Linking evolution and dynamics of microbial ecosystems through theory and model organisms OR Exploring the evolution and metabolic balance of marine ecosystems across multiple scales of space, time and biological organization OR The Need for conjoint studies in microbial ecology and evolution: the unity of biological organization across multiple scales of space, time, and phylogeny Using model systems and theory to explore the evolution and metabolic balance of marine ecosystems across multiple scales of space, time and biological organization CO-PI’s: , Nigel Goldenfeld, and Carl Woese This project to understand the structure, dynamics and energy flow in Prochlorococcus and its phages in the oligotrophic oceans will address the way in which collective interactions determine population structure, metabolic balance and response to exogeneous perturbations in marine microbial ecosystems. Synopsis The last decade of research on Prochlorococcus has significantly advanced our understanding of the forces that shape the diversity and dynamics of marine microbial populations, and revealed the power of model systems for advancing microbial oceanography. The next decade calls for the development of tightly integrated experiments, observations, and theory. Here we propose an alliance that will contemporaneously blend these branches of scientific inquiry through a single focus on the Prochlorococcus model system. TOO PUSHY; more subtlety needed Background A major challenge in microbial oceanography is understanding how the interactions between microbes and their biotic and abiotic environment manifests itself in emergent biogeochemical processes. A powerful way to address this challenge is through theory-driven study of model organisms. As the simplest, and most abundant, well-defined1, and well-studied, microbe in the oceans, Prochlorococcus is a model worthy of continued development. Research on Prochlorococcus to date has revealed, for example, that: • Closely related cells (< 3% different in rRNA sequence) can display vastly different physiological properties: Environmental conditions that are lethal to one, can be optimal for another. • 3% seems about like E. coli vs Salmona sp. In any case some standard of comparison is needed 1 It can be “seen” in real time in the field using flow cytometry, and all Prochlorococcus cells differ by < 3% in rRNA sequence. • These ‘ecotypes’ display different relative abundances along gradients imposed by spatial and temporal environmental variability. • The degree to which ribotype can predict abundance along environmental gradients of light, temperature, and nutrients in the field is a function of both the degree of sequence identity used to define a taxon and the environmental variable. • The global gene pool of the roughly 1027 Prochlorococcus cells in the ocean is enormous: The cells share a core of 1200 genes, and on average carry 800 additional genes, 200 of which are unique to that cell. • A significant fraction of the non-core gene pool is located in hypervariable genomic islands which carry signatures of horizontal gene transfer, hold clues as to important selective agents in different environments, and to mechanisms of adaptation and evolution on environmentally relevant time scales. • Phage carry an arsenal of host-like genes that they employ to redirect host metabolism serving as an exchangeable expanded gene pool that is subject to different selective pressures than the host homolog. • [Something about how finding nitrate reductase genes in DNA fragments of wild Prohlorococcus instantly changed our image of the global nitrogen cycle: Suddenly a very significant fraction of the global chlorophyll – ie. Prochlorococcus -- could utilize nitrate. Our culture collection, which contained only strains that lacked this gene, had given us an entirely wrong impression of what Prochlorococcus was capable of. What else are we missing?] These and other observations have helped solidify a paradigm shift in our thinking about the very nature of microbial systems and their evolution. Letting go of ‘Darwinian thinking’ in which communities are shaped by the relative fitness of individual cells through ‘bottom up’ (growth limitation) and ‘top down’ processes (cell death), we view Prochlorococcus-and-its-phage (and by extension the entire microbial system) as a single entity shaped and sustained by complex information exchange mechanisms. Understanding the design of this system and how it responds to exogenous forces is essential for developing predictive models of ocean biogeochemical processes. Moreover this “emerging picture of microbes as gene-swapping collectives demands a revision of such concepts as organism, species, and evolution itself” [your words… I love them!]