15  Phylogenetics and the Tree of Life

15.1 Learning Objectives

By the end of this chapter, you should be able to:

  1. Interpret phylogenetic trees and understand their components and terminology
  2. Describe different types of data used to construct phylogenetic trees
  3. Explain the principles and methods of phylogenetic inference
  4. Compare different tree-building methods and their assumptions
  5. Use phylogenetic trees to test evolutionary hypotheses
  6. Describe major patterns in the tree of life
  7. Apply phylogenetic thinking to problems in medicine, conservation, and biotechnology
  8. Critically evaluate phylogenetic studies and their limitations

15.2 Introduction

Phylogenetics is the study of evolutionary relationships among organisms. By reconstructing the branching patterns of evolution, we can understand how different species are related, trace the history of traits, and make predictions about unknown characteristics. This chapter explores how biologists build and use phylogenetic trees—the family trees of life. Understanding phylogenetics is essential for making sense of biological diversity, studying evolutionary processes, and applying evolutionary knowledge to practical problems from disease tracking to conservation planning.

15.3 Introduction to Phylogenetics

15.3.1 What is a Phylogeny?

  • Phylogeny: Evolutionary history of a group of organisms
  • Phylogenetic tree: Diagram representing evolutionary relationships
  • Cladogram: Tree showing branching order but not evolutionary rates
  • Phylogram: Tree with branch lengths proportional to evolutionary change
  • Chronogram: Tree with branch lengths proportional to time

15.3.2 Historical Background

  • Pre-Darwin: Natural systems of classification (Linnaeus)
  • Post-Darwin: Evolutionary classification (Haeckel’s trees)
  • Modern era: Computational methods, molecular data

15.3.3 Why Study Phylogenetics?

  1. Understand evolutionary relationships
  2. Classify organisms (systematics)
  3. Study evolutionary processes
  4. Trace origin and spread of diseases
  5. Conservation priorities
  6. Discover new drugs (bioprospecting)
  7. Study coevolution

15.4 Tree Terminology and Interpretation

15.4.1 Tree Components

Nodes: Branch points

  • Internal nodes: Common ancestors
  • Terminal nodes (tips): Extant taxa

Branches: Lineages between nodes

Root: Common ancestor of all taxa in tree

Clade (monophyletic group): Ancestor and all descendants

Outgroup: Taxon outside group of interest (rooting reference)

15.4.2 Reading Trees

Key points:

  1. Tips can be rotated around nodes without changing relationships
  2. Only branching order matters, not left-right order
  3. Trees show relative relationships, not absolute time (unless scaled)
  4. Extinct taxa can be included as tips or internal nodes

Common misconceptions:

  • “Lower” vs. “higher” organisms
  • Linear progression
  • “Primitive” vs. “advanced”

15.4.3 Types of Groups

Monophyletic (clade): Common ancestor + all descendants

Paraphyletic: Common ancestor + some descendants

Polyphyletic: Members from different ancestors

Examples:

  • Monophyletic: Mammals, birds
  • Paraphyletic: Reptiles (without birds), fish
  • Polyphyletic: “Warm-blooded animals” (birds + mammals)

15.5 Character Data for Phylogenetics

15.5.1 Morphological Characters

Traditional approach: Before molecular methods

Advantages:

  • Can study fossils
  • Functional information
  • Often cheaper

Disadvantages:

  • Subjectivity
  • Homoplasy (convergent evolution)
  • Limited number of characters

Scoring: Discrete (presence/absence) or continuous

15.5.2 Molecular Characters

Revolutionized phylogenetics since 1960s

DNA sequences:

  • Nuclear: Single-copy genes, multi-copy genes
  • Organellar: Mitochondrial, chloroplast
  • Ribosomal RNA: Highly conserved, useful for deep divergences

Protein sequences: Amino acid alignments

Molecular markers: RFLPs, microsatellites, SNPs

Choice of gene depends on:

  • Evolutionary rate: Fast for recent divergences, slow for deep
  • Function: Housekeeping vs. specialized
  • Size: Enough informative sites

15.5.3 Character Evolution

Homology: Similarity due to common ancestry

  • Orthology: Genes diverged after speciation
  • Paralogy: Genes diverged after duplication

Homoplasy: Similarity not due to common ancestry

  • Convergence: Independent evolution of similar traits
  • Reversal: Return to ancestral state

Synapomorphy: Shared derived character (evidence for clade)

Symplesiomorphy: Shared ancestral character (not informative for grouping)

