Biofilm Stages Research Articles

Biofilm Eradication and Inhibition of Methicillin-Resistant Staphylococcus Clinical Isolates by Curcumin-Chitosan Magnetic Nanoparticles.

Biofilm-producing methicillin-resistant Staphylococcus aureus (MRSA) and coagulase-negative staphylococci (MR-CoNS) pose clinical challenges in treating healthcare-associated infections. As alternative antimicrobial options are needed, in this study, we aimed to determine the effect of curcumin-chitosan magnetic nanoparticles (Cur-Chi-MNP) on the biofilms of staphylococcal clinical isolates. MRSA and CoNS clinical isolates were identified using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. Antimicrobial susceptibility testing was performed using the broth microdilutions. Nanoparticles were synthesized by the co-precipitation of magnetic nanoparticles (MNP) and encapsulated by the ionotropic gelation of curcumin (Cur) and chitosan (Chi). Biofilm inhibition and eradication by nanoparticles, with and without the addition of oxacillin (OXA), were assessed in Staphylococcus strains. Cur-Chi-MNP showed antimicrobial activity against planktonic cells of MRSA and MR-CoNS strains and inhibited MRSA biofilm. The addition of OXA to Cur-Chi-MNP increased the biofilm inhibition and eradication activity against all staphylococcal strains (P = 0.0007), and higher biofilm activity was observed in the early biofilm stages. Cur-Chi-MNP showed antimicrobial and biofilm inhibitory activities against S. aureus. Addition of OXA increased biofilm inhibition and eradication activity against all staphylococcal strains. A combination treatment of Cur-Chi-MNP and OXA could potentially be used to treat staphylococcal biofilm-associated infections in the early stages before the establishment of biofilm bacterial cells.

Metamorphosis alters concentrations of bioaccumulated pharmaceuticals and endocrine disruptors in caddisflies: Implications for transfer of aquatic–terrestrial subsidies

Abstract Aquatic insects serve as vital inter‐habitat linkages between aquatic and terrestrial ecosystems, as their life cycle includes both aquatic and terrestrial stages. They facilitate the flow of energy and nutrients and play an important role in the transport of waterborne contaminants to terrestrial environments. This study focused on the under‐investigated environmental fate of emerging contaminants (ECs), particularly pharmaceuticals (PhACs) and endocrine‐disrupting compounds (EDCs), which originate from wastewater effluents and accumulate in aquatic insects. Using an in situ study of a highly abundant caddisfly species (Trichoptera), Silo nigricornis (Pictet, 1834), inhabiting a wastewater‐impacted drainage ditch, we examined the bioaccumulation and bioamplification of PhACs and EDCs across the aquatic and terrestrial life stages of the caddisfly. We observed variations in the highest concentrations of PhACs and EDCs in different sample types, including water, biofilm and different life stages of S. nigricornis. Adult S. nigricornis exhibited the highest total concentrations of ECs among the different life stages, with 17 different ECs bioaccumulated in caddisfly tissues. Most of these ECs had higher concentrations in terrestrial adults compared to aquatic larvae and pupae. The total concentration of ECs increased significantly with development, revealing a 41% higher concentration in adult terrestrial caddisflies compared to pupae and larvae. Bioamplification factors provided insights into the increased body burden of the majority of compounds during at least one stage of metamorphosis in S. nigricornis. Evidence of bioamplification was observed in both metamorphosis stages, with certain compounds, such as antibiotics (azithromycin and tilmicosin), endocrine disruptors (TCPP – tris[1‐chloro‐2‐propyl]phosphate and parabens; methylparaben, propylparaben), consistently exhibiting increased concentrations from larval to pupal and from pupal to adult stages. The observed pattern of increasing body burden of contaminants during the caddisfly life cycle emphasises the effects of metamorphosis on the concentration of PhACs and EDCs in adult holometabolous aquatic insects. Our results emphasise the importance of adult caddisflies for the transport of waterborne contaminants such as PhACs and EDCs from aquatic to terrestrial environments. Therefore, our results emphasise the need to include adult trichopterans in the assessment of habitat quality, as their influence extends beyond the boundaries of the aquatic ecosystem.

