Rescuing the damselfish in distress: rescue or depression?

Conservation management

Managing and conserving threatened and endangered species in the wild is a difficult process. There are a large number of possible threats, outcomes, and it’s often not clear which of these (or how many of these) are at play at any one given time. Thankfully, there are also a large number of possible conservation tools that we might be able to use to protect, bolster and restore species at risk.

Using genetics in conservation

Naturally, we’re going to take a look at the more genetics-orientated aspects of conservation management. We’ve discussed many times the various angles and approaches we can take using large-scale genetic data, some of which include:
• studying the evolutionary history and adaptive potential of species
• developing breeding programs using estimates of relatedness to increase genetic diversity
identifying and describing new species for government legislation
• identifying biodiversity hotspots and focus areas for conservation
• identifying population boundaries for effective management/translocations

Genetics flowchart.jpg
An example of just some of the conservation applications of genetics research that we’ve talked about previously on The G-CAT.

This last point is a particularly interesting one, and an area of conservation research where genetics is used very often. Most definitions of a ‘population’ within a species rely on using genetic data and analysis (such as Fst) to provide a statistical value of how different groups of organisms are within said species. Ignoring some of the philosophical issues with the concept of a population versus a species due to the ‘speciation continuum’ (read more about that here), populations are often interpreted as a way to cluster the range of a species into separate units for conservation management. In fact, the most commonly referred to terms for population structure and levels are evolutionarily-significant units (ESUs), which are defined as a single genetically connected group of organisms that share an evolutionary history that is distinct from other populations; and management units (MUs), which may not have the same degree of separation but are still definably different with enough genetic data.

Hierarchy of structure.jpg
A diagram of the hierarchy of structure within a species. Remember that ESUs, by definition, should be evolutionary different from one another (i.e. adaptively divergent) whilst MUs are not necessarily divergent to the same degree.

This can lead to a particular paradigm of conservation management: keeping everything separate and pure is ‘best practice’. The logic is that, as these different groups have evolved slightly differently from one another (although there is often a lot of grey area about ‘differently enough’), mixing these groups together is a bad idea. Particularly, this is relevant when we consider translocations (“it’s never acceptable to move an organism from one ESU into another”) and captive breeding programs (“it’s never acceptable to breed two organisms together from different ESUs”). So, why not? Why does it matter if they’re a little different?

Outbreeding depression

Well, the classic reasoning is based on a concept called ‘outbreeding depression’. We’ve mentioned outbreeding depression before, and it is a key concept kept in mind when developing conservation programs. The simplest explanation for outbreeding depression is that evolution, through the strict process of natural selection, has pushed particularly populations to evolve certain genetic variants for a certain selective pressure. These can vary across populations, and it may mean that populations are locally adapted to a specific set of environmental conditions, with the specific set of genetic variants that best allow them to do this.

However, when you mix in the genetic variants that have evolved in a different population, by introducing a foreign individual and allowing them to breed, you essentially ‘tarnish’ the ‘pure’ gene pool of that population with what could be very bad (maladaptive) genes. The hybrid offspring of ‘native’ and this foreign individual will be less adaptive than their ‘pure native’ counterparts, and the overall adaptiveness of the population will decrease as those new variants spread (depending on the number introduced, and how negative those variants are).

Outbreeding depression example figure.jpg
An example of how outbreeding depression can affect a species. The original red fish population is not doing well- it is of conservation concern, and has very little genetic diversity (only the blue gene in this example). So, we decide to introduce new genetic diversity by adding in green fish, which have the orange gene. However, the mixture of the two genes and the maladaptive nature of the orange gene actually makes the situation worse, with the offspring showing less fitness than their preceding generations.

You might be familiar with inbreeding depression, which is based on the loss of genetic diversity from having too similar individuals breeding together to produce very genetically ‘weak’ offspring through inbreeding. Outbreeding depression could be thought of as the opposite extreme; breeding too different individuals introduced too many ‘bad’ alleles into the population, diluting the ‘good’ alleles.

Inbreeding vs outbreeding figure.jpg
An overly simplistic representation of how inbreeding and outbreeding depression can reduce overall fitness of a species. In inbreeding depression, the lack of genetic diversity due to related individuals breeding with one another makes them at risk of being unable to adapt to new pressures. Contrastingly, adding in new genes from external populations which aren’t fit for the target population can also reduce overall fitness by ‘diluting’ natural, adaptive allele frequencies in the population.

Genetic rescue

It might sound awfully purist to only preserve the local genetic diversity, and to assume that any new variants could be bad and tarnish the gene pool. And, surprisingly enough, this is an area of great debate within conservation genetics.

The counterpart to the outbreeding depression concerns is the idea of genetic rescue. For populations with already severely depleted gene pools, lacking the genetic variation to be able to adapt to new pressures (such as contemporary climate change), the situation seems incredibly dire. One way to introduce new variation, which might be the basis of new adaptation, bringing in individuals from another population of the same species can provide the necessary genetic diversity to help that population bounce back.

Genetic rescue example figure.jpg
An example of genetic rescue. This circumstance is identical to the one above, with the key difference being in the fitness of the introduced gene. The orange gene in this example is actually beneficial to the target population: by providing a new, adaptive allele for natural selection to act upon, overall fitness is increased for the red fish population.

The balance

So, what’s the balance between the two? Is introducing new genetic variation a bad idea, and going to lead to outbreeding depression; or a good idea, and lead to genetic rescue? Of course, many of the details surrounding the translocation of new genetic material is important: how different are the populations? How different are the environments (i.e. natural selection) between them? How well will the target population take up new individuals and genes?

Overall, however, the more recent and well-supported conclusion is that fears regarding outbreeding depression are often strongly exaggerated. Bad alleles that have been introduced into a population can be rapidly purged by natural selection, and the likelihood of a strongly maladaptive allele spreading throughout the population is unlikely. Secondly, given the lack of genetic diversity in the target population, most that need the genetic rescue are so badly maladaptive as it is (due to genetic drift and lack of available adaptive alleles) that introducing new variants is unlikely to make the situation much worse.

Purging and genetic rescue figure.jpg
An example of how introducing maladaptive alleles might not necessarily lead to decreased fitness. In this example, we again start with our low diversity red fish population, with only one allele (AA). To help boost genetic diversity, we introduce orange fish (with the TT allele) and green fish (with the GG allele) into the population. However, the TT allele is not very adaptive in this new environment, and individuals with the TT gene quickly die out (i.e. be ‘purged’). Individual with the GG gene, however, do well, and continue to integrate into the red population. Over time, these two variants will mix together as the two populations hybridise and overall fitness will increase for the population.

