Mechanisms of Microbial Biofilm Resistance and New Methods for Biofilm Control

A special issue of Antibiotics (ISSN 2079-6382). This special issue belongs to the section "Antibiofilm Strategies".

Deadline for manuscript submissions: 31 October 2026 | Viewed by 9564

Special Issue Editor


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Guest Editor
Department of Biomolecular Sciences, University of Urbino “Carlo Bo”, 61032 Urbino, Italy
Interests: biofilm; cystic fibrosis; Pseudomonas aeruginosa; persisters; antibiotic resistance; efflux pumps

Special Issue Information

Dear Colleagues,

Biofilm growth represents a key microbial strategy in terms of colonization, pathogenicity, and resistance, allowing microorganisms to persist both in the environment and within a host during infection. Within biofilms, microbial cells exhibit peculiar features that differ from planktonic ones, and special phenotypes are induced that can endure antimicrobial pressure. Indeed, bacterial cells growing in biofilms can be 10 to 1000 times more resistant to antibiotics, and this constitutes one of the main reasons for the lack of eradication of infections. Moreover, biofilms favor DNA exchange between cells, increasing the spread of antimicrobial resistance.

The development of molecular and microscopy techniques has provided novel and valuable information about biofilm formation, architecture, and responses to antibiotic treatment; still, a number of details remain to be elucidated regarding the regulatory pathways controlling the physiology of sessile cultures. Furthermore, there is an urgent need for novel antibiofilm approaches and/or molecules, to achieve their eradication and infection clearance.

This Special Issue will focus on novel advances in the study of biofilm lifestyles, the features characterizing sessile cells, their resistance to antibiotics, and the identification of new compounds endowed with antibiofilm activity.

Dr. Gianmarco Mangiaterra
Guest Editor

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Keywords

  • quorum sensing
  • chronic infections
  • antibiotic resistance
  • persistence
  • efflux pumps

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Published Papers (3 papers)

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Research

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28 pages, 5089 KB  
Article
Sulfated Chitosan Induces Membrane Disruption, Aggregation, and Antibiofilm Activity in Piscirickettsia salmonis: A Biomimetic Strategy as an Antimicrobial Alternative in Aquaculture
by Darwuin Arrieta-Mendoza, Alejandro A. Hidalgo, Andrónico Neira-Carrillo and Sergio A. Bucarey
Antibiotics 2026, 15(5), 435; https://doi.org/10.3390/antibiotics15050435 - 27 Apr 2026
Viewed by 475
Abstract
Background: Sulfated chitosan (ChS) is a chemically modified polysaccharide derived from chitin that mimics heparan sulfate (HS) structures and has emerged as a promising antimicrobial biomaterial. Piscirickettsia salmonis, the etiological agent of Salmonid Rickettsial Septicemia (SRS), represents the main driver of antibiotic [...] Read more.
Background: Sulfated chitosan (ChS) is a chemically modified polysaccharide derived from chitin that mimics heparan sulfate (HS) structures and has emerged as a promising antimicrobial biomaterial. Piscirickettsia salmonis, the etiological agent of Salmonid Rickettsial Septicemia (SRS), represents the main driver of antibiotic use in Chilean aquaculture. Objective: In this study, the in vitro antibacterial activity of ChS against P. salmonis was evaluated. Methods: Elemental characterization by SEM-EDS and FTIR analysis confirmed successful sulfation of the polymer, with a degree of sulfation ranging from 0.92 to 0.95. Additionally, X-ray diffraction (XRD) analysis revealed a reduction in polymer crystallinity, indicating a transition toward a more amorphous structure associated with increased molecular flexibility and functional group accessibility. Results: Antibacterial assays revealed a minimum inhibitory concentration (MIC) of 1500 µg/mL and a minimum bactericidal concentration (MBC ≥ 1500 µg/mL). LIVE/DEAD™ fluorescence imaging showed the formation of bacterial aggregates with increasing size, frequency, and red fluorescence compared to controls over the exposure to ChS, indicating progressive membrane damage. This was supported by a reduction (p < 0.05) in the Green/Red fluorescence ratio of 37–46% between 5 h and 96 h of exposure, corresponding to alteration of the cell membrane. Scanning electron microscopy revealed pronounced morphological alterations by ChS, including surface disruption and loss of cellular integrity. This was more severe compared to commercial chitosan (ChC). Also, ChS reduced (p < 0.05) biofilm formation (>50% at day 6 and 34.8% at day 8). Conclusions: These results demonstrated that ChS disrupts the cell membrane and reduces biofilm formation in P. salmonis, thereby affecting viability. This is the first report of the antibacterial effect of ChS, an HS analogue, against P. salmonis. Full article
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15 pages, 1000 KB  
Article
Deciphering the Emergence of Biofilm-Independent Colistin Persistence and Resistance in A. baumannii: Toxin–Antitoxin Omics and Novel T/A mRNA-asRNA Balance Regulatory Models
by Eleonora Chines, Ludovica Boscarelli, Gaia Vertillo Aluisio, Maria Santagati, Maria Lina Mezzatesta and Viviana Cafiso
Antibiotics 2026, 15(4), 337; https://doi.org/10.3390/antibiotics15040337 - 26 Mar 2026
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Abstract
Background: Persistence represents a critical evolutionary reservoir for the development of antimicrobial resistance in Acinetobacter baumannii (Ab). Understanding the basal mechanisms that enable this survival strategy is crucial for elucidating how high-risk clones evolve resistance during therapy. Methods: High-dose [...] Read more.
Background: Persistence represents a critical evolutionary reservoir for the development of antimicrobial resistance in Acinetobacter baumannii (Ab). Understanding the basal mechanisms that enable this survival strategy is crucial for elucidating how high-risk clones evolve resistance during therapy. Methods: High-dose colistin time-kill assays were performed in ten ST2 clinical colistin-susceptible (COL-S) Carbapenem-Resistant Ab (CRAB) developing in vivo stable and full-colistin resistance to detect persisters. Genomics and basal transcriptomics of chromosomal/plasmid toxin–antitoxin systems (T/As) were performed, as duplicates for each sample, in two ST2 COL-S CRAB to investigate the genomics and basal T/A transcriptomic backgrounds. Results: Phenotypically, all strains showed a persistent subpopulation (~1% survival at 8 h) under 5× COL MIC exposure. Genomics identified 10 type-II and one type-IV T/A systems. Basal transcriptomics revealed active expression patterns mainly of GNAT superfamily T/A systems, with consistently low toxin mRNA levels associated with toxin- or antitoxin-directed asRNAs in chromosomal modules. This architecture defined new dual-combined regulatory models in which asRNAs acted as primary T/A mRNA balance modulators, putatively impacting on the T/A mRNA ratio. Conversely, the plasmid-encoded BrnT/A module showed a highly balanced expression. Conclusions: Our findings revealed, for the first time, the role of the type-II GNAT T/A superfamily as putative molecular switchers via a fine-tuning transcript balance regulation, impacting the transition from a metabolically active cell state to a dormant one in developing colistin persistence and in vivo resistance CRAB. Full article
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Review

