Jonas Flohr from Portsmouth reports back on his AMI-sponsored summer studentship at Durham University investigating how metals influence bacterial ecosystems.
Jonas (21) is studying BSc Biological Sciences at Durham University and undertook a placement with Dr Karrera Djoko at the Department of Biosciences at Durham University.
Metal ions are essential nutrients required for bacterial growth but are toxic when in excess. Metal abundance influences bacterial physiology, bacteria-bacteria interactions, and thus the ecology of bacterial communities. Together with colleagues from Durham Mathematical Sciences, Dr Djoko sought to elucidate whether fundamental ecological principles also apply to metal-responsive bacterial communities.
Baseline behaviour
Before conducting experiments involving more complex communities, however, it was necessary to understand the baseline behaviour of individual species in response to metal availability, and this was the aim of my project.
I used Escherichia coli as an experimental system, given that its metal homeostasis has been well-studied. To represent different ‘species’, I chose 12 mutant strains from the established KEIO library, each mutant lacking a different gene involved in either iron, copper, or nickel homeostasis. The mutants thus each have distinct metal-dependent growth traits but uniform background metabolisms.
So, what did I do exactly?
1. Design primers and confirm the genotype of mutants by diagnostic PCR.
2. Establish growth of each mutant in M9-Glucose medium.
3. Determine the minimum initial population seeding density at which all mutants grow, which will be important when conducting invasion assays with communities consisting of two or more species (will be done in future work, not during my project). This concentration was used for all subsequent experiments.
4. Measure the growth of the mutants and wild-type over 24 hours, at different metal concentrations.
My project has formed the framework for future experiments involving pairwise communities (this will allow us to test for interspecific interactions, such as competition and cross-feeding), followed by more complex communities (up to 25 different species). The data generated will be used by mathematicians at Durham Mathematical Sciences to build models to test whether fundamental ecological principles also apply to metal-responsive bacterial communities.
Predicting response to change
Bacteria rarely live in isolation but instead as part of complex communities. These communities occupy a vast array of environments, including soils, the ocean, and humans. In land and water ecosystems, bacteria play key roles in nutrient and energy cycling; in humans, they assist in a diversity of processes, including digestion and immune regulation, amongst others, whilst an imbalance in community composition (known as dysbiosis) is associated with numerous disease states. The functioning of a microbial community depends on which species are present and their abundance, and this is dictated by a complex network of intra- and inter-species interactions which may permit or prevent coexistence.
Metals are important nutrients for bacteria but are toxic in excess. They thus have a profound influence upon bacterial physiology and bacterial interactions. In this way, metal availability shapes bacterial community assembly and function. By better understanding how metal availability influences these processes, we will be able to predict how bacterial communities respond to changes in metal abundance, or even engineer microbial communities to improve their function, with benefits in medicine, industry, and agriculture.
Metal response fingerprint
My project was the first step towards gaining a better understanding of how metal abundance shapes bacterial ecosystems. Despite there only being time to study the response of individual ‘species’ to metal abundance, it was hugely rewarding to see that different mutants responded differently to changes in metal abundance, with each mutant having its own metal response ‘fingerprint’. This demonstrated that our aim of using different mutants to represent ‘species’ which differ in their metal homeostasis (but share uniform background metabolisms) had been a success.
I started the project with an ambitious list of things that I wanted to achieve and results I wished to collect. It was sometimes difficult to accept that research does take time, and I learnt that it is better to conduct thorough experiments which yield valuable results rather than trying to produce as many results as possible. In other words: quality over quantity, even if the former may be more time-consuming and require a little more patience.
Trial and error
I also learnt the importance of trial and error, accepting that things might not work as planned the first time round, upon which it is necessary to troubleshoot what could be improved and then repeat things again.
I also became aware of how easy it is for cultures to become contaminated, highlighting the importance of maintaining a sterile environment.
I am very fortunate to be able to continue with my project in the same lab as part of a Level 3 Research Project Module in my final year at Durham University, allowing me to begin studying pairwise interactions.
Least favourite job in the lab?! Refilling ethanol spray bottles!
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