That said, outbreeding depression is not an entirely trivial concept and there are always limitations in genetic rescue procedures. For example, it would be considered a bad idea to mix two different species together and make hybrids, since the difference between two species, compared to two populations, can be a lot stronger and not necessarily a very ‘natural’ process (whereas populations can mix and disjoin relatively regularly).

The reality of conservation management

Conservation science is, at its core, a crisis discipline. It exists solely as an emergency response to the rapid extinction of species and loss of biodiversity across the globe. The time spent trying to evaluate the risk of outbreeding depression – instead of immediately developing genetic rescue programs – can cause species to tick over to the afterlife before we get a clear answer. Although careful consideration and analysis is a requirement of any good conservation program, preventing action due to almost paranoid fear is not a luxury endangered species can afford.

Origination of adaptation: the old and the new (genes)

Adaptation is arguably the most critical biological process in the evolution of species. The process of evolution by natural selection is the cornerstone of evolutionary biology (and indeed, all of contemporary biology!) and adaptation remains fundamental to the process. We know that adaptation is based on the idea that some genetic variants are ‘better’ adapted than others, and thus are unequally shared across a population. But where does this genetic variation come from?

The accumulation of new genetic variation

The classic way for new genetic variants to appear is often thought of as mutation: changes in a single base in the DNA are caused by various external processes such as chemical, physical or environmental influences (such as the sci-fi classics like UV rays or toxic chemicals). Although these forms of mutations happen very rarely and certainly don’t have the same effects comic books would leave you to believe, new mutations can occur relatively rapidly depending on the characteristics of the species. However, the most common way for new mutations to occur is actually part of the DNA replication process: copying DNA is not always perfect and even though the relevant proteins essentially run a spellcheck, sometimes the copy is not 100% perfect and new mutations occur.

Adaptation of mutation figure
An example of how adaptation can occur from a new mutation. In this example, we have one gene (TTXTT), with initial only one allele (variant), TTATT. In the second generation (row), a mutation occurs in one individual which creates a new, second allele: TTGTT. This allele is favoured over the TTATT allele, and in the next generation it’s frequency increases as the alternative allele frequency decreases (the pattern is shown in the frequency values on the right side).

It is important to remember that only mutations that are present in the reproductive cells (sperm and eggs) can be inherited and passed on, and thus be a source for adaptation. Mutations in other tissues of the body, such as within the skin, are not spread across the entire body of the subject and thus aren’t passed on to offspring.

Standing genetic variation

Alternatively, genetic variation might already be present within a species or population. This is more likely if population sizes are large and populations are well connected and interbreeding. We refer to this diverse initial gene pool as ‘standing genetic variation’: that is, the amount of genetic variation within the population or species before the selective pressure requiring adaptation. Standing genetic variation can be thought of as the ‘diversity of choices’ for natural selection to act upon: the variants are readily available, and if a good choice exists it will be favoured by natural selection and become more widespread within the population or species (i.e. evolve).

Adaptation of standing variation figure.jpg
A slightly more complex example of how adaptation can occur from standing variation, this time with two different genes. One codes for fur colour, with two different alleles: GCATA codes for orange fur, and GCGTA codes for grey fur. The other gene codes for ear tufts, with TTCCT coding for tufts and TCCCT coding for no tufts. Natural selection favours both orange fur and tufted ears, and cats with these traits reproduce more frequently than those without (see graph below). These cats probably look familiar.
Graph of standing variation.jpg
The frequency of all four alleles (i.e. either allele for both genes) over the generations in the above figure. Clearly, we can see how adaptation rapidly favours orange fur and tufted ears over grey fur and non-tufted ears with the shifts in frequencies over the different alleles.

We’ve discussed standing genetic variation before on The G-CAT, but often in a different light (and phrasing). For example, when we’ve talked about founder effect: that is, when a population is formed from only a few different individuals which causes it to be very genetically depauperate. In populations under strong founder effect, there is very little standing genetic variation for natural selection to act upon. This has long been an enigma for many pest species: how have they managed to proliferate so widely when they often originate from so few individuals and lack genetic diversity?

Adaptive variation

Adaptation may not require new genetic variants to be generated from mutation. If there are a large number of alleles within the gene pool to start with, then natural selection may favour one of those variants over others and allow adaptation to start immediately. Compared to the rate at which new mutations occur, are potentially corrected for in DNA repair, are potentially erased by genetic drift, and then put under selective pressure, adaptation from standing genetic variation can occur very quickly.

Rate of adaptation figure.jpg
A rough example of the speed of adaptation depending on how the adaptive allele originated: whether it was already present (in the form of standing variation), or whether it was created by a new mutation. As one would expect, there is a significant lag delay in adaptation in the mutation scenario, based on the time it takes for said adaptive mutation to be created through relatively random processes. Thus, a positively selected allele from standing variation can allow a species to adapt much faster than waiting for a positive mutation to occur.

Conserving genetic variation

Given the adaptive potential provided by maintaining a good amount of standing genetic variation, it is imperative to conserve genetic diversity within populations in conservation efforts. This is why we often equate genetic diversity with ‘adaptive potential’ of species, although the exact amount of genetic diversity required for adaptive potential depends on a large number of other factors. Clearly, in some instances species show the ability to adapt to new pressures or novel environments even without a large amount of standing genetic variation.

It is important to remember that standing genetic variation consists of two types: neutral genetic diversity, which is not necessarily under selection at the time, and adaptive genetic diversity, which is directly under selection (although this can be either for or against the given variant). However, currently neutral genetic variants may become adaptive variants in the future if selective pressures change: although those different variants aren’t necessarily beneficial or detrimental at the moment, that may change in the future. Thus, conserving both types of genetic diversity is important for the survivability and longevity of populations under conservation programs.

Other types of adaptation

Although genetic diversity is clearly critically important for adaptive potential, alternative mechanisms for adaptation also exist. One of these relies less on the actual genetic variants being different, but rather how individual genes are used. This can happen in a few different ways, but mostly commonly this is through alternative splicing: when a gene is being ‘read’ and a protein is produced, different parts of the gene can be used (and in different order) to make a completely different protein.

