One of the main sources of MPs contamination in fish farms is aquafeed. The present study investigated, for the first time through a comparative approach, the effects of different-sized fluorescent MPs included in a diet intended for zebrafish (Danio rerio). A comparison based on fish developmental stage (larval vs. juvenile), exposure time, and dietary MPs’ size and concentration was performed. Four experimental diets were formulated, starting from the control, by adding fluorescent polymer A (size range 1–5 µm) and B (size range 40–47 µm) at two different concentrations (50 and 500 mg/kg). Zebrafish were sampled at 20 (larval phase) and 60 dpf (juvenile stage). Whole larvae, intestine, liver and muscles of juveniles were collected for the analyses. Polymer A was absorbed at the intestinal level in both larvae and juveniles, while it was evidenced at the hepatic and muscular levels only in juveniles. Hepatic accumulation caused an increase in oxidative stress markers in juveniles, but at the same time significantly reduced the number of MPs able to reach the muscle, representing an efficient barrier against the spread of MPs. Polymer B simply transited through the gut, causing an abrasive effect and an increase in goblet cell abundance in both stages.

Aquafeed’s contamination by microplastics can pose a risk to fish health and quality since they can be absorbed by the gastrointestinal tract and translocate to different tissues. The liver acts as a retaining organ with the consequent triggering of oxidative stress response. The present study aimed to combine the use of natural astaxanthin with natural-based microcapsules to counteract these negative side effects. European seabass juveniles were fed diets containing commercially available fluorescent microplastic microbeads (1–5 μm; 50 mg/kg feed) alone or combined with microencapsulated astaxanthin (AX) (7 g/kg feed; tested for half or whole feeding trial—30 or 60 days, respectively). Fish from the different dietary treatments did not evidence variations in survival and growth performance and did not show pathological alterations at the intestinal level. However, the microplastics were absorbed at the intestinal level with a consequent translocation to the liver, leading, when provided solely, to sod1, sod2, and cat upregulation. Interestingly, the dietary implementation of microencapsulated AX led to a mitigation of oxidative stress. In addition, the microcapsules, due to their composition, promoted microplastic coagulation in the fish gut, limiting their absorption and accumulation in all the tissues analyzed. These results were supported by in vitro tests, which demonstrated that the microcapsules promoted microplastic coagula formation too large to be absorbed at the intestinal level and by the fact that the coagulated microplastics were released through the fish feces.

Microplastics (MPs) in fish can cross the intestinal barrier and are often bioaccumulated in several tissues, causing adverse effects. While the impacts of MPs on fish are well documented, the mechanisms of their cellular internalization remain unclear. A rainbow-trout (Oncorhynchus mykiss) intestinal platform, comprising proximal and distal intestinal epithelial cells cultured on an Alvetex scaffold, was exposed to 50 mg/L of MPs (size 1–5 µm) for 2, 4, and 6 h. MP uptake was faster in RTpi-MI compared to RTdi-MI. Exposure to microplastics compromised the cellular barrier integrity by disrupting the tight-junction protein zonula occludens-1, inducing significant decreases in the transepithelial-electrical-resistance (TEER) values. Consequently, MPs were internalized by cultured epithelial cells and fibroblasts. The expression of genes related to endocytosis (cltca, cav1), macropinocytosis (rac1), and tight junctions’ formation (oclna, cldn3a, ZO-1) was analyzed. No significant differences were observed in cltca, oclna, and cldn3a expression, while an upregulation of cav1, rac1, and ZO-1 genes was detected, suggesting macropinocytosis as the route of internalization, since also cav1 and ZO-1 are indirectly related to this mechanism. The obtained results are consistent with data previously reported in vivo, confirming its validity for identifying MP internalization pathways. This could help to develop strategies to mitigate MP absorption through ingestion.

