Feeding protocol with live feed and inert diet to determine the ideal age/size for weaning mullet Mugil liza (Valenciennes, 1836).

The Lebranche mullet Mugil liza is a marine fish of great importance for artisanal and industrial fishing, as well as aquaculture. The use of live feeds during the larviculture phase of marine fish is a significant component of production costs. The present study evaluated the effects of the feeding transition on different larval stages of M. liza, from the combined supply of live feed (Brachionus rotundiformis + Artemia franciscana) and live + inert feed (Artemia franciscana + inert diet) until the weaning phase to only the inert diet. A total of 3240 M. liza larvae (weight 0.0133 ± 0.0062 g and length 0.793 ± 0.160 cm) were distributed among the 12 experimental units (n = 270), resulting in four groups with three replicates each. Treatment groups consisted of feed transition with A. franciscana (enriched metanauplii) to commercial inert feed starting weaning at four different larval ages: 28, 31, 34, and 37 days post hatching. Zootechnical performance indexes and intestinal histomorphometry were evaluated. Mortality, condition factor, and length variation coefficient did not show significant differences between treatments. Final weight, final length, weight gain, and length gain were significantly greater in larvae that started weaning at 31, 34, and 37 days post hatching. Weight coefficient of variation was significantly higher in larvae that started weaning at 28 days (67.51 ± 11.70) compared to 37 days (34.40 ± 7.30). In intestinal histology, villi height (180.3 ± 4.4) was significantly higher in larvae that started weaning at 37 days post hatching. Considering the evidence found in the present study, it is recommended to start weaning M. liza on the 31st day post-hatching, using a 2-day co-feeding protocol (31st and 32nd days). From the 33rd day after hatching, M. liza larvae can receive only commercial feed.

Effects of dietary supplementation with inactivated Lactobacillus plantarum on growth performance, haemato-biochemical parameters, liver fatty acids profile and intestinal microbiome of Nile tilapia.

This study investigated the effects of dietary supplementation with inactivated Lactobacillus plantarum for Nile tilapia (Oreochromis niloticus). Three treatments, in quintuplicate, were established: a control group, fish fed a diet without additives; LP group, fish fed a diet supplemented with live probiotic; and IP group, fish fed a diet supplemented with inactivated probiotic. Final weights (49.40 ± 3.15 g) and weight gains (38.20 ± 3.23 g) were increased in tilapia in the IP group. Feed conversion (1.32 ± 0.04) decreased significantly in the IP group. Haemato-biochemical parameters were significantly influenced by dietary supplementation. Erythrocyte count (262.74 ± 69.28 × 106 µL-1) was significantly low, while albumin (1.79 ± 1.12 g dL-1) and cholesterol (254.14 ± 98.49 mg dL-1) were high in the control group. Dietary supplementation modified the tilapia microbiome. Rhodobacter was abundant in fish intestines from the control and IP groups. Phreatobacter was abundant in the IP and LP groups, while Aurantimicrobium and Bosea were abundant in the LP group. Oleic acid (C18:1n9) was significantly increased in the LP (3.25 ± 0.49%) and IP (3.02 ± 0.30%) groups. Hexadecatrienoic acid (C16:3n4) was significantly increased (0.04 ± 0.01%) in the IP group, while Cis 11,14,17-eicosatrienoic acid (C20:3n3) (0.31 ± 0.03%) and adrenic acid (C22:4n6) (0.11 ± 0.02%) were significantly decreased in the LP group. Additionally, monounsaturated fatty acids (MUFA) were significantly increased (4.83 ± 0.35%) in the LP group compared to that in the control group. Collectively, these results indicate the potential of inactivated L. plantarum for use in commercial feed, leading to the conclusion that both inactivated and live L. plantarum can improve the Nile tilapia metabolism, altering haematological and biochemical markers.

Effects of a multi-strain Bacillus probiotic on the intestinal microbiome, haemato-immunology, and growth performance of Nile tilapia.

The study evaluated dietary supplementation with a feed additive composed of multi-strain Bacillus for Nile tilapia Oreochromis niloticus. In vitro and in vivo assays employing culture-based microbiological methods and metagenomics were performed. Additionally, the study assessed the haemato-immunology, intestinal microbiome, and growth performance of the animals. For this, 30 juvenile Nile tilapia were used in the in vitro assay and 180 (60 + 120) in the in vivo assays. In the in vitro assay, we found evidence of adhesion of the probiotic bacteria to the intestinal mucus of fish, corroborated in the 15-day in vivo assay, in which the count of B. licheniformis was significantly higher in fish fed with probiotic when compared to fish of the control group. Furthermore, in the 50-day in vivo trial, a metagenomic analysis provided evidence for the modulation of the intestine microbiome of Nile tilapia by dietary supplementation of the probiotic. In addition, there was an increase in species richness, higher abundance of potentially probiotic autochthonous species and a lower abundance of Aeromonas sp. when the animals were fed the supplemented diet. Finally, no significant differences were observed in growth performance and haemato-immunological analyses, suggesting no harm to fish health when the product was supplemented for 15 and 50 days. The in vitro results indicate that the multi-strain probiotics were able to adhere to the intestinal mucus of Nile tilapia. Additionally, a modulation of the intestinal microbiome was evidenced in the in vivo assay.

