Monday, September 17, 2018

Getting the wrong tree when reticulations are ignored

One issue that has long intrigued me is what happens when someone constructs a phylogenetic tree under circumstances where there are reticulate evolutionary events in the actual (ie. true) phylogeny itself. That is, a network is required to accurately represent the phylogeny, but a tree is used as the model, instead. How accurate is the tree?

By this, I mean that, if the phylogeny can be thought of as a "tree with reticulations", do we simply get that tree but miss the reticulations, or do we get a different (ie. wrong) tree?

Sometimes, people refer to this situation as having a "backbone tree" — the phylogeny is basically tree-like, but there are a few extra branches, perhaps representing occasional hybridizations or horizontal gene transfers. The phylogenetic tree can then be treated as a close approximation to the true phylogeny, representing the diversification events but not the (rarer) reticulation events.

I have argued against this approach (2014. Systematic Biology 63: 628-638.). Instead of seeing a network as a generalization of a tree, we should see a tree as a simplification of a network. If we do this, then we would construct a network every time; and sometimes that network would be a tree, because there are no reticulation events in the phylogeny. It cannot work the other way around, because we can never get a network if all we ask for is a tree!

Presumably, if there are no reticulations then we should get the same answer (phylogenetic tree) irrespective of whether we simply construct a tree or instead construct a network that turns out to be a tree. But what about the "backbone tree" situation? Here, it has always seemed to me to be possible that we do not get the same tree. If this is so, then constructing a tree and then adding a few reticulations to it (as is often done in the literature) would not work — we would be adding reticulations to the wrong backbone tree.

There are two possible ways in which we can get the wrong backbone tree: the topology might be incorrect, or the branch-lengths might be incorrect (or both). For example, if there are true reticulations and yet we do not include them in our model, I have argued that the branches will be too short (2014. Systematic Biology 63: 847-849.) — two taxa will be genetically similar because of the reticulation events, but the tree-building algorithm can only make them similar on the tree by shortening the branches (not by adding a reticulation).

Fortunately, for at least one tree-building model Luay Nakhleh and his group have now done some simulations to answer my questions. You may not yet have noticed their results, because they are not necessarily in the most obvious place; so I will highlight them here. The analyses involve the Multispecies Coalescent (MSC) model, which accounts for incomplete lineage sorting during the tree-like part of evolution, as compared to the Multispecies Network Coalescent (MSNC) which adds reticulations (eg hybridization) to the model.

Dingqiao Wen, Yun Yu, Matthew W. Hahn, Luay Nakhleh (2016) Reticulate evolutionary history and extensive introgression in mosquito species revealed by phylogenetic network analysis. Molecular Ecology 25: 2361-2372.

This paper compares a tree-based analysis (construct a tree first then add reticulations) with a network-based analysis (construct a network) for an empirical genomic dataset. The two results differ.

Dingqiao Wen, Luay Nakhleh (2018) Coestimating reticulate phylogenies and gene trees from multilocus sequence data. Systematic Biology 67: 439-457.

Tucked away in the Supplementary Information are the results of a set of simulations comparing the MSC (using *Beast) and the MSNC (using PhyloNet), with (section 3) and without (section 2) reticulations. The basic conclusion is that, in the presence of reticulation, tree-based methods either get the tree completely wrong, or they get the tree topology right but the branch lengths are "forced" to be very short. A summary of the latter result is shown in the figure above. In the absence of reticulation, both methods produce the same tree.

R.A. Leo Elworth, Huw A. Ogilvie, Jiafan Zhu, and Luay Nakhleh (ms.) Advances in computational methods for phylogenetic networks in the presence of hybridization. (chapter for a forthcoming book]

A summary of the group's work to date. Section 6.3 summarizes the results from the paper 2.

Monday, September 10, 2018

Limitations of the new book about HGT networks

This is a joint post by David Morrison and Ajith Harish.

There has been a flurry of reviewing activity recently about the new book:

The Tangled Tree: a Radical New History of Life
David Quammen. 2018. Simon & Schuster.

This book has received glowing reviews, including:

The book is intended for the general public, rather than for specialists, explaining the "new view" of evolutionary history that includes extensive horizontal gene transfer (HGT), especially in the microbial world. Quammen describes himself as a science, nature and travel writer, so his book is more than just a record of science, and is as much about the people involved as about the scientific theory. In particular, it contains a biography of Carl Woese.

Quammen’s recent New York Times feature article The scientist who scrambled Darwin’s Tree of Life is a very good primer to his book. For us, it indicates that the book has many overlaps with Jan Sapp's earlier book The New Foundations of Evolution: on the Tree of Life (2009. Oxford University Press). The publisher’s advertised selling point of that book is: "This is the first book on (and first history of) microbial evolutionary biology, and that it puts forth a new theory of evolution", with HGT being the new theory. In this sense, the "radical new view" is simply that genetic material can be transferred without sexual reproduction, an idea that goes back rather a long way in history (see The history of HGT), and which is often seen as anti-Darwinian.

