Dense Biofilm Research Articles

An ex vivo study of infections of vascular grafts and endografts with scanning electron microscopy

ObjectiveVascular graft and endograft infections (VGEIs) are complicated by high morbidity, mortality, and recurrence rates, notably due to biofilm formation on the graft surface, hardly dislodgeable by the sole anti-infectious treatment. The characteristics of this biofilm are still poorly documented. The aim of this study was to evaluate ex vivo biofilm on removed infected vascular grafts and endografts (VGEs). MethodsExplanted VGEs were prospectively collected from 2019 to 2022 at Bordeaux University Hospital, France. Two samples per graft were used for scanning electron microscopy imaging; one was sonicated, and both grafts’ sides were imaged. ResultsA total of 26 patients were included, 18 with VGEI, eight without any infection (endoleak and/or thrombosis), and 29 VGEs were collected. Microbial documentation was obtained in 83% of VGEIs. A thick layer of fibrin was visible on almost all grafts, mixed with a dense biofilm matrix on infected grafts visible as early as 1 month after the onset of infection. Bacteria were not always visualized on infected grafts’ surface (80% on outer side and 85% on luminal side) but were surprisingly present on one-third of non-infected grafts. There was no significant difference between biofilm, fibrin, and microorganisms’ distribution between the two grafts’ sides. However, there were clear differences between infected and non-infected grafts, since immune cells, bacteria and biofilm were more frequently visualized on both sides of infected grafts (P < .05). Bacteria and immune cells although still visible, were significantly less present after sonication; the number of other elements including biofilm was not significantly different. ConclusionsThe persistence of a thick layer of fibrin and biofilm embedding microorganisms on both sides of infected VGE even after 1 month of infection could be the explanation for the low success rates of conservative management and the usual need for graft removal to treat VGEIs.

An enteric ultrastructural surface atlas of the model insect Manduca sexta

The tobacco hornworm is a laboratory model that is particularly suitable for analyzing gut inflammation, but a physiological reference standard is currently unavailable. Here, we present a surface atlas of the healthy hornworm gut generated by scanning electron microscopy and nano-computed tomography. This comprehensive overview of the gut surface reveals morphological differences between the anterior, middle, and posterior midgut, allowing the screening of aberrant gut phenotypes while accommodating normal physiological variations. We estimated a total resorptive midgut surface of 0.42 m2 for L5d6 larvae, revealing its remarkable size. Our data will support allometric scaling and dose conversion from Manduca sexta to mammals in preclinical research, embracing the 3R principles. We also observed non-uniform gut colonization by enterococci, characterized by dense biofilms in the pyloric cone and downstream of the pylorus associated with pore and spine structures in the hindgut intima, indicating a putative immunosurveillance function in the lepidopteran hindgut.

Effects of cations on biofilms in gravity-driven membrane system: Filtration performance and mechanism investigation

The gravity-driven membrane (GDM) system is desirable for energy-efficient water treatment. However, little is known about the influence of cations on biofilm properties and GDM performance. In this study, typical cations (Ca2+ and Na+) were used to reveal the combined fouling behavior and mechanisms. Results showed that Ca2+ improved the stable flux and pollutant removal efficiency, while Na+ adversely affected the flux. Compared with GDM control, the concentration of pollutants was lower in Ca-GDM, as indicated by the low biomass, proteins, and polysaccharides. A heterogeneous and loose biofilm was observed in the Ca-GDM system, with roughness and porosity increasing by 43.06 % and 32.60 %, respectively. However, Na+ induced a homogeneous and dense biofilm, with porosity and roughness respectively reduced by 17.48 % and 22.04 %. The richness of bacterial communities increased in Ca-GDM systems, while it decreased in Na-GDM systems. High adenosine triphosphate (ATP) concentration in Ca-GDM system was consistent with the abundant bacteria and their high biological activity, which was helpful for the efficient removal of pollutants. The abundance of Apicomplexa, Platyhelminthes, Annelida and Nematoda increased after adding Ca2+, which was related to the formation of loose biofilms. Computational simulations indicated that the free volumes of the biofilms in Ca-GDM and Na-GDM were 13.7 and 13.2 nm3, respectively. The addition of cations changed intermolecular forces, Ca2+ induced bridging effects led to large and loose floc particles, while the significant dehydration of hydrated molecules in the Na-GDM caused obvious aggregation. Overall, microbiological characteristics and contaminant molecular interactions were the main reasons for differences in GDM systems.

