Biofilm Extracellular Polymeric Substances Research Articles

Microbial Biofilm Inhibition Using Magnetic Cross-Linked Polyphenol Oxidase Aggregates.

Microbial biofilm accumulation poses a serious threat to the environment, presents significant challenges to different industries, and exhibits a large impact on public health. Since there has not been a conclusive answer found despite various efforts, the potential green and economical methods are being focused on, particularly the innovative approaches that employ biochemical agents. In the present study, we propose a bio-nanotechnological method using magnetic cross-linked polyphenol oxidase aggregates (PPO m-CLEA) for inhibition of microbial biofilm including multidrug resistant bacteria. Free PPO solution showed only 55-60% biofilm inhibition, whereas m-CLEA showed 70-75% inhibition, as confirmed through microscopic techniques. The carbohydrate and protein contents in biofilm extracellular polymeric substances (EPSs) were reduced significantly. The m-CLEA demonstrated reusability up to 5 cycles with consistent efficiency in biofilm inhibition. Computational work was also done where molecular docking of PPO with microbial proteins associated with biofilm formation was conducted, resulting in favorable binding scores and inter-residual interactions. Overall, both in vitro and in silico results suggest that PPO interferes with microbial cell attachment and EPS formation, thereby preventing biofilm colonization.

Comparative evaluation of anti-biofilm and anti- adherence potential of plant extracts against Streptococcus mutans: A therapeutic approach for oral health

Dental caries predominantly attributed to the cariogenic nature of Streptococcus mutans, continue to pose a substantial global challenge to oral health. In response to this challenge, this study aimed to evaluate the effectiveness of leaf extracts (LEs) and essential oils (EOs) derived from different medicinal plants in inhibiting the growth of Streptococcus mutans biofilm. In vitro and in silico approaches were employed to identify active compounds and assess their inhibitory effects on S. mutans. Minimum Inhibitory Concentration (MIC) and Minimum Bactericidal Concentration (MBC) were measured to determine the anti-biofilm and anti-adherence activity against S. mutans. Biofilm viability (CFU/mL) and extracellular polymeric substance (EPS) concentration were quantified. GC-MS analysis was utilized to identify active compounds in the most effective plant extracts exhibiting anti-S. mutans activity. A high-throughput screening focused on the interaction between these compounds and the target enzyme SortaseA (SrtA) using molecular docking was performed. Results indicated that Cymbopogon citratus displayed the highest efficacy in reducing S. mutans biofilm formation and adhesion activity, achieving 90 % inhibition at an MIC value of 12 μg/mL. Among the 12 bioactive compounds identified, trans-Carvyl acetate exhibited the lowest binding energy with SrtA (-6.0 Kcal/mole). Trans-Carvyl acetate also displayed favorable pharmacokinetic properties. This study provides novel insights into the anti-S. mutans properties of C. citratus and suggests its potential as a therapeutic approach for oral health. Further research is needed to explore the combined effect of plant extracts for enhanced protection against dental caries.

Alginate Gel Encapsulated with Enzybiotics Cocktail Is Effective against Multispecies Biofilms.

The development of new and effective antibacterials for pharmaceutical or cosmetic skin care that have a low potential for the emergence and expansion of bacterial resistance is of high demand in scientific and applied research. Great hopes are placed on alternative agents such as bactericidal peptidoglycan hydrolases, depolymerases, etc. Enzybiotic-based preparations are being studied for the treatment of various infections and, among others, can be used as topical formulations and dressings with protein-polysaccharide complexes. Here, we investigate the antibiofilm properties of a novel enzybiotic cocktail of phage endolysin LysSi3 and bacteriocin lysostaphin, formulated in the alginate gel matrix and its ability to control the opportunistic skin-colonizing bacteria Staphylococcus aureus, Pseudomonas aeruginosa, and Klebsiella pneumoniae, as well as mixed-species biofilms. Our results propose that the application of SiL-gel affects different components of biofilm extracellular polymeric substances, disrupts the matrix, and eliminates the bacteria embedded in it. This composition is highly effective against biofilms composed of Gram-negative and Gram-positive species and does not possess significant cytotoxic effects. Our data form the basis for the development of antibacterial skin care products with a gentle but effective mode of action.

