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Biofilms and Bacterial Virulence

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A special issue of International Journal of Molecular Sciences (ISSN 1422-0067). This special issue belongs to the section "Molecular Microbiology".

Deadline for manuscript submissions: closed (31 January 2020) | Viewed by 34538

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Special Issue Editors

Dr. Shiro Jimi

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Guest Editor

Central Lab for Pathology and Morphology, Faculty of Medicine, Fukuoka University, Fukuoka, Japan
Interests: biofilm infection; biofilm virulence; biofilm prevention; risks of biofilm formation; eradication methods of biofilm; assay system using different substrates; inter-bacterial biofilm regulation system; MRSA

Dr. Atsushi Togawa

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1. Division of Infectious Diseases, Fukuoka University Hospital, Fukuoka, Japan
2. Division of Medical Oncology, Hematology, and Infectious Diseases, Department of Medicine, Fukuoka University, Fukuoka, Japan
Interests: clinical infectious diseases; biofilm infection; Klebsiella pneumoniae infection; travel medicine

Dr. Motoyasu Miyazaki

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Department of Pharmacy, Fukuoka University Chikushi Hospital, Fukuoka, Japan
Interests: antimicrobial resistance; risks of biofilm formation; epidemiology of MRSA; infection control

Special Issue Information

Dear Colleagues,

Biofilm infections are serious problems in clinical settings. Some bacteria, including Staphylococcus and Pseudomonas aeruginosa, have the ability to form biofilms during their life cycle. Biofilms contribute to bacterial survival and virulence. Most biofilm investigators believe that bacterial properties change when they settle on a surface and form biofilms, which includes the acquisition of nonspecific resistance to anti-bacterial compounds. Biofilms enhance drug resistance more than one hundred times compared to bacteria in the planktonic state, as a consequence of the biofilm’s structural rigidity and/or phenotypic alterations by environmental selection, or gene reconstitution within the confines of the biofilms. The precise nature of these changes is still being unraveled. Even if these changes are understood, eradication of biofilms with drugs is not feasible because the high concentrations required may adversely affect us. For such a disparate situation, the only recourse is to go back to the basics. We may have overlooked something important, such as the exact biofilm risk and bacterial virulence, particularly in the body. This can verify the previous approaches of biofilm investigations.

In this Special Issue, we take a fresh look at this subject with the aim of challenging previous methods and concepts of biofilms. We welcome your articles in this exciting biomedical field.

Dr. Shiro Jimi
Dr. Atsushi Togawa
Dr. Motoyasu Miyazaki
Guest Editors

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Keywords

Biofilm-forming bacteria
Virulence
Biofilm eradication
Substrates
Anti-biofilm compounds
Biofilm assay system

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Published Papers (7 papers)

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Research

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17 pages, 12073 KiB

Open AccessArticle

PchE Regulation of Escherichia coli O157:H7 Flagella, Controlling the Transition to Host Cell Attachment

by Elisa Andreozzi and Gaylen A. Uhlich

Int. J. Mol. Sci. 2020, 21(13), 4592; https://doi.org/10.3390/ijms21134592 - 28 Jun 2020

Cited by 10 | Viewed by 2688

Abstract

Shiga toxins and intimate adhesion controlled by the locus of enterocyte effacement are major enterohemorrhagic Escherichia coli (EHEC) virulence factors. Curli fimbriae also contribute to cell adhesion and are essential biofilm components. The transcriptional regulator PchE represses the expression of curli and their adhesion to HEp-2 cells. Past studies indicate that pchE also represses additional adhesins that contribute to HEp-2 cell attachment. In this study, we tested for pchE regulation of several tissue adhesins and their regulators. Three adhesin-encoding genes (eae, lpfA1, fliC) and four master regulators (csgD, stpA, ler, flhDC) were controlled by pchE. pchE over-expression strongly up-regulated fliC but the marked flagella induction reduced the attachment of O157:H7 clinical isolate PA20 to HEp-2 cells, indicating that flagella were blocking cell attachments rather than functioning as an adhesin. Chemotaxis, motor, structural, and regulatory genes in the flagellar operons were all increased by pchE expression, as was PA20 motility. This study identifies new members in the pchE regulon and shows that pchE stimulates flagellar motility while repressing cell adhesion, likely to support EHEC movement to the intestinal surface early in infection. However, induced or inappropriate pchE-dependent flagellar expression could block cell attachments later during disease progression. Full article

