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Challenges in Discovering Innovations Related to Biofilms: Virulence, Spread, Control...
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Challenges in Discovering Innovations Related to Biofilms: Virulence, Spread, Control Strategies and Treatment

A special issue of Pharmaceuticals (ISSN 1424-8247). This special issue belongs to the section "Pharmacology".

Deadline for manuscript submissions: 25 March 2026 | Viewed by 4241

Special Issue Editor

Dr. Tsvetelina Paunova-Krasteva

E-Mail Website
Guest Editor
Institute of Microbiology Bulgarian Academy of Sciences, Sofia, Bulgaria
Interests: microbiology; biofilm formation; biofouling; antibiofilm agents; antimicrobial resistance; quorum sensing; virulence; antimicrobials

Special Issue Information

Dear Colleagues,

The development of a thematic focus on biofilms presents a significant challenge given the vast scope and scientific interest in understanding these complex microbial communities. This Special Issue aims to encourage researchers to publish innovations that could provide deeper insights into the biofilm network. The aim is to highlight current advances in the field of biofilms, with particular attention to virulence mechanisms, biofilm formation and structure, as well as resistance to antimicrobial agents. New developments targeting biofilm dispersion and removal, especially through the application of natural or synthetic inhibitors (such as plant extracts, phytochemicals, nanoparticles, probiotics, biosurfactants, antimicrobial peptides, microbial enzymes, etc.), will also be a key focus of this issue. Additionally, strategies for controlling and treating infectious and environmental biofilms remain an evolving topic that holds significant promise for future research and is also slated for consideration. Addressing these priority scientific questions focused on biofilm research would reveal significant advancements in the field.

Dr. Tsvetelina Paunova-Krasteva
Guest Editor

Manuscript Submission Information

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Submitted manuscripts should not have been published previously, nor be under consideration for publication elsewhere (except conference proceedings papers). All manuscripts are thoroughly refereed through a single-blind peer-review process. A guide for authors and other relevant information for submission of manuscripts is available on the Instructions for Authors page. Pharmaceuticals is an international peer-reviewed open access monthly journal published by MDPI.

Please visit the Instructions for Authors page before submitting a manuscript. The Article Processing Charge (APC) for publication in this open access journal is 2900 CHF (Swiss Francs). Submitted papers should be well formatted and use good English. Authors may use MDPI's English editing service prior to publication or during author revisions.

Keywords

  • biofilm
  • structure
  • pathogenesis
  • virulence
  • tolerance
  • biofilm infections
  • dispersion
  • control strategies
  • quorum sensing

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Published Papers (5 papers)

