DOI: 10.5061/dryad.5dv41ns6p
title: Association of SNPs in Microtus arvalis and clade infections by TULV-CEN.S and TULV-EST.S

description: The natural host ranges of many viruses are restricted to very specific
 taxa. Little is known about the molecular barriers between species that
 lead to the establishment of this restriction or generally prevent virus
 emergence in new hosts. Here, we identify genomic polymorphisms in a
 natural rodent host associated with a strong genetic barrier to the
 transmission of the European Tula orthohantavirus (TULV). We analyzed the
 very abrupt spatial transition between two major phylogenetic clades in
 TULV across the comparatively much wider natural hybrid zone between
 evolutionary lineages of their reservoir host, the common vole (Microtus
 arvalis). A genomic scan of 79 225 Single Nucleotide Polymorphisms (SNPs)
 in 323 TULV infected host individuals detected 30 SNPs that were
 associated with specific TULV clades in two replicate sampling transects.
 Focusing the analysis on 199 voles with evidence of genomic admixture at
 the individual level (0.1 - 0.9) supported statistical significance for
 all 30 loci. Host genomic variation at these SNPs explained up to 37.6% of
 clade-specific TULV infections. Genes in the vicinity of associated SNPs
 are involved in functions related to immune response or membrane
 transport. This study demonstrates the relevance of natural hybrid zones
 as systems not only for studying processes of evolutionary divergence and
 speciation, but also for the detection of evolving genetic barriers for
 specialized parasites.
description: Host genotyping : Genotyping by Sequencing
 (GBS) (Elshire et al., 2011) was carried out for all TULV infected common
 voles and additional samples from locations where no TULV infection was
 detected on the Illumina NextSeq platform at Cornell University.
 Restriction enzymes PstI and MspI were used to generate the libraries in
 96-well plates. SNPs were identified and individuals genotyped
 simultaneously using the GBS v2 pipeline (part of the Tassel 5 software)
 (Glaubitz et al., 2014), using a chromosome-level <i>M.
 arvalis</i> genome assembly as the reference sequence (Gouy et al.,
 in prep). Default parameters were used, except that a minimum of five
 reads were required to identify a unique tag.  Genome
 wide association study using GEMMA: The Genome-wide
 Efficient Mixed Model Association (GEMMA) analysis was conducted with the
 v.0.98.1 software (Zhou & Stephens, 2012). We ran GEMMA’s linear
 model on default parameters for the whole dataset of 323 infected
 individuals, as well as separately for the Bavaria transect (190 infected
 individuals) and Porcelain transect (133 infected individuals).
 Association strength of individual SNPs was estimated with GEMMA by
 calculating the Wald test p-value. Only SNPs, which were significantly (p
 &lt; 0.05) associated with clade specific TULV infections across all
 three GWAS (Full data set, Bavaria transect only, Porcelain transect only)
 were considered for further analyses. In order to assess the potential
 impact of imputation on the GWAS results, we removed any of the 323
 individuals, which had any missing data among the significant SNPs, which
 resulted in 165 remaining individuals. In order to further assess the
 possible impact of spatial autocorrelation of vole lineages and virus
 clades, we ran a separate GWAS including only the 199 admixed individuals
 with cluster membership between 0.1 and 0.9.