Juvenile Sea Cucumber Apostichopus Japonicus Research Articles

Bacillus licheniformis reduces mortality, improves symptoms, and controls residual pathogenicity in juvenile sea cucumbers Apostichopus japonicus infected with skin ulceration syndrome

Mass mortality caused by skin ulceration syndrome (SUS) is the bottle-neck for the sustainable aquaculture of the sea cucumber Apostichopus japonicus. In the present study, probiotic Bacillus licheniformis (0.25 × 109 CFU/g) was used as the treatment for A. japonicus infected with the SUS that caused by Vibrio harveyi. We found that B. licheniformis significantly reduced the number of infected sea cucumbers 5 days and 7 days after the treatment (group B), compared to those without B. licheniformis treatment (group C) (P < 0.001; P < 0.001). Further, the sea cucumbers fed B. licheniformis had significantly lower mortality at the end of the experiment (<10%) than that of those without the B. licheniformis treatment (>60%) (P < 0.001). These results suggest that the treatment of B. licheniformis is an effective method to reduce the mass mortality resulted from SUS in sea cucumber aquaculture. Further, 3–5 days of treatment significantly improved the adverse symptoms of SUS on the physiology and behavior of sea cucumbers, including the righting behavior, adhesion behavior, food consumption, fecal output and mobility. This indicates B. licheniformis treatment has the advantage in the recovery of sea cucumbers after SUS. Moreover, there was no significant difference observed in the physiology and behavior of sea cucumbers between the SUS infected sea cucumbers after the 7-day treatment of B. licheniformis and the healthy individuals. SUS infected sea cucumbers effectively returned to a stage of normalcy. Further, we found a significantly lower infected rate in sea cucumbers exposed to the culture water of group B (∼5%) than that of those in exposure to the culture water of group C (∼60%). This indicates that the treatment of B. licheniformis efficiently controls the residual pathogenicity of SUS in culture water. The present study demonstrated the effectiveness of B. licheniformis treatment as an environmentally friendly approach to reducing mortality, improving symptoms, and controlling residual pathogenicity in sea cucumber aquaculture.

Effects of Scallop Visceral Mass and Mantle as Dietary Supplements on the Growth, Immune Response and Intestinal Microflora of Juvenile Sea Cucumber Apostichopus japonicus.

Scallop visceral mass and mantle are aquatic byproducts and waste, but they have high contents of protein. In this study, scallop visceral mass and mantle were used as supplements in the diet of juvenile sea cucumber (A. japonicus) and their effects on the growth, fatty acid and amino acid compositions, the non-specific immune responses and the intestinal microflora of A. japonicus were investigated through a 40 d feeding experiment. The results showed that dietary supplementation of scallop visceral mass significantly accelerated the specific growth rate (SGR) of juvenile A. japonicus by 3 times within 20 days, and also raised the contents of ω-3 fatty acids including EPA and DHA and the ω-3/ω-6 ratio of the sea cucumber tissue, which is favorable to the health and commercial value of the sea cucumber. Furthermore, it was found that the supplementation of scallop visceral mass and mantle stimulated the expression of immune-related genes and enhanced the immune defense in A. japonicus. Scallop visceral mass and mantle supplementation also increased the microbial diversity and the abundance of beneficial microbes including Bifidobacteriaceae, Streptomycetaceae, Clostridiaceae and Rhizobiales in the gut of A. japonicus. This study reveals the beneficial effects of dietary supplementation of scallop visceral mass and mantle on the growth of juvenile A. japonicus, which might be a promising way to reutilize this scallop waste and raise its economic value.