Alternate splicing figure.jpg
An extreme example of alternate splicing of one gene. We start with a single gene, composed of 5 (AE) main gene elements (exons). Different environmental pressures (like fire risk, flooding, cold weather or predators, for example) cause the organism to use different combinations of these exons to make different proteins (right side; AD). Actual alternate splicing is not usually this straight-forward (one gene doesn’t conveniently split into four forms depending on the threat), but the process is generally the same.

Believe it or not, we’ve sort of discussed the effects of alternative splicing before. Phenotypic plasticity occurs when a single organism can have very different physiological traits depending on the environment: even though the genes are the same, they are utilised in different ways to make a different body shape. This is how some species can look incredibly different when they live in different places even if they’re genetically very similar. That said, for the vast majority of species maintaining good levels of genetic diversity is critical for the survivability of said species.

It takes (at least) two: coevoultion and species interactions

The environmental context of adaptation

We’ve talked many times before about how species evolve in response to some kind of environmental pressure, which favours (or disfavours) certain traits within that species. Over time, this drives changes in the frequencies of species traits and alters the overall average phenotype of that species (sometimes slowly, sometimes rapidly).

While we usually talk about the environment in terms of abiotic conditions such as temperature or climate, biotic factors are equally important: that is, the parts of the environment which are themselves also alive. Because of this, changes in one species can have profound repercussions on other species linked within the ecosystem. Thus, the evolution of one species is intrinsically linked to the evolution of other relevant species within the ecosystem: often, these connected evolutionary pathways battle with one another as each one changes. Let’s take a look at a few different examples of how evolution of one species may impact the evolution of another.

Predator-prey coevolution

One of the most obvious ways the evolution of two different species can interact is in predator and prey relationships. Naturally, prey species evolve to be able to defend themselves from predators in various ways, such as crypsis (e.g. camouflage), toxicity or behavioural changes (such as nocturnalism or group herding). Contrastingly, predators will evolve new and improved methods for detecting and hunting prey, such as enhanced senses, venom and stealth (through soft-padded feet, for example).

There are millions of possible examples of predator-prey coevolution that could be used as examples here, based on the continual drive for one species to get the upper hand over the other. But one that comes to mind is of a creature that I learnt about while on holiday in Scandinavia: the pine marten, and how it affects squirrels.

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This photo is one that I took whilst on a lunch break at a bakery in the Norwegian mountains, of a small critter running among the rocks by the lakeside. Not sure exactly what species it was, I asked the tour director who excitedly told me that it was a pine marten. After doing a bit of research on them (and trying to figure out what the difference between a pine marten, a stoat, and a weasel is), I’ve discovered that it’s actually more likely to be a stoat than a pine marten, based on size and colour. But pine martens are still an intriguing species in their own right (and also found in Norway, so the confusion is understandable).

The pine marten is a species in the mustelid family, along with otters, weasels, stoats, and wolverines. Like many mustelids, they are carnivorous mammals which feed on a variety of different prey items like rodents, small birds and insects. One of the more abundant species that they prey upon are squirrels: both red squirrels and grey squirrels are potential food for the cute yet savage pine marten.

However, within the distribution of pine martens (across much of Europe), red squirrels are the native species and grey squirrels are invasive, originating from North America. Because of the long-lasting relationship between red squirrels and pine martens, they’ve co-evolved: most notably, by red squirrels changing to a mostly arboreal lifestyle and avoiding the ground as much as possible. Grey squirrels, however, have not had the evolutionary history to learn this lesson and are easy food for a smart pine marten. Thus, in regions where pine martens have been conserved or reintroduced, they are actively controlling the invasive grey squirrel population, which in turn boosts the native red squirrel population by reduction of competition. The coevolutionary link between red squirrels and pine martens is critical for combating the invasive species.

 

Martens and squirrels figure.jpg
The relationship between pine marten abundance and the abundance of both red (native) and grey (invasive) squirrels. On the left, without pine martens the invasive species runs rampant, outcompeting the native species. However, as pine martens increase in the ecosystem, the grey squirrels are predated on much more than the red squirrels due to their naivety, leading to the ‘natural’ balance on the right.
Martens and squirrels stats.jpg
A diagram of how the abundance of squirrels changes relative to the number of pine martens. The invasive grey squirrels are significantly depleted by pine marten presence, which in turn allows the native red squirrels to increase in population size after being freed from competition.

Host-parasite coevolution

In a similar vein to predator and prey coevolution, pathogenic species and their unfortunate hosts also undergo a sort of ‘arms race’. Parasites must keep evolving new ways to infect and transmit to hosts as the hosts evolve new methods of resisting and avoiding the infecting species. This spiralling battle of evolutionary forces is dubbed as the ‘Red Queen hypothesis’, formulated in 1973 by Leigh Van Valen and used to describe many other forms of coevolution. The name comes from Lewis Carroll’s Through the Looking Glass, and one quote in particular:

‘Now, here, you see, it takes all the running you can do, to keep in the same place’.

The quote references how species must continually adapt and respond to the evolution of other species just keep existing and prevent extinction. Species that remain static and stop evolving will inevitably go extinct as the world around them changes.

Mimicry

Plenty of other strange and unique mechanisms of coevolution exist within nature. One of them is mimicry, the process by which one species attempts to look like another to protect itself. The most iconic group known for this is butterflies: many species, although they may be evolutionarily very different, share similar colouration patterns and body shapes as mimics. Depending on the nature of the copy, mimicry can be classified into two broad categories. In either case, the initial ‘reference’ species is toxic or unpalatable to predators and uses a type of colour signal to communicate this: think of the bright yellow colours of bees and wasps or the red of ladybirds. Where the two categories change is in the nature of the ‘mimic’ species.

Müllerian mimicry

If the mimic is also toxic or unpalatable, we call this Müllerian mimicry (after Johann Friedrich Theodor Müller). By sharing the same colouration patterns and both being toxic, the two mimicking species boost the potential for the signal to be learnt by predators. If a predator eats either species, it will associate that colour pattern with toxicity and neither species are as likely to be preyed upon in the future. In this sense, it is a cooperative coevolutionary relationship between the two physically similar species.

Mullerian mimicry figure
A (somewhat familiar) example of Müllerian mimicry with two species of butterflies, the monarch and the viceroy. Although this has traditionally been thought of as a textbook case of Batesian mimicry (see below), the toxicity of both species likely makes it a scenario of Müllerian mimicry instead. Since both butterflies share the same pattern and both are toxic, it sends a strong signal to predators such as wasps to avoid them both.

