Sessile Bacterial Cells Research Articles

Validation of the traditional antimicrobial use of the Iranian medicinal plant Thymus daenensis Celak through phytochemical characterization of its bioactive constituents

Thymus daenensis Celak (Lamiaceae family), known as denaian thyme, is an Iranian endemic plant, commonly used for its carminative, expectorant, antibacterial and antifungal properties. Previous studies report the chemical profile of the essential oil of T. daenensis aerial parts, but little is known about its non volatile constituents. Herein, phytochemical and biological investigation of the polar extract of T. daenensis aerial parts to provide further insight into traditional use of this plant has been accomplished. High-performance liquid chromatography coupled to linear ion-trap and orbitrap high-resolution mass spectrometry (LC-ESI/LTQOrbitrap/MS/MS) analysis of MeOH extract was performed to guide the isolation of specialized metabolites and successive characterization by NMR analysis. MeOH extract displayed antioxidant activity evaluated by DPPH (EC50 = 48.99 ± 1.47 μg/mL) and TEAC assay (1.37 mg/mL). Successively, the biofilm inhibitory activity of extract and isolated compounds against mature biofilms of Escherichia coli, Pseudomonas aeruginosa, Acinetobacter baumannii, Listeria monocytogenes, and Staphylococcus aureus, and their influence on the metabolism of sessile bacterial cells were evaluated. Two previously unreported thymol derivatives have been identified. The biofilm inhibitory activity of isolated compounds highlighted a promising antimicrobial action for the tested compounds. In particular, vanillic acid, (3S,5R,6R,7E,9S)-megastigm-7-ene-3,5,6,9-tetrol 3-O-β-D-glucopyranoside, thymoquinol-2-O-β-D-glucopyranoside and the never reported compound daenol resulted capable of exerting inhibitory activity vs all pathogenic strains. All compounds tested at a concentration of 10 μg/mL inhibited the metabolism of the sessile cells of E. coli at a percentage ranging between 37.7% and 77.39%, and of L.monocytogenes at a percentage ranging between 21.79% and 71.17%.

Phytochemical Investigation and Biofilm-Inhibitory Activity of Bachtiari Savory (Satureja bachtiarica Bunge) Aerial Parts.

Satureja bachtiarica is an endemic plant from the Lamiaceae family, growing in the Zagros mountain range in Iran. Even if S. bachtiarica is reported to possess many biological activities, little is known about its chemical composition. For this reason, in the present research, a phytochemical investigation of this species was carried out. To have a preliminary metabolite profile of S. bachtiarica, the n-BuOH extract was analyzed using LC-ESI/LTQOrbitrap/MS/MS in negative ion mode, allowing the identification of specialized metabolites belonging to flavonoid, monoterpene, indol, phenylpropanoid, phenolic, lignan, coumarin, biphenyl, and triterpene classes. The LC-MS/MS analysis guided the isolation of compounds, and their structures were characterized using spectroscopic methods including 1D- and 2D-NMR experiments and HRMSn analysis. In this way, a compound never reported before belonging to the biphenyl class was identified. Total flavonoid content of the extract along with the antioxidant activity were assessed. Based on the traditional uses of S. bachtiarica suggesting potential antibacterial properties, an evaluation of the biofilm inhibitory activity of the extract and isolated compounds against mature biofilms of Acinetobacter baumannii, Escherichia coli, Listeria monocytogenes, Pseudomonas aeruginosa, and Staphylococcus aureus, as well as their influence on the metabolism of sessile bacterial cells, was conducted. The results evidenced that some compounds including parmentin B, biphenyls, and 1-(1H-indole-3-carboxylate)-β-D-glucopyranoside might inhibit some changes occurring in the bacterial cells, which increases their virulence. In particular, biphenyl derivatives at a concentration of 80 μg/mL were capable of limiting remarkably the mature biofilms of A. baumannii and L. monocytogenes remarkably at a percentage ranging between 52.76% and 75.02%, and they reached an inhibition percentage of 69.28 % against E. coli. Biphenyl derivatives were also effective in exerting an inhibitory action against the mature biofilm of P. aeruginosa (inhibition ranging from 59.38% to 81.08%) and Staphylococcus aureus (inhibition percentage reached 82.94%). Of note, the biphenyl derivatives resulted in being capable of acting on the metabolism of the cells within the biofilm of all five pathogens.

