Number Of Biofilm Cells Research Articles

Unveiling the Antifouling Performance of Different Marine Surfaces and Their Effect on the Development and Structure of Cyanobacterial Biofilms.

Since biofilm formation by microfoulers significantly contributes to the fouling process, it is important to evaluate the performance of marine surfaces to prevent biofilm formation, as well as understand their interactions with microfoulers and how these affect biofilm development and structure. In this study, the long-term performance of five surface materials—glass, perspex, polystyrene, epoxy-coated glass, and a silicone hydrogel coating—in inhibiting biofilm formation by cyanobacteria was evaluated. For this purpose, cyanobacterial biofilms were developed under controlled hydrodynamic conditions typically found in marine environments, and the biofilm cell number, wet weight, chlorophyll a content, and biofilm thickness and structure were assessed after 49 days. In order to obtain more insight into the effect of surface properties on biofilm formation, they were characterized concerning their hydrophobicity and roughness. Results demonstrated that silicone hydrogel surfaces were effective in inhibiting cyanobacterial biofilm formation. In fact, biofilms formed on these surfaces showed a lower number of biofilm cells, chlorophyll a content, biofilm thickness, and percentage and size of biofilm empty spaces compared to remaining surfaces. Additionally, our results demonstrated that the surface properties, together with the features of the fouling microorganisms, have a considerable impact on marine biofouling potential.

Fabrication of Photoactive Electrospun Cellulose Acetate Nanofibers for Antibacterial Applications

The aim of the study was to investigate the process of electrostatic fabrication of cellulose acetate (CA) nanofibers containing methylene blue (MB) as a photosensitizer. The electrical, physicochemical, and biocidal properties of the prepared material were given. CA nanofibers were prepared by electrospinning method using a solvent mixture of acetone and distilled water (9:1 vv−1) and different concentrations of CA (i.e., 10–21%). Additionally, methylene blue was implemented into the polymer solution with a CA concentration of 17% to obtain fibers with photo-bactericidal properties. Pure electrospun CA fibers were more uniform than fibers with MB (i.e., ribbon shape). Fiber diameters did not exceed 900 nm for the tested polymer solutions and flow rate below 1.0 mL h−1. The polymer properties (i.e., concentration, resistivity) and other parameters of the process (i.e., flow rate, an applied voltage) strongly influenced the size of the fibers. Plasma treatment of nanofibers resulted in reduced biofilm formation on their surface. The results of photo-bactericidal activity (i.e., up to 180 min) confirmed the high efficiency of inactivation of Staphylococcus aureus cells using fibers containing methylene blue (i.e., with and without plasma treatment). The most effective reduction in the number of biofilm cells was equal to 99.99 ± 0.3%.

Influence of sugars on biofilm formation of Staphylococcus epidermidis

The problem of biofilm formation by clinical strains of opportunistic bacteria is one of the most significant for medicine, because in a state of biofilm bacteria become more resistant to environmental factors, including antibiotics, a situation that can cause failure of treatment. Among opportunistic pathogens staphylococci are of special interest. Knowledge about the peculiarities of biofilm formation of these strains, in particular the polysaccharide biosynthesis, can be used for creation of a strategy of prophylaxis of different lesions that bind with staphylococci. The effect of different concentrations of the most widespread sugars (glucose, sucrose, lactose, galactose) on the activity of biofilm formation by strains of Staphylococcus epidermidis was investigated. Strains of S. epidermidis (n = 7) were isolated from the reproductive tract of women with dysbiosis. The cultures were grown in universal synthetic media with concentration of one of the listed sugars (0.5–3.0%) during 72 h. Results were obtained colorimetrically. We studied the number of cells in biofilm and the index of biofilm formation. The largest number of cells in the biofilm was observed when the culture incubated in a medium with 2.0% of glucose (increase of 25.3 times compared to control). The amount of CFU in the control biofilm was 9.96 lg CFU/mL. The glucose concentration of 3.0% inhibited the biofilm formation: the number of cells in the biofilm was 569 times less compared to the control. The highest value of biofilm formation index was 7.2, which was 1.3 times higher than the control (5.4). In the presence of lactose and galactose in nutrient medium in concentrations from 1.0% a decrease in the number of cells and biofilm formation index were observed. The received data show that process of biofilm formation is significantly dependent on external sources of sugars, which can indicate the possibility of their use as antibiofilm drug compounds, which inhibit membrane transport of sugars in bacteria.

