Managing genes in conservation and industry

The fundamentals of population genetics

Many times in the past, we’ve discussed the importance of genetic diversity within populations as a foundation for adaptation and evolution. It includes both adaptive variation (which encompasses genetic variation directly under natural selection), as well as neutral variation (which is predominantly generated and maintained by non-selective forces such as demographic history and genetic drift). This pool of genetic variation acts as the underlying architecture for evolution by natural selection, and is a critically important component for future and ongoing evolution.

This all sounds important from an academic perspective: that population genetics can reveal a significant amount of information about the processes and outcomes of evolution and provide novel insights into concepts that have been around for ages. But how can this information be applied to real scenarios? With the ever-growing availability of massive genetic datasets for an increasing number of species, the sheer volume of information in existence that can be used is monumental.

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You’re perfect, you’re beautiful, you look like a model (species)

What is a ‘model’?

There are quite literally millions of species on Earth, ranging from the smallest of microbes to the largest of mammals. In fact, there are so many that we don’t actually have a good count on the sheer number of species and can only estimate it based on the species we actually know about. Unsurprisingly, then, the number of species vastly outweighs the number of people that research them, especially considering the sheer volumes of different aspects of species, evolution, conservation and their changes we could possibly study.

Species on Earth estimate figure
Some estimations on the number of eukaryotic species (i.e. not including things like bacteria), with the number of known species in blue and the predicted number of total species on Earth in purpleSource: Census of Marine Life.

This is partly where the concept of a ‘model’ comes into it: it’s much easier to pick a particular species to study as a target, and use the information from it to apply to other scenarios. Most people would be familiar with the concept based on medical research: the ‘lab rat’ (or mouse). The common house mouse (Mus musculus) and the brown rat (Rattus norvegicus) are some of the most widely used models for understanding the impact of particular biochemical compounds on physiology and are often used as the testing phase of medical developments before human trials.

So, why are mice used as a ‘model’? What actually constitutes a ‘model’, rather than just a ‘relatively-well-research-species’? Well, there are a number of traits that might make certain species ideal subjects for understanding key concepts in evolution, biology, medicine and ecology. For example, mice are often used in medical research given their (relative) similar genetic, physiological and behavioural characteristics to humans. They’re also relatively short-lived and readily breed, making them ideal to observe the more long-term effects of medical drugs or intergenerational impacts. Other species used as models primarily in medicine include nematodes (Caenorhabditis elegans), pigs (Sus scrofa domesticus), and guinea pigs (Cavia porcellus).

The diversity of models

There are a wide variety and number of different model species, based on the type of research most relevant to them (and how well it can be applied to other species). Even with evolution and conservation-based research, which can often focus on more obscure or cryptic species, there are several key species that have widely been applied as models for our understanding of the evolutionary process. Let’s take a look at a few examples for evolution and conservation.

Drosophila

It would be remiss of me to not mention one of the most significant contributors to our understanding of the genetic underpinning of adaptation and speciation, the humble fruit fly (Drosophila melanogaster, among other species). The ability to rapidly produce new generations (with large numbers of offspring with very short generation time), small fully-sequenced genome, and physiological variation means that observing both phenotypic and genotypic changes over generations due to ‘natural’ (or ‘experimental’) selection are possible. In fact, Drosphilia spp. were key in demonstrating the formation of a new species under laboratory conditions, providing empirical evidence for the process of natural selection leading to speciation (despite some creationist claims that this has never happened).

Drosophila speciation experiment
A simplified summary of the speciation experiment in Drosophila, starting with a single species and resulting in two reproductively isolated species based on mating and food preference. Source: Ilmari Karonen, adapted from here.

Darwin’s finches

The original model of evolution could be argued to be Darwin’s finches, as the formed part of the empirical basis of Charles Darwin’s work on the theory of evolution by natural selection. This is because the different species demonstrate very distinct and obvious changes in morphology related to a particular diet (e.g. the physiological consequences of natural selection), spread across an archipelago in a clear demonstration of a natural experiment. Thus, they remain the original example of adaptive radiation and are fundamental components of the theory of evolution by natural selection. However, surprisingly, Darwin’s finches are somewhat overshadowed in modern research by other species in terms of the amount of available data.

Darwin's finches drawings
Some of Darwin’s early drawings of the morphological differences in Galapagos finch beaks, which lead to the formulation of the theory of evolution by natural selection.

