In search for adaptive variation across genomes of the widespread Eurasian lynx and critically endangered Iberian lynx.

The aim of the project was to investigate the similarities and differences between the genetic variability
observed with neutral markers unaffected by natural selection, and the functional diversity influenced by
selection. This was done on the basis of genome variability analyzes of two closely related species of large
carnivores: the Eurasian lynx (Lynx lynx) and the Iberian lynx (L. pardinus). As these species differ
significantly in terms of population demographics (the Iberian lynx is on the verge of extinction, and the
Eurasian lynx, although its populations vary widely in terms of demographics, is one of the most widely
distributed cats in the world), they provide a good model for genetic analysis of basics of adaptation
processes and their relationship with populations declines. The research was based on the assumption that the
diversity of the environmental and climatic conditions inhabited by the Eurasian lynx may generate few but
significant changes in the functional part of its genome as well as in the methylome (epigenetic variability)
resulting from the adaptation of animals to different local conditions. The project idea was to study these
processes in three complementary steps based on re-sequencing of the lynx genome:
1) Tracing the demographic and evolutionary history and reconstruction of the phylogeography of lynx
based on the analysis of the mitogenome sequence and neutral sequences of the nuclear genome.
2) Genome re-sequencing in search of traces of natural selection related to environmental factors.
3) Study of genome methylation in search of epigenetic variability and its role in biological processes,
including the initial stages of speciation and inter-population divergence.
Thanks to the cooperation of many biologists of the international team of the project managed to obtain and
analyze samples of several dozen lynx from twelve different populations within the Eurasian range, living in
extremely different conditions from the forests of the temperate zone of north-eastern Poland, through the
Gobi desert, the taiga of northern Siberia to the far-east coast of Russia. The samples were sequenced and
annotated to the previously studied genome of the Iberian lynx. A great success of the project is the
reconstruction of the evolutionary history and demographics of the Eurasian lynx throughout the Pleistocene,
starting from the period of speciation from a common ancestor with the Canadian lynx (L. canadensis) and
examining the level of genome variability of this species throughout the Eurasian range of the species. These
studies have shown an extremely low level of genomic variation in this species, which testifies to the
strength and rate of genetic erosion caused by both climate change and civilization development. On the
other hand, this low variability and genomic homogeneity of the species, visible on huge stretches of the
continent inhabited by this species, shows that despite the extremely diverse climatic and environmental
conditions, the lynx is probably an animal with wide adaptability. Our research has shown that although
adaptive variability is highly probable in this species, it affects a very small (about 0.01%) part of the
genome. Bioinformatic analyzes aiming at identifying specific genes and linking them to adaptations to the
environment are not yet complete, but approximately 260 SNPs have been identified that are very likely to
be affected by natural selection. Preliminary results indicate the possibility of selective influence of the mean
annual temperature on the adaptive variability of the lynx. This result is all the more interesting as the
influence of temperature was also revealed as a factor related to the color variability of these predators.
An attempt was also made to investigate the epigenetic variability of both lynx species by analyzing the
degree of methylation of various genome fragments, which may occur earlier than changes in the genome
and thus constitute an important adaptive mechanism. Studies have confirmed the existence of differences in
methylation levels both between species and between the studied populations. These differences have been
linked to biological processes (e.g. protein binding, intracellular signaling pathway, transferase activity)
caused by differentiated methylation genes and it has been shown that the differences between the Iberian
and Eurasian lynx concern 20 processes, while between the two populations of the Eurasian species they
concern 27 processes. The functional and evolutionary significance of the identified differentiation is
currently being analyzed.
Owing to a possibility of studying and comparing whole genomes from populations of two sister lynx
species with contrasting demographic histories, the project also provided valuable insight into the
mechanisms of the purging phenomenon and balancing selection in populations exposed to high levels of
inbreeding and genomic load. It has been shown that the genomic load of some deleterious mutations is
lower in the Iberian lynx than in the Eurasian lynx, and that despite passing through a genetic bottleneck, the
MHC genes being the elements of immune system retained their functional variability. These results also
suggest that despite the extensive genetic erosion of such populations, there may be some reduction in
inbreeding depression that may enable its survival. The use of genomic technologies in this project will
enable, among others, the development of conservation biology, especially in relation to studying the
adaptive potential of large carnivores’ populations being in critical danger.