Analysis of the microbial community structure in a membrane bioreactor during initial stages of filtration

Membrane biofouling was investigated during the early stages of filtration in a laboratory-scale membrane bioreactor operated on molasses wastewater. The bacterial diversity and composition of the membrane biofilm and activated sludge were analyzed using terminal restriction fragment length polymorphism coupled with 16S rRNA clone library construction and sequencing. The amount of extracellular polymeric substances produced by bacteria was investigated using spectroscopic methods. The results reveal that the bacterial community of activated sludge differs significantly from that of the membrane biofilm, especially at the initial phase. Phylogenetic analysis based on 16S rRNA gene sequences identified 25 pioneer OTUs responsible for membrane surface colonization. Also, the relationship between the identified bacterial strains and the system specifications was explored.     

Molecular characterization of the bacterial communities in the different compartments of a full-scale reverse-osmosis water purification plant

The origin, structure, and composition of biofilms in various compartments of an industrial full-scale reverse-osmosis (RO) membrane water purification plant were analyzed by molecular biological methods. Samples were taken when the RO installation suffered from a substantial pressure drop and decreased production. The bacterial community of the RO membrane biofilm was clearly different from the bacterial community present at other locations in the RO plant, indicating the development of a specialized bacterial community on the RO membranes. The typical freshwater phylotypes in the RO membrane biofilm (i.e., Proteobacteria, Cytophaga-Flexibacter-Bacteroides group, and Firmicutes) were also present in the water sample fed to the plant, suggesting a feed water origin. However, the relative abundances of the different species in the mature biofilm were different from those in the feed water, indicating that the biofilm was actively formed on the RO membrane sheets and was not the result of a concentration of bacteria present in the feed water. The majority of the microorganisms (59% of the total number of clones) in the biofilm were related to the class Proteobacteria, with a dominance of Sphingomonas spp. (27% of all clones). Members of the genus Sphingomonas seem to be responsible for the biofouling of the membranes in the RO installation.     

Nitric Oxide Treatment for the Control of Reverse Osmosis Membrane Biofouling

Biofouling remains a key challenge for membrane-based water treatment systems. This study investigated the dispersal potential of the nitric oxide (NO) donor compound, PROLI NONOate, on single- and mixed-species biofilms formed by bacteria isolated from industrial membrane bioreactor and reverse osmosis (RO) membranes. The potential of PROLI NONOate to control RO membrane biofouling was also examined. Confocal microscopy revealed that PROLI NONOate exposure induced biofilm dispersal in all but two of the bacteria tested and successfully dispersed mixed-species biofilms. The addition of 40 μM PROLI NONOate at 24-h intervals to a laboratory-scale RO system led to a 92% reduction in the rate of biofouling (pressure rise over a given period) by a bacterial community cultured from an industrial RO membrane. Confocal microscopy and extracellular polymeric substances (EPS) extraction revealed that PROLI NONOate treatment led to a 48% reduction in polysaccharides, a 66% reduction in proteins, and a 29% reduction in microbial cells compared to the untreated control. A reduction in biofilm surface coverage (59% compared to 98%, treated compared to control) and average thickness (20 μm compared to 26 μm, treated compared to control) was also observed. The addition of PROLI NONOate led to a 22% increase in the time required for the RO module to reach its maximum transmembrane pressure (TMP), further indicating that NO treatment delayed fouling. Pyrosequencing analysis revealed that the NO treatment did not significantly alter the microbial community composition of the membrane biofilm. These results present strong evidence for the application of PROLI NONOate for prevention of RO biofouling.     