Potential therapeutic benefits of unconventional oils: assessment of the potential in vitro biological properties of some Rubiaceae, Cucurbitaceae, and Brassicaceae seed oils.

Seed oils are versatile in the food sector and for pharmaceutical purposes. In recent years, their biological properties aroused the interest of the scientific world. We studied the composition of fatty acids (FAs) and some in vitro potential therapeutic benefits of five cold-pressed commercial oils obtained from broccoli, coffee, green coffee, pumpkin, and watermelon seeds. In particular, we assayed the antioxidant activity (using diphenyl-1-picrylhydrazyl (DPPH) and azino-bis (3-ethylbenzothiazoline-6-sulfonic acid (ABTS) assays). In addition, through the fatty acid composition, we calculated the atherogenicity index (AI) and thrombogenicity index (TI) to evaluate the potential impact of such oils on cardiovascular diseases. Furthermore, we assessed the in vitro anti-inflammatory capacity of the oils (evaluated through their effectiveness in preventing protein degradation, using bovine serum albumin as protein standard) and the ability of the oils to inhibit in vitro activity of three among the essential enzymes, cholinesterases and tyrosinase, involved in the Alzheimer's and Parkinson's neurodegenerative diseases. Finally, we evaluated the capacity of the oils to inhibit the biofilm of some pathogenic bacteria. The unsaturated fatty acids greatly predominated in broccoli seed oil (84.3%), with erucic acid as the main constituent (33.1%). Other unsaturated fatty acids were linolenic (20.6%) and linoleic (16.1%) acids. The saturated fatty acids fraction comprised the palmitic (6.8%) and stearic acids (0.2%). Broccoli seed oil showed the best AI (0.080) and TI (0.16) indexes. The oils expressed a good antioxidant ability. Except for the watermelon seed oil, the oils exhibited a generally good in vitro anti-inflammatory activity, with IC50 values not exceeding 8.73 micrograms. Broccoli seed oil and green coffee seed oil showed the best acetylcholinesterase inhibitory activity; coffee seed oil and broccoli seed oil were the most effective in inhibiting butyrylcholinesterase (IC50 = 15.7 μg and 20.7 μg, respectively). Pumpkin and green coffee seed oil showed the best inhibitory activity against tyrosinase (IC50 = 2 μg and 2.77 μg, respectively). In several cases, the seed oils inhibited the biofilm formation and the mature biofilm of some gram-positive and gram-negative bacteria, with Staphylococcus aureus resulting in the most sensitive strain. Such activity seemed related only in some cases to the capacity of the oils to act on the sessile bacterial cells' metabolism, as indicated by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) colorimetric method.

The inhibition effects of Cu and Ni alloying elements on corrosion of HSLA steel influenced by Halomonas titanicae

Halomonas titanicae accelerated steel corrosion by dissimilatory Fe(III) reduction under anaerobic environments, and their adhesion was the key to achieving extracellular electron transfer between cells and Fe(III). This work investigated the inhibition effects of Cu and Ni alloying elements on corrosion of high strength low alloy (HSLA) steel affected by H. titanicae. It was found that both the addition of Cu (1.3%) and high content of Ni (7.2%) brought better corrosion resistance than the steel containing 4.8% Ni via decreasing the amount of sessile bacterial cells. And the inhibition efficiency of Cu with the lower content was stronger than that of Ni with the higher content. Biofilm inhibition mechanisms varied from Cu to Ni alloying elements, and the former was achieved via bactericidal Cu ions released from steel. While for the HSLA steel with high Ni content, the formation of nickel oxides including NiFe2O4 and Ni(OH)2 refined the grains of corrosion products and decreased the bacterial attachment.

