Aggregation Of Microbial Cells Research Articles

The Aerobic Granules Process for Wastewater Treatment: From Theory to Engineering

Aerobic granules are small, dense aggregates of microbial cells that form naturally in aerobic wastewater treatment systems. They are characterized by their spherical shape, strong structural integrity, and ability to rapidly settle. These granules are formed through a self-immobilization process where different microbial species coalesce to degrade organic and inorganic compounds in wastewater. This study summarizes the development of aerobic granulation technology in wastewater treatment and the mechanism of aerobic granules’ formation, analyzes the characteristics and the factors affecting the aerobic granules’ formation, and presents practical engineering examples of its application from pilot-scale to full-scale operation.

Use of Alteromonas sp. Ni1-LEM Supernatant as a Cleaning Agent for Reverse-Osmosis Membranes (ROMs) from a Desalination Plant in Northern Chile Affected by Biofouling.

Biofouling refers to the undesirable growth of microorganisms on water-submerged surfaces. Microfouling, the initial state of biofouling, is characterized by aggregates of microbial cells enclosed in a matrix of extracellular polymeric substances (EPSs). In seawater desalination plants, filtration systems, such as reverse-osmosis membranes (ROMs), are affected by microfouling, which decreases their efficiency in obtaining permeate water. The existing chemical and physical treatments are expensive and ineffective; therefore, controlling microfouling on ROMs is a considerable challenge. Thus, new approaches are necessary to improve the current ROM cleaning treatments. This study demonstrates the application of Alteromonas sp. Ni1-LEM supernatant as a cleaning agent for ROMs in a desalination seawater plant in northern Chile (Aguas Antofagasta S.A.), which is responsible for supplying drinking water to the city of Antofagasta. ROMs treated with Altermonas sp. Ni1-LEM supernatant exhibited statistically significant results (p < 0.05) in terms of seawater permeability (Pi), permeability recovery (PR), and the conductivity of permeated water compared with control biofouling ROMs and those treated with the chemical cleaning protocol applied by the Aguas Antofagasta S.A. desalination plant.

Predicting neurodegenerative disease using the unique gut microbiota composition of transgenic mice modeling amyloid‐β plaques and neurofibrillary tangles

AbstractBackgroundThe gut microbiota, the aggregates of microbial cells that inhabit the gastrointestinal tract, communicates bidirectionally with the brain via immune, neural, metabolic, and endocrine pathways, known as the gut‐brain axis. The gut‐brain axis is suspected to contribute to the development of Alzheimer’s disease (AD). We hypothesize that altered gut microbiota composition contributes to the development of AD pathologies and neuroinflammation via the gut‐brain axis.MethodTo characterize the gut microbiota of 3xTg‐AD mice modeling plaque deposition and hyperphosphorylated tau, fecal samples were collected fortnightly from 4 to 52 weeks of age (n=57 3xTg‐AD mice, n=31 wild‐type). The V4 region of the 16S rRNA gene was amplified and sequenced on the Illumina MiSeq. Data were analyzed using QIIME 2. Targeted reverse transcription qPCR assays were used to assess inflammation in the hippocampus and colon at 8, 24, and 52 weeks of age. Fold change was calculated using ΔΔCt.ResultOur results show altered microbial communities in 3xTg‐AD mice when compared to wild‐type [(PERMANOVA (8 weeks, p=0.001), (24 weeks, p=0.039), (52 weeks, p=0.058)]. Using q2‐longitudinal, we identified a temporal increase in Bacteroides acidifaciens and Turicibacter spp. in 3xTg‐AD mice (r‐squared = 0.658615). Using Random Forest, we successfully predicted strain in 3xTg‐AD mice 100% of the time, and in WT mice 92.85% of the time, improving accuracy over baseline assignment by 1.3 fold. Colonic expression of GFAP was increased at 24 week 3xTg‐AD mice compared to 52 week 3xTg‐AD mice (p=0.009, Mann‐Whitney). Colonic gene expression of IL‐6 was increased in 52 week 3xTg‐AD mice compared to 52 week WT mice (p=0.015, Mann‐Whitney). Hippocampal expression of GFAP was increased at 52 week 3xTg‐AD mice compared to 52 week WT (p=0.049, Mann‐Whitney). Finally, hippocampal expression of Mrc1 was elevated at 24 weeks in 3xTg‐AD mice compared to 52 weeks (p= 0.004, Mann‐Whitney).ConclusionWe have identified changes in the gut microbiota and immune response that may be predictive of the development of AD pathologies. Future shallow shotgun metagenomics sequencing will assess strain‐level features and functions of the gut microbiota in AD.