Influence of membrane surface properties on the behavior of initial bacterial adhesion and biofilm development onto nanofiltration membranes

In order to investigate biofouling problems, the fundamental behaviors of initial bacterial adhesion and biofilm development on four different nanofiltration (NF) membranes were evaluated using Pseudomonas aeruginosa PAO1 as a model bacterial strain. Initial cell adhesion was considerably higher on an aromatic polyamide-based NF membrane with a hydrophobic and rough surface, whereas cell aggregation on a polypiperazine-based NF membrane with a relatively hydrophilic and smooth surface was lower. Moreover, significant differences in the structural heterogeneity of the biofilms were observed among the four NF membranes. This study shows that the surface roughness and hydrophobicity of a membrane play an important role in determining initial cell adhesion, aggregation and favorable localization sites for colony formation. In addition, it was found that biofilm development was strongly influenced by the surface morphology of a membrane.     

Dynamic bacterial communities on reverse-osmosis membranes in a full-scale desalination plant

To better understand biofouling of seawater reverse osmosis (SWRO) membranes, bacterial diversity was characterized in the intake water, in subsequently pretreated water and on SWRO membranes from a full-scale desalination plant (FSDP) during a 9 month period. 16S rRNA gene fingerprinting and sequencing revealed that bacterial communities in the water samples and on the SWRO membranes were very different. For the different sampling dates, the bacterial diversity of the active and the total bacterial fractions of the water samples remained relatively stable over the sampling period whereas the bacterial community structure on the four SWRO membrane samples was significantly different. The richness and evenness of the SWRO membrane bacterial communities increased with usage time with an increase in the Shannon diversity index of 2.2 to 3.7. In the oldest SWRO membrane (330 days), no single operational taxonomic unit (OTU) dominated and the majority of the OTUs fell into the Alphaproteobacteria or the Planctomycetes. In striking contrast, a Betaproteobacteria OTU affiliated to the genus Ideonella was dominant and exclusively found in the membrane used for the shortest time (10 days). This suggests that bacteria belonging to this genus could be one of the primary colonizers of the SWRO membrane. Knowledge of the dominant bacterial species on SWRO membranes and their dynamics should help guide culture studies for physiological characterization of biofilm forming species.     

A new concept in polymeric thin-film composite nanofiltration membranes with antibacterial properties

A new, thin film, biofouling resistant, nanofiltration (NF) membrane was fabricated with two key characteristics, viz. a low rate of silver (Ag) release and long-lasting antibacterial properties. In the new approach, nanoparticles were embedded completely in a polymeric thin-film layer. A comparison was made between the new thin-film composite (TFC), NF membrane and thin-film nanocomposite (TFN), and antibacterial NF membranes. Both types of NF membrane were fabricated by interfacial polymerization on a polysulphone sublayer using m-phenylenediamine and trimesoyl chloride as an amine monomer and an acid chloride monomer, respectively. Energy dispersive X-ray (EDX) microanalysis demonstrated the presence of Ag nanoparticles. Scanning electron microscopy (SEM) and atomic force microscopy (AFM) were used to study the cross-sectional and surface morphological properties of the NF membranes. Permeability and salt rejection were tested using a dead-end filtration cell. Ag leaching from the membranes was measured using inductively coupled mass spectrometry (ICP–MS). Morphological studies showed that the TFC NF membranes had better thin-film formation (a more compact structure and a smoother surface) than TFN NF membranes. Performance experiments on TFC NF membranes revealed that permeability was good, without sacrificing salt rejection. The antibacterial properties of the fabricated membranes were tested using the disk diffusion method and viable plate counts. The antibiofouling properties of the membranes were examined by measuring the quantity of bacterial cells released from the biofilm formed (as a function of the amount of biofilm present). A more sensitive surface was observed compared to that of a typical antibacterial NF membrane. The Ag leaching rates were low, which will likely result in long-lasting antibacterial and biofouling resistant properties.     

