Control Biofilm Formation Research Articles

Cadaverine Is a Switch in the Lysine Degradation Pathway in Pseudomonas aeruginosa Biofilm Identified by Untargeted Metabolomics.

There is a critical need to accurately diagnose, prevent, and treat biofilms in humans. The biofilm forming P. aeruginosa bacteria can cause acute and chronic infections, which are difficult to treat due to their ability to evade host defenses along with an inherent antibiotic-tolerance. Using an untargeted NMR-based metabolomics approach, we identified statistically significant differences in 52 metabolites between P. aeruginosa grown in the planktonic and lawn biofilm states. Among them, the metabolites of the cadaverine branch of the lysine degradation pathway were systematically decreased in biofilm. Exogenous supplementation of cadaverine caused significantly increased planktonic growth, decreased biofilm accumulation by 49% and led to altered biofilm morphology, converting to a pellicle biofilm at the air-liquid interface. Our findings show how metabolic pathway differences directly affect the growth mode in P. aeruginosa and could support interventional strategies to control biofilm formation.

Halogenated Dihydropyrrol-2-One Molecules Inhibit Pyocyanin Biosynthesis by Blocking the Pseudomonas Quinolone Signaling System.

Quorum-sensing (QS) systems of Pseudomonas aeruginosa are involved in the control of biofilm formation and virulence factor production. The current study evaluated the ability of halogenated dihydropyrrol-2-ones (DHP) (Br (4a), Cl (4b), and F (4c)) and a non-halogenated version (4d) to inhibit the QS receptor proteins LasR and PqsR. The DHP molecules exhibited concentration-dependent inhibition of LasR and PqsR receptor proteins. For LasR, all compounds showed similar inhibition levels. However, compound 4a (Br) showed the highest decrease (two-fold) for PqsR, even at the lowest concentration (12.5 µg/mL). Inhibition of QS decreased pyocyanin production amongst P. aeruginosa PAO1, MH602, ATCC 25619, and two clinical isolates (DFU-53 and 364707). In the presence of DHP, P. aeruginosa ATCC 25619 showed the highest decrease in pyocyanin production, whereas clinical isolate DFU-53 showed the lowest decrease. All three halogenated DHPs also reduced biofilm formation by between 31 and 34%. The non-halogenated compound 4d exhibited complete inhibition of LasR and had some inhibition of PqsR, pyocyanin, and biofilm formation, but comparatively less than halogenated DHPs.

The Anticancer Agent 3,3'-Diindolylmethane Inhibits Multispecies Biofilm Formation by Acne-Causing Bacteria and Candida albicans.

ABSTRACTThe Gram-positive anaerobic bacterium Cutibacterium acnes is a major inhabitant of human skin and has been implicated in acne vulgaris formation and in the formation of multispecies biofilms with other skin-inhabiting organisms like Staphylococcus aureus and Candida albicans. Indoles are widespread in nature (even in human skin) and function as important signaling molecules in diverse prokaryotes and eukaryotes. In the present study, we investigated the antibacterial and antibiofilm activities of 20 indoles against C. acnes. Of the indoles tested, indole-3-carbinol at 0.1 mM significantly inhibited biofilm formation by C. acnes without affecting planktonic cell growth, and the anticancer drug 3,3′-diindolylmethane (DIM) at 0.1 mM (32 μg/mL) also significantly inhibited planktonic cell growth and biofilm formation by C. acnes, whereas the other indoles and indole itself were less effective. Also, DIM at 0.1 mM successfully inhibited multispecies biofilm formation by C. acnes, S. aureus, and C. albicans. Transcriptional analyses showed that DIM inhibited the expressions of several biofilm-related genes in C. acnes, and at 0.05 mM, DIM inhibited hyphal formation and cell aggregation by C. albicans. These results suggest that DIM and other indoles inhibit biofilm formation by C. acnes and have potential use for treating C. acnes associated diseases.IMPORTANCE Since indoles are widespread in nature (even in human skin), we hypothesized that indole and its derivatives might control biofilm formation of acne-causing bacteria (Cutibacterium acnes and Staphylococcus aureus) and fungal Candida albicans. The present study reports for the first time the antibiofilm and antimicrobial activities of several indoles on C. acnes. Of the indoles tested, two anticancer agents, indole-3-carbinol and 3,3′-diindolylmethane found in cruciferous vegetables, significantly inhibited biofilm formation by C. acnes. Furthermore, the most active 3,3′-diindolylmethane successfully inhibited multispecies biofilm formation by C. acnes, S. aureus, and C. albicans. Transcriptional analyses showed that 3,3′-diindolylmethane inhibited the expressions of several biofilm-related genes including lipase, hyaluronate lyase, and virulence-related genes in C. acnes, and 3,3′-diindolylmethane inhibited hyphal formation and cell aggregation by C. albicans. Our findings show that 3,3′-diindolylmethane offers a potential means of controlling acne vulgaris and multispecies biofilm-associated infections due to its antibiofilm and antibiotic properties.

