Oreochromis Niloticus Research Articles

A biofloc system avoids the adverse effects of diets with suboptimal protein levels on zootechnical performance, intestinal histomorphometry, and protein metabolism of Nile tilapia juvenile fed Spirulina biomass (Arthrospira platensis) as an alternative protein source.

This study aimed to evaluate the effect of the biofloc technology (BFT) system and the replacement of fish meal with Spirulina biomass on productive performance, intestinal histomorphometry, plasma biochemistry, and oxidative stress of Nile tilapia juveniles (Oreochromis niloticus) fed suboptimal levels of protein. Two factors were evaluated: production systems (clear water × BFT) and replacement of fish meal with Spirulina (0, 33, 66 e 100%). The design was in a 2 × 4 randomized factorial scheme with four replications, and the fish were evaluated for 48days. Four isoproteic (28% crude protein) diets were formulated with gross energy values close to 4300kcalkg-1. Nile tilapia juveniles (0.23 ± 0.01g) were distributed in 16 circular tanks (70 L) at seven fish/tank. The diets were formulated with protein levels approximately 20% below that required for the species and life stage. No interaction was observed between the factors evaluated (production systems × Spirulina inclusion). Rearing the fish in the BFT system avoided the adverse effects of diets with suboptimal protein levels on performance, intestinal histomorphometry, and protein metabolism. Lower values lower lipid peroxidation and higher antioxidant capacity were observed in fish reared in the BFT system, showing evidence of improvements in antioxidant responses and lower levels of physiological oxidative stress. Spirulina completely replaced fish meal in the diets of Nile tilapia juveniles without adverse effects on intestinal morphometry, protein metabolism, and antioxidant response. Replacing 66% of fish meal with Spirulina improved the productive performance, regardless of the rearing system.

Report on Mortalities of Nile Tilapia (Oreochromis niloticus) Due to Tilapinevirus tilapiae and Co-infecting Bacteria in Winter Season

Background: Tilapinevirus tilapiae, commonly known as Tilapia lake virus, is a major viral infection of cultured tilapia, causing severe rates of fatalites and has evolved as a serious hazard to the tilapia in the world. Predisposing factors, notably temperature and the development stage of the host play a decisive role in influencing the extent of pathogenesis and the mortality rate associated with the disease. The seasonality of TiLV outbreaks has been observed in multiple countries, with a higher frequency occurring during the summer months. Methods: In the current investigation, the occurrence of TiLV disease was examined in fry and fingerlings of tilapia during the winter season (January - February, 2020) in Pune District, Maharashtra, India. The disease was observed in fish reared in open cages (Case I) and in nursery rearing farm (Case II). Infected tilapia displayed generalized gross clinical signs. TiLV was identified and confirmed in both instances using a semi-nested reverse transcriptase PCR by amplifying TiLV genome segment 3 and further confirmed by histopathogical investigation. Result: The research demonstrated the vulnerability of early developmemtal stages of O. niloticus to TiLV disease, suggesting the potential occurrence of this infection during the winter season, characterized by lower temperatures. Additionally, microbial investigations exposed the prevalence of Aeromonas spp., predominantly associated with TiLV-positive fish. These results align with previous publications, underscoring the potential role of bacterial infectious agents (Notably aeromonads), in exacerbating TiLV disease during outbreaks.

Integration of fruit in the formulation of feed for Oreochromis niloticus breeding in the region of Vakinankaratra, Madagascar

To promote the nutrition of tilapia in farming, four foods were created using traditional raw materials and containing different quantities of locally produced banana flour. These foods, which contain 36% crude proteins and 8% lipids, were tested on tilapia fry with an initial weight of 8 grams. After 50 days of the experiment, the final weights of the fish fluctuated between 33.28 grams and 41.4 grams, depending on the treatments used. Foods containing 3% and 6% banana flour showed the highest growth and food conversion rates, with respective specific growth (SGR) rates of 3.98 and 3.62, as well as conversion rates (CI) of 1.8 and 1.69. Meanwhile, the food with 10% banana flour obtained a SGR of 2.92 and a CI of 2.01, while the control obtained a SGR of 3.3 and CI of 1.7. Thanks to this test, we were able to observe the impact of bananas on the premature development of tilapia. We will continue to study the use of local agri-food byproducts and waste in feeding this fish.