Insights into the response of nitrogen metabolism to sulfamethoxazole contamination in constructed wetlands with varied substrates

This study conducted an analysis of the variations in nitrogen metabolism pathways within constructed wetlands (CWs) using zeolite (CW-Z), ceramsite (CW-C), and lava (CW-L) under high concentration sulfamethoxazole (SMX) stress. The introduction of SMX hindered the formation of hydrogen bonds on the substrate surfaces; however, these surfaces still maintained a dense and thick biofilm. CW-Z exhibited superior removal efficiencies for ammonium nitrogen (NH4+-N) and nitrate nitrogen (NO3–-N) compared to CW-C and CW-L, with removal rates of 92.54 ± 2.88 % and 89.39 ± 6.74 %, respectively. Interestingly, the proportion of genes involved in nitrification, denitrification and nitrate reduction genes in CW-C (36.05 %) were higher than that in CW-C (29.81 %) and CW-L (29.70 %) but the interactions among nitrogen functional bacteria in CW-Z were much more complex. Further analysis of the nitrogen metabolism pathway indicated that under CW-Z enhanced dissimilatory nitrate reduction SMX stress, while CW-L enhanced assimilatory nitrate reduction process compared to CW-C.

Enterococcus mundtii A2 biofilm and its anti-adherence potential against pathogenic microorganisms on stainless steel 316L.

Pathogenic bacterial biofilms present significant challenges, particularly in food safety and material deterioration. Therefore, using Enterococcus mundtii A2, known for its antagonistic activity against pathogen adhesion, could serve as a novel strategy to reduce pathogenic colonization within the food sector. This study aimed to investigate the biofilm-forming ability of E. mundtii A2, isolated from camel milk, on two widely used stainless steels within the agri-food domain and to assess its anti-adhesive properties against various pathogens, especially on stainless steel 316L. Additionally, investigations into auto-aggregation and co-aggregation were also conducted. Plate count methodologies revealed increased biofilm formation by E. mundtii A2 on 316L, followed by 304L. Scanning electron microscopy (SEM) analysis revealed a dense yet thin biofilm layer, playing a critical role in reducing the adhesion of L. monocytogenes CECT 4032 and Staphylococcus aureus CECT 976, with a significant reduction of ≈ 2 Log CFU/cm2. However, Gram-negative strains, P. aeruginosa ATCC 27853 and E. coli ATCC 25922, exhibit modest adhesion reduction (~ 0.7 Log CFU/cm2). The findings demonstrate the potential of applying E. mundtii A2 biofilms as an effective strategy to reduce the adhesion and propagation of potentially pathogenic bacterial species on stainless steel 316L.

Characteristics of fungi formation in urban sewer at different flow conditions: Distribution, metabolism, and pathogenicity

Fungi are the significant components of the sewer ecology system which can consume substances and exhibit pathogenicity. However, the characteristics of fungi formation and metabolism in the complex sewer environment have not been revealed in depth. In this study, gradient flow conditions were conducted in a pilot sewer and the formation characteristics of fungi were synthetically investigated. The results showed that the low flow rate at 0.1–0.4 m/s led to the loose morphology of biofilms, while the overly loose environment did not allow fungi communities to thrive in sewer. The dense biofilms were found at the middle flow condition (0.4–0.6 m/s), and the fungal communities with degradation functions were exuberant at this condition (such as Tremellales with relative abundance of 6.18% and Talaromyces with relative abundance of 6.51%). In particular, eleven kinds of fungi with known pathogenicity of the sewer biofilm were found in this study, and it is worth noting that the abundance of pathogenic fungi at medium flow rates is significantly higher than that at other flow conditions (higher than 10 %). While, excessive flow shear force (0.8–1.2 m/s) led to biofilm shedding which caused hindering the proper generation of fungi. In summary, the pollutant transformation and pathogenic exposure conducted by fungi communities could affect the sewer management process significantly, and this study could provide research foundation for wastewater quality prediction and management of pathogenic risk in sewer systems.

