The relationship between water quality and bacterial composition may determine the sustainability level in an integrated multi-trophic aquaculture (IMTA) system. In this study, the effects of different shrimp stocking densities on water quality and bacterial community structure were studied in a sea cucumber-shrimp-crab-fish IMTA system. The results showed correlations between bacterial community structure and water quality parameters (dissolved oxygen, salinity, pH, ammonia nitrogen, nitrite nitrogen, and phosphate phosphorus) under different stocking densities of Penaeus japonicus . Temperature, salinity, nitrite nitrogen, ammonia nitrogen, and phosphate phosphorus were significant factors ( p < 0.05) influencing the bacterial community in the samples. Nitrite nitrogen showed the greatest effect on bacterial community diversity and richness. Flavobacteriaceae and Cryomorphaceae play important roles in the denitrification process, and these bacteria were significantly more abundant in low-density ponds than in high-density ponds. This study provides practical knowledge on shrimp stocking density management and its effect on IMTA system biosecurity.

Integration of biological activities of the algal and bacterial communities enhances the bioremediation potency of aquaculture systems. The effects of nitrogen removal mediated by Ulva lactuca on the bacterial community structure and the abundance of nitrogen cycle functional genes were investigated using an integrated multi-trophic aquaculture (IMTA) system mainly composed of sea cucumber (Stichopus japonicus), shrimp (Penaeus japonicus), and crab (Portunus trituberculatus). The experimental treatments were separated into two groups: control group (C: without U. lactuca) and algae treatment (A: with U. lactuca). Microbial diversity and abundance indexes, including the Sobs, Shannon, Ace, and Chao1 indexes, were higher in the U. lactuca treatment group in both water and sediment. Flavobacteriaceae and Rhodobacteraceae were the dominant families in both the U. lactuca and control treatment groups in October and November, respectively. In sediment, Bacillaceae was the dominant family in the U. lactuca treatment group throughout the experimental period, whereas Desulfocapsaceae was the dominant family in the control group in October and November. Moreover, the nitrogen cycle functional genes nifH, amoA, nxrB, norB, and nrfA were more abundant in the U. lactuca treatment group than in the control group. Results of water quality and its correlation with bacterial community were comprehensively investigated, revealing that U. lactuca influenced the bacterial community structure and nitrogen cycle by increasing DO in the IMTA system. In conclusion, U. lactuca co-cultured in an IMTA system could represent a novel approach for enhancing nitrogen removal, based on the interaction between the algal and bacterial communities.

Abstract To study nitrogen removal by Ulva lactuca and its effects on the structure of the bacterial community and abundance of nitrogen cycle functional genes in the ‘sea cucumber–shrimp–crab–fish’ integrated multi-trophic aquaculture (IMTA) system, we analysed bacterial community composition by Illumina MiSeq high-throughput sequencing and nitrogen cycling functional gene levels by real-time quantitative PCR in water and sediment with (Y) and without the presence of U. lactuca (N). Diversity and abundance indices, including Sobs, Shannon, Ace, and Chao1, were higher in Group Y water and sediment. In the water, the predominant phyla were Proteobacteria in Group Y and Cyanobacteria in Group N in September. Flavobacteriaceae and Rhodobacteraceae were the dominant families in Group Y and Group N in October and November, respectively. In the sediment, Bacillaceae was the dominant family in Group Y throughout the experimental period, whereas Desulfocapsaceae was the dominant family in Group N in October and November. Regarding nitrogen cycle functional genes, millions of Amx, nosZ, nxrB, and nrfA genes were detected in both water and sediment. The abundances of nifH, amoA, nxrB, norB, and nrfA genes were higher in Group Y than those in Group N, which can improve ammonia removal without nitrogen loss. In this study, a potential novel approach was proposed to enhance the development of sustainable aquaculture, promote the nitrogen cycle, and remediate and optimise the water environment by cultivating bacteria and algae, according to the effects of U. lactuca on water quality, bacterial communities, and abundance of functional nitrogen cycling genes in IMTA systems.

