effects of dietary selenium (Se) on abalone Haliotis discus hannai Ino against the toxicity of waterborne copper (Cu). A 60‐day feeding trial was conducted in a static water system for abalone (initial weight: 3.17 ± 0.01 g) exposed to 0.02 mg L^?1 of waterborne Cu. The animals were fed one of the three experimental diets with 0.10, 1.31 and 4.20 mg kg^?1 of Se from Na₂SeO₃·5H₂O respectively. Results showed that the abalone fed 1.31 mg kg^?1 of dietary Se had the lowest Cu concentration in shell, muscle, mantle, gill, hepatopancreas and serum. Meanwhile, the significant lowest contents of malondiadehyde and protein carbonyl in hepatopancreas were also found in the treatment with 1.31 mg kg^?1 of dietary Se (P < 0.05). In addition, this treatment had significant higher glutathione content and thioredoxin reductase activity in hepatopancreas (P < 0.05). However, the activity of Se‐dependent glutathione peroxidase (Se‐GPx) was significantly decreased in the treatment with 4.20 mg kg^?1 of dietary Se (P < 0.05). In this treatment, the protein carbonyl content in hepatopancreas was significantly higher than that in the group with 1.31 mg kg^?1 of dietary Se (P < 0.05). In conclusion, in terms of anti‐oxidation and Cu accumulation, the protective effects of dietary Se on abalone against waterborne Cu were dose‐dependent. The 1.31 mg kg^?1 of dietary Se had this effect, but not 4.20 mg kg^?1 of dietary Se. Moreover, the latter increased the oxidative stress in abalone exposed to the waterborne Cu.     

Influences of dietary antimicrobial peptide APSH‐07 on the growth performance,immune response and vibriosis resistance of abalone Haliotis discus hannai Ino

A 115‐day feeding trial and subsequently a 10‐day challenge test with Vibrio parahaemolyticus were conducted to investigate the effects of dietary antimicrobial peptide APSH‐07 on growth performance, immune response, antioxidative status and vibriosis resistance of abalone Haliotis discus hannai (initial body weight: 2.06 ± 0.01 g; initial shell length: 25.42 ± 0.18 mm). Four artificial diets were designed with 0 (artificial diet control), 7.5, 15.0 and 22.5 mg/kg of APSH‐07, respectively. The brown alga Laminaria japonica was used as the live food control. Results showed that the specific growth rates of abalone in the groups with 7.5 and 15.0 mg/kg dietary APSH‐07 were significantly higher than those in the controls (p < .05). The total haemocyte counts and respiratory burst activity in haemolymph of abalone in the group with 7.5 mg/kg of dietary APSH‐07 were significantly higher than those in the groups with 0 and 22.5 mg/kg of dietary APSH‐07 (p < .05). The gene expression levels of Mn‐superoxide dismutase and thioredoxin peroxidase 2, and the activities of glutathione peroxidase in the group with 7.5 mg/kg of dietary APSH‐07 were significantly higher than those in the other groups (p < .05). Cumulative mortality of abalone after the challenge test was significantly decreased in the group with 7.5 mg/kg of dietary APSH‐07 supplementation. Supplementation of 22.5 mg/kg dietary APSH‐07 significantly increased the cumulative mortality. In conclusion, 7.5 mg/kg of dietary APSH‐07 supplementation had the better growth performance, higher antioxidation, immune and disease resistance capacity of abalone. Excessive supplementation of dietary antimicrobial peptide APSH‐07 (22.5 mg/kg) had significantly negative effects. Further studies are needed to determine the optimal level of dietary APSH‐07 supplementation for abalone.     