Biodegradation of atrazine with biochar-mediated functional bacterial biofilm: Construction, characterization and mechanisms

The abuse and residue of herbicides in the black soil area had seriously affected the soil structure, function and crop growth, posing severe threats to agricultural soil environment and public health. Given the limitation of routine microbial remediation, innovative and eco-friendly functional bacterial biofilm which could adapt under adverse conditions was developed on the biochar to investigate its enhanced bioremediation and metabolic characteristics of typical herbicide atrazine. Results revealed that the atrazine degrading strain Acinetobacter lwoffii had competitive advantage in soil indigenous microorganisms and formed dense biofilms on the biochar which was beneficial to cell viability maintenance and aggregations. Metatranscriptomics and RT-qPCR analysis demonstrated that the biochar-mediated biofilm improved the frequency of intercellular communications through quorum sensing and two-component signal regulation systems, and enhanced the atrazine biodegradation efficiency through horizontal gene transfer in co-metabolism mode, providing important scientific basis for the biological remediation of farmland soil non-point source pollution.

2D covalent organic framework/reduced graphene oxide hybrid hydrogel for simultaneous power generation and organic contamination degradation in microbial fuel cell

Microbial fuel cells (MFCs) are promising devices that convert chemical energy into electrical energy via extracellular electron transfer between bacteria and electrodes. However, inefficient extracellular electron transfers and bacteria/electrode interactions often limit their performance. Herein, we demonstrate a hybrid hydrogel assembled by a 2D covalent organic framework (COF) and reduced graphene oxide (rGO) nanosheets as anode materials for MFCs. The 2D COF nanosheets effectively prevent the stacking of rGO nanosheets, resulting in the hybrid hydrogel with a high specific surface area (313.2 m2/g), abundant mesopores and macropores, and high electrical conductivity (492 S/m). The COF material also brings many quinone groups as redox mediators to facilitate electron transfer between bacteria and electrodes. The COF/rGO hybrid hydrogel enhances the growth of Shewanella decolorationis S12 cells, even under the influence of different organic dyes. Dense biofilms are formed on the hybrid hydrogel's surface and inside its macropores, further improving extracellular electron transfers. Assembled MFCs using COF/rGO anodes deliver a maximum power density of 905.1 mW/m2 and can operate for 600 h. They also enable simultaneous power generation and fast organic contamination degradation in electrolytes. This work shows the potential of using 2D nanoarchitectures to promote high-performance MFCs.

Biofilm forming ability of coagulase-positive staphylococci isolated from animals in Ukraine

Staphylococcal biofilms are an important virulence factor that allows for effective infectious effects and colonization of the animal body. This study was devoted to the evaluation of the biofilm forming ability of different strains of Staphylococcus pseudintermedius and Staphylococcus aureus isolated from animals in Ukraine. In addition, the presence of extracellular adhesin genes icaA and icaD in S. pseudintermedius strains was determined. The density of the biofilms was determined by culturing microorganisms in 96-well plates and staining the resulting structures with crystal violet. The genes responsible for biofilm formation were identified by classical polymerase chain reaction (PCR) using primers selected from the literature. The data obtained from this study showed a direct correlation between the density of the biofilm and the number of viable cells involved in its formation. Thus, 23.1% of S. pseudintermedius strains and 25% of S. aureus strains isolated from dogs demonstrated the ability to form a dense biofilm, while 46.2% of S. pseudintermedius strains and 50% of S. aureus strains formed a weak biofilm. The origin of the isolates had no significant effect on the biofilm characteristics. Coagulase-positive staphylococci obtained from cats did not form dense biofilms. 42.9% of methicillin-resistant Staphylococcus aureus (MRSA) isolates from cows with mastitis had moderate to strong biofilm forming properties. Isolates that showed resistance to three or more antibiotic groups tended to form denser biofilms. In addition, 73.3% of the studied S. pseudintermedius strains were found to contain the icaA gene, and 90% – the icaD gene. A genotypic profile combining both icaA and icaD genes was present in 66.7% of the bacteria, while one strain lacked both genes. Understanding the biofilm forming properties of staphylococcal isolates is important in the context of developing optimal treatment strategies and effective antibiotic use, which will contribute to better control of infections caused by these microorganisms.