Integrated multitrophic aquaculture (IMTA) maximises the nitrogen cycle between system components, including bacteria. In order to maximise the bacterial role in nitrogen elimination in an IMTA system, we investigated the effect of bivalve culture on water quality and bacterial community structure in overlying water and sediment in the “shrimp-crab-bivalve-fish” IMTA system. The bacterial composition in overlying water and sediment was measured by Illumina -MiSeq high-throughput sequencing technology. The results show that dissolved oxygen was higher in the bivalve culture area. Ammonia and nitrite in the bivalve culture area were lower than those in the nonbivalve culture area; however, the nitrate and phosphate in the bivalve culture area were higher than those in the nonbivalve culture area. The Chao1, Shannon, and Ace indexes were higher in the bivalve area. More bacteria with nitrification and denitrification functions were detected in bivalve culture areas, such as Ruegeria (1.05%–4.79%), Thalassobius (0.11%–0.69%), Limibaculum (0.07%–0.69%), HIMB11 (0.13%–0.21%), and Rubellimicrobium (0.01%–0.16%). More Cyanobacteria were detected in bivalve culture areas with higher phosphate concentrations. To sum up, bivalves can release phosphorus through bioturbation, increasing the abundance of Cyanobacteria, which release dissolved oxygen into overlying water through photosynthesis, enhance nitrification (mainly ammonia oxidation), and improve the ammonia nitrogen removal capacity of the system. Meanwhile, bivalves can increase bacterial diversity and abundance by regulating dissolved oxygen. This study provided insight into bivalve interaction with bacterial activity in the IMTA system.

Astaxanthin (Axn) is a xanthophyll carotenoid that has previously been shown to suppress hepatic inflammation, reduce oxidative liver damage, and improve metabolic profiles. Exopalaemon carinicauda ( E. carinicauda ) is an economically important fishery species in China that has been found to exhibit increased body weight following Axn feeding as compared to a standard diet. In this study, dietary Axn can significantly decreased MDA content, T-AOC and significantly increased SOD, GSH and CAT activities in shrimp hepatopancreas. Moreover, transcriptome and metabolome of E. carinicauda after Axn feeding were investigated to identify the mechanism of the effect of Axn on E. carinicauda . The transcriptomic data revealed that a total 99 different expression genes (DEGs) were identified between the Axn and control groups, of which 47 and 52 were upregulated and downregulated, respectively. DEGs of E. carinicauda such as catherpsin, actin and PARP after Axn feeding were associated with apoptosis and immune system. The metabolomic analysis revealed that A total of 73 different expression metabolites (DEMs) were identified in both metabolites, including 30 downregulated metabolites and 43 upregulated metabolites. And Axn participate in metabolism processes in hepatopancreas of E. carinicauda , including the TCA cycle, amino acid metabolism and lipid metabolism. The multiple comparative analysis implicated that Axn can improve the antioxidant capacity of hepatopancreas and the energy supply of hepatopancreas mitochondria, and then improve the ability of anti-apoptosis. Collectively, all these results will greatly provide new insights into the molecular mechanisms underlying tolerance of adverse environment in E. carinicauda .

