Resistance Of Bacterial Biofilms Research Articles

Efficacy of Usnea sp. Extracts in Preventing Biofilm Formation by Bacillus Species Isolated from Soaking Liquor Samples

Bacteria forms biofilm to be resistant to antibacterial agents and other unfavorable environment as compared to planktonic bacterial cells. Due to resistance of bacterial biofilms to commonly used antimicrobial agents and adverse effects of these biofilms in different industries, potential natural compounds which can inhibit bacterial biofilms have attracted more attention in recent years. Lichens are known to have unique secondary metabolites with various biological activities including anti-biofilm properties. Therefore, Bacillus toyonensis, Bacillus mojavensis, Bacillus subtilis, Bacillus amyloliquefaciens, Bacillus velezensis, Bacillus cereus, and Bacillus licheniformis, isolated from soak liquor samples in the previous study, were tested for their ability to form biofilm in this study. Biofilm-forming Bacillus species were detected as B. subtilis, B. amyloliquefaciens, and B. velezensis. The anti-biofilm effect of the acetone extracts of Usnea sp. was evaluated at various concentrations against these biofilm-forming isolates. The anti-biofilm effect of acetone extracts of Usnea sp. against B. subtilis and B. amyloliquefaciens was observed at the concentration of 5 µg/mL by inhibition ratios of 62.75% and 72.72%, respectively. In addition, biofilm formation of B. velezensis was inhibited by the treatment with 1.25 µg/mL extracts at a 62.69% inhibition rate. Biofilm formations of B. amyloliquefaciens and B. velezensis were also suppressed by the extracts at varying percentages of inhibition ranging between 10.11-43.69% and 21.25-46.35%, respectively. This study may provide an alternative approach to overcome the biofilm formation and bacterial resistance to the antibacterial agents in the leather industry.

Біоплівки. Сучасний стан і перспективи антимікробної терапії

Огляд присвячений аналізу сучасних даних щодо механізмів стійкості біоплівок до антимікробних засобів та пошуку нових активних речовин з антибіоплівковою активністю. Проведений аналіз літературних джерел показав, що біоплівка є гетерогенною багатоклітинною структурою, стійкість якої до впливу зовнішніх факторів (у т. ч. антимікробних речовин) визначається будовою позаклітинного матриксу, наявністю сигнальної системи (Quorum sensing), а також здатністю до переходу в метаболічно неактивний стан. Для підвищення ефективності антибіоплівкової терапії як перспективні розглядають речовини, які: 1) руйнують позаклітинний матрикс або підвищують його проникність; 2) впливають на систему Quorum sensing і синтез автоіндукторів; 3) впливають на метаболічно неактивні клітини або викликають їхню реверсію до активного стану. З метою подолання стійкості біоплівок окрім пошуку таргетних молекул також розглядають можливість застосування не антимікробних препаратів, створення молекул, подібних до білків людини та мікроорганізмів, а також застосування вірусів бактерій – бактеріофагів. Цей огляд може бути підґрунтям для дослідників, які прагнуть вивчати антибіоплівкові властивості нових речовин і розробляти потенційні лікарські засоби.

Alginate-caseinate based pH-responsive nano-coacervates to combat resistant bacterial biofilms in oral cavity

Biofilm associated microbial resistance is a major concern in oral health and hygiene. Nano-antimicrobials (NAMs) having natural antibiotic replacers e.g. nisin are the possible solution to treat oral infections. In this study, we have developed pH responsive, mucoadhesive protein-polysaccharide [sodium caseinate (NaC)-sodium alginate (AL)] coacervate based nano-carrier systems (PPC-NCS) to prevent and eradicate oral biofilms associated pathogens e.g. Enterococcus faecium, Staphylococcus epidermidis and Enterococcus faecalis. PPC-NCS were formed at pH 5 with highest encapsulation efficiency of 75.1 ± 1.2%, then at pH 6 (31 ± 1.1%) and 7 (15 ± 2%). SEM of PPC-NCS displayed smooth and round morphology. Particle size of PPC-NCS was observed to be 244 nm by DLS, with negative zeta potential (−47 ± 4.31 mV). In FTIR analysis, merging of OH stretching peak of NaC (2889.53 cm−1) and CH stretching of AL (2920 cm−1) with appearance of a new peak at 2062 cm−1 was observed that confirmed covalent and hydrogen bonding between two compounds. Antimicrobial and antibiofilm assays demonstrated better control of nisin loaded PPC-NCS against selected pathogens, while preventing and eradicating bacterial biofilms successfully without any cytotoxic effect. These results recommend that coacervates based NAMs are suitable carriers for pH-triggered release of antimicrobials in the buccal cavity to control biofilm associated oral infections.

