Dual-Targeting Approach Degrades Biofilm Matrix and Enhances Bacterial Killing

Biofilm formation is a key virulence factor responsible for a wide range of infectious diseases, including dental caries. Cariogenic biofilms are structured microbial communities embedded in an extracellular matrix that affords bacterial adhesion-cohesion and drug tolerance, making them difficult to treat using conventional antimicrobial monotherapy. Here, we investigated a multitargeted approach combining exopolysaccharide (EPS) matrix-degrading glucanohydrolases with a clinically used essential oils–based antimicrobial to potentiate antibiofilm efficacy. Our data showed that dextranase and mutanase can synergistically break down the EPS glucan matrix in preformed cariogenic biofilms, markedly enhancing bacterial killing by the antimicrobial agent (3-log increase versus antimicrobial alone). Further analyses revealed that an EPS-degrading/antimicrobial (EDA) approach disassembles the matrix scaffold, exposing the bacterial cells for efficient killing while concurrently causing cellular dispersion and “physical collapse” of the bacterial clusters. Unexpectedly, we found that the EDA approach can also selectively target the EPS-producing cariogenic bacteria Streptococcus mutans with higher killing specificity (versus other species) within mixed biofilms, disrupting their accumulation and promoting dominance of commensal bacteria. Together, these results demonstrate a dual-targeting approach that can enhance antibiofilm efficacy and precision by dismantling the EPS matrix and its protective microenvironment, amplifying the killing of pathogenic bacteria within.

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A Real-Time Thermal Sensor System for Quantifying the Inhibitory Effect of Antimicrobial Peptides on Bacterial Adhesion and Biofilm Formation

Sensors ◽

10.3390/s21082771 ◽

2021 ◽

Vol 21 (8) ◽

pp. 2771

Author(s):

Tobias Wieland ◽

Julia Assmann ◽

Astrid Bethe ◽

Christian Fidelak ◽

Helena Gmoser ◽

...

Keyword(s):

Antimicrobial Peptides ◽

Real Time ◽

Bacterial Adhesion ◽

Biofilm Formation ◽

Pathogenic Bacteria ◽

Bovine Mastitis ◽

Inhibitory Effect ◽

Thermal Sensor ◽

Bacterial Cells ◽

Static Conditions

The increasing rate of antimicrobial resistance (AMR) in pathogenic bacteria is a global threat to human and veterinary medicine. Beyond antibiotics, antimicrobial peptides (AMPs) might be an alternative to inhibit the growth of bacteria, including AMR pathogens, on different surfaces. Biofilm formation, which starts out as bacterial adhesion, poses additional challenges for antibiotics targeting bacterial cells. The objective of this study was to establish a real-time method for the monitoring of the inhibition of (a) bacterial adhesion to a defined substrate and (b) biofilm formation by AMPs using an innovative thermal sensor. We provide evidence that the thermal sensor enables continuous monitoring of the effect of two potent AMPs, protamine and OH-CATH-30, on surface colonization of bovine mastitis-associated Escherichia (E.) coli and Staphylococcus (S.) aureus. The bacteria were grown under static conditions on the surface of the sensor membrane, on which temperature oscillations generated by a heater structure were detected by an amorphous germanium thermistor. Bacterial adhesion, which was confirmed by white light interferometry, caused a detectable amplitude change and phase shift. To our knowledge, the thermal measurement system has never been used to assess the effect of AMPs on bacterial adhesion in real time before. The system could be used to screen and evaluate bacterial adhesion inhibition of both known and novel AMPs.