15.6 Tree-Building Methods

15.6.1 Distance Methods

Principle: Convert sequence data to distance matrix, build tree

Neighbor-Joining (NJ):

  • Fast, works well with many taxa
  • Produces unrooted tree
  • Example: Used in early molecular phylogenetics

UPGMA (Unweighted Pair Group Method with Arithmetic Mean):

  • Assumes constant evolutionary rate (molecular clock)
  • Produces rooted tree
  • Less used now due to clock assumption

15.6.2 Character-Based Methods

Maximum Parsimony (MP):

  • Principle: Minimize number of evolutionary changes
  • Advantages: Intuitive, no model assumptions
  • Disadvantages: Long-branch attraction problem
  • Searches: Exhaustive, branch-and-bound, heuristic

Maximum Likelihood (ML):

  • Principle: Find tree that maximizes probability of observed data given evolutionary model
  • Advantages: Statistical framework, model-based
  • Disadvantages: Computationally intensive
  • Models: Jukes-Cantor, Kimura 2-parameter, more complex

Bayesian Inference:

  • Principle: Find tree with highest posterior probability
  • Advantages: Provides confidence measures (posterior probabilities)
  • Disadvantages: Computationally intensive, priors influence results
  • Software: MrBayes, BEAST

15.6.3 Choosing a Method

Consider:

  • Data type: Morphological vs. molecular
  • Number of taxa: Some methods scale poorly
  • Evolutionary model: Violations affect some methods more
  • Computational resources
  • Goals of study

Best practice: Use multiple methods, compare results

15.7 Phylogenetic Trees

15.7.1 Support Measures

Bootstrap (Felsenstein, 1985):

  • Method: Resample data with replacement, build many trees
  • Interpretation: % of trees containing clade
  • Thresholds: >70% moderate, >95% strong support

Jackknife: Similar but resamples without replacement

Posterior probability (Bayesian): Probability clade is true

Decay index (Bremer support): How many steps until clade collapses

15.7.2 Tree Comparisons

Robustness: Similar results from different methods/data

Congruence: Agreement between different genes/traits

Conflict: May indicate different evolutionary histories (e.g., hybridization, horizontal gene transfer)

15.7.3 Sources of Error

Long-branch attraction: Fast-evolving lineages grouped artificially

Inadequate sampling: Missing taxa, characters

Model misspecification: Wrong evolutionary model

Homoplasy: Convergent evolution misleads

15.8 Molecular Clock and Dating

15.8.1 Concept

Molecular clock hypothesis: Rate of molecular evolution relatively constant over time

First proposed: Zuckerkandl and Pauling (1962)

Evidence: Roughly linear relationship between genetic divergence and time

15.8.2 Calibration

Using fossils: Date of divergence from fossil record

  • Challenges: Fossil may not be earliest, dating uncertainty
  • Best practices: Multiple calibrations, account for uncertainty

Geological events: Vicariance events (e.g., Isthmus of Panama)

Known historical events: Human migrations, domestication events

15.8.3 Relaxed Molecular Clocks

Allow rate variation among lineages

Bayesian methods: Incorporate rate variation in model

Divergence time estimation: BEAST, MCMCTree

15.8.4 Applications

Dating evolutionary events: Origin of groups, biogeographic patterns

Evolutionary rates: Compare rates among lineages, genes

Ancestral sequence reconstruction

15.9 Applications of Phylogenetics

15.9.1 Classification and Systematics

Phylogenetic classification: Groups should be monophyletic

Linnaean system vs. phylogenetic: Rankless vs. ranked classifications

PhyloCode: Proposed alternative emphasizing clades

15.9.2 Comparative Biology

Independent contrasts: Account for phylogenetic non-independence

Ancestral state reconstruction: Infer traits of ancestors

Correlated evolution: Test if traits evolve together

15.9.3 Medicine and Public Health

Pathogen phylogenetics:

  • Outbreak tracing: HIV, SARS-CoV-2, influenza
  • Drug resistance: Evolution of resistance genes
  • Vaccine design: Antigenic evolution

Evolutionary medicine: Phylogenetic perspective on disease

15.9.4 Conservation Biology

Evolutionary distinctiveness: Phylogenetic diversity measures

Conservation prioritization: Protect phylogenetically distinct species

Identifying units: Evolutionary Significant Units (ESUs)