Microbial colonization and chemically influenced selective enrichment of bacterial pathogens on polycarbonate plastic.

Plastic pollution in aquatic environments poses significant concerns due to its potential to serve as a refuge for aquatic pathogens. However, the role of plastic surfaces and microbial biofilm interfaces in facilitating pathogen development remains poorly understood. In this study, a microcosm setup was employed to investigate the interactions between plastics and the microbial community and examine the differences in bacterial community composition and potential pathogen occurrences between the plastisphere-biofilm and surrounding seawater. Community composition analysis combined with SEM observations over time indicated that biofilm extracellular polymeric substance formation over 14 days had a link with the relative abundance and succession patterns of pathogen taxa. Colony clusters were observed on biofilms from day 7 and coincided with higher bacterial pathogen dominance. On day 14, pathogen abundance overall decreased with a potentially degrading biofilm. Pseudomonas and Pseudoalteromonas were the dominant potential pathogen groups observed in the microcosm. When further subjected to chemical treatment as an imposed environmental stress over time, biofilm-associated Psuedoalteromonas sharply increased in abundance after three days of exposure, but quickly diminished by 14days in favor of genera such as Acinetobacter, Pseudomonas, and Staphylococcus. These results suggest that environmental plastisphere-biofilms can promote the early selection, enrichment, and spread of pathogenic bacteria in the aquatic environment and could be later worsened under chemical and long-term pressure. This study provided new insights into the succession of pathogens in plastisphere biofilms, contributing to the understanding of pathogen risks involved in emerging plastisphere biofilms in light of global plastic pollution.

Combination treatment of slightly acidic electrolyzed water with electron beam irradiation to mono- and mixed biofilms of Vibrio parahaemolyticus and Pseudomonas fluorescens

Effective antimicrobial measures are needed to control cross-contamination and rapid spoilage of food caused by Vibrio parahaemolyticus (V. parahaemolyticus) and Pseudomonas fluorescens (P. fluorescens). This study evaluated the combined effect of electron beam irradiation (EBI) combined with slightly acidic electrolytic water (SAEW) treatment on suspended bacteria and single and dual-species biofilms of V. parahaemolyticus and P. fluorescens. Colony counting results showed that EBI combined with SAEW treatment effectively removed biofilm cells. Confocal laser scanning microscopy and ATP leakage analysis showed that EBI combined with SAEW treatment damaged the membrane integrity of biofilm cells and caused ATP leakage. Field emission scanning electron microscopy (SEM) observations showed that EBI with or without SAEW significantly disrupted the three-dimensional reticular structure of mono/co-cultured biofilm (extracellular polymeric substances) EPS, producing reticular perforation damage. Combined treatment with 60 mg/L SAEW and 0.5 kGy (kGy) EBI caused serious damage to the three-dimensional structure of the biofilms. The detection results of biofilm protein and DNA showed that EBI combined with SAEW could effectively destroy the extracellular polymeric matrix of biofilm, but the primary structure of the protein had no significant effect. These results suggest that SAEW(60 mg/L) combined with EBI(0.5 kGy) is effective in reducing microbial and biofilm attachment during the large yellow croaker storage processing.

Candida causes recurrent vulvovaginal candidiasis by forming morphologically disparate biofilms on the human vaginal epithelium

BackgroundRecurrent vulvovaginal candidiasis (RVVC) is a recalcitrant medical condition that affects many women of reproductive age. The importance of biofilm formation by Candida in RVVC has been recently questioned. This study aimed to elucidate the fundamental growth modes of Candida in the vagina of patients with RVVC or sporadic vulvovaginal candidiasis (VVC) and to assess their roles in the persistence of RVVC. MethodsVaginal tissues were sampled from twelve patients clinically and microbiologically diagnosed as RVVC or VVC at a post-antifungal-treatment and asymptomatic period. High-resolution scanning electron microscopy, fluorescence in situ hybridization in combination with Candida-specific 18S rRNA probes and viable fungal burden were used to qualitatively and quantitatively evaluate Candida growth in the human vagina. The presence of Candida biofilm extracellular polymeric substances was examined using confocal laser scanning microscopy and biopsy sections pre-stained with Concanavalin A. Histopathological analysis was carried out on infected vaginal tissues stained with hematoxylin and eosin. Lastly, the susceptibility of epithelium-associated Candida biofilms to fluconazole at the peak serum concentration was evaluated. ResultsCandida species grew on the vaginal epithelium of RVVC patients as morphologically disparate biofilms including monolayers, microcolonies, and macro-colonies, in addition to sporadic adherent cells. Candida biofilm growth on the vaginal epithelium was associated with mild lymphocytic infiltration of the vaginal mucosa. These epithelium-based Candida biofilms presented an important characteristic contributing to the persistence of RVVC that is the high tolerance to fluconazole. ConclusionsIn summary, our study provides direct evidence to support the presence of Candida biofilms in RVVC and an important role of biofilm formation in disease persistence.