Immobilization of biosynthesized gallium nanoparticles in Polyvinylpyrrolidone/Sodium alginate films: Potent bactericidal protection against food spoilage bacteria

The increasing threat of spoilage bacterial infections, driven by the resistance of bacteria to many antimicrobial treatments, is a significant worldwide public health problem, especially concerning food preservation. To tackle these difficulties, this research investigates the possibility of using packaging sheets that include antimicrobial agents and increasing the prolonged storage time by preventing the bioburden of foodborne pathogens. This approach uses metal nanoparticles' ability to prevent harmful bacteria that cause food spoiling. Gallium nanoparticles (GaNPs) were created using a water-based extract from Andrographis paniculata leaves as a bioreducing agent. The GaNPs were added to a film made of sodium alginate (SA) and polyvinylpyrrolidone (PVP). The study showed that incorporating GaNPs into polymer films resulted in films with a desirable contact angle and decreased water vapor permeability. Significantly, the developed films demonstrated increased efficiency against E.coli O157 compared to other species. Also, it exhibited increased vulnerability to bacterial strains at the biofilm stage, surpassing PVP-SA/GaNPs-0. Remarkably, the toxicity tests showed that the films exhibited no cytotoxicity. Overall, the films indicated their potential for avoiding bacterial bioburden, prolonging the shelf life of perishable products, and contributing to diverse antimicrobial applications in the food industry.

Decoding the role of oxidative stress resistance and alternative carbon substrate assimilation in the mature biofilm growth mode of Candida glabrata

BackgroundBiofilm formation is viewed as a vital mechanism in C. glabrata pathogenesis. Although, it plays a significant role in virulence but transcriptomic architecture and metabolic pathways governing the biofilm growth mode of C. glabrata remain elusive. The present study intended to investigate the genes implicated in biofilm growth phase of C. glabrata through global transcriptomic approach.ResultsFunctional analysis of Differentially expressed genes (DEGs) using gene ontology and pathways analysis revealed that upregulated genes are involved in the glyoxylate cycle, carbon-carbon lyase activity, pre-autophagosomal structure membrane and vacuolar parts whereas, down- regulated genes appear to be associated with glycolysis, ribonucleoside biosynthetic process, ribosomal and translation process in the biofilm growth condition. The RNA-Seq expression of eight selected DEGs (CgICL1, CgMLS1, CgPEP1, and CgNTH1, CgERG9, CgERG11, CgTEF3, and CgCOF1) was performed with quantitative real-time PCR (RT-qPCR). The gene expression profile of selected DEGs with RT-qPCR displayed a similar pattern of expression as observed in RNA-Seq. Phenotype screening of mutant strains generated for genes CgPCK1 and CgPEP1, showed that Cgpck1∆ failed to grow on alternative carbon substrate (Glycerol, Ethanol, Oleic acid) and similarly, Cgpep1∆ unable to grow on YPD medium supplemented with hydrogen peroxide. Our results suggest that in the absence of glucose, C. glabrata assimilate glycerol, oleic acid and generate acetyl coenzyme-A (acetyl-CoA) which is a central and connecting metabolite between catabolic and anabolic pathways (glyoxylate and gluconeogenesis) to produce glucose and fulfil energy requirements.ConclusionsThe study was executed using various approaches (transcriptomics, functional genomics and gene deletion) and it revealed that metabolic plasticity of C. glabrata (NCCPF-100,037) in biofilm stage modulates its virulence and survival ability to counter the stress and may promote its transition from commensal to opportunistic pathogen. The observations deduced from the present study along with future work on characterization of the proteins involved in this intricate process may prove to be beneficial for designing novel antifungal strategies.