PBP4 and PBP5 are involved in regulating exopolysaccharide synthesis during Escherichia coli biofilm formation.

Escherichia coli low-molecular-mass (LMM) Penicillin-binding proteins (PBPs) help in hydrolysing the peptidoglycan fragments from their cell wall and recycling them back into the growing peptidoglycan matrix, in addition to their reported involvement in biofilm formation. Biofilms are external slime layers of extra-polymeric substances that sessile bacterial cells secrete to form a habitable niche for themselves. Here, we hypothesize the involvement of Escherichia coli LMM PBPs in regulating the nature of exopolysaccharides (EPS) prevailing in its extra-polymeric substances during biofilm formation. Therefore, this study includes the assessment of physiological characteristics of E. coli CS109 LMM PBP deletion mutants to address biofilm formation abilities, viability and surface adhesion. Finally, EPS from parent CS109 and its ΔPBP4 and ΔPBP5 mutants were purified and analysed for sugars present. Deletions of LMM PBP reduced biofilm formation, bacterial adhesion and their viability in biofilms. Deletions also diminished EPS production by ΔPBP4 and ΔPBP5 mutants, purification of which suggested an increased overall negative charge compared with their parent. Also, EPS analyses from both mutants revealed the appearance of an unusual sugar, xylose, that was absent in CS109. Accordingly, the reason for reduced biofilm formation in LMM PBP mutants may be speculated as the subsequent production of xylitol and a hindrance in the standard flow of the pentose phosphate pathway.

Antimicrobial effectiveness of etidronate powder (Dual Rinse® HEDP) and two EDTA preparations against Enterococcus faecalis: a preliminary laboratory study.

This study aimed to evaluate the antimicrobial and antibiofilm activity of two chelating agents: ethylenediaminetetraacetic acid (EDTA) combined or not with detergents, and etidronic acid combined with sterile saline. The bacterial inhibitory and bactericidal concentrations (MIC and MBC, respectively) were determined on Enterococcus faecalis ATCC 4083 strain. Antimicrobial tests were performed on a biofilm model after treatment with the chelating agents at different times (1, 3, and 5min) using a biofilm eradication concentration (MBEC) and confocal laser scanning microscope (CLSM) assays. Quantification of cell biomass and percentage of live and dead cells in the biomass was assessed for each group. The normality of the distributions for each variable was assessed using the D'Agostino and Pearson's omnibus normality test. The comparison of bacterial viability among groups and between any two groups was performed using the non-parametric Kruskal-Wallis one-way analysis of variance and the Dunn's test, respectively. No significant between-group difference was observed regarding biomass reduction. On the other hand, EDTA combined with detergents displayed a substantial increase of the dead bacteria ranging between 35 and 43%; whereas, the number of cells killed in the control group and in the other treated groups always ranged between 1 and 6%, at all experimental times. The addition of detergents to EDTA can improve its anti-biofilm activity by reducing EPS production and enhancing the killing of sessile bacterial cells. Clinical relevance EDTA presents a relevant antimicrobial activity when combined with surface-active agents.

Targeting the Bacterial Membrane with a New Polycyclic Privileged Structure: A Powerful Tool To Face Staphylococcus aureus Infections

In this paper, a small series of anthracene-maleimide-based compounds was prepared and evaluated to assess the antimicrobial potential of this polycyclic core, a scaffold previously unexplored for new antibiotic development. Some of the new compounds showed appreciable anti-Staphylococcus aureus activity, together with good safety profiles. In particular, compound 13 proved to be the most promising of the series, showing remarkable antimicrobial activity toward planktonic and sessile bacterial cells within a mature preformed biofilm. The mechanism of action seems to be related to the ability of this compound to interfere with bacterial membrane functionality, probably through the targeting of key enzymes responsible for membrane redox homeostasis and energy production. The data reported confirm the ability of this polycyclic nucleus to behave as a new "privileged structure", suitable to be further exploited in the antimicrobial field.

