SYSTEMATICS BASED ON EVOLUTIONARY

GENERAL BIOLOGY 2 RELATIONSHIP

THE CLADISTIC SYSTEM

AND
NEW TRENDS: MOLECULAR PHYLOGENETICS

Learning Competency
The learner describes the species diversity and cladistics including the types of evidence and procedures that
can be used to establish evolutionary relationship.
(STEM_BIO11/12-IIIh-j-16)

Specific Learning Objectives

 Familiarize with different activities or practices (naming, describing, identifying, classifying) used in
taxonomy.
 Distinguish distinct characteristics of a taxa in relation to other groups.
 Describe species diversity and the procedure of the cladistics system that is used to establish
evolutionary histories.
 Explain how modern systematics uses DNA sequences data to answer the perplexing concepts of
speciation and evolution.

The Cladistic System

Cladistics or phylogenetic systematics is a method that groups

organisms based on their shared derived characters. It is said that only
shared derived characters can provide information on the phylogeny of the
organism. This is an important work published in 1966 by Willi Hennig
(Figure 1.1). The grouping mainly focuses on taxa that share many
derived characters as opposed to those that do not have shared derived
characters.

Relationships are represented as a branching hierarchical tree

called a cladogram. The cladogram is constructed such that the number of
changes from one-character state to the next is minimized (Lipscomb,
1998). The rule of parsimony applies to this any hypothesis that requires
fewer assumptions is a more defensible hypothesis. The cladistic Figure 1.1 Photo of German
system is an objective and quantitative method of modern classification, zoologist Willi Hennig, made
and it is to its tenets that the vast majority of modern systematists by his son Gerd Henning
observe.

In the cladistic system, organisms are classified

Figure 1.2 Example of a cladogram
exclusively on the basis of how recent are the
descendants from their common ancestor. Therefore,
always remember that the synapomorphies is the main
basis for the determination of taxonomic/phylogenetic
relationships. The presence of many shared derived
characters among taxa is considered strong evidence of
their common descent. Differences in the degree or rate
of change among branches of organisms are irrelevant to
their classification (Krempels and Lee, 2013 p. 10).

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Constructing the Cladogram

In order to construct phylogenies that show evolutionary relationships, systematists consider the
synapomorphies and symplesiomorphies as well as the homoplasies (Figure 1.2). This requires good
analytical skills to clearly show their evolutionary relationships. Phylogenies can be represented as tree-like
diagrams called phylogenetic or evolutionary tree showing how various taxa branched out from their
ancestors and from each other (Figure 1.3) (Krempels and Lee, 2013)

Initially, character states need to be determined as primitive and/or derived. Some of the procedure to
classify these characters was proposed by Hennig (1966) and others, but the outgroup comparison method is
the primary one being used today. In outgroup comparison, if a taxon that is not a member of the group of
organisms being classified has a character state that is the same as some of the organisms in the group then
that character state can be considered to be plesiomorphic. The outside taxon is called the outgroup and the
organisms being classified are the ingroup (Lipscomb1998).

The two lineages branching from the same ancestor arose at the same geological time. Many people
hold the misconception that Homo sapiens is the most highly evolved" species, or even the most recently
evolved. Neither is true. Always remember the following rules (Krempels and Lee, 2013):

Rule 1: The branches at every node can be rotated. The branches do not infer any sort of order; they indicate
only how recent are the common descenaans

Rule 2: Two lineages branching from a single ancestral node are known as sister taxa. Further specialization
after a branch point is inappropriate. Therefore, it would be improper to say that humans evolved more
recently than chimpanzees, or that humans should be placed in their own family simply because they seem so
different from chimpanzees. Note that common ancestry is the main basis of taxonomic groupings and not
based on subjective perceptions of specialization.

Rule 3: There is no such thing as a "most highly evolved species." All extant species descended from
successful ancestors, and evolved to survive and reproduce in the context of their specific environment.
Evolution is a process. It has neither a goal nor a subjective value system.