Bill Hanage in his review of Sapp’s book (2010. The trouble with trees. Science 327: 645-646) argues that the book neither puts forward a new theory nor is the debate actually about horizontal gene transfer, and the Tree of Life is thus far from settled. There are many other interesting points discussed in that review. Furthermore, even after almost 10 years, Hanage’s review of Sapp’s 2009 book can be substituted verbatim as a review of Quammen’s 2018 book! This PDF shows how the book review would read if the author and book names in Hanage’s review were to be substituted [reproduced with the permission of the original author].

The debate allegedly involving HGT is, at heart, about explaining the pattern of extensively mixed genetic material found in the akaryotes. However, simply looking at a pattern does not tell you about the process that created the pattern. In order to study processes, we need a model, in this case a model about how evolution occurs. The "HGT model" is that the Last Universal Common Ancestor (LUCA) of life was a relatively simple organism genetically, and that subsequent evolutionary history has involved complexification of that ancestor, both by diversification and by HGT.

What the two books do not explore is the other major model for the current distribution of genetic material among akaryotes. This alternative scenario is that the LUCA was genetically complex, and that the subsequent evolutionary history involved independent losses of parts of the genetic material — the sporadically shared material is basically coincidental. All that this model requires is that there be evolutionary history prior to the LUCA, during which it became a complex organism from its simple beginnings — the LUCA is merely as far back as we can see into the past, with the prior history being unrecoverable by us (ie. we cannot see past the LUCA bottleneck).

Over the past couple of decades, a number of papers have explored the evidence for the latter idea, from both the RNA and protein perspectives, including:
  • Anthony Poole, Daniel Jeffares, David Penny (1999) Early evolution: prokaryotes, the new kids on the block. BioEssays 21: 880-889.
  • Christos A. Ouzounis, Victor Kunin, Nikos Darzentas, Leon Goldovsky (2006) A minimal estimate for the gene content of the last universal common ancestor — exobiology from a terrestrial perspective. Research in Microbiology 157: 57-68.
  • Miklós Csűrös István Miklós (2009) Streamlining and large ancestral genomes in Archaea inferred with a phylogenetic birth-and-death model. Molecular Biology and Evolution 26: 2087-2095.
  • Kyung Mo Kim, Gustavo Caetano-Anollés (2011) The proteomic complexity and rise of the primordial ancestor of diversified life. BMC Evolutionary Biology 11: 140.
  • Ajith Harish, Charles G. Kurland (2017) Akaryotes and Eukaryotes are independent descendants of a universal common ancestor. Biochimie 138: 168-183.
Finally, even from the perspective of phylogenetic networks, Quammen's book is very one-sided. In particular, the other processes that lead to reticulate evolution (eg. introgression and hybridization) are pretty much ignored. That is, the focus is on akaryotes not eukaryotes. The latter are also of phylogenetic interest.

Monday, September 3, 2018

More on networks for placing fossils, such as Eocene lantern fruits

A colleague pointed me to a paper published last year in Science about a spectacular fossil find: an Eocene Physalis-fruit with a preserved lampion. In an recent post, I advocated Neighbor-nets as nice and quick tools to place fossils phylogenetically. In this post, I'll will exemplify this once more, and argue why this would have been even more informative than what the authors showed as graphs.

The study and the data

In their 2017 paper, Wilf et al. (Science 355: 71–75) describe a new fossil find, which, by itself, rejects the often-too-young molecular dating estimates for Solanceae, the potato-tomato family, the "Nightshades". The Nightshades include many well-known plants, in addition to potato/tomato (the latter is phylogenetically a subclade of the potatoes) — we have e.g. the tobacco genus (Nicotiana), and also the genus Physalis, which includes several species commercialized as fruits (e.g. P. peruviana, also known as Cape gooseberry or goldenberry) and ornamental plants (e.g. P. alkekengi, the Chinese Lantern).

Just by looking at the pictures showing the fossil (Wilf et al.'s text-Fig. 1), anyone who ever ate a physalis, would agree that it was produced by a member of the genus. However, science is not usually about common sense, but about formal reconstructions. Thus, the authors placed their fossil using a total evidence tree approach: they scored 13 morphological traits as binary or ternary characters, concatenated these data with a molecular data set and inferred trees under maximum parsimony (their text-Fig. 2, below) and maximum likelihood (the tree can be found in the supporting information).

Wilf et al.'s total evidence tree showing the (quoted from the legend)
"Phylogenetic relationships of Physalis infinemundi sp. nov. and selected Solanaceae species" (their Fig. 2). Strict consensus of 2835 most parsimonious trees of 3510 steps (CI = 0.438, RI = 0.726)."