The association between initial adhesion and cyanobacterial biofilm development.

Although laboratory assays provide valuable information about the antifouling effectiveness of marine surfaces and the dynamics of biofilm formation, they may be laborious and time-consuming. This study aimed to determine the potential of short-time adhesion assays to estimate how biofilm development may proceed. The initial adhesion and cyanobacterial biofilm formation were evaluated using glass and polymer epoxy resin surfaces under different hydrodynamic conditions and were compared using linear regression models. For initial adhesion, the polymer epoxy resin surface was significantly associated with a lower number of adhered cells compared with glass (-1.27 × 105cells.cm-2). Likewise, the number of adhered cells was significantly lower (-1.16 × 105cells.cm-2) at 185 than at 40rpm. This tendency was maintained during biofilm development and was supported by the biofilm wet weight, thickness, chlorophyll a content and structure. Results indicated a significant correlation between the number of adhered and biofilm cells (r = 0.800, p < 0.001).Moreover, the number of biofilm cells on day 42 was dependent on the number of adhered cells at the end of the initial adhesion and hydrodynamic conditions (R2 = 0.795, p < 0.001).These findings demonstrate the high potential of initial adhesion assays to estimate marine biofilm development.

The biofilm-forming ability of six Bifidobacterium strains on grape seed flour

Biofilms are aggregates of microorganisms that adhere to the surface of certain substances and produce extracellular polymers to improve their resistance to stress. However, only some Bifidobacterium strains can form biofilms on abiotic surfaces. The study evaluated the biofilm-forming abilities of six Bifidobacterium strains on grape seed flour (GSF) with a particle size of 80–120 μm. Bifidobacterium animalis, Bifidobacterium bifidum, and Bifidobacterium adolescentis formed weak biofilms, whereas Bifidobacterium pseudo, Bifibacterium breve, and Bifidobacterium longum formed strong biofilms on GSF. B. pseudo cells formed biofilm particles with a diameter of 2 mm, and their cell numbers reached 2.04 × 109 CFU/g at 32 h. There were almost no viable cells in the culture without GSF after 60 h; however, the number of biofilm cells on GSF was still over 106 CFU/g, indicating that biofilms improve cell survival. Field-emission scanning electron microscopy revealed the following stages in the bifidobacterial biofilm formation process: adsorption to GSF, secretion of extracellular substance, biofilm maturation and dispersal, and cell death. Collectively, these results strongly suggest that GSF is beneficial for Bifidobacterial biofilm growth, providing a theoretical basis for the production of highly resistant probiotics in the food industry.

Heterogeneous response of Geobacillus stearothermophilus biofilms to calcium

Geobacillus stearothermophilus contamination of milk powder products originates from bacteria released from biofilms in dairy manufacturing plants. Calcium, a major component of milk minerals, was previously found to affect G. stearothermophilus biofilm growth on surfaces exposed to milk formulations. This study endeavoured to reproduce the effect of calcium on biofilm formation of several G. stearothermophilus dairy isolates and one reference strain in modified TSB media and gain more insights into the role of calcium in biofilm formation of G. stearothermophilus. Calcium presence increased biofilm cell numbers of strain A1, but reduced cell numbers of the reference strain and showed no significant effect on strain P3. The quantity of extracellular polymeric substances (EPS), in particular extracellular protein, varied between strains. Unlike the consistent effect of calcium in milk formulations found previously, current findings suggest that strain to strain differences exists in biofilm formation of G. stearothermophilus in the presence of calcium.

Eradication of Acinetobacter baumannii Planktonic and Biofilm Cells Through Erythrosine-Mediated Photodynamic Inactivation Augmented by Acetic Acid and Chitosan.

