Toxins 2021, 13, 710. https://doi.org/10.3390/toxins13100710 www.mdpi.com/journal/toxins Article Aflatoxin B1 Toxicity in Zebrafish Larva (Danio rerio): Protective Role of Hericium erinaceus Davide Di Paola 1,† , Carmelo Iaria 1,† , Fabiano Capparucci 1 , Marika Cordaro 2 , Rosalia Crupi 3 , Rosalba Siracusa 1 , Ramona D’Amico 1 , Roberta Fusco 1 , Daniela Impellizzeri 1 , Salvatore Cuzzocrea 1,4, *, Nunziacarla Spanò 2, *, Enrico Gugliandolo 3,‡ and Alessio Filippo Peritore 1,‡ 1 Department of Chemical, Biological, Pharmaceutical, and Environmental Science, University of Messina, 98166 Messina, Italy; davide.dipaola@unime.it (D.D.P.); carmelo.iaria@unime.it (C.I.); fabiano.capparucci@unime.it (F.C.); rsiracusa@unime.it (R.S.); rdamico@unime.it (R.D.); rfusco@unime.it (R.F.); dimpellizzeri@unime.it (D.I.); aperitore@unime.it (A.F.P.) 2 Department of Biomedical and Dental Sciences and Morphofunctional Imaging, University of Messina, 98166 Messina, Italy; cordarom@unime.it 3 Department of Veterinary Science, University of Messina, 98166 Messina, Italy; rcrupi@unime.it (R.C.); egugliandolo@unime.it (E.G.) 4 Department of Pharmacological and Physiological Science, Saint Louis University School of Medicine, Saint Louis, MO 63130, USA * Correspondence: salvator@unime.it (S.C.); nunziacarla.spano@unime.it (N.S.) † The first two authors contributed equally to this study. ‡ These authors shared senior authorship. Abstract: Aflatoxin B1 (AFB1), a secondary metabolite produced by fungi of the genus Aspergillus, has been found among various foods as well as in fish feed. However, the effects of AFB1 on fish development and its associated toxic mechanism are still unclear. In the present study, we confirmed the morphological alterations in zebrafish embryos and larvae after exposure to different AFB1 doses as well as the oxidative stress pathway that is involved. Furthermore, we evaluated the potentially protective effect of Hericium erinaceus extract, one of the most characterized fungal extracts, with a focus on the nervous system. Treating the embryos 6 h post fertilization (hpf) with AFB1 at 50 and 100 ng/mL significantly increased oxidative stress and induced malformations in six-day post-fertilization (dpf) zebrafish larvae. The evaluation of lethal and developmental endpoints such as hatching, edema, malformations, abnormal heart rate, and survival rate were evaluated after 96 h of exposure. Hericium inhibited the morphological alterations of the larvae as well as the increase in oxidative stress and lipid peroxidation. In conclusion: our study suggests that a natural extract such as Hericium may play a partial role in promoting antioxidant defense systems and may contrast lipid peroxidation in fish development by counteracting the AFB1 toxicity mechanism. Keywords: aflatoxin B1; oxidative stress; Hericium erinaceus Key Contribution: AFB1 exposure induced malformation; bradycardia; and mortality on zebrafish larvae. At the basis of the AFB1 mechanism, there was an increase in oxidative stress related to an increase of apoptosis. Hericium erinaceus treatment was able to reduce both the AFB1-induced oxidative stress than apoptosis pathway in zebrafish larvae. 1. Introduction Aquaculture is a key aspect of the world food business, supplying about half of all seafood intended for human consumption. The use of plant-based proteins instead of fishmeal in commercial feed formulations has acquired general favor in recent years [1]. However, a growing reliance on plant-based ingredients has raised the potential of Citation: Paola, D.D.; Iaria, C.; Capparucci, F.; Cordaro, M.; Crupi, R.; Siracusa, R.; D’Amico, R.; Fusco, R.; Impellizzeri, D.; Cuzzocrea, S.; et al. Aflatoxin B1 Toxicity in Zebrafish Larva (Danio rerio): Protective Role of Hericium erinaceus. Toxins 2021, 13, 710. https://doi.org/10.3390/ toxins13100710 Received: 7 September 2021 Accepted: 5 October 2021 Published: 8 October 2021 Publisher’s Note: MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affiliations. Copyright: © 2021 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses /by/4.0/).