Responses of growth performance, immunity, disease resistance of shrimp and microbiota in Penaeus vannamei culture system to Bacillus subtilis BSXE-1601 administration: Dietary supplementation versus water addition.

This study evaluated the effects of Bacillus subtilis BSXE-1601, applied either as dietary supplementation or water addition, on growth performance, immune responses, disease resistance of Penaeus vannamei, and microbiota in shrimp gut and rearing water. During the 42-day feeding experiment, shrimp were fed with basal diet (CO and BW group), basal diet supplemented with live strain BSXE-1601 at the dose of 1 × 109 CFU kg-1 feed (BD group) and 15 mg kg-1 florfenicol (FL group), and basal diet with strain BSXE-1601 added to water at the concentration of 1 × 107 CFU L-1 every five days (BW group). Results showed that dietary supplementation of strain BSXE-1601 significantly promoted growth performance of shrimp, both in the diet and water, enhanced disease resistance against Vibrio parahaemolyticus (P < 0.05). The BD and BW groups exhibited significant increases in acid phosphatase, alkaline phosphatase, lysozyme, peroxidase, superoxide dismutase activities, phenonoloxidase content in the serum of shrimp compared to the control (P < 0.05). Meanwhile, the expression of immune-related genes proPO, LZM, SOD, LGBP, HSP70, Imd, Toll, Relish, TOR, 4E-BP, eIF4E1α, eIF4E2 were significantly up-regulated compared to the control (P < 0.05). When added in rearing water, strain BSXE-1601 induced greater immune responses in shrimp than the dietary supplement (P < 0.05). Chao1 and Shannon indices of microbiota in rearing water were significantly lower in BD group than in the control. The microbiota in rearing water were significantly altered in BD, BW and FL groups compared to the control, while no significant impacts were observed on the microbiota of shrimp gut. When supplemented into the feed, strain BSXE-1601 obviously reduced the number of nodes, edges, modules in the ecological network of rearing water. The results suggested that dietary supplementation of BSXE-1601 could be more suitable than water addition in the practice of shrimp rearing when growth performance, non-specific immunity, disease resistance against V. parahaemolyticus in shrimp were collectively considered.

A Comparative Study on Effects of Three Butyric Acid-Producing Additives on the Growth Performance, Non-specific Immunity, and Intestinal Microbiota of the Sea Cucumber Apostichopus japonicus.

The providers of butyric acid, Clostridium butyricum (CB), sodium butyrate (SB), and tributyrin (TB), have been extensively studied as aquafeed additives in recent years. However, no comparative study has been reported on the probiotic effects of CB, SB, and TB as feed additives on sea cucumber (Apostichopus japonicus). A 63-day feeding trial was performed to assess the effects of dietary live cells of C. butyricum (CB group, the basal diet supplemented with 1% CB), sodium butyrate (SB group, the basal diet supplemented with 1% SB), and tributyrin (TB group, the basal diet supplemented with 1% TB) on the growth, non-specific immunity, and intestinal microbiota of A. japonicus with a basal diet group as the control. Results indicated that all three additives considerably increased A. japonicus growth, with dietary CB having the optimal growth-promoting effect. Of the seven non-specific enzyme parameters measured in coelomocytes of A. japonicus (i.e., the activities of phagocytosis, respiratory burst, superoxide dismutase, alkaline phosphatase, acid phosphatase, catalase, and lysozyme), dietary CB, SB, and TB considerably increased the activities of six, five, and six of them, respectively. The immune genes (Aj-p105, Aj-p50, Aj-rel, and Aj-lys) expression in the mid-intestine tissues of A. japonicus was significantly increased by all three additives. The CB group had the highest expression of all four genes. Additionally, the relative expression of Aj-p105, Aj-p50, and Aj-lys genes was significantly up-regulated in the three additive groups after stimulation with inactivated Vibrio splendidus. Dietary CB enhanced the intestinal microbial diversity and richness in A. japonicus while dietary TB decreased them. Meanwhile, dietary CB, SB, and TB significantly enhanced the abundance of Firmicutes, unclassified_f_Rhodobacteraceae, and Proteobacteria, respectively, while dietary CB and SB reduced the abundance of Vibrio. Dietary CB and SB improved the stability of microbial ecosystem in the intestine of A. japonicus. In contrast, dietary TB appeared to have a negative effect on the stability of intestinal microbial ecosystem. All three additives improved the intestinal microbial functions associated with energy production and immunity regulation pathways, which may contribute directly to growth promotion and non-specific immunity enhancement in A. japonicus. Collectively, in terms of enhancing growth and non-specific immunity, as well as improving intestinal microbiota, dietary live cells of C. butyricum exhibited the most effective effects in A. japonicus.

