Vet World   Vol.18   December-2025  Article - 15 

Multiphase antibiofilm potential of shrimp-shell–derived chitosan nanoparticles against Aeromonas hydrophila isolated from tropical aquaculture environments

Rozi Rozi^1,2 , Wiwiek Tyasningsih³ , Jola Rahmahani³ , Eduardus Bimo Aksono Herupradoto⁴ , Muchammad Yunus⁵ , Mohammad Anam Al Arif⁶ , Suryo Kuncorojakti⁷ , Putri Desi Wulan Sari⁸ , Annas Salleh⁹ , and Suwarno Suwarno¹⁰

Background and Aim: Biofilm-forming Aeromonas hydrophila represents a critical constraint in aquaculture, driving recurrent infections, environmental persistence, and antimicrobial resistance. Sustainable alternatives to antibiotics are urgently needed. This study evaluated the multiphase antibiofilm activity of chitosan nanoparticles (ChNPs) synthesized from Litopenaeus vannamei shrimp shells against clinical A. hydrophila isolates from Indonesian gourami (Osphronemus gouramy), focusing on their effects during biofilm adhesion, planktonic proliferation, and mature biofilm degradation. 

Materials and Methods: Between February 2024 and March 2025, diseased gourami were sampled from aquaculture sites in Surabaya, Indonesia. Three wild-type A. hydrophila isolates (A1G1, A2G1, A3G1) were confirmed via biochemical and 16S rRNA sequencing. ChNPs were synthesized through ionic gelation of deacetylated chitosan with sodium tripolyphosphate and characterized by Scanning Electron Microscopy (SEM), dynamic light scattering, and Fourier Transform Infrared Spectroscopy (FTIR) analyses. Antibiofilm efficacy was tested at concentrations of 15–45 μg mL⁻¹ using crystal violet staining (optical density [OD]₅₉₅) for adhesion and degradation phases, and turbidity (OD₆₀₀) for planktonic inhibition. Data were analyzed using one- and two-way analysis of variance with Tukey’s post hoc test. 

Results: ChNPs exhibited spherical morphology (≈641 nm; ζ = +51 mV) and stable ionic crosslinking. They significantly inhibited adherent biomass formation (p < 0.05), reducing OD₅₉₅ from 0.787 to 0.317 in the most responsive strain A3G1 (> 59 % inhibition). Planktonic growth (OD₆₀₀) declined dose-dependently (63 % inhibition at 45 μg mL⁻¹), with significant strain–concentration interactions (p < 0.01). Mature biofilm degradation reached 63% at 45 μg mL⁻¹, approaching the level of the antibiotic-treated control. SEM and FTIR data supported electrostatic disruption and extracellular polymeric substance penetration as probable mechanisms. 

Conclusion: Shrimp-shell–derived ChNPs effectively suppressed A. hydrophila biofilms at multiple developmental stages, demonstrating a potent, biodegradable alternative for the control of aquaculture pathogens. Their integration into eco-friendly, antibiotic-free disease management aligns with circular bioeconomy and One Health frameworks. Further in vivo validation and formulation optimization are warranted. 

Keywords: Chitosan nanoparticles, Aeromonas hydrophila, biofilm inhibition, planktonic suppression, aquaculture biocontrol, One Health, circular bioeconomy.

How to cite this article: Rozi R, Tyasningsih W, Rahmahani J, Aksono Herupradoto EB, Yunus M, Anam Al Arif M, Kuncorojakti S, Wulan Sari PD, Salleh A, and Suwarno S (2025) Multiphase antibiofilm potential of shrimp-shell–derived chitosan nanoparticles against Aeromonas hydrophila isolated from tropical aquaculture environments, Veterinary World, 18(12): 3870–3887.

Received: 27-07-2025   Accepted: 10-11-2025   Published online: 13-12-2025

Corresponding author:  Suwarno Suwarno    E-mail: suwarno@fkh.unair.ac.id

DOI: 10.14202/vetworld.2025.3870-3887

Copyright: Rozi, et al. This article is an open access article distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver (http:// creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.