The Most Probable Curve method - A robust approach to estimate kinetic models from low plate count data resulting in reduced uncertainty

A novel method is proposed for fitting microbial inactivation models to data on liquid media: the Most Probable Curve (MPC) method. It is a multilevel model that makes a separation between the “true” microbial concentration according to the model, the “actual” concentration in the media considering chance, and the actual counts on the plate. It is based on the assumptions that stress resistance is homogeneous within a microbial population, and that there is no aggregation of microbial cells. Under these assumptions, the number of colonies in/on a plate follows a Poisson distribution with expected value depending on the proposed kinetic model, the number of dilutions and the plated volume.The novel method is compared against (non)linear regression based on a normal likelihood distribution (traditional method), Poisson regression and gamma-Poisson regression using data on the inactivation of Listeria monocytogenes. The conclusion is that the traditional method has limitations when the data includes plates with low (or zero) cell counts, which can be mitigated using more complex (discrete) likelihoods. However, Poisson regression uses an unrealistic likelihood function, making it unsuitable for survivor curves with several log-reductions. Gamma-Poisson regression uses a more realistic likelihood function, even though it is based mostly on empirical hypotheses. We conclude that the MPC method can be used reliably, especially when the data includes plates with low or zero counts. Furthermore, it generates a more realistic description of uncertainty, integrating the contribution of the plating error and reducing the uncertainty of the primary model parameters. Consequently, although it increases modelling complexity, the MPC method can be of great interest in predictive microbiology, especially in studies focused on variability analysis.

Recovery of Iron-Dependent Autotrophic Denitrification Activity from Cell-Iron Mineral Aggregation-Induced Reversible Inhibition by Low-Intensity Ultrasonication.

Iron-dependent autotrophic denitrification (IDAD) has garnered increasing interests as an efficient method for removing nitrogen from wastewater with a low carbon to nitrogen ratio. However, an inevitable deterioration of IDAD performance casts a shadow over its further development. In this work, the hidden cause for such a deterioration is uncovered, and a viable solution to this problem is provided. Batch test results reveal that the aggregation of microbial cells and iron-bearing minerals induced a cumulative and reversible inhibition on the activity of IDAD sludge. Extracellular polymeric substances were found to play a glue-like role in the cell-iron mineral aggregates, where microbial cells were caged, and their metabolisms were suppressed. Adopting low-intensity ultrasound treatment efficiently restored the IDAD activity by disintegrating such aggregates rather than stimulating the microbial metabolism. Moreover, the ultrasonication-assisted IDAD bioreactor exhibited an advantageous nitrogen removal efficiency (with a maximum enhancement of 72.3%) and operational stability compared to the control one, demonstrating a feasible strategy to achieve long-term stability of the IDAD process. Overall, this work provides a better understanding about the mechanism for the performance deterioration and a simple approach to maintain the stability of IDAD.

Imaging Flow Cytometry to Study Biofilm-Associated Microbial Aggregates.