Nitrogen removal capability and mechanism of a novel heterotrophic nitrifying-aerobic denitrifying strain H1 as a potential candidate in mariculture wastewater treatment.

The nitrogen removal performance and mechanisms of Bacillus subtilis H1 isolated from a mariculture environment were investigated. Strain H1 efficiently removed NH4+-N, NO2--N, and NO3--N in simulated wastewater with removal efficiencies of 85.61%, 90.58%, and 57.82%, respectively. Strain H1 also efficiently degraded mixed nitrogen (NH4+-N mixed with NO2--N and/or NO3--N) and had removal efficiencies ranging from 82.39 to 89.54%. Nitrogen balance analysis revealed that inorganic nitrogen was degraded by heterotrophic nitrification-aerobic denitrification (HN-AD) and assimilation. 15N isotope tracing indicated that N2O was the product of the HN-AD process, while N2 as the final product was only detected during the reduction of 15NO2--N. The nitrogen assimilation and dissimilation pathways by strain H1 were further clarified using complete genome sequencing, nitrification inhibitor addition, and enzymatic activity measurement, and the ammonium oxidation process was speculated as NH4+ → NH2OH → NO → N2O. These results showed the application prospect of B. subtilis H1 in treating mariculture wastewater.

Influence of ammonia nitrogen management strategies on microbial communities in biofloc-based aquaculture systems.

Controlling ammonia nitrogen is very important in intensive aquaculture. This study evaluated how different management strategies, i.e., chemoautotrophic (control), heterotrophic bacterial enhancement using carbon in glucose or polyhydroxy butyrate-hydroxy valerate (PHBV), and mature biofloc application, affect water quality and microbial community structure and composition. The management strategies were examined during the domestication and fish culture stages. In the domestication stage, the average NO2--N concentration, pH, and DO in the glucose-added groups were significantly lower than those in the control and PHBV groups. All water quality parameters differed significantly among treatment groups in the culture stage. Carbon additions decreased both bacterial richness and diversity in the fish culture stage. Both principal coordinate analysis and hierarchical cluster analysis grouped the 33 bacteria community samples from the two stages into four clusters, which were closely related to management strategy. The dominant taxa of the clusters were identified using linear discriminant analysis effect size (LEfSe). The biomarkers of Cluster I included Marinomonas, Photobacterium, and Vibrio. Porticoccus and Clade-1a were identified as the biomarkers of Cluster II. Marivia, Leucothrix, and Phaeodactylibacter were identified as the biomarkers of Cluster IV. The Cluster I biomarkers were positively correlated with NO2--N, while those of Cluster IV were positively correlated with NO3--N. The redundancy analysis showed that the bacterial communities and biomarkers were influenced by water quality parameters. Quantitative real-time PCR analysis revealed significant differences in the abundances of the amoA and nxrB genes among treatments and between the two stages. The abundance of the amoA gene was higher in the control group than in the carton-added treatments at the ends of both stages. This study provides an important theoretical basis for the selection of efficient ammonia nitrogen control strategies in aquaculture systems.

Nitrogen removal capability and mechanism of a novel heterotrophic nitrification-aerobic denitrification bacterium Halomonas sp. DN3.