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Antibiotics Application Strategies to Control Biofilm Formation in Pathogenic Bacteria

Current Pharmaceutical Biotechnology ◽

10.2174/1389201020666191112155905 ◽

2020 ◽

Vol 21 (4) ◽

pp. 270-286 ◽

Cited By ~ 2

Author(s):

Fazlurrahman Khan ◽

Dung T.N. Pham ◽

Sandra F. Oloketuyi ◽

Young-Mog Kim

Keyword(s):

Biofilm Formation ◽

Pathogenic Bacteria ◽

Extracellular Polymeric Substances ◽

Quorum Quenching ◽

Resistance Mechanisms ◽

Bacterial Biofilm ◽

Bacterial Cells ◽

High Concentration ◽

Biofilm Inhibition ◽

Abiotic Surfaces

Background: The establishment of a biofilm by most pathogenic bacteria has been known as one of the resistance mechanisms against antibiotics. A biofilm is a structural component where the bacterial community adheres to the biotic or abiotic surfaces by the help of Extracellular Polymeric Substances (EPS) produced by bacterial cells. The biofilm matrix possesses the ability to resist several adverse environmental factors, including the effect of antibiotics. Therefore, the resistance of bacterial biofilm-forming cells could be increased up to 1000 times than the planktonic cells, hence requiring a significantly high concentration of antibiotics for treatment. Methods: Up to the present, several methodologies employing antibiotics as an anti-biofilm, antivirulence or quorum quenching agent have been developed for biofilm inhibition and eradication of a pre-formed mature biofilm. Results: Among the anti-biofilm strategies being tested, the sub-minimal inhibitory concentration of several antibiotics either alone or in combination has been shown to inhibit biofilm formation and down-regulate the production of virulence factors. The combinatorial strategies include (1) combination of multiple antibiotics, (2) combination of antibiotics with non-antibiotic agents and (3) loading of antibiotics onto a carrier. Conclusion: The present review paper describes the role of several antibiotics as biofilm inhibitors and also the alternative strategies adopted for applications in eradicating and inhibiting the formation of biofilm by pathogenic bacteria.

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Application of Bacteriophages on Shiga Toxin-Producing Escherichia coli (STEC) Biofilm

Antibiotics ◽

10.3390/antibiotics10111423 ◽

2021 ◽

Vol 10 (11) ◽

pp. 1423

Author(s):

Nicola Mangieri ◽

Roberto Foschino ◽

Claudia Picozzi

Keyword(s):

Escherichia Coli ◽

Shiga Toxin ◽

Biofilm Formation ◽

Pathogenic Bacteria ◽

Antimicrobial Agents ◽

Bacterial Cells ◽

Abiotic Surfaces ◽

Continuous Presence ◽

Different Strains ◽

Better Than

Shiga toxin-producing Escherichia coli are pathogenic bacteria able to form biofilms both on abiotic surfaces and on food, thus increasing risks for food consumers. Moreover, biofilms are difficult to remove and more resistant to antimicrobial agents compared to planktonic cells. Bacteriophages, natural predators of bacteria, can be used as an alternative to prevent biofilm formation or to remove pre-formed biofilm. In this work, four STEC able to produce biofilm were selected among 31 different strains and tested against single bacteriophages and two-phage cocktails. Results showed that our phages were able to reduce biofilm formation by 43.46% both when used as single phage preparation and as a cocktail formulation. Since one of the two cocktails had a slightly better performance, it was used to remove pre-existing biofilms. In this case, the phages were unable to destroy the biofilms and reduce the number of bacterial cells. Our data confirm that preventing biofilm formation in a food plant is better than trying to remove a preformed biofilm and the continuous presence of bacteriophages in the process environment could reduce the number of bacteria able to form biofilms and therefore improve the food safety.

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Effects of Baseplates of Orthodontic Appliances with in situ generated Silver Nanoparticles on Cariogenic Bacteria: A Randomized, Doubleblind Cross-over Clinical Trial

The Journal of Contemporary Dental Practice ◽

10.5005/jp-journals-10024-1678 ◽

2015 ◽

Vol 16 (4) ◽

pp. 291-298 ◽

Cited By ~ 8

Author(s):

Roghayeh Ghorbanzadeh ◽

Babak Pourakbari

Keyword(s):