Adverse effects of polystyrene nanoplastics on sea cucumber Apostichopus japonicus and their association with gut microbiota dysbiosis

The mariculture environment is a sink of microplastics (MPs) due to its enclosed nature and mass use of plastics. Nanoplastics (NPs) are MPs with a diameter <1 μm that have a more toxic effect on aquatic organisms than other MPs. However, little is known about the underlying mechanisms of NP toxicity on mariculture species. Here, we performed a multi-omics investigation to explore gut microbiota dysbiosis and associated health problems induced by NPs in juvenile sea cucumber Apostichopus japonicus, a commercially and ecologically important marine invertebrate. We observed significant differences in gut microbiota composition after 21 days of NP exposure. Ingestion of NPs significantly increased core gut microbes, especially Rhodobacteraceae and Flavobacteriaceae families. Additionally, gut gene expression profiles were altered by NPs, especially those related to neurological diseases and movement disorders. Correlation and network analyses indicated close relationships between transcriptome changes and gut microbiota variation. Furthermore, NPs induced oxidative stress in sea cucumber intestines, which may be associated with intraspecies variation in Rhodobacteraceae in the gut microbiota. The results suggested that NPs were harmful to the health of sea cucumbers, and they highlighted the importance of the gut microbiota in the responses to NP toxicity in marine invertebrates.

Effects of Selenium Yeast Addition on the Growth, Intestinal Health, Immune Status and Body Composition of Juvenile Sea Cucumber Apostichopus japonicus before and after Aestivation

This study was performed to investigate the effects of selenium yeast (Se-yeast) on the growth, intestinal health, immune status and body composition of juvenile Apostichopus japonicus before and after aestivation. Five experimental diets were formulated with increasing addition of Se-yeast (0, 0.5, 1.0, 1.5 and 2.0 mg/kg), with the diet without Se-yeast as the control. Each diet was randomly assigned to three tanks of juvenile A. japonicus (initial body weight: 2.96 ± 0.04 g). The whole experiment lasted for 135 days, which included a 45-day feeding experiment before aestivation, a 60-day aestivation phage and a 30-day feeding experiment after aestivation. The results showed that weight gain rate (WGR) was significantly increased by the increasing addition of Se-yeast before aestivation. After aestivation, WGR was markedly elevated by 1.0 mg/kg Se-yeast but was inhibited by 1.5–2.0 mg/kg Se-yeast. The evisceration rate (ER) of A. japonicus was obviously inhibited by the relatively higher addition level of Se-yeast (1.5–2.0 mg/kg) before aestivation. After aestivation, the ER was significantly inhibited by Se-yeast at an addition of 1.5 mg/kg. Before aestivation, the highest activities of all digestive enzymes were obtained by Se-yeast addition at a level equal to or above 1.0 mg/kg Se-yeast. After aestivation, the activities of most digestive enzymes were maximized by Se-yeast at the addition level of no more than 1.0 mg/kg. Before aestivation, the activities of nitric oxide synthase and alkaline phosphatase significantly increased by the increasing addition of Se-yeast. After aestivation, immune related parameters exhibited the highest or comparable values when the addition level of Se-yeast was equal to or less than 1.0 mg/kg. Notably, the selenium content in the body wall of A. japonicus increased significantly as Se-yeast addition increased in the diets. These results showed that Se-yeast at the addition level of 1.5–2.0 mg/kg before aestivation and 1.0–1.5 mg/kg after aestivation was beneficial for the promotion of growth, intestinal health and immune status of juvenile A. japonicus.

An investigation on dietary chromium picolinate supplementation in the juvenile sea cucumber Apostichopus japonicus: Growth, digestive enzyme activity, growth-related genes expression, immune and antioxidant capacity