Batesian mimicry

In contrast, the mimic might not actually be toxic or unpalatable, and simply copying a toxic species. This is referred to as Batesian mimicry (after Henry Walter Bates), and involves a mimic species relying on the association of colour and toxicity to have been learnt by predators through the ‘reference’ species. Although the mimic is not toxic, it is essentially piggy-backing on the hard evolutionary work that has already been done by the actually toxic species. In this case, the coevolutionary relationship is more parasitic as the mimic benefits from the ‘reference’ but the favour is not returned.

Batesian mimicry figure
An example of Batesian mimicry, with hoverflies and wasps. Hoverflies are not at all toxic, and are generally harmless; however, by mimicking the clear bright yellow warning systems of more dangerous species like wasps and bees, they avoid being eaten by predators such as birds.

Coevolution of species and the importance of species interactions

There are countless of other species interactions which could drive coevolutionary relationships in nature. These can include various forms of symbiosis, or the response of different species to ecosystem engineers: that is, species that can change and shape the environment around them (such as corals in reef systems). Understanding how a species evolves within its environment thus needs to consider how many other local species are also evolving and responding in their own ways.

 

 

Notes from the Field: Octoroks

Scientific name

Octorokus infletus

Meaning: Octorokus from [octorok] in Hylian; infletus from [inflate] in Latin.

Translation: inflating octorok; all varieties use an inflatable air sac derived from the swim bladder to float and scan the horizon.

Varieties

Octorokus infletus hydros [aquatic morphotype]

Octorokus infletus petram [mountain morphotype]

Octorokus infletus silva [forest morphotype]

Octorokus infletus arctus [snow morphotype]

Octorokus infletus imitor [deceptive morphotype]

All octoroks.jpg
The various morphotypes of inflating octoroksA: The water octorok, considered the morphotype closest to the ancestral physiology of the species. B: The forest octorok, with grass camouflage. C: The deceptive octorok, which has replaced its tufted vegetation with a glittering chest as bait. D: The mountainous octorok, with rock camouflage. E: The snow octorok, with tundra grass camouflage.

Common name

Variable octorok

Taxonomic status

Kingdom Animalia; Phylum Mollusca; Class Cephalapoda; Order Octopoda; Family Octopididae; Genus Octorokus; Species infletus

Conservation status

Least Concern

Distribution

The species is found throughout all major habitat regions of Hyrule, with localised morphotypes found within specific habitats. The only major region where the variable octorok is not found is within the Gerudo Desert, suggesting some remnant dependency of standing water.

Octorok distribution.jpg
The region of Hyrule, with the distribution of octoroks in blue. The only major region where they are not found is the Gerudo Desert in the bottom left.

Habitat

Habitat choice depends on the physiology of the morphotype; so long as the environment allows the octorok to blend in, it is highly likely there are many around (i.e. unseen).

Behaviour and ecology

The variable octorok is arguably one of the most diverse species within modern Hyrule, exhibiting a large number of different morphotypic forms and occurring in almost all major habitat zones. Historical data suggests that the water octorok (Octorokus infletus hydros) is the most ancestral morphotype, with ancient literature frequently referring to them as sea-bearing or river-traversing organisms. Estimates from the literature suggests that their adaptation to land-based living is a recent evolutionary step which facilitated rapid morphological radiation of the lineage.

Several physiological characteristics unite the variable morphological forms of the octorok into a single identifiable species. Other than the typical body structure of an octopod (eight legs, largely soft body with an elongated mantle region), the primary diagnostic trait of the octorok is the presence of a large ‘balloon’ with the top of the mantle. This appears to be derived from the swim bladder of the ancestral octorok, which has shifted to the cranial region. The octorok can inflate this balloon using air pumped through the gills, filling it and lifting the octorok into the air. All morphotypes use this to scan the surrounding region to identify prey items, including attacking people if aggravated.

inflated octorok
A water morphotype octorok with balloon inflated.

Diets of the octorok vary depending on the morphotype and based on the ecological habitat; adaptations to different ecological niches is facilitated by a diverse and generalist diet.

Demography

Although limited information is available on the amount of gene flow and population connectivity between different morphotypes, by sheer numbers alone it would appear the variable octorok is highly abundant. Some records of interactions between morphotypes (such as at the water’s edge within forested areas) implies that the different types are not reproductively isolated and can form hybrids: how this impacts resultant hybrid morphotypes and development is unknown. However, given the propensity of morphotypes to be largely limited to their adaptive habitats, it would seem reasonable to assume that some level of population structure is present across types.

Adaptive traits

The variable octorok appears remarkably diverse in physiology, although the recent nature of their divergence and the observed interactions between morphological types suggests that they are not reproductively isolated. Whether these are the result of phenotypic plasticity, and environmental pressures are responsible for associated physiological changes to different environments, or genetically coded at early stages of development is unknown due to the cryptic nature of octorok spawning.

All octoroks employ strong behavioural and physiological traits for camouflage and ambush predation. Vegetation is usually placed on the top of the cranium of all morphotypes, with the exact species of plant used dependent on the environment (e.g. forest morphotypes will use grasses or ferns, whilst mountain morphotypes will use rocky boulders). The octorok will then dig beneath the surface until just the vegetation is showing, effectively blending in with the environment and only occasionally choosing to surface by using the balloon. Whether this behaviour is passed down genetically or taught from parents is unclear.

Management actions

Few management actions are recommended for this highly abundant species. However, further research is needed to better understand the highly variable nature and the process of evolution underpinning their diverse morphology. Whether morphotypes are genetically hardwired by inheritance of determinant genes, or whether alterations in gene expression caused by the environmental context of octoroks (i.e. phenotypic plasticity) provides an intriguing avenue of insight into the evolution of Hylian fauna.

Nevertheless, the transition from the marine environment onto the terrestrial landscape appears to be a significant stepping stone in the radiation of morphological structures within the species. How this has been facilitated by the genetic architecture of the octorok is a mystery.

 

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.

From mutation to speciation: the genetics of species formation

The genetics of speciation

Given the strong influence of genetic identity on the process and outcomes of the speciation process, it seems a natural connection to use genetic information to study speciation and species identities. There is a plethora of genetics-based tools we can use to investigate how speciation occurs (both the evolutionary processes and the external influences that drive it). One clear way to test whether two populations of a particular species are actually two different species is to investigate genes related to reproductive isolation: if the genetic differences demonstrate reproductive incompatibilities across the two populations, then there is strong evidence that they are separate species (at least under the Biological Species Concept; see Part One for why!). But this type of analysis requires several tools: 1) knowledge of the specific genes related to reproduction (e.g. formation of sperm and eggs, genital morphology, etc.), 2) the complete and annotated genome of the species (to be able to find and analyse the right genes properly) and 3) a good amount of data for the populations in question. As you can imagine, for people working on non-model species (i.e. ones that haven’t had the same history and detail of research as, say, humans and mice), this can be problematic. So, instead, we can use other genetic information to investigate and suggest patterns and processes related to the formation of new species.

