Phylogeographic insights into the existence of cryptic glacial refugia

Phylogeography is not a recent field of study, it was first coined in 1987 (Avise et al. 1987), but was around long before it had a name. Phylogeography “is not merely an extension of phylogenetic principles to the intraspecific level” (Schaal et al. 1998) rather it is the study of the processes that influence the dispersal of particular lineages through the construction of geneaologies, which are mapped to a particular location (Avise et al. 1987). Phylogeography is an indispensable tool in many biological fields and although it is often more concerned with describing the movements of a particular species, using discrete differences between individual populations, rather than trying to surmise possible allopatric speciation events, some studies do apply evolutionary theory to its results in an attempt to identify the origin of a species. The most notable application is the creation of two opposing theories of the origin of Homo sapiens, with phylogeographic evidence so far supporting the Out-of-Africa model (Stan, Stoneking & Wilson 1987).

How is it carried out?There are many tools available to the phylogeographer. The method of constructing the genealogy very much depends on the target species and the type of relationship one wants to infer. Plant genealogies are usually inferred through the comparison of chloroplast genes or genome fragments, as they have high substitution rates and they show little recombination. Characteristics shared by animal mitochondrial genomes, which makes them the genome of choice in animal studies. Both organelles are found to be inherited either paternally or maternally in many species of both kingdoms. It is common

What are glacial refugia?A refugium is the name given to an area of habitat where a substantial number of species have retreated to during an event that drove them from their usual habitat. The oft stated case is that of glaciation, during which substantial climate change on a global scale has turned warm habitats into frozen ones as ice caps advanced towards the equator. This led to a massive movement of species southward and brought many, once distantly separate, individuals together. Genetically this meant that any slow steps individual populations had ‘taken’ toward speciation as they existed in different environments were now all in a common gene pool. Many species would probably have become extinct as they were out-competed by other species that were already far better adapted to the new environment they suddenly found themselves in. Sometimes it is easy to mistake a dispersal event for a refugium, in that a species may have evolved, out-competed its neighbours and spread, rather than just be returning to its original habitat, but it is this homogenising effect is what is so characteristic of refugia. It leads to an extremely genetically diverse concentration of individuals of a particular species and this is what identifies a refugium. This is because, once the event, such as glaciation, is over, the species disperse again and often each region of recolonisation will have a founder, who is of a particular genetic makeup picked from the diverse pool of the refugium. This leads to discrete regions with particular gene haplotypes or polymorphisms. This genetic uniformity is what separates a recolonisation from a refugium. The best example of this is the study of the Palmaria palmata seaweed which discovered a marine refugium that at first appeared cryptic but was later solved through the rediscovery of a unique geological feature.

Glacial refugia is the current hotspot for phylogeographic studies as improvement in our comprehension of ice ages and the development of more effective molecular markers is leading to a more accurate reconstruction of the past distribution of biota, which is of high ecological importance for conservation. The best way to demonstrate the insights phylogeography has provided into glacial refugia is to look at the results it has produced. The Pleistocene glaciations were most likely the greatest historical events to befall most extant species in evolutionary terms and as such had massive consequences on their evolution and subsequent dispersion. The advancement of the glaciers forced many species to exist as fringe populations along the edge of glacier fronts and meltwater during retreat created massive proglacial lakes, which provided enormous opportunities for aquatic species to disseminate across vast distances.

The first study I have chosen to look at is one that attempts to identify a number of refugia and recolonisation routes by looking at the phylogeography of European white oaks. Plants are useful in this case because the wealth of fossilised pollen provides plenty of opportunity to uncover the location of a species at a particular time. In this study, along with the palynological (pollen) evidence, chloroplast and mitochondrial molecular markers were used. This was because these genomes are known to be maternally inherited in oaks, meaning that it should parallel seed dispersal routes (Dumolin et al. 1995). Oaks themselves are increasingly common in studies, not simply because of their own importance in an ecological sense, but also because they possess a number of characteristics that increase the accuracy and efficiency of any phylogeographic study they are subjected to. Firstly, they show high overall diversity but low levels of within population chloroplastDNA (cpDNA) diversity; this makes differentiating populations a much simpler task. Secondly, there dispersion is widespread meaning most studies on them can be applied as an overlay to other genera and not only this, the fact that they play host to a large number of other species would suggest that there dispersal may have been shadowed by the organisms that depended on them. Also extensive introgressive hybridisation necessitates the study of many species simultaneously. Finally, they produce large quantities of pollen, which has provided much palynological evidence.