颗粒酶(granzyme,Gzm)B是免疫炎症反应必不可少的介质,可激活半胱天冬酶3,进而诱导靶细胞的凋亡.本研究通过PCR扩增和RACE技术获得了半滑舌鳎(Cynoglossus semilaevis)颗粒酶B基因(CsGzmBl)的全长cDNA序列,并对其序列特征和表达水平进行了分析.结果显示,CsGzmBl cDNA全长为923bp,开放阅读框长度为780bp,编码259个氨基酸(前19个氨基酸残基为信号肽序列),5'非编码区为49bp,3'非编码区为94bp.CsGzmBl的基因组结构比较保守,由5个外显子和4个内含子组成.CsGzmBl蛋白包含2个N端糖基化位点、1个催化三联体"His63Asp 112Ser207"、1个"PHSRPYMA"结构域及6个保守的半胱氨酸.荧光定量PCR结果显示,CsGzmBl在半滑舌鳎健康成鱼不同组织中均有表达,其中,在脾脏中表达量最高,头肾、中肾、肝脏和鳃中表达量次之,在肌肉中表达量最低.与对照组0h相比,CsGzmBl在哈维氏弧菌(Vibrio harveyi)感染后的不同时间点的脾脏、肠、肝脏、皮肤、鳃和肾脏中的表达水平均有不同程度的上调.研究表明,CsGzmBl基因在半滑舌鳎抵御哈维氏弧菌感染过程中发挥重要作用.

Astaxanthin (Axn), a feed additive, is becoming increasingly important for modulating the metabolism, growth, development, and reproduction of aquatic organisms in aquaculture. In this study, Exopalaemon carinicauda (E. carinicauda) is an economically important fishery species in China that has been found to exhibit increased body weight following Axn feeding as compared to a standard diet. The antioxidant, transcriptomic, and metabolomic analyses of the response of E. carinicauda after Axn feeding were investigated. Axn could reduce the content of malondialdehyde and increase the activities of various antioxidant enzymes, which also proved that axn can improve the antioxidant capacity Transcriptomic analysis suggested that synthesis and secretion of immune proteins, cytoskeleton structure, and apoptosis signaling were altered after Axn feeding. The metabolic response to axn mainly includes the up regulation of different amino acids and the change of unsaturated fatty acids. Combined transcriptomic and metabolomic data indicated that amino acid metabolic pathways were upregulated in the muscles after Axn feeding. For good measure, energy metabolism pathways were upregulated in the muscles to improve ATP and unsaturated fatty acid production. This study provides key information to increase our understanding of the effects of Axn in shrimp.

Growth and energy budget of marine amphipod juvenile Eogammarus possjeticus at different temperatures (20°C, 24°C, 26°C, 28°C, 30°C, 32°C and 34°C) were investigated in this study. The results showed that the cumulative mortality rate increased significantly with rising temperature (p<0.01), and exceeded 50% after 24 h when temperature was above 30°C. With the temperature increasing from 20°C to 26°C, the ingestion rate and absorption rate increased, but decreased significantly above 28°C (p<0.01), indicating a decline in feeding ability at high temperatures. The specific growth rate increased with rising temperature, but decreased significantly (p<0.01) after reaching the maximum value at 24°C. Similarly, the oxygen consumption and ammonia emission rates also showed a trend of first increase and then decrease. However, the O:N ratio decreased first and then increased with rising temperature, indicating that the energy demand of E. possjeticus juvenile transferred from metabolism of carbohydrate and lipid to protein. In the energy distribution of amphipods, the proportion of each energy is different. With rising temperature, the ratio of the energy deposited for growth accounted for ingested gross energy showing a trend of decrease, while the energy lost to respiration, ammonia excretion, and feces accounted for ingested gross energy being showed a trend of increase. It seemed that rising temperature increased the metabolism and energy consumption of the amphipods and, meanwhile, decreased the energy used for growth, which may be an important reason for the slow growth and small body size of the amphipods during the summer high-temperature period.