Antibiofilm Efficacy and Mode of Action of Etlingera elatior Extracts Against Staphylococcus aureus

Staphylococcus aureus represents a major bacterial human pathogen that causes a wide variety of clinical manifestations. Various medicinal plants have been used to control its infection, however, the effect of Etlingera elatior on S. aureus biofilm is still uncertain. Thus, the objective of this study was to evaluate the antibacterial and antibiofilm efficacy of E. elatior extracts against S. aureus. Phytochemical screening was carried out to determine the presence of phenols, tannins, saponins, and alkaloids in different extracts (acetone, methanol, ethanol, and aqueous) of E. elatior. Antibacterial activities were determined by disk diffusion assay, minimum inhibitory concentration assay (MIC), and minimum bactericidal concentration assay (MBC) while antibiofilm activities were determined by crystal violet assay and Fourier-transform infrared (FTIR) spectroscopy. All the extracts were found to contain phenols, tannins, saponins, and alkaloids. Only acetone extract showed a high amount of saponins. Among all the extracts, acetone extract showed the widest inhibition zone (21.23±0.2 mm), lowest MIC (20 mg/mL), and lowest MBC (50 mg/mL) values. The acetone extract also showed the highest antibiofilm activities at all biofilm stages (6 hr: 12%-31%; 12 hr: 20%-36%; 18 hr: 27%-32%; 24 hr: 5%-46%). Further analysis with FTIR spectroscopy revealed spectral changes associated with proteins (1700–1400 cm-1), phospholipids, and polysaccharides (1300–700 cm-1) in S. aureus biofilm following the treatment with 200 mg/mL of E. elatior extracts. In conclusion, E. elatior is a potential source of antibacterial and antibiofilm agents to control S. aureus infections. Changes in the composition of proteins, phospholipids, and polysaccharides may mediate the biofilm inhibition by E. elatior extracts. The acetone extract of E. elatior may be useful for various applications such as antimicrobial topical cream and wound dressing.

Elimination of Pathogen Biofilms via Postbiotics from Lactic Acid Bacteria: A Promising Method in Food and Biomedicine.

Pathogenic biofilms provide a naturally favorable barrier for microbial growth and are closely related to the virulence of pathogens. Postbiotics from lactic acid bacteria (LAB) are secondary metabolites and cellular components obtained by inactivation of fermentation broth; they have a certain inhibitory effect on all stages of pathogen biofilms. Postbiotics from LAB have drawn attention because of their high stability, safety dose parameters, and long storage period, which give them a broad application prospect in the fields of food and medicine. The mechanisms of eliminating pathogen biofilms via postbiotics from LAB mainly affect the surface adhesion, self-aggregation, virulence, and QS of pathogens influencing interspecific and intraspecific communication. However, there are some factors (preparation process and lack of target) which can limit the antibiofilm impact of postbiotics. Therefore, by using a delivery carrier and optimizing process parameters, the effect of interfering factors can be eliminated. This review summarizes the concept and characteristics of postbiotics from LAB, focusing on their preparation technology and antibiofilm effect, and the applications and limitations of postbiotics in food processing and clinical treatment are also discussed.

Chestnut Honey Is Effective against Mixed Biofilms at Different Stages of Maturity.