(This article belongs to the Special Issue Biofilms and Bacterial Virulence)

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22 pages, 5774 KiB

Open AccessArticle

Attenuation of Pseudomonas aeruginosa Quorum Sensing by Natural Products: Virtual Screening, Evaluation and Biomolecular Interactions

by Lin Zhong, Vinothkannan Ravichandran, Na Zhang, Hailong Wang, Xiaoying Bian, Youming Zhang and Aiying Li

Int. J. Mol. Sci. 2020, 21(6), 2190; https://doi.org/10.3390/ijms21062190 - 22 Mar 2020

Cited by 47 | Viewed by 4899

Abstract

Natural products play vital roles against infectious diseases since ancient times and most drugs in use today are derived from natural sources. Worldwide, multi-drug resistance becomes a massive threat to the society with increasing mortality. Hence, it is very crucial to identify alternate strategies to control these ‘super bugs’. Pseudomonas aeruginosa is an opportunistic pathogen reported to be resistant to a large number of critically important antibiotics. Quorum sensing (QS) is a cell–cell communication mechanism, regulates the biofilm formation and virulence factors that endow pathogenesis in various bacteria including P. aeruginosa. In this study, we identified and evaluated quorum sensing inhibitors (QSIs) from plant-based natural products against P. aeruginosa. In silico studies revealed that catechin-7-xyloside (C7X), sappanol and butein were capable of interacting with LasR, a LuxR-type quorum sensing regulator of P. aeruginosa. In vitro assays suggested that these QSIs significantly reduced the biofilm formation, pyocyanin, elastase, and rhamnolipid without influencing the growth. Especially, butein reduced the biofilm formation up to 72.45% at 100 µM concentration while C7X and sappanol inhibited the biofilm up to 66% and 54.26% respectively. Microscale thermophoresis analysis revealed that C7X had potential interaction with LasR (K_D = 933±369 nM) and thermal shift assay further confirmed the biomolecular interactions. These results suggested that QSIs are able to substantially obstruct the P. aeruginosa QS. Since LuxR-type transcriptional regulator homologues are present in numerous bacterial species, these QSIs may be developed as broad spectrum anti-infectives in the future. Full article

(This article belongs to the Special Issue Biofilms and Bacterial Virulence)

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14 pages, 3096 KiB

Open AccessArticle

Azalomycin F_5a Eradicates Staphylococcus aureus Biofilm by Rapidly Penetrating and Subsequently Inducing Cell Lysis

by Ganjun Yuan, Pingyi Li, Xuejie Xu, Peibo Li, Qiwang Zhong, Su He, Houqin Yi, Wenfang Yi, Yingying Guan and Zezhang Tom Wen

Int. J. Mol. Sci. 2020, 21(3), 862; https://doi.org/10.3390/ijms21030862 - 29 Jan 2020

Cited by 12 | Viewed by 3485

Abstract

Antimicrobial resistance has emerged as a serious threat to public health. Bacterial biofilm, as a natural lifestyle, is a major contributor to resistance to antimicrobials. Azalomycin F_5a, a natural guanidine-containing polyhydroxy macrolide, has remarkable activities against Gram-positive bacteria, including Staphylococcus aureus, a major causative agent of hospital-acquired infections. To further evaluate its potential to be developed as a new antimicrobial agent, its influence on S. aureus biofilm formation was evaluated using the crystal violet method, and then its eradication effect against mature biofilms was determined by confocal laser scanning microscopy, the drop plate method, and regrowth experiments. The results showed that azalomycin F_5a could significantly inhibit S. aureus biofilm formation, and such effects were concentration dependent. In addition, it can also eradicate S. aureus mature biofilms with the minimum biofilm eradication concentration of 32.0 μg/mL. As extracellular deoxyribonucleic acid (eDNA) plays important roles in the structural integrity of bacterial biofilm, its influence on the eDNA release in S. aureus biofilm was further analyzed using gel electrophoresis. Combined with our previous works, these results indicate that azalomycin F_5a could rapidly penetrate biofilm and causes damages to the cell membrane, leading to an increase in DNase release and eventually eradicating S. aureus biofilm. Full article

