BEACON Researchers at Work: Nutrition limitation's role in phytoplankton evolution

This week’s BEACON Researchers at Work blog post is by MSU graduate student Megan Larsen.

Photo of Megan LarsenWe all know that the world is more complex than the simplified systems and questions we use in laboratories. And yet, as scientists we have to ask these simplified questions in order to understand the more complex interactions in the natural world.

One of the qualities that most drew me to science, and particularly microbiology, was being able to strip a complex system down to its core, and then reassemble that puzzle with newly acquired pieces to provide a clearer view of the system. I started my scientific career in biochemistry at Nebraska Wesleyan University, learning the chemical structures of compounds involved in cellular processes such photosynthesis and metabolism.  While learning about these processes, I was also learning broad ecological concepts and theories about resource competition and predator-prey dynamics. Now, as a graduate student in Jay Lennon’s lab at the Kellogg Biological Station, my research centers on synthesizing each of the parts I learned as an undergraduate into a population and community context involving aquatic microbes. 

Within a given environment, species interactions result in complex interactions, often fueled by the local environmental nutrient conditions. Nutrient composition is one of the most important regulatory factors governing population and community assembly in nature. In many cases, microbial organisms mirror environmental conditions internally, thus the cellular stoichiometry or nutrient composition within the cell reflects that of the environment and strongly influences growth and reproduction. Furthermore, bacterial populations are constrained by phage predation (phage are basically viruses that infect bacteria). Unlike their bacterial hosts, however, infecting phage require a specific nutrient composition to reproduce. Therefore, nutrient limiting conditions also, albeit indirectly, reduce phage reproduction.

I am interested in a series of different questions that range from population ecology to molecular and genetic mechanisms of resistance in bacteria. The central focus of my dissertation rests on understanding how stoichiometry (the balance between the organisms and their resources in the environment) influences the ecological and evolutionary feedbacks in species interactions, specifically between phytoplankton, known as cyanobacteria, and their viruses. In order to address these questions, I have setup a model system with the planktonic bacteria known as Synechococcus and a T4-like phage. Synechococcus, a unicellular cyanobacteria ranging in size between 0.5 – 2 µm, can be found throughout the world’s aquatic ecosystems and plays an integral role in net primary productivity and global nutrient cycling. In order to study ecological and evolutionary feedbacks, I decided to work with these ecologically important organisms in long-term continuous cultures, or chemostats. This experimental system allows me to continuously monitor changing population densities and host-phage evolution through time.

Photo of flasksRecent work has shown that phosphorus limitation strongly impacts both bacterial ecology and evolution, likely due to its biological importance in nucleic acid synthesis. In phage, phosphorus limitation has been shown to reduce reproductive capacity, likely due to the inability to produce enough nucleic acid for each new progeny. In a comparative study contrasting the effects of nitrogen (N) and phosphorus (P) limitation (based on N:P stoichiometric ratios), I found that long-term population dynamics and evolution of both host and phage are highly influenced by the local stoichiometric conditions.  Population densities were strongly altered, impacting not only the patterns but also population stability through time with P-limited dynamics more stable than N-limited.

Photo of two flasksPhage resistance in this particular chemostat experiment was selected for rapidly (within 9 days of exposure), regardless of the nutrient treatment, but spread through the population at different rates between the different nutrient limited environments. Infectivity patterns, generated by challenging bacterial and phage isolates through time and across treatments, are also strongly tied to the population dynamics suggesting that rapid evolution of bacteria is stoichiometrically dependent. Bacterial resistance to phage is easily detected with challenge experiments where phage strains are added to a given bacterial strain to assess their infection ability. In my assays with 96 well plates, [seen here], I am able to determine when resistance to infecting phage developed simply based on color; when a bacterial strain is susceptible, cells are unable to grow and the wells remain clear as compared to a resistant strain which grows into a dense pink culture.

Photo of assay platesThe qualitative differences in the infectivity patterns suggest that bacterial isolates are likely developing context dependent resistance mechanisms. In order to elucidate these potential differences, I will be identifying the underlying genetic mutations associated with simultaneous adaptation to nutrient limiting conditions and phage predation with whole genome sequencing. In genomic studies of a closely related organism, Prochlorococcus, researchers have found the presence of genomic islands, pieces of rapidly mutating, non-conserved DNA sequences that confer both resistance and adaptation to phosphorus limiting conditions. It is possible that similar adaptations exist in Synechococcus which may explain the bacterial susceptibility differences between the nutrient treatments.

Collectively, these results demonstrate that ecological and rapid evolutionary feedbacks between bacteria and phage are context dependent, strongly regulated by the local nutrient environment in the laboratory. These patterns may also hold true in natural settings. Furthermore, these results suggest that species interactions in N- or P- limited environments, such as the Pacific and Atlantic Oceans, respectively, may be drastically different due to the stoichiometric conditions alone.  As predicted by recent mathematical theory, nutrient limitation can act as a stronger selective force than predation
by phage, but this is contingent on the identity of the limiting nutrient and its biological significance. When considering all the pieces to this puzzle, it is crucial to consider eco-evolutionary feedbacks, mediated by environmental nutrient stoichiometry, when explaining and predicting microbial populations. 

If you’re interested in the work I do or have questions, feel free to send me an email (larsenm9 at msu dot edu) or check out my website.

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