Chlorella vulgaris algae ameliorates chlorpyrifos toxicity in Nile tilapia with special reference to antioxidant enzymes and Streptococcus agalactiae infection

BackgroundChlorpyrifos (CPF) is a widely used pesticide in the production of plant crops. Despite rapid CPF biodegradation, fish were exposed to wastewater containing detectable residues. Recently, medicinal plants and algae were intensively used in aquaculture to replace antibiotics and ameliorate stress impacts.Methods and resultsAn indoor experiment was conducted to evaluate the deleterious impacts of CPF pollution on Nile tilapia health and the potential mitigation role of Chlorella vulgaris algae. Firstly, the median lethal concentration LC50 − 72 h of CPF was determined to be 85.8 µg /L in Nile tilapia (35.6 ± 0.5 g body weight) at a water temperature of 27.5 °C. Secondly, fish were exposed to 10% of LC50 − 72 h for six weeks, and tissue samples were collected and examined every two weeks. Also, Nile tilapia were experimentally infected with Streptococcus agalactiae. Exposed fish were immunosuppressed expressed with a decrease in gene expressions of interleukin (IL) 1β, IL-10, and tumor necrosis factor (TNF)-α. Also, a decline was recorded in glutathione peroxidase (GPx), superoxide dismutase (SOD), and catalase (CAT) gene expression in the head kidney tissue. A high mortality rate (MR) of 100% was recorded in fish exposed to CPF for six weeks and challenged with S. agalactiae. Fish that received dietary C. vulgaris could restore gene expression cytokines and antioxidants compared to the control. After six weeks of CPF exposure, fish suffered from anemia as red blood cell count (RBCs), hemoglobin (Hb), and packed cell volume (PCV) significantly declined along with downregulation of serum total protein (TP), globulin (GLO), and albumin (ALB). Liver enzymes were significantly upregulated in fish exposed to CPF pollution, alanine aminotransferase (ALT) (42.5, 53.3, and 61.7 IU/L) and aspartate aminotransferase (AST) (30.1, 31.2, and 22.8) after 2, 4, and 6 weeks, respectively. On S. agalactiae challenge, high MR was recorded in Nile tilapia exposed to CPF (G3) 60%, 60%, and 100% in week 2, week 4, and week 6, and C. vulgaris provided a relative protection level (RPL) of 0, 14.29, and 20%, respectively.ConclusionsIt was concluded that CPF pollution induces immunosuppressed status, oxidative stress, and anemic signs in Nile tilapia. In contrast, C. vulgaris at a 50 g/kg fish feed dose could partially ameliorate such withdrawals, restoring normal physiological parameters.

Star1 gene mutation reveals the essentiality of 11-ketotestosterone and glucocorticoids for male fertility in Nile Tilapia (Oreochromis niloticus)

Steroidogenic acute regulatory protein (Star) plays an essential role in the biosynthesis of corticosteroids and sex steroids by mediating the transport of cholesterol from the outer to the inner membrane of mitochondria. Two duplicated Star genes, namely star1 and star2, have been identified in non-mammalian vertebrates. To investigate the roles of star genes in fish steriodogenesis, we generated two mutation lines of star1−/− and star1−/−/star2−/− in Nile tilapia (Oreochromis niloticus). Previous studies revealed that deficiency of star2 gene caused delayed spermatogenesis, sperm apoptosis and sterility in male tilapia. Our present data revealed that mutation of star genes impaired male fertility. Disordered seminiferous lobules and spermatic duct obstruction were found in the testis of both types of mutants. Moreover, significant decline in semen volume, sperm abnormality and impaired fertility were also detected in star1−/− and star1−/−/star2−/− males. In star1−/− male fish, lipid accumulation, up-regulation of steroidogenic enzymes, and significant decline of androgens were found. Additionally, hyperplasic interrenal cells, elevated steroidogenic gene expression level and decline of serum glucocorticoids were detected in star1 mutants. Intriguingly, either 11-KT or cortisol supplementation successfully rescued the impaired fertility of the star1−/− mutants. Taken together, these results further indicate that Star1 might play critical roles in the production of both 11-KT and glucocorticoids, which are indispensable for the maintenance of male fertility in fish.