Improving the growth and immune responses are important to enhance the production efficiency of cultured juvenile sea cucumber Apostichopus japonicus. Here, 300 sea cucumbers (2.31 ± 0.01 g) were fed artificial diets with four concentrations of dietary chromium picolinate (Cr-Pic) supplementation (0, 0.3, 0.6 and 0.9 mg/kg) for 56 days. The growth performance, digestive enzyme activity (lipase, amylase and pepsin), expressions of growth-related genes (MAPK-7 and c-myc), and immune and antioxidant capacity were subsequently assessed. The present study found that final body weight, body weight gain and specific growth rate first increased and then declined with the increase of dietary Cr-Pic supplementation, while the highest values were observed in group 0.6 mg/kg. Consistently, significantly greater feed intake and significantly lower feed conversion ratio occurred in group 0.6 mg/kg than those in control group. Significantly better activities of lipase and amylase, and significantly higher expressions of genes MAPK-7 and c-myc were detected in group 0.6 mg/kg than those in control group. These results suggest that dietary Cr-Pic supplementation greatly improves the food utilization and expressions of growth-related genes, which consequently promotes the growth of sea cucumbers. Further, the activities of immune enzymes (alkaline phosphatase and lysozyme) and antioxidant enzymes (superoxide dismutase and total antioxidant capacity) in group 0.6 mg/kg were all significantly higher than those in control group. This indicates that dietary Cr-Pic supplementation significantly facilitates the immune and antioxidant capacity of sea cucumbers. In conclusion, we suggest dietary Cr-Pic supplementation with 0.6 mg/kg is the optimal level contributing to the growth and non-specific immune responses in the juvenile sea cucumber.

Macroalgal substitution effect in diet on growth, body composition, and stress resistance of juvenile sea cucumber (Apostichopus japonicus) subjected to air and low salinity exposures

Macroalgal substitution effect in diet on growth, body composition, and stress resistance of juvenile sea cucumber (Apostichopus japonicus) subjected to air and low salinity exposures

Biosynthesis and Isotopic Routing of Dietary Protein by Sea Cucumber Apostichopus japonicus (Selenka): Evidence from Compound-Specific Carbon Stable Isotope Analysis.

Compound-specific stable isotope analysis of individual amino acids (AAs) has been widely used in studies on food webs, resource tracing, and biochemical cycling. In the present study, juvenile sea cucumbers Apostichopus japonicus were fed the microalga Cylindrotheca fusiformis (CF) or Sargassum thunbergii (ST) during a 130-day experiment. The δ13C values of individual AAs in the experimental diet and body wall of sea cucumbers were determined to calculate the variability in carbon isotopic fractionation (Δ13C) and elucidate the isotopic routing of essential AAs and biosynthesis of nonessential AAs. The results showed that the sea cucumbers fed with diet CF had higher specific growth and ingestion rates but relatively lower feed conversion efficiency compared to those fed with diet ST. The experimental diets were generally less abundant in nonessential AAs (i.e., glycine, serine, aspartic acid, and arginine) but more abundant in essential AAs (i.e., isoleucine, leucine, lysine, phenylalanine, and histidine) than body walls. The fluctuations in the δ13C values of total AAs analyzed were 19.8 ± 4.6‰ for diets and 21.3 ± 2.7‰ for body walls. Serine and threonine were 13C-enriched AAs, while leucine and phenylalanine were 13C-depleted AAs. The diet ST treatment exhibited more positive Δ13C values of nonessential AAs (e.g., glycine, alanine, aspartic acid, and proline) compared to diet CF. There were significant negative relationships between Δ13C values and differences in nonessential AA percent abundance between the experimental diets and body walls of sea cucumber (for diet CF: y = -0.79 - 0.56x, r2 = 0.47; diet ST: y = 0.75 - 0.29x, r2 = 0.51), which implied the flexibility in the routing of various dietary macronutrients (protein, lipids, and carbohydrates) by sea cucumber. This study can greatly provide a new understanding of nutrient utilization and metabolism routing during juvenile sea cucumber culturing.

Adverse effects of dietary virgin (nano)microplastics on growth performance, immune response, and resistance to ammonia stress and pathogen challenge in juvenile sea cucumber Apostichopus japonicus (Selenka)