Clinical Trial ◽

Silver Nanoparticles ◽

Biofilm Formation ◽

Orthodontic Appliances ◽

Bacterial Cells ◽

Streptococcus Sobrinus ◽

Double Blind ◽

Biofilm Inhibition ◽

Cariogenic Bacteria

ABSTRACT Aim Polymethyl-methacrylate (PMMA) is commonly used primarily for baseplates of orthodontic appliances (BOA). The activities of cariogenic bacteria in biofilm on these surfaces may contribute to dental caries, gingival inflammation and periodontal disease. The PMMA incorporated with nanoparticles of silver (NanoAg-I-PMMA) and NanoAg in situ in PMMA (NanoAg-IS-PMMA) have been shown to control the growth of cariogenic bacteria, but clinical trial of anti-cariogenic application of these novel materials in orthodontics has not been evaluated. The main aim of the study is to compare the clinical effectiveness of using NanoAg-IS-PMMA and NanoAg-I-PMMA for construction of new BOA in inhibiting the planktonic growth and biofilm formation of the cariogenic bacteria. Materials and methods Twenty four patients with a median age of 12.6 years (7-15) harboring Streptococcus mutans, Streptococcus sobrinus and Lactobacillus acidophilus as well as Lactobacillus casei participated in the randomized, doubleblind, cross-over study. The experimental BOA, NanoAg-ISBOA and NanoAg-I-BOA, contained 0.5% w/w NanoAg while the control BOA was standard PMMA. Antibacterial effect of NanoAg-IS-BOA and NanoAg-I-BOA was assessed against test cariogenic bacteria by planktonic and biofilm bacterial cells growth inhibition. Results The average levels of test cariogenic bacteria in saliva decreased about 2 to 70 fold (30.9-98.4%) compared to baseline depending on the microorganism type and test BOA. Biofilm inhibition analysis demonstrated that NanoAg-I-BOA and NanoAg-IS-BOA inhibited the biofilm of all test bacteria by 20.1 to 79.9% compared to BOA. NanoAg-IS-BOA had a strong anti-biofilm effect against S. mutans, S. sobrinus and L. casei. However, NanoAg-I-BOA showed only slight antibiofilm effects on test bacteria. Most notably, at all period of the clinical trial, NanoAg-IS-BOA showed a higher antibacterial activity than NanoAg-I-BOA. Conclusion Based on the novel data that presented here, the NanoAg-IS-BOA had strong antimicrobial activity in the planktonic phase and subsequent biofilm formation of the cariogenic bacteria. Clinical significance Wearing of NanoAg-IS-BOA has the potential to minimize dental plaque formation and caries during orthodontic treatment. How to cite this article Ghorbanzadeh R, Pourakbari B, Bahador A. Effects of Baseplates of Orthodontic Appliances with in situ generated Silver Nanoparticles on Cariogenic Bacteria: A Randomized, Double-blind Cross-over Clinical Trial. J Contemp Dent Pract 2015;16(4):291-298.

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Multiplex CRISPRi System Enables the Study of Stage-Specific Biofilm Genetic Requirements in Enterococcus faecalis

mBio ◽

10.1128/mbio.01101-20 ◽

2020 ◽

Vol 11 (5) ◽

Cited By ~ 2

Author(s):

Irina Afonina ◽

June Ong ◽

Jerome Chua ◽

Timothy Lu ◽

Kimberly A. Kline

Keyword(s):