Is reproductive isolation naturally selected for or just a consequence?

A fundamental aspect of studies of speciation is a “chicken or the egg”-type paradigm: does natural selection directly select for rapid reproductive isolation, preventing interbreeding; or as a secondary consequence of general adaptive differences, over a long history of evolution? This might be a confusing distinction, so we’ll dive into it a little more.

Of the two proposed models of speciation, the by-product of natural selection (the second model) has been the more favoured. Simply put, this expands on Darwin’s theory of evolution that describes two populations of a single species evolving independently of one another. As these become more and more different, both in physical (‘phenotype’) and genetic (‘genotype’) characteristics, there comes a turning point where they are so different that an individual from one population could not reasonably breed with an individual from the other to form a fertile offspring. This could be due to genetic incompatibilities (such as different chromosome numbers), physiological differences (such as changes in genital morphology), or behavioural conflicts (such as solitary vs. group living).

Certainly, this process makes sense, although it is debatable how fast reproductive isolation would occur in a given species (or whether it is predictable just based on the level of differentiation between two populations). Another model suggests that reproductive isolation actually might arise very quickly if natural selection favours maintaining particular combinations of traits together. This can happen if hybrids between two populations are not particularly well adapted (fit), causing natural selection to favour populations to breed within each group rather than across groups (leading to reproductive isolation). Typically, this is referred to as ‘reinforcement’ and predominantly involves isolating mechanisms that prevent individuals across populations from breeding in the first place (since this would be wasted energy and resources producing unfit offspring). The main difference between these two models is the sequence of events: do populations ecologically diverge, and because of that then become reproductively isolated, or do populations selectively breed (enforcing reproductive isolation) and thus then evolve independently?

Reinforcement figure.jpg
An example of reinforcement leading to speciation. A) We start with two populations of a single species (a red fish population and a green fish population), which can interbreed (the arrows). B) Because these two groups can breed, hybrids of the two populations can be formed. However, due to the poor combination of red and green fish genes within a hybrid, they are not overly fit (the red cross). C) Since natural selection doesn’t favour forming hybrids, populations then adapt to selectively breed only with similar fish, reducing the amount of interbreeding that occurs. D) With the two populations effectively isolated from one another, different adaptations specific to each population (spines in red fish, purple stripes in green fish) can evolve, causing them to further differentiate. E) At some point in the differentiation process, hybrids move from being just selectively unfit (as in B)) to entirely impossible, thus making the two populations formal species. In this example, evolution has directly selected against hybrids first, thus then allowing ecological differences to occur (as opposed to the other way around).

Reproductive isolation through DMIs

The reproductive incompatibility of two populations (thus making them species) is often intrinsically linked to the genetic make-up of those two species. Some conflicts in the genetics of Population 1 and Population 2 may mean that a hybrid having half Population 1 genes and half Population 2 genes will have serious fitness problems (such as sterility or developmental problems). Dramatic genetic differences, particularly a difference in the number of chromosomes between the two sources, is a significant component of reproductive isolation and is usually to blame for sterile hybrids such as ligers, zorse and mules.

However, subtler genetic differences can also have a strong effect: for example, the unique combination of Population 1 and Population 2 genes within a hybrid might interact with one another negatively and cause serious detrimental effects. These are referred to as “Dobzhansky-Müller Incompatibilities” (DMIs) and are expected to accumulate as the two populations become more genetically differentiated from one another. This can be a little complicated to imagine (and is based upon mathematical models), but the basis of the concept is that some combinations of gene variants have never, over evolutionary history, been tested together as the two populations diverge. Hybridisation of these two populations suddenly makes brand new combinations of genes, some of which may be have profound physiological impacts (including on reproduction).

DMI figure
An example of how Dobzhansky-Müller Incompatibilities arise, adapted from Coyne & Orr (2004). We start with an initial population (center top), which splits into two separate populations. In this example, we’ll look at how 5 genes (each letter = one gene) change over time in the separate populations, with the original allele of the gene (lowercase) occasionally mutating into a new allele (upper case). These mutations happen at random times and in random genes in each population (the red letters), such that the two become very different over time. After a while, these two populations might form hybrids; however, given the number of changes in each population, this hybrid might have some combinations of alleles that are ‘untested’ in their evolutionary history (see below). These untested combinations may cause the hybrid to be infertile or unviable, making the two populations isolated species.

DMI table
The list of ‘untested’ genetic combinations from the above example. This table shows the different combinations of each gene that could be made in a hybrid if these two populations interbred. The red cells indicate combinations that have never been ‘tested’ together; that is, at no point in the evolutionary history of these two populations were those two particular alleles together in the same individual. Green cells indicate ones that were together at some point, and thus are expected to be viable combinations (since the resultant populations are obviously alive and breeding).

How can we look at speciation in action?

We can study the process of speciation in the natural world without focussing on the ‘reproductive isolation’ element of species identity as well. For many species, we are unlikely to have the detail (such as an annotated genome and known functions of genes related to reproduction) required to study speciation at this level in any case. Instead, we might choose to focus on the different factors that are currently influencing the process of speciation, such as how the environmental, demographic or adaptive contexts of populations plays a role in the formation of new species. Many of these questions fall within the domain of phylogeography; particularly, how the historical environment has shaped the diversity of populations and species today.

Phylogeo of speciation
An example of the interplay between speciation and phylogeography, taken from Reyes-Velasco et al. (2018). They investigated the phylogeographic history of several different groups of species within the frog genus Ptychadena; in this figure, we can see how the different species (indicated by the colours and tree on the left) relate to the geography of their habitat (right).

A variety of different analytical techniques can be used to build a picture of the speciation process for closely related or incipient species. A good starting point for any speciation study is to look at how the different study populations are adapting; is there evidence that natural selection is pushing these populations towards different genotypes or ecological niches? If so, then this might be a precursor for speciation, and we can build on this inference with other complementary analyses.

