Single cell ecology

Many microorganisms live in communities that are spatially structured, for example in biofilms. Such communities exhibit activities and functions that are shaped by metabolic interactions between the community member. Interactions are expected to mainly occur between cells that are close in space. As a consequence, the nature and strength of the interactions that will occur will depend on the spatial arrangement of different types of microbial cells. In turn, the spatial arrangement is expected to be shaped by metabolic interactions, which determine regions where a given cell type grows well. The Ackermann group’s goal here is to better understand this interplay between the spatial arrangement of different types of microbial cells and the interactions that arise between them. Working with synthetic consortia of different E. coli strains with well-defined metabolic interactions, Martin can quantify the spatial range over which interactions occur and understand the consequences for the spatial self-organisation of these multi-genotype systems. The goal of this work is to contribute to identifying general principles that govern how different types of microorganisms organise in space, and how this spatial self-organisation shapes the activities and functions of microbial systems.

The mechanical properties of cells have long been established as a sensitive and label-free biomarker. While mechanical cell assays have been traditionally limited to low throughput or small sample size, the introduction of real-time deformability cytometry (RT-DC) increased analysis rates to up to 1,000 cells per second. RT-DC has demonstrated its relevance in basic and fundamental life science research, e.g. by observing the activation of immune cells and describing the membrane dynamics of Malaria pathogenesis. However, linking immune cell activation to underlying tissue alterations has not been possible so far. Here, the concept of virtual fluidic channels is introduced to bridge the gap between single cell rheology and tissue mechanics. Virtual channels can be created in almost any microfluidic geometry and can be tailored dynamically towards hydrodynamic stress distributions sufficient to probe the rheology of arbitrary cell sizes. Using spheroids as a tissue model, results from virtual channel measurements indicate that the Young’s modulus of single cells exceeds the one of spheroids and that their elasticity increases with size. The availability of a high-throughput assay for mechanical spheroid characterization might lead to a better understanding of tissue rheology and help to study the interplay of virus infiltration and tissue regeneration.

The bacterial and archaeal tree of life has undergone significant expansion, chiefly from candidate phyla obtained through genome-resolved metagenomics and, at smaller scale, via single-cell sequencing efforts. Following this path, viral diversity is being uncovered at a rapid pace. Tanja Woyke discusses how the combination of flow cytometry followed by genome amplification and sequencing can provide a means to cataloguing uncultivated microbial and viral diversity. Focusing on Nanoarchaeota symbionts attached to their hosts, she illustrates that this approach further allows the assignment of novel putative host associations, facilitating the exploration of cell-cell interactions and fine-scale genomic diversity.

Recent developments in community and single-cell genomic approaches have provided an unprecedented amount of information on the ecology of microbes in the aquatic environment. However, linkages between each specific microbe’s identity and their in situ level of activity (be it growth, division, or just metabolic activity) are much more difficult to obtain. One of the ultimate goals of marine microbial ecology would be integrate three levels: the genomic (including identity) one, the activity/growth one, and the morphology/visualisation, and all this for as many individual cells as possible, as a means to understand how each environmental characteristic determines the types of different microbes in nature and their activity or growth, alongside with information on morphology and cell-to-cell associations. Recent reviews have stressed ways for capturing the activity level of different genomic entities, but often one of the three legs, that of visualization, has not been well covered by the available methods. A review of current methodologies that have been applied to marine microbes, particularly prokaryotes, will be presented combined with a discussion of the difficulties in identifying and categorizing activity and growth, in doing so at with the minimal manipulation of the environment, and the level of within-population single-cell variability in activity that occurs in natural marine environments.

Data-rich, molecular analyses are becoming important sources of knowledge about the composition and activities of microorganisms in diverse environments. Yet, today only a small fraction of these meta-omics data (short DNA, RNA and peptide sequences) can be accurately assigned to specific lineages and metabolic pathways, due to the lack of adequate reference genome databases. To address this challenge, the Stepanauskas group sequenced 20,000 individual bacteria, archaea and viral particles from the tropical and subtropical epipelagic using an unbiased, Big Data approach. This GORG-Tropics database provides some of the first insights into the global pangenomes, infections, biogeography and sizes of many of the bacterial and archaeal lineages that dominate surface ocean. The group shows that GORG-Tropics enables accurate taxonomic and functional assignments of the majority of meta-omics data from this global environment. Intriguingly, the group found that every sequenced cell was genomically unique, and that a large fraction of the global bacterioplankton coding potential was present in a single drop of seawater. GORG-Tropics expands the capacity of oceanography’s cyber-infrastructure and enhances the interpretation of diverse marine omics studies. It also demonstrates that relevant genomic representation of complex microbiomes is an achievable goal and may be applied in other environments.

One challenge associated with developing a functional understanding of microbial ecosystems is the magnitude of the diversity encompassed by the word “microbial”. Most ecology is biased towards a few macroscopic plants and animals. In contrast, microbial ecosystems are made up of more than ten-times the diversity of eukaryotes alone, in addition to bacteria, archaea, and viruses. Microbial ecology is also heavily biased, towards bacteria (and sometimes archaea). So we are faced with a need to develop a fuller picture of these networks, but to do so both a quantitative and qualitative problem, because the diversity is so great that neither the technical approaches nor the basic biology required to understand one group is readily translatable to others. For microbial eukaryotes there are paths forward to solving many technical challenges though single cell methods, and advances in both genomics and transcriptomics will be discussed. But this still leaves a thornier conceptual problem of how such data will (or will not) reveal ecological roles of microbial eukaryotes because translating genomics to ecological function is largely restricted to metabolism. For most microbial eukaryotes, complex cellular structures and resulting behaviours are much more significant, but we lack strong conceptual frameworks to derive these traits from genomic data alone.

For decades and longer scientists have described diverse unicellular eukaryotic organisms in aquatic ecosystems. The introduction of molecular approaches to ocean ecology has expanded our view of the diversity of marine protists and their importance in these ecosystems, yet many key taxa remain uncultured. Hence it has been difficult to examine their evolutionary relationships beyond what is possible with marker gene analyses. Likewise, it has not been possible to characterise their nuclear and organellar genomes. Moreover, lack of cultures has obviated deeper exploration of their interactions with other biological entities in the sea. In 2005 the Worden group began using at-sea flow cytometry to sort wild protists in order to generate both targeted (population) metagenomes and eukaryotic single-cell metagenomes. In addition to providing insights into the evolution and distribution of wild phytoplankton species these studies are increasingly revealing co-associations between the targeted eukaryote and other entities. Here, Alexandra will discuss a suite of findings from studies in the tropical Atlantic and eastern North Pacific Oceans. Her long term goal is to use these approaches to facilitate research that goes deeper into the biology of key uncultured groups and their ecological interactions. Collectively, these types of studies are critical for establishing a baseline against which future changes in microbial community structure can be assessed and for understanding the diversity and evolution of marine protists.