Open Veterinary Journal, (2026), Vol. 16(6): 3966-3985

Research Article

10.5455/OVJ.2026.v16.i6.65

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Efficacy of biological products in mitigating clinical signs, T-2 mycotoxicosis, and vibriosis in gilthead seabream (Sparus aurata L.)

Ahmed Said Al-Souti¹, Mahmoud A. Elnakeeb^2*, Ahmed A. El-bahlol², Mohamed M. Elnawsany²,
Saad M. Alsaiad², Mohamed E. Abou El Atta³ and Tamer Mohammed Monir Abdelrahiem³

¹Department of Marine Science and Fisheries, College of Agricultural and Marine Sciences, Sultan Qaboos University, Al-Khoud, Sultanate of Oman

²Fish Production Department, Faculty of Agriculture, Al-Azhar University, Nasr City, Egypt

³Department of Fish Health and Management, Central Laboratory for Aquaculture Research, Agricultural Research Center, Abbassa, Egypt

*Corresponding Author: Mahmoud A. Elnakeeb. Assistant Professor, Fish Production Department, Faculty of Agriculture, Al-Azhar University, Cairo, Egypt. Email: mahmoud.biotech [at] azhar.edu.eg; mahmoud.biotech [at] gmail.com

Submitted: 16/03/2026 Revised: 25/05/2026 Accepted: 07/06/2026 Published: 30/06/2026

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Abstract

Background: T-2 toxin, a type A trichothecene, exhibits hepatotoxic, immunosuppressive, and oxidative effects in fish.

Aim: This study examines the protective effects of Mycofix Plus and Mycosorb A+ commercial mycotoxin binders (MBs) against T-2-induced toxicity in gilthead seabream.

Methods: In a 6-week feeding trial, 180 fingerlings (initial weight 100 g) were allocated into 6 groups in triplicate. The dietary groups consisted of a control, binder-only diets, a T-2 toxin-challenged group, and contaminated diets supplemented with either binder (1 g kg⁻¹). Measurements included clinical manifestations, innate immunity, oxidative status, enzymatic activity, histological changes, and survival following a V. harveyi challenge.

Results: T-2 toxin-exposed fish exhibited severe clinical signs, organ damage, and immunosuppression of the innate immune response compared with control fish (p < 0.05). These changes were accompanied by a significant increase in cortisol and oxidative stress indicators (glutathione reductase and lipid peroxidase). In the disease-resistance trial, the T-2 group showed the highest susceptibility to V. harveyi (mortality: 90%). However, the inclusion of Mycofix Plus and Mycosorb A+ in the diets significantly counteracted the impact of the toxin, improving the antioxidant status and reducing clinical symptoms. Notably, these treatments restored survival rates to 70%–80% and provided substantial relative protection (57.14%–71.43%) compared to fish receiving the T-2 toxin alone.

Conclusion: Mycofix^® Plus and Mycosorb^® A+ are MBs used as functional feed additives in gilthead seabream to reduce T-2 toxin toxicity and enhance disease resistance to V. harveyi.

Keywords: Gilthead seabream, Immunosuppression, Mycotoxin binders, T-2 toxin, Vibrio harveyi.

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Introduction

Mariculture in the northern part of Egypt, especially in the Mediterranean area and around the Suez Canal, is increasingly contributing to fish production in the country (Paruğ et al., 2024). Among the marine species cultivated, the gilthead sea bream (Sparus aurata) is one of the most valuable species (Eissa et al., 2025) in terms of economic benefits due to its high market demand and ability to thrive in intensive culture systems (Mhalhel et al., 2023). Although mariculture in Egypt remains less developed than freshwater aquaculture, it has shown steady expansion in response to the scarcity of freshwater and the need to sustainably exploit marine resources (Shaalan et al., 2018).

Despite this growth, the intensification of sea bream farming is constrained by several environmental and nutritional stressors that adversely affect fish health, growth performance, and survival (Rennie et al., 2019; Dara et al., 2023; Taylor et al., 2025). Contamination of aquafeeds with mycotoxins represents a major and often underestimated risk (Pulvirenti et al., 2024; Gruber-Dorninger et al., 2025; Lorusso et al., 2025). Trichothecenes, a large group of mycotoxins produced by Fusarium species, are of concern in aquaculture due to their known toxicity and common occurrence in plant-based ingredients of aquafeeds (Vulić et al., 2025).

T-2 toxin is one of the most toxic mycotoxins to animals, including fish (Meneely et al., 2023). It is described as a highly lipophilic molecule that dissolves in nonpolar solvents, thereby enhancing its bioavailability (Zhang et al., 2022). There has been a surge in the formulation of fish feeds with plant-based compounds, leading to T-2 toxin contamination in aquafeeds due to their susceptibility to fungal contamination during cultivation, storage, and processing (Lorusso et al., 2025). Previous studies have shown that aquaculture feeds are prone to contamination with mycotoxins, thereby reducing feed efficiency, nutrient utilization, and growth performance, as well as affecting vital organs, especially the liver and kidneys (Gruber-Dorninger et al., 2025).

T-2 toxin has been well documented at a concentration of 500 μg kg⁻¹ as a hepatotoxin and hepatocarcinogen in a variety of animal species, including fish, with the liver being the principal target organ (Qiu et al., 2016; Yin et al., 2020; Kihal et al., 2022). Toxins trigger pathological changes, impair liver function, and cause significant changes in liver enzyme activity, indicating impaired metabolic function (Abdelrahiem et al., 2025). T-2 toxin has also been reported to induce immunosuppression and oxidative stress and to increase the risk of disease in fish, posing a major threat to fish welfare and mariculture sustainability (Kihal et al., 2022; Goda et al., 2025). In light of the emerging issues of mycotoxin contamination of aquafeeds in marine aquaculture, there is a pressing need to develop strategies to mitigate the effects of mycotoxins in fish, which have become a major concern for both researchers and the aquafeed industry (Agriopoulou et al., 2020).

Mycotoxin binders (MBs) reduce toxicity by adsorbing mycotoxins with sufficient affinity to prevent their interaction with animal tissues and to limit their absorption across the gastrointestinal tract (Puvača et al., 2023). Numerous adsorbent materials have been tested, including activated carbon, aluminosilicates, which cover a wide variety of materials from clay and bentonite to zeolite and other phyllosilicates, complex indigestible carbohydrates, which cover cellulose and polysaccharides derived from yeast and bacterial cell walls, and cholestyramine, polyvinylpyrrolidone, and their derivatives (Arafa et al., 2025). Several commercial mycotoxin-binding products, such as Mycofix Plus, Mycosorb A+, Agresol, and CAP T2, have been investigated for their efficacy in aquafeeds (Puvača et al., 2024; Anandaraj et al., 2025; Zahran et al., 2026). However, the binding ability of these products varies widely depending on the type of binder used, its source, level of inclusion, and the target mycotoxin’s chemical structure (Kihal et al., 2022). Some of these products may also negatively influence growth performance due to a lack of specificity in binding mycotoxins because they can also bind other nutrients such as vitamins and minerals (Hussain et al., 2024).