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46 pages, 4712 KB  
Review
Biofilms Exposed: Innovative Imaging and Therapeutic Platforms for Persistent Infections
by Manasi Haval, Chandrashekhar Unakal, Shridhar C. Ghagane, Bijay Raj Pandit, Esther Daniel, Parbatee Siewdass, Kingsley Ekimeri, Vijayanandh Rajamanickam, Angel Justiz-Vaillant, Kathy-Ann A. Lootawan, Fabio Muniz De Oliveira, Nivedita Bashetti, Tatheer Alam Naqvi, Arun Shettar and Pramod Bhasme
Antibiotics 2025, 14(9), 865; https://doi.org/10.3390/antibiotics14090865 - 28 Aug 2025
Cited by 14 | Viewed by 7683
Abstract
Biofilms constitute a significant challenge in the therapy of infectious diseases, offering remarkable resistance to both pharmacological treatments and immunological elimination. This resilience is orchestrated through the regulation of extracellular polymeric molecules, metabolic dormancy, and quorum sensing, enabling biofilms to persist in both [...] Read more.
Biofilms constitute a significant challenge in the therapy of infectious diseases, offering remarkable resistance to both pharmacological treatments and immunological elimination. This resilience is orchestrated through the regulation of extracellular polymeric molecules, metabolic dormancy, and quorum sensing, enabling biofilms to persist in both clinical and industrial environments. The resulting resistance exacerbates chronic infections and contributes to mounting economic burdens. This review examines the molecular and structural complexities that drive biofilm persistence and critically outlines the limitations of conventional diagnostic and therapeutic approaches. We emphasize advanced technologies such as super-resolution microscopy, microfluidics, and AI-driven modeling that are reshaping our understanding of biofilm dynamics and heterogeneity. Further, we highlight recent progress in biofilm-targeted therapies, including CRISPR-Cas-modified bacteriophages, quorum-sensing antagonists, enzyme-functionalized nanocarriers, and intelligent drug-delivery systems responsive to biofilm-specific cues. We also explore the utility of in vivo and ex vivo models that replicate clinical biofilm complexity and promote translational applicability. Finally, we discuss emerging interventions grounded in synthetic biology, such as engineered probiotic gene circuits and self-regulating microbial consortia, which offer innovative alternatives to conventional antimicrobials. Collectively, these interdisciplinary strategies mark a paradigm shift from reactive antibiotic therapy to precision-guided biofilm management. By integrating cutting-edge technologies with systems biology principles, this review proposes a comprehensive framework for disrupting biofilm architecture and redefining infection treatment in the post-antibiotic era. Full article
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