Recently, the functional microorganisms capable of eliminating nitrogenous waste have been applied in mariculture systems. As a potential candidate for treating mariculture wastewater, strain DN3 eliminated 100% of ammonia and nitrate and 96.61%-100% of nitrite within 72 h, when single nitrogen sources at concentrations of 0-50 mg/L. Strain DN3 also exhibited the efficient removal performance of mixed-form nitrogen (ammonia, nitrate, and nitrite) at salinity 30 ‰, C/N ratio 20, and 180 rpm. The nitrogen assimilation pathway dominated inorganic nitrogen metabolism, with less nitrogen (14.23%-25.02% of TN) lost into the air via nitrification and denitrification, based on nitrogen balance analysis. Moreover, the bacterial nitrification pathway was explored by enzymatic assays and inhibition assays. These complex nitrogen assimilation and dissimilation processes were further revealed by bacterial genome analysis. These results provide important insight into nitrogen metabolism of Halomonas sp. and theoretical support for treating mariculture wastewater with strain DN3.

A comparative study on effects of dietary three strains of lactic acid bacteria on the growth performance, immune responses, disease resistance and intestinal microbiota of Pacific white shrimp, Penaeus vannamei.

The present study evaluated the growth performance, immune responses, disease resistance and intestinal microbiota in Penaeus vannamei fed diets supplemented with three strains of lactic acid bacteria (LAB). The basal diet (control, CO) supplemented with Lactobacillus plantarum W2 (LA), Pediococcus acidilactici Nj (PE), Enterococcus faecium LYB (EN) and florfenicol (FL), respectively, formed three LAB diets (1 × 1010 cfu kg-1) and a florfenicol diet (15 mg kg-1, positive control), were fed to shrimp for 42 days. Results indicated that specific growth rate, feed efficiency rate, and disease resistance of shrimp against Vibrio parahaemolyticus in the treatment groups were significantly improved versus the control (P < 0.05). Compared with the control, acid phosphatase, alkaline phosphatase, phenonoloxidase, total nitric oxide synthase, peroxidase, superoxide dismutase activities, total antioxidant capacity, and lysozyme content in the serum and the relative expression levels of SOD, LZM, proPO, LGBP, HSP70, Imd, Toll, Relish, TOR, 4E-BP, eIF4E1α and eIF4E2 genes in the hepatopancreas of LAB groups were enhanced to various extents. Intestinal microbiota analysis showed that the LA and EN groups significantly improved microbial diversity and richness, and LAB groups significantly altered intestinal microbial structure of shrimp. At the phylum level, the Verrucomicrobiota in the LA and PE groups, the Firmicutes in the EN group, and the Actinobacteriota in the PE and EN groups were enriched. Moreover, the CO group increased the proportion of potential pathogens (Vibrionaceae and Flavobacteriaceae). The potential pathogen (Vibrio) was reduced, and potential beneficial bacteria (Tenacibaculum, Ruegeria and Bdellovibrio) were enriched in response to dietary three strains of LAB. When the intestinal microbiota homeostasis of shrimp is considered, L. plantarum and E. faecium showed better effects than P. acidilactici. However, due to the concerns on the possible potential risks of E. faecium strains to human health, L. plantarum W2 is more suitable for application in aquaculture than E. faecium LYB. Considering collectively the above, Lactobacillus plantarum W2 could be applied as better probiotic to improve the growth performance, non-specific immunity, disease resistance and promote intestinal health of P. vannamei.

Effects of dietary yeast culture on health status in digestive tract of juvenile Pacific white shrimp Litopenaeus Vannamei.

A feeding trial was conducted to investigate the effects of dietary yeast culture (YC) on health status in digestive tract of juvenile Pacific white shrimp Litopenaeus Vannamei. Shrimps (initial weight: 3.33 ± 0.06 g) were fed with graded levels of dietary YC (control, 0.3%, 0.5% and 1.0%). Results of the present study showed that villus height and the ratio between villus height and crypt depth in the digestive tract of juvenile shrimp was significantly increased by dietary 0.5% and 1.0%YC (P < 0.05). Besides, dietary 0.5% and 1.0%YC significantly activities of phenoloxidase (PO), lysozyme (LZ), acid phosphatase (ACP) and alkaline phosphatase (AKP) (P < 0.05), significantly up-regulated mRNA levels of prophenoloxidase (propo), lysozyme (lz), anti-lipopolysaccharide factor (alf), crustin and penaienadin (P < 0.05) and down-regulated mRNA levels of caspase-1, nuclear factor κB p65 (nf-κbp65) myeloid differentiation primary response protein (myd88) and toll like receptor (tlr) in the digestive tract of juvenile shrimp (P < 0.05). Compared with the control, dietary 0.5%YC increased Chao1 index in the digestive tract of juvenile shrimp. In addition, compared with the control, dietary 0.5% and 1.0%YC significantly increased relative abundance of Lactobacillus (P < 0.05). It can be concluded that dietary YC made positive contribution to health status in digestive tract of juvenile shrimp through improving morphology and microbiota, enhancing immune function, and inhibiting inflammation of digestive tract.