Hypogonadotropic hypogonadism in male tilapia lacking a functional rln3b gene

Relaxin 3 is a neuropeptide that plays a crucial role in reproductive functions of mammals. Previous studies have confirmed that rln3a plays an important role in the male reproduction of tilapia. To further understand the significance of its paralogous gene rln3b in male fertility, we generated a homozygous mutant line of rln3b in Nile tilapia. Our findings indicated that rln3b mutation delayed spermatogenesis and led to abnormal testes structure. Knocking out rln3b gene resulted in a decrease in sperm count, sperm motility and male fish fertility. TUNEL detection revealed a small amount of apoptosis in the testes of rln3b−/− male fish at 390 days after hatching (dah). RT-qPCR analysis demonstrated that mutation of rln3b gene caused a significant downregulation of steroid synthesis-related genes such as cyp17a1, cyp11b2, germ cell marker gene, Vasa, and gonadal somatic cell marker genes of amh and amhr2. Furthermore, we found a significant down-regulation of hypothalamic-pituitary-gonadal (HPG) axis-related genes, while a significantly up-regulation of the dopamine synthetase gene in the rln3b−/− male fish. Taken together, our data strongly suggested that Rln3b played a crucial role in the fertility of XY tilapia by regulating HPG axis genes.

Arsenic bioaccumulation and biotransformation in different tissues of Nile tilapia (Oreochromis niloticus): A comparative study between As(III) and As(V) exposure and evaluation of antagonistic effects of selenium

The speciation of arsenic in fish has been widely investigated, but bioaccumulation and biotransformation of inorganic As in different tissues of Nile tilapia (Oreochromis niloticus) are not fully understood. The present study aimed to investigate the bioaccumulation of As in Nile tilapia, as well as to evaluate the distribution of the main arsenic species (As(III), As(V), MMA, DMA, and AsB) in liver, stomach, gill, and muscle, after controlled exposures to As(III) and As(V) at concentrations of 5.0 and 10.0 mg L−1 during periods of 1 and 7 days. Total As was determined by inductively coupled plasma mass spectroscopy (ICP-MS). For both exposures (As(III) and As(V)), the total As levels after 7-day exposure were highest in the liver and lowest in the muscle. Overall, the Nile tilapia exposed to As(III) showed higher tissue levels of As after the treatments, compared to As(V) exposure. Speciation of arsenic present in the tissues employed liquid chromatography coupled to ICP-MS (LC-ICP-MS), revealing that the biotransformation of As included As(V) reduction to As(III), methylation to monomethylarsonic acid (MMA) and dimethylarsinic acid (DMA), and subsequent conversion to nontoxic arsenobetaine (AsB), which was the predominant arsenic form. Finally, the interactions and antagonistic effects of selenium in the bioaccumulation processes were tested by the combined exposure to As(III), the most toxic species of As, together with tetravalent selenium (Se(IV)). The results indicated a 4–6 times reduction of arsenic toxicity in the tilapia.

Utilizing outer membrane proteins (OMPs) as a vaccine to manage motile Aeromonas septicemia (MAS) in Nile tilapia (Oreochromis niloticus) with evaluating the vaccine immunoprotective effectiveness