The aim of the research was to design an advanced analytical tool for the precise characterization of microbial aggregates from biofilms formed on food-processing surfaces. The approach combined imaging flow cytometry with a machine learning-based interpretation protocol. Biofilm samples were collected from three diagnostic points of the food-processing lines at two independent time points. The samples were investigated for the complexity of microbial aggregates and cellular metabolic activity. Thus, aggregates and singlets of biofilm-associated microbes were simultaneously examined for the percentages of active, mid-active, and nonactive (dead) cells to evaluate the physiology of the microbial cells forming the biofilm structures. The tested diagnostic points demonstrated significant differences in the complexity of microbial aggregates. The significant percentages of the bacterial aggregates were associated with the dominance of active microbial cells, e.g., 75.3% revealed for a mushroom crate. This confirmed the protective role of cellular aggregates for the survival of active microbial cells. Moreover, the approach enabled discriminating small and large aggregates of microbial cells. The developed tool provided more detailed characteristics of bacterial aggregates within a biofilm structure combined with high-throughput screening potential. The designed methodology showed the prospect of facilitating the detection of invasive biofilm forms in the food industry environment.

New Advances in Aerobic Granular Sludge Technology Using Continuous Flow Reactors: Engineering and Microbiological Aspects

Aerobic granular sludge (AGS) comprises an aggregation of microbial cells in a tridimensional matrix, which is able to remove carbon, nitrogen and phosphorous as well as other pollutants in a single bioreactor under the same operational conditions. During the past decades, the feasibility of implementing AGS in wastewater treatment plants (WWTPs) for treating sewage using fundamentally sequential batch reactors (SBRs) has been studied. However, granular sludge technology using SBRs has several disadvantages. For instance, it can present certain drawbacks for the treatment of high flow rates; furthermore, the quantity of retained biomass is limited by volume exchange. Therefore, the development of continuous flow reactors (CFRs) has come to be regarded as a more competitive option. This is why numerous investigations have been undertaken in recent years in search of different designs of CFR systems that would enable the effective treatment of urban and industrial wastewater, keeping the stability of granular biomass. However, despite these efforts, satisfactory results have yet to be achieved. Consequently, it remains necessary to carry out new technical approaches that would provide more effective and efficient AGS-CFR systems. In particular, it is imperative to develop continuous flow granular systems that can both retain granular biomass and efficiently treat wastewater, obviously with low construction, maintenance and exploitation cost. In this review, we collect the most recent information on different technological approaches aimed at establishing AGS-CFR systems, making possible their upscaling to real plant conditions. We discuss the advantages and disadvantages of these proposals and suggest future trends in the application of aerobic granular systems. Accordingly, we analyze the most significant technical and biological implications of this innovative technology.

Metatranscriptome Analysis of Bisphenol A-Exposed Aerobic Granular Sludge

Understanding of the metabolic pathways connected with a removal of micropollutant bisphenol A (BPA) may help to better design effective wastewater treatment processes. The aim of this study was to determine changes in gene expression in an aerobic granular sludge (AGS) community exposed to BPA. In the study, AGS adapted to BPA degradation was used. In this sludge, BPA was dosed; as a control sample, granules without BPA addition were used. mRNA was isolated from both samples and sequenced using the Illumina platform. Metatranscriptome analysis of AGS exposed to BPA indicated direct biodegradation as the main mechanism of BPA removal from wastewater. High expression of genes coding pilus and flagellin proteins in the BPA-exposed biomass indicated that exposition to BPA stimulated aggregation of microbial cells and formation of biofilm. In the BPA-exposed biomass, nitrogen was mainly used as an energy source, as indicated by the presence of genes coding nitrification enzymes and urease. Moreover, exposition to BPA stimulated expression of genes coding proteins responsible for xenobiotic degradation, including enzymes responsible for benzoate degradation. These results increase knowledge about BPA metabolism in complex microbial communities in wastewater treatment systems and indicate that AGS is suitable for efficiently removing BPA from wastewater.

Proteomic Studies of the Biofilm Matrix including Outer Membrane Vesicles of Burkholderia multivorans C1576, a Strain of Clinical Importance for Cystic Fibrosis.

