A short detective story on the reliability of this peculiar cross.
At the moment, I am exploring the wonderful world of woodpecker hybrids. Checking the Handbook of Avian Hybrids of the World by Eugene McCarthy, I came across a peculiar record: Chestnut-colored Woodpecker (Celeus castaneus) x Golden-olive Woodpecker (Piculus rubiginosus). These species look quite different and diverged more than 11 million years. Never say never when it comes to hybridization in birds, but I was skeptical about this cross. The explanation in the Handbook refers to Miller et al. (1957) who note “an old paper reports a hybrid from Atoyac.” So, I decided to retrace this old paper and verify its reliability.
The study by Miller et al. (1957) turned out to be a checklist of Mexican birds. Below the record of Celeus castaneus, I noticed a footnote that refers to the subspecies Piculus rubiginosus yucatanensis. Although the footnote is located beneath Celeus castaneus, it has nothing to do with this species. So, I assume that McCarthy accidently linked the footnote to Celeus castaneus and inferred a possible hybrid with Piculus rubiginosus. Hence, not a reliable record.
This ornithological detective story highlights the importance of checking the reliability of bird hybrids. Always return to the original source and assess the evidence directly. In this case, it turned out that McCarthy probably made a mistake while reading the report by Miller and colleagues. When preparing a manuscript in tinamou hybrids, I also noticed that several hybrid records were based on anecdotes and speculation about potential interbreeding. Hardly convincing evidence. That is why it is always a good idea to read the original source when assessing the reliability of avian hybrids.
McCarthy, E. M. (2006). Handbook of Avian Hybrids of the World. Oxford university press.
Miller, A. H., Friedmann, H., Griscom, L., & Moore, R. T. (1957). Distributional check-list of the birds of Mexico. Part II. Pacific Coast Avifauna, 33(1), 435.
Cheap molecular markers can help to identify the origin of birds that died as bycatch.
“Conserving biodiversity, then, means more than preventing the extinction of a species. It also means preventing loss of genetic diversity within that species.” This statement comes from a recent article at The Conversation by Laura Bertola, and I wholeheartedly agree. However, in order to protect this genetic diversity, we need to know how it is distributed across a species. Is there one genetically homogeneous population, or are there many small pockets with unique genetic variation? This question is particularly relevant for seabirds, such as albatrosses, which can explore large oceanic regions without any physical barriers. You might expect to find little genetic differentiation between different breeding colonies, because birds can easily soar between islands. Or perhaps there are subtle genetic differences between colonies as birds tend to stay loyal to their breeding grounds. Depending on the situation, different conservation measures are needed to protect these birds. A recent study in the journal Conservation Genetics to a closer look at two endangered species: the Atlantic Yellow-nosed Albatross (Thalassarche chlororhynchos) and the Indian Yellow-nosed Albatross (Thalassarche carteri).
Dilini Abeyrama and her colleagues collected blood samples from different breeding colonies across the range of both species (see map below). Genetic analyses based on a set of 13 microsatellites uncovered two clear clusters, corresponding to the Atlantic and the Indian Yellow-nosed Albatross. Within each species, the researchers detected more fine-scale patterns of population structure. In the Atlantic Yellow-nosed Albatross, for example, Gough Island birds were genetically distinct from birds that breed on the Inaccessible and Nightingale Islands. The separation between the Atlantic and the Indian Yellow-nosed Albatross was also obvious in the mitochondrial data, revealing two clusters separated by six mutations. Based on these patterns, the researchers argue that both species should be treated as separate units for conservation and management. Several island populations will probably need to be considered as distinct conservation units as well, but the population structure within each species requires further investigation with genomic data.
These findings cannot only inform future conservation measures, they also have a direct practical application. Every year, albatrosses die as bycatch in fishery activities. Because Atlantic and Indian Yellow-nosed Albatross are morphologically similar, it is often not possible to identify the right species (let alone pinpoint their island of origin). Here, the genetic data come in handy.
The species-specific primers developed in our study allowed us to differentiate Atlantic and Indian yellow-nosed albatrosses with a high accuracy and low cost. Using this knowledge, we can develop a rapid molecular test to identify the bycatch yellow-nosed albatross samples which cannot be identified from their morphology.