The irresponsible overuse of antibiotics has increased the occurrence of resistant bacterial strains, which represents one of the biggest patient safety risks today. Due to antibiotic resistance and biofilm formation in bacteria, it is becoming increasingly difficult to suppress the bacterial strains responsible for various chronic infections. Honey was proven to inhibit bacterial growth and biofilm development, offering an alternative solution in the treatment of resistant infections and chronic wounds. Our studies included chestnut honey, valued for its high antibacterial activity, and the bacteria Pseudomonas aeruginosa, methicillin-resistant Staphylococcus aureus, and S. epidermidis, known to form multi-species biofilm communities. Minimum inhibitory concentrations (MIC) of chestnut honey were determined for each bacterial strain. Afterwards, the mixed bacterial biofilms were treated with chestnut honey at different stages of maturity (incubation times: 2, 4, 6, 12, 24 h). The extent of biofilm inhibition was measured with a crystal violet assay and demonstrated by scanning electron microscopy (SEM). As the incubation time increased and the biofilm became more mature, inhibition rates decreased gradually. The most sensitive biofilm was the combination MRSA-S. epidermidis, with a 93.5% inhibition rate after 2 h of incubation. Our results revealed that chestnut honey is suitable for suppressing the initial and moderately mature stages of mixed biofilms.

The microbial community of coral reefs: biofilm composition on artificial substrates under different environmental conditions

Artificial reefs are used as tools for the restoration of degrading coral reefs by providing new settlement substrates. The initial recruitment process consists of the formation of microbial biofilms shortly after deployment. The aim of the present study is to compare biofilm composition and development on artificial substrates at two different coral-reef sites. These locations differ in their environmental conditions, including their level of anthropogenic impact. Substrate and seawater samples were collected four times during the first 6 months after deployment, using a new methodology termed ‘tab-by-tab’. DNA was extracted, sequenced, and sorted for both prokaryotic (16s) and eukaryotic (18s) genes. No difference was found between the planktonic communities in the water, yet significant differences were observed in the biofilm communities on the structures at the two sites. Moreover, differences were recorded in coral recruitment rates, which are known to be affected by biofilm composition. Our findings suggest a potential role of environmental conditions in the early biofilm stages (first few months), which in turn may impact the succession and development of coral-reef communities and the success of artificial reefs as restoration tools.

In Vitro Antibacterial Activity of a Novel Acid-Activated Antimicrobial Peptide against Streptococcus mutans.

Dental caries is an oral disease associated with infection by microbial biofilm. The metabolic activity of cariogenic bacteria results in a pH decrease in the plaque biofilm, causing tooth demineralization. This acidic environment favors the growth of cariogenic bacteria that are highly resistant to strong acids, which, in turn, produce more acid resulting in a further decrease in the pH of the plaque biofilm. Therefore, the strategy of utilizing the acidic dental plaque microenvironment to prevent and treat dental caries has become a hot research topic in recent years, such as the development of pH-sensitive drug delivery systems. Design of a new acid-activated antibacterial peptide. To design and synthesis an acid targeted antimicrobial peptide with the GWHHFFHFFHFF sequence. Minimum inhibitory concentration (MIC) and minimum bacterial concentration (MBC) testing confirmed its antibacterial activity. Propidium iodide (PI) staining was used to detect nucleic acid leakage. Determination of anti-biofilm activity by biofilm inhibition assay. A phototoxicity study confirmed the phototoxicity of PPIX-P12. MIC and MBC testing confirmed that P12 possessed acid-activated anti-Streptococcus mutans activity. Bactericidal kinetic experiments and propidium iodide (PI) staining experiments showed that P12 killed planktonic S. mutans UA159 cells leading to the leakage of nucleic acids in the acidic medium. Moreover, P12 showed acid-activated anti-biofilms at the early and mature biofilm stages. P12 was conjugated with the phototherapeutic agent protoporphyrin IX (PpIX) to construct the protoporphyrin derivative PpIX-P12. In vitro experiments revealed that PpIX-P12 displayed better antibacterial activity in pH 5.5 medium than in pH 7.2 medium. In conclusion, we designed an acid-activated AMP, which had no antimicrobial activity at neutral pH, but had antimicrobial activity at an acidic pH.

Single Microcolony Diffusion Analysis in Pseudomonas aeruginosa Biofilms.