It has been well documented that micro- and nanoplastics are emerging pollutants in aquatic environments, and their potential toxic effects has attracted widespread concerns. Here, we evaluated the adverse effects of dietary polystyrene nanoplastics and microplastics (PS-N/MPs) on growth performance, oxidative stress induction, immune response, ammonia detoxification, and bacterial pathogen resistance of sea cucumber Apostichopus japonicus. After collection and acclimation, sea cucumbers were randomized into 3 groups (i.e., control, 100 nm PS-NPs and 20 µm PS-MPs at 100 mg kg−1 diet) for 60-day feeding experiment. Every group contained 360 sea cucumbers which were equally divided into 3 aquaria as biological triplicates. The results showed that the specific growth rate and final weight of the sea cucumbers fed with diets containing PS-N/MPs were significantly lower than those of control group. Dietary virgin PS-N/MPs significantly increased the reactive oxygen species production and malondialdehyde content in coelomic fluid, causing oxidative stress and damage to the growth and development of A. japonicus. During the experiment, 100 nm PS-NPs significantly induced the depletion in cellular and humoral immune parameters. The calculated IBR values based on multi-level biomarkers revealed the size-dependent toxic differences of PS-NPs > PS-MPs. The relative expression levels of GDH and GS mRNA showed first rise and then fall trends after exposure to ammonia, and 100 nm PS-NPs had a more profound impact on suppressing ammonia detoxification compared with 20 µm PS-MPs. Moreover, the expression of Hsp90, Hsp70, CL, TLR, and CASP2 genes were all down-regulated by ammonia exposure. Taken together of IBR results, ammonia stress test and pathogen challenge, we deduced that dietary 100 nm PS-NPs are more potentially hazardous than 20 µm PS-MPs. These findings provide valuable information for understanding the size-dependent toxic effects of PS-N/MPs and early risk warning on marine invertebrates.

Effects of Supplementary Selenium and Vitamin E on the Growth Performance, Antioxidant Enzyme Activity, and Gene Expression of Sea Cucumber Apostichopus japonicus.

A 60-day feeding experiment was conducted to evaluate the effects of single selenomethionine (Se) and its mixture with vitamin E (VE) on the growth, antioxidant enzyme activities, and gene expression of juvenile sea cucumber Apostichopus japonicus. The design of the experiment contained two factors and 5 × 2 levels by means of adding various levels of Se and VE in the feed, i.e., combination of 0, 0.3, 0.6, 0.9, or 1.2mg Se kg-1 and 0 or 200mg VE kg-1. The results revealed that the specific growth rate and weight gain rate were the highest in the group with 0.3mg Se kg-1 and 200mg VE kg-1, followed by the group with 0.6mg Se kg-1 without VE. Se significantly improved the activities of amylase and protease with VE also imposed positive effect on the amylase activity. Glutathione peroxidase (GPX) activity was highest in the group with 1.2mg Se kg-1 and lowest with the basal diet. The activity of catalase (CAT) was increased while glutathione reductase (GR) activity was decreased in response to the addition of Se. No significant interactive effects of Se and VE on the enzyme activities were found except superoxide dismutase (SOD) activity. While relative expressions of GPX, CAT, and SOD genes were significantly responsive to the addition of dietary Se, VE significantly promoted the gene expression of SOD. The results suggested that Se and VE might have beneficial effects on the growth and antioxidant responses of A. japonicus.

Bioaccumulation, Biotransformation, and Multicompartmental Toxicokinetic Model of Antibiotics in Sea Cucumber (Apostichopus japonicus).

Extensive application of antibiotics leads to their ubiquitous occurrence in coastal aquatic environments. However, it remains largely unknown whether antibiotics can be bioaccumulated and biotransformed in major mariculture organisms such as sea cucumbers and toxicokinetic models for Echinodermata are lacking. In this study, laboratory exposure experiments on juvenile sea cucumber (Apostichopus japonicus) were performed for seven antibiotics (sulfadiazine, sulfamethoxazole, trimethoprim, enrofloxacin, ofloxacin, clarithromycin, and azithromycin). Field sea cucumber and surrounding seawater samples were also analyzed. Results show that the sea cucumbers tend to accumulate high concentrations of the antibiotics with kinetic bioconcentration factors (BCFs) up to 1719.7 L·kg-1 for ofloxacin. The BCFs determined in the laboratory agree well with those estimated from the field measurements. Seven biotransformation products (BTPs) of the antibiotics were identified, four of which were not reported previously in aquatic organisms. The BTPs were mainly found in the digestive tract, indicating its high capacity in the biotransformation. A multicompartmental toxicokinetic model based on the principles of passive diffusion was developed, which can successfully predict time-course concentrations of the antibiotics in different compartments of the juvenile sea cucumbers. The findings may offer a scientific basis for assessing health risks and guiding healthy mariculture of sea cucumbers.