AbstractMotile aeromonads are an opportunistic and heterogenic bacteria that cause motile Aeromonas septicemia (MAS) which is accompanied with variety of clinical signs and lesions in fish. It has been found that their outer membrane proteins (OMPs) are highly immunogenic and may be used as a vaccine candidate. The present study was undertaken to evaluate OMP protection against Aeromonas hydrophila (A. hydrophila) strain ATCC 7966 (A5 and A8) and A. hydrophila strain DSM 30,187 (A6). Pathogenicity tests revealed that out of 12 isolated strains, seven were highly pathogenic, three were moderate, and two were avirulent. SDS-PAGE analysis of OMPs showed several protein bands (7–22) of different molecular weights ranging from 14 to 346 kDa. The common protein band was 39 kDa. It seems that pathogenicity and the number of protein bands are related in the highly pathogenic strains. Challenged A. hydrophila for Nile tilapia (O. niloticus) previously actively immunized with OMPs (A4, A5, A6, A8, and A11) showed a significant increase in relative percent survival (RPS) ranging from 50 to 100%. Immunological parameters such as immunoglobulin M (IgM), antioxidants (SOD, GSH, and CAT), and nitric oxide (NO) and expression of pro-inflammatory cytokines, interleukin 1β (IL-1β), and tumor necrosis factor-α (TNFα) showed a significant increase in OMP-immunized fish. Challenged A. hydrophila for O. niloticus previously passively immunized with OMPs (A4, A5, A6, A8, and A11) in fish-raised sera showed a significant increase in RPS (20 to 100%) in the vaccinated groups. It is concluded that OMPs from Aeromonas spp. have been identified as suitable target for protection development, especially in the case of heterogenic bacteria.

Physiological Changes in Nickel-exposed Nile Tilapia Oreochromis niloticus during Exposure and Recovery Periods

Heavy metals are a significant problem in aquatic ecosystems as they are toxic and tend to accumulate, immediately affecting fish physiology. The present investigation was carried out to evaluate the sub-lethality of Nickel chloride on fingerlings of Nile tilapia, Oreochromis niloticus on long-term exposure to it is below safe concentrations1/fifth (9.39 ppm) and 1/10th (4.69 ppm). The physiological studies revealed a significant reduction in Oxygen Consumption Rate (OCR), Ammonia-N Excretion Rate (AER), Oxygen Rate (O: R) and Food Consumption Rate (FCR) and behavioural changes during accumulation and depuration phases. Limited recovery was obtained in all the physiological parameters after depuration for 28 days from the end of the accumulation period.

Database construction and comparative genomics analysis of genes involved in nutritional metabolic diseases in fish

Nutritional metabolic diseases in fish frequently arise in the setting of intensive aquaculture. The etiology and pathogenesis of these conditions involve energy metabolic disorders influenced by both internal genetic factors and external environmental conditions. The exploration of genes associated with nutritional and metabolic disorder has sparked considerable interest within both the aquaculture scientific community and the industry. High-throughput sequencing technology offers researchers extensive genetic information. Effectively mining, analyzing, and securely storing this data is crucial, especially for advancing disease prevention and treatment strategies. Presently, the exploration and application of gene databases concerning nutritional and metabolic disorders in fish are at a nascent stag. Therefore, this study focused on the model organism zebrafish and five primary economic fish species as the subjects of investigation. Using information from KEGG, OMIM, and existing literature, a novel gene database associated with nutritional metabolic diseases in fish was meticulously constructed. This database encompassed 4583 genes for Danio rerio, 6287 for Cyprinus carpio, 3289 for Takifugu rubripes, 3548 for Larimichthys crocea, 3816 for Oreochromis niloticus, and 5708 for Oncorhynchus mykiss. Through a comparative systems biology approach, we discerned a relatively high conservation of genes linked to nutritional metabolic diseases across these fish species, with over 54.9 % of genes being conserved throughout all six species. Additionally, the analysis pinpointed the existence of 13 species-specific genes within the genomes of large yellow croaker, tilapia, and rainbow trout. These genes exhibit the potential to serve as novel candidate targets for addressing nutritional metabolic diseases.

Urbanization effects on growth and otolith asymmetry in Chrysichthys nigrodigitatus and Oreochromis niloticus within tropical coastal lagoon watersheds