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.

Hospital and urban wastewaters shape the matrix and active resistome of environmental biofilms

Understanding the dynamics of antibiotic resistance gene (ARG) transfer and dissemination in natural environments remains challenging. Biofilms play a crucial role in bacterial survival and antimicrobial resistance (AMR) dissemination in natural environments, particularly in aquatic systems. This study focused on hospital and urban wastewater (WW) biofilms to investigate the potential for ARG dissemination through mobile genetic elements (MGEs). The analysis included assessing the biofilm extracellular polymeric substances (EPS), microbiota composition as well as metatranscriptomic profiling of the resistome and mobilome. We produced both in vitro and in situ biofilms and performed phenotypic and genomic analyses. In the in vitro setup, untreated urban and hospital WW was used to establish biofilm reactors, with ciprofloxacin added as a selective agent at minimal selective concentration. In the in situ setup, biofilms were developed directly in hospital and urban WW pipes.We first showed that a) the composition of EPS differed depending on the growth environment (in situ and in vitro) and the sampling origin (hospital vs urban WW) and that b) ciprofloxacin impacted the composition of the EPS. The metatranscriptomic approach showed that a) expression of several ARGs and MGEs increased upon adding ciprofloxacin for biofilms from hospital WW only and b) that the abundance and type of plasmids that carried individual or multiple ARGs varied depending on the WW origins of the biofilms. When the same plasmids were present in both, urban and hospital WW biofilms, they carried different ARGs. We showed that hospital and urban wastewaters shaped the structure and active resistome of environmental biofilms, and we confirmed that hospital WW is an important hot spot for the dissemination and selection of antimicrobial resistance. Our study provides a comprehensive assessment of WW biofilms as crucial hotspots for ARG transfer. Hospital WW biofilms exhibited distinct characteristics, including higher eDNA abundance and expression levels of ARGs and MGEs, highlighting their role in antimicrobial resistance dissemination. These findings emphasize the importance of understanding the structural, ecological, functional, and genetic organization of biofilms in anthropized environments and their contribution to antibiotic resistance dynamics.

Extracellular matrix-degrading enzymes as a biofilm control strategy for food-related microorganisms.

Biofilm is one of the major problems in food industries and is difficult to be removed or prevented by conventional sanitizers. In this review, we discussed the extracellular matrix-degrading enzymes as a strategy to control biofilms of foodborne pathogenic and food-contaminating bacteria. The biofilms can be degraded by using the enzymes targeting proteins, polysaccharides, extracellular DNA, or lipids which mainly constitute the extracellular polymeric substances of biofilms. However, the efficacy of enzymes varies by the growth medium, bacterial species, strains, or counterpart microorganisms due to a high variation in the composition of extracellular polymeric substances. Several studies demonstrated that the combined treatment using conventional sanitizers or multiple enzymes can synergistically enhance the biofilm removal efficacies. In this review, the application of the immobilized enzymes on solid substrates is also discussed as a potential strategy to prevent biofilm formation on food contact surfaces.

Reducing the matrix effect in mass spectral imaging of biofilms using flow-cell culture.

