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This study evaluated the protein requirement of Litopenaeus vannamei post-larvae during the initial rearing phase in a biofloc system. Five different diets were evaluated with increasing concentrations of crude protein: 31.28, 36.29, 41.57, 46.34, and 51.74 g 100 g-1 CP. Post-larvae (0.16 ± 0.01 g) were stocked at a density of 450 PL m-3 in 400 L tanks. Water quality parameters were maintained within the limits recommended for shrimp farming. After 38 days, a regression analysis revealed that levels of CP content (65.29 í 72.83%), EE (10.45 í 11.65%) and body N (10.45 í 11.64%) increased with increasing protein levels in the diet. A similar trend was observed in the biofloc sludge with respect to CP and N. Survival exceeded 80%, and the shrimp with diets containing 31.28 to 46.34g 100 g-1 CP presented an increase in final weight (1.52 í 2.61 g), productivity (0.69 í 1.10 Kg m-3), weight gain (1.38 í 2.44 g), and feeding efficiency (77.28 í 101.68%), whereas these indices decreased to51.74 g 100 g-1 CP. Crude protein content from 44.26 to 47.12 g 100 g-1 provided the best growth performance during the initial rearing phase of Pacific white shrimp in a biofloc system.

Shrimp farming is one profitable sector of aquaculture and the Pacific white shrimp (Litopenaeus vannamei) is the most cultivated species worldwide. Nutrition is considered as one of the greatest challenges in shrimp farming due to its dependence on fishmeal (FM) as an ingredient in the feed. The main goal of the present work was to analyse if the substitution of FM by yellow mealworm (Tenebrio molitor) meal (TM) influences digestive enzyme expression, haemolymph parameters or the intestinal microbiota of shrimp juveniles. Shrimps (average initial weight 4.42±0.02 g) were fed five experimental diets where FM protein was gradually substituted by TM protein (0, 25, 50, 75, and 100%). The shrimps were fed four times per day with an amount equivalent to approximately 6% of their biomass for 42 days. The total haemocyte count, protein concentration and prophenoloxidase activity in the haemolymph were not significantly altered (P>0.05) when FM was substituted by TM. However, the agglutinating activity of shrimp serum determined against dog erythrocytes was higher in the shrimp group fed 25% TM. The absolute and specific activities of trypsin, chymotrypsin, lipase, α-amylase and the patterns of proteolytic activities were not affected by the dietary treatments. The shrimp gut bacterial microbiota profile was similar with the genera Pseudoalteromonas, Rubritalea, Ruegeria, Tenacibaculum, and Vibrio detected. These results show that TM is viable alternative for the replacement of FM, without any deleterious effects on expression of key digestive enzymes, gut microbiota or the immune system of the Pacific white shrimp.

Shrimp farming in biofloc technology system (BFT) has been considered a sustainable alternative to conventional production, reducing environmental impacts due to zero water exchange. Besides increasing productivity, as microorganisms represent additional feed for shrimp, BTF may prevent diseases. White spot disease is the most devastating viral disease of farmed shrimp, causing large socio-economic impacts. Our aim was to compare survival rates and transcription profiles of target genes in different tissues of juvenile specimens of the Pacific white shrimp, Litopenaeus vannamei, reared either in BFT or in clear seawater (CSW), after challenging with white spot syndrome virus (WSSV). Shrimp survival rate was higher among animals kept in CSW, which also presented a lower viral load after 48 h post-infection (p.i.). We could also address possible individual patterns in the sole survivor at 72 h p.i. in biofloc, probably a less vulnerable shrimp to WSSV. There was no significant difference regarding the transcription profile of target genes between systems before WSSV infection; gene transcription was apparently modulated by WSSV infection. Changes in gene profile were more frequently observed in the hepatopancreas. Out of the eleven selected target genes, those coding for Calreticulin, β-tubulin, and Prophenoloxidase showed the most significant upregulation, followed by QM, SOD, and Ubiquitin. Although BFT is considered more biosecure, shrimp kept in this system are not immune to disease, especially against highly virulent pathogens, such as WSSV.

This study evaluated the use of microalgae (Aurantiochytrium sp.) meal as a substitute for fish oil in the diet of juvenile Pacific white shrimp (Litopenaeus vannamei) reared in a clear‐water system. Dietary treatments at five replacement levels (0%, 25%, 50%, 75%, 100%) were performed in triplicate. After 46 days, only a slight difference in shrimp final weight was observed among treatments (0.61 g). An increase in final weight was observed with replacement of up to 50% fish oil for microalgae meal, while the optimal percentage of replacement estimated was 44.7%. Feed conversion rate (FCR) of animals fed a diet of Aurantiochytrium sp.meal to replace up to 50% fish oil decreased, and the optimal percentage of replacement estimated was 49.3%. The fatty acids profile in shrimp muscle tissue demonstrated an increase in docosahexaenoic acid (DHA) from 10.03% to 14.28% with increased replacement of fish oil by microalgae meal in the diet. Therefore, the partial replacement of fish oil by microalgae meal resulted in improved shrimp growth and FCR, and total replacement of dietary fish oil had no negative effects on these parameters. In addition, inclusion of microalgae meal raises the level of DHA in shrimp muscle.

This study aimed to evaluate the effect of different stocking densities of Nile tilapia, Oreochromis niloticus, integrated with Pacific white shrimp, Litopenaeus vannamei, reared in a biofloc system for 57 days. The performance of both species and the ecological efficiency of the system were evaluated. Four levels of tilapia stocking density were evaluated: 0, 8, 16 and 24 fish per tank (90 L useful volume). The initial weight was 4.8 ± 0.1 g and 9.6 ± 0.1 g for shrimp and fish, respectively. The shrimp were fed according to the feed table, and the fish were fed with 1% of fish biomass, stimulating the tilapia to seek food in the biofloc. Results show no difference between average shrimp weight (14.9 ± 0.6 g) and survival (93.0% ± 1.0%). Similarly, fish obtained a final mean weight of 61.9 ± 3.8 g and survival of 91.1 ± 7.9%. Total yield was higher based on the increase in fish density. The sludge produced per animal biomass (sludge:biomass ratio) decreased as fish density increased (y = − 0.0083xx + 0.5995 r2 = 0.87). Nitrogen recovery rose linearly with the increase in fish density (y = 0.036xx + 0.2725, r2 = 0.84), as did phosphorus recovery (y = 0.00711xx + 0.1395, r2 = 0.91). These results demonstrate the feasibility of increasing yield up to 31.2% by integrating L. vannamei and O. niloticus in a biofloc system. Also, the decrease in sludge:biomass ratio and the higher nitrogen and phosphorus recovery increased the sustainability of L. vannamei rearing in a biofloc system.

This work aimed to evaluate growth performance, gut histology, digestive enzyme activity and resistance to challenge with White Spot Syndrome Virus (WSSV) of Litopenaeus vannamei fed 0.5, 2 and 4% of dry biomass from the brown seaweeds Sargassum filipendula and Undaria pinnatifida. Shrimp (4.23 ± 0.15 g) were reared for 38 days in 800 L tanks (40 animals per tank) in triplicate under constant aeration (O2 > 5 mg L−1), controlled temperature (29.06 ± 0.32 °C) and 100% daily water exchange (static). Animals were fed five times per day, and the amount of feed was calculated in accordance with a pre-established feed conversion rate method. To estimate survival and growth, all animals from each tank were weighed and counted every week. At the end of the experiment, the hepatopancreas from each of six shrimp per treatment was collected to assess the digestive enzymes, and the anterior abdominal segment from each of four shrimp per treatment was isolated to evaluate the gut histology. For WSSV challenge, 40 animals per treatment were infected and kept for 72 h to determine mortality. Growth performance was not affected by seaweed addition in the diet. However, the lowest level of both seaweeds increased shrimp gut epithelial surface for nutrient absorption, and all levels of S. filipendula and 4% of U. pinnatifida increased amylase activity, while lipase activity was only stimulated by 4% of U. pinnatifida. Proteolytic enzymes were not affected by the addition of dietary seaweed. Additionally, 4% of U. pinnatifida resulted in lower mortality after WSSV challenge (48%) compared to control (74%). Therefore, seaweed addition improved shrimp digestive capacity and resistance to WSSV challenge, but without impairing growth performance.