Based on the graph, one can confirm that the fossil (arrow; pictured, too) is part of the core Physalis, but its position within this core clade is unresolved. The Decay index shown indicates that moving the entire branch would require just one step more. Not overly re-assuring regarding the total length of the tree (3510 steps) and underlying data (the used matrix has 7070 characters!)

The molecular data were selected from an earlier study (Särkinen et al., BMC Evol. Biol., 2013), but the total evidence matrix is not provided (see this post on why we want to publish our phylogenetic data). But at least the "...morphological matrix developed in this paper is tabulated in the supplementary materials."

This file includes two sheets: the first shows the "raw scores", including four continuous characters, and the second shows the "character scoring" used for the analysis, where the continuous characters were scored (binned) as ternary and binary characters. The iinformation provided is partly wrong, likely to be the result of copy & paste errors (this is another reason why it should be obligatory for phylogenetic studies to provide the data as aligned-FASTA or NEXUS file). A corrected version of the "character scores" sheet based on the "raw scores" sheet is included in the figshare submission for this post.

By just filtering this matrix for same-as-in-the-fossil characters, we can identify two extant species that are identical to the fossil in all scored characters: Physalis acutifolia and P. lanceolata. Both are part of the Physalis core clade in Wilf et al.'s total evidence tree, but their position is as unresolved as that of the fossil.

Enlarged part of the above figure, showing the absolute character difference (0 to 5 out of 13 covered characters) between the fossil and other members of the Physalis core clade.

The reason for this becomes clear in the total-evidence maximum-likelihood tree. Here, the fossil is resolved as the sister of P. lanceolata (maximum likelihood bootstrap support: ML-BS < 70, the actual value would have been nice), to which it is identical, both being deeply nested in the Physalis core clade. However, the other identical species (morphologically), P. acutifolia, is placed in the first diverging subclade of the core clade (ML-BS < 70, along with most of the backbone of this clade). The "low" support may have two possible reasons:
  • the fossil, with 99.8% missing data, acts as a 'rogue' taxon; or
  • the genetic data provides little discriminating or ambiguous signals.
Solanaceae genera can be tricky, and the gene sample lacks high-divergent sequence regions. Since the molecular data are not documented, I can't assess how significant this separation is, but it appears to be supported by at least some mutations: the tree-wise distance is about 0.04 expected substitutions; and the two morphologically indistinct (regarding the scored characters) species are genetically distinct (to some degree).

Extract from Wilf et al.'s Fig. S1, showing the Physalinae subtree with the core Physalis clade and the deeply nested fossil P. infinemundi (in bold font). Support is only shown for branches with a ML-BS support ≥70.

Trees may fail to show the obvious, but networks won't

Just by using the Neighbour-net to visualize the signal in the morphological partition, we can directly argue that the fossil is likely to be part of the core Physalis. Thus, being Eocene of age, rejects the much-too-young age estimates in e.g. the dated tree by Särkinen et al. (the reference for the molecular data used by Wilf et al.)

Neighbour-net splits graph based on the morphological data partition included in Wilf et al.'s "supermatrix".

In contrast to the little information that comes along with the tree shown above (soft-ish polytomy, weak Decay index, potentially decreased ML-BS support), the splits graph highlights the ambiguity (incompatibility) of the morphological signal. The graph shows little tree-likeness, and members of the same (sub)tribe show little coherence (C = Capsiceae, H = Hyoscyameae, J = Juanulloeae, S = Solaneae; W = Withaninae; all represented by de-facto molecular clades with ML-BS ≥ 77 in Wilf et al.'s supplement Fig. S1). There is one notable exception: members of the core Physalis (red dots) are sufficiently distinct from anything else, forming a highly supported clade (ML-BS = 98 in Wilf et al.'s fig. S1),.

The network also shows that the fossil is identical to both P. acutifolia and P. lanceolata.

Neighbour-net after reducing the taxon set to the phylogenetic neighbourhood of the fossil specimen. Filled fields indicate sister/sibling species supported by a ML-BS >= 80 in Wilf et al.'s "total evidence" ML tree.

By focusing on the phylogenetic neighborhood of the fossil, we end up with a spider-web-like graph. Which means that the morphological partition has little consistent signal for recognizing potential relatives: the same features are likely to have evolved in parallel (all members of this neighborhood a likely to share a common origin) — 50 million years (and more) is a long time for a lineage to end up with a similar fruit (see also the maximum-parsimony character reconstructions on the parsimony strict-consensus tree provided in the supplement to Wilf et al.'s study).

Data and graphs

The Splits-NEXUS files for the Neighbor-nets and NEXUS-versions of Wilf et al.'s Data S1, as well as additional graphics (network with labeled bubbles) can be found on figshare.