Effects of aerial exposure on oxidative stress, antioxidant and non-specific immune responses of juvenile sea cucumber Apostichopus japonicus under low temperature

Desiccation is a commonly stressful situation experienced by sea cucumber during transportation without/less water. The present study was conducted to investigate the effects of aerial exposure on the survival, oxidative damage, antioxidant capacity, immune-related response and gene expression of Apostichopus japonicus at different low temperatures. After acclimation, sea cucumbers were randomly divided into 3 groups, which were exposed to 5 °C, 10 °C and 15 °C in the closed laboratory condition, respectively. Each group has three parallel replicates. During the experiment, coelomic fluid and respiratory tree of A. japonicus were sampled at the time points of 0, 3, 6, 12, 24 and 48 h post-desiccation for further analysis. The results showed that the survival rates of sea cucumber significantly decreased as time prolonged, and those of 5 °C at 6–48 h of desiccation were significantly higher than 15 °C. Most oxidative damage parameters (e.g., O2− production, MDA, LPO and PC contents) significant increased after 6–12 h of desiccation. Antioxidant enzyme activities and T-AOC in coelomic fluid firstly increased and then decreased during aerial exposure, indicating that sea cucumber could adjust antioxidant defense to reduce the concentrations of ROS and MDA as a strategy for protecting organisms from oxidative damage in the early stage (0–6 h) of desiccation. The relative expression levels of Hsp90 and Hsp70 mRNA in respiratory tree of sea cucumber exhibited similar rise-fall trends with antioxidant parameters, while immune enzyme activities of ACP, AKP, LSZ and T-NOS, and gene expression of TLR, Rel and p105 all significantly decreased as time prolonged. Overall, low temperature postponed the process of ROS formation and the depression of antioxidant and non-specific immune responses of sea cucumber within a certain extent, which implied that it might play a positive role in improvement of desiccation tolerance. This study not only contribute to better understand the adaption mechanisms of A. japonicus to desiccation stress, but also provide valuable information for sea cucumber culture and transportation.

Colonization and Probiotic Effect of Metschnikowia sp. C14 in the Intestine of Juvenile Sea Cucumber, Apostichopus japonicus

Viable cell count was used to determine whether Metschnikowia sp. C14 can colonize the intestine of juvenile sea cucumber Apostichopus japonicus . Sea cucumber individuals were divided into two groups, which were fed the control diet for 38 days or the C14-supplemented diet at 10 5 cells g −1 diet for 28 days, then the control diet from day 29 to day 38. The number of C14 cells in the intestine of sea cucumber fed the C14-supplemented diet significantly increased from day 7 to day 28, and decreased from day 29 to day 38. Sea cucumber fed with the diet containing C14 showed a significant increase in trypsin activity and lipase activity from day 21 to day 33 compared with the control. Feeding C14 significantly improved the phagocytic activity and respiratory burst in coelomocytes from day 21 to day 35 and from day 14 to day 38, respectively. In addition, there was an obvious enhancement in lysozyme activity (from day 21 to day 38 or day 33), phenoloxidase activity (from day 21 to day 28) and total nitric oxide synthase activity (from day 14 to day 38) in coelomic fluid supernatant and/or coelomocyte cell lysate supernatant compared with the control. There were significant positive correlations between the number of C14 cells colonizing the intestine and trypsin activity of the intestine, lysozyme activity of the coelomic fluid supernatant and coelomocyte lysate supernatant from sea cucumber. These data suggested that the number of C14 cells should be maintained at 10 5 cfu (colony-forming units) g −1 intestine material for the maximum benefit.