Knowing how birds from different breeding colonies move across the ocean can provide insights into the behavior and ecology of these species. Invaluable knowledge that can be used to improve the conservation of these iconic species.
Abeyrama, D. K., Dempsey, Z. W., Ryan, P. G., & Burg, T. M. (2021). Cryptic speciation and population differentiation in the yellow-nosed albatross species complex. Conservation Genetics, 22(5), 757-766.
Or could the genetic patterns be explained by incomplete lineage sorting?
Last year, I published a short paper on tinamou hybrids in which I introduced a scoring scheme to assess the reliability of hybrid records (see this blog post for a summary). In short, I assigned points for different criteria, namely:
Observation of a putative hybrid with photographic evidence or a detailed description (1 point)
Thorough morphological analyses in which the putative hybrid is compared with potential parental species (2 points)
Genetic analyses of the putative hybrid with reference material from potential parental species (3 points)
I decided to put most weight on genetic evidence, because it can be difficult to confidently identify hybrids based on morphology. A specimen with aberrant plumage might be a hybrid, but could also be a color morph. Genetic analyses can often resolve such morphological mysteries. This is nicely illustrated by a recent study in the journal PLoS ONE in which researchers provide some genetic evidence for hybridization between two Sporophila species.
Scoring Plumage Patterns
An extensive survey of Copper Seedeater (Sporophila bouvreuil) and Pearly-bellied Seedeater (Sporophila pileata) across Brazil uncovered a large variation in plumage patterns. The researchers noted that across several Brazilian states “more than one plumage color occurred”, which they explained by age-related changes in coloration. However, the presence of some intermediate plumage patterns points to the possibility of hybrid individuals. To test this hypothesis, Cesar Medolago and his colleagues turned to genetic data by sequencing the mitochondrial gene COI and several microsatellites across a putative contact zone.
First, the researchers scored the plumage coloration of individuals birds from 1 to 4. Individuals were considered as parental species with a score of 1 or 4, and intermediates when scoring 2 or 3. Next, they constructed an evolutionary tree with the mitochondrial data and checked the positions of the intermediate birds in this phylogeny. In general, intermediate birds with score 2 were closer to Pearly-bellied Seedeater, whereas intermediate birds with score 3 were found in a cluster with Copper Seedeater. However, the combination of plumage pattern and mtDNA was not perfect, suggesting that some individuals might represent backcrosses.
Incomplete Lineage Sorting
Based on the microsatellites, the researchers determined the parents at several nests. The analyses revealed all possible parental combinations, including Pearly-bellied Seedeater couples (3 nests), Copper Seedeater couples (3 nests), and mixed couples (6 nests). In four nests, the male was a Pearly-bellied Seedeater and the female a Copper Seedeater. And in two nests, it was the other way around. These findings suggest ongoing hybridization between these two species.
There is, however, an important issue to discuss here. The genetic patterns seem to suggest hybridization, but there is another possibility: incomplete lineage sorting. Imagine a bowl filled with red and blue marbles (representing an ancestral population). As you pour this bowl into two smaller ones (representing the two Seedeater species), you will not get a perfect division between red and blue marbles. A similar process occurred during the speciation process of these birds, some genetic variation present in the ancestral population was incompletely sorted into the two lineages. The sharing of mitochondrial or nuclear variants can thus be explained by incomplete lineage sorting. And indeed, a genomic study on the genus Sporophila suggested that a recent radiation resulted in high levels of incomplete lineage sorting.
Nonetheless, the authors from the present study make a convincing case for the possibility of hybridization. They nicely summarized the supporting evidence at the beginning of the discussion:
The main evidence include: i) the widely disjunct distribution of the two species with records of individuals with intermediate plumage patter concentrated near the contact zone; ii) the similar proportion of haplotypes belonging to pileata and bouvreuil clades only in the area embedded within the contact zone, the only of our three areas in which we found the two typical parental plumage patterns together; iii) the presence of males with intermediate plumage in both of the well-supported clades; iv) the decreasing frequencies of males with intermediate plumage and of mtDNA haplotypes that are exclusive of the bouvreuil clade in the direction of the core area of pileata, and v) the fact that intermediate plumage patterns occur frequently in nature only in this pair of southern capuchinos, differently from the other congeners that do reproduce in sympatry.