Evaluation of Potential Probiotic Properties of a Strain of Lactobacillus plantarum for Shrimp Farming: From Beneficial Functions to Safety Assessment.

In recent years the safety of probiotics has received increasing attention due to the possible transfer and spread of virulence factors (VFs) and antibiotic resistance genes (ARGs) among microorganisms. The safety of a strain of Lactobacillus plantarum named W2 was evaluated in phenotype and genotype in the present study. Its probiotic properties were also evaluated both in vivo and in vitro, including adherence properties, antibacterial properties and beneficial effects on the growth and immunity of Pacific white shrimp, Penaeus vannamei. Hemolysis tests, antibiotic resistance tests and whole genome sequence analysis showed that W2 had no significant virulence effects and did not carry high virulence factors. W2 was found to be sensitive to chloramphenicol, clindamycin, gentamicin, kanamycin and tetracycline, and to be resistant to ampicillin and erythromycin. Most ARGs have no transfer risk and a few have transfer risk but no significant enrichment in human-associated environments. The autoaggregation of W2 was 82.6% and the hydrophobicity was 81.0%. Coaggregation rate with Vibrio parahaemolyticus (24.9%) was significantly higher than Vibrio's autoaggregation rate (17.8%). This suggested that W2 had adhesion potential to mucosal/intestinal surfaces and was able to attenuate the adherence of V. parahaemolyticus. In addition, several adhesion-related protein genes, including 1 S-layer protein, 1 collagen-binding protein and 9 mucus-binding proteins were identified in the W2 genome. W2 had efficiently antagonistic activity against 7 aquatic pathogenic strains. Antagonistic components analysis indicated that active antibacterial substances might be organic acids. W2 can significantly promote the growth of shrimp when supplemented with 1 × 1010 cfu/kg live cells. Levels of 7 serological immune indicators and expression levels of 12 hepatopancreatic immune-related genes were up-regulated, and the mortality of shrimp exposed to V. parahaemolyticus was significantly reduced. Based on the above, L. plantarum W2 can be applied safely as a potential probiotic to enhance the growth performance, immunity capacity and disease resistance of P. vannamei.

Profiling intestinal microbiota of Metaplax longipes and Helice japonica and their co-occurrence relationships with habitat microbes.

Intestinal microbiota plays key roles in maintaining the health and homeostasis of the host. However, information about whether the formation of intestinal microbiota of wild aquatic animals is associated with habitat microbes is not fully understood. Here, intestine samples were collected from two wild crab species and sediment samples were collected from the habitat environment. The total DNA of each sample was extracted, and the V3-V4 regions of 16S rRNA were sequenced using the MiSeq platform. The purpose of this study was to investigate the composition and diversity of intestinal microbiota and habitat microbes, and bacterial community relationships between wild crab intestine and habitat sediment. In the present study, the composition and diversity of intestinal microbiota of the two crab species were different from the habitat microbes. In contrast, a similar composition and diversity of the intestinal microbiota were observed between two crab species. Moreover, the bacterial community relationships between crab intestine and habitat sediment were associated with intestinal regions. Further network analysis revealed that the network structure of the intestinal microbiota was not only associated with intestinal regions, but also with the crab species. Additionally, although the compositions of bacterial functions were similar between crab intestine and sediment, no significant correlation in bacterial functions was observed between crab intestine and sediment. The findings of the present study would contribute to understanding the relationship between intestinal microbiota of wild aquatic animal and habitat microbes, and providing new insights into the intestinal microbiota of wild aquatic animals.