Simultaneous Antibacterial and Antibiofilm Activities of three Types of Silver Nanoparticles against Human Bacterial Pathogens.

Emergence of antibiotic resistant bacteria result in an approximately 550,000 deaths from bacterial infections annually. Several studies reported interesting antibacterial and anti-biofilm activities of silver nanoparticles synthesized by different physical, chemical and biological methods as an alternative to antibiotics. However, the resultant nanoparticles were varying in size, shape, and had different antibacterial and antibiofilm activities with different concentrations. Moreover, previous studies had investigated the antibacterial and antibiofilm activities in separate protocols and didn’t investigate the real-time or dual effect of silver nanoparticles on both planktonic and sessile cells within single protocol. This study aimed to synthesize silver nanoparticles through three methods and analyzing the simultaneous antibacterial and antibiofilm activities against planktonic and sessile bacterial cells. Three methods were applied to analyze silver nanoparticles and used to investigate the dual effect against bacterial biomass and biofilm formation. The results showed that synthesized silver nanoparticles cause significant inhibition to bacterial cell biomass and bacterial biofilm formation when compared with controls at low concentration. Significant Higher antibiofilm activity than antibacterial activity was observed at very low concentration (0.0125 µg/ ml). The antibacterial and antibiofilm activity do not differ according to the type of bacteria. Whereas, the antibacterial effect differs significantly according to the methods of silver nanoparticles synthesis.

Elucidating the effect of anti-biofilm activity of bioactive compounds extracted from plants

Biofilms are dense population of sessile bacterial cells that adhere to the surface, forming a matrix composed of exopolysaccharide, proteins and DNA. This matrix is termed as extracellular polymeric substance and provides stability to the cells adhering to it to form biofilms. It also provides nutrients and thus helps in the pathogenesis of biofilm-associated infections and resistance. Biofilms promote bacterial persistence by resisting host immune responses and antibiotic treatment. Antibiotics are rendered ineffective when biofilms form due to their relative impermeability, the variable physiological status of microorganisms, and subpopulations of persistent strains. Another factor that results in the development of antibiotic resistance within the biofilm is the adaptations that take place within the genes present in the cells dwelling within the biofilm. These adaptations decrease the sensitivity of the bacterial cells toward the antibiotics and develop resistance. Hence, an alternative antimicrobial strategy of making use of plant-based products has been observed to be useful to cure various ailments, as compared to conventional therapy. In this review, we have listed the various biofilm-forming bacteria and the bioactive compounds being produced from the aerial parts of plants having antibiofilm activity and evaluated them against different biofilm-producing bacterial strains.

MAPLE fabricated coatings based on magnetite nanoparticles embedded into biopolymeric spheres resistant to microbial colonization

The aim of this study was to obtain improved coatings for advanced surfaces with increased biocompatibility and resistance to microbial colonization and biofilm formation. The prepared magnetite nanoparticles functionalized with gentamicin (Fe3O4@G) have been embedded into poly(lactic-co-glycolic acid) (PLGA) spheres by oil-in-water emulsion. The PLGA-Fe3O4@G spheres were deposited on glass and silicone surfaces by Matrix Assisted Pulsed Laser Evaporation (MAPLE) technique. The obtained thin coatings were analyzed by Scanning Electron Microscopy (SEM) and Infrared Microscopy (IRM). The antimicrobial and antibiofilm efficiency of coatings was tested with respect to Gram-positive (Staphylococcus aureus) and Gram-negative (Pseudomonas aeruginosa) clinical strains by viable cells counts assay, performed at different time intervals. The obtained results proved that coatings based on PLGA-Fe3O4@G spheres exhibited an efficient antimicrobial activity against both adherent and sessile bacterial cells. Besides their excellent anti-adherence and antibiofilm effect, the obtained MAPLE-deposited coatings were highly biocompatible, allowing the normal development and growth of cultured human amniotic fluid stem cells. This approach could be successfully applied for the optimization of medical surfaces in order to control and prevent microbial colonization and further development of biofilm associated infections.