Development of Antibiofilm Therapeutics Strategies to Overcome Antimicrobial Drug Resistance.

A biofilm is a community of stable microorganisms encapsulated in an extracellular matrix produced by themselves. Many types of microorganisms that are found on living hosts or in the environment can form biofilms. These include pathogenic bacteria that can serve as a reservoir for persistent infections, and are culpable for leading to a broad spectrum of chronic illnesses and emergence of antibiotic resistance making them difficult to be treated. The absence of biofilm-targeting antibiotics in the drug discovery pipeline indicates an unmet opportunity for designing new biofilm inhibitors as antimicrobial agents using various strategies and targeting distinct stages of biofilm formation. The strategies available to control biofilm formation include targeting the enzymes and proteins specific to the microorganism and those involved in the adhesion pathways leading to formation of resistant biofilms. This review primarily focuses on the recent strategies and advances responsible for identifying a myriad of antibiofilm agents and their mechanism of biofilm inhibition, including extracellular polymeric substance synthesis inhibitors, adhesion inhibitors, quorum sensing inhibitors, efflux pump inhibitors, and cyclic diguanylate inhibitors. Furthermore, we present the structure–activity relationships (SAR) of these agents, including recently discovered biofilm inhibitors, nature-derived bioactive scaffolds, synthetic small molecules, antimicrobial peptides, bioactive compounds isolated from fungi, non-proteinogenic amino acids and antibiotics. We hope to fuel interest and focus research efforts on the development of agents targeting the uniquely complex, physical and chemical heterogeneous biofilms through a multipronged approach and combinatorial therapeutics for a more effective control and management of biofilms across diseases.

Role of Efflux Pump-Mediated Antibiotic Resistance in Quorum Sensing-Regulated Biofilm Formation by Salmonella Typhimurium.

This study was designed to assess the influence of efflux pump activity on the biofilm formation in Salmonella Typhimurium. Salmonella enterica subsp. enterica serovar Typhimurium ATCC 19585 (STWT) and clinically isolated S. Typhimurium CCARM 8009 (STCI) were treated with ceftriaxone (CEF), chloramphenicol (CHL), ciprofloxacin (CIP), erythromycin (ERY), norfloxacin (NOR), and tetracycline (TET) in autoinducer-containing media in the absence and presence of phenylalanine-arginine β-naphthylamide (PAβN) to compare efflux pump activity with biofilm-forming ability. The susceptibilities of STWT and STCI were increased in the presence of PAβN. ERY+PAβN showed the highest decrease in the minimum inhibitory concentration (MIC) of ERY from 256 to 2 μg/mL against STWT and STCI. The antimicrobial activity of NOR against planktonic cells was significantly increased in the presence of PAβN, showing the lowest numbers of STWT (3.2 log CFU/cm2), and the TET+PAβN effectively inhibited the growth of STCI (5.2 log CFU/cm2). The lowest biofilm-forming abilities were observed at NOR+PAβN against STWT (biofilm-forming index, BFI < 0.41) and CEF+PAβN against STCI (BFI = 0.32). The bacteria swimming motility and relative fitness varied depending on the antibiotic and PAβN treatments. The motility diameters of STWT were significantly decreased by NOR+PAβN (6 mm) and TET+PAβN (15 mm), while the lowest motility of STCI was observed at CIP+PAβN (8 mm). The significant decrease in the relative fitness levels of STWT and STCI was observed at CIP+PAβN and NOR+PAβN. The PAβN as an efflux pump inhibitor (EPI) can improve the antimicrobial and anti-biofilm efficacy of antibiotics against S. Typhimurium. This study provides useful information for understanding the role of efflux pump activity in quorum sensing-regulated biofilm formation and also emphasizes the necessity of the discovery of novel EPIs for controlling biofilm formation by antibiotic-resistant pathogens.