The application of nitric oxide to control biofouling of membrane bioreactors

A novel strategy to control membrane bioreactor (MBR) biofouling using the nitric oxide (NO) donor compound PROLI NONOate was examined. When the biofilm was pre‐established on membranes at transmembrane pressure (TMP) of 88–90 kPa, backwashing of the membrane module with 80 μM PROLI NONOate for 45 min once daily for 37 days reduced the fouling resistance (R_f) by 56%. Similarly, a daily, 1 h exposure of the membrane to 80 μM PROLI NONOate from the commencement of MBR operation for 85 days resulted in reduction of the TMP and R_f by 32.3% and 28.2%. The microbial community in the control MBR was observed to change from days 71 to 85, which correlates with the rapid TMP increase. Interestingly, NO‐treated biofilms at 85 days had a higher similarity with the control biofilms at 71 days relative to the control biofilms at 85 days, indicating that the NO treatment delayed the development of biofilm bacterial community. Despite this difference, sequence analysis indicated that NO treatment did not result in a significant shift in the dominant fouling species. Confocal microscopy revealed that the biomass of biopolymers and microorganisms in biofilms were all reduced on the PROLI NONOate‐treated membranes, where there were reductions of 37.7% for proteins and 66.7% for microbial cells, which correlates with the reduction in TMP. These results suggest that NO treatment could be a promising strategy to control biofouling in MBRs.     

Bacterial community diversity in paper mills processing recycled paper

Paper mills processing recycled paper suffer from biofouling causing problems both in the mill and final product. The total bacterial community composition and identification of specific taxa in the process water and biofilms at the stock preparation and paper machine areas in a mill with recycled paper pulp was described by using a DNA-based approach. Process water in a similar mill was also analyzed to investigate if general trends can be found between mills and over time. Bacterial community profiles, analyzed by terminal-restriction fragment length polymorphism (T-RFLP), in process water showed that the dominant peaks in the profiles were similar between the two mills, although the overall composition was unique for each mill. When comparing process water and biofilm at different locations within one of the mills, we observed a separation according to location and sample type, with the biofilm from the paper machine being most different. 16S rRNA gene clone libraries were generated and 404 clones were screened by RFLP analysis. Grouping of RFLP patterns confirmed that the biofilm from the paper machine was most different. A total of 99 clones representing all RFLP patterns were analyzed, resulting in sequences recovered from nine bacterial phyla, including two candidate phyla. Bacteroidetes represented 45% and Actinobacteria 23% of all the clones. Sequences with similarity to organisms implicated in biofouling, like Chryseobacterium spp. and Brevundimonas spp., were recovered from all samples even though the mill had no process problems during sampling, suggesting that they are part of the natural paper mill community. Moreover, many sequences showed little homology to as yet uncultivated bacteria implying that paper mills are interesting for isolation of new organisms, as well as for bioprospecting.     

Membrane biofouling characterization: effects of sample preparation procedures on biofilm structure and the microbial community

Ensuring the quality and reproducibility of results from biofilm structure and microbial community analysis is essential to membrane biofouling studies. This study evaluated the impacts of three sample preparation factors (ie number of buffer rinses, storage time at 4°C, and DNA extraction method) on the downstream analysis of nitrifying biofilms grown on ultrafiltration membranes. Both rinse and storage affected biofilm structure, as suggested by their strong correlation with total biovolume, biofilm thickness, roughness and the spatial distribution of EPS. Significant variations in DNA yields and microbial community diversity were also observed among samples treated by different rinses, storage and DNA extraction methods. For the tested biofilms, two rinses, no storage and DNA extraction with both mechanical and chemical cell lysis from attached biofilm were the optimal sample preparation procedures for obtaining accurate information about biofilm structure, EPS distribution and the microbial community.     

Feasibility of supercritical CO₂ treatment for controlling biofouling in the reverse osmosis process

Physical cleaning and/or chemical cleaning have been generally used to control biofouling in the reverse osmosis (RO) process. However, conventional membrane cleaning methods to control biofouling are limited due to the generation of by-products and the potential for damage to the RO membranes. In this study, supercritical carbon dioxide (SC CO(2)) treatment, an environmentally friendly technique, was introduced to control biofouling in the RO process. SC CO(2) (100 bar at 35°C) treatment was performed after biofouling was induced on a commercial RO membrane using Pseudomonas aeruginosa PA01 GFP as a model bacterial strain. P. aeruginosa PA01 GFP biofilm cells were reduced on the RO membrane by >8 log within 30 min, and the permeate flux was sufficiently recovered in a laboratory-scale RO membrane system without any significant damage to the RO membrane. These results suggest that SC CO(2) treatment is a promising alternative membrane cleaning technique for biofouling in the RO process.     

