Citation: Musa, A.; Abdelgawad, M.A.; Shaker, M.E.; El-Ghorab, A.H.; Parambi, D.G.T.; Hamed, A.A.; Sayed, A.M.; Hassan, H.M.; Aboseada, M.A. Screening and Molecular Docking of Bioactive Metabolites of the Red Sea Sponge Callyspongia siphonella as Potential Antimicrobial Agents. Antibiotics 2022, 11, 1682. https:// doi.org/10.3390/antibiotics11121682 Academic Editor: Melissa M. Cadelis Received: 31 October 2022 Accepted: 21 November 2022 Published: 23 November 2022 Publisher’s Note: MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affil- iations. Copyright: © 2022 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 (https:// creativecommons.org/licenses/by/ 4.0/). antibiotics Article Screening and Molecular Docking of Bioactive Metabolites of the Red Sea Sponge Callyspongia siphonella as Potential Antimicrobial Agents Arafa Musa 1, * , Mohamed A. Abdelgawad 2, * , Mohamed E. Shaker 3 , Ahmed H. El-Ghorab 4 , Della Grace Thomas Parambi 2 , Ahmed A. Hamed 5 , Ahmed M. Sayed 6 , Hossam M. Hassan 7 and Mahmoud A. Aboseada 6 1 Department of Pharmacognosy, College of Pharmacy, Jouf University, Sakaka 72341, Aljouf, Saudi Arabia 2 Department of Pharmaceutical Chemistry, College of Pharmacy, Jouf University, Sakaka 72341, Aljouf, Saudi Arabia 3 Department of Pharmacology, College of Pharmacy, Jouf University, Sakaka 72341, Aljouf, Saudi Arabia 4 Department of Chemistry, College of Science, Jouf University, Sakaka 72341, Aljouf, Saudi Arabia 5 National Research Centre, Microbial Chemistry Department, 33 El-Buhouth Street, Dokki, Giza 12622, Egypt 6 Department of Pharmacognosy, Faculty of Pharmacy, Nahda University, Beni-Suef 62513, Egypt 7 Department of Pharmacognosy, Faculty of Pharmacy, Beni-Suef University, Beni-Suef 62513, Egypt * Correspondence: akmusa@ju.edu.sa (A.M.); mhmdgwd@ju.edu.sa (M.A.A.) Abstract: Marine sponges create a wide range of bioactive secondary metabolites, as documented throughout the year. Several bioactive secondary metabolites were isolated from different mem- bers of Callyspongia siphonella species. This study aimed for isolation and structural elucidation of major metabolites in order to investigate their diverse bioactivities such as antimicrobial and anti- biofilm activities. Afterwards, a molecular docking study was conducted, searching for the possible mechanistic pathway of the most bioactive metabolites. Extraction, fractionation, and metabolomics analysis of different fractions was performed in order to obtain complete chemical profile. Moreover, in vitro assessment of different bioactivities was performed, using recent techniques. Additionally, purification, structural elucidation of high features using recent chromatographic and spectroscopic techniques was established. Finally, AutoDock Vina software was used for the Pharmacophore-based docking-based analysis. As a result, DCM (dichloromethane) fraction exerted the best antibacterial activity using disc diffusion method; particularly against S. aureus with an inhibition zone of 6.6 mm. Compound 11 displayed a considerable activity against both MRSA (Methicillin-resistant Staphyl- lococcus aureus) and Staphyllococcus aureus with inhibition ratios of 50.37 and 60.90%, respectively. Concerning anti-biofilm activity, compounds 1 and 2 displayed powerful activity with inhibition ratios ranging from 39.37% to 70.98%. Pharmacophore-based docking-based analysis suggested elon- gation factor G (EF-G) to be a probable target for compound 11 (siphonellinol C) that showed the best in vitro antibacterial activity, offering unexplored potential for new drugs and treatment candidates. Keywords: marine sponges; Callyspongia siphonella; anti-biofilm; antimicrobial; molecular docking; isolation; metabolites; pharmacophore-based docking 1. Introduction Marine sponges have recently been regarded as a very promising scope for the dis- covery of bioactive natural chemical substances pertaining to their primary and secondary metabolites diversity [1]. Numerous marine sponges regularly maintain a remarkable lack of habitation by tiny creatures and plants, as is well known (fouling organisms). It has been proposed that they accumulate physiologically active chemicals that prevent fouling organisms from settling, and thus being able to protect ships hulls, bridges and aquaculture materials from serious problems [2]. Furthermore, the diffusion of these chem- icals in the tissues of marine sponges may improve the efficacy of the retention processes Antibiotics 2022, 11, 1682. https://doi.org/10.3390/antibiotics11121682 https://www.mdpi.com/journal/antibiotics