Parentage assignment in hybrid abalones (Haliotis rufescens × Haliotis discus hannai) based on microsatellite DNA markers

Parentage analysis in aquaculture determines genealogical relationships between broodstock and progeny when the parents are unknown. Thus, parentage analysis is a useful tool to establish pedigree reports in molecular‐assisted selection programs. Here, we evaluated 10 heterologous microsatellite markers for parentage assignment in abalone hybrids produced from 43 abalone broodstocks of red abalone (Haliotis rufescens) and Japanese abalone (H. discus hannai). The allele frequencies, exclusion probabilities and broodstock contributions were calculated using CERVUS, PAPA and GERUD software. The polymorphic information content (PIC) values showed that most of the microsatellite loci were highly informative (>0.7) and more than 90% of parentage assignment was possible with a minimum of 5–6 microsatellite markers. Parentage assignment for hybrid and pure‐red progeny showed a better performance than pure‐Japanese progeny. This result could be due to the high level of allele loss in the parental genotypes. In addition, results indicated that only two sires contributed over 80% and 90% of red and hybrid progenies, respectively. This study gives a new molecular tool to support marker‐assisted selection in abalone hybrids produced in Chile.     

The potential for utilizing fouling macroalgae as feed for abalone Haliotis discus hannai

In order to assess the potential of utilizing fouling macroalgae as feed for abalone Haliotis discus hannai, the species, biomass and total stock of fouling macroalgae attached on the aquaculture facility‐lines were investigated and assessed at Sungo Bay, China, in August 2007. The nutritional value of four fouling macroalgae (Codium fragile, Sargassum muticum, Ulva pertusa and Laminaria japonica) and one cultured macroalgae (Gracilaria lemaneiformis) as the diet of abalone was assessed by analysis of the chemical composition of the macroalgae and by measuring bioavailability and digestibility to abalone H. discus hannai under laboratory conditions in August 2007. The results showed that the biomass of fouling macroalgae at the two sites was 243.61 ± 26.23 and 1078.15 ± 50.28 g m^?1 respectively. The contents of protein of G. lemaneiformis and C. fragile were 18.30% and 17.60%, respectively, which were significantly higher than the others. The five macroalgae were accepted by the abalone, and growth energies (P) and growth efficiency (%) were positive. However, significant differences were found among the different macroalgae. The growth efficiencies of abalone show a negative relationship with the levels of dietary lipid and a positive relationship with the protein contents. These results suggest that fouling macroalgae have a great potential to be used as diets of abalone.     

Effect of dietary organic selenium on survival,growth, antioxidation,immunity and gene expressions of selenoproteins in abalone Haliotis discus hannai

No studies have been focused on the inclusion of organic selenium in formulated feed of abalone when organic minerals are accepted by aquaculture feed industry. This experiment evaluated the effect of organic selenium on the growth, antioxidation, immunity and gene expressions of selenoproteins in abalone. Graded levels of organic selenium were included to the basal diet at 0.00, 0.15, 0.30 and 0.60 ppm providing 157.83, 322.51, 437.59 and 596.59 ug/kg of selenium in the diet. Accordingly, the diets were named as Se‐0.00, Se‐0.15, Se‐0.30 and Se‐0.60 respectively. Results had shown that survival rate, weight gain rate and shell length increase rate were lowest for Se‐0.60 group (p < 0.05). The concentration of selenium in the whole body generally increased with increasing level of dietary organic selenium (p < 0.05). The enzyme activities related to antioxidation increased initially with low levels of dietary selenium and decreased with higher levels, with the highest value in Se‐0.30 group (p < 0.05). The enzyme activities of immunity increased with the addition of dietary selenium. With the increase of dietary selenium, Myostain expression decreased initially and increased afterwards, with lowest value for Se‐0.30 group (p < 0.05). However, the expressions of other genes related to growth, antioxidation and selenoproteins firstly increased and then decreased, with the highest value recorded for Se‐0.15 group. In conclusion, the inclusion rate of 0.15 – 0.30 ppm Se in diet could promote growth, antioxidation, immunity and gene expressions of selenoproteins in abalone; however, excess selenium with 0.60 ppm inclusion lever in diet had toxic effects.     

