Major new microbial groups expand diversity and alter our understanding of the tree of life

I still believe in revolutions. And sometimes they just happen, almost unnoticed. One such revolution happened on a boring 11th of April 2016 when Laura Hug et al. published their new tree of life in the journal of Nature Microbiology. Many textbook trees of life are centered on eukaryotic evolution and underestimate the true global diversity of organisms. This is because it is easier to find large critters than small ones and also because many environments have been under-sampled in the past. Now we have the tools to capture microorganisms from extreme and elusive places, for example hot vents on ocean floors, soils in the rainforest, or shrinking ice on the poles. The only thing we need to find new microbes is an environmental sample (e.g. water or soil), a DNA extraction kit, a sequencing machine, and a computer. Oh yes, and a few clever scientists like Laura and her colleagues. They added new genomic data from more than 1000 uncultivated and little known organisms to the current tree of life. The addition of these sequences to the tree revealed an astounding diversity and reminds us that we do not know much yet about the true diversity of life on this planet.

Cindy Castelle and Jillian Banfield (who is an author of the Hug paper, too) just published a review about the implications of this new tree of life for the rest of us in the journal Cell. In this blog post, I will summarize this review with an emphasis on a few topics that I find very exciting. In summary, Hug et al.’s publication quietly suggested a change in the structure of the tree of life and enriched our understanding of the biology, evolution and metabolic roles of microorganisms.

First a little bit of information about the methodological approach (skip this paragraph if you only want to read about results and implications):

Hug et al. recovered more than 1000 microbial genomes of previously underrepresented bacterial and archaeal lineages. They collected environmental samples, extracted DNA from them, sequenced them, and manually binned the sequences into genomes in their lab. This last step is very labor-intensive and not completely automated yet. I imagine a room full of postdoctoral researchers sitting in front of expensive computers. This room has a glass window where you can look at them working and a sticker on the glass that says: “Please do not tap on the glass, postdoctoral researchers are very sensitive.” If you sample the environment you will end up with DNA from bacteria, archaea, eukaryotes, phages, viruses and plasmids. For their analysis they concentrated on archaeal and bacterial genomes. Once they had their microbial genomes taped together, they built a phylogenetic tree using a concatenated set of 16 ribosomal protein sequences from each organism. These genes are assumed to be subject to the same evolutionary processes. They are also co-located in the same genomic region of archaea and bacteria, which reduces binning errors. They included one representative of each genus (3,083 organisms) in the tree. The placement and evolution of eukaryotes in the tree of life is still not fully resolved yet. More about this later. To place this group in the tree, they used a nuclear-encoded ribosomal protein that is independent of cellular structures.

A current view of the tree of life suggested by Hug et al. It shows the total diversity represented by 3083 sequenced genomes.

New major groups:

I assume you just had a first glance at the tree. I would say the most striking new group is the purple one of the candidate phyla radiation (CPR). This group includes a new radiation of a whole phylum! CPR had first been discovered as OP11 in hot springs of the Yellowstone National Park by Hugenholtz et al. in 1998. Soon it became clear that this group harbors many other phyla (Rinke et al., 2013). During the next couple of years many more major lineages were added to this group (Brown et al., 2015). Kelly Wrighton, a previous member of Jillian Banfield’s lab, was the first one who studied the CPR at the whole genome level, and this already in 2012! Using genomes that had been assembled from environmental samples at extreme environments (acetate-amended aquifers), it could even be shown that some CPRs use an alternative genetic code (Wrighton et al., 2012; Campbell et al., 2013)! The CPR contains at least 73 subgroups and represents a monophyletic radiation of phyla. Now digest that.

It is worth mentioning that Parks et al. point out that the relative diversity of the CPR group varies substantially depending on which protein markers are studied (Parks et al., 2017).

What is a phylum anyways? And what is a superphylum?

In my own words, a phylum is a group of organisms that are more similar to each other than they are to any other groups. Such a group can then be called a phylum. Ernst Haeckel coined this term in 1866 from the Greek word phylon meaning ‘race’ or ‘stock’. Similarity is measured as genetic relatedness. The more genetic data we have about organisms, the better we can quantify such relatedness. As more data become available we might even split some phyla or merge others. Some phyla show a clear and reliable grouping among each other based on genetic sequence information. They can be grouped into a ‘superphylum’. It is not clear how many phyla and superphyla there are. Moreover, there are no fixed rules for naming them. Usually it helps us humans to group things in order to better understand them. A phylogenetic tree is the most important organizing principle in biology (Hug et al., 2016). It shows how we are related to each other, where we came from, and it lets us reconstruct evolutionary processes.

When you look at this new tree you realize that this new superphylum of the CPR shows an enormous extension of evolution that has occurred within. The purple part of the tree accounts for a large proportion of its total diversity. It has a comparable width of diversity to the rest of all bacteria. It is not clear yet, whether this is due to early divergence or rapid evolution (or both).

Hug et al. demonstrated that cultivation-independent genomics, that means the retrieval of genomes without culturing the microbes first in the laboratory, has expanded the domain of bacteria over five times. For the domain of archaea the expansion is similar.

