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Bioethanol production from lignocellulosic biomass is seen as an alternative source of energy. However, large-scale production of bioethanol from lignocellulosic biomass is still not feasible due to the high cost of cellulase and lack of cellulases with a high specific activity that can act on crystalline cellulose. The study aimed at screening for microbial cellulolytic enzymes using a proteomic approach. The objectives were to screen for microbial cellulases with a high specific activity and separate the cellulolytic enzymes using a combination of zymography and two dimensional (2-D) gel electrophoresis followed by tryptic digestion, matrix assisted laser desorption ionisation-time of flight (MALDI-TOF) and bioinformatics analysis. Fungal and bacterial isolates were cultured in M9 minimal and Mandel media for a period of 168 hours at 60°C and 30°C with cellobiose and Avicel as carbon sources. Microbial cells were separated from the supernatants through centrifugation and the crude enzymes from the cultures were used for the determination of cellulase activity, zymography, SDS-PAGE and two-dimensional gel electrophoresis. Five isolates, with lytic action on carbon sources studied were a bacterial strain, (BARK) and fungal strains (VCFF1, VCFF14, VCFF17 and VCFF18). Peak cellulase production by the isolates was found to be 3.8U/ml, 2.09U/ml, 3.38U/ml, 3.18U/ml and 1.95U/ml, respectively. Beta-glucosidase zymography resulted in a dark brown band and clear zones against a dark background for endoglucanase. Affinity precipitation of the VCFF17 isolate’s crude enzyme resulted in seven glycoside hydrolases with a carbohydrate binding module (CBM). The presence of the CBM in the glycoside hydrolases produced by the VCFF17 confer the isolate’s potential to be used in the hydrolysis of plant biomass for bioethanol production. Two-dimensional gel protein maps resulted in the separation and quantitative expression of different proteins by the microbial isolates. MALDI-TOF ana, National Research Foundation (NRF)

Water is crucial to all life forms on earth. Still, millions of people are suffering because of lack of fresh water. One of the most important reasons is industrial pollution. That is why more effective and economical water treatment systems must be developed and studied worldwide. In this study, a new design of continuous electrocoagulation systems is introduced and compared with widely studied batch process systems. This novel design improves the controllability of water flow and electricity. In the treatment of paint industry wash water, a batch system and a novel continuous electrocoagulation system with an aluminum (Al) anode and an iron (Fe) cathode were used. Two parallel analyses of both processes were conducted, and the average efficiency of chemical oxygen demand and Al removal were 68% and 79.8% in batch and 69.7% and 62.1% in the continuous system, respectively. The number of microbes decreased by 99.6% in the batch and by 99.8% in the continuous system. The calculated operational costs in this experiment were 1.63 €/m³ for the batch and 1.19 €/m³ for the continuous system. The novel continuous electrocoagulation process was as efficient as the batch process in this study.

Waterborne microbial pathogen detection via nucleic acid analysis on portable microfluidic devices is a growing area of research, development, and application. Traditional polymerase chain reaction (PCR)-based nucleic acid analysis detects total extracted DNA, but cannot differentiate live and dead cells. A propidium monoazide (PMA) pretreatment step before PCR can effectively exclude DNA from nonviable cells, as PMA can selectively diffuse through compromised cell membranes and intercalate with DNA to form DNA-PMA complex upon light exposure. The complex strongly inhibits the amplification of the bound DNA in PCR, and thus, only cells with intact cell membranes are detected. Herein, this study reports the development of a microfluidic device to carry out PMA pretreatment 'on-chip'. Chip design was guided by computer simulations, and prototypes were fabricated using a high-resolution 3D printer. The optimized design utilizes split and recombine mixers for initial PMA-sample mixing and a serpentine flow channel containing herringbone structures for dark and light incubation. On-chip PMA pretreatment to differentiate live and dead bacterial cells in buffer and natural pond water samples was successfully demonstrated.