Food Safety Interventions to Control Listeria monocytogenes in the Fresh Apple Packing Industry: A Review.

Listeria monocytogenes, in fresh and ready-to-eat produce such as whole fresh apples, is of concern as there is no "kill step" in their packing process that would eliminate the pathogenic bacteria. Recent listeriosis outbreaks revealed that insufficient cleaning and sanitation practices in fresh apple packing houses may lead to contamination of fruit with L. monocytogenes. This article discusses three fundamental aspects for ensuring microbiological safety of fresh apples: protection of fresh apples from microbial contamination during the packing process, decontamination intervention techniques, and the challenges in removal of L. monocytogenes from fresh apples. Currently used and novel methods of fresh produce decontamination are discussed and evaluated on their usefulness for the apple packing process. Additionally, present regulatory requirements, possible routes of produce contamination, and bacteria attachment and survival mechanisms are described. Optimum methods for microbial decontamination of whole fresh apples are still to be determined. Critical aspects that should be considered in developing the interventions include apple morphology, conditions and scale of the packing process, and influence of the interventions on apple quality. Evaluation of the currently used and emerging decontamination methods indicated that the hurdle technology and rotating use of sanitizers to avoid development of bacterial biofilm resistance may give the best results, although not conclusively.

Sodium Oleate Increases Ampicillin Sensitivity in Methylophilus quaylei Biofilms on Teflon and Polypropylene

Drug combination is a new therapy to improve antibiotic deficiency treatment towards biofilm resistance. This study was conducted to determine the potential effect of sodium oleate to inhibit established biofilms of two strains, methylotrophic bacteria Methylophilus quaylei in combination with ampicillin. Minimum inhibitory concentration (MIC) of ampicillin was determined and added in combination with sodium oleate and examined on planktonic and established biofilms of two strains M. quaylei were characterized by different properties of cell surface hydrophobicity. The effect on biofilms was evaluated by the number of colony forming units (CFUs), crystal violet assay, light and scanning electron microscopy. The study demonstrates that sodium oleate has a promoting activity against planktonic growth of M. quaylei strains and has a slight inhibitory effect on biofilm. Addition of sodium oleate enhances the bactericidal effect of ampicillin against biofilm cells. Combination of ampicillin 0.1 mg/ml (MIC) and sodium oleate 0.03 mg/ml showed a remarkable destruction effect on established biofilms. Combination of ampicillin 0.1 mg/ml (MIC) and sodium oleate 0.03 mg/ml showed a remarkable destruction effect on established biofilms. Overall, results indicated that sodium oleate in combination with ampicillin enhances the inhibition of M. quaylei biofilms and this combination can be utilized for combating bacterial biofilm resistance. Overall, results indicated that sodium oleate in combination with ampicillin enhances the inhibition of M. quaylei biofilms and this combination can be utilized for combating bacterial biofilm resistance.