Identifying pioneer bacterial species responsible for biofouling membrane bioreactors

More effective control of membrane biofouling in membrane bioreactors (MBRs) lies in the fundamental understanding of the pioneer microorganisms responsible for surface colonization that leads to biofilm formation. In this study, the composition of the planktonic and sessile microbial communities inhabiting four laboratory-scale MBR systems were compared using amplified ribosomal DNA restriction analysis (ARDRA) and 16S ribosomal DNA gene sequencing. The ARDRA results suggest that the microbial communities on membrane surfaces could be very different from the ones in the suspended biomass. Phylogenetic analysis based on the 16S rRNA gene sequences provided a list of bacteria that might be the pioneers of surface colonization on microfiltration membranes. The results further suggested that research on the mechanisms of cell attachment in such an engineering environment could be critical for future development of appropriate biofouling control strategies.     

Non-destructive approaches for assessing biofouling of household reverse osmosis membranes

Abstract

This study determined economic non-destructive methods to assess biofouling in point of use reverse osmosis (RO) membrane treatment systems. Three parallel household RO membrane units were operated under controlled feed water conditions to promote biofouling, inorganic fouling and a combination of both. Operational and biological parameters were monitored throughout the systems’ lifespan. Membrane autopsies assessed the degree and type of fouling. Statistical models determined statistically relevant parameters for fouling types that were validated with membrane autopsies. Permeate flow rates decreased differently with biofouling vs inorganic fouling. Large increases in permeate conductivity were noted in membranes suffering from biofouling and not in inorganically fouled membranes. The concentration of cell clumps from detached biofilm in the retentate increased in membranes experiencing biofouling and no increase was seen for inorganically fouled membranes. A combination of these methods could be used to conveniently assess the types of fouling experienced by RO systems.     

Feasibility of supercritical CO2 treatment for controlling biofouling in the reverse osmosis process

Physical cleaning and/or chemical cleaning have been generally used to control biofouling in the reverse osmosis (RO) process. However, conventional membrane cleaning methods to control biofouling are limited due to the generation of by-products and the potential for damage to the RO membranes. In this study, supercritical carbon dioxide (SC CO₂) treatment, an environmentally friendly technique, was introduced to control biofouling in the RO process. SC CO₂ (100 bar at 35°C) treatment was performed after biofouling was induced on a commercial RO membrane using Pseudomonas aeruginosa PA01 GFP as a model bacterial strain. P. aeruginosa PA01 GFP biofilm cells were reduced on the RO membrane by >8 log within 30 min, and the permeate flux was sufficiently recovered in a laboratory-scale RO membrane system without any significant damage to the RO membrane. These results suggest that SC CO₂ treatment is a promising alternative membrane cleaning technique for biofouling in the RO process.     

Microbial fouling of reverse-osmosis membranes used in advanced wastewater treatment technology: chemical, bacteriological, and ultrastructural analyses.

Biofouling of reverse-osmosis membranes was investigated at an advanced wastewater treatment facility. Cellulose diacetate membranes operated for approximately 4,000 h became uniformly coated with a mucilaginous fouling layer. The fouling material was approximately 93% water by weight, and nearly 90% of the dehydrated residue was organic in composition. Calcium, phosphorous, sulfur, and chlorine were the major inorganic constituents detected. Protein and carbohydrate represented as much as 30 and 17%, respectively, of the dry weight of the biofilm. Bacteriological plate counts indicated up to 5.6 X 10(6) CFU/cm2 of membrane surface. Accumulation of [3H]glucose in the biofilm and measurement of ATP indicated that the fouling bacteria were metabolically active in situ. The genus Acinetobacter and the Flavobacterium-Moraxella group were the major generic groups associated with the feedwater surface of the membrane, whereas species of the generic groups Acinetobacter, Pseudomonas-Alcaligenes, and Bacillus-Lactobacillus predominated on the permeate water surface. Electron microscopy revealed that the biofilm on the feedwater surface of the membrane was 10 to 20 microns thick and was composed of several layers of compacted bacterial cells, many of which were partially or completely autolyzed. The bacteria were firmly attached to the membrane surface by an extensive network of extracellular polymeric fibrils. Polyester (Texlon) support fibers located on the permeate surface of the reverse osmosis membranes were sparsely colonized, suggesting bacterial regrowth in the product water collection system.     

