How strong is reproductive isolation between Golden-winged and Blue-winged warbler?

A summary of a recent debate in the journal Ecology and Evolution.

The future of the Golden-winged Warbler (Vermivora chrysoptera) is threatened by habitat loss. In addition, it runs the risk of being outcompeted and “out-hybridized” by the invading Blue-winged Warbler (V. cyanoptera). The interactions between these two closely related species have a long history of scientific research (summarized on the Parulidae page). However, it is still unclear how strong the level of reproductive isolation between these warblers is. Recent work by David Toews and his colleagues pointed to six genomic regions that are highly divergent between Golden-winged and Blue-winged Warblers, of which four are likely involved in feather development or pigmentation. These findings suggest that differences in plumage patterns could act as a strong reproductive barrier. The strength of this potential barrier can be tested by quantifying the frequency of mixed pairings between different plumage types, and following the reproductive success of hybrids (if there are any).

A recent study in the journal Ecology and Evolution performed these measurements and reported strong reproductive isolation between Golden-winged and Blue-winged Warblers. However, another team of researchers questioned these results and indicated potential pitfalls in the analyses, to which the original authors responded. In this blog post, I will try to summarize the main arguments in this interesting debate.

Strong Reproductive Isolation?

Let’s start with the first study. To determine the degree of reproductive isolation between Golden-winged Warbler and Blue-winged Warbler, John Confer and his colleagues aggregated data on social pairing from nine studies. Apart from the two pure phenotypes, the researchers also considered two hybrid phenotypes: the “Brewster’s Warbler” and the “Lawrence’s Warbler”. These phenotypes were initially described as distinct species before they were recognized as hybrids. According to the model of Kenneth Parkes (1951), “Brewster’s Warblers” are first-generation hybrids between genetically pure Golden-winged and Blue-winged Warblers, while the “Lawrence’s Warbler” can be produced by crossing two first-generation hybrids.

The analyses revealed a low level of hybridization. Only 14 out of 1680 (0.9%) Golden-winged Warblers and 14 out of 583 (2.4%) Blue-winged Warblers paired up with another phenotype. These patterns indicate high levels of behavioral isolation between the different plumage phenotypes. Next, the researchers turned to the breeding success of the hybrid phenotypes. The pairing success of “Brewster’s Warblers” (54%) was significantly smaller compared to the pure Golden-winged (83%) and Blue-winged Warblers (77%). These percentages suggest some degree of sexual selection against hybrids. Putting it all together, the researchers calculated a reproductive isolation score of 0.96. Given that a score of 1 corresponds to complete reproductive isolation, this number indicates strong reproductive isolation.

The different phenotypes considered in the study: Golden-winged Warbler (V. chrysoptera; GWWA), Blue-winged Warbler (V. cyanoptera; BWWA), “Brewster’s” Warbler (hybrid; BRWA) and “Lawrence’s” Warbler (hybrid LAWA). From: Confer et al. (2020) Ecology and Evolution.

Three Points of Critique

A few months later, David Toews and his colleagues published a critique on this conclusion of strong reproductive isolation, raising three main issues. First, the plumage classification scheme in the original study is not suitable to determine hybrid ancestry in these warblers. Recent genetic work by Marcella Baiz and her colleagues showed that none of the six “Brewster’s Warblers” that they analyzed were first-generation hybrids (see this blog post for the details). Moreover, many warblers that look like pure phenotypes might actually contain some genetic ancestry from past hybridization. The original study did not take these “cryptic hybrids” into account.

A second issue that was not considered in the analyses concerns extra-pair copulations in which birds mate with other individuals besides their social partner. This phenomenon has been well-documented in Vermivora warblers and could significantly contribute to hybridization between Golden-winged and Blue-winged Warblers.

Finally, Toews et al. (2021) pointed out that behavioral isolation is not always sufficient to maintain complete reproductive isolation. For example, recent simulations by Darren Irwin showed that assortative mating on its own cannot prevent populations from merging, some form of postzygotic isolation is needed (see this blog post for the whole story). Although “Brewster’s Warblers” have lower pairing success compared to pure phenotypes, their reproductive output might still be too high to prevent genetic exchange. Hence, the authors of the critique argue that “extensive mixing in areas of sympatry is more consistent with low levels of total reproductive isolation—that is, both low pre-and postmating isolation—and results in high gene flow.”