Pacific white shrimp, Litopenaeus vannamei, is a tropical species known to be intolerant to cold. In our earlier work, increased thermal shock resistance caused by Sargassum filipendula raised unanswered questions about the mechanisms responsible for this resistance. Therefore, in the present work, we used MALDI-TOF mass spectrometry (MS) to assess protein and lipid profiles of intact and lysed hemocytes from Pacific white shrimp fed 0.5% of S. filipendula and submitted to thermal shock. To accomplish this, shrimp (13.0 ± 0.4 g) were reared during 15 days in 800 L tanks (35 shrimps tank− 1), in duplicate, under constant aeration and temperature (28.5 °C), 100% of daily water exchange and feeding four times day− 1. At day 16, animals were submitted to thermal shock by passing from seawater at 28.5 °C to 11.5 °C, where they remained for 1 h, and then returning to 28.5 °C. Mortality was monitored during 24 h. Hemolymph of 10 animals per treatment (control and 0.5% S. filipendula) was collected before and 15 min after thermal shock for MALDI-TOF MS analyses. Hemocytes were processed in both intact and lysed cells. Samples were mixed (1:1) with 2,5-dihydroxy-benzoic acid matrix for lipid analysis and sinapinic acid for protein. Shrimp fed 0.5% S. filipendula showed higher signal intensity of phosphatidylcholines with unsaturated fatty acids and lower signal intensity of lysophosphatidylcholines, after thermal shock. For proteins, S. filipendula increased signals that could be associated with antimicrobial peptides and histone H2A, whereas reduced signals were related to Cytochrome c oxidase subunits. Based on these results, it can be concluded that S. filipendula increased membrane fluidity and antimicrobial defense, diminished apoptotic signaling, and modulated DNA regulation, lipid and energy metabolism of shrimp hemocytes. These mechanisms could explain, at least in part, the higher thermal shock resistance of these shrimp fed a diet with 0.5% S. filipendula.

This study investigated the muscle quality of the shrimp Litopenaeus vannamei fed on a diet containing different proportions of mealworm meal (MW) (0, 25, 50, 75 and 100%) as a substitute for fishmeal, which is the normal diet used in shrimp commercial production. The proximate composition, fatty acid profile, colour and texture of the shrimps were evaluated. Moisture, protein, and ash content of shrimp muscle were not significantly altered when fishmeal was replaced by MW (p>0.05). However, the replacement resulted in a linear increase in lipid content (p

A 13-day nursery trial was conducted to evaluate the performance of young Litopenaeus vannamei post-larvae (from PL 6 to PL 18 ) reared in both biofloc and microalgae-based systems at a stocking density of 67 PLs L −1 . The effects of different concentrations of total suspended solids (TSS) on PL performance were also evaluated. One experimental group was reared in a conventional microalgae-based system with daily water exchange and daily addition of microalgae (herein called microalgae treatment). The other two experimental groups were reared using biofloc technology (BFT) with daily dextrose addition and no water exchange, but in the “Biofloc-500” treatment, TSS were maintained at around 500 mg L −1 , while in the “Biofloc-700” treatment, TSS were maintained at around 700 mg L −1 . Water quality variables remained within the appropriate range for larval culture. In microalgae treatment, ammonia control was likely associated with its assimilation into microalgae biomass and daily water exchange. In biofloc tanks, however, the addition of dextrose stimulated the production of bacterial biomass from ammonia. This system required only 12.9% of the water used by the microalgae treatment since water was not exchanged during the culture. The nursery of young PLs resulted in similar ( P > 0.05) performance in all treatments: survival >94%, PL length ∼ 11.5 mm, and PL dry weight ∼ 1.2 mg. In addition, the salinity stress test (>90.0%) was not significantly different among treatments. Our results indicate that BFT can be as effective as the microalgae-based system for the nursery of young L. vannamei post-larvae. We also found that post-larvae performance was similar ( P > 0.05) between biofloc treatments, indicating that organisms can tolerate environments with large quantities of solids.

This study evaluated the performance, immunology, and survival of the Pacific white shrimp Litopenaeus vannamei to experimental challenge to Vibrio alginolyticus based on the use of the probiotic Lactobacillus plantarum and the combined use of probiotic and butyrate. Four different diets resulted from the addition of additives: butyrate, probiotic, butyrate + probiotic, and control (no additives). The attractiveness of the diets was assessed by the percentage of positive choices and rejections, using a dual-choice Y-maze format aquarium. The shrimps were fed during four weeks and performance parameters, intestinal microbiota, and immunological parameters were all evaluated. Subsequently, the shrimps were challenged with V. alginolyticus and after 48 h, survival and immunological parameters were evaluated. The results showed increased attractiveness and intake, but only with diets supplemented with sodium butyrate. However, other diets were not rejected. No difference in performance or immunological parameters was observed among the different diets. Also, among the treatments, no difference in Vibrio spp., or total heterotrophic bacteria counts, was found in the intestinal tract. However, the lactic acid bacteria count was higher in the intestinal tract of shrimps fed diets supplemented with probiotic. After bacterial challenge, shrimp fed all diets had a greater survival when compared with the control group. Lactobacillus plantarum and sodium butyrate increase the resistance of shrimp to infection with V. alginolyticus, but do so without affecting performance, immunological parameters, or Vibrio spp., and total heterotrophic bacteria counts in the intestinal tract.

Mealworm meal (MM) was evaluated for its potential as a future protein source for the farmed shrimp Litopenaeus vannamei . The apparent digestibility coefficients (ADC) were determined using a reference diet and a test diet that contained 85% reference diet and 15% MM, both contained 0.5% chromic oxide as an inert marker. Subsequently, considering the digestible values of MM, after six weeks of culture under clear water using five diets containing different levels of fishmeal replaced by MM (0%, 25%, 50%, 75% and 100%), the growth parameters and the whole shrimp body composition were evaluated. The values of the ADC were: 45.9% for dry matter, 66.5% for energy and 76.1% for crude protein while the ADC for essential amino acids ranged from 72% to 86%. Methionine was the first limiting amino acid in MM. Weight gain, specific growth rate, feed intake, feed conversion, survival and protein retention were not affected when fishmeal was replaced by MM (P > 0.05). The protein content of the shrimp body showed no significant differences (P > 0.05) between the treatments. However, lipid content of the shrimp body increased from 1.13% to 1.88% when fishmeal was replaced by MM. These results suggest that mealworm meal can be utilized as an alternative protein source for L. vannamei juveniles, although methionine should be added as a supplement.

This study evaluated the midgut microbiology, hemato-immunological parameters and the resistance to thermal shock of the white-leg shrimp Litopenaeus vannamei fed with diets containing 0.5, 2, and 4% of the brown seaweeds Undaria pinnatifida and Sargassum filipendula. Shrimp were reared in clear seawater for 15 days in seven 800-L tanks with 100 shrimp (5.9 ± 0.2 g), under constant aeration and temperature (29 °C), with 80% daily water exchange. Animals were fed four times per day (6% of biomass) during the experimental period. At day 16, bacterial count of midgut and hemato-immunological parameters were assessed before the thermal shock; during thermal shock, shrimp were transferred from seawater at 29 to 12.5 °C for 1 h, and then back to 29 °C, and kept for 48 h to estimate the cumulative mortality. Animals fed with 4% of U. pinnatifida had lower Vibrio spp. quantity in the midgut and showed an overall improvement of evaluated hemato-immunological parameters; however, only phenoloxidase activity was statistically significant. Those fed with 0.5 and 2% of S. filipendula showed lower cumulative mortality after thermal shock, in comparison with control and those fed with 4% of this species. Increasing levels of U. pinnatifida had negative effects on thermal shock resistance. In conclusion, U. pinnatifida enhances shrimp immunity and specifically inhibit Vibrio spp., while S. filipendula increases shrimp resistance to thermal variation.

Higher rates of production per unit area, avoiding or minimizing significant environmental damages, is a challenge for aquaculture to contribute with an ascendant demand for food due to the increase in world population. The present study assessed the hatchery performance of Litopenaeus vannamei between the mysis 1 and postlarva 5 stages, in a zero-exchange biofloc system under 12.5:1 fixed C:N ratio with dextrose in four stocking densities: 200, 250, 300 and 350 larvae per liter (D200, D250, D300, D350 respectively). Water quality and performance parameters were compared among treatments. The mean values of the evaluated water quality parameters were appropriate for this production stage in all treatments. Fertilization with dextrose efficiently controlled ammonia levels not reaching the average concentrations considered toxic for the specie. Lower values of pH and higher values of volatile solids were finding in D350. There was no difference between groups in means of survival and dry weight of postlarva 5. Therefore, the use of biofloc systems without water exchange with dextrose as a carbon source in 12.5:1C:N ratio resulting in adequate production indexes and water quality during the mysis 1 to postlarva 5 hatchery phase of L. vannamei . Based in these results the density limit to a viable BFT hatchery was not reached.

