2/13/17

Phylogenetic trees

Bas E. Dutilh
Systems Biology: Bioinformatic Data Analysis
Utrecht University, February 14th 2017

After this lecture, you can…

… recognize different types of nodes in phylogenetic trees
… interpret branch lengths in the light of evolutionary time
… collapse branches with low support values
… rotate branches and interpret radial and polar trees
… infer the phenotype of ancestors using parsimony
… distinguish orthologous and paralogous genes in a tree
… explain how gene functions evolve
… transfer functional annotations between genes
… reconciliate gene trees and species trees
… identify gene losses and horizontal transfer events
… list the causes of phylogenetic inconsistencies

1
 2/13/17

Phylogeny
• Term coined by Ernst
Haeckel (1866)
– Phylon (fulon)
• Tribe
• Race
– Genus
• Birth
• Origin
• At every node in the tree, a
new lineage is born
• All lineages in a tree are
related because they
descend from the same root
• Tree topology shows how
the lineages are related

Phylogenetic trees
The horizontal lines are branches
• A phylogenetic tree represents and represent evolutionary
the phylogeny of species or lineages changing over time.

sequences
The vertical lines represent nodes
or evolutionary splits. Line length
has no meaning; lines just show
which branches are connected.

The branch length represents the

evolutionary time between two nodes.
Unit: substitutions per sequence site.
http://epidemic.bio.ed.ac.uk/how_to_read_a_phylogeny

2
 2/13/17

Nodes in a tree
• Tips (sometimes called “leaves” or “terminal nodes”)
– Present day species or sequences
– The only things we can directly measure
– Contain information used to build the tree
• Ancestral nodes (or “internal nodes”)
– Last common ancestor of
its daughter lineages
• Root (sometimes)
– Last common
ancestor of
whole tree
– If the tree is
Time axis: away from root
rooted, then
the time axis is
defined away
from the root http://epidemic.bio.ed.ac.uk/how_to_read_a_phylogeny

The molecular clock

• The concept of a molecular
clock states that the number of
mutations between two
sequences increases linearly
with evolutionary time
– Zuckerkandl and Pauling found
that the number of amino
acid differences in hemoglobin
correlates with fossil dates
• Sequence space is so large, that
sequences almost never
converge in evolution
• The molecular clock holds in
some cases

3
 2/13/17

Evolutionary time
• The evolutionary divergence between
lineages can be measured by using
evolving characters
– For molecular phylogenies, the unit is
generally: mutations per sequence site

• Using the principle of the molecular clock

in reverse, we can estimate the dates of
evolutionary events (splits in the tree)
– We need to have a few known dates for this
to calibrate the tree
– For example from the fossil record or known
dates of infection

Introduction of cholera in Haiti in 2010

Evolutionary time

Introduction of cholera in Haiti in 2010

4
 2/13/17

Evolutionary distance
• The evolutionary distance between two nodes can be
calculated as the sum of all the horizontal branch lengths
between them
– For example the distance between
virus3 and virus7 is:

= 11 x
0.77 mutations/site

http://epidemic.bio.ed.ac.uk/how_to_read_a_phylogeny

Topology and branch lengths

• Topology (branching order)
quickly shows you the
evolutionary relationships

• Branch lengths show you how

many mutations occurred in
the evolutionary time
between lineages 0.1 substitutions
per sequence site

• As a whole, the phylogenetic

tree quickly shows you the
most distinctive clusters
– …it is not always this clear J

5
 2/13/17

A case story
• In August 1994 a nurse in Lafayette, LA, tests negative for HIV
• A few weeks later, she breaks off a messy 10 year affair with a doctor
• Three weeks later, while suffering from chronic fatigue symptoms, the
doctor gives his ex-mistress a vitamin B-12 shot, somewhat against her will
• In January 1995, the nurse tests positive for both HIV and hepatitis C.
Investigation reveals no obvious means of infection (positive test for a
sexual partner, accident with a patient, et cetera). The vitamin B-12 shot
becomes suspicious
• The doctor’s office records from the day are conveniently missing but
eventually found by police buried in the back of a closet. The records show
that the doctor had withdrawn blood samples from a known HIV patient
and a known hepatitis C patient the same day as the vitamin B-12 shot.
The record keeping is not in line with standard office procedure and there
is no information as to what happened to either blood sample
• The nurse never had contact with either patient
• Seemingly strong, but otherwise circumstantial, evidence that the doctor
deliberately infected the nurse with HIV and hepatitis C