Blueberry extract inhibits quorum-sensing regulators and controls Vibrio parahaemolyticus biofilms and virulence

Anti-biofilm and anti-virulence effects of blueberry extract (BE) at sub-inhibitory of concentrations (SICs) against Vibrio parahaemolyticus were investigated in this study. When three V. parahaemolyticus strains were exposed to BE at 6.25 mg/mL and 9.38 mg/mL, swarming and twitching motility were suppressed by 40.4%–57.1% and 20.8%–39.1%, respectively. Hydrophobicity and auto-aggregation were decreased by 63.2%–94.5% and 43.8%–88.4%, respectively. The decrease in AI-2 production ranged from 24.8% to 76.8%. The biofilm formation index remained less than 0.64 after 24−72h incubation. Microscopic analyses further evidenced that the dense and thick biofilm architecture were significantly weaken. Meanwhile, BE differentially repressed the expression of the critical genes involved in flagellum (mshA), pili (oxyR), QS (aphA, opaR, and luxS), EPS (cpsA and cpsF), and T3SS (vcrD1, vopD1, and vopS). These results suggested that BE could suppress the motility and cellular surface properties by controlling QS regulators, which further inhibited biofilm formation of V. parahaemolyticus. Thus, BE may be a new alternative natural antibiofilm agent in the future.

Competitive interactions facilitate resistance development against antimicrobials.

While the evolution of antimicrobial resistance is well studied in free-living bacteria, information on resistance development in dense and diverse biofilm communities is largely lacking. Therefore, we explored how the social interactions in a duo-species biofilm composed of the brewery isolates Pseudomonas rhodesiae and Raoultella terrigena influence the adaptation to the broad-spectrum antimicrobial sulfathiazole. Previously, we showed that the competition between these brewery isolates enhances the antimicrobial tolerance of P. rhodesiae. Here, we found that this enhanced tolerance in duo-species biofilms is associated with a strongly increased antimicrobial resistance development in P. rhodesiae. Whereas P. rhodesiae was not able to evolve resistance against sulfathiazole in monospecies conditions, it rapidly evolved resistance in the majority of the duo-species communities. Although the initial presence of R. terrigena was thus required for P. rhodesiae to acquire resistance, the resistance mechanisms did not depend on the presence of R. terrigena. Whole genome sequencing of resistant P. rhodesiae clones showed no clear mutational hot spots. This indicates that the acquired resistance phenotype depends on complex interactions between low-frequency mutations in the genetic background of the strains. We hypothesize that the increased tolerance in duo-species conditions promotes resistance by enhancing the selection of partially resistant mutants and opening up novel evolutionary trajectories that enable such genetic interactions. This hypothesis is reinforced by experimentally excluding potential effects of increased initial population size, enhanced mutation rate, and horizontal gene transfer. Altogether, our observations suggest that the community mode of life and the social interactions therein strongly affect the accessible evolutionary pathways toward antimicrobial resistance.IMPORTANCEAntimicrobial resistance is one of the most studied bacterial properties due to its enormous clinical and industrial relevance; however, most research focuses on resistance development of a single species in isolation. In the present study, we showed that resistance evolution of brewery isolates can differ greatly between single- and mixed-species conditions. Specifically, we observed that the development of antimicrobial resistance in certain species can be significantly enhanced in co-culture as compared to the single-species conditions. Overall, the current study emphasizes the need of considering the within bacterial interactions in microbial communities when evaluating antimicrobial treatments and resistance evolution.

Fabric Fiber as a Biofilm Carrier for Halomonas sp. H09 Mixed with Lactobacillus rhamnosus GG.

Biofilm bacteria have stronger resistance to the adverse external environment compared to planktonic bacteria, and biofilms of non-pathogenic bacteria have strong potential for applications in food. In this experiment, Halomonas sp. H09 and Lactobacillus rhamnosus GG, which have film-forming ability in monoculture and better film-forming ability in mixed culture than the two strains alone, were selected as the target strains for mixed culture. According to SEM observation and bacterial dry weight measurement, the target strain formed a dense biofilm on a 0.1g/L chitosan-modified cellulose III carrier. Furthermore, the presence of extracellular polymeric substances in biofilms was verified by EDS and FTIR. The results showed that 0.1g/L chitosan-modified cellulose III was an ideal carrier material for immobilization of Halomonas sp. H09 with Lactobacillus rhamnosus GG biofilm. This research provided a basis for the selection of non-pathogenic mixed-bacteria biofilm carriers.