(This article belongs to the Special Issue Biofilms and Bacterial Virulence)

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8 pages, 1543 KiB

Open AccessCommunication

Biofilm-Forming by Carbapenem Resistant Enterobacteriaceae May Contribute to the Blood Stream Infection

by Kenichiro Yaita, Kenji Gotoh, Ryuichi Nakano, Jun Iwahashi, Yoshiro Sakai, Rie Horita, Hisakazu Yano and Hiroshi Watanabe

Int. J. Mol. Sci. 2019, 20(23), 5954; https://doi.org/10.3390/ijms20235954 - 26 Nov 2019

Cited by 14 | Viewed by 3914

Abstract

Bloodstream infection (BSI) due to carbapenem-resistant Enterobacteriaceae (CRE) has a high mortality rate and is a serious threat worldwide. Ten CRE strains (eight Enterobacter cloacae, one Klebsiella pneumoniae and one Citrobacter freundii) were isolated from the blood of nine patients, a percentage of whom had been treated with indwelling devices. The steps taken to establish cause included minimum inhibitory concentration (MIC) tests, a pulsed-field gel electrophoresis (PFGE), biofilm study, a multiplex PCR for resistant genes of carbapenemases and extended-spectrum beta-lactamases (ESBLs), and plasmid incompatibility typing. All strains showed a tendency toward resistance to multiple antibiotics, including carbapenems. Frequently isolated genes of ESBLs and carbapenemases include bla_TEM-1 (four strains), bla_SHV-12 (four strains) and bla_IMP-1 (six strains). A molecular analysis by PFGE was used to divide the XbaI-digested genomic DNAs of 10 CRE strains into eight patterns, and the analysis showed that three E. cloacae strains detected from two patients were either identical or closely related. The biofilm production of all CRE strains was examined using a microtiter biofilm assay, and biofilm growth in continuous flow chambers was observed via the use of a confocal laser scanning microscope. Our study indicates that biofilm formation on indwelling devices may pose a risk of BSI due to CRE. Full article

(This article belongs to the Special Issue Biofilms and Bacterial Virulence)

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12 pages, 1247 KiB

Open AccessArticle

Detection and Quantification of eDNA-Associated Bacterial Membrane Vesicles by Flow Cytometry

by Valentina Puca, Eva Ercolino, Christian Celia, Giuseppina Bologna, Luisa Di Marzio, Gabriella Mincione, Marco Marchisio, Sebastiano Miscia, Raffaella Muraro, Paola Lanuti and Rossella Grande

Int. J. Mol. Sci. 2019, 20(21), 5307; https://doi.org/10.3390/ijms20215307 - 25 Oct 2019

Cited by 23 | Viewed by 4909

Abstract

Bacteria generate membrane vesicles, which are structures known as extracellular vesicles (EVs), reported to be involved in different pathogenic mechanisms, as it has been demonstrated that EVs participate in biofilm formation, cell-to-cell communication, bacteria–host interactions, and nutrients supply. EVs deliver nucleic acids, proteins, and polysaccharides. It has been reported that Helicobacter pylori (H. pylori) and Lactobacillus reuteri (L. reuteri), of both planktonic and biofilm phenotypes, produce EVs carrying extracellular DNA (eDNA). Here, we used polychromatic flow cytometry (PFC) to identify, enumerate, and characterize EVs as well as the eDNA-delivering EV compartment in the biofilm and planktonic phenotypes of H.pylori ATCC 43629 and L. reuteri DSM 17938. Biofilm formation was demonstrated and analyzed by fluorescence microscopy, using a classical live/dead staining protocol. The enumeration of EVs and the detection of eDNA-associated EVs were performed by PFC, analyzing both whole samples (cells plus vesicles) and EVs isolated by ultracentrifugation confirm EVs isolated by ultracentrifugation. PFC analysis was performed relying on a known-size beaded system and a mix of three different fluorescent tracers. In detail, the whole EV compartment was stained by a lipophilic cationic dye (LCD), which was combined to PKH26 and PicoGreen that selectively stain lipids and DNA, respectively. Fluorescence microscopy results displayed that both H. pylori and L. reuteri produced well-structured biofilms. PFC data highlighted that, in both detected bacterial species, biofilms produced higher EVs counts when paralleled to the related planktonic phenotypes. Furthermore, the staining with PicoGreen showed that most of the generated vesicles were associated with eDNA. These data suggest that the use of PFC, set according to the parameters here described, allows for the study of the production of eDNA-associated EVs in different microbial species in the same or several phases of growth, thus opening new perspectives in the study of microbial derived EVs in clinical samples. Full article