Photodynamic inactivation (PDI) is an attractive treatment modality for multidrug-resistant bacterial infections. The effectiveness of photosensitization by anionic photosensitizers such as erythrosine B can be further enhanced by the addition of biological or chemical molecules. This study aimed to investigate of the enhancement effect of acetic acid and chitosan on erythrosine-mediated PDI of Acinetobacter baumannii in planktonic and biofilm forms. The planktonic cell growth of three A. baumannii strains was subjected to PDI by using erythrosine B (50 µM) in 0.01% acetic acid and green laser light (530 nm) at fluence of 40 J/cm2. The phototoxic effect of erythrosine B (100 µM) in combination with chitosan (12.5 mg/ml) (in a solution of acetic acid) at fluence of 80 J/cm2 on biofilms was also evaluated. Finally, the cytotoxicity and phototoxicity of the mentioned mixture were assessed on human fibroblasts. Planktonic cells of all three studied A. baumannii strains were almost eradicated by erythrosine B-mediated PDI in the presence of acetic acid. Also, PDI combined with chitosan resulted in a marked decrease in the number of viable biofilm cells (> 3 log10 CFU). At the same experimental conditions, only 15% of the fibroblasts were photoinactivated. The results showed that PDI by using erythrosine B in acetic acid is very effective against A. baumannii planktonic cells and could eliminate them significantly. Also, chitosan enhanced the anti-biofilm efficacy of erythrosine B-mediated PDI against A. baumannii, suggesting that combination therapy may be useful in targeting biofilms.

Phosphate Dosing in Drinking Water Distribution Systems Promotes Changes in Biofilm Structure and Functional Genetic Diversity.

Water utilities treat drinking water by adding phosphate to prevent metal dissolution from water pipe work systems and particularly lead poisoning. Phosphate can be a limiting nutrient for microbial biofilms in DWDS, yet its effects on these microbial consortia are not well understood. This research presents results from phosphate dosing experiments using a real scale chlorinated DWDS, comparing standard phosphate concentrations of United Kingdom drinking water (1 mgP/L) with a double dose (2 mgP/L) commonly used in plumbosolvency treatment. Biofilm development during phosphate treatment experiments was monitored using a holistic approach by combining metagenomics analysis, flow cytometry and SEM characterisation. The increase of phosphate levels in drinking water, reduced biofilm cell numbers and promoted the presence of poorly distributed biofilms on inner pipe surfaces. Metagenomics analysis using genetic markers (16S rRNA and ITS2) showed that phosphate influenced biofilm community structure, particularly fungal composition. Whole metagenome sequencing showed that phosphate enrichment favoured the presence of sequencing reads associated to ATPases, ion transporters and DNA-interacting proteins, whilst reads associated to nitrogen metabolism were predominant in control samples. This research brings new knowledge regarding the influence of phosphate treatment on the composition and structure of biofilms within DWDS, and the implications that this might have for the management of these systems.

Cell death and biomass reduction in biofilms of multidrug resistant extended spectrum β-lactamase-producing uropathogenic Escherichia coli isolates by 1,8-cineole.

Escherichia coli is the most frequent agent of urinary tract infections in humans. The emergence of uropathogenic multidrug-resistant (MDR) E. coli strains that produce extended spectrum β-lactamases (ESBL) has created additional problems in providing adequate treatment of urinary tract infections. We have previously reported the antimicrobial activity of 1,8-cineole, one of the main components of Rosmarinus officinalis volatile oil, against Gram negative bacteria during planktonic growth. Here, we evaluated the antibiofilm activity of 1,8-cineole against pre-formed mature biofilms of MDR ESBL-producing uropathogenic E. coli clinical strains by carrying out different technical approaches such as counting of viable cells, determination of biofilm biomass by crystal violet staining, and live/dead stain for confocal microscopy and flow cytometric analyses. The plant compound showed a concentration- and time-dependent antibiofilm activity over pre-formed biofilms. After a 1 h treatment with 1% (v/v) 1,8-cineole, a significant decrease in viable biofilm cell numbers (3-log reduction) was observed. Biofilms of antibiotic-sensitive and MDR ESBL-producing E. coli isolates were sensitive to 1,8-cineole exposure. The phytochemical treatment diminished the biofilm biomass by 48-65% for all four E. coli strain tested. Noteworthy, a significant cell death in the remaining biofilm was confirmed by confocal laser scanning microscopy after live/dead staining. In addition, the majority of the biofilm-detached cells after 1,8-cineole treatment were dead, as shown by flow cytometric assessment of live/dead-stained bacteria. Moreover, phytochemical-treated biofilms did not fully recover growth after 24 h in fresh medium. Altogether, our results support the efficacy of 1,8-cineole as a potential antimicrobial agent for the treatment of E. coli biofilm-associated infections.

