Towards understanding the role of amyloid adhesins in activated sludge bioflocculation

status: Published onlin

Bioaugmentation of a structural extracellular polymeric substance (EPS) producer to improve activated sludge bioflocculation : lessons learned

Structural extracellular polymeric substances (EPSs) contribute to the bioflocculation performance of activated sludge systems. This research investigates the potential of bioaugmentation of a structural EPS producer, Azoarcus communis, as a bioflocculation improvement or remediation approach. An antibiotic-resistant and fluorescent protein-producing mutant was constructed to monitor the survival, persistence, and location of the augmented strain in the membrane bioreactor. Preliminary batch tests against a kaolin clay model system and deflocculated sludge revealed the flocculation potential of this strain. Morphological image analysis and fluorescence microscopy suggest that most of the bacteria augmented in suspension were initially attached to the sludge flocs with, however, only a limited fraction getting incorporated within the activated sludge floc biomass. This limited bioaugmentation prevented assessing its impact on bioflocculation and might be explained by metazoan and protozoan grazing, together with competition with indigenous organisms and sub-optimal growth conditions in the reactor for the engineered strain

Full-electric microfluidic platform to capture, analyze and selectively release single cells

Current single-cell technologies require large and expensive equipment, limiting their use to specialized labs. In this paper, we present for the first time a microfluidic device which demonstrates a combined method for full-electric cell capturing, analyzing, and selectively releasing with single-cell resolution. All functionalities are experimentally demonstrated on Saccharomyces cerevisiae. Our microfluidic platform consists of traps centered around a pair of individually accessible coplanar electrodes, positioned under a microfluidic channel. Using this device, we validate our novel Two-Voltage method for trapping single cells by positive dielectrophoresis (pDEP). Cells are attracted to the trap when a high voltage (VH) is applied. A low voltage (VL) holds the already trapped cell in place without attracting additional cells, allowing full control over the number of trapped cells. After trapping, the cells are analyzed by broadband electrochemical impedance spectroscopy. These measurements allow the detection of single cells and the extraction of cell parameters. Additionally, these measurements show a strong correlation between average phase change and cell size, enabling the use of our system for size measurements in biological applications. Finally, our device allows selectively releasing trapped cells by turning off the pDEP signal in their trap. The experimental results show the techniques potential as a full-electric single-cell analysis tool with potential for miniaturization and automation which opens new avenues towards small-scale, high throughput single-cell analysis and sorting lab-on-CMOS devices. Single-cell capture and analysis with full-electric microfluidic device

Stirred culture of cartilaginous microtissues promotes chondrogenic hypertrophy through exposure to intermittent shear stress

peer reviewedCartilage microtissues are promising tissue modules for bottom up biofabrication of implants leading to bone defect regeneration. Hitherto, most of the protocols for the development of these cartilaginous microtissues have been carried out in static setups, however, for achieving higher scales, dynamic process needs to be investigated. In the present study, we explored the impact of suspension culture on the cartilage microtissues in a novel stirred microbioreactor system. To study the effect of the process shear stress, experiments with three different impeller velocities were carried out. Moreover, we used mathematical modeling to estimate the magnitude of shear stress on the individual microtissues during dynamic culture. Identification of appropriate mixing intensity allowed dynamic bioreactor culture of the microtissues for up to 14 days maintaining microtissue suspension. Dynamic culture did not affect microtissue viability, although lower proliferation was observed as opposed to the statically cultured ones. However, when assessing cell differentiation, gene expression values showed significant upregulation of both Indian Hedgehog (IHH) and collagen type X (COLX), well known markers of chondrogenic hypertrophy, for the dynamically cultured microtissues. Exometabolomics analysis revealed similarly distinct metabolic profiles between static and dynamic conditions. Dynamic cultured microtissues showed a higher glycolytic profile compared with the statically cultured ones while several amino acids such as proline and aspartate exhibited significant differences. Furthermore, in vivo implantations proved that microtissues cultured in dynamic conditions are functional and able to undergo endochondral ossification. Our work demonstrated a suspension differentiation process for the production of cartilaginous microtissues, revealing that shear stress resulted to an acceleration of differentiation towards hypertrophic cartilage