Mechanism of the Potential Therapeutic Candidate Bacillus subtilis BSXE-1601 Against Shrimp Pathogenic Vibrios and Multifunctional Metabolites Biosynthetic Capability of the Strain as Predicted by Genome Analysis.

The global shrimp industry has suffered bacterial diseases caused mainly by Vibrio species. The typical vibriosis, acute hepatopancreatic necrosis disease (AHPND), has resulted in mass mortality and devastating economic losses. Thus, therapeutic strategies are highly needed to decrease the risk of vibriosis outbreaks. Herein, we initially identified that the growth of the causative agent of AHPND, Vibrio parahaemolyticus (VP AHPND ) and other vibrios in Pacific white shrimp (Litopenaeus vannamei) was inhibited by a Bacillus subtilis strain BSXE-1601. The natural products amicoumacins A, B, and C were purified from the cell-free supernatant from the strain BSXE-1601, but only amicoumacin A was demonstrated to be responsible for this anti-Vibrio activity. Our discovery provided the first evidence that amicoumacin A was highly active against shrimp pathogens, including the representative strain VP AHPND . Furthermore, we elucidated the amicoumacin A biosynthetic gene cluster by whole genome sequencing of the B. subtilis strain BSXE-1601. In addition to amicoumacin A, the strain BSXE-1601 genome harbored other genes encoding bacillibactin, fengycin, surfactin, bacilysin, and subtilosin A, all of which have previously reported antagonistic activities against pathogenic strains. The whole-genome analysis provided unequivocal evidence in support of the huge potential of the strain BSXE-1601 to produce diverse biologically antagonistic natural products, which may facilitate further studies on the effective therapeutics for detrimental diseases in shrimp.

Temporal bacterial community succession during the start-up process of biofilters in a cold-freshwater recirculating aquaculture system.

Start-up of biofilters plays crucial roles in the successful operation of recirculating aquaculture system, and the nature is bacterial community succession. We explored the pattern of bacterial temporal succession during the start-up process of biofilters in a commercial cold-freshwater recirculating aquaculture system (RAS). The whole succession process was divided into three distinct phases: incubation, growth and stability. Phylogenetic diversity and evenness of the bacterial community increased during the start-up process, whereas richness reached its peak at the growth phase. Seven biomarkers were identified, namely Cytophagales, Gemmatimonadales, Sphingomonadales, Sphingobacteriales, Rhizobiales, Clostridiales and Nitrospirales. The relative abundances of these functional bacteria increased, while those with a competitive growth advantage declined. The network interactions were dramatically altered from fairly simple to most complex, and then decreased in complexity during start-up. Positive relationships decreased, while competition increased. The shift in predicted function exhibited a trend from simple to diverse, and converged to idiosyncratic configuration.

Transcriptome signatures of the Pacific white shrimp Litopenaeus vannamei hepatopancreas in response to stress in biofloc culture systems.

Comparative transcriptome analysis via high throughput sequencing was applied to gain knowledge on the immune response in Litopenaeus vannamei reared in biofloc technology systems (BFT). Two types of carbon sources, namely, traditional carbon sources (molasses) and biodegradable polymers [hydroxybutyric acid-co-3-hydroxyvaleric acid (PHBV)] were used in BFT systems. Clear water systems without the addition of carbon sources were treated as the control. Water quality assays showed that the average concentrations of several stress factors, including nitrite, nitrate and TSS, were the highest in molasses-based BFT systems. After sequencing and comparing the transcriptome profiles of the L. vannamei hepatopancreas, 743 and 201 genes were significantly differentially expressed in molasses- and PHBV-based BFT systems, respectively. GO enrichment analysis, which was performed using the differentially expressed genes, revealed seven significantly over-represented GO terms in molasses-based BFT systems, including catabolic process, hydrolase activity, cellular localization, organic substance metabolic process, cellular metabolic process, establishment of localization and response to stress. The captured key genes were mainly involved in the pathways including cellular stress response, immune response and pathogen recognition. However, no GO terms were significantly over-represented in PHBV-based BFT systems compared with control. This study indicates that shrimp are subject to stress in BFT systems when molasses serves as the carbon source. Thus, PHBV may be a better alternative.