Exploring the effects of phage cocktails in preventing Vibrio infections in juvenile sea cucumber (Apostichopus japonicus) farming

Vibrio infection poses a threat to the health of juvenile sea cucumber (Apostichopus japonicus) farming. In the present study, three phages, PVA1, PVc1 and PVS3, are respectively lytic to Vibrio alginolyticus, V. cyclitrophicus and V. splendidus and were used to make a phage cocktail with a ratio of 1:1:1 (V/V/V). Bioinformatic analyses showed that gene related toxins and virulence proteins were not detected in the genome of the three phages. Based on these results, a small pilot study was carried out to measure the effectiveness of a three-phage cocktail on sea cucumber farming. During the 48-day cultivation time, the total Vibrio counts in phage treatment groups decreased and phage particles were detected in the coelomic fluids of sea cucumbers over the cultivation time. In-vitro studies of those incubated with phages showed no significant cell viability loss (P > 0.05) compared to the control group. The level of superoxide dismutase and lysozyme in the phage immersion group remained lower than the control group (P < 0.05), while the activities of nitric oxide synthase and acid phosphatase remained stable. At the end of 48-day cultivation time, the total weight gain of the sea cucumbers in the phage cocktail immersion treatment group increased significantly (P < 0.01) compared to the control group, while remained lower than the antibiotic treatment group (P < 0.05). No significant differences were detected in the viscera body wall ratio and feed conversion among the three groups (P > 0.05). After the metagenomic analysis of sea cucumber intestinal tracts, the Vibrio abundance in the phage cocktail group decreased compared to control group, while there was no effect on the abundance of the other genera. Hence, all results of this small pilot study indicate that using a phage cocktail is a safe and feasible way to combat Vibrio infections in juvenile sea cucumber farming.

The effects of benzo[a]pyrene on the composition of gut microbiota and the gut health of the juvenile sea cucumber Apostichopus japonicus Selenka

The gut microbiota is essential for health and physiological functions in the host organism. However, the toxicological evaluation of environmental pollutants on the gut microbiota is still insufficient. In the present study, the juvenile sea cucumber Apostichopus japonicus was exposed for 14 days to Benzo[a]pyrene (BaP), which is a model polycyclic aromatic hydrocarbon (PAH), at four different concentrations (0, 0.5, 5, and 25 μg/L). We analyzed the intestinal microbial community of A. japonicas using 16S rRNA gene amplicon sequencing. Our results demonstrate that BaP exposure caused alterations to the microbiome community composition in sea cucumbers. At the phylum level, Planctomycetes were significantly more abundant in BaP exposure groups at 14 d compared with the control group, and the abundance of Proteobacteria and Bacteroidetes increased while the abundance of Firmicutes decreased following BaP exposure. At the genus level, multiple beneficial and autochthonous genera declined in the BaP treatment groups compared to the control, including Lactococcus, Bacillus, Lactobacillus, Enterococcus, Leuconostoc and Weissella; however, a bloom of alkane-degrading bacteria was found in BaP-exposed guts and included Lutibacter, Pseudoalteromonas, Polaribacter, Rhodopirellula and Blastopirellula. Furthermore, histological morphology, enzymatic activity and gene expression analysis revealed that BaP exposure also negatively impacted gut structure and function and presented as inflammation or atrophy, oxidative stress and immune suppression in sea cucumber intestines. Collectively, these findings provide insights into the toxic effects of BaP exposure on A. japonicas associated with intestinal microbiota and health.