Penaeus vannamei is one of the largest yield of shrimp aquaculture in the world, which has efficient feed conversion rate, fast growth rate and exhibits high tolerance to poor culture water conditions and with high market demand. However, bottlenecks in culture exist, such as environmental issues caused by culture intensification and quantities of artificial feeds increase, and disease outbreaks occur with increased frequency. We assessed effects of feeding P. vannamei for 35 days with live, artificial and mixed feeds on its feeding activity, growth and energy budget. Our results indicate that growth of mixed feed group to be significantly higher than that of live or artificial feed group (P < 0.05), which the SGRw, SGRl and SGRe of mixed feed group are 2.38 ± 0.15, 0.80 ± 0.13 and 2.43 ± 0.09 (%·d−1), respectively. Survival and growth performance of live feed group are higher than those of artificial feed group (P < 0.05). Feed consumption rate and energy conversion rate of live or mixed feed group are higher than those of artificial feed group (P < 0.05), which feed conversion efficiency of mixed feed group is higher than that of live or artificial feed group (P < 0.05). Diet has a significant effect on energy flow in P. vannamei (P < 0.05). The proportion of energy from consumed feed directed towards growth is the highest in live feed group and the lowest in artificial feed group. With the exception of respiratory energy, the proportion of energy consumed by growth, excretion, molting and feces in mixed feed group was intermediate level of that in the live or artificial feed group. Live feed has better promotion on the survival and growth of P. vannamei. Considering the ecological and economic benefits, the mixed feeds is favorable to the rapid growth of shrimps, which is suitable for the cultivation of this species.

The pore-forming protein perforin is one of the effectors of cell-mediated killing via the granule exocytosis pathway. In this study, a genome-wide association study was conducted in Vibrio harveyi disease-resistant and disease-susceptible families of half smooth tongue sole (Cynoglossus semilaevis) to determine the genes accounting for host resistance, and a perforin homologue was identified, designated perforin-1 like (CsPRF1l). The full-length cDNA of CsPRF1l is 1835 bp, and encodes 514 amino acids. The CsPRF1l gene consists of 10 exons and 9 introns, spanning approximately 7 kb. The amino acid sequence of CsPRF1l shows 60.35, 54.03, 41.92, and 34.17% identities to Morone saxatilis PRF1l, Oryzias melastigma PRF1l, Danio rerio PRF1.5 and Homo sapiens PRF, respectively. Sequence analysis revealed the presence of membrane attack complex/perforin (MACPF) and C2 domains in CsPRF1l. Quantitative real-time PCR showed that CsPRF1l presented a higher intestinal expression level in disease-resistant families than in susceptible families. Tissue expression pattern analysis showed that CsPRF1l is present in most of the tested tissues and highly expressed in the intestine, brain, stomach and gills. After challenge with V. harveyi, CsPRF1l mRNA was markedly upregulated in the liver, spleen, kidney, intestine, gills and skin. In addition, the recombinant CsPRF1l protein exhibited obvious antimicrobial activity against V. harveyi in vitro and in a zebrafish model. Collectively, these data indicate that CsPRF1l modulates host immune defense against V. harveyi invasion and provide clues about the efficacy of rCsPRF1l in fish that will give rise to useful therapeutic applications for V. harveyi infection in C. semilaevis.

The effects of three environmental factors, salinity, carbonate alkalinity, and pH, on the survival, feeding, and respiratory metabolism of Eogammarus possjeticus (Amphipoda: Gammaridae) were investigated experimentally. The results show that E. possjeticus could tolerate a broad salinity range. The 24-h lowest median lethal salinity was 2.70, and the highest was 47.33. The 24-h median lethal alkalinity and pH were 23.05 mmol/L and 9.91, respectively; both values decreased gradually with time. Different values of salinity, carbonate alkalinity, and pH resulted in significant differences in the cumulative mortality (P<0.05). The ingestion rate and feed absorption efficiency were significantly affected by the coupling of the three environmental factors (P<0.05). With increases in carbonate alkalinity, salinity, and pH, both ingestion rate and feed absorption efficiency exhibited a downward trend, indicating a decline in feeding ability under high salinity and more alkaline water conditions. The coupling of salinity, carbonate alkalinity, and pH also had a significant effect on respiration and excretion (P<0.05). The oxygen consumption rate increased first and then decreased with increasing carbonate alkalinity. Under the same carbonate alkalinity values, the oxygen consumption rate increased with increasing salinity. Under the same carbonate alkalinity and salinity, the oxygen consumption rate initially increased and then decreased with increasing pH. The O:N ratio first increased and then decreased with increasing carbonate alkalinity. When carbonate alkalinity was less than 6 mmol/L, the O:N ratio increased with increasing salinity and decreased with increasing pH. The results demonstrate that changes in salinity, carbonate alkalinity, and pH had a measurable impact on the osmotic pressure equilibrium in E. possjeticus and affected the energy supply mode (i.e. ratio of metabolic substrate).