Variations in biofilms harbouring Listeria monocytogenes in dual and triplex cultures with Pseudomonas fluorescens and Lactobacillus plantarum produced under a model system of simulated meat processing conditions, and their resistance to benzalkonium chloride

Contamination of foods with Listeria monocytogenes is a frequent occurrence that results in expensive recalls and on occasion, deadly outbreaks of foodborne listeriosis. Biofilm production is thought to play a key role in the persistence of these bacteria in food processing facilities. To date, research on L. monocytogenes biofilms has predominantly involved pure cultures grown under conditions that do not represent actual food processing environments. In nature, multiple microbial species comprise biofilm communities that develop under constantly changing conditions. This study investigated changes in the characteristics of biofilms harbouring L. monocytogenes (Lm) when in the presence of two different commensal bacteria, Lactobacillus plantarum (Lp) and Pseudomonas fluorescens (Pf), commonly isolated from ready-to-eat (RTE) meat products. Using CDC biofilm reactors, four different strain combinations (Lm monoculture; Lm + Lp; Lm + Pf; and Lm + Lp + Pf) were grown on stainless steel (SS) and high-density-polyethylene (PE-HD) coupons following a dynamic 12-day regimen of simulated meat processing conditions (SMPC) that exposed cells to periods of nutrient cycling and starvation. Viable numbers of L. monocytogenes 568 (Lm568) in pure culture biofilms at the end of the regimen were 5.8 and 6.4 log10 CFU/cm2 on SS and PE-HD, respectively. In dual cultures with P. fluorescens (ATCC 13525), Lm568 biofilm cell numbers rose by 1.0 and 1.8 log on SS and PE-HD, respectively. Enhanced L.monocytogenes numbers were also observed in the triplex biofilms with Lb. plantarum and P. fluorescens (ATCC 1024) (1.4 and 0.5 log for SS and PE-HD, respectively), whereas the same trend was not observed when L. monocytogenes was paired with Lb. plantarum. Imaging biofilms by scanning electron microscopy (SEM) revealed important architectural changes to the biofilms related to the specific strain combinations. Integration of L.monocytogenes cells into P. fluorescens biofilms significantly (P < 0.05) increased the survival of the pathogen to benzalkonium chloride. These results demonstrate the important role other commensal bacteria play in aiding the survival of foodborne pathogens; hence these factors must be considered when evaluating the effectiveness of biofilm mitigation strategies.

Combination of Miconazole and Domiphen Bromide Is Fungicidal against Biofilms of Resistant Candida spp.

The occurrence and recurrence of mucosal biofilm-related Candida infections, such as oral and vulvovaginal candidiasis, are serious clinical issues. Vaginal infections caused by Candida spp., for example, affect 70 to 75% of women at least once during their lives. Miconazole (MCZ) is the preferred topical treatment against these fungal infections, yet it has only moderate antibiofilm activity. Through screening of a drug-repurposing library, we identified the quaternary ammonium compound domiphen bromide (DB) as an MCZ potentiator against Candida biofilms. DB displayed synergistic anti-Candida albicans biofilm activity with MCZ, reducing the number of viable biofilm cells 1,000-fold. In addition, the MCZ-DB combination also resulted in significant killing of biofilm cells of azole-resistant C. albicans, C. glabrata, and C. auris isolates. In vivo, the MCZ-DB combination had significantly improved activity in a vulvovaginal candidiasis rat model compared to that of single-compound treatments. Data from an artificial evolution experiment indicated that the development of resistance against the combination did not occur, highlighting the potential of MCZ-DB combination therapy to treat Candida biofilm-related infections.