Biofilm formation potential of Bacillus toyonensis and Pseudomonas aeruginosa on the stainless steel test surfaces in a model dairy batch system

Bacillus toyonensis (a Gram-positive bacterium) and Pseudomonas aeruginosa (a Gram-negative bacterium) isolated from the different surfaces of a dairy plant in our previous study were selected as the test bacteria for the present study. These two test bacteria were investigated in terms of their attachment on the stainless steel test surfaces in a model dairy batch system. After incubation at 5°C and 20°C for 6h, 12h, and 24h, stainless steel plates were examined using cultural counts, profilometer, scanning electron microscopy (SEM), and fluorescent microscopy. Also, the test plates were subjected to a cleaning/disinfection procedure used in the dairy plant. Tests were employed before and after the cleaning/disinfection procedures. Cell wall characteristics and holding temperature were found to be significant for the attachment of the test bacteria to stainless steel test surfaces. In the study, the effect of the holding temperature varied depending on the type and characteristics of the bacteria. The adhesion ability of P. aeruginosa was higher than that of B. toyonensis. Increases in the holding temperature may increase the adhesion ability of the bacteria. Milk growth medium was found to be more successful in preventing the attachment ability of P. aeruginosa compared to B. toyonensis. This indicates that the chemical characteristic of the contact material may affect adhesion. The adhered bacterial cells were entirely removed by means of the cleaning/disinfection treatment. Therefore, the adhesion of bacterial cells could be explained as "initial phase of biofilm formation." It can be concluded that the microorganism cell adhesion on the surface is followed by biofilm formation, and this situation lasts for many years. These results reveal the importance of controlling biofilm formation in dairy plants from the beginning.

Biofilm combating in the food industry: Overview, non‐thermal approaches, and mechanisms

In the food processing industry, hygiene is the key measure to eliminate the pathogens which cause food spoilage. These spoilage microorganisms are persistent sourcing a problem to the processing industry. It stresses the whole environment of food processing. Processing facilities suffer from microbes that suites the microbiota of raw material. These microorganisms grow in the form of biofilm, which is slimy and are hard to eradicate. The conventional methods of eradication of these biofilms can be tiresome and have their limitation. Novel research frontier pertaining to food has shown unrealized potential for eradicating biofilm that can be employed in the food industry. Potential novel technologies that can control biofilm formation include Plasma-activated water, electrolyzed oxidizing water, probiotics, quorum quenching techniques, electrical effects, and cold plasma. The review focus on biofilms in different food industries and the mechanism of various novel approaches which help in controlling biofilm formation. Novelty impact statement Plasma-activated water causes lipid oxidation, membrane disturbance, and oxidative damages breaking the bacterial cell’s defense mechanism. Ultrasonication increases cell permeability due to more effortless penetration of antimicrobial agents into the cells. Electrolyzed oxidizing water could effectively remove biofilm after a short exposure time.

SRNA chaperone Hfq controls bioluminescence and other phenotypes through Qrr1-dependent and -independent mechanisms in Vibrio fischeri.

