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A special issue of Microorganisms (ISSN 2076-2607). This special issue belongs to the section "Biofilm".

Deadline for manuscript submissions: closed (30 June 2023) | Viewed by 21724

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Dr. Claudia Vuotto

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Microbial Biofilm Laboratory (LABIM), IRCCS "Fondazione Santa Lucia", Via Ardeatina 306, 00179 Rome, Italy
Interests: microbial biofilms; healthcare-associated infections; biomaterial-associated infections; antimicrobial resistance, bioactive compounds; anti-biofilm compounds; probiotics; microbiota
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Dr. Mariagrazia Di Luca

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Department of Biology, University of Pisa, Via San Zeno 37, 56127 Pisa, Italy
Interests: medical biofilms; persister cells; implant-associated infections; antimicrobial resistance; antimicrobial tolerance; bacteriophages; phage therapy; antimicrobials
Special Issues, Collections and Topics in MDPI journals

Special Issue Information

Dear Colleagues,

A biofilm is defined as a community of surface-associated microbial cells enclosed in a self-produced extracellular polymeric substance, named the EPS-matrix. A significant percentage of Healthcare-Associated Infections (HAIs) diagnosed in developed countries are attributable to the ability of bacteria and/or fungi to adhere to biotic or abiotic surfaces, and to colonize them. These infections are frequently found in mucosal and soft tissues, but are more often associated with the temporary or permanent implantation of medical devices. The onset and spread of biofilm-related HAI on both tissues and medical devices constitute a serious public health problem, with a strong impact on hospitalization duration and costs, as well as on patient management. Cells forming biofilms acquire peculiar traits with respect to the planktonic mode of growth, showing an over-production of extracellular polymeric substances, an over- or under-expression of specific genes, a change in metabolic activity and a reduction in the growth rate. These physiological characteristics bring a gain in terms of fitness by protecting cells from the attack of the host immune response, resisting phagocytosis and other body's defence systems, and by considerably increasing the levels of transient tolerance of microorganisms towards antimicrobial compounds. This increased tolerance creates a situation that is not in itself encouraging, and is even more worrying when it comes to dealing with clinical isolates that already shown an intrinsically complex resistance profile, thus resulting in the onset of chronic and/or relapsing infections.

It is, therefore, easy to understand how the study of the biofilm mode of growth, and the possibilities of preventing and/or counteracting it, are more crucial than ever.

There is still plenty to learn regarding biofilms, so the aim of this Special Issue is to collect updated reviews as well as original research articles based on:

The mechanisms underlying the formation of pathogenic biofilms;
The resistance/tolerance of biofilm-embedded microorganisms to antimicrobials;
The interaction of viruses and bacteria within sessile communities;
The development of effective tools to detect biofilm-associated infections;
The identification of new anti-biofilm strategies.

Dr. Claudia Vuotto
Dr. Mariagrazia Di Luca
Guest Editors

Manuscript Submission Information

Manuscripts should be submitted online at www.mdpi.com by registering and logging in to this website. Once you are registered, click here to go to the submission form. Manuscripts can be submitted until the deadline. All submissions that pass pre-check are peer-reviewed. Accepted papers will be published continuously in the journal (as soon as accepted) and will be listed together on the special issue website. Research articles, review articles as well as short communications are invited. For planned papers, a title and short abstract (about 100 words) can be sent to the Editorial Office for announcement on this website.

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. Microorganisms 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 2700 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

monospecies biofilms
multispecies biofilms
phenotypic and genotypic characterization
host immune response
laboratory methods or protocols
diagnostic tools
tolerance
medical devices
anti-biofilm compounds
anti-fouling biomaterials
anti-biofilm coatings
static and dynamic in vitro studies
in vivo studies

Published Papers (11 papers)

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Research

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17 pages, 3566 KiB

Open AccessArticle

Antimicrobial Peptides Originating from Expression Libraries of Aurelia aurita and Mnemiopsis leidyi Prevent Biofilm Formation of Opportunistic Pathogens

by Lisa Ladewig, Leon Gloy, Daniela Langfeldt, Nicole Pinnow, Nancy Weiland-Bräuer and Ruth A. Schmitz