. In concert with this paradigm shift has been the proliferation of theory designed to unveil the mechanisms that underlay the structure and dynamics of microbial systems and their phage. What is rare, however, is contemporary partnerships between theorists and experimentalists in which the theory guides experiments and field observations and vice versa. That is the one of the goals of this work. Driving Questions: (1) What controls Prochlorococcus genomic diversity and ecotype abundance along gradients, and in fluctuating environments with strong (e.g near Bermuda), and weak (eg. near Hawaii) external forcing? With a 6 year historical data set for Prochlorococcus ecotypes (along with all of the ancillary data) at these sites, and archived DNA samples, we are in a good position to address this question. Classical ecological theory is unable to account in detail for the observed phenotypic diversity, being based upon the assumed role of periodic selective sweeps. Further, our previous work using the Prochlorococcus- cyanophage model system has established that it is too simplistic to view their microbe-phage interactions as purely antagonistic: the populations co-evolve and appear to establish a collective state whose detailed features are still unexplored. We can test these ideas explicitly in Procholorococcus, as well as construct detailed individual-based models emanating from the observational data that are spatially-extended, and properly treat demographic fluctuations to model predation. Related questions concern the distribution of ecotypes along a temperature/photon gradient, which appear to track growthoptima curves. Is this an expected outcome of (antagonistic) phage-predation, (cooperative) communal dynamics, or something else? What is the sensitivity of communities to noisy environments – can external variability in time and spatial heterogeneity drive diversity and quantitatively account for the observed abundance distributions? (2) What accounts for the population stability of Prochlorococcus-cyanophage communities? Thus far there is no evidence for lysogeny in this system, but we have reason to suspect that it must be there, and envision a number of approaches to reveal it – particularly though single cell sequencing. We also propose experimental evolution studies designed to understand the rates of co-evolution of host-and phage, greatly facilitated by next-gen sequencing. Are mutation rates, for example, elevated in cells that are exposed steadily to viral attack? ISN’T LYSOGENY JUST AN EASILY RECOGNIZED FORM OF STRESS RESPONSE (3) How do we link population dynamics with energy flow in the Procholorococcus system? A major challenge in ecology is linking population dynamics and biogeochemistry. The function of life is to saturate every thermodynamic gradient in a biogeochemical system, and it can be argued that we do not need to understand the details of “who” is doing it. But we suspect that the key to dynamics and adaptability is in these details, and insights can be gained through studying Prochlorococcus which – in a sense – defines the boundaries of the light harvesting zone of the oceans. Can thermodynamic ideas be used together with the determined microbe-phage interactions to understand the abundance tails? RANDOM SCRAPS BELOW HERE Gaps in our Knowledge/Proposed Work (just listing random thoughts- not well formed at all!) • Need a genetic system for Prochlorococcus to directly test hypotheses, and get at function of hypothetical genes (the most highly expressed gene in the wild is a hypothetical!) • Need to develop ways to understand the co-evolution of host and phage... and which phage can infect which host in the wild • Need to find the lysogens. They have to be there. (Relevant: Your paper modeling sysem with and without lysogeny (I certainly cannot not understand that paper!)?) • Experimental Evolution Experiments in the lab and the field: Sequence - Perturb- Select - Sequence • Directed/selected isolation of Genomic variants • Single cell genomics and population genetics in fluctuating environments. • Programmed cell death equivalent • Cell Cell communication Why this would not be funded through conventional mechanisms ONE GOOD REASON IS THAT IT CALLS INTO SERIOUS QUESTION, SOME “WELL ESTABLILSHED” EVOLUTIONARY DOGMA, AND UNLESS THE WORK IS TREATED AS A SYNTHETIC WHOLE, IT WILL INVITE REJECTION BY CONVENTIONAL WISDOM [Other things the rfi asks us to address, but I don’t think it is necessary as it is obvious] • What is well-characterized about the system (physics, chemistry, community) [this could be what is well characterized about Prochlorococcus/Phage system] • Results must be generalizable <<<<==== Nigel, I took you as saying “but not beyond photosynthetic metabolism and Prochlorococcus”. But I think that this statement gives us the go- ahead to go beyond that, to make some telling suggestions as to the overall evolutionary dynamic itself;. • Please both of you express yourselves on this; because that consensus is what I will follow. • ecosystem must have a tractable level of complexity • ecosystem must be experimentally manipulable