The impact of spatial structure of CRISPR-phage coevolution

It is becoming increasingly clear that bacteriophage (phage) can play a key role in shaping microbial community composition and function. This project aims to study the importance of one of the most widespread immune mechanisms, known as CRISPR-Cas, for bacteria-phage interactions in natural environments. Nearly half of all bacteria encode CRISPR-Cas, and this immune system is therefore thought to be particularly important in determining the role of phage in regulating bacterial populations dynamics and evolution. In the face of bacteria that evolve CRISPR immunity, the importance of phage will strongly depend on whether or not it can coevolve to overcome immunity, which could subsequently lead to an arms race of defense and counter-defense. Although lab based studies frequently find that CRISPR resistant bacteria drive phage extinct, studies on wild bacterial populations support the idea that phage can coevolve with CRISPR in nature. We hypothesize that this discrepancy between CRISPR-phage coevolution in nature and the lab is due to the lack of ecological complexity in lab-based studies, which may be critical for coevolution to occur. By studying CRISPR-phage interactions in laboratory media and semi-natural environments we will be able to identify the factors that determine the importance and coevolutionary consequences of CRISPR-Cas immune mechanisms in nature.

Our preliminary data and existing suggest that spatial structure will be a particularly important factor in determining whether or not phage can coevolve with CRISPR, but the underlying mechanism remains unclear. In the proposed study, we will examine this by generating novel theory and performing experimental tests both in laboratory media and in sterilized soil with different levels of spatial structure. Apart from making an important contribution to our fundamental understanding of CRISPR-phage coevolution in natural environments, this research can also provide important insights into the potential role that CRISPR-phage interactions play in the ecology and evolution of soil microbiomes. Soils remain the most poorly understood ecosystems on Earth, even though it has been estimated that the biological services that soils provide are worth more than $20 trillion globally and are critical for many processes that humans depend on, including the production of food and various compounds, the purification of water, disposal of waste and cycling of nutrients.

In this study, we will use a systems biology approach where we will generate mathematical models that predict the effects of spatial structure for bacterial and phage evolution, their coexistence and coevolution. We will then test these predictions using laboratory evolution experiments, where we will tease apart the different effects of spatial structure on CRISPR-phage interactions in highly controlled environments. In the third objective, we will monitor CRISPR-phage coevolution in soil mesocosms containing different soils with different structural properties and examine CRISPR-phage coevolution across the different soils and under different mixing regimes. Altogether, the data from this research will help to understand how a simple ecological factor - spatial structure - impacts CRISPR-phage coevolution and therefore bacterial and phage population dynamics, which can have important consequences for ecosystem functioning. We will use a powerful combination of mathematics, molecular and evolutionary biology, and microbial ecology to resolve an important discrepancy between experimental and correlational studies on CRISPR-phage coevolution.

The proposed research will demonstrate the importance and consequences of CRISPR systems for bacteria-phage coevolution in environments that are spatially structured, such as soil. We identified the following beneficiaries:

(i) Postdoctoral Research Associates
Advancing science and achieving impact from it in the long term is critically dependent on training young academics. During this project two postdoctoral researchers will be trained in modelling and experimental design, data analysis, data presentation, and manuscript writing.

(ii) The wider academic community.
The proposed study has the potential to increase our understanding of fundamental areas in science and this cross-disciplinary work will be disseminated to the wider academic community through publication in peer-reviewed journals and presentation at international conferences.

(iii) Industry
Bacteria with CRISPR-mediated phage resistance are increasingly being applied in the protection of fermentations in industry and can be used to protect bacteria thriving in more "open" systems such as the rhizosphere and the gut. These applications will clearly benefit from a more detailed understanding of CRISPR-phage coevolution across different environments, including soil. During the course of this project, we will develop existing and novel partnerships with UK companies aiming to establish CASE partnerships to develop the results from this project into direct societal and commercial benefits. Specifically, we will be working with Ed Fuchs of Folium Science, and establish contacts with fertilizer manufacturer and seed companies based in the UK, such as Symbio and Thompson & Morgan.

(iv) Soil stakeholders
The interaction between soil microbes and phage can have important long-term consequences for soil respiration and soil fertility, which has direct relevance to individuals and organizations linked to agriculture, such as farmers, government agencies, policy makers and NGOs such as Natural England, Defra. We will communicate our findings with these groups by inviting them to a London-based mini-conference that we plan to organise in the autumn of 2018, and by using media that are not restricted to members of the scientific community, such as Microbiology Today and newspaper articles.

(v) Clinicians
The model organism that we use in this study, Pseusomonas aeruginosa, is an important opportunistic human pathogen that frequently colonizes the lungs of cystic fibrosis patients and wounds of burn patients. Given widespread antibiotics resistance, phage therapy is increasingly considered as an alternative strategy to treat P. aeruginosa infections. We have existing links with clinicians based in various European countries and will share our results in order to allow development of strategies that enhance the therapeutic potential of phage therapy when the pathogen encodes a CRISPR system.

(vi) General public
It is important for the advancement of science that young people become interested in science and that the general public is informed of scientific progress. To this end, we will use social media and participate in public engagement activities. The University has excellent links with schools, and as part of this project we will welcome work experience students from local schools. To inform the general public of scientific progress, we will tailor findings from our research for coverage by the popular press. The PI and co-Is have completed public engagement courses and always been active in communicating research. The University has excellent support for public engagement, having previously been awarded RCUK Catalyst Funding and with ongoing support through our Impact and Engaged Research Network.