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Research

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24 pages, 7480 KB  
Article
Interference of Pseudomonas aeruginosa Virulence Factors by Different Extracts from Inula Species
by Tsvetelina Paunova-Krasteva, Petya D. Dimitrova, Tsvetozara Damyanova, Dayana Borisova, Milena Leseva, Iveta Uzunova, Petya A. Dimitrova, Viktoria Ivanova, Antoaneta Trendafilova, Ralitsa Veleva and Tanya Topouzova-Hristova
Pharmaceuticals 2025, 18(12), 1824; https://doi.org/10.3390/ph18121824 - 29 Nov 2025
Viewed by 348
Abstract
Objectives: Pseudomonas aeruginosa is an opportunistic pathogen of high clinical relevance due to its ability to form biofilms, its inherent virulence regulated by quorum-sensing systems, and its multidrug resistance. In the present study, we evaluated the inhibitory potential of nine extracts from [...] Read more.
Objectives: Pseudomonas aeruginosa is an opportunistic pathogen of high clinical relevance due to its ability to form biofilms, its inherent virulence regulated by quorum-sensing systems, and its multidrug resistance. In the present study, we evaluated the inhibitory potential of nine extracts from Inula species (chloroform and methanolic fractions, including a sesquiterpene lactone-enriched fraction) against biofilm formation and virulence-associated traits of P. aeruginosa PAO1 and three multidrug-resistant clinical isolates, as well as their cytotoxicity, biocompatibility, and ability to affect cytokine and nitric oxide production in infected skin explants. Methods: The following methods were applied: fractionation and extraction of plant extracts; cytotoxicity assessment on HFF cells; crystal violet assay for determining antibiofilm activity; fluorescence microscopy for evaluating biofilm viability; electron microscopy for assessing the 3D structure of biofilms and morphological alterations; inhibition assays of pyocyanin pigment, protease activity, bacterial motility, interleukin-17, and nitric oxide production; histological analysis of mouse skin explants. Results: Quantitative analyses of antibiofilm activity revealed that five of the tested extracts inhibited biofilm formation by more than 50%. Structural and functional analyses using confocal laser scanning microscopy and scanning electron microscopy demonstrated a substantial reduction in biofilm thickness, exfoliation of biofilm biomass, the presence of isolated bacterial clusters, metabolically inactive cell populations, and morphological abnormalities associated with cell elongation, invaginations, and polar deformations as a consequence of treatment. In addition, the plant extracts strongly affected virulence factors regulated by quorum sensing. The methanolic fractions from I. britannica and I. bifrons significantly suppressed pyocyanin synthesis. In contrast, the chloroform fractions from I. helenium and I. spiraeifolia produced the largest inhibition zones in assays for extracellular protease activity. Furthermore, all chloroform extracts suppressed bacterial motility, with the lowest swarming diameter observed for the chloroform and lactone-enriched fractions from I. britannica. The chloroform extracts of I. helenium and I. bifrons, methanolic extracts of I. britannica, and chloroform and methanolic extracts of I. spiraeifolia showed relatively low toxicity to normal diploid human fibroblasts. Methanolic and chloroform fractions from I. britannica disrupted biofilm integrity and reduced IL-17A and nitric oxide production in infected skin explants. Conclusions: All these findings indicate a possible synergistic action of the chemical constituents within the fractions on quorum-sensing regulation, biofilm formation, cellular viability, and modulation of host inflammatory responses. Full article
(This article belongs to the Special Issue Challenges in Discovering Innovations Related to Biofilms: Virulence, Spread, Control Strategies and Treatment)
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14 pages, 3857 KB  
Article
Coating Doyle Nasal Silicone Splints with a Sustained Release Varnish Containing Antibiotics Provides Long-Term Protection from Staphylococcus aureus: An In Vitro Study
by Ahmad Siag, Ronit Vogt Sionov, Irith Gati, Michael Friedman, Doron Steinberg and Menachem Gross
Pharmaceuticals 2025, 18(11), 1746; https://doi.org/10.3390/ph18111746 - 17 Nov 2025
Viewed by 364
Abstract
Background/Objectives: Doyle nasal silicone splints are commonly used in nasal surgeries to maintain the shape of the nasal passage and prevent scar tissue formation. However, these implants are prone to bacterial colonization, particularly by Staphylococcus aureus, which is associated with severely [...] Read more.
Background/Objectives: Doyle nasal silicone splints are commonly used in nasal surgeries to maintain the shape of the nasal passage and prevent scar tissue formation. However, these implants are prone to bacterial colonization, particularly by Staphylococcus aureus, which is associated with severely recurrent and recalcitrant cases of infected sinonasal cavities. The aim of this study was to develop a sustained-release varnish (SRV) with antibacterial properties that can be applied to Doyle splints to provide an antibacterial environment for an extended period. Methods: Doyle nasal splints (1 cm × 1 cm segments) were coated with SRV containing one of the three antibiotics: augmentin, ciprofloxacin, or chloramphenicol. A placebo varnish without antibiotics served as a control. The coated splints were exposed daily to a fresh culture of S. aureus, and antibacterial activity was assessed by monitoring bacterial growth. Antibiofilm activity was determined using an MTT metabolic assay. Antibacterial activity was further studied by the kinetic disk diffusion assay, where the stents were transferred daily to new, freshly coated S. aureus plates. Biofilm formation on the coated splints was visualized by high-resolution scanning electron microscopy (HR-SEM). Results: Doyle segments coated with augmentin, ciprofloxacin, or chloramphenicol effectively inhibited S. aureus planktonic growth for 9 ± 1, 18 ± 1, and 21 ± 1 days, respectively. Biofilm formation was prevented for 10 ± 1, 18 ± 1, and 21 ± 1 days, and bacterial clearance occurred for 14 ± 1, 52 ± 1, and >65 days, respectively. HR-SEM images showed the prevention of biofilm formation on the coated segments. Conclusions: Our findings demonstrate that coating Doyle nasal silicon splints with SRV containing augmentin, ciprofloxacin, or chloramphenicol provides long-term antibacterial and antibiofilm activity, with SRV–chloramphenicol being superior. Further studies are needed to confirm the in vivo efficacy of this approach. Full article
(This article belongs to the Special Issue Challenges in Discovering Innovations Related to Biofilms: Virulence, Spread, Control Strategies and Treatment)
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19 pages, 2009 KB  
Article
Quinazoline-Derivatives of Imino-1,2,3-Dithiazoles Promote Biofilm Dispersion of Pseudomonas aeruginosa
by Mathieu Gonzalez, Anne-Sophie Tareau, Daphnée de Crozals, Corentin Layec, Nathan Broudic, Magalie Barreau, Adrien Forge, Olivier Lesouhaitier, Corinne Fruit, Sylvie Chevalier, Thierry Besson and Ali Tahrioui
Pharmaceuticals 2025, 18(11), 1733; https://doi.org/10.3390/ph18111733 - 14 Nov 2025
Viewed by 413
Abstract
Background/Objectives: Biofilm-associated infections pose a major clinical challenge since bacteria within biofilms exhibit highly antibiotic tolerance. Pseudomonas aeruginosa forms persistent biofilms that cause chronic infections in vulnerable patients, including those with cystic fibrosis, burns, or medical implants. Such biofilm-associated chronic infections require prolonged [...] Read more.
Background/Objectives: Biofilm-associated infections pose a major clinical challenge since bacteria within biofilms exhibit highly antibiotic tolerance. Pseudomonas aeruginosa forms persistent biofilms that cause chronic infections in vulnerable patients, including those with cystic fibrosis, burns, or medical implants. Such biofilm-associated chronic infections require prolonged treatments that promote antimicrobial resistance. To address this, recent strategies focus on enhancing biofilm dispersion. Methods: Thirty-six N-arylimino-1,2,3-dithiazoles were screened for their biofilm dispersal activity using a crystal violet assay. Their cytotoxicity was assessed on A549 and HaCat eukaryotic cells. Moreover, their influence on bacterial growth and virulence was investigated. Lastly, fluorescence anisotropy was used to measure membrane fluidity to obtain the first insights on the mechanism of action of these chemicals. Results: Our results showed that quinazoline-derivatives of imino-1,2,3-dithiazoles display biofilm dispersion activity. These compounds do not increase virulence through pyocyanin production, do not modify the growth kinetics of P. aeruginosa, and do not show cytotoxicity towards eucaryotic cells. Conclusions: These findings highlight the potential use of N-arylimino-1,2,3-dithiazole-derived compounds as safe and effective dispersal agents of P. aeruginosa biofilms. Full article
(This article belongs to the Special Issue Challenges in Discovering Innovations Related to Biofilms: Virulence, Spread, Control Strategies and Treatment)
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Review