Microorganisms 2023, 11(9), 2184; https://doi.org/10.3390/microorganisms11092184 - 29 Aug 2023

Cited by 1 | Viewed by 950

Abstract

The demand for novel antimicrobial compounds is rapidly growing due to the rising appearance of antibiotic resistance in bacteria; accordingly, alternative approaches are urgently needed. Antimicrobial peptides (AMPs) are promising, since they are a naturally occurring part of the innate immune system and display remarkable broad-spectrum activity and high selectivity against various microbes. Marine invertebrates are a primary resource of natural AMPs. Consequently, cDNA expression (EST) libraries from the Cnidarian moon jellyfish Aurelia aurita and the Ctenophore comb jelly Mnemiopsis leidyi were constructed in Escherichia coli. Cell-free size-fractionated cell extracts (<3 kDa) of the two libraries (each with 29,952 clones) were consecutively screened for peptides preventing the biofilm formation of opportunistic pathogens using the crystal violet assay. The 3 kDa fraction of ten individual clones demonstrated promising biofilm-preventing activities against Klebsiella oxytoca and Staphylococcus epidermidis. Sequencing the respective activity-conferring inserts allowed for the identification of small ORFs encoding peptides (10–22 aa), which were subsequently chemically synthesized to validate their inhibitory potential. Although the peptides are likely artificial products from a random translation of EST inserts, the biofilm-preventing effects against K. oxytoca, Pseudomonas aeruginosa, S. epidermidis, and S. aureus were verified for five synthetic peptides in a concentration-dependent manner, with peptide BiP_Aa_5 showing the strongest effects. The impact of BiP_Aa_2, BiP_Aa_5, and BiP_Aa_6 on the dynamic biofilm formation of K. oxytoca was further validated in microfluidic flow cells, demonstrating a significant reduction in biofilm thickness and volume by BiP_Aa_2 and BiP_Aa_5. Overall, the structural characteristics of the marine invertebrate-derived AMPs, their physicochemical properties, and their promising antibiofilm effects highlight them as attractive candidates for discovering new antimicrobials. Full article

(This article belongs to the Special Issue Feature Papers in Microbial Biofilms)

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17 pages, 4523 KiB

Open AccessArticle

Klebsiella aerogenes Adhesion Behaviour during Biofilm Formation on Monazite

by Arya Van Alin, Melissa K. Corbett, Homayoun Fathollahzadeh, M. Christian Tjiam, Andrew Putnis, Jacques Eksteen, Anna H. Kaksonen and Elizabeth Watkin

Microorganisms 2023, 11(5), 1331; https://doi.org/10.3390/microorganisms11051331 - 18 May 2023

Viewed by 1200

Abstract

The adsorption behaviour of micro-organisms during the initial attachment stage of biofilm formation affects subsequent stages. The available area for attachment and the chemophysical properties of a surface affect microbial attachment performance. This study focused on the initial attachment behaviour of Klebsiella aerogenes on monazite by measuring the ratio of planktonic against sessile subpopulations (P:S ratio), and the potential role of extracellular DNA (eDNA). eDNA production, effects of physicochemical properties of the surface, particle size, total available area for attachment, and the initial inoculation size on the attachment behaviour were tested. K. aerogenes attached to monazite immediately after exposure to the ore; however, the P:S ratio significantly (p = 0.05) changed in response to the particle size, available area, and inoculation size. Attachment occurred preferentially on larger-sized (~50 µm) particles, and either decreasing the inoculation size or increasing the available area further promoted attachment. Nevertheless, a portion of the inoculated cells always remained in a planktonic state. K. aerogenes produced lower eDNA in response to the changed surface chemical properties when monazite was replaced by xenotime. Using pure eDNA to cover the monazite surface significantly (p ≤ 0.05) hindered bacterial attachment due to the repulsive interaction between the eDNA layer and bacteria. Full article

(This article belongs to the Special Issue Feature Papers in Microbial Biofilms)

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16 pages, 1356 KiB

Open AccessArticle

Antibacterial and Antibiofilm Effect of Unifloral Honeys against Bacteria Isolated from Chronic Wound Infections

by Viktória L. Balázs, Lilla Nagy-Radványi, Erika Bencsik-Kerekes, Regina Koloh, Dina Szabó, Béla Kocsis, Marianna Kocsis and Ágnes Farkas