High-throughput sequencing of 16S rRNA amplicons characterizes gut microbiota shift of juvenile sea cucumber Apostichopus japonicus feeding with three antibiotics

Sea cucumber Apostichopus japonicus is an important marine economic species in Asian countries due to its profound nutritional and medicinal value. So far, with the rapid development of intensified artificial aquaculture of sea cucumbers, the use of antibiotics is still an inexpensive and dispensable way to treat pathogenic infections, especially during the nursery phase. However, there is little information on the effects of antibiotics on the intestinal microbiota of sea cucumber. Therefore an Illumina based sequencing method was used to examine the intestinal bacterial composition of juvenile A. japonicas following diets with three typical antibiotics (tetracycline, erythromycin, and norfloxacin) under 15, 30, and 45 d. The findings reveal that different antibiotics have distinct effects on the growth performance of juvenile sea cucumbers. However, the richness and diversity of microbiota were barely affected by antibiotics but the community composition alterations indicated that the three antibiotics exhibited their respective patterns of reshaping the intestinal bacteria of juvenile sea cucumbers. In common, the abundance of some sensitive genera with helpful functions, such as Thalassotalea, Shewanella, Sulfitobacter, and Halomonas decreased significantly with exposure to antibiotics and the abundance of multiple potential pathogenic- and suspected antibiotic-resistant microorganisms like Arcobacter, Leucothrix, and Clostridium_sensu_stricto_1 was found increased significantly in the antibiotic groups. These results suggest that low doses of antibiotics could affect the composition of the intestinal microbiota of sea cucumbers and might increase the risk of infection of the hosts. This study could help us to explore how antibacterial compounds modify the gut microbiota of sea cucumbers and provide theoretical guidance in hatchery management by scientific antibiotic use in sea cucumber mariculture.

Growth, dispersal, and survival of juvenile Japanese sea cucumber Apostichopus japonicus cultured in a rearing facility with air-pocket fences

Growth, dispersal, and survival of juvenile Japanese sea cucumber Apostichopus japonicus cultured in a rearing facility with air-pocket fences

Relationships between body weight and other morphological traits in young sea cucumbers Apostichopus japonicas

Correlation, multiple regression, and path analyses were used to investigate the relationships between body weight and three other morphological traits in juvenile Japanese sea cucumbers Apostichopus japonicus. We measured live body weight (BW), body length (BL), numbers of papillae (NP), and numbers of tube feet (NF) at 60, 80, 100, and 130 days post-hatching (dph). We calculated path correlation coefficients, correlation indices (R2), and coefficients of determination with BW as the dependent variable and the other morphological traits as independent variables. The coefficient of variation for BW was high across all age groups, and all measured morphological traits were significantly correlated (P < 0.01). BL had the greatest direct effect on BW across all age groups (60 dph, 0.526; 80 dph, 0.404; 100 dph, 0.620; and 130 dph, 0.681), while NF had the greatest indirect effect on BW across all age groups (60 dph, 0.528; 80 dph, 0.452; 100 dph, 0.666; and 130 dph, 0.603). Regression analyses between morphological traits and BW indicated that R2 was greater than 0.85 only in the 100-dph specimens. The indirect effects of the other measured morphological traits on BW were age-dependent. The optimal regression equations, as determined with stepwise regression, were, for 60-dph specimens: BW60 =10 (-3.04+0.092BL+0.014NP+0.014NF) (R2 =0.632); for 80-dph specimens: BW80 =10 (- 3.035+0.056BL+0.017NP+0.02NF) (R2 =0.686); for 100-dph specimens: BW100 =10 (- 3.742+0.069BL+0. 633 × lg (NP)+0. 464 × lg (NF)) (R2 =0.893); and for 130-dph specimens: BW130 =10 (- 2.472+0.065BL+0.012NP) (R2 =0.774). Our work clarified the correlation between various morphological traits and body weight of a commercially-important sea cucumber species (A. japonicus). Our predictive models for body weight might be useful for the aquaculture and selective breeding of A. japonicus. These models might also provide theoretical support for the indirect selection of traits that are difficult to select directly.