The range of Golden-winged (orange) and Blue-winged (blue) Warblers. Areas of overlap are highlighted in light blue. From: Toews et al. (2021).

Counterarguments

Recently, the authors of the original study – this time led by Cody Porter – replied to the critique by Toews et al. (2021). First, with regard to the unsuitability of the plumage classification scheme, they explain that the complex genetic ancestry of the warblers (including cryptic hybrids) is actually not that relevant for their question. The focus of their study concerns different plumage phenotypes, not the whole genomic context. They write: “In essence, our study could be viewed as testing whether the six major genomic differences between V. chrysoptera and V. cyanoptera (which largely correspond to plumage differences; Toews et al., 2016) promote reproductive isolation.”

Second, they argue that extra-pair copulations were unlikely to bias their results, referring to the findings of Vallender et al. (2007). This study found only 3 cases of extra-pair copulations (ca. 1.5%) between different phenotypes. In two cases a hybrid female mated with a Golden-winged Warbler and in one case a Golden-winged Warbler female mated with a hybrid.

The third point of critique focuses on the contribution of behavioral isolation to the level of reproductive isolation. You need some degree of postzygotic isolation to prevent species from merging. Toews et al. (2021) argued that the reproductive success of the hybrids is still too high, facilitating gene flow between the species. The authors counter this argument by highlighting the 26% reduction in the pairing success of phenotypic hybrids compared to both parental forms and the fact that only 1.2% of birds with a “pure” phenotype paired with an individual of the alternative phenotype. These numbers “appear to fall well within the parameters for a stable hybrid zone according to Irwin’s (2020) simulations.”

The Verdict

You might be wondering who won this debate? I don’t think this is the right question to ask here. Both groups of authors approached the scientific conundrum of reproductive isolation from a different perspective. The original study focused on behavioral isolation on the phenotypic level, whereas the critique used the genomic patterns of introgression as a starting point. At first sight, the strong reproductive isolation between plumage phenotypes seems incompatible with the largely homogeneous genomes of these warblers. However, reproductive isolation is not complete (remember the score of 0.96), which seems to allow for enough gene flow to homogenize the majority of the genome. Only the genomic regions containing “plumage genes” are able to withstand this homogenizing force.

Similar patterns have been described in other avian systems, such as Hooded Crow (Corvus cornix) and Carion Crow (C. corone) or Taiga Bean Goose (Anser fabalis) and Tundra Bean Goose (A. serrirostris). A few divergent genomic regions seem to be sufficient for a high level of reproductive isolation. We need more studies that quantify reproductive isolation at the phenotypic level and provide a link with the genomic underpinnings of the isolation barriers. Studying the evolution of reproductive isolation from different perspectives – behavioral, morphological and genetic – will fuel healthy debates and will provide more insights into the origin of species.

References

Confer, J. L., Porter, C., Aldinger, K. R., Canterbury, R. A., Larkin, J. L., & Mcneil Jr, D. J. (2020). Implications for evolutionary trends from the pairing frequencies among golden‐winged and blue‐winged warblers and their hybrids. Ecology and Evolution10(19), 10633-10644.

Toews, D. P., Baiz, M. D., Kramer, G. R., Lovette, I. J., Streby, H. M., & Taylor, S. A. (2021). Extensive historical and contemporary hybridization suggests premating isolation in Vermivora warblers is not strong: A reply to Confer et al. Ecology and Evolution.

Porter, C. K., Confer, J. L., Aldinger, K. R., Canterbury, R. A., Larkin, J. L., & McNeil Jr, D. J. (2021) Strong yet incomplete reproductive isolation in Vermivora is not contradicted by other lines of evidence: A reply to Toews et al. Ecology and Evolution.

Featured image: Golden-winged warbler (Vermivora chrysoptera) © Bettina Arrigoni | Wikimedia Commons

These papers have been added to the Parulidae page.

Genomic study unveils the true identity of Brewster’s and Lawrence’s Warbler

Are they first generation hybrids, backcrosses or something else?