Ulva prolifera plays an important part in most green tides around the world. To explore its possible use, the immunostimulatory effect of orally administered U. prolifera polysaccharides in white shrimp, Litopenaeus vannamei , was investigated in this study. Shrimp were divided into four groups: a group fed with the basal diet as the control group (control), and three treatment groups fed respectively with a basal diet supplemented with polysaccharides of cold water extract (PC), polysaccharides of hot water extract (PH), and the filtered residue after the extraction of polysaccharides (R). The highest final weight was found in the group PC (9.31 ± 0.54 g), followed by R (9.11 ± 0.63 g), PH (8.94 ± 0.61 g), and the control group (8.65 ± 0.53 g); the final weight of shrimp in group PC and group R was significantly greater than that in the control group ( P < 0.05). The minimum feed conversion ratio (FCR) was found in group PC (1.33 ± 0.02), followed by group R (1.40 ± 0.04), PH (1.45 ± 0.04), and the control group (1.54 ± 0.04), and FCR in PC and PH was significant lower than that in control group ( P < 0.05). After the feeding trial, the shrimp were challenged with Vibrio parahaemolyticus . Compared to group PH, R, and the control group, the hepatopancreas lysozyme (LSZ) and phenoloxidase (PO) activities of shrimp in group PC were significantly higher from 24 to 96 h. Most shrimp died in 24–72 h after infection; the shrimp in group PC, PH, R, and the control group at 96 h had cumulative mortality of infection of 17.14, 40.00, 34.29, and 54.29%. There were no significances in cumulative mortality between group PH and R ( P > 0.05); however, they were significantly lower than that in the control group ( P < 0.05) while they were significantly higher than that in group PC ( P < 0.05). The highest immune protective rate was found in group PC, followed by group R and PH. There were no significances in immune protective rate between group PH and R; however, they were significantly lower than that in group PC. These results suggest that dietary U. prolifera polysaccharides can promote shrimp growth and enhance non-specific immune and disease resistance enhancement against V. parahaemolyticus in L. vannamei . The findings from this present study will help to promote the application of U. prolifera in aquaculture.

设置26、28和30℃3个水温处理组,研究温度对凡纳滨对虾工厂化养殖水体中浮游微藻优势种演替的影响.结果表明:(1)28℃处理组虾池水体中的无机氮含量与30℃处理组的差异不显著(P>0.05),3个水温处理组间的活性磷含量差异不显著(P>0.05).(2)养殖前期(5月15日—29日),3个水温处理组虾池水体中浮游微藻的密度呈增大的趋势;5月29日之后,26和28℃处理组的微藻密度相对稳定,而30℃处理组呈上下波动的趋势.26、28和30℃处理组分别检测出浮游微藻3门15种、3门16种和4门17种.(3)试验期间,各水温处理组虾池水体中均未检测出轮虫、枝角类和桡足类等浮游动物.(4)26℃处理组虾池水体中浮游微藻的优势种为绿藻门的普通小球藻、卵形衣藻及硅藻门的牟氏角毛藻、新月菱形藻;普通小球藻始终处于主导地位.28℃处理组的浮游微藻总密度呈升高的趋势;优势种为绿藻门的普通小球藻、卵形衣藻及硅藻门的牟氏角毛藻;普通小球藻始终处于主导地位.30℃处理组的浮游微藻密度呈"M"型趋势;前期优势种较少,以普通小球藻和牟氏角毛藻为主,普通小球藻的优势度最高;中期之后,牟氏角毛藻取代普通小球藻,成为最主要的优势种;养殖中期,铜绿微囊藻也演替为优势种;随着养殖的进行,牟氏角毛藻逐渐成为优势种,优势度逐渐增大,在养殖的后期演替为唯一优势种.(5)26、28和30℃处理组虾池水体中浮游微藻的多样性指数分别为1.20～1.64、1.07～1.69和1.26～1.83;随着养殖的进行,26和28℃处理组的多样性指数呈先增大后降低的趋势,30℃处理组呈先增大后降低再增大的趋势.(6)随着养殖的进行,各处理组水体中的Simpson优势集中性指数均呈先增大后降低的趋势;整个养殖期间,26、28和30℃处理组Simpson优势集中性指数的平均值分别为0.32、0.32和0.28.以上研究结果表明,温度是影响凡纳滨对虾工厂化养殖水体中浮游微藻优势种演替的主要原因.