Zebra finches

Even as far as birds go, one species clearly outshines the rest in terms of research. The zebra finch is one of the most highly researched vertebrate species, particularly as a model of song learning and behaviour in birds but also as a genetic model. The full genome of the zebra finch was the second bird to ever be sequenced (the first being a chicken), and remains one of the more detailed and annotated genomes in birds. Because of this, the zebra finch genome is often used as a reference for other studies on the genetics of bird species, especially when trying to understand the function of genetic changes or genes under selection.

Zebra finches.jpg
A pair of (very cute) model zebra finches. Source: Michael Lawton via Smithsonian.com.

 

Fishes

Fish are (perhaps surprisingly) also relatively well research in terms of evolutionary studies, largely due to their ancient origins and highly diverse nature, with many different species across the globe. They also often demonstrate very rapid and strong bouts of divergence, such as the cichlid fish species of African lakes which demonstrate how new species can rapidly form when introduced to new and variable environments. The cichlids have become the poster child of adaptive radiation in fishes much in the same way that Darwin’s finches highlighted this trend in birds. Another group of fish species used as a model for similar aspects of speciation, adaptive divergence and rapid evolutionary change are the three-spine and nine-spine stickleback species, which inhabit a variety of marine, estuarine and freshwater environments. Thus, studies on the genetic changes across these different morphotypes is a key in understanding how adaptation to new environments occur in nature (particularly the relatively common transition into different water types in fishes).

cichlid diversity figure
The sheer diversity of species and form makes African cichlids an ideal model for testing hypotheses and theories about the process of evolution and adaptive radiation. Figure sourced from Brawand et al. (2014) in Nature.

Zebra fish

More similar to the medical context of lab rats is the zebrafish (ironically, zebra themselves are not considered a model species). Zebrafish are often used as models for understanding embryology and the development of the body in early formation given the rapid speed at which embryonic development occurs and the transparent body of embryos (which makes it easier to detect morphological changes during embryogenesis).

Zebrafish embryo
The transparent nature of zebrafish embryos make them ideal for studying the development of organisms in early stages. Source: yourgenome.org.

Using information from model species for non-models

While the relevance of information collected from model species to other non-model species depends on the similarity in traits of the two species, our understanding of broad concepts such as evolutionary process, biochemical pathways and physiological developments have significantly improved due to model species. Applying theories and concepts from better understood organisms to less researched ones allows us to produce better research much faster by cutting out some of the initial investigative work on the underlying processes. Thus, model species remain fundamental to medical advancement and evolutionary theory.

That said, in an ideal world all species would have the same level of research and resources as our model species. In this sense, we must continue to strive to understand and research the diversity of life on Earth, to better understand the world in which we live. Full genomes are progressively being sequenced for more and more species, and there are a number of excellent projects that are aiming to sequence at least one genome for all species of different taxonomic groups (e.g. birds, bats, fish). As the data improves for our non-model species, our understanding of evolution, conservation management and medical research will similarly improve.

Lost in a forest of (gene) trees

Using genetics to understand species history

The idea of using the genetic sequences of living organisms to understand the evolutionary history of species is a concept much repeated on The G-CAT. And it’s a fundamental one in phylogenetics, taxonomy and evolutionary biology. Often, we try to analyse the genetic differences between individuals, populations and species in a tree-like manner, with close tips being similar and more distantly separated branches being more divergent. However, this runs on one very key assumption; that the patterns we observe in our study genes matches the overall patterns of species evolution. But this isn’t always true, and before we can delve into that we have to understand the difference between a ‘gene tree’ and a ‘species tree’.

A gene tree or a species tree?

Our typical view of a phylogenetic tree is actually one of a ‘gene tree’, where we analyse how a particular gene (or set of genes) have changed over time between different individuals (within and across populations or species) based on our understanding of mutation and common ancestry.

However, a phylogenetic tree based on a single gene only demonstrates the history of that gene. What we assume in most cases is that the history of that gene matches the history of the species: that branches in the genetic tree mirror when different splits in species occurred throughout history.

The easiest way to conceptualise gene trees and species trees is to think of individual gene trees that are nested within an overarching species tree. In this sense, individual gene trees can vary from one another (substantially, even) but by looking at the overall trends of many genes we can see how the genome of the species have changed over time.