Microorganisms 2023, 11(2), 509; https://doi.org/10.3390/microorganisms11020509 - 17 Feb 2023

Cited by 7 | Viewed by 2089

Abstract

Honey is known as an alternative remedy for the treatment of wounds. To evaluate the potential of five Hungarian honey types against wound-associated bacteria, in vitro microbiological assays were conducted on Pseudomonas aeruginosa, Staphylococcus epidermidis and methicillin-resistant Staphylococcus aureus (MRSA). Minimum inhibitory concentration (MIC) was determined with the broth macrodilution method, and biofilm degradation capacity was tested with a crystal violet assay. To understand the underlying mechanisms, the effects of honey treatments were assessed on bacterial membrane integrity and quorum sensing (QS). The highest antibacterial activity, indicated by the lowest MIC values, as well as the highest biofilm inhibition rates and membrane disruption, was displayed by chestnut and linden honeys. The most sensitive bacterium was S. epidermidis. Bacterial membrane degradation took place 40 min after treatment with honey solutions of at least a 40% concentration. Each honey sample exhibited anti-QS activity, which was most pronounced in the case of chestnut honey. It was concluded that the antibacterial, biofilm-inhibiting and anti-QS activities of linden and chestnut honeys were superior to those of acacia, goldenrod and milkweed honeys. In addition to the floral source, the antibacterial effect of honey is influenced by the microbial species treated. The use of honey in wound treatment can be justified by its diverse antibacterial mechanisms. Full article

(This article belongs to the Special Issue Feature Papers in Microbial Biofilms)

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15 pages, 2393 KiB

Open AccessArticle

eDNA Provides a Scaffold for Autoaggregation of B. subtilis in Bacterioplankton Suspension

by Iztok Dogsa, Rok Kostanjšek and David Stopar

Microorganisms 2023, 11(2), 332; https://doi.org/10.3390/microorganisms11020332 - 28 Jan 2023

Viewed by 1278

Abstract

The self-binding of bacterial cells, or autoaggregation, is, together with surface colonization, one of the first steps in the formation of a mature biofilm. In this work, the autoaggregation of B. subtilis in dilute bacterial suspensions was studied. The dynamics of cell lysis, eDNA release, and bacterial autoaggregate assembly were determined and related to the spatial autocorrelation of bacterial cells in dilute planktonic bacterial suspensions. The non-random distribution of cells was associated with an eDNA network, which stabilized the initial bacterial cell-cell aggregates. Upon the addition of DNase I, the aggregates were dispersed. The release of eDNA during cell lysis allows for the entrapment of bacterial drifters at a radius several times the size of the dying bacteria. The size of bacterial aggregates increased from 2 to about 100 μm in diameter in dilute bacterial suspensions. The results suggest that B. subtilis cells form previously unnoticed continuum of autoaggregate structures during planktonic growth. Full article

(This article belongs to the Special Issue Feature Papers in Microbial Biofilms)

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13 pages, 2811 KiB

Open AccessArticle

Fungal Glycoside Hydrolases Display Unique Specificities for Polysaccharides and Staphylococcus aureus Biofilms

by Jeremy R. Ellis, James J. Bull and Paul A. Rowley

Microorganisms 2023, 11(2), 293; https://doi.org/10.3390/microorganisms11020293 - 23 Jan 2023