Enterococcus Faecalis ◽

Biofilm Formation ◽

Pathogenic Bacteria ◽

Expression System ◽

Multidrug Resistant ◽

Essential Genes ◽

Host Bacterium ◽

Content Type ◽

Wide Range ◽

Life Threatening

ABSTRACT Enterococcus faecalis is an opportunistic pathogen, which can cause multidrug-resistant life-threatening infections. Gaining a complete understanding of enterococcal pathogenesis is a crucial step in identifying a strategy to effectively treat enterococcal infections. However, bacterial pathogenesis is a complex process often involving a combination of genes and multilevel regulation. Compared to established knockout methodologies, CRISPR interference (CRISPRi) approaches enable the rapid and efficient silencing of genes to interrogate gene products and pathways involved in pathogenesis. As opposed to traditional gene inactivation approaches, CRISPRi can also be quickly repurposed for multiplexing or used to study essential genes. Here, we have developed a novel dual-vector nisin-inducible CRISPRi system in E. faecalis that can efficiently silence via both nontemplate and template strand targeting. Since the nisin-controlled gene expression system is functional in various Gram-positive bacteria, the developed CRISPRi tool can be extended to other genera. This system can be applied to study essential genes, genes involved in antimicrobial resistance, and genes involved in biofilm formation and persistence. The system is robust and can be scaled up for high-throughput screens or combinatorial targeting. This tool substantially enhances our ability to study enterococcal biology and pathogenesis, host-bacterium interactions, and interspecies communication. IMPORTANCE Enterococcus faecalis causes multidrug-resistant life-threatening infections and is often coisolated with other pathogenic bacteria from polymicrobial biofilm-associated infections. Genetic tools to dissect complex interactions in mixed microbial communities are largely limited to transposon mutagenesis and traditional time- and labor-intensive allelic-exchange methods. Built upon streptococcal dCas9, we developed an easily modifiable, inducible CRISPRi system for E. faecalis that can efficiently silence single and multiple genes. This system can silence genes involved in biofilm formation and antibiotic resistance and can be used to interrogate gene essentiality. Uniquely, this tool is optimized to study genes important for biofilm initiation, maturation, and maintenance and can be used to perturb preformed biofilms. This system will be valuable to rapidly and efficiently investigate a wide range of aspects of complex enterococcal biology.

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Anti-biofilm property of essential oils from Cymbopogon sp. against pathogenic bacteria in single-culture and co-culture

Ciência e Natura ◽

10.5902/2179460x39957 ◽

2020 ◽

Vol 42 ◽

pp. e1

Author(s):

Alessandra Farias Millezi ◽

Vanessa Schuh ◽

Janaina Schuh ◽

Taciara Penno do Amaral

Keyword(s):

Essential Oils ◽

Biofilm Formation ◽

Pathogenic Bacteria ◽

Microtiter Plate ◽

Bacterial Cells ◽

Viable Cells ◽

Single Culture ◽

Cymbopogon Flexuosus ◽

Colony Forming Units ◽

Cymbopogon Martinii

In this study, we evaluated whether the essential oils (EOs) from Cymbopogon flexuosus and Cymbopogon martinii can prevent production of biofilms either in single or combined culture of Staphylococcus aureus and Pseudomonas aeruginosa. Biofilm formation was assessed by microtiter-plate test with further quantification of viable cells and biofilm biomass. The evaluated EOs at 0.78 % significantly (P 0.05) reduced only the viable cells of S. aureus that inhabited biofilm. However, in single-and co-culture assays, both oils significantly (P 0.05) decreased the amount of biofilm biomass. Biofilm reductions between 52-83% and 60-93% were achieved for the treatments with EOs from C. flexuosus and C. martinii, respectively. Although the biomass reductions of simgle and co-cultivated biofilms were significant, the same was not true for viable cells, except for S. aureus. Considering that the remaining colony forming units can reconstitute the EPS matrix, studies with higher concentrations than those used in this research are suggested in order to obtain greater logarithmic reductions of viable bacterial cells.

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Rhizobiales commensal bacteria promote Arabidopsis thaliana root growth via host sulfated peptide pathway

10.1101/2021.05.25.444716 ◽

2021 ◽

Author(s):

Jana Hucklenbroich ◽

Tamara Gigolashvili ◽

Anna Koprivova ◽

Philipp Spohr ◽

Mahnaz Nezamivand Chegini ◽

...