Genotypic and Phenotypic Characteristics Associated with Biofilm Formation by Human Clinical Escherichia coli Isolates of Different Pathotypes.

Bacterial biofilm formation is a widespread phenomenon and a complex process requiring a set of genes facilitating the initial adhesion, maturation, and production of the extracellular polymeric matrix and subsequent dispersal of bacteria. Most studies on Escherichia coli biofilm formation have investigated nonpathogenic E. coli K-12 strains. Due to the extensive focus on laboratory strains in most studies, there is poor information regarding biofilm formation by pathogenic E. coli isolates. In this study, we genotypically and phenotypically characterized 187 human clinical E. coli isolates representing various pathotypes (e.g., uropathogenic, enteropathogenic, and enteroaggregative E. coli). We investigated the presence of biofilm-associated genes ("genotype") and phenotypically analyzed the isolates for motility and curli and cellulose production ("phenotype"). We developed a new screening method to examine the in vitro biofilm formation ability. In summary, we found a high prevalence of biofilm-associated genes. However, we could not detect a biofilm-associated gene or specific phenotype correlating with the biofilm formation ability. In contrast, we did identify an association of increased biofilm formation with a specific E. coli pathotype. Enteroaggregative E. coli (EAEC) was found to exhibit the highest capacity for biofilm formation. Using our image-based technology for the screening of biofilm formation, we demonstrated the characteristic biofilm formation pattern of EAEC, consisting of thick bacterial aggregates. In summary, our results highlight the fact that biofilm-promoting factors shown to be critical for biofilm formation in nonpathogenic strains do not reflect their impact in clinical isolates and that the ability of biofilm formation is a defined characteristic of EAEC.IMPORTANCE Bacterial biofilms are ubiquitous and consist of sessile bacterial cells surrounded by a self-produced extracellular polymeric matrix. They cause chronic and device-related infections due to their high resistance to antibiotics and the host immune system. In nonpathogenic Escherichia coli, cell surface components playing a pivotal role in biofilm formation are well known. In contrast, there is poor information for their role in biofilm formation of pathogenic isolates. Our study provides insights into the correlation of biofilm-associated genes or specific phenotypes with the biofilm formation ability of commensal and pathogenic E. coli Additionally, we describe a newly developed method enabling qualitative biofilm analysis by automated image analysis, which is beneficial for high-throughput screenings. Our results help to establish a better understanding of E. coli biofilm formation.

Anti-biofilm formation of a novel stainless steel against Staphylococcus aureus

Staphylococcus aureus (S. aureus) is a bacterium frequently found proliferating on metal surfaces such as stainless steels used in healthcare and food processing facilities. Past research has shown that a novel Cu-bearing 304 type stainless steel (304CuSS) exhibits excellent antibacterial ability (i.e. against S. aureus) in a short time period (24h.). This work was dedicated to investigate the 304CuSS's inhibition ability towards the S. aureus biofilm formation for an extended period of 7days after incubation. It was found that the antibacterial rate of the 304CuSS against sessile bacterial cells reached over 99.9% in comparison with the 304SS. The thickness and sizes of the biofilms on the 304SS surfaces increased markedly with period of contact, and thus expected higher risk of bio-contamination, indicated by the changes of surface free energy between biofilm and the steel surfaces. The results demonstrated that the 304CuSS exhibited strong inhibition on the growth and adherence of the biofilms. The surface free energy of the 304CuSS after contact with sessile bacterial cells was much lower than that of the 304SS towards the same culture times. The continuously dissolved Cu(2+) ions well demonstrated the dissolution ability of Cu-rich precipitates after exposure to S. aureus solution, from 3.1ppm (2days) to 4.5ppm (7days). For this to occur, a hypothesis mechanism might be established for 304CuSS in which the Cu(2+) ions were released from Cu-rich phases that bond with extracellular polymeric substances (EPS) of the microorganisms. And these inhibited the activities of cell protein/enzymes and effectively prevented planktonic bacterial cells attaching to the 304CuSS metal surface.