Mechanism of Antimicrobial Resistance in Bacterial Biofilms

Biyofilm, mikroorganizmaların canlı veya cansız bir
 yüzeye yapışarak kendi ürettikleri ekzopolimerik yapıda jelsi bir tabaka içine
 gömülü halde birbirlerine, bir katı yüzeye ya da bir ara yüzeye tutunarak
 oluşturdukları topluluk olarak tanımlanır. Gıda endüstrisinde biyofilm oluşumu
 gıdalarda bozulmalara, gıda işleme ekipmanlarında hasarlara ve patojen
 kontaminasyonları ile hastalıklara neden olduğu için istenmeyen bir durumdur. Biyofilmler
 antimikrobiyel maddelere karşı planktonik hücrelerden daha dirençlidirler. Biyofilmlerdeki
 bakterilerin antimikrobiyellere karşı gösterdikleri dirençten birçok olası
 mekanizma sorumludur. Biyofilmlerdeki direnç mekanizmasını etkileyen başlıca
 faktörler ekzopolisakkarit matriksi, enzimler, heterojenlik, hücresel
 persistans, metabolik aktivite, genetik adaptasyon, kitle algılama, stres cevapları,
 dış membran yapısı ve sızdırma pompaları’dır. Direnç mekanizmalarının tam
 olarak anlaşılması biyofilm oluşumunun önlenmesi ve kontrol edilmesinde fayda
 sağlayacaktır. Sonuç olarak etkili bir biyofilm kontrolü için araştırmacılar öncelikle
 direnç mekanizmaları üzerinde yoğunlaşmalı ve daha sonra koruyucu önlemlerin
 alınması gerekmektedir. Bu derlemede, gıda endüstrisindeki bakteriyel biyofilmlerin
 antimikrobiyel maddelere karşı gösterdikleri direnç mekanizması üzerinde
 durulmuştur.

A multiplatform approach to investigate the structure and architecture of the biofilms of Pseudomonas aeruginosa and Staphylococcus aureus in response to antimicrobial treatment

Bacterial biofilms are highly complex communities, composed of highly structured extracellular polymers and subpopulations of differentiated cells, such as persisters. These contribute to the resistance of bacterial biofilms to antibiotics, creating a significant issue in the treatment of infections and resulting in elevated levels of mortality and morbidity. Here, we use a microfluidic system coupled with time-lapse microscopy and fluorescent dyes for exopolysaccharide (EPS) and extracellular DNA (eDNA) to investigate how the architecture of the growing biofilm is ordered. We show that in Pseudomonas aeruginosa EPS is deposited first in the initial stages of microcolony development, but that eDNA then acts as a leading edge for further microcolony expansion. We explore how this assembly is perturbed by the introduction of different antimicrobial agents. Further, working with partners in the European Association of National Metrology Institutes (EURAMET) we are developing cross platform methods for the label-free localisation of antimicrobial agents and bacterial components within the biofilm. These platforms include 3D OrbiSIMS (secondary ion mass spectrometry) and Raman spectroscopy. Here we demonstrate the localisation of key biofilm components such as Pseudomonas Quinolone Signal (PQS) molecules in a 3D chemical map of the biofilm.

Synergistic in vitro effects of indocyanine green and ethylenediamine tetraacetate-mediated antimicrobial photodynamic therapy combined with antibiotics for resistant bacterial biofilms in diabetic foot infection.

Antibiotic resistance has emerged as one of the most important determinants in diabetic foot infections outcomes. Antimicrobial Photodynamic therapy(A-PDT) or Photodynamic antimicrobial chemotherapy (PACT) has been proposed as an alternative approach for inactivating bacteria, especially resistant bacterial biofilms. This research investigated the synergistic effects of PACT mediated by the photosensitizer indocyanine green (ICG) and ethylenediamine tetraacetate (EDTA) combined with antibiotics against common pathogens of diabetic foot ulcer infection, including Staphylococcus aureus and Pseudomonas aeruginosa, in vitro. Planktonic bacteria and biofilms of S. aureus and P. aeruginosa were incubated with ICG and EDTA, and then exposed to laser treatment. Quantitative viable counting estimates the phototoxic effects on S. aureus and P. aeruginosa. The susceptibility of methicillin-resistant S. aureus (MRSA) and multidrug-resistant P. aeruginosa (MRPA) to PACT treatment was detected by disk diffusion and micro-broth dilution methods. Confocal microscopy was used to detect the morphology of biofilms treated with PACT and antibiotics. The resazurin assay was used to quantify the metabolic activity of bacteria in biofilms. PACT mediated by ICG and EDTA led to a more pronounced antibacterial effect in S. aureus and P. aeruginosa compared with ICG alone-mediated PACT. P. aeruginosa was more sensitive to ICG and EDTA-mediated PACT than S. aureus. After PACT treatment, the susceptibility of MRSA and MRPA to antibiotics increased. Furthermore, PACT combined with antibiotic treatment significantly contributed to killing bacteria in the biofilm and disrupting biofilm structure. ICG and EDTA-mediated PACT combined with antibiotics synergistically enhanced the effects of sterilization and biofilm destruction.