The interactions between soil microorganisms and soil minerals play a crucial role in the formation and evolution of minerals and the stability of soil aggregates. Due to the heterogeneity and diversity of the soil environment, the under-standing of the functions of bacterial biofilms in soil minerals at the microscale is limited. A soil mineral-bacterial biofilm system was used as a model in this study, and it was analyzed by time-of-flight secondary ion mass spectrometry (ToF-SIMS) to acquire molecular level information. Static culture in multi-wells and dynamic flow-cell culture in microfluidics of biofilms were investigated. Our results show that more characteristic molecules of biofilms can be observed in SIMS spectra of the flow-cell culture. In contrast, biofilm signature peaks are buried under the mineral components in SIMS spectra in the static culture case. Spectral overlay was used in peak selection prior to performing Principal component analysis (PCA). Comparisons of the PCA results between the static and flow-cell culture show more pronounced molecular features and higher loadings of organic peaks of the dynamic cultured specimens. For example, fatty acids secreted from bacterial biofilm extracellular polymeric substance are likely to be responsible for biofilm dispersal due to mineral treatment up to 48h. Such findings suggest that the use of microfluidic cells to dynamically culture biofilms be a more suitable method for reducing the matrix effect arisen from the growth medium and minerals as a perturbation fac-tor for improved spectral and multivariate analysis of complex mass spectral data in ToF-SIMS. These results show that the interaction mechanism between biofilms and soil minerals at the molecular level can be better studied using the flow-cell culture and advanced mass spectral imaging techniques like ToF-SIMS.

Long-term operation and biofouling of graphene oxide membrane in practical water treatment: Insights from performance and biofilm characteristics

The widespread research of graphene oxide (GO) membranes has revealed the need to investigate their long-term performance and associated biofouling behavior in practical water treatment conditions. In this study, we have evaluated the long-term operation of GO membranes from the perspective of filtration performance and biofilm characteristics, based on two representative GO membranes (V-GO and P-GO, fabricated by vacuum and pressure filtration, respectively) with different surface properties and a natural surface water, using a gravity driven membrane system. The results showed that the GO membranes were capable of achieving sustainable water purification over 45 days of operation, associated with a complete rejection for biopolymers and desirable removal for fluorescent organic matters. Furthermore, the V-GO membrane with a rougher surface, reduced hydrophilicity and higher sheet-edge exposure than that of the P-GO membrane formed a highly heterogeneous and porous biofilm with high permeability, despite its greater biofilm thickness and extracellular polymeric substances (EPS) contents. In contrast, the P-GO membrane biofilm exhibited a thin but dense structure, and therefore an increased hydraulic resistance (∼1.5 times greater than V-GO membrane). The microbial community analysis revealed that bacteria related to biofilm formation was richer in V-GO biofilm. While bacteria associated with the degradation of organic matters and the aggregation of microorganisms accounted for a greater proportion in P-GO biofilm. These factors were responsible for forming a thick biofilm for V-GO, and a thin but high resistance biofilm for P-GO membrane. Our findings highlight the sustainable water purification performance of the GO membranes, and the P-GO membrane can alleviate biofilm formation but not necessarily reduce biofouling during the long-term filtration, while this was opposite in case of the V-GO membrane.

Electroactive microorganisms synthesizing iron sulfide nanoparticles for enhanced hexavalent chromium removal in microbial fuel cells

Microbial fuel cells (MFCs) have been considered a promising technology for Cr6+ removal, but they are limited by Cr6+-reducing biocathodes with low extracellular electron transfer (EET) and poor microbial activity. In this study, three kinds of nano-FeS hybridized electrode biofilms, obtained through synchronous biosynthesis (Sy-FeS), sequential biosynthesis (Se-FeS) and cathode biosynthesis (Ca-FeS), were applied as biocathodes for Cr6+ removal in MFCs. The Ca-FeS biocathode exhibited the best performance due to the superior properties of biogenic nano-FeS (e.g., more synthetic amount, smaller particle size, better dispersion). The MFC with the Ca-FeS biocathode achieved the highest power density (42.08 ± 1.42 mW/m2) and Cr6+ removal efficiency (99.18 ± 0.1 %), which were 1.42 and 2.08 times as high as those of the MFC with the normal biocathode, respectively. The synergistic effects of nano-FeS and microorganisms enhanced the bioelectrochemical reduction of Cr6+, first realizing deep reduction of Cr6+ to Cr0 in biocathode MFCs. This significantly alleviated the cathode passivation caused by Cr3+ deposition. In addition, the hybridized nano-FeS as “armor” layers protected the microbes from toxic attack by Cr6+, improving the biofilm physiological activity and extracellular polymeric substances (EPS) secretion. The hybridized nano-FeS as “electron bridges” facilitated the microbial community to form a balanced, stable and syntrophic ecological structure. This study proposes a novel strategy through the cathode in-situ biosynthesis of nanomaterials to fabricate hybridized electrode biofilms with enhanced EET and microbial activity for toxic pollutant treatment in bioelectrochemical systems.