Diffusion phenomena play an important role during various stages of biofilm development. Hence, quantification of the diffusion coefficient of molecules provides important information necessary to understand the variability in diffusion profiles at different biofilm stages. Imaging fluorescence correlation spectroscopy (FCS) enables the visualization of 3D diffusion profiles of biofilms. In this protocol, we explain how to grow biofilms for investigation using imaging FCS and how to acquire and analyze the data.

Multi-omics analysis of the biofilm forming mechanism of Bifidobacterium longum

Biofilm is considered to be a protective lifestyle formed by bacteria to adapt to the environment. To study the biofilm formation mechanism of Bifidobacterium longum, we used comparative genes to find their biofilm functional genes, then exogenously added signal molecules and inhibitors to verify related genes by using transcriptome and targeted metabolome. Finally, a time series transcriptome and metabolome were performed to completly analyze the biofilm formation pathways on wheat fiber. These results indicated B. longum mainly regulated biofilm formation through AI-2/QS, cAMP, and LuxC/LuxE TCS system by regulating Tad IV pilin and extracellular substances secretion. In the initial biofilm stage of B. longum, cAMP reduced cell motility through Tad IV pilin to promote biofilm state transition and enhanced cell repair through LuxC/LuxE TCS. With increasing cell numbers, AI-2/QS promoted biofilm growth by enhancing the transport of extracellular matrix by the Tat and Sec secretion systems. During the biofilm disperse stage, AI-2/QS enhanced the stress response to external stimuli. In addition, seventeen WGCNA modules were identified and two of them positively correlated with pH value, indicating biofilm formation associated with acid stress. These results can provide a theoretical basis for the formation mechanism of Bifidobacterium biofilm on food-grade particles.

Ecological mechanisms of biofilm development in the hybrid sludge-biofilm process: Implications for process start-up and optimization

The hybrid sludge-biofilm processes have been widely applied for the construction or upgradation of biological wastewater treatment process. Ecological mechanisms of biofilm development remain unclear in the hybrid ecosystem, because of the intricate interactive effects between sludge and biofilms. Herein, the establishment principles of biofilms with distinct coexisting sludge amounts were uncovered by varying sludge retention times (SRTs) from 5 to 40 days in the hybrid process. With the increasing of SRTs, biofilm biomass decreased with the increase of suspended sludge, resulting in lower biofilm proportion. As estimated by the Gompertz growth model, the increased sludge amounts (i.e., higher SRTs of 20 and 40 days) prolonged the initial colonization stage and decreased the specific development rate of biofilms when compared to lower sludge amounts with the shorter SRTs (i.e., 5 and 10 days). Null model analysis demonstrated that deterministic homogenous selection could facilitate the colonization and accumulation of biofilms with less coexisting sludge (SRT of 10 days). However, stochastic ecological drift and homogenizing dispersal dominated the colonization and accumulation stages of biofilms with more coexisting sludge (SRT of 20 days), respectively. The ecological networks reflected that positively-related taxa presented taxonomic relatedness, whereas high inconsistency of taxonomic relatedness was observed among aggregate forms or development stages as affected by varied SRTs. The high incidence of intra-taxa co-occurrence patterns suggested that taxa with similar ecological niches could be specifically selected in biofilms when being exposed with less coexisting sludge. This study uncovered ecological mechanisms of biofilm development driven by varying the SRTs of suspended sludge, which would help to propose appropriate strategies for the efficient start-up and optimization of the hybrid sludge-biofilm system.

Kinetic modeling for miniaturize single-chambered microbial fuel cell: effects of biochemical reaction on its performance.