The food web perspective on aquatic biofilms

AbstractBiofilms, the complex communities of microbiota that live in association with aquatic interfaces, are considered to be hotspots of microbial life in many aquatic ecosystems. Although the importance of attached algae and bacteria is widely recognized, the role of the highly abundant biofilm‐dwelling micrograzers (i.e., heterotrophic protists and small metazoans) is poorly understood. Studies often highlight the resistance of bacterial biofilms to grazing within the microbial food web and therefore argue that the micrograzers have a modulating role (i.e., have effects on biofilm phenotype) rather than a direct trophic role within biofilms. In the present review, we show that this view comes too short, and we establish a conceptual framework of biofilm food webs consisting of three major elements. (1) Energy pathways and subsidization from plankton. As inhabitants of interfaces, biofilm‐dwelling grazers potentially access both planktonic organisms and surface‐associated organisms. They can play an important role in importing planktonic production into the biofilm food web and thus in the coupling of the planktonic and benthic food webs. Nevertheless, specialized grazers are also able to utilize significant amounts of autochthonous biofilm production. (2) Horizontal complexity of the basal food web. While bacteria and algae within biofilms are edible in general, food quality and accessibility of both bacteria and algae can differ considerably between different prey phenotypes occurring during biofilm formation with respect to morphology, chemical defense, and nutrient stoichiometry. Instead of considering bacteria and algae within biofilms to be generally resistant to feeding by micrograzers, we suggest considering a horizontal food‐quality axis to be at the base of biofilm food webs. This food quality gradient is probably associated with increasing costs for the micrograzers. (3) Vertical food web complexity and food chain length. In addition to the consumption of bacteria and algae, many predatory micrograzers exist within biofilm food webs. With the help of video microscopy, we were able to demonstrate the existence of a complex food web with several trophic levels within biofilms. Our conceptual framework should assist in integrating food web concepts and processes into whole‐biofilm budgets and in understanding food‐web‐related interactions within biofilms.

Eradication of Candida Albicans Biofilm By Electrochemical Scaffold Producing Hypochlorous Acid

Biofilms are present in virtually all chronic wounds and contribute to the delay of wound healing. Biofilms are communities of bacterial and/or fungal cells embedded in an extracellular polymeric substance. Conventional treatment methods utilizing topical applications or systemic delivery of antibiotics are often inadequate because biofilms are inherently resistant to most of these approaches since the majority of antibiotics are active against metabolically-active, but not dormant, cells. The present study examines a new technology that can simultaneously eradicate biofilms on the wound surface and promote wound healing. The electrochemical scaffold (e-scaffold) is a platform technology that allows controlled, continuous electrochemical production of low concentrations of biocides. The proposed novel e-scaffold consists of a three-electrode system, the working and counter electrodes are made of flexible carbon fabric, are accompanied by a Ag/AgCl reference electrode, controlled using a custom potentiostat. The e-scaffold works by electrochemically oxidizing chloride ions on the working electrode to chlorine gas when a constant potential of 1.5 VAg/AgCl is applied, and produces hypochlorous acid via chlorine dissolution. A phosphate buffer saline solution is added to the e-scaffold to provide chloride ions. While e-scaffold effectiveness in eradicating bacterial and antibiotic resistant bacterial biofilms was demonstrated in previous studies, its effectiveness against fungal biofilms has not been demonstrated. The goal of this work is to demonstrate the efficacy of the e-scaffold against in vitro Candida albicans biofilm. Fungal biofilms, such as C. albicans, differ from bacterial biofilms because fungal cells are eukaryotic cells and polymorphic, making fungal biofilms difficult to treat. After treating the C. albicans biofilm in an in vitro model with electrochemically produced hypochlorous acid using e-scaffold for 24 hours, the biofilm was completely eradicated. The results suggest that the electrochemical scaffold is a viable, new technology that can simultaneously eradicate antibiotic resistant fungal biofilms from the surfaces.