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27 pages, 1688 KB  
Review
Bacteriophages and Their Enzymes: Allies Against Microbial Biofilms
by Fohad Mabood Husain, Andaleeb Zahra, Asghar Ali, Mohan Kamthan, Nasser A. Al-Shabib, Zeba Farooqui, Naved Ahmad, Thamer Albalawi, Pravej Alam and Nayla Munawar
Pharmaceuticals 2025, 18(12), 1771; https://doi.org/10.3390/ph18121771 - 21 Nov 2025
Viewed by 529
Abstract
Bacterial biofilms pose a substantial challenge in healthcare and industrial and environmental settings because of their resilience and antibiotic resistance. Biofilm formation is a complex process involving microbial communities encased in an extracellular matrix that contributes to increased resistance and persistent infections. This [...] Read more.
Bacterial biofilms pose a substantial challenge in healthcare and industrial and environmental settings because of their resilience and antibiotic resistance. Biofilm formation is a complex process involving microbial communities encased in an extracellular matrix that contributes to increased resistance and persistent infections. This review explores the emerging roles of bacteriophages and their derived enzymes as promising alternatives or adjunct therapies to combat bacterial biofilms. Bacteriophages, viruses that infect bacteria, exhibit marked specificity and diverse mechanisms for targeting and lysing bacterial cells within biofilms. Enzymes, including endolysins and depolymerases, have demonstrated efficacy in disrupting biofilm matrices. Moreover, the potential synergy between bacteriophages and antibiotics enhances their antimicrobial activity, providing a multifaceted approach for combating biofilm-associated infections. This review critically evaluates the current research, highlighting the successes and limitations of bacteriophage-based strategies in biofilm control, and underscores the potential of these alternatives in shaping future therapeutic interventions against biofilm-related bacterial infections. Full article
(This article belongs to the Special Issue Challenges in Discovering Innovations Related to Biofilms: Virulence, Spread, Control Strategies and Treatment)
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37 pages, 4016 KB  
Review
Recent Trends in Bioinspired Metal Nanoparticles for Targeting Drug-Resistant Biofilms
by Devaraj Bharathi and Jintae Lee
Pharmaceuticals 2025, 18(7), 1006; https://doi.org/10.3390/ph18071006 - 5 Jul 2025
Cited by 3 | Viewed by 2147
Abstract
Multidrug-resistant (MDR) biofilm infections characterized by densely packed microbial communities encased in protective extracellular matrices pose a formidable challenge to conventional antimicrobial therapies and are a major contributor to chronic, recurrent and device-associated infections. These biofilms significantly reduce antibiotic penetration, facilitate the survival [...] Read more.
Multidrug-resistant (MDR) biofilm infections characterized by densely packed microbial communities encased in protective extracellular matrices pose a formidable challenge to conventional antimicrobial therapies and are a major contributor to chronic, recurrent and device-associated infections. These biofilms significantly reduce antibiotic penetration, facilitate the survival of dormant persister cells and promote horizontal gene transfer, all of which contribute to the emergence and persistence of MDR pathogens. Metal nanoparticles (MNPs) have emerged as promising alternatives due to their potent antibiofilm properties. However, conventional synthesis methods are associated with high costs, complexity, inefficiency and negative environmental impacts. To overcome these limitations there has been a global push toward the development of sustainable and eco-friendly synthesis approaches. Recent advancements have demonstrated the successful use of various plant extracts, microbial cultures, and biomolecules for the green synthesis of MNPs, which offers biocompatibility, scalability, and environmental safety. This review provides a comprehensive overview of recent trends and the latest progress in the green synthesis of MNPs including silver (Ag), gold (Au), platinum (Pt), and selenium (Se), and also explores the mechanistic pathways and characterization techniques. Furthermore, it highlights the antibiofilm applications of these MNPs emphasizing their roles in disrupting biofilms and restoring the efficacy of existing antimicrobial strategies. Full article
(This article belongs to the Special Issue Challenges in Discovering Innovations Related to Biofilms: Virulence, Spread, Control Strategies and Treatment)
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