Keyword(s):

Arabidopsis Thaliana ◽

Root Growth ◽

Pathogenic Bacteria ◽

Redox Regulation ◽

Growth Promotion ◽

Primary Root ◽

Redox Status ◽

Commensal Bacteria ◽

Wide Range ◽

Physiological Impact

Root-associated commensal bacteria that belong to the order Rhizobiales, which also contains symbiotic and pathogenic bacteria, promote primary root growth of Arabidopsis thaliana. Yet, its underlying molecular mechanism and physiological impact remained unclear. Here, we conducted a transcriptomic analysis of A. thaliana roots inoculated with root-associated commensal bacteria of Rhizobiales and sister lineages and revealed common and strain/lineage-specific transcriptional responsea, possibly mediated by WRKY and ANAC family of transcription factors. We revealed that the observed common response was also partially triggered by a wide range of non-pathogenic bacteria, fungi, and a multi-kingdom synthetic community (SynCom). This response was characterized by a down-regulation of genes related to intracellular redox regulation, suggesting distinctive redox status between pathogenic and non-pathogenic interactions. By integrating this analysis with developmental and cell biological analyses, we identified a crucial role for the sulfated peptide pathway mediated by TYROSYLPROTEIN SULFOTRANSFERASE (TPST) in Rhizobiales root growth promotion (RGP) activity. Conversely, none of the known sulfated peptide pathway appeared to be required for this activity, suggesting a novel sulfated protein pathway targeted by Rhizobiales RGP. Finally, we show that TPST is needed for RGP exerted by Rhizobiales but not Pseudomonadales isolates, delineating lineage-specific mechanisms to manipulate host root development.

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Staphylococcus aureus biofilm matrix under bone environment influence

10.5194/biofilms9-42 ◽

2020 ◽

Author(s):

Fabien Lamret ◽

Jennifer Varin-Simon ◽

Sophie Gangloff ◽

Fany Reffuveille

Keyword(s):

Staphylococcus Aureus ◽

Bacterial Adhesion ◽

Biofilm Formation ◽

Matrix Composition ◽

Magnesium Concentration ◽

Bone Microenvironment ◽

Low Oxygen ◽

The Matrix ◽

Main Components ◽

Biofilm Matrix

Bone and joint infections linked to implanted materials are mostly due to Staphylococcus aureus. Deciphering the biofilm structure appears to be a promising strategy to develop antibiofilm molecules in order to curb infection occurrence and the bacterial recurrence. Indeed, the characterization of biofilm architecture and physiology could help to find new therapeutic targets through notable quantification of the matrix main components. Our hypothesis is that the very complex and interconnected bone microenvironment influences the bacterial adhesion and biofilm maturation and so its composition. To identify the main factors influencing biofilm formation in the bone microenvironment, we determined biofilm biomass and the number of live adhered bacteria in a static model, completed with microscopy approaches to support our results. Different factors of bone microenvironment were tested: starvation, low oxygen rate, excess of magnesium, and presence of bone cell products. Our first results showed that MSSA or MRSA strains did not have the same behaviors under the tested conditions. However, for both types of strains, excess of magnesium combined to paucity of amino acids and oxygen increased the most the proportion of adhered Staphylococcus aureus (a 6 to 43 fold-increase, p = < 0.01). But biofilm biomass quantification and bacterial adhesion results showed divergent profiles leading us to think that matrix could be involved in such contrasts. Scanning electron microscopy highlighted several structures of matrix produced by these bacteria: well-known slime aspect, but also fibrous appearance, and no matrix production was revealed under some conditions. Indeed, all strains produced few matrix when cultured with control medium and oxygenated condition. Only CIP 53.154 strain built a strong slime-like matrix in response to oxygen depletion. However, both MSSA CIP 53.154 and SH1000 strains developed fibrous structures under anaerobic conditions associated with amino acid starvation, high magnesium concentration with or without glucose. MRSA USA300 strain did not seem to produce a matrix under our conditions, which is supported by the literature. Further investigations of the biofilm matrix are needed to conclude on the matrix nature, which surrounds bacteria under our conditions. The bone microenvironment is complex but our results show that the parameters that mimicked this specific environment influenced the bacterial adhesion and probably the biofilm matrix composition of several strains of Staphylococcus aureus. Further investigations will help to understand how the different factors influence biofilm formation through quantification of the matrix main components by fluorescence microscopy and enzyme digestion. Our final aim is to develop an in vitro model mimicking this specific microenvironment in order to screen different antimicrobial molecules, which could target the biofilm matrix.