In this study, Nernst growth model equations are used to explain the anodic biofilm (ABF) modeling, linear sweep voltammetry (LSV) at various growth stages of biofilm, and polarization curve modeling for its electron generation behavior in a miniaturized single-chambered microbial fuel cell (SMFC). Kinetic constants of various growth model equations were determined using non-linear regression analysis. Maximum specific growth rate (μmax) at anodic surface is observed 0.016h-1 at a glucose concentration of 12g L-1, whereas retardation in μmax is observed 14g L-1 or more in SMFC. LSV results showed maximum current density of 6720.56mAm-2. Anode performance in SMFC is examined through polarization curve resulting maximum open-circuit voltage (OCV), minimum charge transfer loss, and ohmic loss for NWG (NiWO4 impregnated on rGO), NiWO4, rGO, and plain CC (carbon cloth) anode. These results demonstrate significant enhancement in performance of MFC to lead towards model-based process controlling for significant scale-up in future.

Understanding the role of sorption and biodegradation in the removal of organic micropollutants by membrane aerated biofilm reactor (MABR) with different biofilm thickness

The role of sorption and biodegradation in a membrane aerated biofilm reactor (MABR) were investigated for the removal of 10 organic micropollutants (OMPs) including endocrine disruptors and pharmaceutical active compounds. The influence of the biofilm thickness on the mechanisms of removal was analyzed via kinetic test at three different stages. At all biofilm stages, biodegradation was demonstrated to dominate the removal of selected OMPs. Higher OMPs rates of removal via biodegradation (Kbiol) were achieved when biofilm increased its thickness from (stage T1) 0.26 mm, to (stage T2) 0.58 mm and (stage T3) 1.03 mm. At stage T1 of biofilm, heterotrophs contribute predominantly to OMPs degradation. Hydrophilic compounds removal (i.e., acetaminophen) continue to be driven by heterotrophic bacteria at the next stages of biofilm thickness. However, for medium hydrophobic neutral and charged OMPs, the combined action of heterotrophic and enriched nitrifying activity at stages T2 and T3 enhanced the overall removal. A degradation pathway based on heterotrophic activity for acetaminophen and combined action of nitrifiers-heterotrophs for estrone was proposed based on identified metabolites. Although biodegradation dominated the removal of most OMPs, sorption was also observed to be essential in the removal of biologically recalcitrant and lipophilic compounds like triclosan. Furthermore, sorption capacity of apolar compound was enhanced as the biofilm thickness grew and increased in EPS protein fraction. Microbial analysis confirmed the higher abundance of nitrifying and denitrifying activity at stage T3 of biofilm, which not only facilitated near complete ammonium removal but also enhanced degradation of OMPs.

Using electrochemical impedance spectroscopy to study biofilm growth in a 3D-printed flow cell system

Biofilm contamination is a widespread issue that can occur anywhere when organisms attach to surfaces in the presence of water. In industrial environments, formation of biofilms can lead to component failure, material degradation, and biofouling or spoilage, which collectively come with significant economic costs. Microfabricated electrochemical impedance spectroscopy (EIS) sensors have emerged as a promising tool for monitoring biofilm as EIS sensors capture information about biofilm growth autonomously in real-time; however, sensors suffer from drift, and the technique lacks temporal interpretation of dynamic biofilm processes. In this work, microfabricated sensors featuring gold micro-interdigitated electrodes (μIDEs) were modified with an electrically conductive polymer layer resulting in EIS measurement variability that was significantly reduced compared to unmodified sensors, and enabled highly stable, time-resolved EIS measurements. EIS characterization of Pseudomonas aeruginosa biofilm in parallel with high-resolution confocal laser scanning microscopy (CLSM) was performed using a novel 3D-printed flow cell system, resulting in distinct changes to EIS data corresponding with consistent biofilm growth. We have shown that EIS microsensors can detect four stages of biofilm: (i) initial biofilm attachment to the sensor substrate, (ii) early-stage irreversible biofilm proliferation characterized by sparse biofilm coverage, (iii) mature biofilm detection characterized by uniform biofilm coverage, and (iv) changes due to detachment and regrowth of biofilm.

Antibacterial potential of riboflavin mediated blue diode laser photodynamic inactivation against Enterococcus faecalis: A laboratory investigation.