Experimental Assessment of the Performance of Two Marine Coatings to Curb Biofilm Formation of Microfoulers

Biofilms formed on submerged marine surfaces play a critical role in the fouling process, causing increased fuel consumption, corrosion, and high maintenance costs. Thus, marine biofouling is a major issue and motivates the development of antifouling coatings. In this study, the performance of two commercial marine coatings, a foul-release silicone-based paint (SilRef) and an epoxy resin (EpoRef), was evaluated regarding their abilities to prevent biofilm formation by Cyanobium sp. and Pseudoalteromonas tunicata (common microfoulers). Biofilms were developed under defined hydrodynamic conditions to simulate marine settings, and the number of biofilm cells, wet weight, and thickness were monitored for 7 weeks. The biofilm structure was analyzed by confocal laser scanning microscopy (CLSM) at the end-point. Results demonstrated that EpoRef surfaces were effective in inhibiting biofilm formation at initial stages (until day 28), while SilRef surfaces showed high efficacy in decreasing biofilm formation during maturation (from day 35 onwards). Wet weight and thickness analysis, as well as CLSM data, indicate that SilRef surfaces were less prone to biofilm formation than EpoRef surfaces. Furthermore, the efficacy of SilRef surfaces may be dependent on the fouling microorganism, while the performance of EpoRef was strongly influenced by a combined effect of surface and microorganism.

Characteristics of a Series of Three Bacteriophages Infecting Salmonella enterica Strains.

Molecular and functional characterization of a series of three bacteriophages, vB_SenM-1, vB_SenM-2, and vB_SenS-3, infecting various Salmonella enterica serovars and strains is presented. All these phages were able to develop lytically while not forming prophages. Moreover, they were able to survive at pH 3. The phages revealed different host ranges within serovars and strains of S. enterica, different adsorption rates on host cells, and different lytic growth kinetics at various temperatures (in the range of 25 to 42 °C). They efficiently reduced the number of cells in the bacterial biofilm and decreased the biofilm mass. Whole genome sequences of these phages have been determined and analyzed, including their phylogenetic relationships. In conclusion, we have demonstrated detailed characterization of a series of three bacteriophages, vB_SenM-1, vB_SenM-2, and vB_SenS-3, which reveal favorable features in light of their potential use in phage therapy of humans and animals, as well as for food protection purposes.

Modeling the elimination of mature biofilms formed by Staphylococcus aureus and Salmonella spp. Using combined ultrasound and disinfectants

Biofilm formation by foodborne pathogens on food processing surfaces has contributed to numerous disease outbreaks and food recalls. We evaluated the following strategies for elimination of mature biofilm formed by Staphylococcus aureus and Salmonella spp. on stainless steel surfaces: acidic electrolyzed water (AEW), ozone water (OW), or ultrasound (40 kHz) alone, and combinations of ultrasound and disinfectants. The dynamics of elimination by combinations were determined using the Weibull and biphasic models. Treatment with AEW alone reduced the number of biofilm cells by approximately 3.0 log cfu/cm2, whereas less than 0.8 log cfu/cm2 of cells reduction was observed in biofilm exposed to OW or ultrasound alone, even with treatment for 20 min. The combination of AEW and ultrasound produced an obvious synergistic effect on biofilm reduction, achieving approximately 4.8 log cfu/cm2 reduction in Salmonella spp. biofilm. Interestingly, the biphasic model was a better fit than the Weibull model for the elimination process of mature biofilm formed by both pathogens and subjected to a combination of ultrasound and AEW, as determined by smaller values of the statistical parameters RMSE and AIC, although both models could evaluate the dynamic processes. Our findings indicated that a combination of ultrasound and AEW could effectively reduce the biofilm formed by pathogens on food contact surfaces, and that the biphasic model could predict the number of residual cells after biofilm exposure to this intervention approach.

Gardnerella vaginalis Enhances Atopobium vaginae Viability in an in vitro Model.