This study evaluated dietary supplementation with sodium butyrate and polyhydroxybutyrate (PHB) on the enzymatic activity, in vitro nutrient digestibility, and intestinal morphology of Pacific white shrimp (Litopenaeus vannamei). The treatments were: shrimp fed diet supplemented with 2% butyrate, shrimp fed diet supplemented with 2% PHB, and shrimp fed unsupplemented diet. Shrimp fed with PHB-supplemented diet showed higher values of intestinal protease, trypsin, chymotrypsin, and amylase. Shrimp fed with butyrate-supplemented diet showed higher intestinal lipase than unsupplemented shrimp. Butyrate increased hepatopancreatic protease, trypsin, and chymotrypsin activity and in vitro protein digestibility. Shrimp fed with PHB-supplemented diet and butyrate had higher digestibility of polysaccharides and lipids. Shrimp fed with PHB-supplemented diet presented increase in the length, width, and perimeter of intestinal villi. Animals fed with both diets retained overall integrity of their intestinal m...

Este trabalho objetivou avaliar viabilidade da suplementação de duas dietas ao camarão Litopenaeus vannamei com Lactobacillus plantarum. Uma usando farinha de peixe como fonte proteica e outra usando concentrado de proteico de soja, bem como o efeito destas formulações na microbiota intestinal de camarão. Para testar a viabilidade probiótica nas dietas, o número de UFC g-1 foi observado semanalmente ao longo de quatro semanas. A viabilidade de Lactobacillus plantarum em relação aos parí­¢metros fí­­sicos das dietas, incluindo estabilidade, flutuabilidade e expansão, foram testados. O efeito das dietas na microbiota e na morfologia do trato intestinal foi determinado por meio de um experimento fatorial 2x2 (duas dietas, com ou sem suplementação), em triplicata, totalizando 12 unidades experimentais, com cinco animais por unidade, alimentados com 3,5% de biomassa por 17 dias. A concentração de bactérias do ácido láctico diminuiu ao longo do tempo, independentemente da fonte de proteí­­na. A dieta com farinha de peixe como fonte proteica, independente da suplementação probiótica, apresentou boa estabilidade e não se desintegrou após quatro horas. Em contrapartida, dieta com concentrado proteico de soja, suplementado ou não, desintegrou-se entre 2,5 e 3 horas, apresentando baixa estabilidade. Todas as dietas apresentaram flutuabilidade de 0%. A taxa de expansão foi maior nas dietas com concentrado proteico de soja, mas sem influência da suplementação probiótica ou interação entre os fatores. No ensaio in vivo, ambas as dietas suplementadas apresentaram maior contagem de bactérias heterotróficas totais; no entanto, nenhuma diferença na contagem foi observada em dietas com diferentes fontes de proteí­­na. Bactérias ácido-lácticas só foram observadas em dietas suplementadas com probiótico. A histologia do trato intestinal mostrou que todos intestinos tinham células intactas, bem desenvolvidas e organizadas, independentemente da dieta. Assim, L. plantarum, quando combinado com diferentes fontes proteicas, produziu efeitos similares na estrutura e microbiota do camarão marinho Litopenaeus vannamei.

The aim of this study was to investigate the effect on fish health indicators of different Nile tilapia (Oreochromis niloticus) stocking densities when reared in an integrated culture with Pacific white shrimp (Litopenaeus vannamei) using biofloc technology for 57 days. Three fish densities were evaluated: 8, 16 and 24 fish tank−1 with four replicates each. The experimental units comprised 800 L (useful volume) tanks for shrimp rearing and 90 L (useful volume) tanks for the culture of tilapia, which had an initial mean weight of 9.64 ± 0.14 g. A submerged pump located inside the shrimp tanks recirculated the water between the two units. The system was managed as a biofloc technology unit, with the addition of organic carbon to control ammonia and the monitoring of water quality, which remained within suitable ranges for both species. On day 57, fish were sampled for the assessment of the hemato-immunological variables. Fish reared under the highest stocking density exhibited significantly higher values of hematocrit and total leukocytes when compared to the two lowest densities (p

This work aimed to evaluate the effects of integrated culture of sea lettuce (Ulva fasciata) on the shrimp (Litopenaeus vannamei) and mullet (Mugil liza) rearing in a biofloc technology (BFT), on water quality, animal performance and seaweed growth, nitrogen and phosphorus recovery, seaweed bioactive compounds. Two groups were compared: i) Ulva group: cultivation system integrating shrimp and mullet with seaweed, ii) control group: the same without seaweed, both groups used BFT. Each group had four replicates; initial biomass was 618 ± 8 g and 246 ± 34 g for the shrimp and mullet, respectively. Additionally, in the Ulva group, 120 g of sea lettuce per tank was included every two weeks. After 55 days, the integration of seaweed in the system did not influence the shrimp and mullet performance. Shrimps grew, on average, 1.22 g week−1, with 90.6% survival, 2.91 kg m−3 yield and 1.84 FCR. Mullets grew, on average, 1.035 g week−1, with 99.17% survival, and 4.08 kg m−3 yield. At the end 317.01 ± 32.02 g of seaweed per experimental unit was harvested. The total final biomass and yield, considering the whole system, were 12.2% higher in the Ulva group than in the control group. Nitrogen and phosphorus recovery were respectively 5.5% and 7.6% higher in the Ulva group than in the control group. The nitrogen, phosphorus, chlorophyll-a and carotenoids contents in the Ulva group were also higher at the end of the experiment. In conclusion, the integration of sea lettuce can increase the total yield, nitrogen and phosphorus recovery in the biofloc system. Furthermore, the integration of sea lettuce in this system can increase the content of chlorophyll-a and carotenoids, without affecting the yield of ulvan extract.

This study aimed to evaluate the effect of different carbon sources, at two fertilization doses, on water quality and shrimp performance in a biofloc system. To accomplish this, marine shrimp Litopenaeus vannamei post-larvae (PL17) were cultured in intensive (150 animals m−2) zero water-exchange biofloc technology (BFT). The experiment was conducted during 39 days, and 12 tanks (0.85 m3) were used. Treatments consisted of two sources of organic carbon, rice bran (RB) or molasses (MO); each supplied at two rates (25 mg L−1 and 100 mg L−1 of total suspended solids). At the end of the fertilization period, water column respiration rate was higher, but net ecosystem production was lower in RB100 and MO100 treatments. These results indicated that the different carbon sources evoked similar metabolic behavior among microbial communities. Ammonia level was significantly higher in the MO100 treatment after the preparation period (7 days). No significant differences were observed in water quality parameters between treatments at the end of the experiment, except for orthophosphate. RB yield was 22% higher and the feed conversion rate 15% lower compared to MO. Therefore, RB could be used as a carbon source for biofloc systems, increasing shrimp yield, while decreasing feed conversion rate.

Este estudo teve por objetivo avaliar o uso suplementar de probiótico (Lactobacillus plantarum) e butirato de sódio, e a interação deles na dieta de pósí larvas de camarão-branco-do-pací­­fico (Litopenaeus vannamei) em fase de pré-berçário. Durante o experimento foram aplicados nas dietas os tratamentos: (1) probiótico 1x108 UFC/g, (2) butirato de sódio 2%, (3) probiótico 1x108 UFC/g + butirato de sódio 2%, (4) controle (dieta base). Todos os tratamentos tiveram a adição de meio de cultura na mesma dosagem utilizada nos tratamentos com probiótico. Foram utilizadas 16 unidades experimentais de 60 L cada uma, com fundo em formato U, povoadas com 2.880 pós-larvas 5. Após quinze dias, independentemente da presença ou não do probiótico, os camarões dos tratamentos com butirato apresentaram maior sobrevivência (p = 0,0039) e menor peso seco individual (p = 0,0043). Não foram observadas alterações morfológicas no intestino das pós-larvas em nenhum dos tratamentos. Portanto, a suplementação com butirato de sódio na ração aumenta a sobrevivência das pós-larvas de camarões na fase de pré-berçário sem causar alteração morfológica em seu intestino.

This study aimed to evaluate the effects of dietary supplementation with sodium butyrate or polyhydroxybutyrate (PHB) on growth performance, as well as changes in intestinal microbiota and hemato‐immunological parameters, of Litopenaeus vannamei reared under a superintensive biofloc system. Twelve 800‐L tanks were each stocked with 250 shrimp/m³ (3.96 ± 0.04 g mean initial weight) and reared over a 6‐wk period. The basal diet and two test diets supplemented with 2% of each feed additive. At the conclusion of the growth trial shrimp fed with the butyrate‐supplemented diet, as compared with the control shrimp, showed higher survival and productivity and lower total bacterial and Thiosulfate‐citrate‐bile salts‐sucrose Agar (TCBS) counts in the intestine. However, no differences were observed in other performance parameters analyzed. Shrimp fed with both supplementation regimens also showed an increase in total and granular hemocytes, as well as an increase in serum agglutination titer. Shrimp offered diets supplemented with sodium butyrate had higher counts of hyaline cells. Thus, for L. vannamei reared in a superintensive biofloc system, it can be concluded that dietary supplementation of sodium butyrate, more so than PHB, acted as an immune system modulator by reducing the concentration of pathogenic bacteria in shrimp gut, thereby increasing survival and productivity.