The broad efficacy of commercial mycotoxin-binding agents in aquaculture feeds has been demonstrated. For instance, Mycofix, a commercial formulation based on clay minerals and specific enzymatic components, reduced mycotoxin bioavailability by more than 70% when included at a low dietary level of 1 g kg⁻¹ (Chefchaou et al., 2019). Several studies have confirmed the effectiveness of Mycofix and similar binders across aquaculture species, indicating their general applicability rather than species-specific action (Koletsi et al., 2021). In parallel, Mycosorb A+ has shown considerable promise as an alternative adsorbent; dietary inclusion at 0.2%–0.4% in aflatoxin T2-contaminated diets resulted in improved hepatic function, immune responses, toxin residues, and intestinal microbial profiles comparable to those of uncontaminated controls (Kihal et al., 2022). These outcomes support the effectiveness of both mineral-based binders and biological adsorbents in mitigating mycotoxin-induced toxicity in aquafeed formulations (Agriopoulou et al., 2020).

Disease outbreaks, particularly vibriosis induced by Gram-negative Vibrio spp., represent a primary impediment to the advancement of global mariculture (Mohamad et al., 2019). Vibrio harveyi is a prominent pathogen that affects numerous teleost species, including S. aurata, often resulting in systemic septicemia and mass mortality (Triga et al., 2024). Consequently, V. harveyi challenge trials are widely employed to evaluate disease resistance and immunological competence in fish subjected to various physiological stressors (Serna-Duque et al., 2023). Despite extensive research on MBs in terrestrial livestock and, to a lesser extent, freshwater fish species, their functional efficacy under concurrent toxicological and infectious stressors in marine aquaculture systems remains poorly understood. Studies integrating chronic dietary exposure to T-2 toxin with a subsequent pathogenic challenge are extremely limited, thereby constraining our understanding of the interaction between mycotoxicosis, immune suppression, and disease susceptibility. Furthermore, comparative evaluations of distinct commercial MBs under such dual-stress conditions are scarce, especially in economically important marine species, such as S. aurata. Thus, the current study combines dietary toxin exposure, physiological and histological analysis, and a controlled V. harveyi challenge to provide a complete picture of the host’s resilience. This study is based on the idea that supplementing the diet with Mycofix Plus and Mycosorb A+ may influence immune function and oxidative stress in different ways. These treatments could help reduce the negative effects of toxin exposure and improve the resistance of S. aurata to V. harveyi infection.

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Materials and Methods

Experimental site and the fish

The current study was conducted in a controlled indoor setup that simulates mariculture conditions using seawater sourced from a commercial fish farm located in Alexandria, Egypt. A cohort of 180 healthy S. aurata fingerlings, with an initial mean body weight of 100 ± 0.16 g, was used in this study. Fish were obtained from a single hatchery batch to ensure uniformity and minimize biological variability. Fingerlings were acclimated under laboratory conditions for 2 weeks before the start of the experiment. During the acclimation period, the fish were fed a basal diet at a rate of 3% body weight per day.

After acclimation, the fish were randomly distributed in glass aquaria measuring 70 × 35 × 50 cm, with 10 fish per aquarium. Continuous aeration was maintained using an electric air compressor. Partial water exchange (approximately one-third of the volume) was conducted daily to maintain the water quality within optimal levels. Water salinity was maintained at 32 ± 0.02 ppt under continuous aeration and partial daily water exchange, while water temperature ranged from 26 °C to 28 °C; dissolved oxygen remained above 5 mg l⁻¹.

Diets and incorporation of T-2 toxin

A basal diet was designed to meet the specific nutrient requirements of seabream fingerlings (NRC, 2011). The diet comprised 42% crude protein, 18% ether extract, 7% crude fiber, 6.5% ash, and 26.5% nitrogen-free extract. The main ingredients included fish meal, soybean meal, yellow maize, wheat bran, sunflower oil, and a vitamin premix. All ingredients were ground into a fine powder, thoroughly homogenized, and extruded into 1-mm diameter pellets using a feed manufacturing machine (Okolie et al., 2019). After extrusion, the pellets were air-dried at room temperature until a constant weight was achieved. The dried diets were stored in airtight containers under cool, dry conditions to prevent moisture uptake and minimize the risk of fungal growth or unintended mycotoxin contamination prior to feeding (Ilesanmi et al., 2024). T-2 toxin, at a concentration of 500 μg kg⁻¹, was incorporated into the toxin-contaminated diets according to the experimental protocols (Matejova et al., 2017). The toxin was made soluble in an appropriate solvent for its even distribution before being incorporated into the diets.

Two commercial MBs were used in this study: (1) Mycofix Plus and (2) Mycosorb A+. Both binders were incorporated into the experimental diets at an inclusion level of 1 g kg⁻¹ feed, either alone or in combination with T-2 toxin, as specified in the experimental design.

Experimental design

The study employed a randomized design with 6 dietary treatments, each in triplicate (Table 1, Fig. 1). Gilthead seabream were stocked at a rate of 10 fish per aquarium, with 3 aquariums per treatment. The fish were fed twice daily at 3% of their body weight for 6 weeks. The neutralizing capacity of Mycofix Plus and Mycosorb A+ in the presence of T-2 toxin was assessed. Feed rations were adjusted weekly according to weight measurements. Water quality parameters were rigorously monitored and maintained within the optimal ranges required for seabream cultivation throughout the experimental period.

Table 1. Experimental groups and S. aurata fingerling dietary treatments for 42 days.

Fig. 1. Experimental groups and dietary treatments of S. aurata fingerlings for 42 days.