Hybridization between Haliotis rufescens and Haliotis discus hannai: evaluation of fertilization,larval development,growth and thermal tolerance

Two introduced abalone species are currently produced in Chile, red abalone Haliotis rufescens and Japanese abalone Haliotis discus hannai. However, red abalone accounts for 99% of total production, while the Japanese abalone has not adapted well to Chilean coastal waters. This study reports the hatching, growth and thermal tolerance performance in interspecific hybrids produced between red (R) and Japanese (J) abalone. Our results show that egg age and sperm concentration were critical factors to produce hybrids. The cross R♀ × J♂ showed a fertilization rate of 55.3 ± 3.5% using 20‐min‐old eggs and sperm concentrations of 14 × 10⁶ cells mL^?1, while the reciprocal cross (J♀ × R♂) was not successful. Further, larval development stages were similar in RR, JJ and RJ hybrid abalones. Among the experimental trials, settlement rate varied from 12.3% to 18.6% and final survival from 20.1% to 31.7%, being the RJ hybrid rates intermediate between parental species. The final shell lengths were similar between RR and RJ hybrids, but significantly higher in JJ abalones. In addition, thermal tolerance was ascertained due its pivotal role for the abalone physiology. Thus, RJ hybrids showed the highest HSP70 gene expression and offers new possibilities to expand Chilean abalone production in warm waters zones.     

Mining expressed sequences for single nucleotide polymorphisms in Pacific abalone Haliotis discus hannai

Although single nucleotide polymorphisms (SNPs) are important resources for population genetics, pedigree analysis and genomic mapping, such loci have not been reported in Pacific abalone so far. In this study, a bioinformatics strategy was adopted to discover SNPs within the expressed sequences (ESTs) of Pacific abalone, Haliotis discus hannai , and furthermore, polymerase chain reaction direct sequencing (PCR-DS) and allele-specific PCR (AS-PCR) were used for SNPs detection and genotype scoring respectively. A total of 5893 ESTs were assembled and 302 putative SNPs were identified. The average density of SNPs in ESTs was 1%. Fifty-two sets of sequencing primers were designed from SNPs flanking ESTs to amplify the genomic DNA, and 13 could generate products of expected size. Polymerase chain reaction direct sequencing of the amplification products from pooled DNA samples revealed 40 polymorphic SNP loci. Using a modified tetra-primer AS-PCR, seven mitochondrial and six nuclear SNPs were typed and characterized among 37 wild abalones. In conclusion, it is feasible to discover SNPs from number limited ESTs and the AS-PCR as a simple, robust and reliable assay could be a primary method for small- and medium-scale SNPs detection in abalones as well as other non-model organisms.     

Nitrogen and phosphorus budget of a Haliotis discus hannai and Apostichopus japonicus polyculture system

The system nitrogen (N), phosphorous (P) budget, N/P utilization rate and the physiological response mechanism of the abalone Haliotis discus hannai (body weight: 12.87 ± 0.82 g) and the sea cucumber Apostichopus japonicus (body weight: 10.85 ± 1.16 g) to different co‐culture environment conditions were examined. Animals were kept in a multilayer, cubic recirculating aquaculture system at different polyculture densities (abalones at 400 ind/m²in monoculture [Group C] and abalones at 400 ind/m²with sea cucumbers at 10 ind/m² [AS1] or 20 ind/m² [AS2]). Each treatment was replicated four times, and the experimental cycle was 90 days. No significant difference in survival rate of abalones was detected when the stocking density of sea cucumbers increased from 10 to 20 ind/m², but the concentrations of total ammonia nitrogen and nitrite nitrogen in the water were significantly higher in AS2 than in AS1. Survival rate, specific growth rate (SGR) of body weight of sea cucumbers and SGR of body weight of abalones were significantly lower in AS2 than in AS1. No significant difference in protease (PES), lipase, amylase and cellulase activities of abalones was identified between Group C and AS1, but the PES and amylase activities of abalones and sea cucumbers in AS1 were significantly higher than those in AS2. In AS1, the N/P output from harvesting of abalones and sea cucumbers and the N/P utilization rates were significantly higher than those in AS2. Although the N/P output from faeces was significantly lower in AS2 than in Group C and AS1, the N/P output from the water layer was significantly higher than that in AS1. The expression levels of Cu/Zn superoxide dismutase (Cu/Zn‐SOD) and heat shock protein 70 (HSP70) of both abalones and sea cucumbers in AS2 were significantly higher than those in AS1. No significant difference in expression of catalase (CAT) and HSP90of abalones was identified among these groups, but the expression levels of CAT and HSP90of sea cucumbers in AS2 were significantly higher than those in AS1. These results indicate that stocking sea cucumbers at 10 ind/m² in the polyculture system will relieve the organic load on the system and improve the N/P utilization rate. It will also increase aquaculture production and improve the ecological and economic benefits of the system.     