Cited by 3 | Viewed by 1715

Abstract

Commercially available cellulases and amylases can disperse the pathogenic bacteria embedded in biofilms. This suggests that polysaccharide-degrading enzymes would be useful as antibacterial therapies to aid the treatment of biofilm-associated bacteria, e.g., in chronic wounds. Using a published enzyme library, we explored the capacity of 76 diverse recombinant glycoside hydrolases to disperse Staphylococcus aureus biofilms. Four of the 76 recombinant glycoside hydrolases digested purified cellulose, amylose, or pectin. However, these enzymes did not disperse biofilms, indicating that anti-biofilm activity is not general to all glycoside hydrolases and that biofilm activity cannot be predicted from the activity on pure substrates. Only one of the 76 recombinant enzymes was detectably active in biofilm dispersion, an α-xylosidase from Aspergillus nidulans. An α-xylosidase cloned subsequently from Aspergillus thermomutatus likewise demonstrated antibiofilm activity, suggesting that α-xylosidases, in general, can disperse Staphylococcus biofilms. Surprisingly, neither of the two β-xylosidases in the library degraded biofilms. Commercial preparations of amylase and cellulase that are known to be effective in the dispersion of Staphylococcus biofilms were also analyzed. The commercial cellulase contained contaminating proteins with multiple enzymes exhibiting biofilm-dispersing activity. Successfully prospecting for additional antibiofilm enzymes may thus require large libraries and may benefit from purified enzymes. The complexity of biofilms and the diversity of glycoside hydrolases continue to make it difficult to predict or understand the enzymes that could have future therapeutic applications. Full article

(This article belongs to the Special Issue Feature Papers in Microbial Biofilms)

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10 pages, 1123 KiB

Open AccessArticle

Attenuation of Pseudomonas aeruginosa Virulence by Pomegranate Peel Extract

by Samuele Peppoloni, Bruna Colombari, Davide Tagliazucchi, Alessandra Odorici, Cristiano Ventrucci, Aida Meto and Elisabetta Blasi

Microorganisms 2022, 10(12), 2500; https://doi.org/10.3390/microorganisms10122500 - 16 Dec 2022

Cited by 3 | Viewed by 1499

Abstract

Pseudomonas aeruginosa is an opportunistic pathogen often responsible for biofilm-associated infections. The high adhesion of bacterial cells onto biotic/abiotic surfaces is followed by production of an extracellular polysaccharidic matrix and formation of a sessile community (the biofilm) by the release of specific quorum-sensing molecules, named autoinducers (AI). When the concentrations of AI reach a threshold level, they induce the expression of many virulence genes, including those involved in biofilm formation, motility, pyoverdine and pyocyanin release. P. aeruginosa embedded into biofilm becomes resistant to both conventional drugs and the host’s immune response. Accordingly, biofilm-associated infections are a major clinical problem underlining the need for new antimicrobial therapies. In this study, we evaluated the effects of pomegranate peel extract (PomeGr) in vitro on P. aeruginosa growth and biofilm formation; moreover, the release of four AI was assessed. The phenolic profile of PomeGr, exposed or not to bacteria, was determined by high-performance liquid chromatography coupled to electrospray ionization mass spectrometry (HPLC-ESI-MS) analysis. We found that bacterial growth, biofilm production and AI release were impaired upon PomeGr treatment. In addition, the PomeGr phenolic content was also markedly hampered following incubation with bacterial cells. In particular, punicalagin, punicalin, pedunculagin, granatin, di-(HHDP-galloyl-hexoside) pentoside and their isomers were highly consumed. Overall, these results provide novel insights on the ability of PomeGr to attenuate P. aeruginosa virulence; moreover, the AI impairment and the observed consumption of specific phenolic compounds may offer new tools in designing innovative therapeutic approaches against bacterial infections. Full article

(This article belongs to the Special Issue Feature Papers in Microbial Biofilms)

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18 pages, 1766 KiB

Open AccessArticle

Analysis of Chemical Structure and Antibiofilm Properties of Exopolysaccharides from Lactiplantibacillus plantarum EIR/IF-1 Postbiotics

by Basar Karaca, Ozan Haliscelik, Mervi Gursoy, Fadime Kiran, Vuokko Loimaranta, Eva Söderling and Ulvi Kahraman Gursoy

Microorganisms 2022, 10(11), 2200; https://doi.org/10.3390/microorganisms10112200 - 07 Nov 2022