A CuS@g-C3N4 heterojunction endows scaffold with synergetic antibacterial effect.

Graphitic carbon nitride (g-C3N4) had aroused tremendous attention in photodynamic antibacterial therapy due to its excellent energy band structure and appealing optical performance. Nevertheless, the superfast electron-hole recombination and dense biofilm formation abated its photodynamic antibacterial effect. To this end, a nanoheterojunction was synthesized via in-situ growing copper sulfide (CuS) on g-C3N4 (CuS@g-C3N4). On the one hand, CuS could form Fermi level difference with g-C3N4 to accelerate carrier transfer and thus facilitate electron-hole separation. On the other hand, CuS could respond near-infrared light to generate localized thermal to disrupt biofilm. Then the CuS@g-C3N4 nanoparticle was introduced into the poly-l-lactide (PLLA) scaffold. The photoelectrochemistry results demonstrated that the electron-hole separation efficiency was apparently enhanced and thereby brought an approximate sevenfold increase in reactive oxygen species (ROS) production. The thermal imaging indicated that the scaffold possesses a superior photothermal effect, which effectively eradicated the biofilm by disrupting its extracellular DNA and thereby facilitated to the entry of ROS. The entered ROS could effectively kill the bacteria by causing protein, K+, and nucleic acid leakage and glutathione consumption. As a consequence, the scaffold displayed an antibacterial rate of 97.2% and 98.5% against E. coli and S. aureus, respectively.

Oleic-acid functionalized mesoporous silica nanoparticles with a hydroxyapatite core enhanced growth of the hydrocarbon degrader Dietzia maris at oil–water interfaces

Rapid biodegradation of poorly water-soluble hydrocarbons as nonaqueous (oil) phases in contaminated aquatic environments is enabled by attachment of hydrocarbon-degrading bacteria to the oil–water interface. Herein, we report the synthesis of nanoparticles comprising a hydroxyapatite (Ca5(PO4)3(OH)) core encapsulated in a mesoporous silica shell and surface-modified with oleic acid (OA–nHAP@MSN) for targeted binding at the oil–water interface and to supply P to bacteria at the interface. P is an essential and often limiting nutrient for bacteria in hydrocarbon-contaminated environments. In microcosm experiments, where the hydrocarbon-degrading bacteria, Dietzia maris strain NWWC4, and OA–nHAP@MSN were inoculated in mineral media in contact with pure liquid hexadecane (sole C source), there was 419.6–fold growth at the hexadecane-water interface, compared to 31.2-fold in identical, but NP-free microcosms. The continuous release of P from the hydroxyapatite core in OA–nHAP@MSN to water was demonstrated in separate experiments in well mixed batch systems and was found to be pH-sensitive. Environmental Scanning Electron Microscopy showed significantly larger cell aggregates and dense biofilms in the OA–nHAP@MSN–amended systems, compared to NP-free systems. Our results demonstrate a strategy for enhancing oil–spill bioremediation using NPs targeting nutrient supply to hydrocarbon-degrading bacteria at oil-water interfaces.

The aggregated biofilm dominated by Delftia tsuruhatensis enhances the removal efficiency of 2,4-dichlorophenol in a bioelectrochemical system

