Searching for a hybrid zone between two Andean warblers

Extensive phenotypic variation across a transect between Ecuador and Colombia.

From the 1930s into 1950s, John Zimmer published numerous extensive monographs about “Studies on Peruvian Birds”. In number 54 of the series, he focused on the bird families Catamblyrhynchidae (now part of the Thraupidae) and Parulidae. While describing specimens of the genus Myioborus, he commented on some “puzzling specimens” that showed characteristics of two subspecies of the Golden-fronted Redstart (M. ornatus chrysops) and the Spectacled Redstart (M. melanocephalus ruficoronatus).

The extreme characters of ruficoronatus strongly suggest those of the ornatus group, and an occasional specimen of o. chrysops shows a noticeable patch of rufous in the center of the crown, strongly suggesting ruficoronatus. It is not impossible, therefore, that the puzzling specimens of one sort and another may represent intergrades or even hybrids between the two groups, but much more material will be necessary before an adequate solution is reached.

Several researchers followed the advice of Zimmer and collected more material on these birds. A recent study in the journal Ornithology reported on their findings. Is there a hybrid zone or not?

Two Subspecies

Before we delve into the possibility of a hybrid zone, we need to clarify the distribution of the M. ornatusM. melanocephalus species complex. First, the Spectacled Redstart (M. melanocephalus) is divided into five subspecies that replace each other when you travel along the Andes from Bolivia to Ecuador (see dots in the map below). The southern subspecies (malaris, melanocephalus and bolivianus) lack a rufous crown, which is present in the two northern subspecies (griseonuchus and ruficoronatus). Second, the Golden-fronted Redstart (M. ornatus) comprises two subspecies (ornatus and chrysops) that occur in different cordilleras in Colombia and Venezuela (see triangles in the map below). The putative hybrid zone concerns interactions between the most northern subspecies of the Spectacled Redstart (ruficoronatus) and the western subspecies of the Golden-fronted Redstart (chrysops). Laura Céspedes-Arias and her colleagues collected samples of these subspecies across a transect running from Ecuador into Colombia.

(A) Map showing the distribution of the subspecies in the M. ornatus-M. melanocephalus species complex. (B) Overview of the sampling transect in the study. From: Céspedes-Arias et al. (2021) Ornithology.

Clinal Patterns

Plumage analyses of over 300 specimens revealed a wide variety of phenotypes, representing different trait combination of both subspecies. It quickly became clear that individuals with intermediate phenotypes were most common along the transect. Some traits, such as head and chest coloration, showed a clear clinal transition from one subspecies into the other (see this blog post for more information on cline theory). All in all, these morphological patterns pointed to a roughly 200 kilometer wide hybrid zone, confirming the suspicion of John Zimmer.

Next, the researchers turned to genetic data by sequencing the mitochondrial gene ND2. In contrast to the plumage traits, this gene did not show a smooth transition. Instead the researchers found extensive haplotype sharing between the subspecies. This pattern can be explained by the recent origin of these subspecies (i.e. incomplete lineage sorting) or by extensive introgression due to hybridization. Genomic analyses will be needed to discriminate between these possibilities. Nonetheless, the most likely scenario seems to entail allopatric divergence leading to differences in plumage traits, followed by secondary contact and extensive hybridization. Another exciting avian hybrid zone to study in more detail.

A sample of the diversity of plumage phenotypes across the hybrid zone, from a typical chrysops (top) over intermediates to a typical ruficoronatus (bottom). From: Céspedes-Arias et al. (2021) Ornithology.


Céspedes-Arias, L. N., Cuervo, A. M., Bonaccorso, E., Castro-Farias, M., Mendoza-Santacruz, A., Pérez-Emán, J. L., Witt, C. C. & Cadena, C. D. (2021). Extensive hybridization between two Andean warbler species with shallow divergence in mtDNA. Ornithology138(1), ukaa065.

Featured image: Spectacled Redstart (Myioborus melanocephalus) © Francesco Veronesi | Wikimedia Commons

Bioacoustics support the decision to split the Elegant Pitta into three species

Playback experiments corroborate acoustic analyses.

An integrative approach to taxonomy is becoming more common. On this blog, I have covered several studies that used multiple lines of evidence to support a taxonomic decision (see for instance this lark species complex or the Gentoo Penguin). Combining multiple datatypes protects some taxonomists from their eagerness to split taxa into multiple species based on a few minor differences. For example, a morphological study on the bean goose complex found that it was possible to discriminate between the taxa fabalis and rossicus using just two measurements: the number of “teeth” in the upper mandible and the maximum height of the lower mandible. Some ornithologists argued that these differences were sufficient to elevate both taxa to species status. But how biologically relevant are these traits? Do they represent a plastic phenotype that is shaped by certain environmental conditions? Or are they heritable adaptations to a particular ecological lifestyle? Such questions need to be addressed before one can properly assess their taxonomic level. Similar reasoning can be applied to the songs and sounds produced by different taxa. Just because two birds sound different does not necessarily mean that they are different species (just think of all the dialects sung by certain songbirds). The biological relevance of bioacoustic differences needs to be investigated in more detail, such as conducting playback experiments. A recent study in the journal Avian Research did just that to evaluate the taxonomy of the Elegant Pitta (Pitta elegans) species complex.