Cited by 1 | Viewed by 1595

Abstract

Previous studies have indicated that the exopolysaccharides of lactic acid bacteria exhibit antibiofilm activity against non-oral bacteria by preventing their initial adhesion to surfaces and by downregulating the expression of genes responsible for their biofilm formation. The aims of this study were to (1) characterize the exopolysaccharides (EPSs) of Lactobacillus plantarum EIR/IF-1 postbiotics, (2) test their antibiofilm effect on dual biofilms, and (3) evaluate their bacterial auto-aggregation, co-aggregation, and hydrocarbon-binding inhibitory activity. The EPSs were characterized by FTIR, HPLC, and thermogravimetric analysis. Bacterial auto- and co-aggregation were tested by Kolenbrander’s method and hydrocarbon binding was tested by Rosenberg’s method. Dual biofilms were formed by culturing Fusobacterium nucleatum ATCC 25586 with one of the following bacteria: Prevotella denticola ATCC 33185, P. denticola AHN 33266, Porphyromonas gingivalis ATCC 33277, P. gingivalis AHN 24155, and Filifactor alocis ATCC 35896. The EPSs contained fractions with different molecular weights (51 and 841 kDa) and monosaccharides of glucose, galactose, and fructose. The EPSs showed antibiofilm activity in all the biofilm models tested. The EPSs may have inhibited bacterial aggregation and binding to hydrocarbons by reducing bacterial hydrophobicity. In conclusion, the EPSs of L. plantarum EIR/IF-1, which consists of two major fractions, exhibited antibiofilm activity against oral bacteria, which can be explained by the inhibitory effect of EPSs on the auto-aggregation and co-aggregation of bacteria and their binding to hydrocarbons. Full article

(This article belongs to the Special Issue Feature Papers in Microbial Biofilms)

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15 pages, 7456 KiB

Open AccessArticle

The Multifunctional Role of Poloxamer P338 as a Biofilm Disrupter and Antibiotic Enhancer: A Small Step forward against the Big Trouble of Catheter-Associated Escherichia coli Urinary Tract Infections

by Lucia Henrici De Angelis, Mariarita Stirpe, Dario Tomolillo, Gianfranco Donelli, Iolanda Francolini and Claudia Vuotto

Microorganisms 2022, 10(9), 1757; https://doi.org/10.3390/microorganisms10091757 - 31 Aug 2022

Cited by 1 | Viewed by 1666

Abstract

Poloxamer 338 (P338), a nonionic surfactant amphiphilic copolymer, is herein proposed as an anti-biofilm compound for the management of catheter-associated urinary tract infections (CAUTIs). P338’s ability to disrupt Escherichia coli biofilms on silicone urinary catheters and to serve as antibiotic enhancer was evaluated for biofilm-producing E. coli Ec5FSL and Ec9FSL clinical strains, isolated from urinary catheters. In static conditions, quantitative biofilm formation assay allowed us to determine the active P338 concentration. In dynamic conditions, the BioFlux system, combined with confocal laser scanning microscopy, allowed us to investigate the P338 solution’s ability to detach biofilm, alone or in combination with sub-MIC concentrations of cefoxitin (FOX). The 0.5% P338 solution was able to destroy the structure of E. coli biofilms, to reduce the volume and area fraction covered by adherent cells (41.42 ± 4.79% and 56.20 ± 9.22% reduction for the Ec5FSL and Ec9FSL biofilms, respectively), and to potentiate the activity of 1\2 MIC FOX in disaggregating biofilms (19.41 ± 7.41% and 34.66 ± 3.75% reduction in the area fraction covered by biofilm for Ec5FSL and Ec9FSL, respectively) and killing cells (36.85 ± 7.13% and 32.33 ± 4.65% increase in the biofilm area covered by dead Ec5FSL and Ec9FSL cells, respectively). Full article

(This article belongs to the Special Issue Feature Papers in Microbial Biofilms)

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22 pages, 4088 KiB

Open AccessArticle

Transcriptomic Analysis Reveals the Role of tmRNA on Biofilm Formation in Bacillus subtilis

by Shanshan Xu, Qianqian Cao, Zengzhi Liu, Junpeng Chen, Peiguang Yan, Bingyu Li and Ying Xu

Microorganisms 2022, 10(7), 1338; https://doi.org/10.3390/microorganisms10071338 - 01 Jul 2022