Bioelectrochemical system is a prospective strategy in organic-contaminated groundwater treatment, while few studies clearly distinguish the mechanisms of adsorption or biodegradation in this process, especially when dense biofilm is formed. This study employed a single chamber microbial electrolysis cell (MEC) with two three-dimensional electrodes for removing a typical organic contaminant, 2,4-dichlorophenol (DCP) from groundwater, which inoculated with anaerobic bacteria derived from sewage treatment plant. Compared with the single biodegradation system without electrodes, the three-dimensional electrodes with a high surface enabled an increase of alpha diversity of the microbial community (increased by 52.6% in Shannon index), and provided adaptive ecological niche for more bacteria. The application of weak voltage (0.6 V) furtherly optimized the microbial community structure, and promoted the aggregation of microorganisms with the formation of dense biofilm. Desorption experiment proved that the contaminants were removed from the groundwater mainly via adsorption by the biofilm rather than biodegradation, and compared with the reactor without electricity, the bioelectrochemical system increased the adsorption capacity from 50.0% to 74.5%. The aggregated bacteria on the surface of electrodes were mainly dominated by Delftia tsuruhatensis (85.0%), which could secrete extracellular polymers and has a high adsorption capacity (0.30 mg/g electrode material) for the contaminants. We found that a bioelectrochemical system with a three-dimensional electrode could stimulate the formation of dense biofilm and remove the organic contaminants as well as their possible more toxic degradation intermediates via adsorption. This study provides important guidance for applying bioelectrochemical system in groundwater or wastewater treatment.

Antibacterial performance of a porous Cu-bearing titanium alloy by laser additive manufacturing.

Porphyromonas gingivalis (P. gingivalis) is the most common species that causes peri-implantitis. It forms an irreversible dense biofilm and causes inflammation. A novel 3D-printed porous TC4-6Cu alloy was fabricated using selective laser melting (SLM) technology for the dental implant, which is anticipated to inhibit biofilm formation. We attempted to investigate the antibacterial ability and antibacterial mechanism of the 3D-printed porous TC4-6Cu alloy against P. gingivalis. This work used scanning electron microscopy (SEM) and laser confocal microscopy (CLSM) to detect the antimicrobial ability of the alloy against sessile P. gingivalis. The results indicated that the 3D-printed porous TC4-6Cu alloy could cause bacterial fragmentation and deformation. Plate antimicrobial counting experiments showed that the antibacterial rates of the alloy against adherent bacteria and planktonic bacteria after 24h were 98.05% and 73.92%, respectively. The minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) of Cu2+ were tested to appraise the antibacterial property of the alloy against planktonic P. gingivalis. The relationship between the antibacterial mechanism of the alloy with oxidative stress was evaluated through ROS fluorescence intensity and protein leakage concentration. The results revealed that the alloy significantly eliminated adherent bacteria and inhibited biofilm formation. Moreover, 3D-printed porous TC4-6Cu alloy demonstrated significant bactericidal ability by inducing the production of reactive oxygen species (ROS), which could result in protein leakage from the bacterial cell membrane. This research may open a new perspective on the development and biomedical applications for dental implantation.

Synergistic Treatment of Multidrug-Resistant Bacterial Biofilms Using Silver Nanoclusters Incorporated into Biodegradable Nanoemulsions.

Multidrug resistance (MDR) in bacteria is a critical global health challenge that is exacerbated by the ability of bacteria to form biofilms. We report a combination therapy for biofilm infections that integrates silver nanoclusters (AgNCs) into polymeric biodegradable nanoemulsions (BNEs) incorporating eugenol. These Ag-BNEs demonstrated synergistic antimicrobial activity between the AgNCs and the BNEs. Microscopy studies demonstrated that Ag-BNEs penetrated the dense biofilm matrix and effectively disrupted the bacterial membrane. The Ag-BNE vehicle also resulted in more effective silver delivery into the biofilm than AgNCs alone. This combinacional system featured disruptionof biofilms by BNEs and enhanced delivery of AgNCs for synergy to provide highly efficient killing of MDR biofilms.

Diversity, metabolome profiling and bioactivities of benthic filamentous cyanobacteria isolated from coastal mangroves of Mayotte

IntroductionCyanobacteria are important members of the dense biofilms that colonize available substrates in mangrove habitats worldwide. However, their taxonomic diversity and biological activities have received little attention.MethodsThe occurrence of cyanobacteria is evaluated in 27 biofilms collected from mangroves in Mayotte. Filamentous cyanobacterial strains were isolated and characterized using 16S rRNA comparative gene sequence analysis. LC-MS/MS experiments were performed on the crude extracts of the faster-growing strains, and construction of their molecular network showed a conspectus of their chemical diversity. Biological activities of the strain extracts were then evaluated using standard assays.Results and discussionIsolation procedures yielded 43 strains representing 22 species-level taxa, of which only three could be assigned to existing species. Some of these strains were among the most abundant cyanobacteria present in biofilms. PCR assays did not support the production of the major cyanotoxins. Analysis of metabolites from 23 strains using both in silico tools ISDB- DNP (In silico Data Base–Dictionary of Natural Products) and MolDiscovery, revealed occurrence of godavarin K, a limonoid natural product previously isolated from the seeds of an Indian mangrove tree. This annotation was further confirmed by the marine database MarinLit, suggesting that cyanobacteria might be an alternative source of godavarin K and its four isomers. While no significant antimicrobial and cytotoxic activities were observed, some strains exhibited anthelmintic and antibiofilm activities that warrant further investigation and may be relevant to biofilm ecology. Mangrove biofilms thus appear to be an untapped reservoir of novel culturable cyanobacterial lineages, with bioactivities relevant to their biofilm lifestyle, which may be of interest for bioinspiration.