15.9.5 Biotechnology

Gene discovery: Find homologs in related species

Protein engineering: Ancestral sequence reconstruction

Bioprospecting: Target related species for drug discovery

15.9.6 Forensics and Agriculture

Food authentication: Identify species in products

Pest management: Understand pest evolution and relationships

Crop improvement: Wild relatives as genetic resources

15.10 The Tree of Life

15.10.1 Three Domains

Woese et al. (1990): Based on ribosomal RNA

  1. Bacteria: Prokaryotes, diverse metabolisms
  2. Archaea: Prokaryotes, many extremophiles
  3. Eukarya: Eukaryotes

Relationships: Archaea and Eukarya more closely related

15.10.2 Major Eukaryotic Groups

Opisthokonta: Animals, fungi, choanoflagellates

Archaeplastida: Plants, green algae, red algae

SAR clade: Stramenopiles, Alveolates, Rhizarians

Excavata: Euglenozoans, diplomonads

Uncertainties: Root of eukaryotic tree debated

15.10.3 Relationships Within Animals

Major phyla: ∼35 recognized

Deuterostomes: Echinoderms, chordates

Protostomes:

  • Ecdysozoa: Arthropods, nematodes (molting)
  • Lophotrochozoa: Mollusks, annelids, flatworms

15.10.4 Plant Phylogeny

Green plants: Chlorophytes, charophytes, land plants

Land plants: Bryophytes, vascular plants

Seed plants: Gymnosperms, angiosperms

Angiosperms: Monocots, eudicots, magnoliids

15.10.5 Ongoing Challenges

Horizontal gene transfer: Complicates prokaryotic trees

Incomplete lineage sorting: Gene trees ≠ species trees

Hybridization: Common in plants, some animals

Sampling: Many groups poorly studied

15.11 Phylogenetic Software and Databases

15.11.2 Databases

Sequence: GenBank, EMBL, DDBJ

Alignment: PFAM, SMART

Trees: TreeBASE, Open Tree of Life

Taxonomy: NCBI Taxonomy, Catalogue of Life

15.11.3 Analysis Pipeline

  1. Sequence acquisition
  2. Alignment
  3. Model selection (jModelTest, ModelTest)
  4. Tree building
  5. Support assessment
  6. Visualization and interpretation

15.12 Chapter Summary

15.12.1 Key Concepts

  1. Phylogenetic trees: Diagram evolutionary relationships
  2. Tree interpretation: Understand components and avoid misconceptions
  3. Character data: Morphological and molecular data used for inference
  4. Tree-building methods: Distance, parsimony, likelihood, Bayesian
  5. Tree assessment: Bootstrap, posterior probabilities, other support measures
  6. Molecular clock: Dating evolutionary events
  7. Applications: Classification, medicine, conservation, biotechnology
  8. Tree of life: Three domains, major eukaryotic groups

15.12.2 Tree-Building Method Comparison

Method Principle Advantages Disadvantages
Neighbor-Joining Distance matrix Fast, good for many taxa No explicit model
Maximum Parsimony Minimize changes Intuitive, no model Long-branch attraction
Maximum Likelihood Probability maximization Statistical, model-based Computationally intensive
Bayesian Posterior probability Confidence measures, flexible Computationally intensive, priors

15.12.3 Support Measures

Measure Range Interpretation
Bootstrap 0-100% % of resampled trees with clade
Posterior probability 0-1 Probability clade is true
Decay index ≥0 Extra steps to collapse clade

15.12.4 Major Eukaryotic Clades

Clade Major Groups Characteristics
Opisthokonta Animals, fungi Posterior flagellum (in some)
Archaeplastida Plants, algae Primary plastids
SAR Diatoms, ciliates, forams Diverse, often with secondary plastids
Excavata Euglenids, trypanosomes Feeding groove

15.12.5 Phylogenetic Applications

Field Application Example
Medicine Outbreak tracing COVID-19 origins
Conservation Prioritization EDGE species
Agriculture Pest relationships Resistance gene transfer
Biotechnology Gene discovery Thermostable enzymes from thermophiles

15.13 Review Questions

15.13.1 Level 1: Recall and Understanding

  1. Draw and label the components of a phylogenetic tree.
  2. What is the difference between monophyletic, paraphyletic, and polyphyletic groups?
  3. List three types of data used to build phylogenetic trees.
  4. What are the main phylogenetic tree-building methods?
  5. How is the molecular clock used to date evolutionary events?

15.13.2 Level 2: Application and Analysis

  1. Given the tree (A,(B,C)), which pair is most closely related?
  2. Why might different genes give different phylogenetic trees for the same species?
  3. How would you choose an outgroup for a phylogenetic analysis?
  4. What does a bootstrap value of 85% mean for a particular clade?
  5. Why is long-branch attraction a problem in phylogenetic inference?