Bacterial vaginosis (BV) is the most common vaginal infection among women of reproductive age. A hallmark of BV is the presence of a highly structured polymicrobial biofilm on the vaginal epithelium, presumably initiated by facultative anaerobes of the genus Gardnerella, which then becomes a scaffold for other species to adhere to. One of the species often found incorporated in Gardnerella mediated biofilms is Atopobium vaginae. Interestingly, A. vaginae is very rarely found without the presence of Gardnerella. However, not much is known regarding the interactions between A. vaginae and Gardnerella species. This study assessed biological interactions between Gardnerella vaginalis and A. vaginae. In our in vitro model, by using specific Gardnerella and A. vaginae Peptide Nucleic Acid (PNA)-Fluorescence In Situ Hybridization (FISH) probes, we confirmed that A. vaginae was able to incorporate a pre-formed G. vaginalis biofilm, accounting for up to 20% of the total number of biofilm cells. However, our findings showed that almost 92% of A. vaginae cells lost viability after 48 h of mono-species planktonic growth, but were able to maintain viability when co-cultured with Gardnerella or after pre-conditioning with cell-free supernatant of Gardnerella cultures. While the in vitro conditions are very different from the in vivo microenvironment, this study contributes to a better understanding of why A. vaginae vaginal colonization rarely occurs in the absence of Gardnerella. Overall, this highlights the importance of microbial interactions between BV-associated bacteria and demands more studies focused on the polymicrobial bacterial communities found in BV.

Biofilm formation by Staphylococcus aureus isolated from food poisoning outbreaks and effect of Butia odorata Barb. Rodr. Extract on planktonic and biofilm cells

The aims of this study were to verify the ability of S. aureus isolates from food poisoning outbreaks to form biofilm, and evaluate the susceptibility of these isolates to Butia odorata extract (BOE) and its effect on S. aureus biofilm. All 13 S. aureus isolates formed biofilm on polystyrene; and 84.6% (11/13) carried icaA and icaB genes, 69.2% (9/13) the icaD gene, and 61.5% (8/13) the icaC gene. All S. aureus isolates were susceptible to BOE, with minimum inhibitory concentration (MIC) values ranging from 2.8 to 11.4 mg.mL-1. Confocal laser scanning microscopy (CLSM) analysis showed that BOE damaged the cell membrane of S. aureus. The addition of 2 X MIC (60 min contact) decreased the number of biofilm cells by 99.99%, and using 4 X MIC (30 and 60 min contact) the reduction was 99.9%. All S. aureus isolates were biofilm formers, which causes concern because these isolates were involved in outbreaks. On the other hand, the isolates were susceptible to BOE, as the extract damaged the cell membrane of S. aureus. Besides that, BOE reduced the number of S. aureus biofilm cells, having the potential to be used as a sanitizer.

Effect of Clove and Thyme Essential Oils on Candida Biofilm Formation and the Oil Distribution in Yeast Cells.

Candida biofilm structure is particularly difficult to eradicate, since biofilm is much more resistant to antifungal agents than planktonic cells. In this context, a more effective strategy seems to be the prevention of biofilm formation than its eradication. The aim of the study was to examine whether the process of initial colonization of materials (glass, polyethylene terephthalate, polypropylene) by food-borne Candida sp. can be impeded by clove and thyme essential oils, used at their minimal inhibitory concentrations. In the presence of clove oil, 68.4–84.2% of the yeast tested showed a statistically significant reduction in biofilm formation, depending on the material. After treatment with thyme oil, statistically significant decrease in biofilm cell numbers was observed for 63.2–73.7% of yeasts. Confocal laser scanning microscopy showed diverse compounds of clove and thyme oils that were disparately located in C. albicans cell, on a cell wall and a cell membrane, in cytoplasm, and in vacuoles, depicting the multidirectional action of essential oils. However, essential oils that were used in sub-inhibitory concentration were sequestrated in the yeast vacuoles, which indicate the activation of Candida defense mechanisms by cell detoxification. Clove and thyme essential oils due to their anti-biofilm activity can be efficiently used in the prevention of the tested abiotic surfaces colonization by Candida sp.