DNA Nanoflower Eye Drops with Antibiotic-Resistant Gene Regulation Ability for MRSA Keratitis Target Treatment.

Methicillin-resistant Staphylococcus aureus (MRSA) biofilm-associated bacterial keratitis is highly intractable, with strong resistance to β-lactam antibiotics. Inhibiting the MRSA resistance gene mecR1 to downregulate penicillin-binding protein PBP2a has been implicated in the sensitization of β-lactam antibiotics to MRSA. However, oligonucleotide gene regulators struggle to penetrate dense biofilms, let alone achieve efficient gene regulation inside bacteria cells. Herein, an eye-drop system capable of penetrating biofilms and targeting bacteria for chemo-gene therapy in MRSA-caused bacterial keratitis is developed. This system employed rolling circle amplification to prepare DNA nanoflowers (DNFs) encoding MRSA-specific aptamers and mecR1 deoxyribozymes (DNAzymes). Subsequently, β-lactam antibiotic ampicillin (Amp) and zinc oxide (ZnO) nanoparticles are sequentially loaded into the DNFs (ZnO/Amp@DNFs). Upon application, ZnO on the surface of the nanosystem disrupts the dense structure of biofilm and fully exposes free bacteria. Later, bearing encoded aptamer, the nanoflower system is intensively endocytosed by bacteria, and releases DNAzyme under acidic conditions to cleave the mecR1 gene for PBP2a down-regulation, and ampicillin for efficient MRSA elimination. In vivo tests showed that the system effectively cleared bacterial and biofilm in the cornea, suppressed proinflammatory cytokines interleukin 1β （IL-1β） and tumor neocrosis factor-alpha （TNF-α）, and is safe for corneal epithelial cells. Overall, this design offers a promising approach for treating MRSA-induced keratitis.

Biofilm Microenvironment-Responsive Self-Assembly Nanoreactors for All-Stage Biofilm Associated Infection through Bacterial Cuproptosis-like Death and Macrophage Re-Rousing.

Bacterial biofilm-associated infections (BAIs) are the leading cause of prosthetic implant failure. The dense biofilm structure prevents antibiotic penetration, while the highly acidic and H2 O2 -rich biofilm microenvironment (BME) dampens the immunological response of antimicrobial macrophages. Conventional treatments that fail to consistently suppress escaping planktonic bacteria from biofilm result in refractory recolonization, allowing BAIs to persist. Herein, a BME-responsive copper-doped polyoxometalate clusters (Cu-POM) combination with mild photothermal therapy (PTT) and macrophage immune re-rousing for BAI eradication at all stages is proposed. The self-assembly of Cu-POM in BME converts endogenous H2 O2 to toxic ·OH through chemodynamic therapy (CDT) and generates a mild PTT effect to induce bacterial metabolic exuberance, resulting in loosening the membrane structure of the bacteria, enhancing copper transporter activity and increasing intracellular Cu-POM flux. Metabolomics reveals that intracellular Cu-POM overload restricts the TCA cycle and peroxide accumulation, promoting bacterial cuproptosis-like death. CDT re-rousing macrophages scavenge planktonic bacteria escaping biofilm disintegration through enhanced chemotaxis and phagocytosis. Overall, BME-responsive Cu-POM promotes bacterial cuproptosis-like death via metabolic interference, while also re-rousing macrophage immune response for further planktonic bacteria elimination, resulting in all-stage BAI clearance and providing a new reference for future clinical application.