Case story continued

• HIV evolves very fast
– This is partly why it has been so difficult to develop a cure
• Can we show that the HIV in the nurse is related to the
HIV from the patient?
1. Take samples of HIV from the nurse
2. Take samples of HIV from the patient
3. Take samples of HIV from other HIV positive people from the
same town
4. Sequence HIV gene sequences
5. Construct a phylogeny of the HIV

6
 2/13/17

HIV phylogeny

} HIV strains
found in patient

} HIV strains
found in victim

} HIV strains
found in other
individuals from
Lafayette

7
 2/13/17

Branch support
• Support values show you how reliable a branching split is
• Mostly displayed as values or circles
– Often between 0 and 1, or 0-100%
• Branches that are not well supported
might be collapsed 0.01
– This means the topology is unclear
– Bifurcating à multifurcating branch

Support*
100%
Note: this is the same split
50% (in an unrooted tree)
1%
*more about this later
http://epidemic.bio.ed.ac.uk/how_to_read_a_phylogeny

Rotating branches
• In the tree on the left, it looks like bat viruses are all
grouped together, but they are not in one lineage!
• Because the vertical dimension has no meaning, branches
can be freely rotated
• The trees below are identical, one branch is rotated:

8
 2/13/17

Phylogenetic mobile of mammals

Diameter: 3.5 m. Location: lobby of the Broad Institute in

Cambridge, Massachusetts, USA. Design: Peter Agoos.

Different portraits of the same tree

• Because the vertical dimension has no meaning, trees can
be displayed in different ways

Radial display
Terminal nodes
Internal nodes
Root

Polar display
Normal display http://epidemic.bio.ed.ac.uk/how_to_read_a_phylogeny

9
 2/13/17

The time axis is always away from the root

Time

Phylogenies allow us to look back in time

Origin of mammals

Origin of vertebrates

Origin of animals

Origin of eukaryotes

Earliest fossils

Origin of life

10
 2/13/17

Ancestral states
• Can we figure out what animal the very first virus infected?
• We know that evolution tends to be conservative
• We can infer ancestral states by
assuming the fewest possible
changes in the tree
• This is called the
parsimony principle

http://epidemic.bio.ed.ac.uk/how_to_read_a_phylogeny

• Multiple parsimonious solutions means that the ancestral

state is ambiguous

11
 2/13/17

Ancestral states
• Can we figure out what animal the very first virus infected?
• We know that evolution tends to be conservative
• We can infer ancestral states by
assuming the fewest possible
changes in the tree
• This is called the
parsimony principle

http://epidemic.bio.ed.ac.uk/how_to_read_a_phylogeny

Orthology* and paralogy

*Not to be confused with Ornithology

Bas E. Dutilh
Systems Biology: Bioinformatic Data Analysis
Utrecht University, February 14th 2017

12
 2/13/17

Reminder: nodes in a phylogenetic tree

• Terminal nodes: the present day sequences
that were used to create the tree
• Ancestral nodes: ancestors of the present-
day sequences
• Root: Last common ancestor of all the
sequences in the tree

Time axis: away from root

http://epidemic.bio.ed.ac.uk/how_to_read_a_phylogeny

Using phylogenetics to study the evolution of genes

13
 2/13/17

The origin of homologs

• Observed branch in phylogenetic tree:

• So what happened at this node? Present-day genomes

• Homologous genes can result from:

– Speciation: when separate species lineages
diverge from a common ancestor Ancestral genome
• This means that the two homologs and are
Orthologs
present in the genome sequences of different
species Paralogs
• These homologs are called orthologs
– Gene duplication: when a gene is duplicated
within a genome and the duplication becomes Present-day genome
fixed in the population
• This means that two the two homologs and
are present in the same genome sequence
• These homologs are called paralogs
Ancestral genome

The evolution of a gene

• Orthologs are derived from a speciation event

– Orthologs are directly related to same ancestral gene
• Paralogs are derived from a gene duplication event
– Paralogs have evolved together in the same genome for a while

14
 2/13/17

Question
Mouse A Mouse C
Human A Mouse D
Mouse B Human C
Tree 1
Human B Human D
Tree 2

• Observe the two simplified gene trees above of two

homologs from mouse and two homologs from human.
1. Which are the speciation nodes and which are the gene
duplication nodes?
2. What kind of homologs are Mouse A and Human A in Tree 1?
3. What kind of homologs are Human C and Human D in Tree 2?
4. What kind of homologs are Mouse D and Human D in Tree 2?
5. Which genes do you think may have the same function in Tree 1?
6. Which genes do you think may have the same function in Tree 2?

How to tell the difference?

• Rule of thumb:
– If the two daughter
branches of a node
contain the same
species, it may be a
gene duplication node
– Otherwise it may be a
speciation node

• There are exceptions

where the rule of
thumb does not hold
– Horizontal gene
transfer (HGT)
– Unrecognized paralogy

15
 2/13/17

Evolution of function

• Orthologs are directly related to same ancestral gene

à Likely perform same ancestral function (evolution is
conservative)
• Paralogs evolved together in the same genome for a while
àFunction might diverge (unlikely that two genes have the exact
same function in one genome)

Gene function prediction

• Transfer of functional annotations between genes is only
reliable between orthologs
• Function is more likely to diverge between paralogs

Annotation
transfer OK

Annotation
transfer not OK

Annotation
transfer OK

16
 2/13/17

Orthology & paralogy are evolutionary concepts

Annotation
transfer OK

• Researchers are often trying to

identify orthologs in model
organisms, with the goal of
transferring functional
annotation
• However, note that orthology
and paralogy are originally
evolutionary definitions that
say nothing about function

Species trees and gene trees

• The phylogenetic tree of a gene family can be much more

complex than the species tree
• It can be challenging to reconcile the gene tree with the
species tree
• It helps if you have prior knowledge of the species tree

17
 2/13/17

Species tree reconciliation

• Using species tree reconciliation
we can deduce where gene
losses occurred

Gene loss

Gene losses Loss in the ancestor of Medaka and Trout

• Species tree reconciliation

allows us to answer questions
like:
– How many copies of this gene
did the last common ancestor
of all fishes have?
– How many copies of this gene
did the last common ancestor
of all mammals have?

Gene loss

18
 2/13/17

Horizontal gene transfer (HGT)

• Sometimes, you need to Mitochondrial branch

assume many losses to

explain a gene tree
• It may be more
parsimonious to assume
just one HGT event

Wolbachia branch

• … or just one
contaminated sample L

Unrecognized paralogy
• Another mechanism that causes of conflict between the
phylogenetic tree of a gene and the species tree is
unrecognized paralogy
species A
ancestor

Gene invention species B

Speciation node (orthologs)
Gene duplication node (paralogs) species C
Gene loss
species A

species B …with all these

processes going on
in thousands of
species C genes evolution can
get very complex!

19
 2/13/17

Phylogenetic inconsistencies
• The phylogenies of different genes from the same genomes
can be inconsistent

• This can be the result of:

– Evolution of the gene is different than the evolution of the genome
• Horizontal gene transfer
• Unrecognized paralogy
– Technical issues
• Bad model of evolution
• Bad alignments
• Bad phylogenies
– Biological noise
• Mutational saturation: multiple mutations at the same sequence site
• Different rates of evolution in different lineages (inconsistent molecular clock)

The “Tree” of Life?

• Due to evolutionary processes
like HGT, gene loss,
endosymbiosis
(mitochondrion, chloroplast)
relationships might better be
represented as a network
than as a tree of life

20