The aim of the present study was to test synthetic flavors as potential feed attractants in zebrafish (Danio rerio) during early development. Six experimental groups were set up in triplicate: (i) a CTRL group fed a zebrafish commercial diet; (ii) a PG group fed a control diet added with Propylene Glycol (PG); (iii) A1⁺ and A2⁺ groups fed a control diet added with 1% of the two attractive flavors (A1⁺ cheese odor made by mixing Propylene Glycol (PG) with the aromatic chemicals trimethyamine, 2-acetylpyrazine, 2-acetylpyridine, and dimethyl sulfide; and A2⁺ caramel odor, made of PG mixed with the aromatic chemicals vanillin, maltol, cyclotene, acetoin, butyric acid, and capric acid with traces of both gamma-octalactone and gamma-esalactone) or the repulsive flavor (A⁻ coconut odor, made by mixing PG with the aromatic chemicals gamma-eptalactone, gamma-nonalactone, delta-esalactone, and vanillin with trace of both delta-octalactone and maltol), respectively; (iv) an ROT group fed the two attractive diets, each administered singularly in a weekly rotation scheme. All the tested synthetic flavors did not affect the overall health of larval and juvenile fish and promoted growth. Due to the longer exposure time, results obtained from the juvenile stage provided a clearer picture of the fish responses: zebrafish fed both attractive diets showed higher appetite stimulus, feed ingestion, and growth, while the brain dopaminergic activity suggested the A2⁺ diet as the most valuable solution for its long-lasting effect over the whole experiment (60-day feeding trial, from larvae to adults). The present study provided important results about the possible use of attractive synthetic flavors for aquafeed production, opening new sustainable and more economically valuable opportunities for the aquaculture sector.

The invasive blue crab is challenging the Mediterranean basin, progressively declining local populations. This reflects a lower prey availability and suitability of dietary nutrients (mainly n-3 polyunsaturated fatty acids, PUFA). The present study aimed to challenge blue crab males and females with a feed source low in n-3 PUFA with respect to one showing a proper fatty acid profile and to investigate the responses in terms of growth, welfare, lipid characterization of target tissues, and reproductive status. Blue crabs were divided into three groups as follows: (i) Marine: crabs fed sardinella (Sardinella aurita) fillet for 60 days; (ii) Mix: crabs fed bovine heart for the first 40 days and sardinella fillet for the following 20 days; and (iii) Terrestrial: crabs fed bovine heart for 60 days. The diet did not alter the health status but reflected the fatty acid profile of muscle and ovary of the blue crabs. In each group, males and females showed a proper hepatopancreas structure, with comparable levels of lipid reserves. This properly supported gonad maturation in both sexes. However, males and females from the group fed the terrestrial diet were characterized by reduced body weight, revealing that blue crabs prioritize reproductive investment rather than growth by directing crucial nutrients to reproductive organs when a suboptimal diet is available.

Dietary microplastics (MPs) can be consumed by fish, crossing through the gastrointestinal tract. MPs smaller than 20 μm can easily translocate to other organs, such as liver, commonly triggering oxidative stress in fish. Given the current unlikelihood of their short-term elimination, strategies to mitigate MPs-related issues on fish are of considerable interest to the scientific community. In the present study, to reduce both the dietary MPs-induced oxidative stress and the accumulation of MPs, the effectiveness of microencapsulated astaxanthin (ASX) was evaluated in zebrafish (Danio rerio). Specifically, zebrafish were reared from larvae to adults (6 months) and fed diets containing MPs different in range-size (polymer A: 1-5 μm; polymer B: 40-47 μm) at different concentrations (50 or 500 mg/kg). After this period, fish from each experimental group were divided in two sub-groups that were fed, for an additional month, with the previous diets or with the same diets containing implemented with microencapsulated ASX (7 g/kg), respectively. Results showed that microencapsulated ASX was able to counteract the negative effects caused by MPs different in size. Particularly, in zebrafish fed diets containing polymer B microbeads, microencapsulated astaxanthin was able to restore the intestinal epithelium, affected by the abrasive role of MPs during gut transit. Differently, in zebrafish fed diets containing polymer A microbeads, absorbed at intestinal level and translocated mainly to the liver, the microencapsulated ASX decreased the oxidative stress response and reduced the MPs accumulation in target organs due to the antioxidant and the coagulant properties of the ASX and microcapsules wall, respectively. Taken together, the results highlighted that the aquafeeds' implementation with microencapsulated astaxanthin is a prospective tool to prevent MPs-related issues in fish.