In this study, we investigated the growth dynamics and otolith shape asymmetry of two fish species, Chrysichthys nigrodigitatus (CN) and Oreochromis niloticus (ON), within urbanized watersheds of the southern lagoon system, Nigeria. Using the von Bertalanffy growth model (VBGM), in addition to sediment metal concentration indices such as the average shale content, index of geoaccumulation (Igeo), contamination factor (CF), pollution load index (PLI), and potential ecological risk (PER) index, contamination levels were classified, and ecological risks were assessed. Notably, a lower growth potential (t0) was observed in CN at Ikorodu than at Epe, with similar trends for ON in the Epe during the dry season. Otolith asymmetry patterns, particularly in the CN at Ikorodu and ON in the Epe during the dry season, exhibited distinct ecological variations, indicating heightened stress levels at Ikorodu. Sediment analyses revealed moderate to strong contamination (Cd, Pb, Ni, and Cr) in both Lagos Lagoon (Ikorodu) and Epe Lagoon, with Ikorodu exhibiting notably high to moderate contamination levels according to the CF index. Elevated PLI values for Cd and Pb in Ikorodu, in addition to greater PER, indicated increased risk, with Cd posing a high risk (61.42%) and Pb posing a moderate risk (49.50%). Additionally, the reduced asymptotic length in the Epe during the dry season suggests that Chrysichthys nigrodigitatus is adaptable to seasonal variations, while divergent growth patterns in both areas indicate the existence of trade-off mechanisms in response to changing conditions. Habitat-specific otolith asymmetry and metal contamination underscore species adaptability, with wider stressor variability in Lagos than in Epe. Furthermore, multidimensional scaling analysis highlights the intricate relationship between otolith shape variables and environmental factors, emphasizing the need for tailored conservation efforts in urbanized watersheds.

Molecular characterization and bivalent vaccine development of Aeromonas hydrophila and Aeromonas veronii in Nile tilapia (Oreochromis niloticus)

In the present study, nine strains of Aeromonas spp. were isolated from diseased Nile tilapia in Thailand and identified based on 16S rRNA and gyrB gene sequences and multilocus sequence typing (MLST) analysis. Phenotypic characterization was performed for all the isolates. For the antimicrobial susceptibility test, all isolates were resistant to amoxicillin, whereas most of the isolates were susceptible to enrofloxacin. Pathogenicity analysis revealed that most A. hydrophila strains were highly virulent to fish, corresponding with the hemolytic activity test demonstrating that clear hemolysis occurred on sheep blood agar. To develop a vaccine for fish against A. veronii and A. hydrophila infection, a bivalent vaccine based on the formalin-killed cells (FKC) of A. veronii A5 and A. hydrophila A9, which showed higher virulence to fish, was prepared. This FKC vaccine was administered to Nile tilapia by intraperitoneal injection. The booster vaccination was conducted at 4 weeks after the first vaccination. The specific antibody titer was evaluated during rearing since the first vaccination, and then at 8 weeks after the first vaccination, the fish were challenged with A. veronii A5 and A. hydrophila A9 via intraperitoneal injection. There is not significance different in relative percentage survival of the vaccinated fish against A. veronii (63.49 ± 19.51%) and A. hydrophila (93.33 ± 6.67%). These results suggest that this candidate bivalent vaccine is effective against both A. veronii and A. hydrophila infection in Nile tilapia and could be an “motile Aeromonas septicemia (MAS)” control measure. However, to obtain the promising vaccine efficacy, the optimization of vaccine formulation might be achieved and be validated vaccine efficacy in field trial.

Poultry manure fertilization of Egyptian aquaculture ponds brings more cons than pros

Aquaculture is a crucial sector for Egyptian food production, providing a cheap source of animal protein while securing income and employment for a substantial part Egypt's population. Nile tilapia (Oreochromis niloticus) is the most commonly produced fish, usually farmed in earthen ponds around the Northern Delta Lakes. A common practice among farms is to fertilize ponds with chicken manure (CM) in order to increase nutrient levels and promote phytoplankton, consumed by the fish. However, with reports of use of antibiotics in Egypt's poultry sector, and that CM contains residues of antibiotics, antibiotic resistant pathogens and antibiotic resistance genes (ARGs) are production benefits large enough to compensate a potential health hazard?Using production data from 501 aquaculture farms and fish pond sediment from 28 ponds we evaluated potential benefits in yields and profitability for farms using CM for fertilization, and used qPCRs to screen sediments for three antibiotic resistance genes coding for resistance to the most commonly used antibiotics in the poultry sector. The analysis showed no significant benefits to fish yields or profitability in farms where CM was applied, but a risk of significantly increased nutrient loads. Meanwhile, we detected increased abundances of tetA and tetW resistance genes in fish pond sediment where CM was applied. With the risk of disseminating ARGs and causing eutrophication of local waterways, we recommend that Egyptian tilapia pond farmers refrain from using CM and adopt best management practices for increasing farm profitability in order to to reduce environmental and health hazards.