Gene tree incongruence figure
A (potentially familiar) depiction of individual gene trees (coloured lines) within the broader species tree (defined b the black boundaries). As you might be able to tell, the branching patterns of the different genes are not the same, and don’t always match the overarching species tree.

Gene tree incongruence

Different genes may have different patterns for a number of reasons. Changes in the genetic sequences of organisms over time don’t happen equally across the entire genome, and very specific parts of the genome can evolve in entirely different directions, or at entirely different rates, than the rest of the genome. Let’s take a look at a few ways we could have conflicting gene trees in our studies.

Incomplete lineage sorting

One of the most prolific, but more complicated, ways gene trees can vary from their overarching species tree is due to what we call ‘incomplete lineage sorting’. This is based on the idea that species and the genes that define them are constantly evolving over time, and that because of this different genes are at different stages of divergence between population and species. If we imagine a set of three related populations which have all descended from a single ancestral population, we can start to see how incomplete lineage sorting could occur. Our ancestral population likely has some genetic diversity, containing multiple alleles of the same locus. In a true phylogenetic tree, we would expect these different alleles to ‘sort’ into the different descendent populations, such that one population might have one of the alleles, a second the other, and so on, without them sharing the different alleles between them.

If this separation into new populations has been recent, or if gene flow has occurred between the populations since this event, then we might find that each descendent population has a mixture of the different alleles, and that not enough time has passed to clearly separate the populations. For this to occur, sufficient time for new mutations to occur and genetic drift to push different populations to differently frequent alleles needs to happen: if this is too recent, then it can be hard to accurately distinguish between populations. This can be difficult to interpret (see below figure for a visualisation of this), but there’s a great description of incomplete lineage sorting here.

ILS_adaptedfigure
A demonstration of incomplete lineage sorting, generously adapted from a talk by fellow MELFU postdocs Dr Yuma (Jonathon) Sandoval-Castillo and Dr Catherine Attard. On the left is a depiction of a single gene coalescent tree over time: circles represent a single individual at a particular point in time (row) with the colours representing different alleles of that same gene. The tree shows how new mutations occur (colour changes along the branches) and spread throughout the descendent populations. In this example, we have three recently separated species, with a good number of different alleles. However, when we study these alleles in tree form (the phylogeny on the right), we see that the branches themselves don’t correlate well with the boundaries of the species. For example, the teal allele found within Species C is actually more similar to Species B alleles (purple and blue) than any other Species B alleles, based on the order and patterns of these mutations.

Hybridisation and horizontal transfer

Another way individual genes may become incongruent with other genes is through another phenomenon we’ve discussed before: hybridisation (or more specifically, introgression). When two individuals from different species breed together to form a ‘hybrid’, they join together what was once two separate gene pools. Thus, the hybrid offspring has (if it’s a first generation hybrid, anyway) 50% of genes from Species A and 50% of genes from Species B. In terms of our phylogenetic analysis, if we picked one gene randomly from the hybrid, we have 50% of picking a gene that reflects the evolutionary history of Species A, and 50% chance of picking a gene that reflects the evolutionary history of Species B. This would change how our outputs look significantly: if we pick a Species A gene, our ‘hybrid’ will look (genetically) very, very similar to Species A. If we pick a Species B gene, our ‘hybrid’ will look like a Species B individual instead. Naturally, this can really stuff up our interpretations of species boundaries, distributions and identities.

Hybridisation_figure
An example of hybridisation leading to gene tree incongruence with our favourite colourful fishA) We have a hybridisation event between a red fish (Species A) and a green fish (Species B), resulting in a hybrid species (‘Species’ H). The red fish genome is indicated by the yellow DNA, the green fish genomes by the blue DNA, and the hybrid orange fish has a mixture of these two. B) If we sampled one set of genes in the hybrid, we might select a gene that originated from the red fish, showing that the hybrid is identical (or very similar) the Species A. D) Conversely, if we sampled a gene originating from the green fish, the resultant phylogeny might show that the hybrid is the same as Species B. C) If we consider these two patterns in combination, which see the true pattern of species formation, which is not a clear dichotomous tree and rather a mixture of the two sets of trees.