Efficacy of Dilute Povidone-Iodine against Multi- Drug Resistant Bacterial Biofilms, Fungal Biofilms and Fungal Spores

To study the in vitro effect of novel low-dose povidoneiodine formulations against established biofilms of multi-drug resistant Staphylococcus aureus, Klebsiella pneumoniae, Pseudomonas aeruginosa and Candida albicans and to test the Minimal Inhibitory Concentration of the same formulations against Candida auris, Trichophton mentagrophytes, Microsporum canis, Candida albicans and Aspergillus fumigatus.

Grazing resistance of bacterial biofilms: a matter of predators' feeding trait.

Biofilm formation in bacteria is considered to be one strategy to avoid protozoan grazing. However, this assumption is largely based on experiments with suspension-feeding protozoans. Here we test the hypothesis that grazing resistance depends on both the grazers' feeding trait and the bacterial phenotype, rather than being a general characteristic of bacterial biofilms. We combined batch experiments with mathematical modelling, considering the bacterium Pseudomonas putida and either a suspension-feeding (i.e. the ciliate Paramecium tetraurelia) or a surface-feeding grazer (i.e. the amoeba Acanthamoeba castellanii). We find that both plankton and biofilm phenotypes were consumed, when exposed to their specialised grazer, whereas the other phenotype remained grazing-resistant. This was consistently shown in two experiments (starting with either only planktonic bacteria or with additional pre-grown biofilms) and matches model predictions. In the experiments, the plankton feeder strongly stimulated the biofilm biomass. This stimulation of the resistant prey phenotype was not predicted by the model and it was not observed for the biofilm feeders, suggesting the existence of additional mechanisms that stimulate biofilm formation besides selective feeding. Overall, our results confirm our hypothesis that grazing resistance is a matter of the grazers' trait (i.e. feeding type) rather than a biofilm-specific property.

Inactivation of bacterial biofilms using visible-light-activated unmodified ZnO nanorods

Various zinc oxide (ZnO) nanostructures are widely used for photocatalytic antibacterial applications. Since ZnO possesses a wide bandgap, it is believed that only UV light may efficiently assist bacterial inactivation, and diverse crystal lattice modifications should be applied in order to narrow the bandgap for efficient visible-light absorption. In this work we show that even unmodified ZnO nanorods grown by an aqueous chemical growth technique are found to possess intrinsic defects that can be activated by visible light (λ = 405 nm) and successfully applied for total inactivation of various highly resistant bacterial biofilms rather than more sensitive planktonic bacteria. Time-resolved fluorescence analysis has revealed that visible-light excitation creates long-lived charge carriers (τ > 1 μs), which might be crucial for destructive biochemical reactions achieving significant bacterial biofilm inactivation. ZnO nanorods covered with bacterial biofilms of Enterococcus faecalis MSCL 302 after illumination by visible light (λ = 405 nm) were inactivated by 2 log, and Listeria monocytogenes ATCL3C 7644 and Escherichia coli O157:H7 biofilms by 4 log. Heterogenic waste-water microbial biofilms, consisting of a mixed population of mesophilic bacteria after illumination with visible light were also completely destroyed.

Understanding the Mechanism of Bacterial Biofilms Resistance to Antimicrobial Agents.