Acid-base disturbances and effects on oxygen uptake rates in Nile tilapia (Oreochromis niloticus) following acute and prolonged CO2 exposure

High levels of dissolved carbon dioxide (CO2) occur nightly in earthen ponds characterized by high respiration rates. Exposure to high CO2 conditions (hypercapnia) leads to acidosis in fish, which can be compensated by an accumulation of HCO3− to recover intra- and extracellular pH levels, with a capacity that appears to be species-specific. For Nile tilapia, a freshwater tropical teleost traditionally produced in earthen ponds, little information is available on the tolerance to dissolved levels of CO2 and associated acid-base disturbances. Here, we investigated first the effects of acute and progressively increasing CO2, from normocapnic conditions to 60 mg CO2 L−1, on oxygen uptake rates (MO2). This was followed by exposure to three concentrations of CO2; 10, 30, and 60 mg L−1 (equivalent to pCO2 of 5.4, 16.2, and 32.4 mmHg) against a normocapnic control (pCO2 0.3 mmHg), to investigate acute (1 h) or prolonged (24 h) effects on standard (SMR) and maximum metabolic rates (MMR), haematology, and extra- and intracellular acid-base status in adult Nile tilapia (mean BM 435 ± 16 g ± SE). Acute exposure to hypercapnia led to concentration-dependent decreases in both SMR and MMR. Fish were able to fully or partially recover MMR and metabolic scope (MS) after 24 h, while depression of SMR persisted at all CO2 levels. Acute exposure to CO2 caused intra- and extracellular pH levels to decrease by up to 0.5 units in a concentration-dependent manner. Only the lowest hypercapnic treatment (pCO2 5.4 mmHg) was able to fully recover within 24 h. Changes in haematological variables appeared minor, being restricted to increasing haematocrit, haemoglobin concentration, and mean cell volume in the highest CO2 treatments after 24 h exposure. Although the Nile tilapia is generally considered a species able to tolerate poor water quality, the modest or slow acid-base regulation following hypercapnic exposure suggests sensitivity to hypercapnia.

Evaluation of the effects of dietary cinnamaldehyde on growth and nutrient use in Nile Tilapia (Oreochromis niloticus)

Cinnamaldehyde is an active organic compound and an aromatic aldehyde substance found in cinnamon oil. It is a new feed supplement for animals that can improve growth, nutrient digestion and absorption, lipid use, and immunity. The present study evaluated the effects of dietary trans-cinnamaldehyde 98 % (CIN) on nutrient utilization, antioxidant capacity, and growth performance in Nile tilapia (Oreochromis niloticus). Juveniles used was weighing at about 19.77 ± 0.10 g received dietary CIN at different doses (0, 0.25, 0.5, 0.75, and 1.0 g/kg feed). The fish were maintained in a glass aquarium of 95 × 45 × 35 cm3 in size at a stocking density of 20 fish per aquarium and fed three times a day to clear satiation. Dietary carbohydrate digestion and absorption were significantly improved in fish fed 0.75 g/kg CIN, as indicated by increased amylase activity and glucose tolerance test. The latter was subsequently confirmed by increases in relative ir, glut4, hx and gs expression that indicated the increase in glucose absorption led to an increase in glucose utilization as an energy source and the synthesis of glycogen as an energy reserve. Interestingly, dietary CIN supplementation also resulted in the downregulation of fas, which facilitates the conversion of glucose to fatty acids, and the upregulation of cpt1a and hsl, indicating increased fatty acid oxidation for energy. CIN at a range of 0.5–0.75 g/kg improved the antioxidant status in the liver, as illustrated by elevated glutathione peroxidase and superoxide dismutase activities (both P < 0.05). Final weight, feed consumption, the specific growth rate, feed efficiency, and protein retention increased were significantly in all CIN groups, with the highest values reached in the 0.5 g/kg CIN (all P < 0.05). Based on a polynomial orthogonal analysis of the fish-specific growth rate, the optimum dosage of CIN inclusion is 0.42 g/kg.