Predation preference for extracellular polysaccharides by paramecia and rotifers may have accelerated the decline of membrane biofilm hydraulic resistance

The hydraulic resistance of biofilm layer on membranes impacts the filtration resistance significantly. The effect of predation by two model microfauna (i.e., paramecia and rotifers) on the hydraulic resistance, structure, extracellular polymeric substance (EPS), and bacterial community of biofilms developed on supporting materials (i.e., nylon mesh) was evaluated in this study. Long-term experiments demonstrated that predation could alter biofilm compositions and accelerated the decline of hydraulic resistance by increasing biofilm heterogeneity and deformation. Importantly, predation preference of paramecia and rotifers on biofilm components were further investigated for the first time by tracking the fluorescence change in the predator bodies after exposure to the stained biofilms. Results indicated that after 12-hour’s incubation, the ratio of extracellular α-polysaccharides (α-PS) to proteins (PN) within the bodies of paramecia and rotifers increased to 2.6 and 3.9, respectively, which was 0.76 in the original biofilms. The ratios of α-PS/live cells within paramecia and rotifers increased to 1.42 and 1.64 from 0.81 in the original biofilms. The ratio of live/dead cells in the predator bodies, however, changed slightly compared to the original biofilms. These results clearly and directly evidenced that both paramecia and rotifers could feed on biofilm EPS and cells, but having a significant preference for PS over PN and cells. Since extracellular PS is recognized as a primary biofilm adhesion agent, the preference for PS could better explain why predation had accelerated the disintegration and hydraulic resistance decline of mesh biofilms.

Deciphering the distribution and microbial secretors of extracellular polymeric substances associated antibiotic resistance genes in tube wall biofilm

Antibiotics and disinfectants have both been proposed to exert selective pressures on the biofilm as well as affecting the emergence and spread of antibiotic resistance genes (ARGs). However, the transfer mechanism of ARGs in drinking water distribution system (DWDS) under the coupling effect of antibiotics and disinfectants has not been completely understood. In the current study, four lab-scale biological annular reactors (BARs) were constructed to evaluate the effects of sulfamethoxazole (SMX) and NaClO coupling in DWDS and reveal the related mechanisms of ARGs proliferation. TetM was abundant in both the liquid phase and the biofilm, and redundancy analysis showed that the total organic carbon (TOC) and temperature were significantly correlated with ARGs in the water phase. There was a significant correlation between the relative abundance of ARGs in the biofilm phase and extracellular polymeric substances (EPS). Additionally, the proliferation and spread of ARGs in water phase were related to microbial community structure. Partial least-squares path modeling showed that antibiotic concentration may influence ARGs by affecting MGEs. These findings help us to better understand the diffusion process of ARGs in drinking water and provide a theoretical support for technologies to control ARGs at the front of pipeline.

Low-rate ferrate dosing damages the microbial biofilm structure through humic substances destruction and facilitates the sewer biofilm control