Sampling

At the termination of the trial, all fish were sampled using a standard procedure following a 24-hours fasting period. The number of fish was estimated, their weight was measured, and any growth variation, clinical symptoms, and morphological changes caused by the diets were evaluated. Three fish per replication aquarium (n=9 fish per treatment) were randomly chosen, sacrificed, and dissected for the inspection of internal organs and lesion sites to conduct a postmortem examination and histopathological investigation. After this, liver and kidney samples were harvested for histopathological examination. Five fish per replication (n=15 fish per treatment) were randomly selected, anesthetized, and blood samples were collected individually to evaluate the levels of nonspecific humoral immunity (NBT, lysozyme, and bactericidal activities), stress response markers (plasma cortisol), oxidative stress markers (glutathione reductase [GR] and lipid peroxidation), antioxidant enzymes (catalase [CAT], superoxide dismutase [SOD], and Glutathione S-transferase [GST]), and metallothionein according to the methodology outlined by Ashry et al. (2024).

Clinical and postmortem examination

Throughout the 6-week trial period, the fish were observed daily for signs of toxicity, behavior, swimming patterns, and appearance (Mello et al., 2025). At the end of the second, fourth, and sixth weeks, fish from each test group were randomly selected and autopsied for examination. The internal organs of the fish, such as the liver, gills, and kidneys, were examined in situ for any gross pathological changes (Shrivastava et al., 2025).

Nonspecific humoral immune assays

Nonspecific humoral immune parameters were analyzed to measure the innate immune potential. Respiratory burst of circulating phagocytes was measured by assessing nitroblue tetrazolium (NBT) reduction. The NBT reduction assay was performed following the method described by Biller-Takahashi et al. (2013). In brief, 15 μl of blood was mixed with NBT solution under specific conditions, and ROS formation was quantified by measuring intracellular formazan granules. Plasma lysozyme activity, based on the Sitós-Bobadilla method (Saera-Vila et al., 2009), was measured by turbidimetry using the lysis of Micrococcus lysodeikticus. After incubation for 0.5 and 4.5 minutes, turbidity was measured at 450 nm using a spectrophotometer. The unit of measurement for plasma lysozyme activity was IU/ml. Bactericidal activity of sera was measured as described by Mizaeva et al. (2023), in which plasma samples were mixed with pathogenic bacterial suspensions, and activity was determined by counting CFU relative to the control.

Analysis of stress and oxidative biomarkers

Physiological stress and oxidative status were evaluated to assess the biological response to treatment. Cortisol levels were determined according to the guidelines provided in the handbook for a commercial enzyme-linked immunosorbent assay kit (Colás-Ruiz et al., 2022). GR activity was determined as the NADPH oxidation rate at 340 nm in the presence of GSSG (Gisbert et al., 2015). The malondialdehyde (MDA) content was determined using the TBARS test, a marker of lipid peroxidation as a symptom of membrane oxidative damage (Janik et al., 2021).

Evaluation of the antioxidant capacity

Key enzyme activities were measured to assess the antioxidant status of the system. The rate of hydrogen peroxide (H₂O₂) breakdown at 240 nm was used to gauge CAT activity (Kim et al., 2017). The ability of SOD to prevent the photochemical reduction of NBT was used to measure SOD activity (Elnakeeb et al., 2025). GST was assessed by measuring the conjugation rate of 1-chloro-2,4-dinitrobenzene (CDNB) and reduced glutathione (GSH) at 340 nm, as described by Gisbert et al. (2015).

Metallothionein profiling

Total, oxidized, and reduced metallothionein (MT) concentrations in whole blood were quantified using a modified spectrophotometric method based on the reaction of thiol groups with Ellman’s reagent (DTNB), as described by Bejaoui et al. (2024). MT levels were expressed as μmol thiol per milligram of protein.

Histopathological examination

At the end of the trial, a set of fish from each treatment group was euthanized, and samples of the target organs, particularly the liver and kidney, were collected. Following fixation in 10% neutral buffered formalin for 24–48 hours, the tissues were rinsed under running water, dehydrated in ethanol, clarified in xylene, and then embedded in paraffin wax (Bisai et al., 2025). Sections of the paraffin block were placed on glass slides and stained with hematoxylin and eosin (H&E) after being cut to a thickness of 5–6 µm. Prepared sections were then examined microscopically for any pathological changes following standard histological procedures (Sivakumar et al., 2022).

Resistance to Bacterial Infection

Vibrio harveyi strain was obtained from naturally affected marine fish exhibiting classical signs of vibriosis and was pathogenically characterized using standard microbiological, biochemical, and molecular biology techniques, including 16S rRNA sequences (Aly et al., 2025). Cultured cells of the selected strain were stored in the microbiology laboratory and periodically subcultured on tryptic soy agar (TSA) supplemented with 2% NaCl. Bacteria were grown in tryptic soy broth containing 2% NaCl at 28 °C for 12–18 hours. The culture was then centrifuged at 3000 × g for 10 minutes at 25 °C, after which the bacteria were suspended in phosphate-buffered saline (PBS) and washed twice. Bacterial concentration was standardized to 1 × 10⁸ CFU ml⁻¹. To prepare the final challenge dose, a 10-fold serial dilution in sterile PBS was performed to obtain a working concentration of 1 × 10⁶ CFU ml⁻¹ per fish. This dose was selected based on a previously determined LD₅₀ value reported by El-Waziry et al. (2025).

At the end of the sixth week of the experiment, a challenge test was conducted using 10 fish per experimental group. The fish were anesthetized with clove oil (Sigma-Aldrich, USA; 50 mg l⁻¹) and intraperitoneally injected with the prepared bacterial suspension (Aly et al., 2024). The infected fish were monitored for 2 weeks for any signs of disease, and the mortality rate was recorded to calculate the relative percentage survival (RPS) using the following equation described by Amend (1981):

Where: SR=survival rate; MR=mortality rate; RPS=relative percent survival.

The RPS values indicate the treatment’s efficacy in improving disease resistance, with higher values signifying greater protection against the pathogen relative to the control.

Data analysis

Statistical analysis was performed using SAS software version 9.1 (Littell WW Stroup and Freund, 2002), and all experimental data were expressed as the arithmetic mean ± standard deviation (M ± SD). To validate the assumptions of normality and homoscedasticity, datasets were evaluated using the Shapiro–Wilk test before further processing. The aquarium (n=3 per treatment) served as the experimental unit for all statistical tests to ensure statistical independence and avoid pseudoreplication. Multiple fish were sampled from each aquarium to assess the immunological, biochemical, and antioxidant properties of the specimens. However, data from individual fish were not treated as separate experimental units; only the average of these data was considered. The treatment effect was separately analyzed at each sampling interval (weeks 2, 4, and 6) using one-way analysis of variance after confirming the assumption of normality and homogeneity of variances. LSD was used in situations where there were differences at p ≤ 0.05 to determine whether there was any separation of means (Gomez and Gomez, 1984). This post hoc approach was selected because of the balanced experimental design and the hypothesis-driven nature of the comparisons, which required maintaining statistical sensitivity to detect biologically relevant differences.