Analysis of microbial community structure in a biofilm on membrane surface in the submerged membrane bioreactor treating domestic wastewater on the basis of respiratory quinone profiles

The objective of this study was to investigate the microbial community structure of the biofouling film formed on hollow-fiber membrane surfaces in the submerged membrane bioreactor (SMBR) with a nitrification-denitrification process. In this experiment, aeration was conducted intermittently (60 min off, 90 min on) cyclic anoxic and oxic conditions in the SMBR. The dominant quinone types of biofilm on the membrane surface in an intermittently aerated SMBR were ubiquinone (UQs)-8, -10, followed by menaquinones (MKs)-8(H4), -8(H2) and -7, but those of suspended microorganisms were UQ-8, UQ-10 followed by MKs-8, -9(H4) and -6. The change in quinone profiles of biofilm on the membrane surface suggested that UQ-9, MK-7, MK-8(H2) and MK-8(H4) contributed to microbiological fouling in the intermittently aerated SMBR treating domestic wastewater. The microbial diversities of suspended microorganisms and biofilm, calculated based on the composition of all quinones, were 9.5 and 10.9, respectively.     

Use of rhamnolipid biosurfactant for membrane biofouling prevention and cleaning

Rhamnolipids were evaluated as biofouling reducing agents in this study. The permeability of the bacterial outer membrane was increased by rhamnolipids while the growth rate of Pseudomonas aeruginosa was not affected. The surface hydrophobicity was increased through the release of lipopolysaccharides and extracellular polymeric substances from the outer cell membrane. Rhamnolipids were evaluated as agents for the prevention and cleaning of biofilms. A high degree of biofilm detachment was observed when the rhamnolipids were used as a cleaning agent. In addition, effective biofilm reduction occurred when rhamnolipids were applied to various species of Gram-negative bacteria isolated from seawater samples. Biofilm reduction using rhamnolipids was comparable to commercially available surfactants. In addition, 20% of the water flux was increased after rhamnolipid treatment (300 μg ml^?1, 6 h exposure time) in a dead-end filtration system. Rhamnolipids appear to have promise as biological agents for reducing membrane biofouling.     

Composition and variability of biofouling organisms in seawater reverse osmosis desalination plants

Seawater reverse osmosis (SWRO) membrane biofouling remains a common challenge in the desalination industry, but the marine bacterial community that causes membrane fouling is poorly understood. Microbial communities at different stages of treatment processes (intake, cartridge filtration, and SWRO) of a desalination pilot plant were examined by both culture-based and culture-independent approaches. Bacterial isolates were identified to match the genera Shewanella, Alteromonas, Vibrio, and Cellulophaga based on 16S rRNA gene sequencing analysis. The 16S rRNA gene clone library of the SWRO membrane biofilm showed that a filamentous bacterium, Leucothrix mucor, which belongs to the gammaproteobacteria, accounted for nearly 30% of the clone library, while the rest of the microorganisms (61.2% of the total clones) were related to the alphaproteobacteria. 16S rRNA gene terminal restriction fragment length polymorphism (T-RFLP) analysis indicated that bacteria colonizing the SWRO membrane represented a subportion of microbes in the source seawater; however, they were quite different from those colonizing the cartridge filter. The examination of five SWRO membranes from desalination plants located in different parts of the world showed that although the bacterial communities from the membranes were not identical to each other, some dominant bacteria were commonly observed. In contrast, bacterial communities in source seawater were significantly different based on location and season. Microbial profiles from 14 cartridge filters collected from different plants also revealed spatial trends.     

