Real-time visualisation and modelling of biofilm inhibition by lactam

In order to meet its environmental targets Unilever has a vested interest in natural and nature-derived product ingredients. Antimicrobials and anti-biofilm technologies find applications in Unilever's formulations, packaging, and processing either as performance actives or preservatives. At the core of Unilever's business, home and personal care liquid formulations (e.g. Surf, Domestos, Dove) aim at improving surface hygiene and freshness through biostatic and anti-biofilm actives. In refreshment and food categories there is a continuous ambition to improve the integrity and lifespan of products with biofilm-ready product processing and packaging technologies. Current microbial control ingredients are receiving increased safety, regulatory and NGO pressure due to their petrochemical profile.

The anti-biofilm potential of a library of over 600 chemicals (called furanone derived lactam analogues) have been assessed that are derived from Delisea pulchra, a marine algae that typically is devoid of bacterial biofilm growth. Some variants have been shown to disrupt quorum sensing (cell-cell signalling) during biofilm growth and maturation, without inducing microbial adaptation. It has been demonstrated that lactam analogues jam quorum sensing, acting as antagonists of signal molecule receptors on gram negative bacteria. From the moment free floating cells start adsorbing reversibly on surfaces, cell-cell crosstalk is initiated that controls events from the early reversible adsorption, anchorage, aggregation and colonisation of the surface to secretion of EPS, virulence and dispersal of the biofilm. To-date it is not fully understood which stage(s) of the process are affected by the lactam analogues.

In order to progress the anti-biofilm lactam analogue treatment to market, and obtain the requisite regulatory approval, it is essential that the role of biological and physical processes is clarified. This project aims to visualise in real-time phenotypic changes to non-planktonic bacteria induced by Unilever's lead lactam analogue. In particular, it will explore the full spectrum of activity of the treatment on early stage biofilm development by visualisation and modelling in 3D, exploring the very earliest stages of non-planktonic cell-surface association to lasting impact on mature biofilm structure and functionality. It will employ unique state-of-the-art holographic and imaging techniques developed by the research investigators at the University of York to measure microscopic 3D flow fields together with quorum sensing activity and, in tandem, will construct a mechanistic mathematical description of the interaction of biological and physical processes. These approaches will aim to identify the limiting factor(s) of lactam analogue biofilm inhibition and inform the design of next generation anti-biofilm technologies.

Natural and nature-derived antimicrobials and anti-biofilm technologies have application in Unilever's formulations, packaging, and processing, either as performance actives or preservatives. Today's petrochemical options are receiving safety, regulatory and NGO pressure and can be ineffective on established biofilms on inert or biological surfaces.
The anti-biofilm potential of a library of 600 chemicals derived from Delisea pulchra, a marine algae that typically is devoid of bacterial biofilm growth, has been assessed. Some disrupt quorum sensing (cell-cell signalling) during biofilm growth and maturation, without inducing microbial adaptation. These lactam analogues can jam quorum sensing, acting as antagonists of signal molecule receptors on Gram negative bacteria. From the moment free-floating cells adsorb reversibly on surfaces, cell-cell crosstalk is initiated controlling anchorage, aggregation and colonisation, involving secretion of EPS, virulence and dispersal. To date the action of lactam analogues is not fully understood.
In order to progress anti-biofilm treatments to market, and obtain the requisite regulatory approval, it is essential that biological and physical processes are clarified. This project aims to visualise real-time phenotypic changes to non-planktonic bacteria induced by Unilever's lead lactam analogue. It will explore the very earliest stages of non-planktonic cell-surface association to lasting impact on mature biofilm structure and functionality. It will employ unique state-of-the-art holographic and imaging techniques developed by the research investigators at the University of York to measure microscopic 3D flows together with quorum sensing activity and will construct a mechanistic mathematical description of the interaction of biological and physical processes. These approaches will aim to identify the limiting factor(s) of lactam analogue biofilm inhibition and inform the design of next generation anti-biofilm technologies.

The work described in this proposal may have enormous impact on the progression of a new class of naturally occurring anti-biofilm agents to market. It aims to demonstrate the feasibility of lactam analogues (Unilever UK Ltd. possesses rights to over 600 such analogues) as components of cleaning products by providing experimental evidence for efficacy and a mechanistic understanding of the process through visualisation and modelling. This project aims to recommend an optimal biofilm treatment stage. Moreover, knowledge of the feasibility of the treatment and its mode of action are absolute requirements for regulatory purposes; this study represents a critical step in the product development process, made possible by the suite of world-leading methods developed by the named investigators at the University of York.

Biofilms are ubiquitous in domestic and industrial processes. Therefore, the development and public availability of anti-biofilm technology would have global economic, social and environmental significance. Given the existing path to market through Unilever (multi-billion pound brands such as Surf, Domestos and Dove), undoubtedly this would generate and/or secure revenue and jobs on a very large scale in the UK. Novel anti-biofilm treatments could be transformative for health and wellbeing in society by the public provision of more effective cleaning products, with improved fabric and personal hygiene potentially leading to a reduction in antimicrobial resistance and healthcare-associated infections. There may be additional benefits to the environment through a decrease in washing temperature (thus contributing to reductions in CO2 emissions) and use of mild/'green' ingredients (releasing fewer harmful chemicals). Furthermore, the technology could have far-reaching impact on a huge range of other products, including biofilm-ready paints and coatings for oil, gas and marine industries, nonwovens and medical textiles, wound-care dressings, and water filtration. With feasibility established for anti-biofilm laundry liquids, further research and development will be required to obtain regulatory, safety and environmental clearances. As part of the scoping for this project, the commercial partner (Unilever) has predicted that a lactam analogue-based laundry liquid product will be viable for launch by 2021.

The proposed theory will use existing simplified approaches, where possible, to capture key mechanisms of the coupled biological-hydrodynamic system. Furthermore, the proposed experiments will take advantage of recently developed, theoretically inspired visualisation and high-throughput measurement techniques. This study will aim to demonstrate the value of combining these methods, to add value in one closely-aligned experimental-theoretical package. The IP in this approach may find application over a wide range of problem areas, beyond those of interest to Unilever, where biological and physical mechanisms are coupled, ranging from infection and immunity (e.g. parasite interactions with surfaces in the body) to biofuels (e.g. algal photobioreactors). Therefore, the work could have additional impact on heathcare and energy, but this will depend very much on the outcome of this first feasibility study.