For example, estimating divergence times between populations can help us suggest whether there has been sufficient time for speciation to occur (although this isn’t always clear cut). Additionally, we could estimate the levels of genetic hybridisation (‘introgression’) between two populations to suggest whether they are reasonably isolated and divergent enough to be considered functional species.

The future of speciation genomics

Although these can help answer some questions related to speciation, new tools are constantly needed to provide a clearer picture of the process. Understanding how and why new species are formed is a critical aspect of understanding the world’s biodiversity. How can we predict if a population will speciate at some point? What environmental factors are most important for driving the formation of new species? How stable are species identities, really? These questions (and many more) remain elusive for a wide variety of life on Earth.

 

The direction of evolution: divergence vs. convergence

Direction of evolution

We’ve talked previously on The G-CAT about how the genetic underpinning of certain evolutionary traits can change in different directions depending on the selective pressure it is under. Particularly, we can see how the frequency of different alleles might change in one direction or another, or stabilise somewhere in the middle, depending on its encoded trait. But thinking bigger picture than just the genetics of one trait, we can actually see that evolution as an entire process works rather similarly.

Divergent evolution

The classic view of the direction of evolution is based on divergent evolution. This is simply the idea that a particular species possess some ancestral trait. The species (or population) then splits into two (for one reason or another), and each one of these resultant species and populations evolves in a different way to the other. Over time, this means that their traits are changing in different directions, but ultimately originate from the same ancestral source.

Evidence for divergent evolution is rife throughout nature, and is a fundamental component of all of our understanding of evolution. Divergent evolution means that, by comparing similar traits in two species (called homologous traits), we can trace back species histories to common ancestors. Some impressive examples of this exist in nature, such as the number of bones in most mammalian species. Humans have the same number of neck bones as giraffes; thus, we can suggest that the ancestor of both species (and all mammals) probably had a similar number of neck bones. It’s just that the giraffe lineage evolved longer bones whereas other lineages did not.

Homology figure
A diagrammatic example of homologous structures in ‘hand’ bones. The coloured bones demonstrate how the same original bone structures have diverged into different forms. Source: BiologyWise.

Convergent evolution

But of course, evolution never works as simply as you want it to, and sometimes we can get the direct opposite pattern. This is called convergent evolution, and occurs when two completely different species independently evolve very similar (sometimes practically identical) traits. This is often caused by a limitation of the environment; some extreme demand of the environment requires a particular physiological solution, and thus all species must develop that trait in order to survive. An example of this would be the physiology of carnivorous marsupials like Tasmanian devils or thylacines: despite being in another Class, their body shapes closely resemble something more canid. Likely, the carnivorous diet places some constraints on physiology, particularly jaw structure and strength.

Convergent evol intelligence
A surprising example of convergent evolution is cognitive ability in apes and some bird groups (e.g. corvids). There’s plenty of other animal groups more related to each of these that don’t demonstrate the same level of cognitive reasoning (based on the traits listed in the centre): thus, we can conclude that cognition has evolved twice in very, very different lineages. Source: Emery & Clayton, 2004.

A more dramatic (and potentially obvious) example of convergent evolution would be wings and the power of flight. Despite the fact that butterflies, bees, birds and bats all have wings and can fly, most of them are pretty unrelated to one another. It seems much more likely that flight evolved independently multiple times, rather than the other 99% of species that shared the same ancestor lost the capacity of flight.

Parallel evolution

Sometimes convergent evolution can work between two species that are pretty closely related, but still evolved independently of one another. This is distinguished from other categories of evolution as parallel evolution: the main difference is that while both species may have shared the same start and end point, evolution has acted on each one independent of the other. This can make it very difficult to diagnose from convergent evolution, and is usually determined by the exact history of the trait in question.

Parallel evolution is an interesting field of research for a few reasons. Firstly, it provides a scenario in which we can more rigorously test expectations and outcomes of evolution in a particular environment. For example, if we find traits that are parallel in a whole bunch of fish species in a particular region, we can start to look at how that particular environment drives evolution across all fish species, as opposed to one species case studies.

Marsupial handedness.jpg
Here’s another weird example; different populations of marsupials (particularly kangaroos and wallabies) show preferential handedness depending on where the population is. That is, different populations of different species of marsupials shows parallel evolution of handedness, since they’re related to one another but have evolved it independently of the other species. Source: Giljov et al. (2015).

Following from that logic, it is then important to question the mechanisms of parallelism. From a genetic point of view, do these various species use the same genes (and genetic variants) to produce the same identical trait? Or are there many solutions to the selective question in nature? While these questions are rather complicated, and there has been plenty of evidence both for and against parallel genetic underpinning of parallel traits, it seems surprisingly often that many different genetic combinations can be used to get the same result. This gives interesting insight into how complex genetic coding of traits can be, and how creative and diverse evolution can be in the real world.

Where is evolution going?

Cat phylogeny
An example of all three types of evolutionary trajectory in a single phylogeny of cats (you know how we do it here at The G-CAT). This phylogeny consists of two distinct genera; one with one species (P. aliquam) and another of three species (the red box indicates their distance). Our species have three main physical traits: coat colour, ear tufts and tail shape. At the ancestral nodes of the tree, we can see what the ancestor of these species looked like for these three traits. Each of these traits has undergone a different type of evolution. The tufts on the ears are the result of divergent evolution, since F. tuftus evolved the trait differently to its nearest relative, F. griseo. Contrastingly, the orange coat colour of F. tuftus and P. aliquam are the result of convergent evolution: neither of these species are very closely related (remembering the red box) and evolved orange coats independently of one another (since their ancestors are grey). And finally, the fluffy tails of F. hispida and F. griseo can be considered parallel evolution, since they’re similar evolutionarily (same genus) but still each evolved tail fluff independently (not in the ancestor). This example is a little convoluted, but if you trace the history of each trait in the phylogeny you can more easily see these different patterns.

So, where is evolution going for nature? Well, the answer is probably all over the place, but steered by the current environmental circumstances. Predicting the evolutionary impacts of particular environmental change (e.g. climate change) is exceedingly difficult but a critical component of understanding the process of evolution and the future of species. Evolution continually surprises us with creative solution to complex problems and I have no doubt new mysteries will continue to be thrown at us as we delve deeper.

All the world in the palm of your hand: whole genome sequencing for evolution and conservation

Building an entire genome

If bigger is better, then biggest is best. Having the genome of a particular study species fully sequenced allows us to potentially look at all of the genetic variation in the entire gene pool: but how do we sequence the entirety of the genome? And what are the benefits of having a whole genome to refer to?