Ethical approval

Specimens were obtained from the Central Laboratory for Aquaculture Research (CLAR), Egypt. They were obtained in accordance with the ethical standards established for conducting biological research. The research design and protocols were ratified officially by the Ethics and Research Committee of the Fish Production Department, Faculty of Agriculture, Al-Azhar University, Cairo, Egypt. They were conducted under the official authorization code (APR-111-fish. Dep. Issue No. 25/202, Dec., 2025). All research protocols were conducted following the established animal welfare regulations. The investigation design and protocols were conducted as per the ARRIVE guidelines (https://arriveguidelines.org; accessed on 20 April 2025).

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Results

Clinical examination of seabream fingerlings exposed to T-2 toxin

External clinical signs

Throughout the experimental period, fingerlings of gilthead sea bream exposed to dietary T-2 toxin exhibited pronounced clinical signs compared to the control and binder-treated groups, as is well documented regarding the impact of dietary mycotoxin contamination on fish health and welfare under intensive aquaculture conditions (Lorusso et al., 2025). After two weeks of T-2 toxin dietary exposure (500 μg kg⁻¹), fingerlings expressed evident abdominal distension due to ascites, which is related to the initial signs of systemic toxicity (Fig. 2A). T-2 toxin has already been shown to cause multi-organ damage, oxidative stress, and immunosuppression in aquatic and terrestrial animals (Matejova et al., 2017; Wang et al., 2020). With prolonged exposure for 4 weeks, severe external hemorrhages were observed, including fin erosion and fin rot, with varying degrees of sloughing observed on dorsal, caudal, and pectoral fins (Fig. 2B), which is consistent with reports regarding the progressive worsening of animal condition, tissue integrity, and external signs of mycotoxicosis in fish and livestock (Diab et al., 2018; Barany et al., 2021).

Fig. 2. Progression of T-2 toxin-induced external pathologies in S. aurata during a 6-week trial (A) Early-stage (week 2) localized skin hemorrhages and pigmentary changes (red arrows). (B) The mid-stage (week 4) manifestation of extensive hemorrhaging (red arrows). (C) Late-stage (week 6) consequences, including marked growth retardation and increased body size variability, are observed in fish (green arrows).

By the end of the 6-week experimental period, fish in the T-2 toxin group displayed marked growth retardation, as reflected by noticeable differences in size among the experimental groups (Fig. 2C). This dwarfism-like condition is consistent with previous reports of impaired growth and development following trichothecene exposure in fish species, including carp and Nile tilapia, in which chronic dietary mycotoxins reduce weight gain and growth efficiency (Matejova et al., 2017; Diab et al., 2018; Barany et al., 2021). On the contrary, seabream fingerlings receiving diets supplemented with Mycofix Plus or Mycosorb A+, either alone or in combination with T-2 toxin, showed a clear attenuation of these external clinical signs, maintaining normal body conformation and fin integrity relative to the toxin-only group. This pattern is consistent with evidence that commercial MBs such as Mycofix and Mycosorb mitigate T-2 and aflatoxin toxicity, improve performance, and alleviate clinical and pathological alterations in different animal species (Jaćević et al., 2020; Kihal et al., 2022; Koynarski et al., 2025.

Postmortem signs

Postmortem examination revealed progressive internal pathological alterations in seabream fingerlings exposed to T-2 toxin, including hepatic congestion, gill damage, and renal abnormalities observed over the experimental period. During the second week of exposure, the fish exhibited hepatic congestion, gill congestion, and exophthalmia, reflecting acute circulatory and inflammatory disturbances (Fig. 3A). These findings are consistent with previous reports of vascular congestion, hemorrhage, and early degenerative changes in the gills and liver following exposure to toxicants and mycotoxins (Shah and Parveen, 2022; Wang et al., 2022).

Fig. 3. Postmortem alterations and tissue lesions in gilthead seabream specimens. (A) Early necropsy results showing severe visceral lesions and abdominal tissue degeneration (blue arrows) together with exophthalmia and eye hemorrhages (red arrow). (B) Hepatic degeneration and accumulation of visceral fat within the abdominal cavity are evident (red arrows), together with severe gill degeneration (blue arrows) after 4 weeks of exposure. (C) Extensive hemorrhage and severe internal organ damage are observed within the abdominal cavity (red arrows) and marked vascular congestion (blue arrows) after 6 weeks of exposure.

More pronounced lesions were evident after 4 weeks of intoxication, including hepatomegaly, characterized by an enlarged liver with a pale whitish appearance, as well as severe gill degeneration marked by sloughing and destruction of the secondary lamellae (Fig. 3B). Similar hepatomegaly, hepatic degeneration, and lamellar sloughing or fusion of gill secondary lamellae have been described as typical responses to chronic toxic insult in fish, including exposure to T-2 toxin and other environmental contaminants (Shah and Parveen, 2022; Wang et al., 2024). At the sixth week, chronic pathological changes were observed, particularly in the kidneys, which appeared enlarged, congested, and mottled with sporadic hemorrhagic patches distributed across the organ surface (Fig. 3C), consistent with previous descriptions of nephrotoxic effects of T-2 and related mycotoxins, including congestion, tubular degeneration, and hemorrhages in renal tissues. These findings are consistent with those of earlier studies reporting severe edema, visceral organ damage, and intestinal or mucosal loss in animals exposed to T-2 toxin and other potent toxicants (Shah and Parveen, 2022; Wang et al., 2024).

Notably, postmortem lesions were markedly less severe or absent in fish receiving dietary supplementation with Mycofix Plus or Mycosorb A+, indicating their protective role in mitigating T-2 toxin-induced organ damage, which is supported by evidence that Mycosorb, Mycofix, and other mycotoxin-binding feed additives significantly reduce T-2-associated histopathological lesions and improve organ integrity in experimentally intoxicated animals (Rodrigues et al., 2017; Singh, 2021).

Temporal effects of T-2 toxin and MBs on nonspecific humoral immune responses

Table 2 summarizes the effects of dietary treatments on nonspecific humoral immune responses. A clear time-dependent pattern was observed, in which fish exposed to T-2 toxin alone exhibited a progressive and significant decline (p < 0.05) in all immune parameters, including NBT activity, lysozyme activity, and bactericidal capacity, with the most pronounced suppression recorded at the 6^th week. This represents a reduction of approximately 30%–40% compared with the control group, indicating severe impairment of innate immune defenses.