肌球蛋白重链和肌球蛋白轻链是粗肌丝的重要组成单位,其表达量的高低影响肌纤维的组成和肌肉生长.为了解析肌球蛋白重链(Myosin heavy chain,MHC)和肌球蛋白轻链(Myosin light chain,MLC)基因在脊尾白虾(Exopalaemon carinicauda)生长发育中的作用,采用RACE技术,克隆了脊尾白虾EcMHC2、EcMLC1全长cDNA序列,采用实时荧光定量PCR方法分析了EcMHC2、EcMLC1在脊尾白虾不同组织、不同幼体发育时期及大小差异个体在不同生长时期的表达特征.研究显示,脊尾白虾EcMHC2基因(GenBank:MG545144)全长5 929 bp,开放阅读框5 727 bp,编码1 909个氨基酸;脊尾白虾EcMLC1基因(GenBank:MG545145)全长1 506 bp,开放阅读框459 bp,编码153个氨基酸.脊尾白虾肌球蛋白重链和肌球蛋白轻链在系统进化上与其它十足目甲壳动物的关系较近.脊尾白虾EcMHC2、EcMLC1在各组织中均有表达,但主要在肌肉组织内表达.在脊尾白虾幼体发育的不同时期,EcMHC2、EcMLC1在溞状幼体Ⅱ期的表达量显著高于溞状幼体的其它时期;仔虾第一天,EcMHC2、EcMLC1的表达量显著高于其它时期.在脊尾白虾生长的四个阶段中,大个体组EcMHC2的表达量均显著的高于小个体组;60、80和120日龄时期,大个体组的EcMLC1表达量显著的高于小个体组;在大个体和小个体组中,EcMHC2、EcMLC1在80日龄的表达量最高.脊尾白虾EcMHC2、EcMLC1在大个体组与小个体组的差异表达提示它们在脊尾白虾生长发育过程中发挥重要作用.

