Diversity and population pattern of nuclear SNPs
Illumina reads from 26 genomes were successfully mapped on the reference
with mean coverage ranging from 18.9x (30VOJ_SW) to 34x (29DZ_SW).
None of the assemblies had to be filtered out due to low coverage and/or
low Phred scores.
Patterns of population structure obtained using nuclear SNPs differed
from genealogies based on maternally inherited genomes (Figure 4). In
contrast to mtDNA, the structure derived by PCA from nuclear SNPs did
not recognize only two mutually exclusive clusters corresponding to SW
and SE mitochondrial lineages but also a third admixed cluster
containing hybrid individuals. The pattern fitted very well geographical
sampling along the anticipated contact zone in a transect running from
north-west (SW population 2HBC_HorBl_SW) to the south-east (SE
population 39RAD_SE). The geographical outlier, sample NE125b_Kot_SW
from north-east Germany, was positioned within the hybrid samples on the
main axis, whilst being separated from the rest of the samples along the
other axis. This pattern may reflect a large geographical distance from
the Czech localities combined with possible occurrence of a contact zone
in NE Germany (Figure 4, map Figure 3).
Bayesian analysis in Admixture found the highest support for the
occurrence of two clusters (see Figure S4 for cross-validation values).
Each of the two clusters corresponded to one of the mtDNA lineages, with
samples from the centre of the contact zone and the German specimen
Ne125b_Kot showing admixed origin (Figure 4d).
ML tree-building method yielded a
similar pattern to PCA (Figure S5). It revealed the two parental
populations as monophyletic clusters and the hybrids forming an
intermediate paraphyletic assemblage (with the long-branched Ne125b_Kot
sample clustering among the hybrids; Figure S5).
Measures of genetic diversity
corresponded very well with the previous methods. When divided into
three populations (SW, SE, and hybrids), the highest pairwise
FST value was detected between the SW and SE clusters
(0.59), compared to the lower values between hybrids and SW or SE
clusters (0.21 and 0.36, respectively, Table S4). Similarly, hybrids
showed approximately 2 fold increase in heterozygosity compared to
either of the SW and SE clusters (Table S4), as expected for individuals
admixed between two lineages.