New groups within the domain of archaea:

Within the archaea there is a new group that was named DPANN, a newly recognized superphylum that is described herein (Castelle et al., 2015). The term DPANN comes from Diapherotrites, Parvarchaeota, Aenigmarchaeota, Nanoarchaeota, and Nanohaloarchaeota. Most DPANN archaea depend on other microbes to survive. They often sit on the surface of other microbes and exchange resources through pili-like structures. They all have small genomes, small cell sizes, and many lack core biosynthetic pathways for nucleotides, amino acids and lipids. I will elaborate more on other archaeal groups further below.

How diverse is the tree of life?

Now when we look at the tail of the tree we see this evanescently small group of eukaryotes. In their defense I must say that Hug et al. did not focus on eukaryotic diversity. Nevertheless, eukaryotic diversity is still not much compared to the other domains. Most probably because we have a relatively young evolutionary history. Nothing compared to the many new branches of bacteria and archaea comprising taxa that have been discovered between 2012-2016. How is this going to continue? Will the increasing diversity plateau at some point? How big would a complete tree of life be?

You might also wonder why the discovery of new taxa happened only so recently but so quickly?

Cindy and Jillian give three major explanations:

  • Because many taxa live on subsurface environments that have not been sampled before. They are difficult to sample.
  • Many new taxa are lower-priority targets because these microbes are not associated with us humans, economically-important animals or crops.
  • Cultivation-independent methods are pretty recent.

Genomic data for uncultivated microbes resulted in key discoveries related to their roles in carbon, nitrogen and sulfur cycling (reviewed in Adam, Borrel, Brochier-Armanet, & Gribaldo, 2017; Spang et al., 2015). It is very relevant for our understanding of the ecology and the evolution of microorganisms and it led to a revolution in the whole field. Or at least in my field. I picked out a few stories from the review.

Story one – Cyanobacteria in human fecal samples:

This has been a long-standing mystery. Many 16S amplicon studies (studies that aimed at looking at bacterial community composition based on the amplification of a bacteria-specific gene) found cyanobacteria in human fecal samples. It was not clear what they were doing there. Were the samples contaminated or are cyanobacteria members of the human gut microbiome? Using metagenomic approaches a complete genome was constructed (Di Rienzi et al., 2013). Then, it was compared to other, available cyanobacterial genomes. Metabolic predictions based on the gene sequences that were found in these cyanobacterial genomes revealed the absence of the whole photosynthetic machinery that you usually find in cyanobacteria. Hence, these relatives in the human gut have a fermentation-based anaerobic metabolism. Phylogenetic analyses placed these bacteria in the candidate phylum Melainabacteria (no, not Melania-bacteria…). It is not fully resolved yet how they relate to the photosynthetic cyanobacteria. However, it looks like photosynthesis evolved after the divergence of cyanobacteria. Moreover, this data supports the hypothesis that aerobic respiration arose after the evolution of photosynthesis on earth. Those Melainabacteria that have been found in human guts can synthesize several B and K vitamins, which suggests that these bacteria are beneficial to their host (Di Rienzi et al., 2013; Soo et al., 2014).

Now, since I already stumbled over their name, here is some more background about it: The name ‘Melainabacteria’ is derived from the title of a Greek nymph of dark waters. Melainabacteria are a candidate phylum. That means they are in the process of being evaluated and might become a real phylum (as discussed above).

Green cyanobacteria. This is how we traditionally think of them. Wikimedia Commons by Christian Fischer. https://commons.wikimedia.org/wiki/File:Cyanobacteria_Aggregation1.jpg

Story two – Episymbiosis:

Many CPR and DPANN live a very unusual lifestyle. A few of them are associated with eukaryotic hosts, however, most of them are likely symbionts of bacteria and archaea. They like to sit on the surface of their hosts (episymbiosis; Luef et al., 2015). A closer look with cryogenic transmission electron microscopy (short cryo-TEM) gave us some insight into their strange lifestyle. They have pili-like structures that extend from their cell surfaces to their host. They might provide access to nutrients and allow for an exchange of metabolites. There could be new types of symbioses that we have not even seen before. They make us rethink concepts related to microbial community structure and functioning. In the near future real-time and low-cost sequencing will become common and genomically-informed ecosystem manipulations will come within reach. What an exciting time to be a researcher!

Freya, the queen of gods. Wikimedia Commons. Scanned from ‘A book of myths’ (1915) G.P. Putnam’s sons; London. Copy at New York Public Library, scanned by Nicole Deyo

When will microbes be (named after) goddesses???

 

Where do we belong ?

