In recent years, research on fish has seen remarkable advancements, especially in aquaculture, ornamental fish industry, and biomedical studies. Immunohistochemistry has become crucial in fish research, aiding in physiological and pathological investigations. However, the use of antibodies originally developed for mammals has raised concerns about their cross-reactivity and specificity in fish. This study systematically evaluated the reactivity of commonly used antibodies for diagnostic purposes, especially in fish pathology, including pan-cytokeratin, vimentin, S-100, glial fibrillary acidic protein, and desmin in the tissue of Sparus aurata, Dicentrarchus labrax, Oncorhynchus mykiss, and Carassius auratus. Western immunoblotting was employed to assess antibody specificity. The results revealed that the pan-cytokeratin and glial fibrillary acidic protein antibodies cross-react with all tested fish species, while S-100 demonstrated specific staining in sea bream, goldfish, and rainbow trout tissues. Conversely, vimentin and desmin antibodies displayed no reactivity. In conclusion, the anti-cytokeratin clone AE1/AE3 and the polyclonal rabbit anti-glial fibrillary acidic protein antibody, which are extensively used in mammals, were validated for fish immunohistochemical studies. Regrettably, D33 anti-desmin and V9 anti-vimentin clones are unsuitable for immunohistochemistry in the tested fish. These findings underscore the need for species-specific antibodies and proper validation for accurate immunohistochemistry analyses in fish research.

The microcotylid Sparicotyle chrysophrii is one of the most harmful parasites for the aquaculture of the gilthead seabream Sparus aurata, for which effective treatments are needed. The efficacy of dietary praziquantel (PZQ, 30–50 mg kg body weight⁻¹ day⁻¹), emamectin benzoate (EMB, 0.015–0.050 mg kg BW⁻¹ day⁻¹), and fenbendazole (FBZ, 40–75 mg kg BW⁻¹ day⁻¹) against S. chrysophrii on gilthead seabream was tested in four different in vivo challenge trials. Specimens of gilthead seabream naturally infected with S. chrysophrii were maintained in three separate recirculating aquaculture systems of 142 L (1 control vs. 2 treated). Fish were fed (ration 1.2% BW day⁻¹) 5 days a week for 4 weeks, alternating weeks with medicated and control feed. On days 0, 14, and 28, samples of 7–9 fish per group were collected and the gills were examined for S. chrysophrii. The gills of hosts treated with EMB and FBZ were evaluated by histopathology. The trials showed that in‐feed PZQ has low palatability, causing rejection which did not allow its efficacy to be adequately investigated. While dietary EMB was associated with adverse effects, such as gill de‐epithelialisation and necrosis, with no demonstrated efficacy against S. chrysophrii. On the other hand, all the fish treated with FBZ lost all parasites, showing 100% efficacy and also suggesting a preventive/repellent action against the monogenean. In conclusion, the present study provides indications for the use of dietary FBZ as a medical device for the treatment/prophylaxis of sparicotylosis in aquaculture.

Meagre (Argyrosomus regius) is one of the fast-growing species considered for sustainable aquaculture development along the Mediterranean and Eastern Atlantic coasts. The emergence of Systemic Granulomatosis (SG), a disease marked by multiple granulomas in various tissues, poses a significant challenge in meagre aquaculture. In the current study, we investigate the association of Mycobacterium spp. and SG in offshore aquaculture facilities in Sardinia, Italy. A total of 34 adult seemingly healthy meagre were arbitrarily collected and analyzed, combining histological, microbiological, molecular, metagenomics, and in situ techniques to investigate the presence of pathogens. Ziehl–Neelsen (ZN), periodic acid–schiff (PAS), and Giemsa stains were performed for the detection of acid-fast bacteria, common parasites, and fungi within granulomas, respectively. Granulomas were detected in 91% (31/34) of fish. The affected organs were kidney (88%), liver (47%), heart (41%), intestine (17.6%), and brain (5%). Acid-fast staining, along with Mycobacterium spp. specific quantitative PCR (qPCR), in situ hybridization (ISH) assay, and microbiological analyses showed negative results for the detection of Mycobacterium spp. and other bacteria implicated in granuloma formation. However, PCR amplification and sequencing of the 65-kDa heat shock protein gene revealed the presence of M. chelonae in 13% of both formalin-fixed and frozen liver tissues. Bacterial isolation failed to detect nontuberculous mycobacteria (NTM) and other bacteria typically associated with granulomas. Consistently, the use of an M. chelonae-specific probe in ISH failed to identify this bacterial species in granulomas. Collectively, results do not support the role of M. chelonae in the development of granulomas and suggest rejecting the hypothesis of a potential link between NTM and SG.