On the production of glycogen by Pseudomonas fluorescens during biofilm development: an in situ study by attenuated total reflection-infrared with chemometrics

Attenuated total reflection Fourier transform infrared (ATR-FTIR) spectroscopy was used to monitor Pseudomonas fluorescens biofilms in situ, non-destructively, in real time, and under fully hydrated conditions. Changes accompanying the metabolic evolution of the sessile bacterial cells from the nascent biofilm monolayer to the beginning of the multi-layered structure in the presence of nutrients were identified via the ATR-FTIR fingerprints of the young biofilm on the ATR crystal. The ATR-FTIR spectra were analysed by classical methods (time evolution of integrated intensities and profile evolution of specific bands), and also by a multivariate curve resolution, Bayesian positive source separation, to extract the pure component spectra and their change of concentration over time occurring during biofilm settlement. This work showed clearly the overproduction of glycogen by sessile P. fluorescens, which had not previously been described by other research groups.

Perfil de sensibilidade de células sésseis e planctônicas de Escherichia coli a antimicrobianos usados no tratamento da mastite bovina

Escherichia coli é um micro-organismo altamente adaptativo e sua habilidade em formar biofilmes pode ser fundamental na resistência a tratamentos com antimicrobianos. A avaliação da concentração mínima inibitória (CMI) vem sendo utilizada para verificar a sensibilidade dos micro-organismos aos antimicrobianos. Entretanto, quando se avaliam células sésseis, a concentração do antimicrobiano requerido para erradicação do biofilme é maior do que a determinada pela CMI. Objetivou-se comparar as CMI com as concentrações mínimas de erradicação de biofilmes (CMEB) de antimicrobianos usados no tratamento da mastite em 27 isolados de E. coli produtores de biofilmes provenientes de mastite. Os isolados foram submetidos a testes de sensibilidade a antimicrobianos usados no tratamento da mastite, tanto para células planctônicas, por meio da CMI, quanto para células sésseis, pela avaliação da CMEB. Os resultados revelaram uma alta sensibilidade: apenas quatro (14,8%) isolados obtiveram valores da CMI elevados, variando de 4 a 10µg/mL, sendo classificados como resistentes. Para os demais isolados (85,2%), os valores foram menores, variando de 0,125 a 2µg/mL, classificados como sensíveis. A avaliação de CMEB indicou que a concentração dos antimicrobianos necessária para eliminar as células sésseis variou de 100µg/mL a 500µg/mL. Os valores de CMEB foram significativamente maiores nos isolados grandes e moderados produtores de biofilmes em relação aos isolados fracos produtores de biofilmes (p<0,001). Não houve correlação entre os valores de CMEB e CMI (p>0,05). A escolha da terapêutica antimicrobiana correta para o tratamento de infecções intramamárias em bovinos relacionadas com a produção de biofilmes parece exigir a aplicação de testes mais específicos. Testes de susceptibilidade antimicrobiana baseados apenas em valores de CMI mostraram-se ineficazes em determinar com precisão a susceptibilidade das células bacterianas sésseis.

Origanum vulgare subsp. hirtum essential oil prevented biofilm formation and showed antibacterial activity against planktonic and sessile bacterial cells.

Essential oils from six different populations of Origanum vulgare subsp. hirtum were compared for their antibiofilm properties. The six essential oils (A to F) were characterized by a combination of gas chromatography with flame ionization detector and gas chromatography with mass spectrometer detector analyses. All oils showed weak activity against the planktonic form of a group of Staphylococcus aureus strains and against a Pseudomonas aeruginosa ATCC 15442 reference strain. The ability to inhibit biofilm formation was investigated at sub-MIC levels of 200, 100, and 50 m g/ml by staining sessile cells with safranin. Sample E showed the highest average effectiveness against all tested strains at 50 m g/ml and had inhibition percentages ranging from 30 to 52%. In the screening that used preformed biofilm from the reference strain P. aeruginosa, essential oils A through E were inactive at 200 m g/ml; F was active with a percentage of inhibition equal to 53.2%. Oregano essential oil can inhibit the formation of biofilms of various food pathogens and food spoilage organisms.