Rule 4. No extant or extinct taxon is considered ancestral to any other extant or extinct taxon. Nodes
represent hypothetical ancestors, not taxonomic units. When an ancestral lineage diverges to become two
separate taxa, the ancestral lineage (hypothetical ancestor) is considered extinct, even if one of the descendant
taxa is (or might be) virtually the same as that hypothetical ancestor. This should be remembered when one
hears the oft-repeated, but incorrect statement that "humans evolved from monkeys" They did not. Humans
and monkeys share a common ancestor

Grouping Taxa Based on Shared Apomorphies

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Using apomorphies, taxa can be grouped in two ways: (1) Hennig argumentation, which was discussed in
detail in Hennig (1966); and (2) Wagner method, which uses an algorithm to search for the tree.

a. The Hennig Tree. Hennig argumentation uses characters one at a time. Use the following
information below as an example (Lipscomb, 1998):

Using the dataset, we can infer the following:

This is the first stage of analyzing the relationships of the taxa. Keep in mind that you have to include and
consider all the characters in the cladogram. This is a relatively easy example by real sets of character data. A
more realistic situation would be having characters that exhibit conflicting relationships among the taxa. If
this happens, then choose the tree that calls for the least assumptions of the changes in the character states
(Lipscomb, 1998). Let us look at the next example and try to construct a cladogram using Hennig
argumentation:

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b. The Wagner Tree. In the Wagner method, taxa is connected one at a time until all the taxa is
included in the tree. When added, each taxon is joined to the tree to minimize the number of character
state changes (Lipscomb, 1998).

THE CAMINALCULES

Here are the characters: (Adapted from Krempels and Lee, 2013)

Character a: "eyes present" (+) versus "eyes absent" (-)

Character b:"body mantle present" (+) versus "body mantle absent" (-)

Character c:"paired, anterior non-jointed appendages present" (+) versus "paired, anterior non-jointed
appendages not present" (-)

Character d: “anterior appendages flipperlike" (+) versus "anterior appendages not flipperlike”(-)

Character e: “eyes stalked” (+) versus “ eyes not stalked” (-)

Character f: “body mantle posterior bulbous”(+) versus “ body mantle posterior not bulbous” (-)

Character g; “eyes fused into one”(+) versus “eyes separate” (-)

Character h: “ forelimbs with digits” (+) versus “forelimbs without digits” (-)

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Rooted vs. Unrooted Trees

Trees can be rooted or unrooted, scaled or unscaled or combination of both (Figure 1.3). A rooted tree
is used when each of the node represents the most recent common ancestor of the taxa branching from it.
Rooted trees are directional, which means that a single common ancestor is present at the root ad all taxa
evolve or radiate from that single common ancestor. In order to choose an outgroup. you have to identify a
taxonomic unit that is closely related to but phylogenetically outside the group of taxa to root a tree.

An unrooted tree is used when there is no hypothetical ancestor, outgroup, or directionality to the tree
and only the putative evolutionary relationships of the taxa on the tree is present in the tree (Krempels and
Lee, 2013 p.12).

Figure 1.3 Types of

phylogenetic trees

Monophyly, Paraphyly, and Polyphyly

A phylogenetic tree is not constructed randomly. A systematist uses different characters to determine
recency of common descent. One of the tasks of a systematist is to convert the tree into the formal
hierarchical Linnean classification by giving a formal taxonomic name to groups that share a common
ancestor. Such groups are called monophyletic taxa and they are recognized because they share unique
derived characters. The tree shows several sets of most closely related taxa that are nested within larger sets.
Because Linnean categories are also internested, these sets of taxa can be converted into Linnean categories
(Lipscomb, 1998, p. 42).

Since the classification system was created due to the absence of a clear guideline, some biologists
(called lumpers) focus on similarities in organisms there by delimiting several species into a single genus.
Other biologists (called splitters) focus on the differences between the species, thus, dividing the species into
several different genera (Lipscomb, 1998 p. 43-44). Systematists can change any classification schemes that
were presented before because of the following reasons based on Lipscomb

(1998):

a. New data - Input from new technologies provides new information in the similarities and differences
among taxa that leads to revision, lumping, or splitting a taxon.