Colonization of the squid Euprymna scolopes by the bacterium Vibrio fischeri depends on bacterial biofilm formation, motility, and bioluminescence. Previous work has demonstrated an inhibitory role for the small RNA (sRNA) Qrr1 in quorum-induced bioluminescence of V. fischeri, but the contribution of the corresponding sRNA chaperone, Hfq, was not examined. We thus hypothesized that V. fischeri Hfq similarly functions to inhibit bacterial bioluminescence as well as regulate other key steps of symbiosis, including bacterial biofilm formation and motility. Surprisingly, deletion of hfq increased luminescence of V. fischeri beyond what was observed for the loss of qrr1 sRNA. Epistasis experiments revealed that, while Hfq contributes to the Qrr1-dependent regulation of light production, it also functions independently of Qrr1 and its downstream target, LitR. This Hfq-dependent, Qrr1-independent regulation of bioluminescence is also independent of the major repressor of light production in V. fischeri, ArcA. We further determined that Hfq is required for full motility of V. fischeri in a mechanism that partially depends on the Qrr1/LitR regulators. Finally, Hfq also appears to function in the control of biofilm formation: loss of Hfq delayed the timing and diminished the extent of wrinkled colony development, but did not eliminate the production of SYP-polysaccharide-dependent cohesive colonies. Furthermore, loss of Hfq enhanced production of cellulose and resulted in increased Congo red binding. Together, these findings point to Hfq as an important regulator of multiple phenotypes relevant to symbiosis between V. fischeri and its squid host.

Quorum sensing regulates transcription of the pilin gene mshA1 of MSHA pilus in Vibrio parahaemolyticus.

Vibrio parahaemolyticus produces two types of IV pili: mannose-sensitive haemagglutinin type IV pili (MSHA) and chitin-regulated pili (ChiRP). Both of them are required for biofilm formation and the pathogen persistence in hosts. However, there are few reports on the regulation of their expression. In the present study, we showed that the master quorum sensing (QS) regulators AphA and OpaR oppositely regulated the transcription of mshA1 encoding the pilin of MSHA pilus in V. parahaemolyticus. At low cell density (LCD), AphA indirectly repressed mshA1 transcription. In contrast, at high cell density (HCD), OpaR bound to the regulatory DNA region of mshA1 to activate its transcription. Oppositely regulation of mshA1 by AphA and OpaR led to a gradual increase in the expression level of mshA1 from LCD to HCD. Thus, regulation of type IV pili production was one of the mechanisms that V. parahaemolyticus adopted to control biofilm formation.

Antimicrobial Properties and Shear Bond Strength of Composite Used in Orthodontics Following the Addition of Curcumin-Reduced Nanographene Oxide

Background: Composites used in the direct bonding of brackets in fixed orthodontics are considered a suitable environment for the multiplication of cariogenic bacteria and the establishment of microbial plaques by creating a distance between the bracket, tooth surface, and surface properties such as porosity. In the present study, we decided to combine curcumin-reduced nano-graphene oxide (rGO-NCUR) with a composite resin used in orthodontics to achieve a composite with optimal shear bond strength (SBS) and antimicrobial properties against Streptococcus mutans as a main cariogenic bacterium. Methods: In the present study, nanoparticle of graphene oxide (NGO) was synthesized by the modified Hummers’ method using graphite powder and reacted by NCUR to regenerate reduced-NGO (rGO-NCUR). Scanning electron microscopy (SEM) was used to confirm the synthesis of NGO, NCUR, and rGO-NCUR. SBS and antimicrobial activity of Transbond XT composite containing 5% rGO-NCUR were performed against S. mutans biofilms. SPSS software, one-way ANOVA test, and Tukey post hoc test with a significance level equal to or less than 0.05 were used to analyze the data. Results: The synthesis of NGO, NCUR, and rGO-NCUR in nanoscale was confirmed by SEM. The amount of SBS in the composite sample with 5% w/w rGO-NCUR was 12.30 ± 0.65 MPa. The optical density of S. mutans in the biofilm structure formed on composite containing 5% w/w rGO-NCUR was significantly reduced (62.10%) compared with the control group, namely, original composites used in orthodontics (P&lt;0.05). Conclusions: Based on the data of the present study, it can be concluded that composite containing 5% w/w rGO-CUR, without adverse effect on the physical-mechanical properties of composites, can be used as an antimicrobial additive of composites used in orthodontics to control biofilm formation and caries around orthodontic brackets.