Whole genome assembly
A very, very simplified overview of whole genome sequencing. Similar to other genomic technologies, we start by fragmenting the genome into much smaller, easier to sequence parts (reads). We then use a computer algorithm which pieces these reads together into a consecutive sequence based on overlapping DNA sequence (like building a chain out of Lego blocks). From this assembled genome, we can then attach annotations using information from other species’ genomes or genetic studies, which can correlate a particular sequence to a gene, a function of that gene, and the resultant protein from these gene (although not always are all of these aspects included).

Well, assembling the whole genome of an organism for the first time is a very tricky process. It involves taking DNA sequence from only a few individuals, breaking them down into smaller fragments and multiplying these fragments into the billions (moreorless the same process used in other genomics technologies: the real difference is that we need the full breadth of the genome so that we don’t miss any spaces). From these fragments, we use a complex computer algorithm which builds up a consensus sequence like a Lego tower; by finding parts of sequences which overlap, the software figures out which pieces connect to one another. Hopefully, we eventually end up with one very long continuous sequence; the genome! Sometimes, we might end with a few very large blocks (called contigs), but this is also useful for analyses (correlated with how many/big blocks there are). With this full genome, we use information from other more completed genomes (such as those from model species like humans, mice or even worms) to figure out which sections of the genome relate to specific genes. We can then annotate these sections by labelling them as clear genes, complete with start and end point, and attach a particular physical function of that gene.

The benefits of whole genomes

Having an entire genome as a reference is an extremely helpful tool in conservation and evolutionary studies. The first, and perhaps most obvious benefit, is the sheer scale of the data we can use. By having the entirety of the genome available, we can use potentially billions of base pairs of sequence in our genetic analyses (for reference, the human genome is >3 billion base pairs long). Even if we don’t sequence the full genome for all of our samples, having a reference genome as basis for assembly our reduced datasets significantly improves the quantity and quality of sequences we can use.

Another very important benefit is the ability to prescribe function in our studies. Many of our processes for obtaining data, even for genomic technologies, use random and anonymous fragments of the genome. Although this is a cost-effective way to obtain a very large amount of data, it unfortunately means that we often have no idea which part of the genome our sequences came from. This means that we don’t know which sequences relate to specific genes, and even if we did we would have no idea what those genes are or do! But with an annotated genome, we can take even our fragmented sequence and check it against the genome and find out what genes are present.

Understanding adaptation

Based on that, it seems pretty obvious about exactly how having an annotated genome can help us in studies of adaptation. Knowing the functional aspect of our genetic data allows us to more directly determine how evolution is happening in nature; instead of only being able to say that two species are evolving differently from one another, for example, we can explicitly look at how they are evolving. Is one evolving tolerance to hotter temperatures? Are they evolving different genes to handle different diets? Are they evolving in response to an external influence, like a viral outbreak or changing climate? What are the physiological consequences of these changes? These questions are critical in understanding past and future evolution, and full genome analysis allows us to delve into them much deeper.

Manhattan plot example
A (slightly edited) figure of full genome comparisons between domestic dogs and wild wolves by Axelsson et al. (2013), with the aim of understanding the evolutionary changes associated with domestication. For avid readers, this figure probably looks familiar. This figure compares the genetic differentiation across the entire genome between dogs and wolves, with some sections of the genome (circled) showing clear differences. As there is an annotated dog genome, the authors then delved into these genes to understand the functional differences between the two. By comparing their genetic differences to functional genes, the authors can more explicitly suggest mechanisms or changes associated with the domestication process (such as adaptation to a starch-heavy and human-influenced diet).

 

 

This includes allowing us to better understand how adaptation actually works in nature. As we’ve discussed before, more traditional studies often assumed that single, or very few, genes were responsible for allowing a species to adapt and change, and that these genes had very strong effects on their physiology. But what we see far more often is polygenic adaptation; small changes in a very large number of genes which, combined together, allow the species to adapt and evolve. By having the entirety of the genome available, we are much more likely to capture all of the genes that are under natural selection in a particular population or species, painting a clearer picture of their evolutionary trajectory.

Understanding demography

The much larger dataset of full genomes is also important for understanding the non-adaptive parts of evolution; the demographic history. Given that selectively neutral impacts (e.g. reductions in population size) are likely to impact all of the genes in the gene pool somewhat equally, having a full genome allows us to more accurately infer the demographic state and historical patterns of species.

For both adaptive and non-adaptive variation, it is also important to consider what we call linkage disequilibrium. Genetic sequences that are physically close to each other in the genome will often be inherited together due to the imprecision of recombination (a fairly technical process, so I won’t delve into this): what this can mean is that if a gene is under very strong selection, then sequences around this gene will also look like they’re under selection too. This can give falsely positive adaptive genes (i.e. sequences that look like genes under selection but are just linked to a gene that is) or can interfere with demographic analyses (since they often assume no selection, or linkage to selection, on the sequences used). With a whole genome, we can actually estimate how far away a base pair has to be before it’s not linked anymore; we call these linkage blocks, and they’re very useful additions to analyses.

Linkage_example
An example of linkage as a process. We start with a particular sequence (top); during recombination, this sequence may randomly break and rearrange into different parts. In this example, I’ve simulated four different ‘breaks’ (dashed coloured lines) due to recombination. Each of these breaks leads to two separate blocks of fragments; for example, the break at the blue line results in the second two sequence blocks (middle). If we focus on one target base pair in the sequence (golden A), then we can see in some fragments it remains with certain bases, but sometimes it gets separated by the break. If we compare how often the golden A is in the same block (i.e. is co-inherited) as each of the other bases, across all 4 breaks, then we see that the bases that are closest to it (the golden A is represented by the golden bar) are almost always in the same block. This makes sense: the further away a base is from our target, the more likely that there will be a break between it. This is shown in the frequency distributions at the bottom: the left figure shows the actual frequencies of co-inheritance (i.e. linkage) using the top example and those 4 breaks. The right figure shows a more realistic depiction of how linkage looks in the genome; it rapidly decays as we move away from the target (although the width and rate of this can vary).

Improving conservation management

In a similar fashion to demography, full genome datasets can improve our estimates of relatedness and pedigrees in captive breeding programs. The massive scale of whole genomes allows us to more easily trace the genealogical history of individuals, allowing us to assign parents more accurately. This also helps with our estimations of genetic relatedness, arguably the most critical aspect of genetic-based breeding programs. This is particularly helpful for species with tricky mating patterns, such as polyamory, brood spawning or difficult to track organisms.