Effect of Clostridium butyricum in different forms on growth performance, disease resistance, expression of genes involved in immune responses and mTOR signaling pathway of Litopenaeus vannamai.

A 42-day feeding trial was conducted to evaluate the effects of diet supplemented with various additives from Clostridium butyricum (fermentation supernatant, FS; live cells, LC; cell-free extract, CE; spray-dried spores, DS; mixture of live cells and supernatant, LCS) on the growth, intestinal morphology, disease resistance, immune gene expression and mTOR signaling-related gene expression in Litopenaeus vannamai. The feeding trial showed that the final weight and specific growth rate of the shrimp were improved significantly while the feed conversion ratio were reduced significantly in LC, CE, DS and LCS groups compared to the control. The villus height and intestinal wall thickness of shrimp's mid-intestine in LC, DS and LCS group increased significantly. After challenge test to Vibrio parahaemolyticus, the cumulative mortalities of the shrimp in LC, CE, DS and LCS groups were significantly lower than that of the control. As compared to the control, the relative expression levels of superoxide dismutase, lysozyme, prophenoloxidase (proPO), Toll, Immune deficiency (Imd), Relish, TOR, 4E-BP, eIF4E1α and eIF4E2 genes in the shrimp of DS and LCS groups enhanced significantly, whereas the relative expression levels of proPO, SOD, Toll, Imd, Relish, elF4E1α and elF4E2 genes were statistically the same between FS group and the control. These results suggested that the spray-dried spores and mixture of live cells and supernatant of C. butyricum exerted better probiotic benefits in modulating immune responses of shrimp. In addition, single supernatant could not be helpful to shrimp while mixture of live cells and supernatant could better improve the immune responses of shrimp in comparison to single live cells. The integration of C. butyricum and their metabolites supplemented into feed could significantly improve growth performance, intestinal morphology, immunity capacity and resistance against V. parahaemolyticus of L. vannamei.

Metabolic responses in the gills of tongue sole (Cynoglossus semilaevis) exposed to salinity stress using NMR-based metabolomics.

Salinity is an important environmental factor affecting fish physiology. Tongue sole (Cynoglossus semilaevis) is a euryhaline species that can survive in a wide range of salinity, and might be used as a promising model animal for environmental science. In this study, by using the nuclear magnetic resonance (1H NMR)-based metabolomics, amino acids analysis and real-time quantitative PCR assay, we investigated the metabolic responses in the gills and plasma of tongue sole subjected to hypo- (0 ppt, S0) and hyper-osmotic stress (50 ppt, S50) from isosmotic environment (30 ppt, S30). The results showed that the metabolic profiles of S50 were significantly different from those of S0 and S30 groups, and a clear overlap was found between the latter two groups. Ten metabolites were significantly different between the salt stress groups and the isosmotic group. Taurine and creatine elevated in both S0 and S50 groups. Choline decreased in S50 group while increased in S0 group. Amino acids and energy compounds were higher in the gills of S50 group. The metabolic network showed that ten metabolic pathways were all found in S50 group, while seven pathways were observed in S0 group. Meanwhile, the transcript levels of the Tau-T and ATP synthase in the gills increased with increasing salinity. Aspartate and methionine exhibited significant differences in the plasma among the groups, but did not show differences in the gills. Comparatively, glutamate exhibited significant differences both in the plasma and the gills. Overall, these findings provide a preliminary profile of osmotic regulation in euryhaline fish.

Effects of l-tryptophan on the growth, intestinal enzyme activities and non-specific immune response of sea cucumber (Apostichopus japonicus Selenka) exposed to crowding stress.