Cited by 4 | Viewed by 1820

Abstract

Bacillus strains are widely distributed in terrestrial and marine environments, and some of them are used as biocontrol organisms for their biofilm-formation ability. In Bacillus subtilis, biofilm formation is fine-tuned by a complex network, a clear understanding of which still requires study. In bacteria, tmRNA, encoded by the ssrA gene, catalyzes trans-translation that can rescue ribosomes stalled on mRNA transcripts lacking a functional stop codon. tmRNA also affects physiological bioprocesses in some bacteria. In this study, we constructed a ssrA mutant in B. subtilis and found that the biofilm formation in the ssrA mutant was largely impaired. Moreover, we isolated a biofilm-formation suppressor of ssrA, in which the biofilm formation was restored to a level even stronger than that in the wild type. We further performed RNAseq assays with the wild type, ssrA mutant, and suppressor of ssrA for comparisons of their transcriptomes. By analyzing the transcriptomic data, we predicted the possible functions of some differentially expressed genes (DEGs) in the tmRNA regulation of biofilm formation in B. subtilis. Finally, we found that the overexpression of two DEGs, acoA and yhjR, could restore the biofilm formation in the ssrA mutant, indicating that AcoA and YhjR were immediate regulators involved in the tmRNA regulatory web controlling biofilm formation in B. subtilis. Our data can improve the knowledge about the molecular network involved in Bacillus biofilm formation and provide new targets for manipulation of Bacillus biofilms for future investigation. Full article

(This article belongs to the Special Issue Feature Papers in Microbial Biofilms)

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Review

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30 pages, 1459 KiB

Open AccessReview

Biosynthesis of Metal and Metal Oxide Nanoparticles Using Microbial Cultures: Mechanisms, Antimicrobial Activity and Applications to Cultural Heritage

by António Carrapiço, Maria Rosário Martins, Ana Teresa Caldeira, José Mirão and Luís Dias

Microorganisms 2023, 11(2), 378; https://doi.org/10.3390/microorganisms11020378 - 02 Feb 2023

Cited by 15 | Viewed by 3783

Abstract

Nanoparticles (1 to 100 nm) have unique physical and chemical properties, which makes them suitable for application in a vast range of scientific and technological fields. In particular, metal nanoparticle (MNPs) research has been showing promising antimicrobial activities, paving the way for new applications. However, despite some research into their antimicrobial potential, the antimicrobial mechanisms are still not well determined. Nanoparticles’ biosynthesis, using plant extracts or microorganisms, has shown promising results as green alternatives to chemical synthesis; however, the knowledge regarding the mechanisms behind it is neither abundant nor consensual. In this review, findings from studies on the antimicrobial and biosynthesis mechanisms of MNPs were compiled and evidence-based mechanisms proposed. The first revealed the importance of enzymatic disturbance by internalized metal ions, while the second illustrated the role of reducing and negatively charged molecules. Additionally, the main results from recent studies (2018–2022) on the biosynthesis of MNPs using microorganisms were summarized and analyzed, evidencing a prevalence of research on silver nanoparticles synthesized using bacteria aiming toward testing their antimicrobial potential. Finally, a synopsis of studies on MNPs applied to cultural heritage materials showed potential for their future use in preservation. Full article

(This article belongs to the Special Issue Feature Papers in Microbial Biofilms)

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14 pages, 1322 KiB

Open AccessReview

Oral Dysbiosis and Neurodegenerative Diseases: Correlations and Potential Causations

by Justine S. Nicholson and Kyle S. Landry

Microorganisms 2022, 10(7), 1326; https://doi.org/10.3390/microorganisms10071326 - 30 Jun 2022

Cited by 6 | Viewed by 2710

Abstract

Biofilms are a heterogenous complex community of vegetative cells and extracellular polymeric substances that can adhere to various surfaces and are responsible for a variety of chronic and acute diseases. The impact of bacterial biofilms on oral and intestinal health is well studied, but the correlation and causations of biofilms and neurodegenerative diseases are still in their infancy. However, the correlations between biofilms and diseases such as Alzheimer’s Disease, Multiple Sclerosis, and even Parkinson’s Disease are starting to demonstrate the role bacterial biofilms have in promoting and exasperating various illnesses. The review article provides insight into the role bacterial biofilms may have on the development and progression of various neurodegenerative diseases and hopefully shine a light on this very important area of research. Full article

(This article belongs to the Special Issue Feature Papers in Microbial Biofilms)

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