Paralogous genes

More confusingly, we can even have events where a single gene duplicates within a genome. This is relatively rare, although it can have huge effects: for example, salmon have massive genomes as the entire thing was duplicated! Each version of the gene can take on very different forms, functions, and evolve in entirely different ways. We call these duplicated variants paralogous genes: genes that look the same (in terms of sequence), but are totally different genes.

This can have a profound impact as paralogous genes are difficult to detect: if there has been a gene duplication early in the evolutionary history of our phylogenetic tree, then many (or all) of our study samples will have two copies of said gene. Since they look similar in sequence, there’s all possibility that we pick Variant 1 in some species and Variant 2 in other species. Being unable to tell them apart, we can have some very weird and abstract results within our tree. Most importantly, different samples with the same duplicated variant will seem similar to one another (e.g. have evolved from a common ancestor more recently) than it will to any sample of the other variant (even if they came from the exact same species)!

Paralogy_figure.jpg
An example of how paralogous genes can confound species tree. We start with a single (purple) gene: at a particular point in time, this gene duplicates into a red and a blue form. Each of these genes then evolve and spread into four separate descendent species (A, B, C and D) but not in entirely the same way. However, since both the red and blue genetic sequences are similar, if we took a single gene from each species we might (somewhat randomly) sequence either the red or the blue copy. The different phylogenetic trees on the right demonstrate how different combinations of red and blue genes give very different patterns, since all blue copies will be more related to other blue genes than to the red gene of the same species. E.g. a blue A and a blue C are more similar than a blue A and a red A.

Overcoming incongruence with genomics

Although a tricky conundrum in phylogenetics and evolutionary genetics broadly, gene tree incongruence can largely be overcome with using more loci. As the random changes of any one locus has a smaller effect of the larger total set of loci, the general and broad patterns of evolutionary history can become clearer. Indeed, understanding how many loci are affected by what kind of process can itself become informative: large numbers of introgressed loci can indicate whether hybridisation was recent, strong, or biased towards one species over another, for example. As with many things, the genomic era appears poised to address the many analytical issues and complexities of working with genetic data.

 

Moving right along: dispersal and population structure

The impact of species traits on evolution

Although we often focus on the genetic traits of species in molecular ecology studies, the physiological (or phenotypic) traits are equally as important in shaping their evolution. These different traits are not only the result themselves of evolutionary forces but may further drive and shape evolution into the future by changing how an organism interacts with the environment.

There are a massive number of potential traits we could focus on, each of which could have a large number of different (and interacting) impacts on evolution. One that is often considered, and highly relevant for genetic studies, is the influence of dispersal capability.

Dispersal

Dispersal is essentially the process of an organism migrating to a new habitat, to the point of the two being used almost interchangeably. Often, however, we regard dispersal as a migration event that actually has genetic consequences; particularly, if new populations are formed or if organisms move from one population to another. This can differ from straight migration in that animals that migrate might not necessarily breed (and thus pass on genes) into a new region during their migration; thus, evidence of those organisms will not genetically proliferate into the future through offspring.

Naturally, the ability of organisms to disperse is highly variable across the tree of life and reliant on a number of other physiological factors. Marine mammals, for example, can disperse extremely far throughout their lifetimes, whereas some very localised species like some insects may not move very far within their lifetime at all. The movement of organisms directly facilitates the movement of genetic material, and thus has significant impacts on the evolution and genetic diversity of species and populations.

Dispersal vs pop structure
The (simplistic) relationship between dispersal capability and one aspect of population genetics, population structure (measured as Fst). As organisms are more capable of dispersing longer distance (or more frequently), the barriers between populations become weaker.

Highly dispersive species

At one end of the dispersal spectrum, we have highly dispersive species. These can move extremely long distances and thus mix genetic material from a wide range of habitats and places into one mostly-cohesive population. Because of this, highly dispersive species often have strong colonising abilities and can migrate into a range of different habitats by tolerating a wide range of conditions. For example, a single whale might hang around Antarctica for part of the year but move to the tropics during other times. Thus, this single whale must be able to tolerate both ends of the temperature spectrum.

As these individuals occupy large ranges, localised impacts are unlikely to critically affect their full distribution. Individual organisms that are occupying an unpleasant space can easily move to a more favourable habitat (provided that one exists). Furthermore, with a large population (which is more likely with highly dispersive species), genetic drift is substantially weaker and natural selection (generally) has a higher amount of genetic diversity to work with. This is, of course, assuming that dispersal leads to a large overall population, which might not be the case for species that are critically endangered (such as the cheetah).