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b. New taxa - discovery of new species with new or conflicting characters leads to the revision of
classification.

c.Misinterpreted data - new monophyletic taxon is created when new studies revealed that the characters
used to group a certain taxon are not unique or are convergent with other characters. Forming new
monophyletic taxon occurs in two instances: it is considered polyphyly when new studies revealed that the
features. The used for the grouping is a convergent characters, the taxon is called polyphyleticphyloge (no
recent common ancestor and were grouped on the basis of homoplasy). On the other hand, it is paraphyly
when incorrectly interpreted plesiomorphicas characters were considered as synapomorphies; a paraphyletic
taxon is created tree. The paraphyletic tree shows the traditional, evolutionary system molecule for
classifying reptiles. This does not reflect the actual evolutionary relationship. This classification can be made
monophyletic by considering Aves (birds) in the order a tree. The polyphyletic tree in Figure 1.7 illustrates a
superficial character as a means of classification (e.g, "warm bloodedness" or "four-chambered heart").

These characters are unreliable means of classifying organisms.

New Trends: Molecular Phylogenetics

It has since been determined that evolution is actually a molecular process based on genetic
information, encoded in DNA, RNA, and proteins. One molecule undergoes diversification into many
variations while one or more of those variants can be selected to be reproduced or amplified throughout a
population over many generations(Dowell, 2008). Indeed, the onset of DNA technology helped systematists
to establish relationships between species and unknown species using genetic structures (Diversity of Lire,
n.a.). As computers become more powerful and more generally accessible, and computer algorithms are
becoming more sophisticated, researchers are able to tackle the immensely complicated stochastic and
probabilistic algorithms. The main goal of molecular phylogenetics is to recover the order of evolutionary
events and place them in evolutionary trees (Dowell, 2008).

These evolutionary events are shaped by homology and represented in pnyogenetic trees as homologous
relationships. Homologous relationship can occur as: (1) paralogs, when homologous sequences become
separated by gene duplication; or as (2) orthologs, when separated by a speciation event. A molecular
phylogenetic tree is derived from biomolecular sequence alignments of DNA, RNA, Or amino acids,
molecular markers, such as single nucleotide polymorphisms (SNPs) or restriction fragment length
polymorphisms (RFLPs), morphology data, or information on gene order and content (Dowell, 2008).

According to Dowell (2008, p. 5-7), the process of creating a molecular

phylogenetic tree is as follows:

1. DNA Isolation. A sample is taken from the specimen to be analyzed. The sample is homogenized and
then subjected to different chemicals that cause the cells to release their contents. Everything is
removed except the long DNA spirals. There are lots of manufacturers that provide isolation kits
which aid for simpler, robust, and faster isolation of the DNA.
2. Amplification of the DNA. The DNA that was isolated is then amplified using the polymerase chain
reaction. This technology is actually about the principle that you can exponentially multiply a single
copy of your DNA. You can actually compare it to a Xerox machine wherein you can replicate a
number of many copies. With the right combination of primer pairs to use, right amount of buffer
solutions, bivalent cations (usually Mg ), and polymerase, the whole method now relies on thermal
cycling, wherein typically PCR consists of a series of 20-40 repeated temperature changes, called
cycles, with each cycle commonly consisting of 2-3 discrete temperature steps. After the complete
PCRcycles, PCR products are purified using commercialized purifications kits and they are ready to
be sequenced (Boundless, 2016).

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3.DNA Sequencing. Now is the right time to know the actual and precise orders of nucleotides (A, T, C, and
G) within the DNA molecule. There were a lot of different sequencing methods that were utilized in the past
decades (i.e., Maxam-Gilbert sequencing that requires radioactive labeling or the more commonly used
Sanger sequencing) (Figure 1.4). But nowadays, there are more robust and high throughput type of next
generation sequencing (NGS) available in the market which gives millions of DNA coming from different
species extracted in just a single environmental samples (Pettersson et al., 2009).