Bacterial inorganic nitrogen control using carbon:nitrogen ratio (C:N) manipulation is a tool for aquaculture systems. The present study assessed the hatchery performance of Litopenaeus vannamei between the mysis 1 and postlarvae 5 stages in a zero-exchange biofloc system under different C:N fertilization levels (10:1, 12.5:1 and 15:1) with dextrose. Water quality, performance parameters and water microbiology were compared among treatments. The mean values of the evaluated water quality parameters were appropriate for this production stage. Fertilization with dextrose efficiently controlled ammonia levels, which did not reach the average concentrations considered toxic for the species. In the 10:1 C:N ratio treatment, the levels of ammonia started to increase early and showed significantly higher levels from the third to the last experimental day. There was no difference among groups in means of survival (>76%) and dry weight (0.26 mg) of L. vannamei production parameters and water quality were maintained without water exchange using a biofloc system supplemented with dextrose. Therefore, the use of biofloc systems without water exchange with dextrose as a carbon source in C:N ratios of 10:1, 12.5:1 and 15:1 results in both adequate production indexes and water quality during the misis 1 to post-larvae 5 hatchery phases of L. vannamei . However, the ratios of 12.5:1 and 15:1 keep lower levels of ammonia.

The expansion of shrimp farming is limited by dependency on fish meal and fish oil as the main sources of protein and lipids. Alternative sources of protein could decrease production costs, making shrimp farming more sustainable. Therefore, this paper aimed to evaluate the growth performance of Litopenaeus vannamei (formerly Penaeus vannamei), also known as Pacific white shrimp, co-fed with the green seaweed Ulva lactuca in different percentages (25, 50, 75, and 100 %) of substitution for commercial feed. Shrimps were cultivated in 30-L plastic boxes under constant aeration, temperature of 26 ± 2 °C, 35‰ salinity, and daily water exchange of 80 % for 28 days. Ten shrimp (3.7 ± 0.2 g) were stocked in each experimental unit and fed according to the treatments. Shrimp fed with no algal substitution were used as control. All treatments and control were performed in triplicate. Algal biomass was kept available for shrimp for 24 h, and commercial feed, calculated as 5 % of mean starting shrimp biomass in each unit, was given three times a day. Results indicate that commercial feed can be replaced by as much as 50 % without significant loss on the growth performance of the marine shrimp L. vannamei when cultivated in a clear water system.

Soybean molasses was evaluated as a partial replacement for sugarcane molasses as a carbon source for biofloc development in the superintensive culture of Pacific white shrimp (Litopenaeus vannamei). A 50-day study was conducted with juvenile (3.2 g) shrimp stocked in 16 800 L tanks at a stocking density of 250 shrimp m−3. Control of total ammonia concentration was performed by the addition of combined mixtures of soybean and sugarcane molasses to the culture water. Three different molasses treatments were evaluated using different soybean-to-sugarcane molasses ratios: 15–85%, 38–62% and 60–40% respectively. The control group was treated only with sugarcane molasses. Water quality, chlorophyll a concentration, heterotrophic bacterial load, Vibrio spp. concentration and zootechnical indexes were all evaluated. Total ammonia concentration was controlled by heterotrophic and chemotrophic pathways. Biofloc formation, as quantified by measuring the total suspended solids, was not altered. The Vibrio spp. concentration showed a significant reduction in treatments with soybean-to-sugarcane molasses ratios of 38–62% and 60–40%. All combined mixtures of soybean and sugarcane molasses could maintain water quality and productivity in the superintensive culture of L. vannamei using the biofloc system. Thus, the potential use of a residue from agroindustry as a carbon source in a biofloc culture is demonstrated.

The objective of this study was to isolate and identify bacteria of the genus Bacillus sp. from a super-intensive culture system with Litopenaeus vannamei microbial flakes, and evaluate their potential of affecting the water quality, performance and immunological parameters when added to the water and diet. Bacillus spp. isolates were evaluated in vitro for the ability to inhibit pathogens, antagonistic activity. After selection, Bacillus spp. was added to the water to assess the influence in the formation and maintenance of microbial flakes in post-larvae culture. In order to verify the performance and hemato-immunological parameters of L. vannamei for 42 days of culture, they were incorporated in formulated diets. Four bacteria were isolated from the microbial flakes super-intensive system: Bacillus thuringiensis CPQBA 571-12 DRM 06 (> 99%), Bacillus licheniformis CPQBA 571-12 DRM 07 (> 99%), B. licheniformis CPQBA 571-12 DRM 08 (> 99%) and Bacillus cereus CPQBA 571-12 DRM 09 (> 99%). B. licheniformis CPQBA 571-12 DRM 07 was the only isolate that showed in vitro inhibitory characteristics against the pathogen Vibrio alginolyticus (BCCM 2068) with a 20 mm diameter inhibition zone and the isolates did not show any antagonistic activity. For the test of formation and maintenance of microbial aggregates generated in the post-larvae culture super-intensive system, a significant difference was observed (p Statement of relevance Bacillus spp. were isolated from biofloc system water and can be supplemented in shrimp (L. vannamei) feed to increase immunocompetence and used as biocontrol for Vibrio spp. in water.

Protein hydrolysates from poultry by-product (CPH) and poultry by-product plus swine liver (PHPPL) were evaluated for digestibility and attractiveness for Litopenaeus vannamei, their effect on growth as a substitute for salmon by-product meal. Initially, the apparent digestibility coefficient (ADC) of the protein hydrolysates was determined and their attractiveness was also evaluated. Subsequently, five diets with 0, 25, 50, 75 and 100 % substitution of the salmon by-product meal protein by PHPPL, were evaluated. The experiment was conducted for 42 days in a clear water system. For the digestibility assay, the protein hydrolysates had high ADC for protein, dry matter, energy, and essential amino acids for L. vannamei. However, PHPPL had better digestibility results than CPH, so it was used in the growth assay. No difference in the attractiveness of the tested ingredients was observed (p ≥ 0.05). In the growth assay, survival was no different (p ≥ 0.05) among shrimp fed the different protein replacement levels. However, we observed an increase in shrimp growth with the treatment of 25 % protein replacement. Diet with 50 % protein replacement promoted growth similar to that of the control diet with a subsequent decline of up to 100 % of protein replacement. Therefore, it can be concluded that up to 25 % of dietary protein replacement with PHPPL can improve shrimp growth.

This study aimed to evaluate the relationship between salinity in the performance of marine shrimp Litopenaeus vannamei and halophyte plant Sarcocornia ambigua in an aquaponic system with biofloc. The experiment was conducted for 57 days, and four treatments were evaluated: 8 psu (practical salinity unity), 16 psu, 24 psu, and 32 psu, with three replicates. Each experimental unit consisted of an 800 L tank for shrimp rearing, a 40 L settling chamber and a hydroponic bench of 0.3 m2 of planting area and density of 40 plants m−2. The tank water was continuously pumped to the settling chamber, and the supernatant was distributed on the hydroponic bench to irrigate the plants, returning to the tank by gravity. The tanks were stocked with 300 shrimp m−3 (1.6 ± 0.1 g). The shrimp were fed four times daily with a commercial diet containing 38% crude protein. Salinity affected shrimp survival, which was lower in 8 psu treatment (56.3 ± 4.7%). No salinity relationships were detected with any of the plant performance parameters; however, the highest biomass was produced at 16 psu, which is close to the isosmotic point of S. ambigua. The lowest concentrations of ammonia and nitrite and the highest concentrations of nitrate were found, through the interpolation of the data, near the 18 psu salinity. So, it is suggested that in this salinity, the absorption of ammonia by the plants is favored. The salinity also affected the concentration of dissolved orthophosphate. There was no relationship between salinity and the production of phenolic compounds and antioxidant activity in plants. The integrated cultivation of L. vannamei with S. ambigua can be carried out between salinities of 16 and 24 psu since the performance of the shrimp is not impaired, and the growth of the plants and the removal of nitrogen and phosphate compounds are favored in this range of salinity.