Medolago, C. A., Costa, M. C., Silveira, L. F., & Francisco, M. R. (2020). Hybridization between two recently diverged neotropical passerines: The Pearly-bellied Seedeater Sporophila pileata, and the Copper Seedeater S. bouvreuil (Aves, Passeriformes, Thraupidae). PloS one, 15(3), e0229714.
Although phylogenetic relatedness plays a major role as well.
Several species of wood warbler (family Parulidae) migrate at night. During their nightly travels, they produce short, simple calls known as “flight calls”. In some species, these flight calls are so similar that they have been grouped into bio-acoustic categories, such as the “Zeep complex” and the “Upsweep complex.” Interestingly, members of these complexes tend to exhibit similar migration strategies. Could there be an evolutionary link between these traits? Given that wood warblers often migrate in mixed species flocks, shared flight calls might facilitate interspecific communication during migration. In a recent Evolution study, Zach Gayk and his colleagues tested this hypothesis by comparing the flight calls and migration strategies of 36 wood warbler species.
The researchers quantified the similarity of flight calls in the different species and correlated this with several features of migration, such as overlap in wintering range and total migration distance. The analyses indicated that phylogenetic relatedness explains a large part of the variation in flight calls (similar to this study on the evolution of bird song). In other words, closely related species produce similar flight calls. After correcting for phylogenetic relationships, some migratory variables were associated with similarity in flight calls. When considering all 36 species, flight calls were similar between species that breed at similar latitudes and that show a temporal overlap in migration timing. A more detailed analysis on the “Zeep complex” found significant effects of migration length and overlap in wintering range on flight call similarity. Taken together, these findings support the hypothesis that migratory similarity is a driving factor in the evolution of flight calls.
Intuitively, these findings make sense. Efficient communication during migration can be a life-saver. Literally. Similar flight calls can help birds to find high-quality stop-over sites, avoid predators, and reduce the chance of getting lost during migration. The researchers suspect that selection for similar flight calls was quite strong during the last 50,000 years of glacial cycles when “shifting migratory routes may have driven the need for increased communication.” However, the link between flight calls and migratory strategies remains to be tested in other bird species.
Finally, I cannot help but think of a possible role for flight calls in hybridization. Wood warblers are known for their propensity to hybridize (see the Parulidae page for an overview). Could this be related to the similarity in flight calls or migration strategies? Disentangling these two factors will be tricky, but worthwhile to explore.
Gayk, Z. G., Simpson, R. K., & Mennill, D. J. (2021). The evolution of wood warbler flight calls: Species with similar migrations produce acoustically similar calls. Evolution, 75(3), 719-730.
Genetic study reconstructs the evolutionary history of this Central American species.
In the classic verbal model of allopatric speciation, a geographic barrier arises and splits a population in two. Over time, these isolated population diverge genetically and slowly evolve into distinct species. So, when you observe a geographic barrier, such as a river or mountain chain, between two species, it is tempting to infer that this barrier played a role in the speciation process. But this does not have to be the case. The barrier might have formed millions of years before the divergence of the species. Other evolutionary, ecological or geological processes might explain the origin of these species. This situation is nicely illustrated by a recent study on the Azure-crowned Hummingbird (Amazilia cyanocephala) which appeared in the Journal of Ornithology.
The Azure-crowned Hummingbird can be found from the north of Mexico to Nicaragua and has been divided into three subspecies: cyanocephala, guatemalensis and chlorostephana. Using a mitochondrial marker, Flor Rodriguez-Gómez and her colleagues detected four genetic lineages within this hummingbird species. Within the subspecies cyanocephala, there is a clear break between populations east and west of the Isthmus of Tehuantepec. A third genetic lineage corresponds to the subspecies guatemalensis and is separated from the other subspecies by the Motagua-Polochic-Jocotán fault system. Finally, the fourth genetic lineage (represented by the subspecies chlorostephana) can be found in the savannah pine forests of the Moskitia region in Honduras.
