Microbially induced calcium carbonate precipitation through CO2 sequestration via an engineered Bacillus subtilis.

BACKGROUND: Microbially induced calcium carbonate precipitation has been extensively researched for geoengineering applications as well as diverse uses within the built environment. Bacteria play a crucial role in producing calcium carbonate minerals, via enzymes including carbonic anhydrase-an enzyme with the capability to hydrolyse CO2, commonly employed in carbon capture systems. This study describes previously uncharacterised carbonic anhydrase enzyme sequences capable of sequestering CO2 and subsequentially generating CaCO3 biominerals and suggests a route to produce carbon negative cementitious materials for the construction industry. RESULTS: Here, Bacillus subtilis was engineered to recombinantly express previously uncharacterised carbonic anhydrase enzymes from Bacillus megaterium and used as a whole cell catalyst allowing this novel bacterium to sequester CO2 and convert it to calcium carbonate. A significant decrease in CO2 was observed from 3800 PPM to 820 PPM upon induction of carbonic anhydrase and minerals recovered from these experiments were identified as calcite and vaterite using X-ray diffraction. Further experiments mixed the use of this enzyme (as a cell free extract) with Sporosarcina pasteurii to increase mineral production whilst maintaining a comparable level of CO2 sequestration. CONCLUSION: Recombinantly produced carbonic anhydrase successfully sequestered CO2 and converted it into calcium carbonate minerals using an engineered microbial system. Through this approach, a process to manufacture cementitious materials with carbon sequestration ability could be developed.

Biofilm inspired fabrication of functional bacterial cellulose through ex-situ and in-situ approaches.

Bacterial cellulose (BC) has been explored for use in a range of applications including tissue engineering and textiles. BC can be produced from waste streams, but sustainable approaches are needed for functionalisation. To this end, BslA, a B. subtilis biofilm protein was produced recombinantly with and without a cellulose binding module (CBM) and the cell free extract was used to treat BC either ex-situ, through drip coating or in-situ, by incorporating during fermentation. The results showed that ex-situ modified BC increased the hydrophobicity and water contact angle reached 120°. In-situ experiments led to a BC film morphological change and mechanical testing demonstrated that addition of BslA with CBM resulted in a stronger, more elastic material. This study presents a nature inspired approach to functionalise BC using a biofilm hydrophobin, and we demonstrate that recombinant proteins could be effective and sustainable molecules for functionalisation of BC materials.

Innovating fire safety with recombinant hydrophobic proteins for textile fire retardancy.

Fire retardancy for textiles is important to prevent the rapid spread of fire and minimize damage to property and harm to human life. To infer fire-resistance on textile materials such as cotton or nylon, chemical coatings are often used. These chemicals are usually toxic, and economically and environmentally unsustainable, however, some naturally produced protein-based fire retardants could be an alternative. A biofilm protein from Bacillus subtilis (BslA) was identified and recombinantly expressed in Escherichia coli with a double cellulose binding domain. It was then applied to a range of natural and synthetic fabric materials. A flame retardancy test found that use of BslA reduced fire damage by up to 51% and would pass fire retardancy testing according to British standards. It is therefore a viable and sustainable alternative to current industrial fire-retardant coatings.

Survival and activity of an indigenous iron-reducing microbial community from MX80 bentonite in high temperature / low water environments with relevance to a proposed method of nuclear waste disposal.

MX80 bentonite clay has been selected as the buffer and backfill in a proposed method for long-term deep geological storage of nuclear waste. Extensive studies have been carried out on the geomechanical properties of the clay; however, it is not clear what effect microbes, specifically iron-reducing bacteria, will have on its ability to function as an affective barrier. Iron-reducing bacteria can reduce structural or external Fe(III) to Fe(II) and have been previously identified in the indigenous microbial community of MX80 bentonite. Experiments to assess bacterial survival at the high temperature and low water conditions likely to exist in the repository were carried out at different temperatures with the addition of steel to represent a nuclear waste canister. The resulting microbial enrichments were analysed, and mineralogical and geomechnical analysis was carried out on the clay. Microbial sequencing revealed that iron-reducing bacteria, and other indigenous species can survive these conditions in MX80 bentonite in either an active or dormant state. Microbial influenced mineralogical changes may lead to a loss of silica from the clay and reduction of Fe(III) to Fe(II). These changes could alter the ability of the clay to act as an effective barrier in nuclear waste disposal. Furthermore, evidence of reduced steel corrosion when microbes were present suggested that microbial activity may lead to either a protective coating on the steel or depletion of oxygen to slow corrosion. The production of such a layer would benefit nuclear waste disposal by inhibiting corrosion of a metal waste canister.