In biofloc technology (BFT) rearing systems, nitrogen compounds, specially ammonia and nitrite, have to be controlled by microbial pathways, mainly through the activity of heterotrophic and chemoautotrophic bacteria. The objective of this work was to assess different water preparation strategies (heterotrophic, chemoautotrophic and mature) in BFT system for nursery of Pacific white shrimp (Litopenaeus vannamei). A 35-day study was conducted with post-larvae shrimp (0.08 g) stocked in twelve 300 L tanks at a stocking density of 2000 shrimp m−3. The water preparation strategies for shrimp rearing that were evaluated in this study included: i) Heterotrophic treatment, where the water received sugar as a carbon source; ii) Chemoautotroph treatment, where ammonium and nitrite salts were added to the water; and iii) Mature treatment, which was created by the addition of a significant amount of water containing mature biofloc from another established BFT system. In both mature and chemoautotrophic treatments, the nitrification process was able to keep toxic nitrogen compounds (ammonia and nitrite) at low levels without the addition of carbohydrates. In contrast, heterotrophic system showed peaks of ammonia and nitrite during the rearing cycle, and the level of these compounds were found to be higher in this treatment (relative to the mature and chemoautotrophic treatments). The chemoautotrophic system exhibited a lower abundance of bacteria from the family Vibrionaceae in the beginning of the experiment compared to the heterotrophic and mature treatments. The combination of low Vibrionaceae abundance and good water quality resulted in improved growth performance in this treatment. These findings demonstrate the importance of manipulating the environment of BFT systems to induce an enrichment of nitrifying bacteria before stocking shrimp. We have also found that the addition of a carbon source to BFT systems is necessary only in emergency situations, when ammonia spikes need to be controlled.

This study evaluated the effects of heterotrophic and mature biofloc systems on yield, water quality, sludge production, water bacterial community, recovery of nutrients and fish health in Litopenaeus vannamei and Oreochromis niloticus integrated culture through a 53-day experiment. Two treatments were evaluated: an organic carbon (C) supplemented system (heterotrophic treatment), in which organic C was supplemented throughout the whole experiment and no biofloc inoculum was added at the beginning, and a mature treatment in which half of the volume of shrimp units was filled with biofloc-rich water from a previous rearing cycle before the culture started. Shrimp (3.34 ± 0.03 g) and fish (5.73 ± 0.06 g) were stocked at 300 shrimp m−3 and 344 fish m−3. The former were fed according to a feeding table, while the latter were fed at the rate of 1% of their biomass to stimulate the ingestion of bioflocs. Shrimp growth performance was unaffected (p > .05), but fish reared in the heterotrophic treatment exhibited better values for all growth performance variables evaluated (p

Litopenaeus vannamei shrimp were reared in a bioflocs system and fed different levels of soybean protein concentrate as a replacement for fishmeal, and both immunological parameters of this marine shrimp and its resistance to Vibrio sp. infection (CPQBA 378-12 DRM01) were evaluated. Four different diets were formulated with 0, 33, 66 and 100 % of soybean protein concentrate as a substitute for fishmeal. Shrimp were reared in a biofloc system in twelve 800 L tanks (250 shrimp m-3) maintained at constant aeration and temperature. After 42 days, 36 animals (14.21 ± 0.89 g) per treatment were challenged with Vibrio sp. (1 x 105 CFU mL-1 - LD10), and hemolymph was collected before and after challenge to perform immunological assays (agglutination titer, concentration of protein and phenoloxidase activity). Shrimp fed with the experimental diets showed no difference in their resistance to infection and haemato-immunological parameters. Thus, rearing L. vannamei in a biofloc system on diets containing either partial or total replacement of fishmeal for soybean protein concentrate did not affect either immunocompetence or susceptibility to infection. RESUMO. O presente trabalho avaliou os parâmetros imunológicos e a resistência contra o Vibrio sp. (CPQBA 378-12 DRM01) de camarões marinhos Litopenaeus vannamei alimentados com diferentes níveis de concentrado proteico de soja em substituição à farinha de peixe e cultivados em sistema de bioflocos. Foram formuladas quatro dietas com 0, 33, 66 e 100 % de concentrado proteico de soja em substituição a farinha de peixe. Os camarões foram cultivados em 12 tanques de 800 L com 250 camarões m-3, sob temperatura e aeração constantes. Após 42 dias, 36 animais (14,21 ± 0,89 g) por tratamento foram desafiados com Vibrio sp. (1 x 105 UFC mL-1 - DL10). Antes e após o desafio foi coletada hemolinfa para avaliação dos parâmetros imunológicos (título aglutinante, concentração de proteína no soro e atividade da fenoloxidase). Não houve diferença na resistência ao desafio e nos parâmetros imunológicos entre os tratamentos. Portanto, o cultivo de L. vannamei em sistema de bioflocos utilizando dietas com substituição total ou parcial com concentrado proteico de soja não afeta sua imunocompetência e susceptibilidade a infecção bacteriana.

We assessed the hatchery performance of Litopenaeus vannamei between the mysis1 and postlarva 5 stages, in a zero-exchange biofloc system. Two sources of organic carbon (molasses and dextrose) were evaluated and water quality, zootechnical parameters, microbiology, and water consumption during production were compared between carbon-supplemented and control groups. The mean values of the evaluated water quality parameters were appropriate for this production stage. Fertilization with molasses and dextrose efficiently controlled ammonia levels and ammonia did not reach the average concentrations that are considered toxic for the species (total ammonia −1 and free ammonia −1 ). The number of heterotrophic bacteria in the water was greater in the molasses and dextrose groups than in the control group. However, there was no difference in Vibrionaceae count between groups. There was no difference between groups in survival (>85%), length (6.15 mm), dry weight (0.17 mg) of postlarvae 5. Treatment with dextrose or molasses required approximately 12% of the water used by the control group. L. vannamei production rates and water quality were maintained without water exchange using a biofloc system supplemented with dextrose or molasses.

This study aimed to assess the final body weight, weekly weight gain, yield, apparent feed efficiency, protein efficiency ratio, and feed intake of the Pacific white shrimp (Litopenaeus vannamei) fed four diets containing different levels of soy protein concentrate (SPC) as a replacement for fishmeal, reared in a super-intensive biofloc system. Diets consisted of replacing 209 g kg-1 fishmeal at 0, 33, 66, and 100% SPC. Shrimp were raised in a biofloc system using twelve experimental units stocked with 250 shrimp m-3 under constant aeration (O2>5mg L-1) and temperature (29±0.5 °C). No significant differences among treatments were observed based on water quality parameters. Shrimp fed diets with 0 and 33% substitution exhibited the highest weekly growth (1.88 and 1.79 g per week) and final weights (15.2 and 14.7 g) compared with shrimp fed the 66 and 100% replacement. A lower feed intake was observed for shrimp fed the 33% SPC diet (3.18 kg per experimental unit) compared with 0% replacement (3.62 kg). Shrimp fed the 33% replacement achieved a similar performance and lower feed intake than animals fed diet without replacement.

The objective of this study was to evaluate the influence of different dietary lipid and fatty acids on the nutritional value of bioflocs used as a feed, as well as shrimp performance and health. A total of 1800 Litopenaeus vannamei juveniles (2.87 ± 0.01 g) were cultured in biofloc technology, with a density of 200 shrimp m−2, and fed with three isoproteic experimental diets at different lipid levels (85 g kg−1, 95 g kg−1 and 105 g kg−1); each treatment was performed in triplicate. After 61 days, no significant difference was observed (P > 0.05) among the water quality parameters. For the shrimp performance, significant difference was observed (P = 0.011) among the values of survival, where treatments with lower lipid levels had higher survival (92.5 ± 3.5% and 91.0 ± 2.5%). Although there are significant differences in survival, no significant differences in the total haemocytes count (THC) were observed. For other growth performance, no differences were observed (P > 0.05). A positive correlation (r = 0.75) has been observed between the dietary oleic acid and bioflocs. The bioflocs showed ‘long-chain’ polyunsaturated fat acids (lcPUFA), especially arachidonic acid. The shrimp showed similar growth and stayed healthy at the end of the experimental period.

The aims of this research were to evaluate the zootechnical performance and dietary cost for the marine shrimp ( Litopenaeus vannamei ) cultured in a superintensive biofloc system and a conventional semi-intensive system using diets containing different protein levels. Four different diets, with crude protein (CP) contents of 24.3, 30.3, 32.9 and 36.7% were produced. In the laboratory, experimental culture in the biofloc system was performed in twelve experimental units at a density of 250 shrimp m − 3 , the four treatments were performed in triplicate. The semi-intensive culture system was applied at the commercial farm in eleven ponds containing 15.5 shrimp m − 2 . After 49 days, the zootechnical performance of the shrimp was rated, and the cost per kilogram of shrimp was produced by both culture systems. In the semi-intensive system, the shrimp fed with a diet containing 32.9% CP showed a higher final weight and weight gain and a lower dietary cost. In the biofloc culture, a higher final weight, weight gain, growth rate and productivity were observed in shrimp fed diets containing 30.3, 32.9 and 36.7% CP compared with those fed a diet containing 24.3% CP. The lowest dietary cost of shrimp farming in the biofloc system was registered in the shrimp fed the 30.3% CP diet, whereas, in the semi-intensive system, the highest final weight, highest weight gain and lowest cost were observed in shrimp fed the 32.9% CP diet.

