Origin of the samples
Apodemus flavicollis mice were captured across the hybrid zone ofPolyplax serrata -specific lineages in 2018 and 2019 in the north-west of the Czech Republic, using wooden snap traps. Field studies were carried out with permits approved by the Committee on the Ethics of Animal Experiments of the University of South Bohemia, by the Ministry of the Environment of the Czech Republic, and by the Ministry of the Agriculture of the Czech Republic (No. 51304/ENV/14-2981/630/14, MZP/2017/630/854, 22395/2014-MZE-17214). Mice were searched for lice by visual checking and combing. Lice were removed and stored in 100% ethanol in the -20°C. Genomic DNA from whole louse specimens was individually extracted using the Qiagen QIAamp DNA Micro Kit (Qiagen, Valencia, CA, USA). Membership to individual louse lineages (SE, SW, or N) was assigned by sequencing a fragment of the mitochondrial cytochrome oxidase subunit I gene (COI, 379 bp) as in Martinů et al (2018). All sequences of the S lineage available from previous work together with the newly gained specimens (Table S1) were collapsed into haplotypes using ALTER (http://www.sing-group.org/ALTER/). Then the specimens were assigned to individual lineages by phylogenetic reconstruction, using maximum likelihood method with 1000 bootstraps (Figure S1). Model GTR+I+G was used as the best-fit model, selected according to a corrected Akaike information criterion using jModelTest2 v2.1.10 (Darriba, Taboada, Doallo, & Posada, 2012; Guindon & Gascuel, 2003). Sample DBab5 (PolyN) from the N lineage was used as an outgroup. The S louse lineage was found on 24 A. flavicollis captured across the contact zone. Twenty-six lice (15 SE and 11 SW), 1 to maximum 3 from each parasitized host, were selected for genomic re-sequencing (Figure 3, Table 1). DNA concentrations were quantified with a Qubit 2.0 Fluorometer (Invitrogen, Carlsbad, CA, USA) using High Sensitivity reagents.