I will wrap this post up with a little bit of Norse mythology. If you zoom into the domain of archaea you will see that some of them were named after Norse gods, such as Loki or Thor. These organisms belong to the group of the Asgard archaea, also including Odin and Heimdall. A team of scientists led by Thijs Ettema from Uppsala University in Sweden came up with these names. These Scandinavian scientists do not get much light during winter. However, this is not the reason for the mysterious naming. Ettema and his colleagues, and collaborators from all over the world, sampled extreme environments for archaea using ‘genome-resolved metagenomics’, the same approach that Hug et al. applied. Ettema was driven by this idea that an Asgardian god, umm, I mean microbe, gave rise to us eukaryotes. He is not the first one who thinks that. This hypothesis is not new. Martin and Müller published it in 1998 in the journal  Science (Martin & Müller, 1998). Somehow a bacterium found its way into an archaeon. They both felt OK with it and survived. The bacterium provided the archaeon with more energy, which helped the latter grow bigger, accumulate more genes and evolve into a eukaryote. The bacterium eventually turned into a mitochondrium, a little machinery that powers eukaryotic cells until today. In 2015, Ettema and his collaborators, led by Anja Spang found an archaeon that was more closely related to eukaryotes than all existing archaea (Spang et al., 2015). They found that little critter in a field of hydrothermal vents between Greenland and Norway at Loki’s Castle and named it Lokiarchaeota. After this discovery many other scientists named their archaea after other Norse gods. The story does not end here. When these Asgardian gods are included in the tree of life, eukaryotes do not cluster anymore as a separate domain. Instead, they fall within the domain of archaea.

Cropped from Fig. 2 in Hug et al. Here each major lineage represents the same amount of evolutionary distance.

This post has been updated on the 20th of March to include the paper of Wrighton et al. (2012). Thank you Jillian Banfield for reading and improving this post.

References:

Adam, P. S., Borrel, G., Brochier-Armanet, C., & Gribaldo, S. (2017). The growing tree of Archaea: new perspectives on their diversity, evolution and ecology. ISME Journal, 11(11), 2407-2425. doi:10.1038/ismej.2017.122
Brown, C. T., Hug, L. A., Thomas, B. C., Sharon, I., Castelle, C. J., Singh, A., . . . Banfield, J. F. (2015). Unusual biology across a group comprising more than 15% of domain Bacteria. Nature, 523(7559), 208-211. doi:10.1038/nature14486
Campbell, J. H., O’Donoghue, P., Campbell, A. G., Schwientek, P., Sczyrba, A., Woyke, T., . . . Podar, M. (2013). UGA is an additional glycine codon in uncultured SR1 bacteria from the human microbiota. Proceedings of the National Academy of Sciences of the United States of America, 110(14), 5540-5545. doi:10.1073/pnas.1303090110
Castelle, C. J., & Banfield, J. F. (2018). Major new microbial groups expand diversity and alter our understanding of the tree of life. Cell, 172(6), 1181-1197. doi:10.1016/j.cell.2018.02.016
Castelle, C. J., Wrighton, K. C., Thomas, B. C., Hug, L. A., Brown, C. T., Wilkins, M. J., . . . Banfield, J. F. (2015). Genomic expansion of domain archaea highlights roles for organisms from new phyla in anaerobic carbon cycling. Current Biology, 25(6), 690-701. doi:10.1016/j.cub.2015.01.014
Di Rienzi, S. C., Sharon, I., Wrighton, K. C., Koren, O., Hug, L. A., Thomas, B. C., . . . Ley, R. E. (2013). The human gut and groundwater harbor non-photosynthetic bacteria belonging to a new candidate phylum sibling to Cyanobacteria. eLife, 2. doi:ARTN e01102 10.7554/eLife.01102
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Luef, B., Frischkorn, K. R., Wrighton, K. C., Holman, H. Y. N., Birarda, G., Thomas, B. C., . . . Banfield, J. F. (2015). Diverse uncultivated ultra-small bacterial cells in groundwater. Nature Communications, 6. doi:ARTN 6372 10.1038/ncomms7372
Martin, W. & Müller, M. (1998). The hydrogen hypothesis for the first eukaryote. Nature, 392(6671), 37-41. doi:Doi 10.1038/32096
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Rinke, C., Schwientek, P., Sczyrba, A., Ivanova, N. N., Anderson, I. J., Cheng, J. F., . . . Woyke, T. (2013). Insights into the phylogeny and coding potential of microbial dark matter. Nature, 499(7459), 431-437. doi:10.1038/nature12352
Soo, R. M., Skennerton, C. T., Sekiguchi, Y., Imelfort, M., Paech, S. J., Dennis, P. G., . . . Hugenholtz, P. (2014). An expanded genomic representation of the phylum Cyanobacteria. Genome Biology and Evolution, 6(5), 1031-1045. doi:10.1093/gbe/evu073
Spang, A., Saw, J. H., Jorgensen, S. L., Zaremba-Niedzwiedzka, K., Martijn, J., Lind, A. E., . . . Ettema, T. J. G. (2015). Complex archaea that bridge the gap between prokaryotes and eukaryotes. Nature, 521(7551), 173-179. doi:10.1038/nature14447
Wrighton, K. C., Thomas, B. C., Sharon, I., Miller, C. S., Castelle, C. J., VerBerkmoes, N. C., . . . Banfield, J. F. (2012). Fermentation, hydrogen, and sulfur metabolism in multiple uncultivated bacterial phyla. Science, 337(6102), 1661-1665. doi:10.1126/science.1224041

Thanks @casettron for your valuable comments on an earlier draft of this post!

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