There is an urgent need to develop sustainable solutions to convert solar energy into energy carriers used in the society. In addition to solar cells generating electricity, there are several options to generate solar fuels. This paper outlines and discusses the design and engineering of photosynthetic microbial systems for the generation of renewable solar fuels, with a focus on cyanobacteria. Cyanobacteria are prokaryotic microorganisms with the same type of photosynthesis as higher plants. Native and engineered cyanobacteria have been used by us and others as model systems to examine, demonstrate, and develop photobiological H-2 production. More recently, the production of carbon-containing solar fuels like ethanol, butanol, and isoprene have been demonstrated. We are using a synthetic biology approach to develop efficient photosynthetic microbial cell factories for direct generation of biofuels from solar energy. Present progress and advances in the design, engineering, and construction of such cyanobacterial cells for the generation of a portfolio of solar fuels, e. g., hydrogen, alcohols, and isoprene, are presented and discussed. Possibilities and challenges when introducing and using synthetic biology are highlighted.

In recent years, the physical properties and interaction forces of microbial cell surfaces have been extensively studied using atomic force microscopy (AFM). A variety of AFM force spectroscopy approaches have been developed for investigating native cell surfaces with piconewton (nanonewton) sensitivity and nanometer lateral resolution, providing novel information on the nanomechanical properties of cell walls, on surface forces such as van der Waals and electrostatic forces, solvation and steric/bridging forces, and on the forces and localization of molecular recognition events. The intention of this article is to survey these different applications and to discuss related methodologies (how to prepare tips and samples, how to record and interpret force curves). (C) 2006 Elsevier B.V. All rights reserved.

Atomic force microscopy (AFM) was used to characterize, under water, the surface morphology and molecular interactions of two types of microbial cells: fungal spores (Phanerochaete chrysosporium) and bacteria (Lactococcus lactis), High-resolution deflection images showed that the spore surface was uniformly covered with patterns of rodlets that were several hundred nanometres in length and had a periodicity of similar to 10 nm, Such surface organization was not detected on the bacterial surface, which showed a sponge-like structure. Force-distance curves revealed very different molecular interactions for the two microorganisms: upon approach, no significant curvature was seen in the contact region for spores, contrary to bacteria, pointing to a difference in cell softness; and upon retraction, no adhesion forces were detected on spores but multiple unbinding events and elongation forces attributed to macromolecular bridging were observed on bacteria. These results show that AFM is a powerful tool for probing the surface properties of native microbial cells on the nanometre scale. Copyright (C) 2000 John Wiley & Sons, Ltd.

Climate change from anthropogenic hydrocarbon fuel use is causing rapid and unprecedented changes to global environmental systems. To prevent environmental collapse, it is vital to develop technologies for producing carbon- negative energy and fuels. This is the grand scientific, industrial, and economic challenge of our time and requires novel energy production and waste valorisation strategies at all levels of society, from global- scale producers to individual households. While there is no individual solution, the unique chemical <> electricity energy conversion possibilities of Microbial Electrochemical Technologies (MET) may offer innovative routes for capturing the residual energy in waste, alongside valorising spent carbon into energy rich fuels and chemicals. However, a range of process- limiting challenges results in MET applicability mostly languishing at the laboratory scale. This thesis aims to address two of these challenges, specifically: (1) MET being bound to the intrinsic physiological properties and growth rates of native electroactive microbes, and 2) the inefficiency of biological extracellular electron transport (EET) between cells and electrodes. The work presented in this thesis employed two distinct strategies to combat these challenges. First, a gene modification strategy was used to increase the electroactive potential of the industrially relevant microorganism Cupriavidus necator. Second, a ‘biohybrid’ strategy was proposed, based on the implantation of functionalised carbon nanotubes within the outer membrane of Cupriavidus necator to augment electron exchange, in addition to the native biological electron transport mechanisms. The first strategy involved structural and aromatic amino acid modifications of Cupriavidus necator’s native type- IV pili, to provide an additional electron transfer mechanism. Cupriavidus necator strains expressing modified pili were characterised using PeakForce TUNA Atomic Force Microscopy and Cyclic Voltammetr