These patterns suggest that the geographic barriers across Central America – the Isthmus of Tehuantepec and the Motagua-Polochic-Jocotán fault system – drove the evolution of the Azure-crowned Hummingbird. But this scenario is not supported by the divergence times between the four genetic lineages. They originated within the last million years, whereas these geographic barriers arose much earlier (the current geological shape of the Isthmus of Tehuantepec probably formed about 6 million years ago). What factors did influence the evolution of the Azure-crowned Hummingbird?
Additional genetic analyses detected signatures of a population expansion before the last glacial maximum (about 20,000 years ago). This finding was corroborated by ecological niche modelling which indicated that the suitable habitat for the Azure-crowned Hummingbird was larger during this period. However, during the last inter-glacial (about 120,000 to 140,000 years ago), the distribution of suitable habitat was more fragmented, with major regions separated by the Isthmus of Tehuantepec and other smaller regions isolated in Honduras and Nicaragua. Together, these patterns point to cycles of contraction and expansion driven by climatic changes. Indeed, the authors conclude that:
Likely, phylogeographic structure in Azure-crowned Hummingbird was promoted by habitat fragmentation during interglacials and, if so, competitive exclusion in zones of contact and habitat specialization played a greater role on differentiation among the three subspecies than isolation by geographic barriers.
Rodríguez-Gómez, F., Licona-Vera, Y., Silva-Cárdenas, L., & Ornelas, J. F. (2021). Phylogeography, morphology and ecological niche modelling to explore the evolutionary history of Azure-crowned Hummingbird (Amazilia cyanocephala, Trochilidae) in Mesoamerica. Journal of Ornithology, 162(2), 529-547.
Researchers follow the evolution of reproductive isolation in two contact zones.
When two species start hybridizing after a period of geographic isolation several scenarios are possible. Barriers of reproductive isolation might break down and the species collapse into one panmictic population. Or new interactions between the hybridizing species might push reproductive isolation towards completion. In most cases, we can only guess what happened in the past by studying present-day patterns of genetic variation. In my own work, for example, I reconstructed the evolutionary history of two Bean Goose species that established secondary contact about 60,000 years ago. It seems that these geese are in the merging into one species, but it is tricky to draw conclusions on a process that could take thousands to millions of years. Sometimes, however, we come across a situation where we can directly study different stages of speciation process. In a recent study in the journal Molecular Ecology, researchers could compare different contact zones between two Empidonax flycatchers to understand how reproductive isolation evolves between these species.
Willow Flycatcher (E. traillii) and Alder Flycatcher (E. alnorum) interbreed across two contact zones in North America: a broad overlapping area in the east (more than 1000 kilometers) and a narrow one (less than 200 kilometers) in the west. Jordan Bemmels, Ashley Bramwell and their colleagues used whole genome data to investigate patterns of introgression at these contact zones. In the western zone, there was clear evidence for introgression (2.4-8.2%), while no admixture was detected in the eastern zone. These patterns indicate that reproductive isolation is strong in the east and still incomplete in the west. Additional analyses revealed that the western contact zone is of recent origin, whereas the formation of the eastern contact zone can be traced back to the last glacial maximum. Birds in the east have thus had more time to build-up reproductive isolation.
The traits underlying reproductive isolation between these flycatchers remain to be determined. These traits can often be identified by looking for signatures of divergent character displacement. For instance, selection for reduced competition could result in differences in habitat use or beak morphology when species adapt to distinct food sources. Selection can also directly contribute to reproductive isolation. If hybridization is maladaptive, traits involved in species recognition will evolve along different trajectories to prevent birds from hybridizing. The researchers tested several traits, such as beak morphology and the colors of crown feathers, but found no clear evidence for character displacement. Moreover, the species did not appear to differ in habitat use or timing of breeding.
However, there are still plenty of other traits that can be tested, such as differences in song or sperm morphology (see this blog post). The researchers also mentioned that “the genomes of the two species are well differentiated, with numerous Fst peaks that occur on almost every chromosome.” These peaks in genetic differentiation could contain some interesting candidate genes that might point to the traits underlying reproductive isolation between Willow Flycatcher and Alder Flycatcher.