Highly dispersive animals often fit the “island model” of Wright, where individual subpopulations all have equal proportions of migrants from all other subpopulations. In reality, this is rare (or unreasonable) due to environmental or physiological limitations of species; distance, for example, is not implicitly factored into the basic island model.

Island model
The Wright island model of population structure. In this example, different independent populations are labelled in the bold letters, with dispersal pathways demonstrated by the different arrows. In the island model, dispersal is equally likely between all populations (including from BD in this example, even though there aren’t any arrows showing it). Naturally, this is not overly realistic and so the island model is used mostly as a neutral, base model.

Intermediately dispersing species

A large number of species, however, are likely to occupy a more intermediate range of dispersal ability. These species might be able to migrate to neighbouring populations, or across a large proportion of their geographic range, but individuals from one end of the range are still somewhat isolated from individuals at the other end.

This often leads to some effect of population structure; different portions of the geographic range are genetically segregated from one another depending on how much gene flow (i.e. dispersal) occurs between populations. In the most simplest scenario, this can lead to what we call isolation-by-distance. Rather than forming totally independent populations, gene flow occurs across short ranges between adjacent ‘populations’. This causes a gradient of genetic differentiation, with one end of the range being clearly genetically different to the other end, with a gradual slope throughout the range. We see this often in marine invertebrates, for example, which might have somewhat localised dispersal but still occupy a large range by following oceanographic currents.

River IDB network
An example of how an isolation-by-distance population network might come about. In this example, we have a series of populations (the different pie charts) spread throughout a river system (that blue thing). The different pie charts represent how much of the genetics of that population matches one end of the river: either the blue end (left) or red end (right). Populations can easily disperse into adjacent populations (the green arrows) but less so to further populations. This leads to gradual changes across the length of the river, with the far ends of the river clearly genetically distinct from the opposite end but relatively similar to neighbouring populations.
River IDB pop structure.jpg
The genetic representation of the above isolation-by-distance example. Each column represents a single population (in the previous figure, a pie chart), with the different colours also representing the relative genetic identity of that population. As you can see, moving from Population 1 to 10 leads to a gradient (decreasing) in blue genes but increase in red genes. The inverse can be said moving in the opposite direction. That said, comparing Population 1 and Population 10 shows that they’re clearly different, although there is no clear cut-off point across the range of other populations.

Medium dispersal capabilities are also often a requirement for forming ‘metapopulations’. In this population arrangement, several semi-independent populations are present within the geographic range of the species. Each of these are subject to their own local environmental pressures and demographic dynamics, and because of this may go locally extinct at any given time. However, dispersal connections between many of these populations leads to recolonization and gene flow patterns, allowing for extinction-dispersal dynamics to sustain the overall metapopulation. Generally, this would require greater levels of dispersal than those typically found within metapopulation species, as individuals must traverse uninhabitable regions relatively frequently to recolonise locally extinct habitat.

Metapopulation structure.jpg
An example of metapopulation dynamics. Different subpopulations (lettered circles) are connected via dispersal (arrows). These different subpopulations can be different sizes and are mostly independent of one another, meaning that a single subpopulation can go locally extinct (the red X) without collapsing the entire system. The different dispersal pathways mean that one population can recolonise extinct habitat and essentially ‘rebirth’ other subpopulations (the green arrows).

Weakly dispersing species

At the far opposite end of the dispersal ability spectrum, we have low dispersal species. These are often localised, endemic species that for various reasons might be unable to travel very far at all; for some, they may spend their entire adult life in a sedentary form. The lack of dispersal lends to very strong levels of population structure, and individual populations often accumulate genetic differences relatively quickly due to genetic drift or local adaptation.

Species with low dispersal capabilities are often at risk of local extinction and are unable to easily recolonise these habitats after the event has ended. Their movement is often restricted to rare environmental events such as flooding that carry individuals long distances despite their physiological limitations. Because of this, low dispersal species are often at greater risk of total extinction and extinction vertices than their higher dispersing counterparts.