In order to reveal the effects of l-tryptophan (Trp) on the physiology and immune response of sea cucumber (Apostichopus japonicus Selenka) exposed to crowding stress, four density groups of sea cucumbers (i.e. 4, 8, 16 and 32 individuals per 40 L water, represented as L, ML, MH and H) were fed with diets containing 0, 1, 3 and 5% l-tryptophan respectively for 75 days. The results showed that the specific growth rates (SGR) of the sea cucumber fed with diet with 3% Trp (L, 2.1; ML, 1.76; MH, 1.2; H, 0.7) were significantly higher than those fed with basal diet without Trp supplementation (P < .05). Peak amylase activity occurred at H stress density at 3% dietary Trp. Trypsin activity was higher in diet 3% in ML and MH densities than the controls, which increased by 66.4% and 53.8%. However, the lipase activity first increased and then decreased from the stocking density L to H, with highest values of 3% Trp group showed the highest value than other groups. Compared to those fed with the basal diet, sea cucumber fed diets with Trp (3%) had significantly higher phagocytic activities (0.28 OD540/106 cells, H) in coelomic fluid and respiratory burst activities (0.105 OD630/106 cells, MH) (P < .05). The results suggested that Trp cannot improve superoxide dismutase (SOD) activity at L, ML and MH densities. The alkaline phosphatase activity (AKP) significantly decreased at H stress density. Under the experimental conditions, the present results confirmed that a diet supplemented with 3% Trp was able to enhance intestinal enzyme activities, non-specific immune response and higher growth performance of A. japonicus.

Genetic adaptation of microbial populations present in high-intensity catfish production systems with therapeutic oxytetracycline treatment.

Microbial communities that are present in aquaculture production systems play significant roles in degrading organic matter, controlling diseases, and formation of antibiotic resistance. It is important to understand the diversity and abundance of microbial communities and their genetic adaptations associated with environmental physical and chemical changes. Here we collected water and sediment samples from a high-intensity catfish production system and its original water reservoir. The metagenomic analysis showed that Proteobacteria, Actinobacteria, Bacteroidetes, Cyanobacteria, and Firmicutes were the top five phyla identified from all samples. The aquaculture production system significantly changed the structure of aquatic microbial populations. Substantial changes were also observed in SNP patterns among four sample types. The gene-specific sweep was found to be more common than genome-wide sweep. The selective sweep analysis revealed that 21 antibiotic resistant (AR) genes were under selection, with most belonging to antibiotic efflux pathways. Over 200 AR gene gains and losses were determined by changes in gene frequencies. Most of the AR genes were characterized as ABC efflux pumps, RND efflux pumps, and tetracycline MFS efflux pumps. Results of this study suggested that aquaculture waste, especially waste containing therapeutic antibiotics, has a significant impact on microbial population structures and their genetic structures.

Molecular ecological network analysis reveals the effects of probiotics and florfenicol on intestinal microbiota homeostasis: An example of sea cucumber.

Animal gut harbors diverse microbes that play crucial roles in the nutrition uptake, metabolism, and the regulation of host immune responses. The intestinal microbiota homeostasis is critical for health but poorly understood. Probiotics Paracoccus marcusii DB11 and Bacillus cereus G19, and antibiotics florfenicol did not significantly impact species richness and the diversity of intestinal microbiota of sea cucumber, in comparison with those in the control group by high-throughput sequencing. Molecular ecological network analysis indicated that P. marcusii DB11 supplementation may lead to sub-module integration and the formation of a large, new sub-module, and enhance species-species interactions and connecter and module hub numbers. B. cereus G19 supplementation decreased sub-module numbers, and increased the number of species-species interactions and module hubs. Sea cucumber treated with florfenicol were shown to have only one connecter and the lowest number of operational taxonomic units (OTUs) and species-species interactions within the ecological network. These results suggested that P. marcusii DB11 or B. cereus G19 may promote intestinal microbiota homeostasis by improving modularity, enhancing species-species interactions and increasing the number of connecters and/or module hubs within the network. In contrast, the use of florfenicol can lead to homeostatic collapse through the deterioration of the ecological network.

Impact of water temperature on the growth and fatty acid profiles of juvenile sea cucumber Apostichopus japonicus (Selenka).