The microbial activities in sewer biofilms are recognized as a major reason for sewer pipe corrosion, malodor, and greenhouse gas emissions. However, conventional methods to control sewer biofilm activities were based on the inhibitory or biocidal effect of chemicals and often required long exposure time or high dosing rates due to the protection of sewer biofilm structure. Therefore, this study attempt to use ferrate (Fe(VI)), a green and high-valent iron, at low dosing rates to damage the sewer biofilm structure so as to enhance sewer biofilm control efficiency. The results showed the biofilm structure started to crush when the Fe(VI) dosage was 15 mg Fe(VI)/L and the damage enhanced with the increasing dosage. The determination of extracellular polymeric substances (EPS) showed that Fe(VI) treatment at 15-45 mgFe/L mainly decreased the content of humic substances (HS) in biofilm EPS. This is because the functional groups, such as C-O, -OH, and C=O, which held the large molecular structure of HS, were the primary target of Fe(VI) treatment as suggested by 2D-Fourier Transform Infrared spectra. As a result, the coiled chain of EPS maintained by HS was turned to extended and dispersed and consequently led to a loosed biofilm structure. The XDLVO analysis suggested that both the microbial interaction energy barrier and secondary energy minimum were increased after Fe(VI) treatment, suggesting that the treated biofilm was less likely to aggregate and easier to be removed by the shear stress caused by high wastewater flow. Moreover, combined Fe(VI) and free nitrous acid (FNA) dosing experiments showed for achieving 90% inactivation, the FNA dosing rate could be reduced by 90% with the exposure time decreasing by 75% at a low Fe(VI) dosing rate and the total cost was substantially decreased. These results suggested that applying low-rate Fe(VI) dosing for sewer biofilm structure destruction is expected to be an economical way to facilitate sewer biofilm control.

Disruption of biofilms in periodontal disease through the induction of phase transition by cationic dextrans.

Biofilm of oral pathogenic microorganisms induced by their multiplication and coaggregation would lead to periodontitis. In biofilms, the extracellular polymeric substances (EPS) as a protective shield encapsulates the individual bacteria, protecting them against attack. To alleviate periodontal disease, disrupting the EPS of pathogenic bacteria is crucial and challenging. Based on the sufficient capacity of disorganizing EPS of our designed cationic dextrans, we hypothesized that these polymers could be competent in relieving periodontitis. We validated that cationic dextrans could induce the phase transition of EPS in biofilms, especially the Porphyromonas gingivalis (P. gingivalis), a keystone periodontal pathogen, thus effectively destroying biofilm in vitro. More importantly, satisfactory in vivo treatment was achieved in a rat periodontal disease model. In summary, the study exploited a practical and effective strategy to treat periodontitis with cationic dextrans' powerful biofilm-controlling potential. STATEMENT OF SIGNIFICANCE: Periodontal disease is closely related to dental plaque biofilms on the tooth surface. The biofilm forms gel structures and shields the bacteria underneath, thus protecting oral pathogens from traditional anti-bacterial reagents. Due to limited penetration into gel, the efficacy of these reagents in biofilm elimination is restricted. Our designed cationic dextran could wipe out the coverage of gel-like EPS to disperse encapsulated bacteria. Such superior capacity endowed them with satisfactory effect in disrupting biofilm. Notably, in a rat periodontitis model, cationic dextrans dramatically suppressed alveolar bone loss and alleviated periodontal inflammation by controlling dental plaque. Given the increasing global concerns about periodontal disease, it's worth expanding the application of cationic dextrans both scientifically and clinically.

Sensitive and Accurate Differentiation of Extracellular Polymeric Substance Hydrolysis Using Flow Cytometry during Biofilm Extracellular Polymeric Substance Extraction

Extracellular polymeric substances (EPS) affect the formation, properties, and function of biofilms. Understanding the roles of EPS in biofilm-related processes depends on reliable extraction and evaluation methods. However, differentiating true EPS from intracellular organic polymers released by cell lysis has proven to be difficult during EPS extraction. In this study, thermal and cation exchange resin (CER) treatments were used to extract biofilm EPS. SYTOX Green (SG) dye was used to stain nucleic acid (NA), and the enhanced fluorescence signal (FS) from the NA-SG complex was sensitively recorded by flow cytometry (FC). Results showed that thermal extraction could achieve effective EPS release, but a higher temperature and longer time caused a significant cell activity decline, protein denaturation, and cell lysis. In contrast, CER extraction had no effects on cell activity and organics denaturation, and it only caused minor cell lysis at a high CER dose. Combining all the results indicates that CER extraction using 70 g-CER/g-VSS for 2 h provided reliable and efficient extraction of biofilm EPS: high EPS extraction efficiency with minimal EPS denaturation, cell lysis, and loss of cell activity. FC combined with SG staining should be applicable for evaluating the effectiveness of EPS extraction from any microbial aggregates.