Accounting for dispersal in population genetics

Incorporating biological and physiological aspects of our study taxa is important for interpreting the evolutionary context of species. Dispersal ability is but one of many characteristics that can influence the ability of species to respond to selective pressures, and the context in which this natural selection occurs. Thus, understanding all aspects of an organism is important in building the full picture of their evolution and future prospects.

An identity crisis: using genomics to determine species identities

This is the fourth (and final) part of the miniseries on the genetics and process of speciation. To start from Part One, click here.

In last week’s post, we looked at how we can use genetic tools to understand and study the process of speciation, and particularly the transition from populations to species along the speciation continuum. Following on from that, the question of “how many species do I have?” can be further examined using genetic data. Sometimes, it’s entirely necessary to look at this question using genetics (and genomics).

Cryptic species

A concept that I’ve mentioned briefly previously is that of ‘cryptic species’. These are species which are identifiable by their large genetic differences, but appear the same based on morphological, behavioural or ecological characteristics. Cryptic species often arise when a single species has become fragmented into several different populations which have been isolated for a long time from another. Although they may diverge genetically, this doesn’t necessarily always translate to changes in their morphology, ecology or behaviour, particularly if these are strongly selected for under similar environmental conditions. Thus, we need to use genetic methods to be able to detect and understand these species, as well as later classify and describe them.

Cryptic species fish
An example of cryptic species. All four fish in this figure are morphologically identical to one another, but they differ in their underlying genetic variation (indicated by the different colours of DNA). Thus, from looking at these fish alone we would not perceive any differences, but their genetic make-up might suggest that there are more than one species…
Cryptic species heatmap example
The level of genetic differentiation between the fish in the above example. The phylogenies on the left and top of the figure demonstrate the evolutionary relationships of these four fish. The matrix shows a heatmap of the level of differences between different pairwise comparisons of all four fish: red squares indicate zero genetic differences (such as when comparing a fish to itself; the middle diagonal) whilst yellow squares indicate increasingly higher levels of genetic differentiation (with bright yellow = all differences). By comparing the different fish together, we can see that Fish 1 and 2, and Fish 3 and 4, are relatively genetically similar to one another (red-deep orange). However, other comparisons show high level of genetic differences (e.g. 1 vs 3 and 1 vs 4). Based on this information, we might suggest that Fish 1 and 2 belong to one cryptic species (A) and Fish 3 and 4 belong to a second cryptic species (B).

Genetic tools to study species: the ‘Barcode of Life’

A classically employed method that uses DNA to detect and determine species is referred to as the ‘Barcode of Life’. This uses a very specific fragment of DNA from the mitochondria of the cell: the cytochrome c oxidase I gene, CO1. This gene is made of 648 base pairs and is found pretty well universally: this and the fact that CO1 evolves very slowly make it an ideal candidate for easily testing the identity of new species. Additionally, mitochondrial DNA tends to be a bit more resilient than its nuclear counterpart; thus, small or degraded tissue samples can still be sequenced for CO1, making it amenable to wildlife forensics cases. Generally, two sequences will be considered as belonging to different species if they are certain percentage different from one another.

Annotated mitogeome
The full (annotated) mitochondrial genome of humans, with the different genes within it labelled. The CO1 gene is labelled with the red arrow (sometimes also referred to as COX1) whilst blue arrows point to other genes often used in phylogenetic or taxonomic studies, depending on the group or species in question.

Despite the apparent benefits of CO1, there are of course a few drawbacks. Most of these revolve around the mitochondrial genome itself. Because mitochondria are passed on from mother to offspring (and not at all from the father), it reflects the genetic history of only one sex of the species. Secondly, the actual cut-off for species using CO1 barcoding is highly contentious and possibly not as universal as previously suggested. Levels of sequence divergence of CO1 between species that have been previously determined to be separate (through other means) have varied from anywhere between 2% to 12%. The actual translation of CO1 sequence divergence and species identity is not all that clear.

Gene tree – species tree incongruences

One particularly confounding aspect of defining species based on a single gene, and with using phylogenetic-based methods, is that the history of that gene might not actually be reflective of the history of the species. This can be a little confusing to think about but essentially leads to what we call “gene tree – species tree incongruence”. Different evolutionary events cause different effects on the underlying genetic diversity of a species (or group of species): while these may be predictable from the genetic sequence, different parts of the genome might not be as equally affected by the same exact process.