Citrus paradisi and Citrus reticulata essential oils interfere with Pseudomonas aeruginosa quorum sensing in vivo on Caenorhabditis elegans

Background: Pseudomonas aeruginosa is a Gram-negative opportunistic pathogen, considered a leading cause of acute and chronic infections in immunocompromised patients. P. aeruginosa uses quorum sensing to control virulence and biofilm formation. To combat this human-resistant pathogen increasing attention has been paid to anti-QS compounds from natural products as potential therapeutic agents. Purpose: To assess the efficacy of C. paradisi (Grapefruit) and C. reticulata (Mandarin) commercial EOs (obtained by cold-pressing and cold-pressing followed by steam distillation, named EOP and EOPD, respectively) and their majority component, limonene, in inhibiting the effect of QS-controlled virulence factors of P. aeruginosa PAO1 and PA14 in a C. elegans infection model. Results: C. paradisi and C. reticulata EOs at 0.125% (v/v) significantly inhibited the in vitro biofilm formation of PAO1 and PA14 strains between 40 and 50%. This EOs concentration, safe for C. elegans according to the survival and brood size assays, rescued the nematodes from P. aeruginosa infections, reducing their death rate by 20–30% in the paralysis assay, and increasing worm lifespan with median survivals of 6 (Grapefruit) and 5 (Mandarin) in the slow killing assay (3 for non-treated worms). Limonene protected nematodes from death in these assays although less effectively than both EOs species. Grapefruit EOs rescued the nematodes in 45–50% from phenazine-death, while mandarin EOs and limonene rescued them in 30%. In the food choice assay, worms preferred the PA14 grown with 0.125% of mandarin and grapefruit EOs or limonene after 3-4 hours (CI=0.11 to 0.18). Conclusion: Our results suggest that C. paradisi and C. reticulata EOPDs, with lower commercial value but similar in vivo effects than EOPs, are sources of anti-QS agents for controlling P. aeruginosa infections, therefore should be considered as potential candidates for the development of novel therapeutics against persistent microorganisms.

Targeting Acyl Homoserine Lactones (AHLs) by the quorum quenching bacterial strains to control biofilm formation in Pseudomonas aeruginosa

Navigating novel biological strategies to mitigate bacterial biofilms have great worth to combat bacterial infections. Bacterial infections caused by the biofilm forming bacteria are 1000 times more resistant to antibiotics than the planktonic bacteria. Among the known bacterial infections, more than 70% involve biofilms which severely complicates treatment options.Biofilm formation is mainly regulated by the Quorum sensing (QS) mechanism. Interference with the QS system by the quorum quenching (QQ) enzyme is a potent strategy to mitigate biofilm. In this study, bacterial strains with QQ activity were identified and their anti-biofilm potential was investigated against the Multidrug Resistant (MDR) Pseudomonas aeruginosa. A Chromobacterium violaceum CV026 and Agrobacterium tumefaciens A136-based bioassays were used to confirm the degradation of different Acyl Homoserine Lactones (AHLs) by QQ isolates. The 16S rRNA gene sequencing of the isolated strains identified them as Bacillus cereus strain QSP03, B. subtilis strain QSP10, Pseudomonas putida strain QQ3 and P. aeruginosa strain QSP01. Biofilm mitigation potential of QQ isolates was tested against MDR P. aeruginosa and the results suggested that 50% biofilm reduction was observed by QQ3 and QSP01 strains, and around 60% reduction by QSP10 and QSP03 bacterial isolates. The presence of AHL degrading enzymes, lactonases and acylases, was confirmed by PCR based screening and sequencing of the already annotated genes aiiA, pvdQ and quiP. Altogether, these results exhibit that QQ bacterial strains or their products could be useful to control biofilm formation in P.aeruginosa.

Gene Sets and Mechanisms of Sulfate-Reducing Bacteria Biofilm Formation and Quorum Sensing With Impact on Corrosion.

