Professor Joe Keddie - Adding functionality to coatings with non-growing metabolically-active bacteria

Presentation - pdf
Yuxiu Chen^a, Simone Krings^b, Joshua R. Booth^c, Stefan A. F. Bon^c, Suzanne Hingley-Wilson^b and Joseph L. Keddie^a
aDepartment of Physics, University of Surrey, Guildford, UK
^bDepartment of Microbial Sciences, University of Surrey, Guildford, UK
^cDepartment of Chemistry, University of Warwick, Coventry, UK

The commonly-used biomimetic strategy attempts to mimic Nature when designing the structure and properties of materials. In the design of coatings with targeting wetting properties, surface textures take inspiration from the lotus leaf or the rose petal. One could also imagine adding functionality – such as responsiveness to the environment, the remediation of pollutants, catalysis of chemical reactions, or even the creation of useful by-products – by copying Nature. As an alternative to this strategy, we envisage directly using Nature (in the form of viable cells) to add these types of functionality to coatings. Specifically, we have successfully made a biocoating, which confines non-growing, metabolically-active bacteria within a synthetic colloidal polymer (i.e. latex) film. A biocoating needs to have a high permeability to allow a high rate of mass transfer for rehydration and the transport of both nutrients and metabolic products. It therefore requires an interconnected porous structure. In this talk, I will describe how we exploited rigid tubular nanoclays (halloysite) and non-toxic latex particles (with a relatively high glass transition temperature) as the colloidal “building blocks” to tailor the porosity inside biocoatings containing Escherichia coli bacteria as a model organism. Electron microscope images revealed inefficient packing of the rigid nanotubes and proved the existence of nanovoids along the halloysite/polymer interfaces. Single-cell observations using confocal laser scanning microscopy provided evidence for metabolic activity of the E. coli within the biocoatings through the expression of yellow fluorescent protein. Whereas there was no measurable permeability in a coating made from only latex particles, the permeability coefficient of the composite biocoatings increased with increasing halloysite content up to a value of 1x10^-4 m h^-1. The effects of this increase in permeability on the cell viability was demonstrated through a specially-developed resazurin reduction assay. Bacteria encapsulated in halloysite composite biocoatings had statistically significant higher metabolic activities in comparison to bacteria encapsulated in a non-optimized coating made from latex particles alone. Enhancing bacterial viability in biocoatings has enormous potential in applications including waste-water treatment and the production of biomass and biofuel gases

Q&A

From  Justin Perry: How long does E. coli live for within these halloyside films?
Answer: Well, there have been other people that have made biocoatings with a variety of bacteria, and if you keep them rehydrated they can survive for weeks, we checked the viability essentially after a couple of days, in future work we definitely want to check but in this experiment we just looked at a few days.

From  Wai Lee : The work on graphene is really interesting. Will the size of graphene affect the packing of the film and thus the optical properties? Also, will the way to prepare the film affect too for example,  layer-by layer rather than mixing?
Answer: you don’t want big sheets, the graphene sheets we used were about 350 nm which is a bit larger than the size of the polymer colloids. I do think we nanotubes would be the same. Big sheets can disrupt the packing and we didn’t want that. In that work, the sample preparation is essential. You have to keep high order, we didn’t do layer by layer so it is hard to comment on that. But this is kind of simpler, a one step process. It is interesting that the crystals form from the top down, so we are not doing a sedimentation, just as it dries forms skin layer at the top.

From  Bob Luigjes : If I understood correctly, the film formation was done at temperatures slightly above latex Tg. Have you looked at the influence of adding coalescing agents on the bacteria?
Answer: I guess I would be a little nervous about adding coalescing agents because it could harm the bacteria. I guess it would be interesting ones that they could use as food, but the short answer is that we haven’t looked at that and just played with the temperature.

From  Craig Allan : Thanks for your talk, have you looked at other bacteria?
Answer: We have just started to use cyanobacteria, which will undergo photsynthesis. We are also looking at bacteria found in soils.

From  Jordan Petkov : More of a comment than question, but osmotic pressure measurements would have given water permeability and not that of the fluorescent molecule, i.e. you might have had the right permeability at lower level of halloyside.
Answer: Thanks. Yes, we are measuring the permeability of the fluorescein (and not water). I'll look into osmotic pressure measurements. That's a good idea.

From  Peter Collins : Have you looked at other minerals with different morphologies?
Answer: So far, we've only tried halloysite. But disk-like particles would probably similarly introduce porosity.

From  Bob Luigjes : If coalescing agents are not an option, did you look at / are you planning to look at latex particles with low Tg, to from films at ambient temperature?
Answer: There is some older work on biocoatings that have film-formed at room temperature. The key problem is that coalescence is quite good, which means that the cooatings are not permeable to water. "Soft" particles will continue to deform and coalesce slowly over time. By using a high-Tg polymer, we retained porosity at ROOM temperature over time.