A biofilm is a group of microorganisms, that causes health problems for the patients with indwelling medical devices via attachment of cells to the surface matrix. It increases the resistance of a microorganism for antimicrobial agents and developed the human infection. Current strategies are removed or prevent the microbial colonies from the medical devices, which are attached to the surfaces. This will improve the clinical outcomes in favor of the patients suffering from serious infectious diseases. Moreover, the identification and inhibition of genes, which have the major role in biofilm formation, could be the effective approach for health care systems. In a current review article, we are highlighting the biofilm matrix and molecular mechanism of antimicrobial resistance in bacterial biofilms.

Influence of Nutrient Availability and Quorum Sensing on the Formation of Metabolically Inactive Microcolonies Within Structurally Heterogeneous Bacterial Biofilms: An Individual-Based 3D Cellular Automata Model.

The resistance of bacterial biofilms to antibiotic treatment has been attributed to the emergence of structurally heterogeneous microenvironments containing metabolically inactive cell populations. In this study, we use a three-dimensional individual-based cellular automata model to investigate the influence of nutrient availability and quorum sensing on microbial heterogeneity in growing biofilms. Mature biofilms exhibited at least three structurally distinct strata: a high-volume, homogeneous region sandwiched between two compact sections of high heterogeneity. Cell death occurred preferentially in layers in close proximity to the substratum, resulting in increased heterogeneity in this section of the biofilm; the thickness and heterogeneity of this lowermost layer increased with time, ultimately leading to sloughing. The model predicted the formation of metabolically dormant cellular microniches embedded within faster-growing cell clusters. Biofilms utilizing quorum sensing were more heterogeneous compared to their non-quorum sensing counterparts, and resisted sloughing, featuring a cell-devoid layer of EPS atop the substratum upon which the remainder of the biofilm developed. Overall, our study provides a computational framework to analyze metabolic diversity and heterogeneity of biofilm-associated microorganisms and may pave the way toward gaining further insights into the biophysical mechanisms of antibiotic resistance.

Evaluation of the antibacterial activity of a conventional orthodontic composite containing silver/hydroxyapatite nanoparticles.

BackgroundOne of the most important complications of fixed orthodontic treatment is the formation of white spots which are initial carious lesions. Addition of antimicrobial agents into orthodontic adhesives might be a wise solution for prevention of white spot formation. The aim of this study was to evaluate the antibacterial properties of a conventional orthodontic adhesive containing three different concentrations of silver/hydroxyapatite nanoparticles.MethodsOne hundred and sixty-two Transbond XT composite discs containing 0, 1, 5, and 10 % silver/hydroxyapatite nanoparticles were prepared and sterilized. Antibacterial properties of these composite groups against Streptococcus mutans, Lactobacillus acidophilus, and Streptococcus sanguinis were investigated using three different antimicrobial tests. Disk agar diffusion test was performed to assess the diffusion of antibacterial agent on brain heart infusion agar plate by measuring bacterial growth inhibition zones. Biofilm inhibition test showed the antibacterial capacity of composite discs against resistant bacterial biofilms. Antimicrobial activity of eluted components from composite discs was investigated by comparing the viable counts of bacteria after 3, 15, and 30 days.ResultsComposite discs containing 5 and 10 % silver/hydroxyapatite nanoparticles were capable of producing growth inhibition zones for all bacterial types. Results of biofilm inhibition test showed that all of the study groups reduced viable bacterial count in comparison to the control group. Antimicrobial activity of eluted components from composite discs was immensely diverse based on the bacterial type and the concentration of nanoparticles.ConclusionsTransbond XT composite discs containing 5 and 10 % silver/hydroxyapatite nanoparticles produce bacterial growth inhibition zones and show antibacterial properties against biofilms.