为了评估浒苔(Ulva prolifera)对养殖废水的净化效果,本文研究了4个温度水平(22.5℃、25.5℃、28.5℃和31.5℃)和3种氮源(NH4Cl、NaNO2和NaNO3)下,浒苔对工厂化养殖水环境的适应能力及其净化效果.结果 显示,96 h内4个温度处理组浒苔对总氨氮(TAN,包括NH4+-N和NH3)的平均吸收速率分别为14.65、14.88、14.48和13.53 μmol/(g.h),144 h内温度各处理组对亚硝态氮(NO2--N)的平均吸收速率分别为11.28、10.48、9.11和8.38 μmol/(g·h),144h内各温度处理组对硝态氮(NO3--N)的平均吸收速率分别为9.41、8.62、8.80和7.35 μmol/(g·h);温度对浒苔的生长速率有极显著的影响(P＜0.01),而氮源对浒苔的生长速率有显著影响(P＜0.05);在相同氮源条件下,浒苔的生长速率随着温度的升高而逐渐降低;在相同温度条件下,氮源为氨氮(NH4+-)时,浒苔的生长速率大于氮源为NO2--N和NO3--N的生长速率;温度和氮源对浒苔叶绿素a的含量影响不显著(P＜0.05),氮源为NH4+-N和NO1--N时,随着温度的升高,浒苔中叶绿素a的含量均有升高的趋势,而氮源为NO3--N时,浒苔中叶绿素a的含量呈先降低再升高的趋势;温度和氮源对浒苔中类胡萝卜素的含量均有极显著影响(P＜0.01).随着温度的升高,各处理组浒苔中类胡萝卜素的含量均呈升高的趋势,其中,在28.5℃和31.5℃条件下,NO3--N处理组浒苔类胡萝卜素的含量明显高于其他各处理组(P＜0.05).研究表明,温度在22.5℃～31.5℃范围内,浒苔可以有效吸收TAN、NO2--N和NO3--N等对虾工厂化养殖废水中的营养盐,浒苔对NH4+-N的吸收速率最大,但随着水温的升高,浒苔对NHH4+-N、NO2--N和NO3--N的吸收速率均呈降低的趋势.

Microbial community and its management are crucial to the stabilization of culture environment for recirculating aquaculture system (RAS). Although several studies have been carried out for the microbial community of RAS, few studies were on the RAS for shrimp. Water recirculation ratio is an important factor for the microbial community and the management of RAS. Therefore, low (LC), medium (MC) and high (HC) recirculation ratio systems were set to explore the microbial community constitution of RAS for Litopenaeus vannamei and study the effect of water recirculation rate on it. The bacterial community of bioreactor was mainly dominated by Proteobacteria (41.6–70.7%), followed with Planctomycetes (12.5–31.0%), Bacteroidetes (10.5–26.0%), Actinobacteria (1.1–4.8%) and Verrucomicrobia (1.4–6.8%) phylum. The most dominant family of bioreactor was Rhodobacteraceae or Planctomycetaceae. The bacterial community of culture water was simpler than bioreactor and dominated by Proteobacteria (61.8–96.4%). The dominant bacterial groups of bioreactor and culture water are also different among the three water recirculation rates, and the proportions of dominant groups showed a trend with the variety of water recirculation rate. Water quality indexes including ammonia and nitrite decreased with the increasing of water recirculation rate. According to the growth performance of L. vannamei, shrimp had better performance of growth rate and final weight in MC and HC, however, shrimp had higher survival and yield in LC. Shrimp survival and yield had an inverse correlation with water recirculation rate. The results demonstrate the microbial community of RAS for shrimp, highlight the importance of further studies on the function of bacterial taxa, and promote the understanding of the effects of water recirculation rate on the microbiota. The findings suggest that water recirculation rate has important impacts on the microbial community, water quality and shrimp growth. Increasing the water recirculation rate could improve the water quality and promote the growth of shrimp. However, the survival rate and yield of L. vannamei are higher under low water recirculation rate. Recirculation rate is an effective method to manage RAS, and its impact on RAS needs further study, especially in the application of low level of water recirculation.

Ulva prolifera is one of the most common macroalgae and is distributed widely off the coast of China. It is well known for its rapid growth rate and good reproduction and it has fast nitrogen removal efficiency. Integration of seaweed cultivation in shrimp farming water is considered a potential aquaculture practice since seaweeds can convert dissolved inorganic nitrogen into biomass and the biomass can be harvested easily. This study investigated the effect of integrating green seaweed (Ulva prolifera) with Litopenaeus vannamei (500 shrimp m−3) at varying levels of water exchange daily on water quality and shrimp growth performance. The four daily water exchange quantities were 5% (T1), 10% (T2), 15% (T3), and 20% (T4). The appropriate range of the stocking density of U. prolifera was 800 mg L−1. No significant differences were observed in total ammonia nitrogen (TAN) concentration between T2 and T3 (P > 0.05) from beginning to end. The concentrations of nitrite and nitrate in all treatment remained constant at low levels from beginning to the end. On day 35, there were no significant differences in survival rate of shrimp among T2, T3, and T4. No significant differences in FCR were observed in group T2, as compared to T1, T3, or T4. No significant differences in cumulative weight of U. prolifera were observed among T1, T2, and T3; however, they were significantly larger than that of T4. The study demonstrates that integrating U. prolifera (800 mg L−1) with L. vannamei (500 shrimp m−3) with 10% water exchange can control the water quality and enhance shrimp growth.

