Weak sperm differentiation in Darwin's finches

The submission contains data sets from a study of sperm evolution in eight species of Darwin's finches from two islands in the Galapagos archipelago. The data sets comprise morphological measurements, sperm size measurements, and genome-wide Single-Nucleotide-Polymorphisms (SNPs). The morphological measurements were used for species confirmation. Sperm size differentiation was analysed in a phylogenetic context where a time-calibrated phylogeny for the species were constructed based on SNP data. The sperm size data were also used to infer the frequency of extrapair paternity. The results indicate that sperm size evolves much more slowly than beak and body size in this radiation, and similar to other songbirds with moderate-to-low levels of extrapair paternity. 

Methods
The field work was carried out in March 2023 on the islands of San Cristobal and Santa Cruz. A total of 106 male birds were sampled for blood and sperm, of which 101 had detectable sperm. Sperm samples were collected by cloacal massage. All birds were released unharmed after sampling and measurements of their wing length and beak dimensions. Sperm samples were fixed in formalin and later photographed at 320x by bright-field microscopy for measurements (10 sperm cells per male). Mean total sperm length per male was used to calculate mean and SD for the populations and pairwise divergences using Hedges' g (Hedges 1981).

A total of 50 blood samples were extracted for DNA and sequenced on the Illumina Novaseq platform (150 bp, paired end). In addition, we downloaded 14 genomes from the sequence-read archive of NCBI (https://www.ncbi.nlm.nih.gov/sra) that originated from San Cristobal (Rubin et al. 2022). After genome assembly and SNP calling, using a high quality Camarhynchus parvulus genome as reference (Rubin et al. 2022), and various filtering, we had a final SNP dataset of 261 754 SNPs scored in all 64 individuals. This data set was the basis for a phylogenetic Principal Component Analysis and an ADMIXTURE analysis (Alexander et al. 2009) for the examination of the population genetic structure. To estimate the divergence time for the major genetic clusters, we performed a Bayesian phylogenetic analysis on a pruned dataset of  50K SNPs using the software SNAPP (Bryant et al. 2012) and 10 individuals representing five clusters or ancestral groups.

Pairwise divergences in sperm length were compared with similar data from other songbird populations (Lifjeld et al. 2024) in relation to their divergence time. 

References:

Alexander, D. H., Novembre, J., & Lange, K. (2009). Fast model-based estimation of ancestry in unrelated individuals. Genome Research,* 19*(9), 1655-1664. https://doi.org/10.1101/gr.094052.109

Bryant, D., Bouckaert, R., Felsenstein, J., Rosenberg, N. A., & RoyChoudhury, A. (2012). Inferring species trees directly from biallelic genetic markers: bypassing gene trees in a full coalescent analysis. Molecular Biology and Evolution,* 29*(8), 1917-1932. https://doi.org/10.1093/molbev/mss086

Hedges, L. V. (1981). Distribution theory for Glass's estimator of effect size and related estimators. Journal of Educational Statistics,* 6*(2), 107-128. https://doi.org/10.3102/10769986006002107

Lifjeld, J. T., Cramer, E. R. A., Leder, E. H., & Voje, K. L. (2024). Sperm as a speciation phenotype in promiscuous songbirds. Evolution,* 79*(1), 134-143. https://doi.org/10.1093/evolut/qpae154

Rubin, C.-J., Enbody, E. D., Dobreva, M. P., Abzhanov, A., Davis, B. W., Lamichhaney, S., Pettersson, M., Sendell-Price, A. T., Sprehn, C. G., Valle, C. A., Vasco, K., Wallerman, O., Grant, B. R., Grant, P. R., & Andersson, L. (2022). Rapid adaptive radiation of Darwin’s finches depends on ancestral genetic modules. Science Advances,* 8*(27), eabm5982. https://doi.org/doi:10.1126/sciadv.abm5982