Effects of supplementation with different zinc-based products on the growth and health of Nile tilapia.

Zinc is one of the essential microelements for the metabolism of animals. Zinc nanoparticles may have higher bioavailability due to their low specific surface area, facilitating absorption by fish. The present study aimed to evaluate the effects of supplementation with different zinc-based products on the growth and health of Nile tilapia Oreochromis niloticus. Zinc, in different sizes (nanoparticles or bulk) and forms (inorganic or organic), were used as a supplement in the tilapia diet at a dose of 15 mg kg feed-1 for 60 days. At the end of the feeding trial, production performance, hemato-immunological parameters, activity of antioxidant system enzymes, exposure to Streptococcus agalactiae and zinc concentration in the muscle were examined. After the bacterial challenge, the mean corpuscular hemoglobin concentration (MCHC) significantly increased in the fish treated with organic zinc, inorganic nano zinc, and organic nano zinc, while in the control group (inorganic zinc), MCHC remained unchanged. Regarding defense cells, dietary inorganic nano zinc increased the number of basophils (1.50 ± 1.10) compared to organic zinc (0.80 ± 0.90). Lymphocyte count increased after the challenge only in the organic zinc treatments (bulk and nanoparticles). Neutrophils decreased in the control (inorganic zinc) (2.20 ± 1.70) and inorganic nano zinc (2.60 ± 2.70) treatments after the challenge. When compared before and after the bacterial challenge, the plasma antimicrobial titer significantly increased after the bacterial challenge in all treatments. No significant differences were observed for total proteins, enzymes (SOD and CAT), cumulative survival and zinc deposition on fillet. In conclusion, organic zinc in nanoparticles or bulk size increased Nile tilapia innate defense during bacterial infection. However, the other parameters evaluated were not affected by zinc particle size or form (organic or inorganic), indicating that further evaluations should be conducted with organic zinc in nanoparticles or bulk size in the tilapia diet.

Biofilm viability and microbial community of non-inoculated moving bed biofilm reactor in Nile tilapia Oreochromis niloticus cultivation.

The aim of this study was to evaluate two moving bed biofilm reactors (MBBR) without nitrifying bacteria inoculation. Biofilms and viable bacterial colonies were evaluated after 124 days. MBBR bioreactors received water from Oreochromis niloticus fish farming and water quality parameters were monitored daily. Four distinct phases with different fish stocking density were established.: phase 1 (2.40 kg m-3), phase 2 (4.95 kg m-3), phase 3 (8.71 kg m-3) and phase 4 (12.23 kg m-3). The successful maturation of the bioreactors occurred around on the 100th experimental day when the nitration rate increased to 57 % in MBBR1 and 38 % in MBBR2. 105 species were identified in the biofilms, which were grouped into 65 genera, three of which were essential: Pseudomonas (21.7 %), Nitrospira (15.1 %) and Gemmobacter (11.2 %). MBBR start-up without bacterial inoculation is time-consuming, however, strengthened by important nitrifying groups.

Lippia sidoides essential oil at concentration of 0.25% provided improvements in microbiota and intestine integrity of Danio rerio.

The study evaluated the effects of dietary supplementation with Lippia sidoides essential oil on the microbiota and intestinal morphology of Danio rerio. For this, 448 fish were randomly distributed in 28 tanks divided into a control group fed a commercial diet without supplementation, a group fed a commercial diet containing grain alcohol and five groups fed a commercial diet containing essential oil of L. sidoides (LSEO) at concentrations of 0.25%, 0.50%, 0.75%, 1.00% and 1.25%. After the period of dietary supplementation, biological materials were collected for microbiological and histological analyses. There were no significant differences regarding the microbiological count between the groups. Diversity of the microbiome was higher in 0.25% group than in control group. LSEO inhibited the growth of potentially pathogenic bacteria. Fish fed LSEO0.25% showed greater intestinal histomorphometric indices. The inclusion of LSEO at 0.25% in the diet of D. rerio provided improvements in fish microbiota and intestine integrity.