The correlation between biofilm biopolymer composition and membrane fouling in submerged membrane bioreactors

Biofouling, the combined effect of microorganism and biopolymer accumulation, significantly reduces the process efficiency of membrane bioreactors (MBRs). Here, four biofilm components, alpha-polysaccharides, beta-polysaccharides, proteins and microorganisms, were quantified in MBRs. The biomass of each component was positively correlated with the transmembrane pressure increase in MBRs. Proteins were the most abundant biopolymer in biofilms and showed the fastest rate of increase. The spatial distribution and co-localization analysis of the biofouling components indicated at least 60% of the extracellular polysaccharide (EPS) components were associated with the microbial cells when the transmembrane pressure (TMP) entered the jump phase, suggesting that the EPS components were either secreted by the biofilm cells or that the deposition of these components facilitated biofilm formation. It is suggested that biofilm formation and the accumulation of EPS are intrinsically coupled, resulting in biofouling and loss of system performance. Therefore, strategies that control biofilm formation on membranes may result in a significant improvement of MBR performance.     

Biofouling of reverse osmosis membranes: effects of cleaning on biofilm microbial communities,membrane performance,and adherence of extracellular polymeric substances

Laboratory-scale reverse osmosis (RO) flat-sheet systems were used with two parallel flow cells, one treated with cleaning agents and a control (ie undisturbed). The cleaning efforts increased the affinity of extracellular polymeric substances (EPS) to the RO membrane and altered the biofilm surface structure. Analysis of the membrane biofilm community composition revealed the dominance of Proteobacteria. However, within the phylum Proteobacteria, γ-Proteobacteria dominated the cleaned membrane biofilm, while β-Proteobacteria dominated the control biofilm. The composition of the fungal phyla was also altered by cleaning, with enhancement of Ascomycota and suppression of Basidiomycota. The results suggest that repeated cleaning cycles select for microbial groups that strongly attach to the RO membrane surface by producing rigid and adhesive EPS that hampers membrane performance.     

Characterization and effect of biofouling on polyamide reverse osmosis and nanofiltration membrane surfaces

Biofouling is a major reason for flux decline in the performance of membrane-based water and wastewater treatment plants. Initial biochemical characterization of biofilm formation potential and biofouling on two commercially available membrane surfaces from FilmTec Corporation were investigated without filtration in laboratory rotating disc reactor systems. These surfaces were polyamide aromatic thin-film reverse osmosis (RO) (BW30) and semi-aromatic nanofiltration (NF270) membranes. Membrane swatches were fixed on removable coupons and exposed to water with indigenous microorganisms supplemented with 1.5 mg l(-1) organic carbon under continuous flow. After biofilms formed, the membrane swatches were removed for analyses. Staining and epifluorescence microscopy revealed more cells on the RO than on the NF surface. Based on image analyses of 5-μm thick cryo-sections, the accumulation of hydrated biofoulants on the RO and NF surfaces exceeded 0.74 and 0.64 μm day(-1), respectively. As determined by contact angle the biofoulants increased the hydrophobicity up to 30° for RO and 4° for NF surfaces. The initial difference between virgin RO and NO hydrophobicities was ～5°, which increased up to 25° after biofoulant formation. The initial roughness of RO and NF virgin surfaces (75.3 nm and 8.2 nm, respectively) increased to 48 nm and 39 nm after fouling. A wide range of changes of the chemical element mass percentages on membrane surfaces was observed with X-ray photoelectron spectroscopy. The initial chemical signature on the NF surface was better restored after cleaning than the RO membrane. All the data suggest that the semi-aromatic NF surface was more biofilm resistant than the aromatic RO surface. The morphology of the biofilm and the location of active and dead cell zones could be related to the membrane surface properties and general biofouling accumulation was associated with changes in the surface chemistry of the membranes, suggesting the validity of the combination of these novel approaches for initial assessment of membrane performance.