Figure 1.4 An example of the result

generated from sanger sequencing which
uses the chain termination protocol and
fluorescent dyes to detect the actual
sequences of DNA

4. Dataset assembly. The real work begins after getting the whole actual DNA sequence. Initially, you must
identify the DNA sequence of interest and assemble dataset consisting of other related sequences. There are a
number of free, online tools available to simplify and streamline this process. DNA sequence of interest can
be retrieved using NCBI BLAST or similar search tools. When evaluating a setoff related sequences retrieved
in a BLAST search, you must pay close attention to the score and E-value. A high score indicates the subject
sequence retrieved with closely related to the sequence used to initiate the query. The smaller the E-value, the
higher the probability that the homology reflects a true evolutionary relationship as opposed to sequence
similarity due to chance. As a general rule, sequences with E-values less than 10-5 are homologs of a query
sequence (NCBI,2016).

5. Sequence alignment. After you selected and retrieved, multiple sequence alignment is done (Figure 1.5).
This is actually arranging a set of sequences in a matrix to identify regions where they are the same
(homologous). Typically, gaps (one or more spaces in an alignment file) must be introduced in several
sequences to represent either insertions or deletions in the molecular code that may have happened over
generations. There are many websites and software programs, such as ClustalW, MSA, MAFFT, and T-
Coffee designed to perform multiple sequences on a given set of molecular data. ClustalW is currently the
most mature and most widely used (Dowell, 2008, p. 5).

Figure 1.5 An example of an aligned data matrix of aligned

sequences
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6.Phylogenetic Tree building. To construct a phylogenetic tree (Figure 1.6),statistical methods are applied
to determine the tree topology and calculate the branch lengths that best describe the phylogenetic
relationships of the aligned sequences in a dataset. Many different methods for building trees exist and no
single method performs well for all types of trees and datasets. The most common computational methods
applied include distance-matrix methods, and discrete data methods, such as maximum parsimony and
maximum likelihood. There are several software packages, such as Paup", PAML, PHYLIP, that apply most
popular methods (Dowell, 2008, p. 6).

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References and Further Readingsi

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5aldaut, Sandra L. "Phylogeny for the faint of heart: a tutorlal" TRENDS In Genetics vol

19, no. 6, 2003, 345-351.Phylogenetic Tree building. To construct a phylogenetic tree (Figure 5.31),6

statistical methods are applied to determine the tree topology and calculate thebranch lengths that best
describe the phylogenetic relationships of the alignedsequences in a dataset. Many different methods for
building trees exist and nosingle method performs well for all types of trees and datasets. The most
commoncomputational methods applied include distance-matrix methods, and discretedata methods, such as
maximum parsimony and maximum likelihood. There areseveral software packages, such as Paup", PAML,
PHYLIP, that apply most popularmethods (Dowell, 2008, p. 6)

Bank, Claudia, Reinhard Bürger, and Joachim Hermisson. "The imits to parapatric

Speciation: Dobzhansky-Muller incompatibilities in a continent-island model"Genetics vol. 191, no. 3, 2012,

845-863.

Bickford, David, David J. Lohman, Navjot S. Sodhi, Peter KL Ng, Rudolf Meier, Kevin

Winker, Krista K. Ingram, and Indraneil Das. "Cryptic species as a window on

diversity and conservation" Trends in ecology & evolution vol. 22, no. 3, 2007, pp.148-155.

Boundless. "Amplifying DNA: The Polymerase Chain Reaction"' Boundless Microbiology

Boundless, 26 May. 2016. Accessed 23 July 2016 from https://goo.gl/g911PS.

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Coyne, Jerry A., and H. Allen Orr. Speciation. Vol. 37. Sunderland, MA: Sinauer Associates,

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Cracraft, Joel. "Speciation and its ontology: the empirical consequences of alternative

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Cronquist, Arthur. "Once again, what is a species?" Symposium on Biosystematics in

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Darwin Charles, R. "On the origin of species by means of natural selection, or thepreservation of favoured
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De Queiroz, Kevin. "Ernst Mayr and the modern concept of species." Proceedings of the

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Computational MethodsAnd Tools For Analyzing Evolutionary Relationships" Math 500, 2008, pp. 18.

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Lipscomb, Diana. "Basics of cladistic analysis."George Washington University, Washington,DC, 1998.

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