Table 2. Immunological responses in S. aurata fingerlings fed with T-2 toxin and MBs for 42 days.

On the contrary, fish fed Mycofix Plus (G2) or Mycosorb A+ (G3) showed a consistent and significant enhancement of immune parameters across all sampling periods, with the highest values recorded at week 6. Notably, G2 and G3 exhibited up to 35%–60% higher lysozyme and bactericidal activity than the control, demonstrating a strong immunostimulatory effect.

Immune parameters were partially restored under combined stress conditions (G5 and G6), with values significantly higher than the T-2 group but still lower than binder-only groups. This indicates that while both binders mitigate T-2-induced immunosuppression, their protective efficacy is incomplete under continuous exposure to toxins.

Effects of T-2 toxin and MBs on stress and oxidative biomarkers

Table 3 presents the effects of dietary treatments on stress and oxidative biomarkers. A pronounced, time-dependent increase in plasma cortisol, GR activity, and MDA levels was observed in fish in the T-2 toxin alone (PC) group. Increased MDA is a hallmark of oxidative damage, and elevated GR activity reflects a compensatory enzymatic response to neutralize toxin-induced free radicals. These effects intensified over time, reaching peak levels at the 6th week, where cortisol and MDA levels were 40%–70% higher than those of the NC group. These findings, specifically the elevated cortisol levels and a sustained stress response, indicate the activation of the hypothalamic–pituitary–interrenal (HPI) axis in T-2 toxin-exposed fish.

Table 3. Stress and antioxidant biomarkers in S. aurata fingerlings fed with T-2 toxin and MBs for 42 days.

Conversely, fish fed diets supplemented with Mycofix Plus (G2) or Mycosorb A+ (G3) maintained significantly lower levels of cortisol and MDA throughout the experiment. The reduction in these biomarkers, particularly with Mycofix Plus during the later stages, demonstrates a high degree of protection against toxin-related physiological stress and oxidative imbalance.

The results for the combined groups (G5 and G6) indicated intermediate values of the measured parameters. These were significantly lower in the T-2 toxin-exposed group than in the control and binder groups; however, they were higher than in the control and binder groups. Therefore, partial protection from the stress caused by toxin action can be assumed.

Effects of T-2 toxin and MBs on antioxidant enzyme activities

The findings regarding the antioxidant enzyme activity are summarized in Table 4. The activities of CAT, SOD, and GST were significantly decreased throughout the experiment. In particular, the most pronounced decreases in activity were registered during week 6 for all studied enzymes in the T-2 toxin group PC. The constant ROS production resulting from T-2 toxin exposure led to a continuous endogenous antioxidant defense system weakening.

Table 4. Antioxidant enzyme activities in S. aurata fingerlings fed with T-2 toxin and MBs for 42 days.

On the contrary, Mycofix Plus (G2) and Mycosorb A+ (G3) treatments led to a greater increase in enzyme activity than the control and T-2 toxin groups. A significant increase in antioxidant enzyme activity was observed at weeks 4 and 6. During these periods, CAT and SOD activities increased by 20%–40%. In other words, the use of Mycofix Plus and Mycosorb A+ protects fish from ROS damage.

Fish in the combined-treatment groups (G5 and G6) exhibited partial restoration compared to the T-2 toxin group but remained significantly lower than those observed in the binder-only groups. Although the binders alleviate oxidative stress, the presence of T-2 toxin continues to suppress antioxidant systems.

Effects of T-2 toxin and MBs on metallothionein profiles in whole blood

The MT content in whole blood for each experimental group is presented in Table 5. Fish exposed to T-2 toxin only PC displayed statistically significant reductions (p < 0.05) in overall, oxidized, and reduced MTs, especially at the 6th week, when a decrease in total MTs exceeded 40% compared with the control group. It may be assumed that T-2 toxin causes serious disturbances in the functioning of thiol-based antioxidant and metal scavenging systems.

Table 5. Metallothionein profiles in S. aurata fingerlings fed with T-2 toxin and MBs for 42 days.

On the contrary, fish from groups G2 and G3, which were given Mycofix Plus and Mycosorb A+, respectively, had significantly higher MT content than other groups. Simultaneously, fish from G3 had the maximum MT content at the 6th week of the experiment. This result may indicate that metallothioneins are more effective in maintaining the redox state and protecting cells.

Fish from the combined treatment groups (G5 and G6) showed partial restoration of MT levels compared with the PC group, but these levels were still significantly lower compared with the groups receiving the binders only. This means that MT synthesis is partially inhibited in the presence of T-2 toxin even after adding binders to feed.

Taken together, these findings confirm that T-2 toxin severely impairs MT-mediated defense systems, whereas MBs restore thiol-dependent antioxidant capacity and enhance resilience against oxidative and toxic stress.

Histopathological findings in seabream fingerlings exposed to T-2 toxin

Histopathological findings indicated significant alterations in liver and kidney tissues of T-2 toxin-exposed seabream fingerlings, while dietary inclusion of MBs reduced the adverse effects of T-2 toxin, consistent with previous findings indicating T-2 toxin and other trichothecene derivatives cause damage to liver and kidney tissues (Fig. 4), although such adverse effects can be minimized using adsorbent products (Koletsi et al., 2021; Gruber-Dorninger et al., 2025).

Fig. 4. Histopathological alterations in the hepatic and renal tissues of S. aurata fingerlings after exposure to dietary T-2 toxin and supplementation with MB. The upper part (A–C) represents histological sections of the liver. In the control group (A), normal histological features are observed with typical cytoplasmic vacuolation. In the T-2 toxin group (B), marked hepatic and pancreatic tissue damage is observed with extensive tissue disintegration (arrowhead). In the MB group (C), extensive vacuolation is observed in the hepatic tissue (arrowhead), indicating protection against tissue necrosis. The lower part (D–F) represents histological sections of the kidney; normal glomerular and tubular structures are observed in the control group (D, arrowhead); tubular tissue damage is observed in the T-2 toxin group (E, arrowhead); and marked improvement in kidney tissue with normal histology is observed in the MB group (F, arrowhead), indicating protection against T-2 toxin-induced tissue damage. (H&E stain; scale bars=50 µm).

The liver of control seabream fingerlings had normally structured liver tissue, consisting of well-organized hepatocytes, whereas those of T-2 toxin-treated fingerlings showed severe damage, including hepatocyte and pancreatic cell disintegration, consistent with previous findings of T-2 toxin hepatotoxicity and hepatocarcinogenicity in fish and other vertebrates (Taroncher et al., 2025). Moreover, there was evident vacuolation of the liver tissue of fingerlings exposed to T-2 toxin and Mycofix Plus, indicating some degree of protection against T-2 toxin-induced damage, although this damage was not completely abolished, aligning with previous findings indicating Mycofix to reduce but not completely abolish mycotoxin-induced liver damage (Chefchaou et al., 2019; Koletsi et al., 2021).