O estudo avaliou os efeitos da suplementação dietária com óleo essencial de Lippia sidoides sobre a microbiota e a morfologia intestinal de Danio rerio. Para isso, 448 peixes foram distribuídos aleatoriamente em 28 tanques divididos em grupo controle alimentado com dieta comercial sem suplementação, grupo alimentado com dieta comercial contendo álcool de cereais e cinco grupos alimentados com dieta comercial contendo óleo essencial de L. sidoides (LSEO) em concentrações de 0,25%, 0,50%, 0,75%, 1,00% e 1,25%. Após o período de suplementação alimentar, foram coletadas amostras biológicas para análises microbiológicas e histológicas. Não houve diferenças significativas quanto à contagem microbiológica entre os grupos. A diversidade do microbioma foi maior no grupo de 0,25% do que no grupo controle. LSEO inibiu o crescimento de bactérias potencialmente patogênicas. Peixes alimentados com LSEO 0,25% apresentaram maiores índices histomorfométricos intestinais. A inclusão de LSEO a 0,25% na dieta de D. rerio proporcionou melhorias na microbiota e na integridade intestinal dos peixes.

Dietary effect of multi-strain prebiotics and probiotics on growth, hemato-biochemical parameters, intestinal histomorphometry, and resistance to hypoxia in juvenile tambaqui (Colossoma macropomum).

This study evaluated the effects of a multi-strain prebiotics and probiotics on the diet of tambaqui Colossoma macropomum. One hundred and fifty juvenile tambaqui (20.2 ± 3.6 g and 10.32 ± 5.78 cm) were randomly distributed in 15 experimental units with a volume of 80 L and fed for 45 days with a diet containing the symbiotic additive at five inclusion levels (0, 2, 4, 6 and 8 g kg feed- 1). At the end of the period, growth performance, survival, hemato-biochemical and intestinal parameters, as well as the fish's resistance to stress were evaluated. The additive did not alter the growth performance, hemato-biochemical and intestinal parameters of the tambaqui. However, the hemato-biochemical parameters of aspartate aminotransferase (AST), plasma protein, hematocrit, glucose, triglycerides and cholesterol showed differences between treatments after the stress challenge. AST increased significantly during challenge and post-challenge. Plasma protein increased significantly during and after the challenge. The hematocrit was highest at 48 h after the challenge and lowest in the 2 g, 6 g and 8 g groups. Glucose was significantly reduced 24 h after the challenge, while triglycerides were lower 24 h and 48 h after the challenge. Cholesterol increased significantly in the challenge. There was an interaction between the factors sampling time and symbiotic concentration for hematocrit and glucose. In hypoxic stress situations, the 2 g concentration was more favorable for the fish. We recommend further studies with 2 g kg feed- 1 in trials lasting more than 45 days.

Dietary supplementation with Lippia sidoides essential oil improves organ integrity but the specific activity of antioxidant enzymes is dose-dependent in Danio rerio.

The nutritional quality of food can affect the health of animals. This study examined the effects of dietary supplementation with Lippia sidoides essential oil (LSEO) on the physiology of Danio rerio. Four hundred fourty-eight fish were divided into 28 tanks and subjected to different dietary treatments: a control group with no supplementation, a group with grain alcohol supplementation and five groups with LSEO at concentrations of 0.25%, 0.50%, 0.75%, 1.00% and 1.25%. After 15 days, histological and enzymatic analyses were conducted. The 0.25% LSEO group exhibited lower glutathione peroxidase and catalase activity compared to the 1.00% group. Additionally, fish in the 0.25% LSEO group showed improved liver, kidney and splenic integrity indices. These findings support the inclusion of 0.25% LSEO in the diet of D. rerio, suggesting potential benefits for fish physiology and encouraging further research on phytotherapeutics in fish diets.

Laboratory-controlled challenges of streptococcosis in Nile tilapia using the oral route (infected-feed) for infection.