Analysing multi-class pesticide residues in tilapia fish: development and validation of LC-MS/MS and GC-MS/MS methods

ABSTRACT Pesticides are an integral part of increasing food production and have remarkable agricultural benefits such as an increase in crop yield. However, they have been identified as some of the most hazardous substances in food, particularly in fish, posing a risk to both ecosystems and human health. Therefore, this study focuses on the development and optimisation of an analytical technique to detect multiple pesticide residues in fish products. The method involves liquid-liquid extraction, followed by analysis using both liquid and gas chromatography coupled with mass spectrometry. A total of 411 pesticides were selected (e.g.: insecticides, fungicides, and herbicides) and the concentrations of their residues in tilapia (Oreochromis niloticus) were determined. The samples were processed according to the optimised QuEChERS (quick, easy, cheap, effective, rugged, and safe) method, including extraction and clean-up procedures. Lipid co-extracts were successfully minimised before instrumental analysis using optimised cleanup. Matrix-matched calibrations were performed at two different levels (0.05,0.01 mg/kg) in tilapia fish to compensate for signal enhancement/suppression. The validation procedure was performed to assess accuracy, precision, limit of quantification (LOQ), and uncertainty according to the SANTE guidelines. The LOQs of this method were 0.01 mg/kg for all selected pesticides except Cyazofamid, Dodine, Fenpyrazamine, Methomyl, Oxadiargyl, Parathion ethyl, Parathion methyl, Prothioconazole Desthio, and Tolylfluanid were 0.05 mg/kg. The suggested method was implemented to investigate the levels of pesticide residues in fresh fish samples (n = 15) procured from local markets in Egypt. Only two samples had residues of the targeted analytes. In light of our findings, the devised methodology was effectively utilised for the detecting and measurement of pesticide residues present in fish samples.

Use of Bacillus subtilis multiplicate in the water used for biofloc formation: Growth, hemato-biochemistry, intestinal bacteria colonies, and bacterial resistance evaluations of Nile tilapia

Probiotics in the biofloc technology (BFT) system improve fish growth and immune response and promote a favorable intestinal microbiota balance. This study aimed to investigate the growth, hemato-biochemical parameters, bacterial resistance, and the number of intestinal bacteria colonies of Nile tilapia (6.23 g) reared in the BFT system with graded levels of Bacillus subtilis multiplicate (BSM) in the water. The multiplication of these microorganisms was performed in a biofactory. Fish were distributed into four different groups with graded BSM concentrations in the water of 0.00, 0.25, 0.50, and 0.75% (or 0; 750; 1500; and 2250 mL; called BSM0 (control), BSM25, BSM50, and BSM75, respectively). The experimental procedure had 45 days for growth performance evaluations and seven additional days of bacterial challenge with Aeromonas hydrophila. The experimental design was randomized into four treatments in triplicate (n = 20 fish per tank; initial stocking density of 0.50 kg m−3). Survival was 100% before and after the bacterial challenge. Increasing BSM in the BFT system had a positive linear effect on nitrite, nitrate, settled solids, hemoglobin, and hematimetric indices. BSM50 showed the highest specific growth rate, weight gain, intestinal total bacteria, and B. subtilis counts and the lowest feed conversion rate. In BSM50 and BSM75, liver glycogen levels and aspartate aminotransferase (AST) activity were higher and lower, respectively, than in the other treatments. Plasma total cholesterol, total proteins, albumin, and skin mucus total proteins and albumin were highest in BSM25. After the bacterial challenge, the number of lesions was lower in BSM75 than in the control group. In conclusion, we recommended BSM50 because it increased the growth performance and bacterial colonization in the intestine. It also decreased liver AST activity in Nile tilapia reared in the BFT system without compromising water quality and hematological and plasmatic parameters.