Sulfate-reducing bacteria (SRB) have a unique ability to respire under anaerobic conditions using sulfate as a terminal electron acceptor, reducing it to hydrogen sulfide. SRB thrives in many natural environments (freshwater sediments and salty marshes), deep subsurface environments (oil wells and hydrothermal vents), and processing facilities in an industrial setting. Owing to their ability to alter the physicochemical properties of underlying metals, SRB can induce fouling, corrosion, and pipeline clogging challenges. Indigenous SRB causes oil souring and associated product loss and, subsequently, the abandonment of impacted oil wells. The sessile cells in biofilms are 1,000 times more resistant to biocides and induce 100-fold greater corrosion than their planktonic counterparts. To effectively combat the challenges posed by SRB, it is essential to understand their molecular mechanisms of biofilm formation and corrosion. Here, we examine the critical genes involved in biofilm formation and microbiologically influenced corrosion and categorize them into various functional categories. The current effort also discusses chemical and biological methods for controlling the SRB biofilms. Finally, we highlight the importance of surface engineering approaches for controlling biofilm formation on underlying metal surfaces.

Capsicumicine, a New Bioinspired Peptide from Red Peppers Prevents Staphylococcal Biofilm In Vitro and In Vivo via a Matrix Anti-Assembly Mechanism of Action.

ABSTRACTStaphylococci are pathogenic biofilm-forming bacteria and a source of multidrug resistance and/or tolerance causing a broad spectrum of infections. These bacteria are enclosed in a matrix that allows them to colonize medical devices, such as catheters and tissues, and that protects against antibiotics and immune systems. Advances in antibiofilm strategies for targeting this matrix are therefore extremely relevant. Here, we describe the development of the Capsicum pepper bioinspired peptide “capsicumicine.” By using microbiological, microscopic, and nuclear magnetic resonance (NMR) approaches, we demonstrate that capsicumicine strongly prevents methicillin-resistant Staphylococcus epidermidis biofilm via an extracellular “matrix anti-assembly” mechanism of action. The results were confirmed in vivo in a translational preclinical model that mimics medical device-related infection. Since capsicumicine is not cytotoxic, it is a promising candidate for complementary treatment of infectious diseases.IMPORTANCE Pathogenic biofilms are a global health care concern, as they can cause extensive antibiotic resistance, morbidity, mortality, and thereby substantial economic loss. So far, no effective treatments targeting the bacteria in biofilms have been developed. Plants are constantly attacked by a wide range of pathogens and have protective factors, such as peptides, to defend themselves. These peptides are common components in Capsicum baccatum (red pepper). Here, we provide insights into an antibiofilm strategy based on the development of capsicumicine, a natural peptide that strongly controls biofilm formation by Staphylococcus epidermidis, the most prevalent pathogen in device-related infections.

Bacterial cellulose synthesis gene regulates cellular c-di-GMP that control Biofilm formation and mussel larval settlement

Bacterial cellulose is a widely dispersed biofilm matrix component, yet the role of cellulose synthesis-encoding gene on the settlement of macrofouling organisms remains elusive. Here, the functional role of the cellulose synthesis gene bcsQ of the bacterium Pseudoalteromonas marina on settlement of Mytilus coruscus larvae was investigated. It was found that the deficiency of the gene bcsQ was associated with biofilm-settlement inducing activity, the secretion of bacterial α- and β-polysaccharides, and cytosolic c-di-GMP content change. The deletion of the gene bcsQ downregulated c-di-GMP level accompanied by reduced production of polysaccharides including cellulose and colanic acid. Bacterial characteristics in the deficient knockout of bcsQ also changed showing a slow cell growth, reducing biofilm forming capacity and increasing biofilm dispersion. The larval settlement rate of M. coruscus onto the ΔbcsQ biofilm was about 46% lower than that of the wild-type strain in comparison. Therefore, the gene bcsQ manipulates cytosolic c-di-GMP that controls secretion of exopolysaccharides and biofilm formation, and indirectly regulates mussel settlement. The present findings shed new light on interactions among function of bacterial cellulose biosynthesis gene, biofilm formation and the settlement of typical fouling organisms.