The study aimed to evaluate the oral infected-feed, intragastric-gavage, and intraperitoneal routes of the Streptococcus agalactiae infection in Nile tilapia (Oreochromis niloticus). For this purpose, 270 juveniles of Nile tilapia, with an average weight of 2 g, were distributed in 18 experimental units of 90 L, acclimatized, and raised for 55 days, until reaching 50 g (median weight). The experimental design was entirely randomized in six treatments, three of which were composed by bacterial infection routes: intraperitoneal 100 µL fish-1 [108 CFU], intragastric 100 µL fish-1 [108 CFU], feed inoculum 100 µL g feed-1 [109 CFU], and three respective control groups. Clinical signs were observed, and mortalities monitored until reaching 50% in the infected groups. Then, tissue samples from the spleen, liver, intestine, brain, and blood were collected from 20 fish per treatment for histopathological and hemato-immunological analyses. In addition, a related mortality curve was established at the end of the experimental challenge. The intraperitoneal and intragastric routes were more aggressive than the oral inoculum, causing greater brain damage, acute hemato-immunological response, and early mortality. While the orally fed inoculum, fish presented brain lesions with less intensity, and a chronic haemato-immunological response, the mortalities occurred twice as long as the other routes. The present research demonstrated that the S. agalactiae oral (feed inoculum) administration can be an innovative methodology to future experimental challenges in aquaculture research.

Effects of sodium butyrate and Lippia origanoides essential oil blend on growth, intestinal microbiota, histology, and haemato-immunological response of Nile tilapia.

This study aimed to verify the effects of dietary supplementation with sodium butyrate and Lippia origanoides, combined and isolated, on the health and zootechnical performance of Nile tilapia juveniles Oreochromis niloticus. A total of 120 fish (5.38 ± 0.65 g) were randomly distributed in 12 experimental units and fed different experimental diets for 30 days, namely: commercial diet without supplementation (Unsupplemented); commercial diet supplemented with 0.5% sodium butyrate (Butyrate); commercial diet supplemented with 0.125% L. origanoides (Lippia) and commercial diet supplemented with a mixture of 0.5% sodium butyrate and 0.125% L. origanoides (Butyrate + Lippia). After preparing the experimental diets there was an increase in the pH of diet Butyrate when compared to the other diets. After 30 days the fish supplemented with Butyrate + Lippia showed reduction significate in the mean corpuscular haemoglobin, concentration of total heterotrophic bacteria in the intestine, and lymphocyte infiltrates in the liver. Besides that, the supplementation with Butyrate + Lippia promoted an increased number of intestinal villi compared to the fish Unsupplemented ones. Additionally, fish fed a diet containing only Lippia presented an increase in the villus perimeter in the posterior region of the gut and in the red blood cell number. Animals supplemented only with sodium butyrate demonstrated increased lactic acid bacterium in the gut and macrosteatosis in the liver, besides decreased melanomacrophages in the spleen. The use of sodium butyrate associated with essential oil had positive effects on the intestinal microbiota, intestinal structure, liver, and spleen integrity, suggesting a greater efficiency of the compounds when used together in the nutrition of Nile tilapia juveniles.

Parasitological and histopathological diagnosis of a non-native fish (Oreochromis sp.) with a noticeable presence in a natural Brazilian river environment.

Nile tilapia is an exotic species produced in Brazilian aquaculture. The aim of this study was to constitute a pathological diagnosis in Oreochromis sp. with a noticeable presence in a river in southern Brazil, and to verify the origin of these fish in this natural habitat, an environment with high sanitary precariousness due to discharges of industrial effluents. A total of 30 fish were collected during three sampling events and their health status was assessed by parasitological fauna survey, parasitological indices and histopathological examination of the gills, liver, spleen and kidney. Five species of monogenea were identified infesting gills and stomach: Cichlidogyrus sclerosus, C. halli, C. thurstonae, Scutogyrus longicornis and Enterogyrus cichlidarum; and 4 species of trichodinidae infesting the body surface and gills: Trichodina magna, T. compacta, T. centrostrigeata and Paratrichodina africana. The results showed differences according to the seasonality of the collection, with higher parasite count and larger tissue alterations in summer. Histopathological analysis fluctuated from moderate to severe, but necrosis prevailed in all organs and in all fish, indicating that these fish did not enjoy full health.

Amyloodiniosis in farmed Sardinella brasiliensis (Steindachner, 1879): Case report of successful therapeutic control with copper sulfate.

Brazilian sardine is emerging as a promising species in Aquaculture. This study describes for the first time a case of parasitic infestation by Amyloodinium in Brazilian sardines Sardinella brasiliensis obtained from natural spawning in captivity. The sardines kept in nurseries were naturally parasitized by the amylodiniosis causative agente the dynoflagellate A. ocellatum with high mortalities above 50%. Fish presented clinical signs characteristic of amyloodiniosis which included easily perceived behavioral changes such as loss of appetite, scraping of the body against objects, walls and bottom, nursery pipes, agglomerations near the aerators and water inlets, presented with accelerated opercular beating and erratic swimming. For therapeutic treatment copper sulfate was used for 10 days. At the end of the treatment period the fish had no clinical signs or presence of the parasite on the body surface, indicating that the application of copper sulfate in concentration of 0.2 mg L-1 of Cu+ was effective to control this parasite, apparently without causing damage to Brazilian sardine.