Effects of erythromycin, nystatin and their combination on Pseudomonas aeruginosa and Candida albicans biofilms

Pseudomonas aeruginosa is frequently found to coexist and interact with Candida albicans in the polymicrobial biofilms. Considering the pathogenicity of polymicrobial biofilms in many persistent infections, this study was aimed to evaluate the combined effects of antibiotics erythromycin and nystatin on P. aeruginosa-C. albicans polymicrobial biofilm under both aerobic and anaerobic conditions. Treatment with single (erythromycin or nystatin) and combined antibiotics caused subtle reductions in the number of viable biofilm cells as well as protein and extracellular DNA content of the biofilm extracellular polymeric substance (EPS). Protein profiling demonstrated differential expression of biofilm proteins following exposure to the antimicrobial treatments. These findings suggest that erythromycin and nystatin, either single or in combination effects EPS composition (protein and extracellular DNA) as well as biofilm protein expression.

Biofilm Removal and Bacterial Re-Colonization Inhibition of a Novel Erythritol/Chlorhexidine Air-Polishing Powder on Titanium Disks.

Air-polishing with low abrasiveness powders is fast arising as a valid and mini-invasive instrument for the management of biofilm colonizing dental implants. In general, the reported advantage is the efficient removal of plaque with respect to the titanium integrity. In the present study, we evaluated the in situ plaque removal and the preventive efficacy in forestalling further infection of an innovative erythritol/chlorhexidine air-polishing powder and compared it with sodium bicarbonate. Accordingly, two peri-implantitis-linked biofilm formers, strains Staphylococcus aureus and Aggregatibacter actinomycetemcomitans, were selected and used to infect titanium disks before and after the air-polishing treatment to test its ability in biofilm removal and re-colonization inhibition, respectively. Biofilm cell numbers and viability were assayed by colony-forming unit (CFU) count and metabolic-colorimetric (2,3-Bis-(2-Methoxy-4-Nitro-5-Sulfophenyl)-2H-Tetrazolium-5-Carboxanilide) (XTT) assay. Results demonstrated that air-polishing performed with either sodium bicarbonate or erythritol/chlorhexidine was effective in reducing bacteria biofilm viability and number on pre-infected specimens, thus showing a similar ability in counteracting existing infection in situ; on the other hand, when air-polished pre-treated disks were infected, only erythritol/chlorhexidine powder showed higher post-treatment biofilm re-growth inhibition. Finally, surface analysis via mechanical profilometry failed to show an increase in titanium roughness, regardless of the powder selected, thus excluding any possible surface damage due to the use of either sodium bicarbonate or erythritol/chlorhexidine.

Control of Staphylococcus aureus biofilms by the application of single and combined treatments based in plant essential oils

Effective and environmentally-friendly alternatives to traditional disinfectants are necessary to reduce the pollution and the emergence of antimicrobial-resistant bacterial strains in food-related environments. In the present study, treatments based in single and combined applications of plant essential oils (EOs) were evaluated for control Staphylococcus aureus biofilms. EOs of Lippia sidoides, Thymus vulgaris and Pimenta pseudochariophyllus showed a higher efficacy than peracetic acid and sodium hypochlorite against S. aureus planktonic cells and 24-h-old biofilms formed on polystyrene and stainless steel under food-related conditions. High concentrations of thymol and chavibetol were detected in these EOs, as well as the presence of other antimicrobial compounds such as carvacrol, eugenol, p-cymene, limonene, α-pinene, α-terpineol, terpinen-4-oil and linalool. L. sidoides oil were particularly effective against S. aureus, but doses higher than 2.75% (v/v) were required to completely eradicate 24-h-old biofilms. Binary combinations of L. sidoides, T. vulgaris and P. pseudochariophyllus allowed decrease significantly doses required to reduce 99.99% the number of biofilm cells. Furthermore, peracetic acid increased its efficacy against S. aureus biofilms by the combined application with these EOs. The most effective treatments against S. aureus biofilms were those combining L. sidoides with T. vulgaris or peracetic acid. Therefore, these EO-based treatments can be considered as an effective and environmentally-friendly alternative to control S. aureus biofilms in food-contact surfaces.