Silver Decorated Myconanoparticles Control Growth and Biofilm Formation in Uropathogenic E. coli.

Nanotechnology involves the synthesis of nanoparticles that have been used in the therapeutic application for treating diseases. In this present study, we have adopted the synthesis of myconanoparticles from the extracellular extract of endophytic fungi Penicillium sclerotiorum (PsNps) and validated its antibacterial potential against antibiotic-resistant uropathogenic E. coli and ATCC (25,922) strain of Escherichia coli. Endophytic fungi were isolated from the healthy leaves of Tamarindus indica. The genomic DNA from endophytic fungi was isolated and the ITS region was amplified by polymerase chain reaction (PCR) using universal fungal primers ITS1 and ITS4 and sequenced for the identification of endophytic fungal isolates. Penicillium sclerotiorum extract was used for the synthesis of silver nanoparticles (PsNps) and was characterized by UV-vis spectroscopy, Fourier transform infrared spectroscopy (FTIR), zeta potential, FE-SEM, and Energy dispersive X-ray analysis (EDAX). Antibacterial activity of PsNps was tested against the antibiotic-resistant uropathogenic E. coli and ATCC (25,922) strain of E. coli. Further experiments were carried out to explore the potential of PsNps in regulating the CTX-M-15 gene. The antimicrobial activity showed that the PsNps inhibited growth, biofilm formation in both the strains of E. coli. The expression of the gene encoding CTX-M-15 was downregulated in a resistant strain of uropathogenic E. coli. Our results suggest that the PsNps could be used as an alternative source for antibiotics. Thus, further studies can be conducted to prove the in vivo potential of PsNps and can be formulated for commercialization.

Antibiofilm Effects of Nanoparticles and Visible Light Illumination Against Listeria monocytogenes.

Listeria monocytogenes bacteria pose a particular risk to the food industry as the species is known to form biofilm and to survive in a wide range of challenging environmental conditions. L. monocytogenes can cause listeriosis, a serious food-borne disease, and effective and safe antibiofilm materials and sanitary methods for food processing environments are intensively sought. A variety of nanoparticle materials have been recognized as safe to use in food environments, which allows the application of nanomaterials also for food safety purposes. Nanoparticles together with light illumination generate reactive oxygen species which inactivate bacteria by breaking down cell membranes, proteins, and DNA. The main objective of this study was to evaluate the efficacy of nanomaterials and blue light illumination for L. monocytogenes ATCC 7644 biofilm inactivation. Biofilm was allowed to form for 72 h on nanocoated stainless steel and aluminum plates, after which the plates were illuminated. Non-coated control plates were used to evaluate the antibiofilm efficacy of nanocoating. Plate count method was used to evaluate bacteria counts after illumination. Nanocoating did not affect initial biofilm formation compared to the control plates. Biofilm was significantly (p < 0.05) reduced on stainless steel, aluminum, and TiO2-coated aluminum plates after 72-h illumination by 1.9, 3.2, and 5.9 log, respectively. Nanocoating with visible light illumination could be an effective and safe method for enhancing food safety in food processing facilities to control biofilm formation. Evidence of antibiofilm properties of nanomaterials together with visible light illumination is limited; hence, future studies with variable light intensities and nanomaterials are needed.

Laser surface texturing inhibits Biofilm formation

Biofilm development on surfaces leads to biofouling, which is an undesirable phenomenon in the marine industry. Marine fouling leads to an increase in fuel consumption and corrosion of propeller blades. Most of the antifouling technologies available today are chemical methods that have detrimental effects on the environment. The current work focused on producing non-toxic and effective antifouling systems to control biofilm formation on the titanium alloy surfaces, which has potentially wide range of applications in the marine industry, such as propeller blades, the hull of the ships, heat exchangers, and offshore piping systems. A laser micro-machining was used in this study for the generation of surface texture patterns on the titanium specimens. The contact angle of the untextured surface was 28°, but the wettability of the textured surface increased with a contact angle of 134° for a square pit and 112° for a triangular pit. Surface roughness of the untextured sample showed Ra = 3.811 μm, but post-laser-texturing, the Ra value increased to 10.66 μm and 9.55 μm for the square pit pattern and triangular pit pattern, respectively. Microbes formed biofilm on the untextured specimen but, the square textured surface showed no apparent biofilm formation, while the biofilm was found in triangular textured surface as seen under the fluorescence and scanning electron microscope. The biofilm biomass was significantly reduced in the square textured surface. This work showed that laser textured surfaces might be potentially used to reduce biofilm development on the surfaces. Unlike most methods used for reducing bacterial adhesion, this kind of physical alteration of the surface topography is non-toxic, and it will help to prolong the life of the components used in the marine environment.