The influence of three antibiotics on the growth, intestinal enzyme activities, and immune response of the juvenile sea cucumber Apostichopus japonicus selenka

The global abuse and misuse of antibiotics in the treatment and prevention of bacterial infections has resulted in the ubiquitous existence of these drugs in aquatic environments, which causes frequent antimicrobial resistance and pollution in ecosystems. However, the chronic effects of antimicrobial agents on aquatic animal growth and health have not been fully evaluated. In the present study, three typical antibiotics (tetracycline, erythromycin, and norfloxacin) were administered orally to juvenile sea cucumbers Apostichopus japonicus for 45 days, to mimic the long-term use of antibiotics. As a result, tetracycline and erythromycin promoted the growth and digestive activity of lipase, pepsin, and trypsin, but norfloxacin did not show significant prompting effect on digestive activity and even retarded the weight gain of the sea cucumbers. The mortality was higher in antibiotic treated groups between the 2nd and 4th days after challenge with Vibrio splendidus. At the same time, lower immune-related parameters were found in antibiotic feeding juveniles, suggesting that the use of antibiotics might weaken the immune defense system of sea cucumbers. This study revealed that antibiotic administration could facilitate the growth of sea cucumbers to varying degrees yet coupled with high risks of impaired immune function and compromised disease resistance.

An Evaluation on the Ratio of Plant to Animal Protein in the Diet of Juvenile Sea Cucumber (Apostichopus japonicus): Growth, Nutrient Digestibility and Nonspecific Immunity

This study was conducted to evaluate the effects of plant/animal (P/A) protein ratios (viz.1:4, 1:3, 1:2, 1:1,2:1, 3:1, 4:1) on growth performance, body composition, apparent digestibility of diets, and nonspecific immunity of juvenile sea cucumber (Apostichopus japonicus). Sea cucumbers were divided into 21 plastic tanks, and each tank was stocked with 15 individuals (initial weight: about 23.73 g). Each feed was allocated to three replicates of sea cucumbers. The feeding experiment lasted for 50 days. Results indicated that weight gain rate (WGR) and body wall weight (BWW) significantly increased as dietary ratio of P/A increased from 1:4 to 3:1, and then decreased significantly with further increase of this ratio (P 0.05). The apparent digestibility of dry matter, protein and lipid increased with ratio of P/A increasing from 1:4 to 2:1 (P < 0.05), and then decreased with further increase of this ratio. Correspondingly, activities of trypsin and amylase were significantly increased as P/A increased from 1:4 to 2:1 (P < 0.05). The activities of SOD and CAT showed a similar trend with WGR, with the highest value observed in the ratio of 1:2 and 1:1, respectively. Results above showed that moderate or relatively higher ratio of P/A protein (1:1–3:1) significantly increased the growth performance, apparent digestibility, and nonspecific immunity of sea cucumber. This will contribute to improving the feed formulation for juvenile cucumbers.

Effects of a Potential Autochthonous Probiotic Bacillus subtilis 2-1 on the Growth and Intestinal Microbiota of Juvenile Sea Cucumber, Apostichopus japonicus Selenka

The effects of Bacillus subtilis 2-1 from the intestine of healthy sea cucumber on the growth, digestive enzyme activities and intestinal microbiota of juvenile sea cucumber (Apostichopus japonicus) were determined in the present study. Sea cucumber was fed with Sargassum thunbergii powder supplemented with B. subtilis 2-1 at different concentrations varying among 0 (control), 105, 107, and 109 CFU g−1 for 8 weeks. Results showed that the growth performance and intestinal amylase and trypsin activities were significantly increased by dietary B. subtilis 2-1 at 109 CFU g−1 (P 0.05). The polymerase chain reaction denaturing gradient gel electrophoresis and 16S rRNA gene sequencing analysis indicated that dietary B. subtilis 2-1 at 105 and 107 CFU g−1 inhibited most of the Proteobacteria including those in genus Vibrio. Dietary B. subtilis 2-1 at 109 CFU g−1 not only decreased the abundance and species of genus Vibrio, but also increased the intensity of genera Psychrobacter and Bacillus. A specific dosage of dietary B. subtilis 2-1 could increase the growth and modulate the intestinal microbiota of sea cucumber; thus it might be a novel probiotic for keeping the health of sea cucumber.