Can histology and haematology explain inapparent Streptococcus agalactiae infections and asymptomatic mortalities on Nile tilapia farms?

The aim of this study was to characterise possible histopathological and haemato-immunological changes after subclinical infection by S. agalactiae S13 serotype Ib. One hundred juveniles of Nile tilapia with average weight of 45 g were distributed in ten 90 L experimental units. After the acclimation period 25 fish were euthanised, and fragments of liver, spleen and posterior mid-intestine tissue were sampled to verify the integrity of the organs and blood samples taken to check the haematological profile. Fifty animals were used to verify the optimal dosage for the challenge. The remaining 25 fish were infected with S. agalactiae S13. After 96 h, tissue fragments from the liver, spleen and posterior mid-intestine and blood samples were collected. The analyses revealed that, 96 h after contagion, S. agalactiae S13 serotype Ib caused subclinical lesions in the liver and spleen that are not commonly described as pathognomonic, in addition to haematological alterations. These results allow a better understanding of sudden recurrent mortalities in Brazilian tilapia farms, since the serotype Ib of S. agalactiae causes inapparent infections and can remain lodged in internal organs and cause irreversible lesions and haemato-immunological alterations, therefore compromising physiological functions vital for the health of fish without revealing external clinical signs in the animals.

Histological effects on the kidney, spleen, and liver of Nile tilapia Oreochromis niloticus fed different concentrations of probiotic Lactobacillus plantarum.

The aims of this study were to evaluate the inclusion of different concentrations of Lactobacillus plantarum in Nile tilapia diet and to verify histological effects on tissues of the animal organs, as well as to verify its effects on growth parameters and possible increase in the immune system. A total of 240 juveniles were distributed in 16 tanks arranged in a recirculation system. One control group and three treated groups (104, 106, and 108 colony-forming unit (CFU) g -1L. plantarum kg feed-1) were established in quadruplicate. After 35 days of supplementation, it was not possible to observe differences in growth rates and hematological parameters. However, in the kidney, there was a reduction in the presence of PAS-positive granular leukocytes (PAS-GL) between the collections (15 and 35 days). The liver had lower number of lesions related to loss of cordonal aspects of fish fed 108 CFU g-1 on day 15. Fish fed 104 and 108 CFU g-1 showed lower degree of congestion at day 35. The probiotic also provided a reduction in the number of melanomacrophage centers in the splenic tissue and an increase in the amount of goblet cells in the gut. The concentration 108 CFU g-1 of probiotic in diets increased the number of goblet cells, improved cordonal aspects, and reduced hepatic congestion, and indicated a possible improvement in the immunophysiological conditions of the fish.

Sylimarin as hepatic protector and immunomodulator in Nile tilapia during Streptococcus agalactiae infection.

This study investigated the use of silymarin, an extract obtained from the milk thistle (Silybum marianum) and its effects as a possible hepatoprotector in Nile tilapia (Oreochromis niloticus). Silymarin was used as feed additive to the diet at a concentration of 0.1% (1 kg per ton of dry ration) with the commercial product named Di-Heptarine S® (16% silymarin phosphatide). A total of 90 juvenile tilapia with approximately 45 days old and mean weight of 0.72 ±â€¯0.04 g were distributed in two groups, one fed with a diet with the hepatoprotector and the other without the additive. At the end of the assay (55 days after feeding), samples of blood were collected for hematological, immunological, histological (liver, spleen and intestine) and enzymatic analysis such as superoxide dismutase (SOD), catalase (CAT) and glutathione S-transferase (GST). After 55 days all fish were challenged with Streptococcus agalactiae serotype Ib to verify the sylimarin effects on the immunological parameters and its protection effect while challenged. During the challenge period another biological material sample was collected for hematological, immunological and histopathological analysis (liver, spleen and intestine). Before the challenge, an increase on the count of thrombocyte was found in the supplemented fish. In the liver, dilation of the sinusoids was observed in unsupplemented fish while supplemented fish the alteration was less severe. No significant alteration was found in SOD, CAT and GST between the groups. Histological changes after the challenge were provoked by bacterial toxins as a result of inflammatory processes. Periacinar degeneration was less intense in unsupplemented fish when compared to supplemented fish. On the other hand, eosinophilic and lymphocytic infiltrate did occur in unsupplemented fish differently from supplemented fish which did not show the alteration. The survival was 28% higher in silymarin supplemented fish when compared to unsupplemented fish that presented no survival. Silymarin supplementation in the diet provided a hepatoprotective and immunomodulatory effect on Nile tilapia.