Bemmels, J. B., Bramwell, A. C., Anderson, S. A., Luzuriaga‐Aveiga, V. E., Mikkelsen, E. K., & Weir, J. T. (2021). Geographic contact drives increased reproductive isolation in two cryptic Empidonax flycatchers. Molecular Ecology, 30(19), 4833-4844.
Although the domestic chicken is one of the most studied birds ever, the evolutionary history of its genus (Gallus) is still a mystery. The four species in this genus – Sri Lanka Junglefowl (G. lafayetti), Grey Junglefowl (G. sonneratii), Green Junglefowl (G. varius) and Red Junglefowl (G. gallus, the ancestor of the domestic chicken) – can be arranged in no less than 15 different topologies. Early genetic studies, using mitochondrial DNA or a few nuclear markers, reported evidence for six of these topologies. And a recent analysis of ultraconserved elements (UCEs) added a seventh possibility to the list. You can see an overview of these seven evolutionary arrangements in the figure below.
The discordant results among previous genetic studies can be explained by several factors. First, introgression can lead to patterns that deviate from the main evolutionary history (i.e. the species tree). Extensive gene flow from one species into another can pull distantly related species together in an evolutionary tree. Second, the random sampling of gene trees can cause issues. It is known that different genes tell different evolutionary stories. If you happen to sample a set of molecular markers that do not follow the species tree, you will draw the wrong conclusions. Third, some genetic markers might have too little phylogenetic information to confidently resolve phylogenetic relationships. These three issues can be addressed with whole genome sequences: there is plenty of data available and detailed analyses of gene trees can detect signatures of introgression and random sampling biases.
Recently, two genomic studies appeared that tried to resolve the Gallus phylogeny. The first study – published in the journal Avian Research – focused on the effects of different datatypes. George Tiley and his colleagues performed phylogenetic analyses on different molecular markers, namely protein-coding exons, introns, UCEs and conserved non-exonic elements (CNEEs). Interestingly, all markers converged upon the same topology in which the Green Junglefowl diverged first, followed by the Red Junglefowl. The remaining two species – Sri Lanka Junglefowl and Grey Junglefowl – are sister species.
Moreover, the researchers took a closer look at the distribution of the gene trees. As expected, the most common gene tree (36%) reflected the topology described above. The remaining topologies can provide important insights into introgression dynamics. If there has been no introgression, two minor gene trees are expected to occur in roughly equal frequencies (similar to flipping a coin). Introgression, however, leads to a bias towards one minor gene tree. Additional analyses on gene tree distributions revealed several of such biases that pointed to introgression between Red Junglefowl and Green Junglefowl, and between Red Junglefowl and Grey Junglefowl.
NJ vs. ML
To answer the question in the title: the Green Junglefowl evolved first. Well, not so fast. Because a second genomic study in the journal Molecular Phylogenetics and Evolution suggests otherwise. Phylogenetic analyses of more than 20,000 gene sequences resulted in two main topologies depending on the applied method. A Neighbor-Joining (NJ) approach indicated that the Green Junglefowl evolved first, whereas a Maximum Likelihood (ML) approach pointed to Red Junglefowl as the first species to diverge. Introgression analyses on the NJ-tree – using the popular D-statistic – revealed extensive gene flow from the Green Junglefowl into the ancestor of the Sri Lanka Junglefowl and Grey Junglefowl: no less than 27.6%. The researchers consider this amount of introgression unlikely, which invalidates the NJ-topology. Hence, the ML-tree with more realistic levels of introgression – and where Red Junglefowl evolved first – represents a more likely scenario.
So, now what? The two genomic studies report contrasting results and we still don’t know which species evolved first. Personally, I find the first study more convincing because it took different datatypes into account and provided a detailed overview of the gene trees. The second study, on the other hand, analyzed all autosomal gene trees in one go and did not report the distribution of these gene trees (I am very curious to see it). My bet would thus be on the Green-Junglefowl-first-scenario.
However, until now we assumed that the evolutionary history of the genus Gallus can be captured in a bifurcating tree. This is not necessarily the case. The high levels of ancient and recent introgression between these species might be better depicted in a phylogenetic network (see for example here). Trying to find the “true” species tree could be seen as a wild goose (or chicken) chase. And the question which junglefowl species evolved first becomes a bit nonsensical.