15.13.3 Level 3: Synthesis and Evaluation

  1. Evaluate the statement: “The tree of life is actually more like a web due to horizontal gene transfer.”
  2. How has molecular phylogenetics changed our understanding of biological classification?
  3. Why is it important to consider phylogeny in comparative studies?
  4. Design a phylogenetic study to resolve relationships in a problematic group.

15.14 Key Terms

  • Phylogeny: Evolutionary history of a group of organisms
  • Phylogenetic tree: Diagram showing evolutionary relationships
  • Clade: Monophyletic group consisting of an ancestor and all its descendants
  • Monophyletic: Group containing a common ancestor and all descendants
  • Paraphyletic: Group containing a common ancestor but not all descendants
  • Polyphyletic: Group that does not contain the common ancestor of all members
  • Synapomorphy: Shared derived character that indicates common ancestry
  • Homoplasy: Similarity not due to common ancestry (convergence or reversal)
  • Outgroup: Taxon used to root a tree, closely related but outside group of interest
  • Molecular clock: Hypothesis that molecular evolution occurs at roughly constant rate
  • Bootstrap: Method for assessing support for nodes in phylogenetic tree
  • Maximum parsimony: Tree-building method that minimizes evolutionary changes
  • Maximum likelihood: Tree-building method that maximizes probability of observed data
  • Bayesian inference: Tree-building method based on posterior probabilities

15.15 Further Reading

15.15.1 Books

  1. Baum, D. A., & Smith, S. D. (2013). Tree Thinking: An Introduction to Phylogenetic Biology. Roberts & Company.
  2. Felsenstein, J. (2004). Inferring Phylogenies. Sinauer Associates.
  3. Page, R. D. M., & Holmes, E. C. (1998). Molecular Evolution: A Phylogenetic Approach. Blackwell Science.

15.15.2 Scientific Articles

  1. Woese, C. R., Kandler, O., & Wheelis, M. L. (1990). Towards a natural system of organisms: proposal for the domains Archaea, Bacteria, and Eucarya. PNAS, 87(12), 4576-4579.
  2. Felsenstein, J. (1985). Confidence limits on phylogenies: An approach using the bootstrap. Evolution, 39(4), 783-791.
  3. Huelsenbeck, J. P., & Ronquist, F. (2001). MRBAYES: Bayesian inference of phylogenetic trees. Bioinformatics, 17(8), 754-755.

15.15.3 Online Resources

  1. Tree of Life Web Project: http://tolweb.org
  2. Phylogeny.fr: http://www.phylogeny.fr
  3. Interactive Tree of Life: https://itol.embl.de
  4. OneZoom Tree of Life: http://www.onezoom.org

15.16 Quantitative Problems

  1. Tree Space: For n taxa, number of possible unrooted trees = (2n-5)!/(2ⁿ⁻³(n-3)!)
    1. Calculate number of possible unrooted trees for 5 taxa.
    2. For 10 taxa.
    3. Why is exhaustive search impossible for large n?
  2. Sequence Evolution: Two sequences of length 1000 bp differ at 150 positions. Using Jukes-Cantor model: d = -(3/4)ln(1 - (4/3)p)
    1. Calculate uncorrected distance (p).
    2. Calculate Jukes-Cantor corrected distance.
    3. If molecular clock is 1% per million years, estimate divergence time.
  3. Bootstrap Calculation: In 1000 bootstrap replicates, a particular clade appears in 850 trees.
    1. What is the bootstrap support?
    2. If the true probability of the clade is 0.9, what is the probability of getting ≥850/1000?
    3. What does this say about interpreting bootstrap values?

15.17 Case Study: HIV Phylogenetics and Transmission Networks

Background: HIV phylogenetics has been used to trace transmission networks and understand epidemic dynamics.

Questions:

  1. How can phylogenetic trees help identify transmission clusters?
  2. What are the ethical considerations in using phylogenetics for public health?
  3. How has phylogenetics contributed to understanding HIV origins and spread?
  4. What are the limitations of phylogenetic approaches in outbreak investigation?

Data for analysis:

  • HIV evolutionary rate: ∼0.001 substitutions/site/year
  • Within-host diversity: High due to error-prone reverse transcriptase
  • Transmission bottlenecks: Typically 1-3 variants transmitted
  • Forensic use: Admissible evidence in some court cases
  • Privacy concerns: Genetic data can reveal relationships

Next Chapter: Human Evolution