Comparison between mountain site mean stridulatory peg number and the mean and variance predicted from interpolation between the two valley transects.

<p>Distance = distance in km directly south from the predicted cline centre.</p

List of tested models for each of stridulatory peg number and echeme length, in order of AIC (best-fitting model first).

<p>w = cline width, c = cline centre relative to distances listed in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0037684#pone.0037684.s001" target="_blank">Table S1</a>, b = elevation in trait variance at the zone centre.</p

Parameter estimates for parental means and variances for stridulatory peg number and log(echeme length) and for cline centre, width and variance elevation for each season.

<p>Numbers in brackets after parameter names indicate season. Estimates are from the model with 2 centres, 3 widths and 2 elevations in variance in which cline centre and width were constrained to be identical for peg number and echeme length. Higher negative values for <i>b</i> indicate a greater elevation in trait variance in the zone centre; zero would indicate no elevation in variance. The first parameter of the quadratic (not shown) = (C. jacobsi variance – C. brunneus variance) – <i>b</i>.</p

Stridulatory peg numbers and fitted clines.

<p>Stridulatory peg numbers (grey diamonds) along (a) transect 1 and (b) transect 2 for each temporally separated collection; and fitted clines (solid lines) from the best-fitting model with combined width and centre estimates for song and pegs ± fitted standard deviation (SD) in peg number (thin dashed lines). Thick vertical dashed lines indicate fitted cline centres.</p

Sampling locations in Spain.

<p>Map of northern Spain indicating cline centres (open diamonds) and widths (unbroken lines) along transects 1 (the LE232, to the west) and 2 (the C615 to the east) in season 3 (the season most closely matching collecting periods by Bridle and colleagues). Collecting sites at the northern and southern ends of each transect are indicated by open circles and the four mountain sites (M1 to M4 moving from north to south) by grey-filled circles. Black circles indicate major towns and cities close to the study area. The existing fitted two-dimensional cline centre for stridulatory peg number <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0037684#pone.0037684-Bridle1" target="_blank">[28]</a> is indicated by a dashed line, and the square indicates the study area in which strong seasonality and habitat associations were previously identified <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0037684#pone.0037684-Bailey2" target="_blank">[27]</a>.</p

Test for gene exchange in sympatry.

<p>FST values between Gargano peninsula populations of Spanish sparrows and sympatric (Lesina) and nearby allopatric Italian sparrows.</p><p>indicates highly significant genotypic divergence, <i>P</i><0.0001.</p

AlpsSparrowdata

All individual-level SNP and plumage data used in cline-fitting. This includes: genotypes of 75 SNPs; plumage trait values (discriminant axis scores for multivariate traits) for crown colour, cheek colour and eyebrow colour; a molecular hybrid index based on 75 SNPs; a molecular hybrid index based on 4 SNPs. See publication for more details

Genetic incompatibilities between the hybrid Italian sparrow and its parent species.

<p>(A) Representative geographic cline (<i>ND2</i>) for the mitochondrion and Z-linked genes shifting significantly between the Italian and Spanish sparrows of mainland Italy/Sicily and Sardinia. Colors refer to posterior likelihood of belonging to group corresponding to the Italian sparrow (>0.9, no color) relative to the Spanish sparrow (<0.1, red). The numbers refer to three transects through the Italian-house sparrow hybrid zone in the Alps. Black dots denote sampling locations. (B) Genomic location (in zebra finch) of genes inferred to be involved in hybrid-parent reproductive isolation. Blue outlines denote genes shifting significantly between the Italian and house sparrow, and red outlines denote genes shifting significantly between the Italian and Spanish sparrow. Markers highlighted in yellow have significantly steeper clines (significant β) than the neutral expectation according to a <i>BGC</i>-analysis (see main text) in addition to being significantly skewed towards either hybrid-parent species boundary (significant α), and hence represent the strongest candidate RI genes. Markers in white have significant α only. Chromosomal location for the Z-linked and Chr. 4A genes are indicated. (C) Geographic clines along transect 2 for the three genes shifting significantly in the Italian-house sparrow hybrid zone in the Alps. Upper panel shows results from the Z-linked genes <i>CHD1Z</i>/<i>CETN3</i>, lower panel shows results for the autosomal gene <i>RPS4</i>. Colors refer to posterior likelihood of belonging to group corresponding to the house sparrow (>0.9, white) relative to the Italian sparrow (<0.1, red). Black dots denote sampling locations.</p

Candidate RI genes revealed by genomic cline analysis.

<p>(A–B) Markers are ordered along chromosomes as in <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1004075#pgen.1004075.s008" target="_blank">Table S3</a>. (A) <i>BGC</i> run 2 estimates of genomic cline center (α) with 95% credibility intervals for all 86 SNP markers. Red dots indicate markers with significant excess house sparrow ancestry, blue dots indicate markers with significant excess Spanish sparrow ancestry, grey dots indicate markers with either significant house or Spanish excess ancestry but where the difference in allele frequency between the parent species are below 0.5, and black dots indicate markers that do not differ from neutral expectations. (B) <i>BGC</i> run 2 estimates of genomic cline rate (β) with 95% credibility intervals for all 86 SNP markers. Orange dots indicate markers with clines significantly steeper than neutral expectations, white dots indicate markers with significantly shallower clines than neutral expectations, grey dots indicate markers with either significantly steeper or shallower clines than neutral expectations but where the difference in allele frequency between the parent species is below 0.5, and black dots indicate markers that do not differ from neutral expectations. (C) Genomic cline center (α) plotted against genomic cline rate (β). Red dots indicate markers that exhibit significant excess house sparrow (<i>P. d.</i>) ancestry, clines steeper than neutral expectations and where the difference in allele frequency between the parent species is greater than 0.5. The red dots that are encircled and named shift at the Italian-Spanish boundary and are hence candidate RI genes. Blue dots indicate markers that exhibit significant excess Spanish sparrow (<i>P. h.</i>) ancestry, clines steeper than neutral expectations and where the difference in allele frequency between the parent species is greater than 0.5. The blue dots that are encircled and named shift at the Italian-house boundary and are hence candidate RI genes. Green dots indicate markers that have allele frequency differences between the parent species greater than 0.5 and clines steeper than neutral expectations but do not exhibit excess house or Spanish sparrow ancestry. These markers are candidates for being incompatibilities within the Italian sparrow. Grey dots indicate markers in which the allele frequency difference between the parent species is less than 0.5. (D). Examples of <i>BGC</i> genomic clines representative of the marker categories described in panel (C). For illustrations of all clines, see <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1004075#pgen.1004075.s002" target="_blank">Figure S2</a>.</p

Phenotypic and genetic makeup of the hybrid Italian sparrow.

<p>Coloration of the map denotes phenotypic distribution as indicated by the bird drawings to the right of the map (blue: house sparrow, turquoise: Italian-house hybrids, yellow: typical Italian sparrow, orange: Italian sparrows with plumage intermediate between typical Italian and Spanish sparrows, red: Spanish sparrow). Bird drawings indicate species-specific male plumage characteristics of the three taxa <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1004075#pgen.1004075-SummersSmith1" target="_blank">[16]</a>. Pie charts denote mean hybrid index at sampling localities where white and black color indicate house and Spanish sparrow genetic contribution, respectively. Locations with evidence of recent gene exchange between Spanish and Italian sparrows are indicated by arrows.</p