The present study determined the changes in the fatty acid (FA) profiles of juvenile sea cucumber Apostichopus japonicus in response to the varied water temperature. Sea cucumbers with similar size (4.02±0.11g) were cultured for 8 weeks at 14°C, 18°C, 22°C and 26°C, respectively. At the end of the experiment, the specific growth rate (SGR) and the profiles of FAs in neutral lipids and phospholipids of the juvenile sea cucumbers cultured at different temperatures were determined. The SGRs of the sea cucumbers cultured at 26°C significantly decreased 46.3% compared to thos cultured at 18°C. Regression analysis showed that the SGR-temperature (T) relationship can be expressed as SGR=-0.0073T(2)+0.255T -1.0231 (R(2)=0.9936) and the highest SGR was predicted at 17.5°C. For the neutral lipids, the sum of saturated FAs (SFAs), monounsaturated FAs (MUFAs) or polyunsaturated FAs (PUFAs) of the sea cucumbers that were cultured at the water temperature from 18°C-26°C did not change significantly, indicating the insensitivity of FA profiles for the neutral lipids of sea cucumbers in response to increasing water temperature. For phospholipids, the sum of PUFAs in the sea cucumbers dramatically decreased with the gradually increased water temperature. The sum of SFAs and MUFAs of sea cucumbers, however, increased with the gradually elevated water temperature. In particular, the contents of highly unsaturated fatty acids (HUFAs), including eicosapentaenoic acid (EPA, 20:5n-3) and docosahexaenoic acid (DHA), in the phospholipids of the sea cucumbers decreased 37.2% and 26.1%, respectively, when the water temperature increased from 14°C to 26°C. In summary, the sea cucumbers A. japonicus can regulate the FA compositions, especially the contents of EPA and DHA, in the phospholipids so as to adapt to varied water temperature.

Effects of Acute and Chronic Heavy Metal (Cu, Cd, and Zn) Exposure on Sea Cucumbers (Apostichopus japonicus).

Acute and chronic toxicity tests were conducted with sea cucumber (Apostichopus japonicus) exposed to heavy metals. Acute toxicity values (96 h LC50) were 2.697, 0.133, and 1.574 mg L(-1) for Zn, Cu, and Cd, respectively, and were ranked in order of toxicity: Cu > Cd > Zn. Under chronic metal exposure the specific growth rates of sea cucumbers decreased with the increase of metal concentration for all the three metals. After acute metal exposure, the oxygen consumption rate (OCR) decreased. The OCRs in all groups were significantly different than control (P < 0.05) except in the group treated with 1.00 mg L(-1) Zn (P < 0.05), where the increase of OCR was observed. The OCRs in groups chronically exposed to metals were significantly lower than that in the control group (P < 0.05). The activity of both pyruvate kinase (PK) and hexokinase (HK) in sea cucumbers followed: respiratory tree > muscle > intestine in natural sea water. After chronic Zn, Cu, and Cd exposure, the change pattern of HK and PK in respiratory tree, muscle, and intestine varied slightly. However, the activity of the enzyme showed a general trend of increase and then decrease and the higher the exposure concentration was, the earlier the highest point of enzyme activity was obtained.

Carbon dioxide and methane fluxes from feeding and no-feeding mariculture ponds.

The CO2 and CH4 fluxes at the water-air interface were measured in shrimp (Marsupenaeus japonicus) monoculture ponds (S) with feed supply and shrimp-sea cucumber (Apostichopus japonicus) polyculture ponds (SS) without feed supply. During farming seasons of the whole year, cumulated CO2-C fluxes were -5.69 g m(-2) (S) and 11.23 g m(-2) (SS), respectively. The cumulated CO2 emissions from S and SS did not differ significantly. The cumulated CH4-C emissions from S (0.57 g m(-2)) were significantly higher than those from SS (0.068 g m(-2)). S absorbed C from the atmosphere with a mean absorption rate of -5.12 g m(-2), whereas SS emitted C to the atmosphere with a mean emission rate of 11.30 g m(-2). Over 20-year horizon, the compressive global warming potentials (cGWPs) were 33.55 (S) and 47.71 (SS), respectively, indicating both feeding and no-feeding mariculture ponds could promote global warming.