This study aimed to evaluate the inactivation potency of riboflavin-mediated blue diodelaser photodynamic inactivation (PDI) against Enterococcusfaecalis at planktonic and biofilm stages and also investigated itseffect on the tooth color change. The antibacterial and antibiofilm activities of riboflavin mediated PDI against E.faecalis were investigated. The numbers of colony-forming units (CFUs)/mL were calculated. Teeth discoloration were evaluated using the CIE L*a*b* based color difference (ΔE). Antibacterial analysis indicated that the blue diode laser irradiation at 12, 18, 24, and 30J/cm2 alone and different concentrations of riboflavin solution (6.25 to 100μM) reduced the number of CFU/mL of E.faecalis, but the reduction was not statistically significant (P>0.05). Depending on the riboflavin concentration and the light dose, there can be as much as a 1-log effect on CFU/mL. In addition, E.faecalis biofilmwas more affected with 30J/cm2 irradiation dosage and 100μM riboflavin than other groups. Meanwhile, bacterial suspensions treated with 5.25% sodiumhypochlorite (NaOCl) showed maximum biofilm inhibition and colony number reduction, compared with the control. The teeth exhibited clinically acceptable color change after riboflavin treatment at concentrationranging from 6.25 to 50μM (ΔE < 3.7). The riboflavin mediated PDI process is somewhat less effective than NaOCl but perhaps less toxic to tissues. It might be feasible to repeat the riboflavin+light treatment to further promote efficacy.

An in vitro study on the efficacy of hydrogen peroxide mediated high-power photodynamic therapy affecting Enterococcus faecalis biofilm formation and dispersal.

Biofilms are involved in failure of root canal treatment due to their high resistance to antimicrobial agents, which make their removal as a big challenge. The present study aims at utilizing hydrogen peroxide (HP) plus high frequency laser reinforced antimicrobial photodynamic therapy (a-PDT) as a complementary therapy against Enterococcus faecalis (E. faecalis) at planktonic and biofilm stages. E. faecalis at planktonic and biofilm stages was treated with the photosensitizer HP, followed by no irradiation or irradiation with a power of 2.5W (ʎ=980nm). The cell viability, anti-biofilm, anti-metabolic potential, and temperature changes were evaluated. The combination of HP and 980nm diode laser intensely boosted antibacterial and anti-biofilm efficacy compared with either component alone, affirming HP reinforcement as a bacteriostatic agent. The maximum effect on biofilm occurs in 5.25% sodium hypochlorite (NaOCl) group. During laser irradiations, the mean of temperature changes remains below 5.6°C. It could be concluded that the HP could improve anti-biofilm efficacy as a photosensitizer in a-PDT.

Metabolomic Insights of Biosurfactant Activity from Bacillus niabensis against Planktonic Cells and Biofilm of Pseudomonas stutzeri Involved in Marine Biofouling

In marine environments, biofilm can cause negative impacts, including the biofouling process. In the search for new non-toxic formulations that inhibit biofilm, biosurfactants (BS) produced by the genus Bacillus have demonstrated considerable potential. To elucidate the changes that BS from B. niabensis promote in growth inhibition and biofilm formation, this research performed a nuclear magnetic resonance (NMR) metabolomic profile analysis to compare the metabolic differences between planktonic cells and biofilms of Pseudomonas stutzeri, a pioneer fouling bacteria. The multivariate analysis showed a clear separation between groups with a higher concentration of metabolites in the biofilm than in planktonic cells of P. stutzeri. When planktonic and biofilm stages were treated with BS, some differences were found among them. In planktonic cells, the addition of BS had a minor effect on growth inhibition, but at a metabolic level, NADP+, trehalose, acetone, glucose, and betaine were up-regulated in response to osmotic stress. When the biofilm was treated with the BS, a clear inhibition was observed and metabolites such as glucose, acetic acid, histidine, lactic acid, phenylalanine, uracil, and NADP+ were also up-regulated, while trehalose and histamine were down-regulated in response to the antibacterial effect of the BS.