Some bird hybrids were initially described as distinct species. I have covered some notable examples on this blog, such as Rawnsley’s Bowerbird (Ptilonorhynchus rawnsleyi) and Argus Bare-eye (Phlegopsis barringeri). In most cases, the species name disappears when the hybrid identity of the bird has been revealed, but sometimes the name stays around. In papers on hybridization dynamics between Golden-winged Warbler (Vermivora chrysoptera) and Blue-winged Warbler (V. pinus), you often come across Brewster’s Warblers and Lawrence’s Warblers. The latter two “species” turned out to be hybrids. In 1893, Sage already expressed his doubt by stating that ‘I am not inclined to believe leucobronchialis [i.e. Brewster’s Warbler] a hybrid, but hope to have more to say on this subject at another time.” However, the names are still used to indicate the characteristics of these birds.

“Lawrence’s” hybrids are similar to Blue-winged Warblers (i.e. yellow overall, with 2 narrow white wing bars) but have the black throat patch and face mask, similar to Golden-winged Warblers. “Brewster’s” hybrids, by contrast, lack a black throat patch, have little to no yellow on the underparts, and commonly have partially separated yellowish wing bars.

Based on these traits, Nichols (1908) and Parkes (1951) speculated that first generation hybrids would look like Brewster’s Warblers, while second generation hybrids and backcrosses would resemble Lawrence’s Warblers. With the advent of genomic data, we can put these hypotheses to the test. In a recent study in the journal The Auk, Marcella Baiz and her colleagues examined the genetic make-up of nine Vermivora warblers.

The different species and hybrids of Vermivora Warblers. From: Baiz et al. (2020) The Auk.

 

Hybrid Triangles

To figure out whether Nichols and Parkes were right, the researchers used triangle plots. Based on two statistics – heterozygosity and hybrid index – you can deduce what kind of hybrid or backcross you are dealing with. Pure individuals are located in the lower corners, while first generation hybrids are at the top. The sides of the triangles (D1 and D2) indicate backcrosses. You would thus expect that Brewster’s Warblers (F1) end up at the top and Lawrence’s Warblers at the sides (backcrosses) of these triangles.

This was, however, not the case. The sequenced individuals were scattered across the triangle and did not follow the predictions by Nichols and Parkes. The Lawrence’s Warbler in this study is not a backcross, but probably a multigenerational hybrid with mostly Blue-winged Warbler ancestry. Similarly, the Brewster’s hybrids are not F1 hybrid, but can trace the majority of their ancestry to either parental species. It thus seems that these hybrid types are quite variable and that F1 hybrids and backcrosses are not easy to distinguish based on the coloration of their underparts.

An example of a triangle plot (left, adapted from Pulido-Santacruz et al. 2018). In this case, you would expect Brewster’s Warblers (F1) at the top and Lawrence’s Warblers at the sides (backcrosses) of these triangles. The results show a different picture, indicating that the hybrids are quite variable (right, from Baiz et al. 2020).

 

Black Throat Patch

The story of the black throat patch is very different. Previous work by David Toews and his colleagues uncovered high genetic differentiation between Golden-winged and Blue-winged Warblers near the gene ASIP. This candidate gene has been linked to plumage differences in other bird species, such as Sporophila Seedeaters, Setophaga Warblers and Lonchura Munias. In this study, the researchers could zoom in on the genomic region where this gene resides. They found genetic variants in front of ASIP, suggesting that mutations in the regulatory sequences – the on-and-off switches – are responsible for the presence or absence of a black throat patch. Gene expression studies are needed to confirm this prediction. So, we moved on from one set of predictions (by Nichols and Parkes) to the next one. In science, we call that progress!

A clear signal of genetic differentiation at the ASIP gene (highlighted in grey). From: Baiz et al. (2020) The Auk.

 

References

Baiz, M. D., Kramer, G. R., Streby, H. M., Taylor, S. A., Lovette, I. J., & Toews, D. P. (2020). Genomic and plumage variation in Vermivora hybrids. The Auk, 137(3), ukaa027.

Featured image: Golden-winged Warbler (Vermivora chrysoptera) © Caleb Putnam | Wikimedia Commons

 

This paper has been added to the Parulidae page.