The results of this study are best observed by applying them to a map demonstrating most likely colonisation routes. Each colour of line represents a different haplotype. It is suddenly apparent that there are three distinct origins of colonisation, or refugia, the Iberian peninsula, southern Italy and the Balkans. This is well in keeping with most other studies of terrestrial organisms, providing further evidence for the existence of glacial refugia.

(Petit et al. 2002)

One study commented on the focus of most phylogeography on the post-glacial recolonisation routes and the lack of information about the effect the actual contraction of the species caused by glaciation had on its genetic diversity.This study looked at this in great detail in brown trout species present in north-western Iberia.

Freshwater fish demonstrate a particular dispersal pattern as they are limited to the

network of streams present; this results in a much reduced gene flow and subsequently high levels of population structure. These qualities are ideal for a phylogeographic study of a possible glacial refugium. The study did demonstrate some support for an increased genetic differentiation between several refugia, in other words the beginnings of allopatric speciation were evidenced. However, the group found it hard to clearly define each refugium as post-glacial introgression both with native and hatchery populations led to a degree of homogeneity. Also legal limitations on sample sizes greatly reduced the strength of the inferences made.

This research paper is a good example of the problems faced by phylogeography, most of them due to anthropogenic factors, such as law, farming and artificial dispersal. These problems do need addressed quickly as more and more reports are showing irreversible damage to many ecosystems throughout the globe.

Another study of glacial refugia in fish used mitochondrial DNA D-loop sequencing (a common technique in animals) of the freshwater fish perch, to look at genetic variability between and among drainages. They found that current perch populations in western and northern Europe were colonized from three main refugia, located in southeastern, northeastern and western Europe. They also found the Baltic Sea to have been recolonised by all three refugia. This would be a good example of a case where it can be hard to discern the origin from the destination, as refugia usually contain the most genetic diversity and subsequent colonisations less so. Further study of the data suggested that the southern European mtDNA lineage is the oldest and therefore likely to be the founder of all present perch lineages. The colonization routes outlaid in the perch study probably also apply to other freshwater species with similar contemporary distribution patterns; another illustration of how any phylogeographic enquiry can have a wider application.

Very few phylogeographic studies are targeted at marine species, usually because sampling is more difficult, and there potential usefulness is overlooked. One quite interesting review looked at the congruence of the environmental factors and phylogeographic patterns displayed by a number of marine species on the Californian coast. It demonstrated the influence the environmental factors had on the evolution of many coastal taxa. It also showed the data reviewed to be consistent with the 3 hypotheses stated by Avise et al. Previously the individual reports had assumed an incongruence as they misidentified the biogeographic boundary, showing the importance of successful phylogeography on a correct model of glaciation.

A new tool in this field is comparative phylogeography and has been hailed as an effective method for assessing the roles of historical events in determining genetic diversity. Looking at the phylogeography of many species comparatively enables broader suppositions to be drawn than species-specific studies currently allow and has led to the appreciation that phylogeography may be used to illustrate the effects of historical events on the processes of population differentiation that may eventually lead to speciation (Avise 1994; Hewitt 1996). Despite the fact that most such studies have examined species with similar distributions (Avise 1992; Turner et al. 1996; Zink 1996), phylogeographic patterns of species from different areas can also be compared to determine the difference in the of effects of large-scale historical events, such as the Pleistocene glaciation.

The advancement of the ice sheets during the late Pleistocene has led many to believe that most high latitude plant and animal communities are of very recent origin, having been established by northward range expansions from southern refugia following the end of the last glacial maximum 14–20 thousand YBP (Blaise et al. 1990; Williams et al. 1998; Hewitt 1999, 2000). An effective use of comparative analysis in one study on bienthic marine gastropods shows that the population structure of the two similar species suggests a very different history. The data implies that the Nucella ostrina species recently reinvaded the northeastern Pacific, but that the N. lamellosa species survived there during the last glacial maximum in a previously uncharacterised refugium; contradictory to common belief, this implies that some northerly species are not recent inhabitants but rather persistent colonies. Though this is often an exception other studies have discovered similar cryptic refugia (Wattier).

Studies which make full use of this comparative phylogeography have provided a great deal of

insight into the effects of glaciation on the genealogy of many species and revealing new previously undiscovered refugia can help better our models of evolution, population structure and historical biogeography.