Mariadassou, M., Suez, M., Sathyakumar, S., Vignal, A., Arca, M., Nicolas, P., … & Tixier-Boichard, M. (2021). Unraveling the history of the genus Gallus through whole genome sequencing. Molecular Phylogenetics and Evolution, 158, 107044.
Tiley, G. P., Pandey, A., Kimball, R. T., Braun, E. L., & Burleigh, J. G. (2020). Whole genome phylogeny of Gallus: introgression and data-type effects. Avian Research, 11(1), 1-15.
The importance of different processes seems to vary per location.
Hybridization often leads to introgression, the exchange of genetic material between the interacting species. The resulting patterns of introgression are determined by the complex interplay between numerous factors. First, local population sizes can affect hybridization rates. When one species is rare, its members will have more difficulty finding a mate and they might settle with a partner from another species. This phenomenon – known as Hubb’s Principle – has been nicely illustrated on the Falkland Islands where a numerical imbalance between Speckled Teal (Anas flavirostris) and Yellow-billed Pintails (Anas georgica) led to hybridization. Next, selection on hybrids might play a role. The selection pressure might be endogenous, affecting the fertility or viability of the hybrids. Or hybrids might have to deal with exogenous selection when they are badly adapted to local environmental conditions. Disentangling all these different factors is a challenging endeavor. But that did not stop Logan Maxwell and colleagues from studying how neutral and selective processes determine introgression patterns between Saltmarsh Sparrow (Ammospiza caudacuta) and Nelson’s Sparrow (Ammospiza nelsoni). Their findings recently appeared in the journal BMC Ecology and Evolution.
The researchers genotyped more than 500 birds at two locations (coastal and inland) along the coast of New England. The genetic data allowed them to determine the number of “pure” individuals, first-generation hybrids and backcrosses. If introgression is mainly determined by neutral processes, such as local population sizes, you would expect the distribution of genotypes to follow a random pattern. This was not the case: the coastal site had less backcrossed Saltmarsh Sparrows than expected while at inland site there were more hybrids, backcrosses and “pure” Saltmarsh Sparrows than predicted. These findings suggest that neutral demographic processes are not sufficient to explain introgression rates in this hybrid zone. Selection plays a role as well.
But is the selection on hybrids endogenous or exogenous? The results suggest that both types of selection are at work. The researchers detected a reduction in hybrid females among the adults. Because there was no difference in viability of females at the egg stage, it seems that hybrid females have a lower chance of survival. This pattern is in accordance with Haldane’s Rule which states that in a hybrid cross the sex with two different sex chromosomes (i.e. the female in birds) will suffer the greatest fitness reduction. In addition, there were more individuals with Saltmarsh Sparrow DNA at the coastal site, suggesting that genetic variants from this species provide an adaptive advantage in that area (see also this blog post). There is thus exogenous selection against Nelson’s Sparrows at the coast.
Finally, the researchers tested whether the birds mated assortatively in the hybrid zone (i.e. finding a partner of the same species). In general, this was certainly the case: the majority of mating events (79%) occurred between the same species. However, the patterns differed by site. Assortative mating was strong at the coastal site, but not at the inland site. The inland population is smaller which could increase the frequency of hybridization there. The exact mechanisms underlying mate choice remain to be determined, but could be related to differences in song and mating behavior.
All in all, the hybridization and introgression dynamics between Saltmarsh and Nelson’s Sparrow are determined by the complex interplay of numerous factors which differ between locations. The authors nicely summarized the situation at the beginning of the discussion-section:
We found that neutral demographic factors—relative abundances of the two species—alone could not explain the observed patterns of introgression between Saltmarsh and Nelson’s Sparrows and that spatial variation in the distribution of parental and offspring genotypes was a result of both exogenous and endogenous selective forces. In addition, sexual selection played a role in maintaining species boundaries through assortative mating. However, these patterns differed on the coastal and inland site, suggesting local differences in the strength of selection.
Maxwell, L. M., Walsh, J., Olsen, B. J., & Kovach, A. I. (2021). Patterns of introgression vary within an avian hybrid zone. BMC Ecology and Evolution, 21(1), 1-18.