Effects of dietary guaiacol on shell biomineralization of juvenile abalone Haliotis discus hannai, Ino

A study was conducted to investigate the effects of guaiacol on shell biomineralization of abalone Haliotis discus hannai Ino. Seven semi-purified diets were formulated with graded levels of guaiacol (0, 5, 10, 50, 100, 500 and 1000 mg kg⁻¹). Abalone (initial weight: 0.29±0.01 g; initial shell length: 8.55±0.27 mm) were fed with these diets in a re-circulated water system for 152 days. The results showed that the contents of calcite in the shell increased significantly with dietary guaiacol levels, but the aragonite content decreased significantly. Dietary guaiacol significantly decreased the contents of calcium and strontium, but significantly increased the magnesium content in the shell. The effects of dietary guaiacol on the contents of phosphorus and zinc in shell were not significant. It was obvious that the ratio of potential acidic/basic amino acids [(Asx+Glx)/(Lys+Arg+His)] in the shell was elevated by dietary guaiacol. In conclusion, dietary guaiacol has effects on both the mineralogy and the amino acid composition in the shell. Furthermore, changes in mineralogy are related to the ratio of (Asx+Glx)/(Lys+Arg+His) in the shell.     

Suitability of cage culture for Pacific abalone Haliotis discus hannai Ino production in China

Pacific abalone (Haliotis discus hannai Ino) aquaculture is a thriving industry in China. This study describes a novel submerged cage culture system for abalone rearing in Fujian, South China. The cage consisted of five vertical slots that were oriented perpendicular to the flow of water. The slots were separated by six vertically connected plastic plates for abalone attachment and shelter at the bottom of the cage. Experiment 1 was designed to determine the appropriate stocking density at the start of the abalone sea‐based production cycle. Eight‐month‐old hatchery reared and size‐graded juveniles were transferred to the sea‐based culture system. For different stocking densities, shell length of juveniles obtained in this novel culture system on 2, 3.5 and 5 months, respectively, was compared with shell lengths obtained in a traditionally multi‐tier basket culture system. In Experiment 2, daily growth rates (DGRs) in shell length and biomass in terms of wet weight of 2‐year‐old abalones reared in cage and tiered basket culture systems were compared over a 6‐month period. Results of Experiment 1 showed that growth of abalone in the cage culture system is density‐dependent; the mean final shell length of juveniles obtained was 6.7–15.9% higher than in tiered baskets system even at the same initial stocking density. In Experiment 2, DGRs in shell length of 53.83–78.38 μm day^?1 obtained in cage system were significantly higher than that in tiered baskets (P < 0.01). And in terms of wet weight biomass, it was 1.48–3.01 times higher in the cage system compared with the traditional system. Abalone survival was more than 87.5% in both culture systems in both experiments. Advantages of the newly established cage culture system included better growth performance of the animals reared and potential improvement of rearing conditions, such as improved water flow velocity and dissolved oxygen.     

碘伏对皱纹盘鲍的急性毒性试验

试验结果表明：碘伏对稚鲍2，42，48，72，96hLC50分别为3．91，0．64，0．38和0．36mg.L^-1有效碘。稚鲍对碘伏很敏感，且忍耐质量浓度范围较窄，不适合带水消毒。     

Production and sinking rates for bio‐deposits of abalone (Haliotis discus hannai Ino)