Pedigrees
An example of how whole genomes can improve our estimation of pedigrees. Say we have a random individual (star), and we want to know how they fit into a particular family tree (pedigree). With only a few genes, we might struggle to pick where in the family it fits based on limited genetic information. With a larger genetic dataset (such as reduced-representation genomics), we might be able to cross off a few potential candidate spots but still have some trouble with a few places (due to unknown parents, polygamy or issues with genetic analysis). With whole genomes, we should be able to much better clarify the whole pedigree and find exactly where our star individual fits in the tree (red circle). It is thanks to whole genomes, we can do those ancestry analyses that have gone viral lately!

The way forwards

While many non-model species are still lacking in the available genomic information, whole genomes are progressively being sequenced for more and more species. As this astronomical dataset grows, our ability to investigate, discover and test theories about evolution, natural selection and conservation will also improve. Many projects already exist which aim specifically to increase the number of whole genomes available for certain taxonomic groups such as birds and bats: these will no doubt prove to be invaluable resources for future studies.

Why we should always pander to diversity

Diversity in the natural world

‘Diversity’ is a term that gets used a lot these days, albeit usually in reference to social changes and structures. However, diversity is not merely a human construct and reflects an extremely important aspect of the natural world at a variety of levels. From the smallest genes to the biggest ecosystems, diversity is a trait that confers a massive range of benefits to individuals, populations, species and even the entire globe. Let’s dissect this diversity down at different scales and see how beneficial it can be.

Hierarchy of diversity
The generalised hierarchy at life, with diversity being an important component of each tier. At the smallest tier, genes underpin all life. The collection of genetic diversity is often summarised into a population (as a single cohesive genetic unit). Several populations can be pooled together into a single (usually) cohesive speciesDifferent species are then components of a larger community (which in turn are components of a broader ecosystem).

Genetic diversity

At the smallest scale in the hierarchy of genetic differentiation, we have the genes themselves. It is a well-established concept that having a diversity of genetic variants (alleles) within a population or species is critical to their future adaptation, evolution and persistance. This is because different alleles will have different benefits (or costs) depending on the environmental pressure that influences them; natural selection might favour one allele over another at one time, but a different one as the pressure changes. Having a higher number of alleles within the population or species means that there is a greater chance at least a few individuals will possess an adaptive gene with the changing environment (which we know can be quite rapid and very, very strong). The diversity serves as a ‘buffer’ against extinction; evolution by natural selection functions best when there are many options to choose from.

Without this diversity, species run the risk of having no adaptive genes at the ready to deal with a selective pressure. Either a new adaptive gene must mutate (or come about in other ways, such as through gene flow from another population or species) or the population/species will suffer and potentially go extinct. As strong selection causes the species to dwindle, it enters what is referred to as the ‘extinction vortex’. Without genetic diversity, they can’t adapt: thus, more individuals die off, causing more genetic diversity to be lost from the population. This pattern is a vicious cycle which can inevitably destroy species (without serious intervention).

Extinction vortex
A very dramatic representation of the extinction vortex.

For this reason, captive breeding programs aim to maintain as much of the genetic diversity of the original population as possible. This reduces the probability of entering a downward extinction spiral from inbreeding depression and helps to maintain populations into the future (both the captive one and the wild population when we reintroduce individuals into the wild).

“Population”  diversity

Because genetic diversity is critically important for species survival, we must also try to preserve the diversity of the entire gene pool of a species. This means conserving highly genetically differentiated populations within a species as a priority, as they may be the only ones that possess the necessary adaptive genes to save the rest of the species. This adaptive genetic variation can then be introduced into other populations in genetic rescue programs and serve as a means to semi-naturally allow the species to evolve. Evolutionarily-significant units (ESUs) are one measure of the invaluable nature of genetically unique populations.

Although many more traditional conservationists strongly believe that ESUs should be managed entirely independently of one another (to preserve their evolutionary ‘pedigree’ and prevent the risk of outbreeding depression), it has been suggested that the benefit of genetic rescue in many cases significantly outweighs this risk of outbreeding depression. For some species, this really is an act of rescue: they are at the edge of extinction, and if we do nothing we condemn them to die out.

Introducing genetic material across populations (or even species!) can generate new functional genes that allow the recipient species to adapt to selective pressures. This might sound very strange, and could be extremely rare, but examples of adaptive genetic material in one species originating from another species through hybridisation do exist in nature. For example, the black coat of wolves is a highly adaptive trait in some populations and is encoded for by the Melanocortin 1 receptor (Mc1r) gene. However, the specific mutation in Mc1r gene that generates the black coat colour actually first originated in domestic dogs; when wild wolves and domestic dogs interbred, this mutation was transferred into the wolf gene pool. Natural selection strongly favoured this new variant, and it very rapidly underwent strong positive selection. Thus, the adaptiveness of black wolves is thanks to a domestic dog mutation!

Species diversity

At a higher level of the hierarchy, the diversity of species within a particular community or ecosystem has been shown to be important for the health and stability of said community. Every species, however small or seemingly unimpressive, plays a role in the greater ecosystem balance, through interactions with other species (e.g. as predator, as prey, as competitor) and the abiotic environment. While some species are known to have very strong impacts on the immediate ecosystem (often dubbed ‘keystone species’, such as apex predators), all species have some influence on the world around them (we’re especially good at it).

Species interactions flowchart

The overall health and stability of an ecosystem, as well as the benefits it can provide to all living things (including humans) is largely determined by the diversity of species. For example, ‘habitat engineers’ are types of species that, by altering the physical environment around them (such as to build a home), directly provide new habitat for other species. They are a fundamental underpinning of many incredibly vibrant ecosystems; think of what a reef system would look like if there were no corals in it. There’d be no anemones growing colourfully; no fish to live in them; no sharks to feed on these non-existent fish. This is just one example of a complex ecosystem that truly relies on its inhabiting species to function.

Ecosystem jenga
Much like Jenga, taking out one block (a species) could cause the entire stack (the ecosystem) to collapse in on itself. Even if it stands up, however, the system will still be weaker without the full diversity to support it.

Protecting our diversity

Diversity is not just a social construct and is an important phenomenon in nature, at a variety of different levels. Preserving the full diversity of life, from genetic diversity within populations and species to full species diversity within ecosystems, is critical to maintaining healthy and robust natural systems. The more diversity we have at each level of this hierarchy, the greater robustness and security we will have in the future.