(This article belongs to the Special Issue Biofilms and Bacterial Virulence)

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19 pages, 4292 KiB

Open AccessArticle

Role of CpxR in Biofilm Development: Expression of Key Fimbrial, O-Antigen and Virulence Operons of Salmonella Enteritidis

by Deeksha Shetty, Juan E. Abrahante, Samuel M. Chekabab, Xuxiaochen Wu, Darren R. Korber and Sinisa Vidovic

Int. J. Mol. Sci. 2019, 20(20), 5146; https://doi.org/10.3390/ijms20205146 - 17 Oct 2019

Cited by 18 | Viewed by 5347

Abstract

Salmonella Enteritidis is a non-typhoidal serovar of great public health significance worldwide. The RpoE sigma factor and CpxRA two-component system are the major regulators of the extracytoplasmic stress response. In this study, we found that the CpxR has highly significant, but opposite effects on the auto-aggregation and swarming motility of S. Enteritidis. Auto-aggregation was negatively affected in the ∆cpxR mutant, whereas the same mutant significantly out-performed its wild-type counterpart with respect to swarming motility, indicating that the CpxR plays a role in biofilm-associated phenotypes. Indeed, biofilm-related assays showed that the CpxR is of critical importance in biofilm development under both static (microtiter plate) and dynamic (flow cell) media flow conditions. In contrast, the RpoE sigma factor showed no significant role in biofilm development under dynamic conditions. Transcriptomic analysis revealed that the cpxR mutation negatively affected the constitutive expression of the operons critical for biosynthesis of O-antigen and adherence, but positively affected the expression of virulence genes critical for Salmonella-mediated endocytosis. Conversely, CpxR induced the expression of curli csgAB and fimbrial stdAC operons only during biofilm development and flagellar motAB and fliL operons exclusively during the planktonic phase, indicating a responsive biofilm-associated loop of the CpxR regulator. Full article

(This article belongs to the Special Issue Biofilms and Bacterial Virulence)

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Review

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22 pages, 2469 KiB

Open AccessReview

Plant-Derived Inhibitors of AHL-Mediated Quorum Sensing in Bacteria: Modes of Action

by Dmitry Deryabin, Anna Galadzhieva, Dianna Kosyan and Galimjan Duskaev

Int. J. Mol. Sci. 2019, 20(22), 5588; https://doi.org/10.3390/ijms20225588 - 8 Nov 2019

Cited by 97 | Viewed by 8331

Abstract

Numerous gram-negative phytopathogenic and zoopathogenic bacteria utilise acylated homoserine lactone (AHL) in communication systems, referred to as quorum sensing (QS), for induction of virulence factors and biofilm development. This phenomenon positions AHL-mediated QS as an attractive target for anti-infective therapy. This review focused on the most significant groups of plant-derived QS inhibitors and well-studied individual compounds for which in silico, in vitro and in vivo studies provide substantial knowledge about their modes of anti-QS activity. The current data about sulfur-containing compounds, monoterpenes and monoterpenoids, phenylpropanoids, benzoic acid derivatives, diarylheptanoids, coumarins, flavonoids and tannins were summarized; their plant sources, anti-QS effects and bioactivity mechanisms have also been summarized and discussed. Three variants of plant-derived molecules anti-QS strategies are proposed: (i) specific, via binding with LuxI-type AHL synthases and/or LuxR-type AHL receptor proteins, which have been shown for terpenes (carvacrol and l-carvone), phenylpropanoids (cinnamaldehyde and eugenol), flavonoid quercetin and ellagitannins; (ii) non-specific, by affecting the QS-related intracellular regulatory pathways by lowering regulatory small RNA expression (sulphur-containing compounds ajoene and iberin) or c-di-GMP metabolism reduction (coumarin); and (iii) indirect, via alteration of metabolic pathways involved in QS-dependent processes (vanillic acid and curcumin). Full article

(This article belongs to the Special Issue Biofilms and Bacterial Virulence)

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