Histological assessment of kidney tissue from fingerlings in the control group showed normal kidney tissue with normally structured glomeruli and renal tubules. On the other hand, histological assessment of kidney tissue from seabream fingerlings exposed to T-2 toxin alone showed focal renal tubular degeneration, indicating severe nephrotoxic effects consistent with reported T-2 toxin-induced vacuolar degeneration, tubular necrosis, and congestion in mammalian kidney tissue (Janik et al., 2021). However, seabream fingerlings exposed to T-2 toxin in combination with Mycofix Plus showed improvement in kidney histology, with normally structured renal tubules, indicating the protective effects of T-2 toxin binder against T-2 toxin-induced kidney damage, consistent with the reported effects of T-2 toxin binder on mycotoxin bioavailability and kidney tissue damage (Agriopoulou et al., 2020).

Histopathological findings corroborate the biochemical and immunological results, confirming that T-2 toxin induces severe hepatic and renal damage, whereas Mycofix Plus and Mycosorb A+ markedly attenuate tissue injury and support organ integrity in seabream fingerlings, supporting the broader evidence that dietary MBs are an effective strategy to mitigate T-2-associated organ toxicity in farmed animals (Nácher-Mestre et al., 2015; Chefchaou et al., 2019; Agriopoulou et al., 2020).

Resistance to V. harveyi infection

The survival and RPS of gilthead seabream fingerlings following experimental challenge with V. harveyi are presented in Table 6. Among fish exposed to dietary T-2 toxin PC, the highest mortality rate (90%) and lowest survival rate (10%) were observed, confirming the immunosuppressive effects of T-2 toxin observed in the immunological assays. Fish receiving diets supplemented with MBs (G2 and G3) showed significantly improved survival, with rates of 70% and 80%, respectively, following bacterial infection.

Table 6. Survival and relative protection levels of S. aurata fingerlings challenged against V. harveyi following dietary exposure to T-2 toxin and MBs.

The calculated RPS values reached 57.14% in G2 and 71.43% in G3, indicating a strong protective effect of both binders against V. harveyi infection. Fish receiving combined treatments of T-2 toxin with Mycofix Plus (G5) or Mycosorb A+ (G6) showed partial disease resistance recovery, with survival rates of 40% and 50% corresponding to RPS values of 14.29% and 28.57%, respectively. However, these values were lower than those recorded in the binder-only groups. These findings demonstrate that dietary exposure to T-2 toxin significantly increases the susceptibility of seabream to bacterial infection, whereas supplementation with MBs enhances disease resistance and improves survival following pathogenic challenge.

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Discussion

This study shows that dietary exposure to T-2 toxin causes severe adverse effects in S. aurata, including clinical manifestations, immunological parameters, oxidative status, and histopathological alterations, while dietary supplementation with Mycofix Plus and Mycosorb A+ MBs significantly reduced these effects. These data underscore the potential risks associated with trichothecenes in fish diets and confirm the efficacy of supplementation with MB in mariculture systems (Gruber-Dorninger et al., 2025; Lorusso et al., 2025).

Clinical and postmortem manifestations of T-2 toxin exposure

Adverse clinical signs, including abdominal distension, fin erosion, and stunted growth, were observed in fish exposed to T-2 toxin, consistent with its cytotoxic and protein synthesis-inhibiting activity (Matejova et al., 2017). The observed dwarfism and body size variability likely result from the negative effects of T-2 toxin on nutrient utilization and growth efficiency, as previously reported in species such as common carp and Nile tilapia (Diab et al., 2018; Barany et al., 2021). These clinical signs indicate systemic toxicity and disruptions in the metabolic and physiological processes of the animals caused by T-2 toxin.

T-2 toxin binds to the 60S ribosomal subunit at the cellular level (Zhang et al., 2022), inhibiting peptidyl transferase activity and consequently blocking protein synthesis (Janik et al., 2021). This leads to impaired tissue regeneration, reduced enzyme production, and metabolic pathway disruption, which collectively explain growth retardation and external deformities (Vörösházi et al., 2024; Abdelrahiem et al., 2025).

Postmortem examination revealed severe organ damage, including enlarged and discolored livers, congested kidneys, and gill lamellae degeneration. These changes indicate cumulative T-2 toxicity in metabolically active organs involved in detoxification, osmoregulation, and immune defense (Shah and Parveen, 2022; Wang et al., 2024). Similar vascular congestion, hemorrhages, and degenerative changes in internal organs have been observed in other aquatic and terrestrial animals after exposure to trichothecenes (Shah and Parveen, 2022).

Such organ-specific damage is closely linked to T-2 toxin-induced mitochondrial dysfunction and cellular apoptosis, particularly in highly metabolically active tissues such as the liver and kidney, where toxin accumulation and oxidative stress are most pronounced (Janik et al., 2021).

Immunosuppressive effects and protective role of MBs

In the current study, non-specific humoral defense mechanisms were found to be severely suppressed in fish exposed to T-2 toxin, as evidenced by decreased respiratory burst activity, plasma lysozyme levels, and bactericidal capacity. A notable outcome of this study is the differential efficacy observed between Mycofix Plus and Mycosorb A+, particularly in terms of immune modulation. Fish receiving Mycosorb A+ consistently exhibited slightly higher lysozyme activity, bactericidal capacity, and respiratory burst responses than those receiving Mycofix Plus. This difference may be attributed to the glucomannan nature of Mycosorb A+ obtained from yeast cell walls, its high binding capacity for trichothecene mycotoxins through hydrogen bond formation, and its complementary structural features (Agriopoulou et al., 2020; Kihal et al., 2022). However, although the efficacy of Mycofix Plus is proven, it depends on its multicomponent composition, including detoxification and adsorption, which may exhibit differential efficiency at varying toxin levels.

