Biological specimens in natural history collections constitute a massive repository of genetic information. Many specimens have been collected in areas in which they no longer exist or in areas where present-day collecting is not possible. There are also specimens in collections representing populations or species that have gone extinct. Furthermore, species or populations may have been sampled throughout an extensive time period, which is particularly valuable for studies of genetic change through time. With the advent of high-throughput sequencing, natural history museum resources have become accessible for genomic research. Consequently, these unique resources are increasingly being used across many fields of natural history. In this paper, we summarize our experiences of resequencing hundreds of genomes from historical avian museum specimens. We publish the protocols we have used and discuss the entire workflow from sampling and laboratory procedures, to the bioinformatic processing of historical specimen data.

Despite large differences in morphology, behavior and lek-mating strategies the birds-of-paradise are known to hybridize occasionally, even across different genera. Many of these bird-of-paradise hybrids were originally described as distinct species based on large morphological differences when compared to recognized species. Nowadays, these specimens are generally recognized as hybrids based on morphological assessments. Having fascinated naturalists for centuries, hybrid specimens of birds-of-paradise have been collected and the specimens kept in Natural History Collections. In the present study, we utilize this remarkable resource in a museomics framework and evaluate the genomic composition of most described intergeneric hybrids and some intrageneric hybrids. We show that the majority of investigated specimens are first-generation hybrids and that the parental species, in most cases, are in line with prior morphological assessments. We also identify two specimens that are the result of introgressive hybridization between different genera. Additionally, two specimens exhibit hybrid morphologies but have no identifiable signals of hybridization, which may indicate that minor levels of introgression can have large morphological effects. Our findings provide direct evidence of contemporary introgressive hybridization taking place between genera of birds-of-paradise in nature, despite markedly different morphologies and lek-mating behaviors.

Hybridisation has sometimes been viewed as an evolutionary dead end due to the purported reduced fitness of hybrids. Alternatively, hybridisation can also be viewed as an evolutionary conduit in which adaptive or neutral variants can be exchanged. Hybridisation has been reported in a wide range of taxa, including organisms under strong sexual selection due to elaborate male courtships and lek-mating. Whether occasional hybridisation can overcome the reproductive barriers in such systems and lead to the establishment of hybrid zones is poorly known. In this study, we address these questions by focusing on one of the most well-known avian radiations where lekking has resulted in an extraordinary assemblage of phenotypic diversity; the birds-of-paradise (Paradisaeidae). We explore this remarkable system by quantifying the genome-wide distribution of introgression. We find multiple signals of historical gene flow between and within two genera of birds-of-paradise. In addition, we present the first empirical genomic evidence of a hybrid zone between two lekking bird-of-paradise species that differ substantially in their sexually selected traits and behaviours. Our findings are surprising evidence of weak reproductive barriers that allow for continuous gene flow between species that are otherwise expected to be highly constrained by their lekking and male courtship behaviours.

Hybridisation can introduce genetic variation at a faster rate than denovo mutations alone, thus it is recognised as facilitating the exploitation of new niches in many species radiations. The birds-of-paradise is a well-known avian species radiation, with most species endemic to the island of New Guinea. The family is known for extraordinary phenotypic diversity and elaborate courtship rituals. Prior genomic research on birds-of-paradise has indicated widespread gene flow between divergent species of the family. However, if genetic material transferred through introgression has facilitated the evolution of phenotypic traits remains unanswered. In this study, we assemble a population level dataset for eleven species from four separate genera of birds-of-paradise. The species were chosen as hybridisation between many of the focal species have been seen on genome wide level, as well as a few contemporary F1-hybrids have been reported among them. In the species comparisons with the highest levels of genome wide d-statistics, we use window based test of treeness to find regions with excess allele sharing and investigate the genes present in these regions and their function. Some of the genes present in the candidate introgressed windows have been shown to be important for phenotypic traits, such as beak and feather morphology. In addition, we find one of the investigated species to have high genome wide levels of admixture to all focal species, while having low overall levels of shared allele proportions with the focal species indicating heavy hybridisation with an out group, and a potential hybrid origin of the species.