Priority changes between biofilm exopolysaccharides synthesis and rhamnolipids production are mediated by ac-di-GMP-specific phosphodiesterase NbdA inPseudomonas aeruginosa.

The synthesis of biofilm exopolysaccharides and rhamnolipids (RLs) are two interrelated processes in Pseudomonas aeruginosa, but how bacteria coordinate these two processes remains unclear. We collected a P.aeruginosa KT1115 with rugose small colony variant (RSCV) phenotype from soil, and used it to study the dynamic regulation mechanism of biofilm polysaccharide and RLs synthesis. The results showed that the overproduction of biofilm exopolysaccharides at biofilm stage ultimately contributed the surge of RLs production at RLs stage. This phenomenon was further verified by comparing PAO1 with its engineered RSCV mutant, PAO1ΔwspF. Further genomic, transcriptomic analyses and gene deletion revealed that downregulation of c-di-GMP level was the key to switch biofilm exopolysaccharides accumulation to RLs surge, by transcriptionally upregulating a c-di-GMP phosphodiesterase NbdA. Overall, this study demonstrates the importance of c-di-GMP in coordinating biofilm exopolysaccharides and RLs synthesis, and provides an inspiration for enhancing RLs production through regulating c-di-GMP level.

Biosynthesis of gold nanoparticles and inhibition of various stages of bacterial biofilms formed by drug-resistant Aeromonas hydrophila, Escherichia coli, and Klebsiella pneumoniae.

Biofilm development by bacteria is a serious issue in many different sectors, including the food, pharmaceutical, water, petroleum, and others. Because biofilms are difficult to remove due to their capacity to survive in bad environments, attempts were undertaken throughout this work to biologically manufacture gold nanoparticles with possible antibiofilm capabilities. During this study, 19 morphologically distinct fungal cultures were isolated from soil samples and used to synthesize gold nanoparticles. Among 19 fungi, 7 were producing gold nanoparticles and these particles were subjected to antimicrobial activity. Among 7 gold nanoparticles, nanoparticles synthesized from S2-10 isolate of fungi exhibited high antimicrobial activity toward biofilm-forming drug-resistant bacteria including Klebsiella pneumoniae (14.6 ± 0.57mm), Aeromonas hydrophila (13.3 ± 0.57mm) and Escherichia coli (10.0 ± 1.0mm), and was used for further studies. The chosen nanoparticle also shown strong anti-biofilm action against test microorganisms on a variety of material substrates, including plastic, metal, and glass. These nanoparticles also exhibited inhibition of microcolony/initial biofilm, mature and preformed biofilm formed by Klebsiella pneumoniae (53.42 ± 4.45%, 62.14 ± 7.45%, and 24.12 ± 9.87%, respectively), Aeromonas hydrophila (43.89 ± 0.95%, 59.79 ± 1.19%, and 54.85 ± 6.50%, respectively) and Escherichia coli (45.28 ± 6.88%, 45.17 ± 3.94%, and 32.39 ± 4.16%, respectively). The physicochemical characterization of potential nanoparticles was conducted using UV-visible Spectrophotometer, X-ray diffractionSpectroscopy (XRD), Energy Dispersive X-ray (EDX) spectroscopy, Fourier Transform Infrared Spectroscopy (FTIR), and Scanning Electron Microscopy (SEM). These physicochemical analyses confirmed the formation of gold nanoparticles which were found to be spherical in shape with the size between 30 and 40nm. Furthermore, the potential fungal isolate S2-10 was characterized as Aspergillus niger isolate CJ14 with the help of morphological and molecular techniques and deposited at National Center for Biotechnology Information (NCBI) (Accession number MN519438.1). Hence, it can be concluded that the gold nanoparticles produced using A. niger isolate CJ14 contain significant antibiofilm properties and can be used for developing antibiofilm products for future applications.