These results are consistent with those of earlier studies on the negative impact of T-2 toxin on leukocyte activity, cytokine production, and phagocytosis (Rocha et al., 2005; Matejova et al., 2017; Modra et al., 2018; Chen et al., 2025). These observations are also supported by the results of Rocha et al. (2005), who reported a significant suppression of cell and humoral immunity caused by T-2 toxin, and by other studies revealing the immunotoxicity of T-2 toxin induced through oxidation, inflammasome activation, and apoptosis in immune organs (Rehberger et al., 2017; Kooij et al., 2025). Immunosuppression is achieved by interfering with the biosynthesis of proteins that contribute to immune processes (Matejova et al., 2017) and by inducing the apoptosis of immune cells, including macrophages and lymphocytes. Dysregulation of cellular signaling pathways, such as MAPK and NF-κB, also contributes to this condition by disrupting cytokine secretion (Bai and Guo, 2025). Consequently, T-2-intoxicated seabream displayed the highest mortality rate following V. harveyi challenge, confirming functional immunosuppression.

On the other hand, the immune systems of fish supplemented with an MB were restored to a greater extent than in the control group. This recovery can be attributed primarily to the adsorption of T-2 toxin within the gastrointestinal tract, thereby reducing systemic exposure and restoring immune cell function (Koynarski et al., 2025; Tayyab et al., 2025). Furthermore, Mycosorb A+ may contribute to immunomodulation by enhancing gut-associated immune responses (Jaćević et al., 2020; Mesgar et al., 2022).

Endocrine stress response and oxidative damage

The elevated cortisol levels of T-2-exposed fish indicate the activation of the hypothalamic–pituitary–interrenal axis and a pronounced stress response (Modra et al., 2018; Kulcsár et al., 2023). This endocrine response is linked to cellular stress signaling due to the adverse effects of toxins, which cause oxidative stress and metabolic disturbances. Increased GSH reductase activity and MDA levels indicate oxidative stress and cell membrane damage (Modra et al., 2020; Kulcsár et al., 2023). On the molecular level, the T-2 toxin causes ROS overproduction due to disturbances in mitochondrial transport, lipid oxidation, proteins, and DNA (Kihal et al., 2022; Papatsiros et al., 2023; Goda et al., 2025). The addition of dietary binders helped alleviate these effects (Cuccato et al., 2025; Raz et al., 2025). Differences in the modulation of the stress and oxidative markers between the two binders were also evident. Mycosorb A+ exhibited a significantly higher decrease in cortisol concentrations and lipid peroxidase during the latter sampling times, implying that it better inhibited stress and oxidative injury. This might be attributed to its superior ability to block toxin uptake, thus avoiding the subsequent triggering of stress pathways and ROS production. The slower or incomplete detoxification process, involving both toxin binding and enzyme-mediated conversion, might explain the relatively mild effect observed with Mycofix Plus.

Antioxidant enzyme modulation

The observed suppression of CAT, SOD, and GST activities in fish treated with T-2 toxin indicates a definite disturbance of the antioxidant mechanism and increased sensitivity to ROS-mediated injury. These effects have been previously reported for rainbow trout as well as in other vertebrates, where feeding on T-2 toxin caused decreased CAT activity and deterioration of antioxidant properties, even with some transient activation of other GSH -dependent enzymes, thereby revealing oxidative stress and exhaustion of enzymatic defense (Deng et al., 2017; Modra et al., 2020; Taroncher et al., 2025). Moreover, higher T-2 levels could lead to even more suppressed SOD activity after a preliminary adaptive response, indicating a breakdown of the antioxidant system when faced with constant ROS generation.

Feeding fish on diets containing Mycofix Plus and Mycosorb A+, on the other hand, helped to improve the activities of antioxidant enzymes and decrease lipid peroxidation, providing considerable protection against ROS-induced oxidative damage. The observed results are due to reduced systemic toxins resulting from digestive fluid adsorption and do not reflect any direct antioxidant properties of the tested mycotoxin deactivators, which help preserve redox balance and prevent excessive ROS formation. Similar increases in CAT and SOD activities, as well as decreases in oxidative damage, were observed in carp, poultry, and other animals supplemented with Mycosorb or other mycotoxin sorbents, supporting their protective effects (Puvača et al., 2024; Koynarski et al., 2025).

The partial improvement observed with the combination of treatments demonstrates a specific, dose- and formulation-dependent effect of MBs. Mycosorb A+ consistently showed higher CAT, SOD, and GST activities than Mycofix Plus, indicating a more effective preservation of redox homeostasis. This confirms that rapid toxin sequestration in the gastrointestinal tract plays a critical role in preventing oxidative damage, whereas delayed detoxification mechanisms may allow transient oxidative stress to occur before it is mitigated. These results are also in agreement with previous reports suggesting that the protective efficiency of the studied adsorbents with respect to T-2 depends on the degree of its binding ability, composition, and inclusion in feed (Deng et al., 2017; Modra et al., 2020). Overall, these data are consistent with the established knowledge of the effects of mycotoxins, including T-2, on antioxidant enzyme systems, such as CAT, SOD, and GST, across various animals.

Metallothionein (MT) response

Metallothioneins (MTs) are widely recognized as sensitive biomarkers of oxidative stress and toxic exposures due to their high content of sulfur-containing amino acids and thiol-mediated metal-binding properties, and they represent crucial components of cellular redox systems in aquatic organisms (Roesijadi, 1992; Abdelghany et al., 2023). In this study, fingerlings of seabream exposed to T-2 toxin showed a persistent decrease in total, oxidized, and reduced MT fractions across all treatment periods, indicating the inhibition of thiol-mediated antioxidant mechanisms. The known effects of trichothecene mycotoxins increase oxidative stress and interfere with sulfhydryl-containing defenses, leading to decreased antioxidant protection (Modra et al., 2018; Puvača et al., 2024). These changes imply that the detoxifying abilities of cells are lost due to MT reduction.

In contrast to the toxin-exposed group, diet supplementation with mycotoxin adsorbents increased MT concentration, indicating the ability of these additives to protect the cell redox status. The mechanism is based on reduced toxin uptake, thereby limiting its effects on cellular metabolism and oxidative stress. In particular, fingerlings consuming Mycosorb A+ had maximum MT concentrations after 6 weeks, implying better efficiency in protecting and restoring redox homeostasis. This is in line with previous research confirming the higher effectiveness of yeast-based organic adsorbents in reducing the impact of T-2 toxin (Jaćević et al., 2020; Wang et al., 2023).

In addition, fish receiving combinations of additives showed only a slight increase in the MT fraction compared with the toxin group, with levels still significantly lower than those in groups with adsorbers. This finding shows that detoxifying activity depends on the dose and type of additives, similar to data from other studies performed with various animal species (Dai et al., 2019; Jaćević et al., 2020). Altogether, these results indicate a strong modulation of MT concentration in fish in accordance with contaminant exposure and oxidative stress. This confirms the applicability of MTs as biomarkers of diet-induced mycotoxin stress and the efficiency of additives in aquaculture systems (Roesijadi, 1992; Bejaoui et al., 2024).