Particle size distribution, organic content and the sinking velocity of bio‐deposits of abalone Haliotis discus hannai Ino were evaluated. The abalone was divided into three size classes according to shell length: Group B: 68.8 ± 2.8 mm, group M: 56.1 ± 2.3 mm and group S: 42.4 ± 2.8 mm. The faecal pellets had an amorphous shape, with a cross‐sectional area ranging from 0.01 to 2.64 mm². The mean cross‐sectional area varied among the size classes: 0.59 (B), 0.21 (M) and 0.12 (S) mm². The bio‐deposit organic content was similar among the groups (B: 18.3 ± 1.1%; M: 19.9 ± 0.9%; S: 19.3 ± 1.0%). Sinking velocity ranged from 0.5 to 2.3 cm s^?1. Larger abalone tended to have larger and thus faster‐sinking faecal pellets. The median sinking velocity for groups B, M and S was 0.9, 0.5 and 0.7 cm s^?1 respectively. The mean daily production rates for groups S, M and B were 2.8 ± 1.2, 3.4 ± 1.3 and 4.3 ± 1.8 g ind.^?1 respectively. Bio‐deposit production rate estimated to be between 134.4 and 206.4 mg m^?2 day^?1 in the bay. Our results suggest that the pellet size was within the appropriate size range for filter‐feeding bivalves.     

用流式细胞仪检测活体鲍倍性

分别取皱纹盘鲍（Ｈａｌｉｏｔｉｓ ｄｉｓｃｕｓ ｈａｎｎａｉ）足肌和血液制样,经ＤＡＰＩ染色后,用ＰａｒｔｅｃＰＡＳ－Ⅲ型流式细胞仪测定其倍性,试验证明三倍体鲍的ＤＮＡ相对含量是二保体的１．５倍,切足法和抽取血液法两种制样方法均得到满意结果,用流式细胞仪法检测鲍倍性适应鲍多倍体产业化的需要。     

叶酸对皱纹盘鲍脂肪酸组成的影响

以皱纹盘鲍幼鲍Haliotis discus hannaiIno(平均壳长:13.90±0.01mm;平均体重:482.2±2.2mg)为研究对象,在室内循环水养殖系统中进行112d的摄食生长实验,以海带为对照,探讨饲料中不同叶酸含量(0.06、1.44、2.86、5.88、14.27、35.71mg/kg饲料)对皱纹盘鲍脂肪酸组成的影响。实验结果表明,(1)叶酸缺乏可显著降低皱纹盘鲍C20∶4n-6和C20∶5n-3的含量(P<0.05),却显著提高C18∶3n-3和C22∶6n-3的含量(P<0.05);(2)海带与实验基础饲料中的脂肪酸组成存在显著差异,海带中的C14∶1、C16∶0、C18∶1n-9、C18∶3n-3、C20∶4n-6、C20∶5n-3含量显著高于饲料组(P<0.05),而C18∶0、C18∶1n-7、C18∶2n-6、C20∶0、C22∶6n-3均显著低于饲料组(P<0.05);(3)海带组鲍鱼的C20∶4n-6、C20∶5n-3显著高于饲料组鲍鱼(P<0.05),而C18∶1n-9、C18∶1n-7、C18∶2n-6、C22∶6n-3则显著低于饲料组鲍鱼(P<0.05)。(4)C16∶0、C16∶1、C18∶0在鲍鱼体内的含量比较稳定,不受饲料中叶酸含量和脂肪酸组成的显著影响(P>0.05)。这些结果说明,饲料中叶酸缺乏导致皱纹盘鲍脂肪酸组成发生改变,很可能抑制C18∶2n-6和C18∶3n-3的碳链延长和去饱和作用,从而减少C20∶4n-6和C20∶5n-3的合成。此外,饲料中的脂肪酸组成也是影响皱纹盘鲍体组织脂肪酸组成的一个重要因素。     

皱纹盘鲍食道的结构与功能

以组织学，组织化学，扫描电镜和透射电镜观察及酶活性测定等方法研究了皱纹盘鲍的食道，食道可分为前，中后，三段，中段又可分为食物通道和食道侧囊，食道粘膜上皮由纤毛柱状细胞，粘液细胞，闰状腺细胞，微绒毛细胞和分泌细胞组成，纤毛柱状细胞参与运输食物和分泌物，并呈现吸收细胞的结构特征；粘液细胞分泌中性和酸性粘多糖；颗粒状腺细胞内充盈分泌颗粒；微绒毛细胞呈现吸收细胞的特征；分泌细胞具有强的蛋白酶等酶活性，能以楔浆分泌形式分泌消化酶，该细胞还具有吸收和细胞内消化作用。食道中段还呈现3种植物多糖酶活性。