Biofilms are aggregates of microbial cells encased in a highly hydrated matrix made up of self-produced extracellular polymeric substances (EPS) which consist of polysaccharides, proteins, nucleic acids, and lipids. While biofilm matrix polysaccharides are unraveled, there is still poor knowledge about the identity and function of matrix-associated proteins. With this work, we performed a comprehensive proteomic approach to disclose the identity of proteins associated with the matrix of biofilm-growing Burkholderia multivorans C1576 reference strain, a cystic fibrosis clinical isolate. Transmission electron microscopy showed that B. multivorans C1576 also releases outer membrane vesicles (OMVs) in the biofilm matrix, as already demonstrated for other Gram-negative species. The proteomic analysis revealed that cytoplasmic and membrane-bound proteins are widely represented in the matrix, while OMVs are highly enriched in outer membrane proteins and siderophores. Our data suggest that cell lysis and OMVs production are the most important sources of proteins for the B. multivorans C1576 biofilm matrix. Of note, some of the identified proteins are lytic enzymes, siderophores, and proteins involved in reactive oxygen species (ROS) scavenging. These proteins might help B. multivorans C1576 in host tissue invasion and defense towards immune system assaults.

Multispecies Interactions in Biofilms and Implications to Safety of Drinking Water Distribution System

In the aquatic environment, microorganisms are predominantly organized as biofilms. Biofilms are formed by the aggregation of microbial cells and are surrounded by a matrix of extracellular polymeric substances (EPS) secreted by the microbial cells. Biofilms are attached to various surfaces, such as the living tissues, indwelling medical devices, and piping of the industrial potable water system. Biofilms formed from a single species has been extensively studied. However, there is an increased research focus on multispecies biofilms in recent years. It is important to assess the microbial mechanisms underlying the regulation of multispecies biofilm formation to determine the drinking water microbial composition. These mechanisms contribute to the predominance of the best-adapted species in an aquatic environment. This review focuses on the interactions in the multispecies biofilms, such as coaggregation, co-metabolism, cross-species protection, jamming of quorum sensing, lateral gene transfer, synergism, and antagonism. Further, this review explores the dynamics and the factors favoring biofilm formation and pathogen transmission within the drinking water distribution systems. The understanding of the physiology and biodiversity of microbial species in the biofilm may aid in the development of novel biofilm control and drinking water disinfection processes.

Biofilm- An Alarming Niche in Dairy Industry

Biofilm is an aggregation of microbial cells interconnected by extracellular polymeric substances which accelerate growth on different material surfaces adversely affect the dairy industry. This polymicrobial community contains altered phenotype which differs them from planktonic microbes physiologically. It affects the quality and safety of raw materials and their products. Thus, producing serious problems directly affecting human health. Colonisation at the surfaces of open and closed piping systems, floors, waste, walls, ceilings of the production halls play a major role as barrier in the selection of effective sanitation agents for their control. As a result, the sector is forced to face a havoc economical loss. In fact, the condition is aggravating day-by-day. As microbes growing in a biofilm are highly resistant to antimicrobial agents and host’s immune system, it is obligatory to employ effective methods to retard the biofilm formation. Therefore, immediate appropriate precautionary measures should be adopted to combat the condition and prevent further complications.

The Bacterial Extracellular Matrix Protein TapA Is a Two-Domain Partially Disordered Protein.

Biofilms are aggregates of microbial cells that form on surfaces and at interfaces, and are encased in an extracellular matrix. In biofilms made by the soil bacterium Bacillus subtilis, the protein TapA mediates the assembly of the functional amyloid protein TasA into extracellular fibers, and it anchors these fibers to the cell surface. We used circular dichroism and NMR spectroscopy to show that, unlike the structured TasA, TapA is disordered. In addition, TapA is composed of two weakly interacting domains: a disordered C-terminal domain and a more structured N-terminal domain. These two domains also exhibited different structural changes in response to changes in external conditions, such as increased temperatures and the presence of lipid vesicles. Although the two TapA domains weakly interacted in solution, their cooperative interaction with lipid vesicles prevented disruption of the vesicles. These findings therefore suggest that the two-domain composition of TapA is important in its interaction with single or multiple partners in the extracellular matrix in biofilms.