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Iranian propolis efficiently inhibits growth of oral streptococci and cancer cell lines

BMC Complementary and Alternative Medicine ◽

10.1186/s12906-019-2677-3 ◽

2019 ◽

Vol 19 (1) ◽

Cited By ~ 1

Author(s):

Fariba Asgharpour ◽

Ali Akbar Moghadamnia ◽

Ebrahim Zabihi ◽

Sohrab Kazemi ◽

Amirmorteza Ebrahimzadeh Namvar ◽

...

Keyword(s):

Chemical Composition ◽

Cell Lines ◽

Cancer Cell ◽

Biofilm Formation ◽

Cytotoxic Effect ◽

Cancer Cell Lines ◽

Oral Streptococci ◽

Cariogenic Bacteria ◽

Wide Range ◽

The Impact

Abstract Background Propolis is a natural bee product with a wide range of biological activities that are related to its chemical composition. The present study investigated the quantification of quercetin (Q) in Ardabil ethanol extract of propolis (AEEP), and then compared its anti-bacterial, anti- biofilm and cytotoxic effects on cancer and normal cell lines. Method In the present study, the chemical composition of AEEP was determined through the high-performance liquid chromatography (HPLC). The AEEP and its main component, quercetin (Q), were evaluated in vitro against 57 oral streptococci by a broth micro-dilution method. The biofilm formation was assessed through the crystal violet staining and MTT assays. The impact of AEEP and Q anti-proliferative effect were evaluated on the fibroblast as normal and cancer cell lines (KB and A431). Results The Q concentration in the composition of AEEP was 6.9% of all its components. The findings indicated that the AEEP and Q were efficient against the cariogenic bacteria and were able to inhibit the S.mutans biofilm adherence at a sub-MIC concentration. Moreover, electron micrographs indicated the inhibition of biofilms compared to control biofilms. In addition, the AEEP and Q indicated a dose-dependent cytotoxic effect on A431 and KB cell lines. On the contrary, they had no cytotoxic effect on fibroblast cells. Conclusion The results indicated that the synergistic impact of main components of AEEP was related to the inhibition of the cancer cell proliferation, cariogenic bacteria and oral biofilm formation. It may play a promising role in the complementary medicine and, it is suggested to be used as food additives.

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THE CHEMISORPTION-RELEASE AND ANTIBACTERIAL POTENTIAL STUDIES OF CIPROFLOXACIN FROM HYDROXYAPATITE-BASED IMPLANTS

Journal of Science and Arts ◽

10.46939/j.sci.arts-20.3-b04 ◽

2020 ◽

Vol 20 (3) ◽

pp. 731-738

Author(s):

RENATA-MARIA VARUT ◽

VALENTIN MANDA ◽

OANA GINGU ◽

GABRIELA SIMA ◽

CRISTINA TEISANU ◽

...

Keyword(s):

Bacterial Adhesion ◽

Biofilm Formation ◽

Pathogenic Bacteria ◽

Inhibitory Concentration ◽

Antibacterial Effect ◽

Antibiotic Concentration ◽

Implant Surface ◽

Antibiotic Release ◽

Calcium Fructoborate ◽

Antibacterial Potential

Implant infections are the result of bacterial adhesion to the implant surface and subsequent biofilm formation at the implant site. A sustained and high antibiotic concentration over minimal inhibitory concentration (MIC) of pathogenic bacteria at the implant site is expected to inhibit bacterial adhesion, colonization, and biofilm formation. In the present study we performed implants based on hydroxyapatite (HAp), HAp reinforced with titanium particles (HAp/Ti) and Hap/Ti with added calcium fructoborate (CaFb) by chemisorption deposition method (HAp/Ti/CaFb). The implants were immersed in ciprofloxacin (CP) solution for 24 hours, then was determined the release profile of antibiotic for 14 days and the antibacterial effect of the three types of composite. The period of antibiotic release may be considered as sufficient to support osteointegration under antibacterial protection.

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