Organic salts may improvement the animal performance, increasing the efficiency of nutrient utilization and modifying the intestinal microbiota. This study aimed to evaluate the effects of sodium butyrate and sodium propionate supplementation at different levels of dietary inclusions in the growth of Litopenaeus vannamei. In total, seven diets were evaluated: a control diet (without supplementation) and three diets from each sodium salt, propionate and butyrate, in concentrations of 0.5%, 1% and 2%. We used 21 tanks of 6000 L stocked with 150 shrimps (2.53 � 0.03 g). The shrimps fed diets supplemented with propionate and butyrate, in all concentrations, increased their final weight. The feed efficiency, nitrogen retention, protein efficiency rate, survival and yield of shrimps fed the diet containing 2% butyrate were higher in comparison with control treatment. The shrimps supplemented with butyrate also showed lower counts of Vibrio sp. in the intestine. The shrimps fed the diet supplemented with butyrate and propionate also showed higher values of serum agglutination titre. Thus, it is possible to conclude that dietary supplementation with propionate and butyrate in different dietary concentrations modify the intestinal microbiota and improves the growth of L. vannamei.

We evaluated an integrated system for rearing Pacific white shrimp (Litopenaeus vannamei) and white mullet (Mugil curema) in an experimental-scale biofloc system. Performance of the animals, water quality, phosphorus and nitrogen retention and mass balance, and fish hematology and immunology were compared among two treatments: 1) shrimp with mullet integrated system (Shrimp + Mullet), and ii) shrimp without mullet system (Shrimp). A tank with 800 L of useful volume was used to rear the shrimp with a density of 250 shrimp m−3. Another tank with 90 L of useful volume was maintained for rearing fish (15 mullet tank−1) in the Shrimp + Mullet treatment. The shrimp treatment also used the 90-L tanks, but without the fish. Water was recirculated into the mullet tanks via a submerged pump and returned by gravity to the shrimp tanks. The shrimp were fed four times a day. A feeding table was used to adjust the amount of feed provided, and the fish were fed only once daily with a ratio of 1% initial biomass to stimulate biofloc consumption (BFT). After 53 days, shrimp growth (final weight 12.56 ± 0.22 g) and survival (91.8% ± 2.9%) were similar between treatments. Fish had adequate survival (91.1% ± 10.2%), and adequate growth for Mugil species (0.71 ± 0.05 g week−1) in the integrated system. In the Shrimp + Mullet treatment, animal productivity was increased by 11.9% by combining Shrimp + Mullet biomass, when compared to the Shrimp treatment. The Shrimp + Mullet treatment also demonstrated a 16.8% increase in phosphorus retention compared to the Shrimp treatment. The hemato-immunological parameters showed that fish kept in a biofloc system had adequate values comparable to those reported for Mugil species. Thus, these experimental-scale results demonstrate that the integration of shrimp and mullet in biofloc systems increases yield and phosphorous retention without compromising fish health and shrimp growth. In addition, we recommend reproducing this integrated system over a longer period and at commercial-scale, supporting an economic analysis.

This study aimed to evaluate the performance of an integrated multitrophic aquaculture (IMTA) system applied to shrimp rearing in biofloc technology (BFT). The IMTA system consisted of shrimp (Litopenaeus vannamei) in a rearing tank (800 L), a tilapia (Oreochromis niloticus) rearing tank (90 L), and a hydroponic bench with 0.33 m2 of planting space for Sarcocornia ambigua culture. A submerged pump constantly pumped water from the shrimp tank to the tilapia tank. Then, by gravity, water flowed through the sarcocornia hydroponic bench and returned to the shrimp tank. The hydroponic bench had enough capacity for 32 plants. Shrimp, tilapia and sarcocornia stock densities were 312 shrimp m−3 (250 shrimps per 800 L tank), 445 tilapia m−3 (40 tilapias per 90 L tank), and 97 sarcocornia plants m−2 (32 plant per system), respectively. The same experimental units were used in the control system which only differed by the absence of sarcocornia. The initial weight was 4.09 ± 0.025 g, 1.16 ± 0.02 g and 1.17 ± 0.175 g for shrimp, fish and sarcocornia, respectively. Shrimps were fed according to the feed table, and the fishes were fed with 1% of fish biomass, stimulating tilapia to use biofloc as a food source. Results show no difference between shrimp and tilapia performance in either treatment. Only IMTA total yield (4.83 ± 0.19 kg m−3) was significantly higher than that in the control system (3.99 ± 0.045 kg m−3). Nitrate was higher in the control system (12.28 ± 1.27 mg L−1) compared to the IMTA system (9.38 ± 1.09 mg L−1).These results demonstrate a yield increase of up to 21.5% by multitrophic integration of L. vannamei, O. niloticus and S. ambigua in a biofloc system.

O objetivo deste trabalho foi avaliar o uso de suplemento prebiótico, probiótico e simbiótico, na dieta de camarões marinhos (Litopenaeus vannamei) e seus efeitos sobre o crescimento, a microbiota intestinal, a resposta imune e a resistência ao desafio experimental com Vibrio alginolyticus. Foram utilizados quatro tratamentos: prebiótico inulina; probiótico Lactobacillus plantarum; simbiótico Lactobacillus plantarum + inulina; e controle. Os camarões foram distribuídos em 16 tanques de dez mil litros de água, povoados com 200 camarões cada, cultivados por seis semanas. Avaliaram-se a microbiologia do trato intestinal dos camarões e a reposta imune, antes e após o desafio com V. alginolyticus. A concentração de Vibrio spp. no trato digestório foi menor em camarões alimentados com dieta suplementada com prebiótico, probiótico e simbiótico, enquanto a concentração de bactérias acidoláticas foi superior somente nos camarões alimentados com probiótico e simbiótico. O título aglutinante do soro contra V. alginolyticus aumentou no grupo probiótico e simbiótico, antes da infecção, e foi maior em todos os tratamentos após infecção com V. alginolyticus, em comparação ao controle. Não foi observada diferença entre os tratamentos quanto aos demais parâmetros avaliados. As dietas probióticas, prebióticas e simbióticas alteram a microbiota intestinal e aumentam o título aglutinante do soro contra V. alginolyticus; contudo, não alteram a resistência ao desafio nem o crescimento dos camarões. The objective of this work was to evaluate the use of prebiotic, probiotic, and symbiotic supplements, in the diet of shrimp (Litopenaeus vannamei), and their effects on growth, intestinal microbiota, immune response, and resistance to experimental challenge with Vibrio alginolyticus. Four treatments were used: prebiotic (inulin); probiotic (Lactobacillus plantarum); symbiotic (Lactobacillus plantarum + inulin); and a control. Shrimp were distributed in 16 tanks with ten thousand liters of water, stocked with 200 shrimp each, cultivated for six weeks. Microbiology of the intestinal tract and the immune response of shrimp were evaluated before and after the challenge with V. alginolyticus. The concentration of Vibrio spp. in the digestive tract was lower in shrimp fed prebiotic, probiotic, and symbiotic-supplemented diet, while concentration of lactic acid bacteria was higher only in shrimp fed probiotic and symbiotic-supplemented diet. The title of agglutinating serum against V. alginolyticus increased in shrimp fed the probiotic and symbiotic-supplemented diet prior to infection, and was higher in all treatments after infection with V. alginolyticus, in comparison to the control. No difference was observed between treatments for the remaining parameters. Probiotic, prebiotic, and symbiotic diets change intestinal microbiota, and increase the agglutinating serum title against V. alginolyticus; however, they change neither the shrimp resistance to challenge by V. alginolyticus nor shrimp growth.

We have performed here a gene expression analysis to determine the developmental stage at the main genes involved in crustacean immune response begin to be expressed and their changes in mRNA abundance during shrimp development. By using a quantitative PCR-based approach, we have measured the mRNA abundance of 24 immune-related genes from different functional categories in twelve developmental stages ranging from fertilized eggs to larval and postlarval stages and also in juveniles. We showed for the first time that the main genes from the RNAi-based post-transcriptional pathway involved in shrimp antiviral immunity are transcribed in all developmental stages, but exhibit a diverse pattern of gene expression during shrimp ontogenesis. On the other hand, hemocyte-expressed genes mainly involved in antimicrobial defenses appeared to be transcribed in larval stages, indicating that hematopoiesis initiates early in development. Moreover, transcript levels of some genes were early detected in fertilized eggs at 0–4 h post-spawning, suggesting a maternal contribution of immune-related transcripts to shrimp progeny. Altogether, our results provide important clues regarding the ontogenesis of hemocytes as well the establishment of antiviral and antimicrobial defenses in shrimp.