Rapid Formation of Microbe-Oil Aggregates and Changes in Community Composition in Coastal Surface Water Following Exposure to Oil and the Dispersant Corexit.

During the Deepwater Horizon (DWH) oil spill, massive quantities of oil were deposited on the seafloor via a large-scale marine oil-snow sedimentation and flocculent accumulation (MOSSFA) event. The role of chemical dispersants (e.g., Corexit) applied during the DWH oil spill clean-up in helping or hindering the formation of this MOSSFA event are not well-understood. Here, we present the first experiment related to the DWH oil spill to specifically investigate the relationship between microbial community structure, oil and Corexit®, and marine oil-snow in coastal surface waters. We observed the formation of micron-scale aggregates of microbial cells around droplets of oil and dispersant and found that their rate of formation was directly related to the concentration of oil within the water column. These micro-aggregates are potentially important precursors to the formation of larger marine oil-snow particles. Therefore, our observation that Corexit® significantly enhanced their formation suggests dispersant application may play a role in the development of MOSSFA events. We also observed that microbial communities in marine surface waters respond to oil and oil plus Corexit® differently and much more rapidly than previously measured, with major shifts in community composition occurring within only a few hours of experiment initiation. In the oil-amended treatments without Corexit®, this manifested as an increase in community diversity due to the outgrowth of several putative aliphatic- and aromatic-hydrocarbon degrading genera, including phytoplankton-associated taxa. In contrast, microbial community diversity was reduced in mesocosms containing chemically dispersed oil. Importantly, different consortia of hydrocarbon degrading bacteria responded to oil and chemically dispersed oil, indicating that functional redundancy in the pre-spill community likely results in hydrocarbon consumption in both undispersed and dispersed oils, but by different bacterial taxa. Taken together, these data improve our understanding of how dispersants influence the degradation and transport of oil in marine surface waters following an oil spill and provide valuable insight into the early response of complex microbial communities to oil exposure.

Binding of heavy metal ions in aggregates of microbial cells, EPS and biogenic iron minerals measured in-situ using metal- and glycoconjugates-specific fluorophores

Aggregates consisting of bacterial cells, extracellular polymeric substances (EPS) and Fe(III) minerals formed by Fe(II)-oxidizing bacteria are common at bulk or microscale chemical interfaces where Fe cycling occurs. The high sorption capacity and binding capacity of cells, EPS, and minerals controls the mobility and fate of heavy metals. However, it remains unclear to which of these component(s) the metals will bind in complex aggregates. To clarify this question, the present study focuses on 3D mapping of heavy metals sorbed to cells, glycoconjugates that comprise the majority of EPS constituents, and Fe(III) mineral aggregates formed by the phototrophic Fe(II)-oxidizing bacteria Rhodobacter ferrooxidans SW2 using confocal laser scanning microscopy (CLSM) in combination with metal- and glycoconjugates-specific fluorophores. The present study evaluated the influence of glycoconjugates, microbial cell surfaces, and (biogenic) Fe(III) minerals, and the availability of ferrous and ferric iron on heavy metal sorption. Analyses in this study provide detailed knowledge on the spatial distribution of metal ions in the aggregates at the sub-μm scale, which is essential to understand the underlying mechanisms of microbe–mineral–metal interactions. The heavy metals (Au3+, Cd2+, Cr3+, CrO42−, Cu2+, Hg2+, Ni2+, Pd2+, tributyltin (TBT) and Zn2+) were found mainly sorbed to cell surfaces, present within the glycoconjugates matrix, and bound to the mineral surfaces, but not incorporated into the biogenic Fe(III) minerals. Statistical analysis revealed that all ten heavy metals tested showed relatively similar sorption behavior that was affected by the presence of sorbed ferrous and ferric iron. Results in this study showed that in addition to the mineral surfaces, both bacterial cell surfaces and the glycoconjugates provided most of sorption sites for heavy metals. Simultaneously, ferrous and ferric iron ions competed with the heavy metals for sorption sites on the organic compounds. In summary, the information obtained by the present approach using a microbial model system provides important information to better understand the interactions between heavy metals and biofilms, and microbially formed Fe(III) minerals and heavy metals in complex natural environments.