A classic example of this is hybridisation. If we have two initial species, which then hybridise with one another, we expect our resultant hybrids to be approximately made of 50% Species A DNA and 50% Species B DNA (if this is the first generation of hybrids formed; it gets a little more complicated further down the track). This means that, within the DNA sequence of the hybrid, 50% of it will reflect the history of Species A and the other 50% will reflect the history of Species B, which could differ dramatically. If we randomly sample a single gene in the hybrid, we will have no idea if that gene belongs to the genealogy of Species A or Species B, and thus we might make incorrect inferences about the history of the hybrid species.

Gene tree incongruence figure
A diagram of gene tree – species tree incongruence. Each individual coloured line represents a single gene as we trace it back through time; these are mostly bound within the limits of species divergences (the black borders). For many genes (such as the blue ones), the genes resemble the pattern of species divergences very well, albeit with some minor differences in how long ago the splits happened (at the top of the branches). However, the red genes contrast with this pattern, with clear movement across species (from and into B): this represents genes that have been transferred by hybridisation. The green line represents a gene affected by what we call incomplete lineage sorting; that is, we cannot trace it back far enough to determine exactly how/when it initially diverged and so there are still two separate green lines at the very top of the figure. You can think of each line as a separate phylogenetic tree, with the overarching species tree as the average pattern of all of the genes.

There are a number of other processes that could similarly alter our interpretations of evolutionary history based on analysing the genetic make-up of the species. The best way to handle this is simply to sample more genes: this way, the effect of variation of evolutionary history in individual genes is likely to be overpowered by the average over the entire gene pool. We interpret this as a set of individual gene trees contained within a species tree: although one gene might vary from another, the overall picture is clearer when considering all genes together.

Species delimitation

In earlier posts on The G-CAT, I’ve discussed the biogeographical patterns unveiled by my Honours research. Another key component of that paper involved using statistical modelling to determine whether cryptic species were present within the pygmy perches. I didn’t exactly elaborate on that in that section (mostly for simplicity), but this type of analysis is referred to as ‘species delimitation’. To try and simplify complicated analyses, species delimitation methods evaluate possible numbers and combinations of species within a particular dataset and provides a statistical value for which configuration of species is most supported. One program that employs species delimitation is Bayesian Phylogenetics and Phylogeography (BPP): to do this, it uses a plethora of information from the genetics of the individuals within the dataset. These include how long ago the different populations/species separated; which populations/species are most related to one another; and a pre-set minimum number of species (BPP will try to combine these in estimations, but not split them due to computational restraints). This all sounds very complex (and to a degree it is), but this allows the program to give you a statistical value for what is a species and what isn’t based on the genetics and statistical modelling.

Vittata cryptic species
The cryptic species of pygmy perches identified within my research paper. This represents part of the main phylogenetic tree result, with the estimates of divergence times from other analyses included. The pictures indicate the physiology of the different ‘species’: Nannoperca pygmaea is morphologically different to the other species of Nannoperca vittata. Species delimitation analysis suggested all four of these were genetically independent species; at the very least, it is clear that there must be at least 2 species of Nannoperca vittata since is more related to N. pygmaea than to other N. vittata species. Photo credits: N. vittata = Chris Lamin; N. pygmaea = David Morgan.

The end result of a BPP run is usually reported as a species tree (e.g. a phylogenetic tree describing species relationships) and statistical support for the delimitation of species (0-1 for each species). Because of the way the statistical component of BPP works, it has been found to give extremely high support for species identities. This has been criticised as BPP can, at time, provide high statistical support for genetically isolated lineages (i.e. divergent populations) which are not actually species.

Improving species identities with integrative taxonomy

Due to this particular drawback, and the often complex nature of species identity, using solely genetic information such as species delimitation to define species is extremely rare. Instead, we use a combination of different analytical techniques which can include genetic-based evaluations to more robustly assign and describe species. In my own paper example, we suggested that up to three ‘species’ of N. vittata that were determined as cryptic species by BPP could potentially exist pending on further analyses. We did not describe or name any of the species, as this would require a deeper delve into the exact nature and identity of these species.

As genetic data and analytical techniques improve into the future, it seems likely that our ability to detect and determine species boundaries will also improve. However, the additional supported provided by alternative aspects such as ecology, behaviour and morphology will undoubtedly be useful in the progress of taxonomy.