Phylogenetic and co-ancestry analyses
Phylogenetic analyses based on GBS data were conducted with the main
objective of identifying the source lineage from which the UCSD
population derived. For this test we excluded once again the UCSD n.b.s.
individuals from the ‘Neutral Dataset’. A maximum likelihood phylogeny
and a neighbor-net were produced using the programs IQ-TREE (Nguyenet al. 2015) and SplitsTree (Bryant & Moulton 2004),
respectively. For the former, the ascertainment bias correction and the
generalized time-reversible (GTR) model were implemented.
We further tested the origin of the UCSD junco population by exploring
pairwise levels of co-ancestry among Oregon junco forms with the program
RADpainter (Malinsky et al. 2018). Starting from the ‘Full
Dataset’, UCSD n.b.s. individuals were excluded. To detect subtler signs
of differentiation, and because this program relies on linkage
disequilibrium to achieve more refined assignment of nearest- neighbor
relationships, we did not apply neutrality and linkage filters in this
analysis. We did, however, increase the per-site missing data threshold
to 0.75 as recommended by the authors, and applied the same MAF and HWE
filters as in the PCA and STRUCTURE analyses, resulting in a matrix of
41,231 SNPs. We ran the analysis with 100K burn-in and 100K sampling
iterations, and a thinning interval of 1,000 to produce a matrix of
pairwise individual co-ancestry estimates, which were averaged over
forms. For estimating 95% confidence intervals around pairwise
co-ancestry values between UCSD and the different Oregon junco forms, we
applied a non-parametric bootstrap procedure. Bootstrapping was carried
out by splitting the SNP matrix in 1,000 blocks and randomly combining
them for each one of the repetitions. A total of 100 analyses were run
for confidence interval estimation.
Breeding latitude of UCSD juncos captured during the non-breeding
season Hydrogen isotopic ratios in bird feathers correspond to those found in
the localities where the feathers were grown, and since juncos molt
their feathers at the end of the breeding season, isotopic ratios can be
used to determine relative differences in breeding latitude (Chamberlainet al. 1996). To determine whether UCSD birds sampled in the
non-breeding season (UCSD n.b.s.) were local breeders or winter visitors
breeding elsewhere, we tested for correlation between deuterium isotope
ratio (δD) and posterior likelihoods of genetic population assignment.
We obtained isotopic data from the 28 individuals sampled in UCSD over
winter, to compare to two confirmed year-round residents used as
controls. A single secondary feather was collected per bird and washed
using a 2:1 chloroform: methanol solution. Approximately 0.50 mg of vane
material was clipped from the proximal end of each feather and the
hydrogen isotope composition was measured by continuous flow isotope
ratio mass spectrometry using a Finnigan TC/EA interfaced to a Finnigan
DeltaPlus XL mass spectrometer (Thermo Scientific, Bremen, Germany), as
described in Wunder et al. (2012). δD values are reported in per
mil notation (‰) relative to V-SMOW, using internal standards (-78 ‰ and
-172 ‰ respectively) calibrated to CFS-CHS-BWB (Wassenaar & Hobson
2003). Benzoic acid, (δD = −61‰) and IAEA-CH-7 (δ2H = −100‰) were also
analyzed within analytical sequences with a precision of <
+/-4‰. A SNP subset of the ‘Neutral Dataset’ was generated for 26 of the
same 30 individuals (four failed to pass missing data filters, Table 1),
and assignment likelihoods were computed with STRUCTURE under the same
conditions, in this case only for K = 2. Correlation was tested by means
of a linear regression between STRUCTURE scores and δD abundance.