In vitro activity of the new water-dispersible Fe3O4@usnic acid nanostructure against planktonic and sessile bacterial cells

A new water-dispersible nanostructure based on magnetite (Fe3O4) and usnic acid (UA) was prepared in a well-shaped spherical form by a precipitation method. Nanoparticles were well individualized and homogeneous in size. The presence of Fe3O4@UA was confirmed by transmission electron microscopy, Fourier transform-infrared spectroscopy, and X-ray diffraction. The UA was entrapped in the magnetic nanoparticles during preparation and the amount of entrapped UA was estimated by thermogravimetric analysis. Fabricated nanostructures were tested on planktonic cells growth (minimal inhibitory concentration assay) and biofilm development on Gram-positive Staphylococcus aureus (S. aureus), Enterococcus faecalis (E. faecalis) and Gram-negative Escherichia coli (E. coli), Pseudomonas aeruginosa (P. aeruginosa) reference strains. Concerning the influence of Fe3O4@UA on the planktonic bacterial cells, the functionalized magnetic nanoparticles exhibited a significantly improved antimicrobial activity against E. faecalis and E. coli, as compared with the Fe3O4 control. The UA incorporated into the magnetic nanoparticles exhibited a very significant inhibitory effect on the biofilm formed by the S. aureus and E. faecalis, on a wide range of concentrations, while in case of the Gram-negative microbial strains, the UA-loaded nanoparticles inhibited the E. coli biofilm development, only at high concentrations, while for P. aeruginosa biofilms, no inhibitory effect was observed. The obtained results demonstrate that the new water-dispersible Fe3O4@UA nanosystem, combining the advantages of the intrinsic antimicrobial features of the UA with the higher surface to volume ratio provided by the magnetic nanocarrier dispersible in water, exhibits efficient antimicrobial activity against planktonic and adherent cells, especially on Gram-positive strains.

Examination of nanoparticle inactivation of Campylobacter jejuni biofilms using infrared and Raman spectroscopies

To investigate inactivation effect and mechanism of zinc oxide nanoparticles (ZnO NPs) activity against Campylobacter jejuni biofilms. ZnO NPs with concentrations of 0, 0·6, 1·2 and 6mmoll(-1) were employed in antimicrobial tests against Camp.jejuni planktonic cells and biofilms. Campylobacter jejuni sessile cells in biofilms were more resistant to a low concentration of ZnO NPs when compared to planktonic cells. The ZnO NPs penetrated the extracellular polymeric substance (EPS) without damage to the EPS and directly interacted with the sessile bacterial cells, as determined using infrared spectroscopy and scanning electron microscopy. Raman spectroscopy shows alterations in quinone structures and damage to nucleic acids following Camp.jejuni treatment with ZnO NPs. The mechanism of DNA damage is most likely due to the generation of reactive oxygen species (ROS). Spectroscopic-based partial least squares regression (PLSR) models could predict the number of surviving sessile cell numbers within a bacterial biofilm (≥log 4 CFU, root mean square error of estimation <0·36) from Fourier transform infrared (FT-IR) spectral measurements. ZnO NPs were found to have antimicrobial activity against Camp.jejuni biofilms. ZnO NPs penetrated the biofilm EPS within 1h without damaging it and interacted directly with sessile cells in biofilms. Alterations in the DNA/RNA bases, which are owing to the generation of ROS, appear to result in Camp.jejuni cell death. ZnO NPs may offer a realistic strategy to eliminate Camp.jejuni biofilms in the environment.