Aiming at assessing the cryopreservation potential of Litopenaeus vannamei sperm cells, 74 spermatophores were manually extracted from the sexually mature individuals. After the toxicity test to define the cryoprotectant concentration, suspensions of spermatic cells were cryopreserved in the groups in freezing solutions comprising different cryoprotectants such as dimethyl sulfoxide (DMSO) and ethylene glycol (EG) at 10% concentration. Each treatment was divided in subgroups for storage in liquid nitrogen during 0, 30, 60 and 90 days, in triplicate. After thawing at 25oC for 40 seconds, cell viability in the suspensions was analyzed under the microscope in eosin-nigrosin stain and flow cytometry. There were no significant differences between the cryoprotectants used. For all the treatments, lower and higher mortalities occurred in the 0 and 90 days, respectively. By applying the eosin-nigrosin technique, lower and higher mortalities were 23.17 and 82.11% for DMSO and 29.94 and 83.72% for EG, while the flow cytometry registered mortalities of 2.42 and 55.13% for DMSO and 0.90 and 55.56% for EG. The Spearman correlation coefficient indicated a positive correlation (R=0.91) between the two techniques used. It was concluded that there was a decrease in cell viability within a longer cryopreservation time. Key words: cryogenics, semen, marine shrimp, cell viability

The objective of this work was to isolate strains of lactic acid bacteria with probiotic potential from the digestive tract of marine shrimp (Litopenaeus vannamei), and to carry out in vitro selection based on multiple characters. The ideotype (ideal proposed strain) was defined by the highest averages for the traits maximum growth velocity, final count of viable cells, and inhibition halo against nine freshwater and marine pathogens, and by the lowest averages for the traits duplication time and resistance of strains to NaCl (1.5 and 3%), pH (6, 8, and 9), and biliary salts (5%). Mahalanobis distance (D²) was estimated among the evaluated strains, and the best ones were those with the shortest distances to the ideotype. Ten bacterial strains were isolated and biochemically identified as Lactobacillus plantarum (3), L. brevis (3), Weissella confusa (2), Lactococcus lactis (1), and L. delbrueckii (1). Lactobacillus plantarum strains showed a wide spectrum of action and the largest inhibition halos against pathogens, both Gram-positive and negative, high growth rate, and tolerance to all evaluated parameters. In relation to ideotype, L. plantarum showed the lowest Mahalanobis (D²) distance, followed by the strains of W. confusa, L. brevis, L. lactis, and L. delbrueckii. Among the analyzed bacterial strains, those of Lactobacillus plantarum have the greatest potential for use as a probiotic for marine shrimp. O objetivo deste trabalho foi isolar cepas de bactérias ácido-lácticas, com potencial probiótico do trato digestório de camarões marinhos e realizar seleção in vitro baseada em múltiplos caracteres. O ideótipo (cepa ideal proposta) foi definido por meio das maiores médias para os caracteres velocidade máxima de crescimento, contagem final de células viáveis e halo de inibição contra nove patógenos de origem continental e marinha, e pelas menores médias para os caracteres tempo de duplicação e tolerância das cepas a NaCl (1,5 e 3%), pH (6, 8 e 9) e sais biliares (5%). Foram estimadas as distâncias de Mahalanobis (D²) entre as cepas avaliadas, e as melhores foram aquelas que apresentaram menor distância do ideótipo. Foram isoladas dez cepas de bactérias identificadas bioquimicamente como Lactobacillus plantarum (3), L. brevis (3), Weissella confusa (2), Lactococcus lactis (1) e L. delbrueckii (1). As cepas de L. plantarum apresentaram amplo espectro de ação e os maiores halos de inibição, tanto para patógenos Gram-positivos quanto negativos, alta taxa de crescimento e tolerância a todos os parâmetros avaliados. Em relação ao ideótipo, L. plantarum apresentou a menor distância de Mahalanobis (D²), seguida pelas cepas de W. confusa, L. brevis, L. lactis e L. delbrueckii. Entre as cepas bacterianas avaliadas, as de L. plantarum apresentam o maior potencial para uso como probiótico para camarões marinhos.

O objetivo deste trabalho foi determinar o método de coleta de fezes mais adequado para camarões marinhos (Litopenaeus vannamei) na determinação do coeficiente de digestibilidade aparente da proteína bruta e da matéria seca de dietas comerciais, e avaliar o efeito da suplementação de Lactobacillus plantarum na dieta sobre o coeficiente de digestibilidade aparente da proteína, da energia e da matéria seca. Foram testados três sistemas de coletas de fezes: decantação, sifonamento e dissecação dos animais. A coleta por decantação foi o sistema mais adequado para a determinação in vivo dos coeficientes de digestibilidade aparente da proteína bruta e da matéria seca das dietas, por apresentar baixo grau de lixiviação das fezes e por ser de fácil execução. Nos camarões alimentados com a dieta suplementada com L. plantarum, foi observado maior coeficiente de digestibilidade aparente da proteína (84,18±0,35%) e da energia (81,28±0,16%), quando comparado ao grupo controle (82,73±1,01 e 78,63±0,93%, respectivamente). O uso de L. plantarum como aditivo probiótico na ração pode trazer benefícios nutricionais a L. vannamei, com melhor aproveitamento dos nutrientes. The objective of this work was to determine the most adequate feces collection method for sea shrimp (Litopenaeus vannamei) in the determination of the apparent digestibility coefficients of crude protein and dry matter of commercial diets, and to evaluate the effect of supplementation of Lactobacillus plantarum to the diet on the apparent digestibility coefficient of protein, energy, and dry matter. Three feces collection methods were tested: decanting, siphoning, and animal dissection. Decanting was the method best suited for the in vivo determination of the apparent digestibility coeffcients of crude protein and dry matter of commercial diets, since it promoted lower nutrient leaching from feces and was easy to perform. Shrimp fed the diet supplemented with probiotic showed higher apparent digestibility of crude protein (84.18±0.35%) and energy (81.28±0.16%) when compared to the control group (82.73±1.01 and 78.63±0.93%, respectively). The use of L. plantarum as a probiotic additive in shrimp diets can provide nutritional benefits to L. vannamei, with better nutrient utilization.

O objetivo deste trabalho foi avaliar a substituição de dois níveis de inclusão de farinha de peixe por outros ingredientes, nas dietas de Litopenaeus vannamei cultivados em sistema bioflocos. Foram avaliadas dietas sem uso de farinha de peixe (dieta A, 100% de substituição), com inclusão de 12,5% de farinha de peixe (dieta B, 40% de substituição) e direta controle com inclusão de 21% (dieta C, 0% de substituição). Nas dietas A e B, o farelo de soja e as farinhas de carne e vísceras foram os principais substitutos proteicos. Foram analisados os índices de desempenho dos camarões e os parâmetros físicos e químicos da água de cultivo. Os camarões alimentados com a dieta B apresentaram maior peso final (11,63±1,38 g), em comparação aos camarões alimentados com a dieta A (peso final, 9,39±0,31 g) e com a dieta C (peso final, 10,20±1,10 g). Os demais parâmetros de desempenho como produtividade, conversão alimentar e sobrevivência não apresentaram diferenças entre os tratamentos. A redução de até 40,0% da farinha de peixe pode ser feita em cultivos superintensivos de L. vannamei com bioflocos, sem interferir em seu desempenho zootécnico e na qualidade de água do cultivo. The objective of this work was to evaluate the substitution of two fish meal inclusion levels by other ingredients, in the diets of Litopenaeus vannamei reared in a biofloc system. The following diets were evaluated: without fish meal (diet A, 100% replacent), with addition of 12.5% of fish meal (diet B, 40% replacent), and a control diet with addition of 21% (diet C, 0% replacent). In diets A and B, soybean meal and meat and viscera meals were the main protein replacement. The performance indexes of shrimps and the physical and chemical parameters of water were analyzed. Shrimp fed diet B had higher final weight (11.63±1.38 g), compared to shrimp fed a diet A (9.39±0.31 g final weight) or C (10.20±1.10 g final weight). The other parameters of performance such as yield, feed conversion and survival did not differ between treatments. The reduction of fish meal by 40.0% can be done in a L. vannamei superintensive biofloc system without interfering on their growth performance or on the water quality of the culture.

This study evaluated the pathogenic characteristics of bacteria isolated from Litopenaeus vannamei during an outbreak at the Laboratory of Marine Shrimp, UFSC, Santa Catarina State, Brazil. Their virulence potential in larvae and juvenile shrimp and the effects on the total haemocyte count, phenoloxidase activity and serum agglutinate titre were examined after experimental infection. Bacterial strains were isolated from larvae and adult shrimps, identified by the AP120E biochemical system as: two strains of Vibrio alginolyticus, three of Aeromonas salmonicida and one of Pasteurella multocida sp. and Pasteurella sp. All the bacterial strains isolated in this study caused mortality in shrimp. One strain of V. alginolyticus was responsible for 97.3 and 88.7% mortality in larvae and juvenil shrimps, respectively. The shrimp immunological system was influenced by experimental infection with V. alginolyticus. Decrease in the total haemocyte count and increase in the phenoloxidase activity and the serum agglutinate titre (p < 0.05) were observed. The results showed the high pathogenicity of V. alginolyticus isolated from larvae and juvenile reared marine shrimp.