Effects of environment-relevant concentrations of antibiotics on seawater Chlorella sp. biofilm in artificial mariculture effluent

Attached cultivation of microalgae using mariculture effluent could simultaneously realize cost-effective biomass production and effluent polishing, and the obtained microalgal biofilm could be readily applied to aquatic animals that feed directly on biofilms. However, the widespread existence of antibiotics in mariculture effluent may have inhibitory effects on microalgae. Existing studies regarding the influences of antibiotics on microalgae were overwhelmingly mostly conducted in suspended freshwater cultures, and the explored antibiotic concentrations were several orders of magnitudes higher than realist effluent, rendering limited guidance for practical applications. This study, therefore, studied three commonly observed antibiotics in mariculture effluent, i.e. sulfamethoxazole (SMX), tetracycline (TL), and clarithromycin (CLA), and explored their effects on attached seawater Chlorella sp. growth, nutrients removal, and antioxidative properties at environment-relevant concentrations (0.1–200 μg/L). Hormesis was shared among the three antibiotics, and highest biofilm density (14.6 g/m2) was observed under 0.1 μg/L SMX. SMX displayed strongest chlorophyll impairment, while SMX, TL, and CLA respectively favored lipid (up to 42.4 %), protein (up to 39.6 %), and polysaccharides (up to 65.4 %) accumulation. Highest correlation between biofilm density and extracellular polymeric substances (EPS) contents as well as highest total nitrogen (TN) removal were observed under TL exposure, while total phosphorus (TP) removal among all groups met the strictest local discharge standard for mariculture effluent (TP < 0.2 mg/L). Antioxidative responses revealed least stress towards attached seawater Chlorella sp. under TL treatment, while best acclimation under SMX exposure. Potential coping mechanisms of this microalgae under each antibiotic exposure were proposed as well. This study expands the knowledge of microalgae biofilm under environment-relevant concentrations of antibiotics.

Tetracycline accumulation in biofilms enhances the selection pressure on Escherichia coli for expression of antibiotic resistance.

Microorganisms are present as either biofilm or planktonic species in natural and engineered environments. Little is known about the selection pressure emanating from exposure to sub-minimal inhibitory concentration of antibiotics on planktonic vs. biofilm bacteria. In this study, an E. coli bioreporter was used to develop biofilms on glass and high-density polyethylene (HDPE) surfaces, and compared with the corresponding planktonic bacteria in antibiotic resistance expression when exposed to a range of μg/L levels of tetracycline. The antibiotic resistance-associated fluorescence emissions from biofilm E. coli reached up to 1.6 times more than those from planktonic bacteria. The intensively developed biofilms on glass surfaces caused the embedded bacteria to experience higher selection pressure and express more antibiotic resistance than those on HDPE surfaces. The temporal pattern of fluorescence emissions from biofilm E. coli was consistent with the biofilm-developing processes during the experimental period. The increased expression of antibiotic resistance from biofilm bacteria could be attributed to the high affinity of tetracycline with extracellular polymeric substances (EPS). The enhanced accumulation of tetracycline in biofilms could exert higher selection pressure on the embedded bacteria. These results suggest that in many natural and engineered systems the higher antibiotic resistance in biofilm bacteria could be attributed partially to the retention antibiotics by the EPS in biofilms.

Microscopic influence mechanisms of polysaccharide on the adsorption and corrosion inhibition performance of imidazoline on metal surface

Microbiologically influenced corrosion caused by biofilm adhesion is the most complex type of corrosion in the oil–gas pipeline. However, the mechanism of biofilm extracellular polymeric substances’ main components that influence the performance of corrosion inhibitors remains unclear. Therefore, molecular dynamics simulations were performed to gain insights into the aggregation and adsorption of alginate and imidazoline, representative compounds of extracellular polysaccharide and corrosion inhibitor, respectively, on iron surface. Results demonstrated that the corrosion inhibition of imidazoline was mainly due to the formation of a regular bilayer structure at high concentration that provided mass transport barrier and effectively expelled the corrosive Cl- and water from metal surface. However, this capability disappeared when high concentration alginates pre-adsorbed on the metal surface. It was mainly attributed to the fact that the alginates significantly decreased the imidazoline coverage on metal surface (up to 84%), wherein the majority of imidazoline molecules suspended in the bulk water. Moreover, high concentration alginate could form porous network structure with permeable channels, which allowed Cl- and water to pass through and approach the metal surface.