Control of planktonic and sessile bacterial cells by essential oils

The antibacterial potential of essential oils (EOs) from Cinnamomum cassia bark and Melaleuca alternifolia and Cymbopogon flexuosus leaves was evaluated against planktonic and sessile cells of enteropathogenic Escherichia coli (EPEC) and Listeria monocytogenes. The EOs were tested singly and in different combinations of equal percentages: mixtures of two (1:1 in v/v) and three EOs (1:1:1 in v/v/v). The minimal inhibitory concentrations (MICs) were determined against planktonic cells and the anti-biofilm activity was verified against bacterial cells adhered in the wells of polystyrene microplates. These initial tests indicated the EO of C. cassia as a potential anti-biofilm agent, and their effect was studied against sessile cells of biofilms formed on stainless steel surface under agitation and static conditions. For both bacterial species, a solution containing 2% (v/v) of C. cassia EO was effective against the biofilm formed under static conditions, because the counts obtained were below the detection level of the plate count method employed. Although the biofilm of L. monocytogenes showed a decreased number of adhered cells after formation under agitating conditions (p<0.05), it was surprisingly more resistant to the EO of C. cassia than the biofilm formed under static conditions (p<0.05). All of the EOs and combinations tested presented antibacterial activity, almost against planktonic cells; however, the EO of C. cassia showed to be the most effective as a potential agent for the production of sanitizers for biofilm control in the food industries.

Cinnamon essential oil and cinnamaldehyde in the control of bacterial biofilms formed on stainless steel surfaces

The antibacterial effects of Cinnamomum cassia essential oil (EO) and cinnamaldehyde were evaluated against single- and mixed-species cultivation of enteropathogenic Escherichia coli (EPEC) and Listeria monocytogenes attached to stainless steel. A central composite rotational design with two variables and eleven assays was used to optimize the concentrations (0.00–1.00% v/v for the EO and 0.00–0.80% v/v for cinnamaldehyde) and contact times (1–21 min). The models generated were validated, and the effectiveness of C. cassia EO and cinnamaldehyde was compared with that of commercially available chemical sanitizers. Cinnamaldehyde and C. cassia EO proved to be efficient alternatives to commercial chemical sanitizers in the reduction or elimination of sessile bacterial cells. The activity of these natural compounds was, in most cases, equivalent or superior to that of the chemical sanitizers tested. However, L. monocytogenes was more resistant than EPEC to C. cassia EO and cinnamaldehyde, and the bacterial association in mixed-species biofilms made them more susceptible to these compounds.

Interaction of bacteria isolated from clinical biofilms with cardiovascular prosthetic devices and eukaryotic cells

Purpose To identify the relationships between some infectious agents implicated in cardiovascular diseases with the cellular substrate and prosthetic devices in the presence of antibiotics. Specific objectives Strains isolation and identification, comparative study of antibiotic resistance of planktonic (disk diffusion, E-test, automatic systems) and sessile (using original experimental models for in vitro development of monospecific biofilms) bacterial cells, virulence assays (adherence and invasion of HeLa cells, slime test, soluble virulence factors expression), dynamic study of biofilm development on inert substrata, under the influence of antibiotics, the influence of cellular and soluble bacterial fractions on HeLa cells (by flow cytometry and real-time PCR). Results The identified strains were isolated from different sources, the etiology being dominated by Gram-negative non-fermentative bacilli, Gram-positive cocci and yeasts, harboring invasion enzymes responsible for development of systemic infections. The isolated strains exhibited a high level of antibiotic resistance to beta-lactams, aminoglycosides and quinolones, and an evident tendency of colonizing the cellular and inert substrate, the degree of colonization depending on the physico-chemical nature of the substrate. By comparison with planktonic ones, the sessile bacterial strains expressed a changed profile of antibiotic resistance, this aspect being very important for the readjustment of the treatment and prevention of infections associated with prosthetic devices. In vitro experiments suggested that different fractions of S. aureus cultures could trigger the release of proinflammatory (TNF-α, IL-1b, IL-6) and anti-inflammatory (IL-8) cytokines and induced apoptosis in HeLa cells.