Towards understanding the role of extracellular polymeric substances in cyanobacterial Microcystis aggregation and mucilaginous bloom formation

The development of mucilaginous cyanobacterial Microcystis blooms is a serious environmental and ecological problem, and information on the bloom-formation mechanism has been lacking until now. The aggregation of microbial cells was attributed to the matrix of extracellular polymeric substances (EPS). In this study, the quantitative role of EPS matrix in Microcystis aggregation and mucilaginous bloom formation was investigated. The results showed that when EPS matrix was extracted, the aggregation abilities decreased by 27.6% and 57.4% for the lab-cultured Microcystis suspension and the field-sampled Microcystis aggregates, respectively. The extended DLVO theory revealed that EPS extraction increased the energy barrier and the values of the second energy minimum, which accounted for the deteriorated aggregation. Further analysis showed an increasing attraction energy of EPS matrix during the Microcystis bloom development, whereas the predominant contribution originated from tightly bound EPS (TB-EPS) and loosely bound EPS (LB-EPS) for the lab-cultured and field-sampled Microcystis samples. The heterogeneous energy contribution of EPS subfractions was found to be associated with the variations in organic contents. Specifically, Microcystis aggregates exhibited a higher organic content of TB-EPS than of LB-EPS compared with the lab-cultured Microcystis suspension, whereas organic content in only the LB-EPS fraction for the bloom samples was significantly higher (p<0.01) than that of the Microcystis aggregates. Based on these results, a conceptual model of EPS function was proposed in which TB-EPS plays an important role in the formation of Microcystis aggregates, after which LB-EPS contributed to the subsequent development from Microcystis aggregates to mucilaginous bloom formation.

Aggregation of yeast cells: direct measurement of discrete lectin-carbohydrate interactions.

Aggregation of microbial cells mediated by specific interactions plays a pivotal role in the natural environment, in medicine and in biotechnological processes. Here we used atomic force microscopy (AFM) to measure individual lectin-carbohydrate interactions involved in the flocculation of yeast cells, an aggregation event of crucial importance in fermentation technology. AFM probes functionalized with oligoglucose carbohydrates were used to record force-distance curves on living yeast cells at a rate of 0.5 micro m s(-1). Flocculating cells showed adhesion forces of 121+/-53 pN, reflecting the specific interaction between individual cell-surface lectins and glucose residues. Similar adhesion forces, 117+/-41 pN, were measured using probes functionalized with the lectin concanavalin A and attributed to specific binding to cell-surface mannose residues. By contrast, specific interaction forces were not observed in non-flocculating conditions, i.e. in the presence of mannose or when using non-flocculating cells, pointing to their involvement in yeast flocculation. The single molecule force spectroscopy measurements presented here provide a means to study a variety of cellular interactions at the molecular level, such as the adhesion of bacteria to animal and plant tissues.

Evolving perspectives of biofilm structure

The application of confocal scanning laser microscopy (CSLM) to biofilm research has provided detailed new information on the structural complexity of fully hydrated living biofilms. Biofilms studied by CSLM are widely reported to be heterogeneous and consist of micro‐colonies or cell clusters (aggregates of microbial cells in an extracellular polysaccharide matrix) separated by interstitial voids and channels. CSLM combined with molecular probing can identify the distribution of individual species within biofilms, while CSLM combined with microelectrodes and particle tracking has demonstrated the significance of structural heterogeneity on mass transfer in biofilms. In this review the contribution that CSLM has made towards the development of conceptual and mathematical biofilm models over the last decade is discussed. The significance of biofilm structural heterogeneity to understanding of mass transfer processes in biofilm systems and the fundamental significance of hydrodynamics in shaping biofilm structure are also discussed. Finally, some of the recent findings which suggest that biofilm structure may be controlled in part by cell signalling, quorum sensing mechanisms, are briefly discussed.