Histopathological alterations

The histopathological alterations observed in this study provide clear evidence of the physiological and biochemical disruptions caused by the exposure of the sea bream fingerlings to T-2 toxin. Extreme hepatocellular vacuolation, hepatocyte disruption, and pancreatic tissue damage, in addition to severe degenerative renal tubular changes and congestion, indicate the toxic effects of the T-2 toxin on the liver and kidneys. The findings are in line with the already established ability of trichothecene mycotoxins to cause oxidative stress-based cytotoxicity resulting in progressive degenerative changes of liver and kidney tissues in vertebrates (Koletsi et al., 2021; Taroncher et al., 2025; Gruber-Dorninger et al., 2025). The described histopathological changes, such as vacuolation and tubular necrosis, form the structural background for the functional disorders identified in the current biochemical and immunological analyses.

On the contrary, the consumption of MBs reduced the degree of histopathological impairment and maintained tissue integrity in both the liver and kidney in most tested cases. In particular, the hepatocytes of the treated fish presented organized cell structure without any visible vacuolation, thus reflecting partial protection against toxin-induced degeneration. The tissues of the kidneys in fish receiving Mycofix Plus and Mycosorb A+ supplements had almost intact glomerular and tubular structures compared with the extremely degenerated structures observed following exposure to T-2 toxin. Thus, the dietary intake of such substances appears to be effective in preventing the cell uptake of toxins and, as a result, their subsequent harmful influence on organ structure.

Despite the presence of residual alterations in the livers of some experimental fish after using binder supplements and exposure to multiple treatments, the overall improvement in tissue integrity indicates a clear protective effect of the tested supplements. Previous research has shown that the use of mycotoxin adsorbent additives leads to a decrease in adverse outcomes for organs in the case of their exposure but does not fully prevent damage to tissue structure (Chefchaou et al., 2019; Agriopoulou et al., 2020). Thus, the improvement in tissue structure is clearly correlated with the positive changes in biochemical and immunological characteristics described above, indicating that oxidative stress mitigation and immune suppression also affect tissue morphology.

Resistance to infection

Long-term feeding of T-2 toxin had significant effects on the innate immune system of gilthead seabream, characterized by decreased NBT capacity, plasma lysozyme concentration, and bactericidal activity. The immunosuppressive effect made gilthead seabream extremely vulnerable to V. harveyi infection (SR=10%; RPS=−28.57%). Cortisol and MDA levels were also elevated in gilthead seabream, indicating increased stress and oxidative damage in this species, which may lead to immunosuppression (Modra et al., 2018; Kulcsár et al., 2023). Histopathology revealed hepatocyte and renal tubular cell disintegration, which is consistent with previous findings of T-2-induced organ toxicity in teleosts (Shah and Parveen, 2022).

Dietary supplementation with Mycofix Plus (G2) or Mycosorb A+ (G3) successfully restored immune function, antioxidant enzyme activities (CAT, SOD, and GST), and MT levels. This physiological recovery led to significantly improved survival during the bacterial challenge (SR=70%–80%; RPS=57%–71%). On the contrary, the combined treatment groups (G5 and G6) exhibited intermediate protection (SR=40%–50%; RPS=14%–29%), indicating only partial mitigation of T-2 toxicity during periods of ongoing exposure (Dai et al., 2019; Kihal et al., 2022).

The differences in survival rate and RPS further indicate that Mycosorb A+ is more effective than Mycofix Plus under these experimental conditions. The survival rates and RPS were higher in the fish treated with Mycosorb A+ than Mycofix Plus. Therefore, it can be inferred that better immunity and antioxidant capacity lead to increased disease resistance. This study shows the significance of the composition and working of binders in their efficacy against mycotoxins’ immunosuppressive effects.

Moreover, the strong correlation between immune biomarkers and survival rates highlights the predictive value of NBT, lysozyme, and bactericidal capacity for assessing disease resistance under mycotoxin-induced stress (Rocha et al., 2005; Matejova et al., 2017). These findings demonstrate that commercial MBs effectively counteract T-2-induced immunosuppression, preserve organ integrity, and enhance resistance to opportunistic bacterial infections. These results support the implementation of these binders as a robust preventive strategy for sustainable aquaculture management.

Limitations

The present study has several limitations that warrant consideration. The evaluation of a single T-2 concentration level precludes the observation of dose-dependent effects, and reliance on a controlled laboratory environment may not fully capture the complex ecological and management dynamics of commercial mariculture facilities. The absence of toxin residue analysis in fish tissues (e.g., liver and muscle) prevents a definitive quantification of reduced T-2 bioavailability; consequently, the observed recovery in physiological and antioxidant parameters infers the adsorptive efficacy of the biological binders. Furthermore, the histopathological assessments presented here are descriptive; the absence of a standardized quantitative scoring system for tissue lesions limits the precise indexing of the degree of pathological mitigation. Accordingly, these findings were interpreted as primary experimental observations requiring further validation through multi-dose designs, long-term trials, and advanced molecular approaches. Further investigation into gut microbiome dynamics and validation at the field scale under commercial farming conditions are required to enhance the practical applicability of these results.

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Conclusion

Dietary exposure of gilthead seabream fingerlings (S. aurata) to T-2 toxin induced pronounced immunosuppression, oxidative stress, and tissue damage, thereby increasing their vulnerability to V. harveyi infection. This study clearly demonstrates the significant risks posed by trichothecene contamination in fish feed. Partial protection against the harmful effects of dietary T-2 was observed with Mycofix Plus and Mycosorb A+ administration, resulting in improvements in immunity, antioxidant status, and tissue structure, as well as increased survival after bacterial challenge. This protective effect is probably due to reduced toxin availability and stimulation of physiological defense processes. Nevertheless, the protection provided by the tested binders remained insufficient under continuous exposure of fish to the toxin, and further investigation is required. Furthermore, the use of a single toxin concentration and the absence of molecular-level analyses limit this study. Future studies should involve dose–response relationship assessment, long-term exposure experiments, and underlying mechanism investigation.

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Acknowledgments

The authors would like to extend their deepest gratitude to the members of the participating research groups for their outstanding support. We thank the Aquaculture Laboratory at the Department of Fish Production, Faculty of Agriculture, Al-Azhar University.

Funding

This study received no external funding.

Authors' contributions

All authors contributed equally.

Conflict of interest

The authors declare no conflicts of interest.

Data availability

The data could be provided to the corresponding author upon request.

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