Intranasal delivery of ciprofloxacin to rats: A topical approach using a thermoreversible in situ gel

Intranasal administration of antibiotics is an alternative and attractive delivery approach in the treatment of local infections such as chronic rhinosinusitis. This topical route has the advantage of delivering high drug concentrations directly to the site of infection when trying to eradicate the highly resistant bacterial biofilms. The purpose of this study was to assess and compare the pharmacokinetic parameters of ciprofloxacin following intranasal and intravenous administrations to rats in plasma, olfactory bulb and nasal mucosa of two different nasal regions. For intranasal administration a thermoreversible in situ gel was used to increase drug residence time in nasal cavity. Ciprofloxacin concentration time-profile in nasal mucosa of the studied anterior region (at naso- and maxilloturbinates level) was markedly higher after intranasal administration (0.24mg/kg) than that following intravenous administration (10mg/kg), while in nasal mucosa of the more posterior region (at ethmoidal turbinates level) ciprofloxacin concentrations were found to be higher after intranasal administration when the different dose administered by both routes is taken into account. A plateau in ciprofloxacin concentration was observed in nasal mucosa of both studied regions after intranasal administration, suggesting a slow delivery of the drug over a period of time using the nasal gel formulation. In plasma and olfactory bulb, concentration of ciprofloxacin was residual after intranasal administration, which demonstrates this is a safe administration route by preventing systemic and particularly central nervous system adverse effects. Dose-normalized pharmacokinetic parameters of ciprofloxacin exposure to nasal mucosa revealed higher values after intranasal delivery not only in the anterior region but also in the posterior nasal region. In conclusion, topical intranasal administration appears to be advantageous for delivering ciprofloxacin to the biophase, with negligible systemic and brain exposure using a 41.7-fold lower dose than intravenous administration. Therefore, it may represent a promising approach in the drug management of chronic rhinosinusitis.

Antibiotics Resistance of Bacterial Biofilms

Antibiotics Resistance of Bacterial Biofilms

Efflux as a Glutaraldehyde Resistance Mechanism in Pseudomonas fluorescens and Pseudomonas aeruginosa Biofilms

A major challenge in microbial biofilm control is biocide resistance. Phenotypic adaptations and physical protective effects have been historically thought to be the primary mechanisms for glutaraldehyde resistance in bacterial biofilms. Recent studies indicate the presence of genetic mechanisms for glutaraldehyde resistance, but very little is known about the contributory genetic factors. Here, we demonstrate that efflux pumps contribute to glutaraldehyde resistance in Pseudomonas fluorescens and Pseudomonas aeruginosa biofilms. The RNA-seq data show that efflux pumps and phosphonate degradation, lipid biosynthesis, and polyamine biosynthesis metabolic pathways were induced upon glutaraldehyde exposure. Furthermore, chemical inhibition of efflux pumps potentiates glutaraldehyde activity, suggesting that efflux activity contributes to glutaraldehyde resistance. Additionally, induction of known modulators of biofilm formation, including phosphonate degradation, lipid biosynthesis, and polyamine biosynthesis, may contribute to biofilm resistance and resilience. Fundamental understanding of the genetic mechanism of biocide resistance is critical for the optimization of biocide use and development of novel disinfection strategies. Our results reveal genetic components involved in glutaraldehyde resistance and a potential strategy for improved control of biofilms.

Extracellular matrix structure governs invasion resistance in bacterial biofilms.

Many bacteria are highly adapted for life in communities, or biofilms. A defining feature of biofilms is the production of extracellular matrix that binds cells together. The biofilm matrix provides numerous fitness benefits, including protection from environmental stresses and enhanced nutrient availability. Here we investigate defense against biofilm invasion using the model bacterium Vibrio cholerae. We demonstrate that immotile cells, including those identical to the biofilm resident strain, are completely excluded from entry into resident biofilms. Motile cells can colonize and grow on the biofilm exterior, but are readily removed by shear forces. Protection from invasion into the biofilm interior is mediated by the secreted protein RbmA, which binds mother-daughter cell pairs to each other and to polysaccharide components of the matrix. RbmA, and the invasion protection it confers, strongly localize to the cell lineages that produce it.