LuxQ-LuxU-LuxO pathway regulates biofilm formation by Vibrio parahaemolyticus

Vibrio parahaemolyticus, a common foodborne pathogen, can form biofilms for survival in various environments and for bacterial transmission. Lux systems in Vibrio species are the typical two-component signal transduction systems, which have been demonstrated to contribute to various phenotypes; however, the functions of each homolog of the Lux system in V. parahaemolyticus in the regulation of biofilm formation remain largely unknown. In this study, we first showed that LuxQ, LuxU, and LuxO are essential for controlling biofilm formation by V. parahaemolyticus, through gene knockout studies. We also found that they acted in the same signaling pathway and their deletion mutants exhibited a similar level of biofilm formation. Furthermore, site-directed mutagenesis revealed that the conserved residues for phosphorylation in LuxQ (D784), LuxU (H56) and LuxO (D47) were critical for their regulatory functions on biofilm formation. Phos-tag™ sodium dodecyl sulfate-polyacrylamide gel electrophoresis showed the phosphorylation of LuxU and LuxQ in vivo. Finally, qPCR analysis displayed that the three mutants had a significant decrease in the transcription level of cps loci and cpsQ compared with the wild type strain, which is consistent with the observed phenotype of biofilm formation. Therefore, we propose that LuxQ and its downstream factors LuxU and LuxO function in the same signaling cascade to control biofilm formation by regulating the expression of cpsQ and cps loci. The results of this study provide new data regarding the role of the LuxQ-LuxU-LuxO pathway in biofilm formation by V. parahaemolyticus and help further understand the complex regulatory functions of Lux pathways.

A LysR Family Transcriptional Regulator Modulates Burkholderia cenocepacia Biofilm Formation and Protease Production.

ABSTRACTQuorum-sensing (QS) signals are widely employed by bacteria to regulate biological functions in response to cell densities. Previous studies showed that Burkholderia cenocepacia mostly utilizes two types of QS systems, including the N-acylhomoserine lactone (AHL) and cis-2-dodecenoic acid (BDSF) systems, to regulate biological functions. We demonstrated here that a LysR family transcriptional regulator, Bcal3178, controls the QS-regulated phenotypes, including biofilm formation and protease production, in B. cenocepacia H111. Expression of Bcal3178 at the transcriptional level was obviously downregulated in both the AHL-deficient and BDSF-deficient mutant strains compared to the wild-type H111 strain. It was further identified that Bcal3178 regulated target gene expression by directly binding to the promoter DNA regions. We also revealed that Bcal3178 was directly controlled by the AHL system regulator CepR. These results show that Bcal3178 is a new downstream component of the QS signaling network that modulates a subset of genes and functions coregulated by the AHL and BDSF QS systems in B. cenocepacia.IMPORTANCE Burkholderia cenocepacia is an important opportunistic pathogen in humans that utilizes the BDSF and AHL quorum-sensing (QS) systems to regulate biological functions and virulence. We demonstrated here that a new downstream regulator, Bcal3178 of the QS signaling network, controls biofilm formation and protease production. Bcal3178 is a LysR family transcriptional regulator modulated by both the BDSF and AHL QS systems. Furthermore, Bcal3178 controls many target genes, which are regulated by the QS systems in B. cenocepacia. Collectively, our findings depict a novel molecular mechanism with which QS systems regulate some target gene expression and biological functions by modulating the expression level of a LysR family transcriptional regulator in B. cenocepacia.