The aim of this study was to assess the effect of a probiotic (Lactobacillus plantarum) supplemented diet on Nile tilapia (Oreochromis niloticus) in a polyculture system with marine shrimp (Litopenaeus vannamei) as regards culture performance, hematology, and gut bacterial microbiota. Ten 20-m² pens were arranged in one earthen pond and stocked with 2 fish (41.9 g) m−2 and 10 shrimp (2.3 g) m−2, in total of 40 Nile tilapias and 200 shrimp per experimental unit. Tilapia groups in five of the experimental units were fed a commercial diet supplemented with L. plantarum and the other five with an unsupplemented commercial diet (control). After 12 weeks of culture, the tilapia groups fed the probiotic-supplemented diet presented values 13.6, 7.5, and 7.1% higher for feed efficiency, yield, and final weight, respectively. Viable culturable heterotrophic bacteria counts were reduced, and the number of lactic acid bacteria was increased in the gut of fish and shrimp fed the probiotic-supplemented diet. Hematological analyses showed higher number of thrombocytes and leukocytes in tilapia fed the supplemented diet. L. plantarum utilized in this study colonized the gut of tilapia and shrimp and resulted in reduced number of total bacteria and increased tilapia final weight and feed efficiency.

Objetivou-se averiguar o efeito da bactéria Lactobacillus plantarum e células inativas das bactérias Vibrio alginolyticus, Aeromonas salmonicida e Pasteurella multocida na sobrevivência larval de Litoenaeus vannamei, no teste de estresse e infecção experimental com Vibrio harveyi. Foram utilizados tanques cônicos de 30 L, povoados com 400 larvas cada, no estádio de pós-larva cinco. Tratamentos em triplicatas foram consistidos de: 1: ração comercial (controle), 2: ração comercial + bacterina via oral na artemia, 3: ração comercial + bacterina via imersão e 4: ração comercial com inóculo de Lactobacillus plantarum. A aplicação da bacterina ocorreu seis horas antes da infecção e do teste de estresse; enquanto o Lactobacillus plantarum foi administrado por 15 dias antes dos desafios. As pós-larvas do tratamento 4 (ração suplementada com L. plantarum) obtiveram maior índice de sobrevivência no teste de estresse (87,86 ± 2,35%), seguido dos tratamentos 2 e 3 (bacterina via imersão e oral) com 81,54±1,50% e 80,16 ± 2,15% respectivamente, superiores ao índice do controle (72,63 ± 3,34%). Já no desafio com V. harveyi, os animais do grupo tratado com a adição de bacterina via imersão apresentaram maior sobrevivência (79,60 ± 7,12%). As pós-larvas dos tratamentos com bacterina via oral na artêmia e alimentadas com o probiótico L. plantarum, apresentaram sobrevivências de 65,60 ± 5,18% e 69,60 ± 10,43 %, respectivamente, sendo superiores ao controle (56,4 ± 5,58%), quando desafiados com V. harveyi. Os resultados demonstram que o uso de ração com L. plantarum e bacterina aumentam a sobrevivência das pós-larvas de L. vannamei frente aos testes de estresse e infecções experimentais com V. harveyi.

The objective of this work was to assess the time-related action of probiotic Lactobacillus plantarum in the bacterial microbiota of the digestive tract of Litopenaeus vannamei, and the relation of total haemocyte count and serum phenol oxidase activity of shrimp challenged with Vibrio harveyi. Shrimps were fed with a probiotic-supplemented diet, for eight days, then shifted to a commercial diet. Shrimps fed only with the commercial diet served as control. Evaluations were made on the 8th day of experiment and repeated two, four, six and eight days later. Total lactic bacteria in the digestive tract was higher until the 4th day of evaluation in the probiotic-supplemented group. Vibrio spp. counts were higher in the control at days zero and two. Until the 4th day of evaluation, the total haemocyte counts in shrimps after challenge with V. harveyi were higher in probiotic-supplemented group than in control group. Significant difference was not observed in phenol oxidase activity. On the 6th day after shifting from supplemented to control diet, all parameters were equal in both groups, suggesting that the time-related action of L. plantarum in shrimp is short. O objetivo deste trabalho foi avaliar o tempo de atuação de Lactobacillus plantarum, na microbiota bacteriana intestinal de Litopenaeus vannamei, e sua relação com a contagem total de hemócitos e a atividade da fenoloxidase após a exposição a Vibrio harveyi. Os camarões foram alimentados com dieta suplementada com probióticos por oito dias e, depois, substituída por dieta comercial. Os camarões alimentados com dieta comercial constituíram o controle. As avaliações foram realizadas no oitavo dia de experimento e repetidas dois, quatro, seis e oito dias depois. A contagem total de bactérias lácticas no intestino foi maior até o quarto dia de avaliação, no grupo alimentado com probióticos. A contagem de Vibrio spp. foi superior no grupo controle nos dias zero e dois. Até o quarto dia, a contagem total de hemócitos dos camarões após a inoculação de V. harveyi foi superior no grupo alimentado com probiótico em relação ao grupo controle. Não foi observada diferença na atividade da fenoloxidase. No sexto dia, após a substituição da dieta suplementada com probióticos por dieta controle, todos os parâmetros avaliados foram iguais nos dois grupos, o que indica que o tempo de atuação de L. plantarum em camarões é curto.

This study evaluated the survival, post-larvae quality, and the population of bacteria in Litopenaeus vannamei after the addition of two strains of lactic-acid bacteria (2 and B6) experimentally infected by Vibrio harveyi. Fifteen hundred nauplii were distributed in 20 L capacity tanks with four replicates. The survival of control animals was lower (21%) than that of animals fed with the strains B6 (50%) and 2 (44%). Total bacterial population in the water and larvae, as well as of the Vibrio ssp. in water was not different among the treatments. No difference was observed in the population of Vibrio ssp. between the control larvae (5.5±0.5 log UFC/mL) and that fed with strain 2 (5.4±0.1 log UFC/mL). Shrimp from control and fed with strain 2 showed significantly higher bacterial population than those fed with strain B6 (1.2±0.2 log UFC/mL). It was detected the lower load of Vibrio ssp. bacteria with potential of pathogenicity after feeding with strain B6.Moreover, these larvae showed more active behavior and low number of necrosis in relation to the control group and to that fed with strain 2.Este trabalho avaliou a adição de duas cepas de bactérias lácticas (2 e B6) na sobrevivência, qualidade de pós-larva e na população de bactérias na larvicultura de Litopenaeus vannamei experimentalmente infectado por Vibrio harveyi. Mil e quinhentos náuplios foram distribuídos em tanques de 20 L com quatro repetições. A sobrevivência dos animais controle foi menor (21%) do que a dos alimentados com as cepas B6 (50%) e 2 (44%). Sobrevivência de misis após desafio com V. harveyi foi maior em B6 do que nos outros tratamentos. A população total de bactérias na água e nas larvas, bem como de Vibrio ssp. na água não foi diferente entre os tratamentos. Não houve diferença, também, entre a população de Vibrio ssp. em larvas do grupo controle (5,5±0,5 log UFC/mL) e larvas alimentadas com a cepa 2 (5,4±0,1 log UFC/mL). Camarões do grupo controle e alimentados com cepa 2 apresentaram maior população de bactérias do que os alimentados com cepa B6 (1,2±0,2 log UFC/mL). Foi comprovada a menor presença de bactérias entéricas com potencial de patogenicidade nos animais alimentados com a cepa B6, apresentando também comportamento mais ativo e menor número de necroses em relação ao controle e cepa 2.

A preliminary study to characterize filamentous bacteria, whose presence is related to high mortality of Litopenaeus vannamei larvae cultured in Santa Catarina State, Brazil, is reported. The extract of infected larvae was diluted in different concentrations, cultured in marine agar (Difco, Marine Agar 2216) and incubated at 30 degrees C for 48 hours. The biochemical characterization included hydrolytic reactions of starch, gelatin and tyrosine, growth in TCBS agar, growth in 0 and 37 per thousand salinity, pigment production in tyrosine agar, production of H2S, nitrate reduction, congo red reaction, oxidase and catalase. The isolated bacteria belong to the species Flexibacter maritimus, Gram-negative bacilli of 0.4-0.5 microm width and 15 microm length. Experiments were carried out on pathogenicity of F. maritimus in post-larvae of L. vannamei. Survival and symptoms in L. vannamei post-larvae 24 hours after inoculation with F. maritimus and its growth in marine agar were evaluated. Mortality was detected around 92,5% as well as symptoms like melanized lesions in several parts of body, discolouration of gills, bad formation of appendages and of the last abdominal segment, low motility and feeding reduction. The experimental infection results suggested that isolated bacteria of the genus Flexibacter are pathogenic to the shrimp Litopenaeus vannamei post-larvae.