A multiplex PCR assay to detect mislabelling in fish products

ABSTRACT Fish substitution in fish products is an important issue in fish markets, as it is a widespread practice. An authentication protocol for Rohu, Thaila and Tilapia was developed by multiplex PCR. Three species-specific and one degenerate common forward primer were designed using the Cytb gene of the mitochondrial genome. These primers for Labeo rohita, Labeo catla and Oreochromis niloticus showed the fragment size of 235 bp, 186 bp and 506 bp on the agarose gel, respectively. The primers for L. rohita and L. catla were sensitive to 0.1 ng of DNA template, while for O. niloticus this value was 1 ng of DNA template. A total of 230 commercial samples (160 fried and 70 processed fish products) were screened, where 60% mislabeling in fried and 30% mislabeling in processed fish were found. This multiplex PCR protocol could give useful insights for food inspection and enforcement of regulatory food control.

The cellular expression patterns of gdnfa and gdnfb in the gonads of Nile tilapia and their differential response to retinoic acid

In mammals, glial cell derived neurotrophic factor (GDNF) plays a critical role in the self-renewal and maintenance of spermatogonial stem cells (SSCs) in testis and oogenesis in ovary, whilst retinoic acid (RA), the key factor of meiosis initiation, can downregulate its expression. Unlike mammals, two Gdnf replication genes are widely present in teleost fishes, however, our understanding of them is still poor. In the present study, two paralogous gdnf from Nile tilapia (Oreochromis niloticus), namely as Ongdnfa and Ongdnfb, were characterized, and then their cellular expression profiles in testis and ovary and responsiveness to RA treatment at the tissue and cellular levels were investigated. In phylogenetic tree, the Gdnfa and Gdnfb from teleost fishes were clustered into two different subclasses, respectively, and then clustered with the homologs from cartilaginous fish and tetrapods, suggesting that OnGdnfa and OnGdnfb are orthologous to GDNF and paralogous to each other. Ongdnfa is expressed in Sertoli cells and Leydig cells in testis and oocytes in ovary. The expression pattern of Ongdnfb is similar to Ongdnfa. In the ex vivo testicular organ culture, RA down-regulated the expression of Ongdnfa, whereas up-regulated the expression of Ongdnfb (P < 0.05), suggesting that they have differential responsiveness to RA signaling. RA treatment of the cultured cells derived from adult Nile tilapia testis which have the expression of RA receptors (RAR), Ongdnfa and Ongdnfb further confirmed the above result. Collectively, our study suggests that Ongdnfa and Ongdnfb have non-germline expression patterns in testis and germline expression patterns in ovary; furthermore, they have differential responsiveness to RA signaling, implying that they might have differential biological functions. This study broadens and enriches our understanding of fish GDNF homologs and lays foundation for the study of their biological functions in the future.

Hyperosmotic Stress Induces Inflammation and Excessive Th17 Response to Blunt T-Cell Immunity in Tilapia.

Despite the advances in study on osmotic physiology in bony fish, the mechanism by which the immune system, especially T-cell immunity, adapts and responds to osmotic stress remains unknown. In the current study, we investigated the response of T cells to hyperosmotic stress in the bony fish Nile tilapia (Oreochromis niloticus). As a euryhaline fish, tilapia was able to adapt to a wide range of salinities; however, hypertonic stress caused inflammation and excessive T-cell activation. Furthermore, hypertonic stress increased the expression of IL-17A in T cells, upregulated the transcription factor RORα, and activated STAT3 signaling, along with IL-6- and TGF-β1-mediated pathways, revealing an enhanced Th17 response in this early vertebrate. These hypertonic stress-induced events collectively resulted in an impaired antibacterial immune response in tilapia. Hypertonic stress elevated the intracellular ROS level, which in turn activated the p38-MK2 signaling pathway to promote IL-17A production by T cells. Both ROS elimination and the p38-MK2 axis blockade diminished the increased IL-17A production in T cells under hypertonic conditions. Moreover, the produced proinflammatory cytokines further amplified the hypertonic stress signaling via the MKK6-p38-MK2 axis-mediated positive feedback loop. To our knowledge, these findings represent the first description of the mechanism by which T-cell immunity responds to hypertonic stress in early vertebrates, thus providing a novel perspective for understanding the adaptive evolution of T cells under environmental stress.