Numerous studies suggest R-spondins (Rspos) plays a role in mammalian sex development and differentiation by activating WNT signaling pathways. However, Rspos are frequently less reported in teleosts. In this study, a molecular characterization and expression analysis was conducted with a new rspondin member in the Chinese tongue sole, rspondin2-like (rspo2l). The length of rspo2l cDNA is 1251 bp with 732 bp of coding sequence. A qRT-PCR analysis revealed that the transcription of rspo2l was distributed in various tissues, with high transcription levels in the liver, skin, and gills which might indicate a possible role in immunity. We next examined a time-course of transcription levels in four immune tissues (gill, liver, spleen, and kidney) after Vibrio harveyi challenge. It was found that rspo2l was up-regulated in the gills, spleen, and kidney and down-regulated in the liver, and the greatest responses occurred at 24 and 48 h after bacterial challenge. An assessment of β-catenin, the key regulator of the canonical WNT signaling pathway, at different time points in four immune organs revealed that its transcription profile was similar to that of rspo2l after bacterial challenge. The results suggest that tongue sole rspo2l might play a role in immune responses after bacterial challenge, while the potential link with the WNT signaling pathway still requires further investigation. This is the first report about the involvement of rspondins in fish immune responses.

STAT plays important roles in innate immunity during JAK/STAT signaling pathway, and STAT5 is particularly focused due to the existence of duplicated forms in fish and mammal. In Chinese tongue sole, stat5bl was suggested to be a candidate related to Vibrio harveyi resistance based on previous QTL screening. In this study, the full length of stat5bl cDNA was cloned and its expression patterns were analyzed. stat5bl was predominantly expressed in immune tissues, where the highest level was observed in liver, followed by skin and gill. Time course expression patterns were examined in six tissues (liver, skin, gill, kidney, intestine, spleen) after V. harveyi infection. stat5bl could be up-regulated by V. harveyi infection in all tissues except liver, despite the timepoints of peak were different. In contrast, stat5bl was significantly downregulated in liver. To elucidate the role of stat5bl in liver, in vitro RNAi were performed using primary liver cell culture. Knockdown of stat5bl could regulate the expression of genes closely related to JAK/STAT pathway. This study would enlarge our understanding of stat5bl in fish immunity.

In mammals, microtubule-dependent trafficking could participate the immune response, where the motor proteins are suggested to play an important role in this process, while the related study in fish was rare. In this study, dctn5, a subunit of dyactin complex for docking motor protein, was obtained by previous immune QTL screening. The full-length cDNAs of two dctn5 transcript variants were cloned and identified (named dctn5_tv1 and dctn5_tv2, respectively). Tissue distribution showed that dctn5_tv1 was widely distributed and high transcription was observed in immune tissue (skin), while dctn5_tv2 was predominantly detected in gonad and very low in other tissues. Time-course expression analysis revealed that dctn5_tv1 could be up-regulated in gill, intestine, skin, spleen, and kidney after Vibrio harveyi challenge. Moreover, recombinant Dctn5_tv1 exhibited high antimicrobial activity against Escherichia coli and Streptococcus agalactiae due to binding to bacteria cells. Taken together, these data suggest Dctn5_tv1 is involved in immune response of bacterial invasion in Chinese tongue sole.