Territorial Calls

Currently, the Elegant Pitta is classified into five subspecies (concinna, maria, vigorsii, elegans, and virginalis). Arya Yue and her colleagues took a closer look at these subspecies to see whether some might represent distinct species. An analysis of territorial calls revealed striking differences between the subspecies. Here is the succinct summary from the results section:

The widespread taxon concinna, resident from Lombok to Alor with the exception of Sumba, exhibited a distinct two-element motif. In contrast, most other taxa, including nominate elegans centered around Timor, maria and virginalis utter a three-element motif in which the first two elements are given in quick succession. The only taxon displaying variability in this trait was the far-eastern vigorsii, centered around the Tanimbar and Kai Islands, in which both two-element and three-element motifs were detected. However, neither of them resembled those given by the other taxa.

The bioacoustic differences were also apparent in the principal component analysis (PCA) where elegans, virginalis and maria formed one cluster distinct from the two other subspecies (concinna and vigorsii).

Principal component analysis plot for territorial calls in the Elegant Pitta species complex. From: Yue et al. (2020) Avian Research.

Playback Trials

However, as I explained at the beginning of this blog post, just because two taxa sound different does not necessarily mean that they are different species. So, how biologically relevant are these differences in territorial calls? One of the co-authors (James Eaton) performed playback experiments over multiple years to see how the different taxa respond to one another. These experiments revealed that the three subspecies from the PCA-cluster (elegans, virginalis and maria) responded aggressively to each others calls, whereas they ignored the calls of the other subspecies (concinna and vigorsii). It would take me too long to describe all the species combinations, so you can check the table below for the results. In essence, the playback trials corroborated the bioacoustic analyses.

An overview of the playback experiments involving different members of the Elegant Pitta species complex. From: Yue et al. (2020) Avian Research.

Three Species

Based on these bioacoustic findings – and additional analyses of plumage patterns and morphology – the authors suggest to split the Elegant Pitta into three species:

  • Temminck’s Elegant Pitta (P. elegans) with subspecies elegans, virginalis and maria
  • Wallace’s Elegant Pitta (P. concinna)
  • Banda Elegant Pitta (P. vigorsii)

However, this classification remains to be confirmed with genetic analyses. If the genetic patterns follow the bioacoustic results, it would indicate that the territorial calls contribute to reproductive isolation between the different taxa. Then they could undoubtedly be considered a biologically relevant and taxonomically informative markers.


Yue, A. Y., Ng, E. Y., Eaton, J. A., & Rheindt, F. E. (2020). Species limits in the Elegant Pitta (Pitta elegans) complex from Wallacea based on bioacoustic and morphometric analysis. Avian Research11(1), 1-12.

Featured image: Elegant Pitta (Pitta elegans) © Abdul Azis Gizan | Wikimedia Commons

Before the invasion: Quantifying the level of pre-introduction selection on two Weaver species

Captured birds go through several selective filters before escaping into the wild.

A few months ago, I published a review paper on hybridization in the Anthropocene. Part of this study quantified the most common mechanisms underlying human-induced hybridization events. It turned out that almost half of the studies I looked at reported hybrids due to the introduction of non-native species (34% intentional and 19% unintentional). These numbers indicate that the invasions of exotic species can have substantial consequences. It is no surprise that many biologists have tried to identify the factors contributing to a successful invasion. However, most of these studies focused on the later stages of biological invasions, such as establishment of escaped individuals or population expansions. The very early stages of the invasion have been largely ignored, even though there might be biases in the capture and transport of exotic species. A recent paper in the journal Evolutionary Applications addressed this knowledge gap by focusing on two avian invaders: the Black-headed Weaver (Ploceus melanocephalus) and the Yellow-crowned Bishop (Euplectes afer).

Selection Filters

At the start of the study, Adrián Baños‐Villalba and his colleagues accompanied several professional bird trappers in Senegal. After catching the birds with traditional clap nets, the researchers took several morphological and behavioral measurements and marked each bird with a unique set of numbered rings. This allowed them to follow the individual birds through the different steps from capture to sale, and determine any selection pressures along the way. After spending about one week in traditional storage cages, the birds were transported over 350 kilometers to a bird-trading company in Dakar. According to this company, the birds were typically kept there for one to three months before export. So, the researchers quantified survival rates thirty days after arrival in Dakar. By comparing the morphological and behavioral traits of the surviving individuals at these different stages, it was possible to estimate the strength of selection on certain traits.

An overview of potential selective filters acting during the pre-introduction stages of an invasion process. From: Baños‐Villalba et al. (2021) Evolutionary Applications.

Micro-evolutionary Changes

The analyses revealed significant changes in several morphological and behavioral traits. During the capture stage, for instance, individuals with a smaller head volume (and perhaps smaller brains) were more likely to be caught. This might reflect some variation in cognitive features and the ability to escape. Later on, however, selection seemed to favor larger brains, possibly because these birds could cope better with novel situations. This example nicely illustrates how pre-introduction selection shapes the captive population and can potentially affect the future establishment success of escaped birds. The authors highlight the relevance of this finding at the end of the discussion:

However, as our results suggest, introduced populations may have already undergone micro-evolutionary changes (assuming traits are heritable) through selective filters (even when these are artificial and human-induced) before reaching the establishment stage, and this likely affects all subsequent changes involved in the adaptation to a new non-native area in the subsequent stages.

These insights can be used to better understand the mechanisms behind successful invasions, and to take appropriate management measures. And this study is also very relevant from a hybridization perspective (the main topic of this blog). While working on this blog post, I came across this paper on mussels: “Pre-introduction introgression contributes to parallel differentiation and contrasting hybridization outcomes between invasive and native marine mussels”. The unchartered territory of pre-introduction selection is waiting to be explored.


Baños‐Villalba, A., Carrete, M., Tella, J. L., Blas, J., Potti, J., Camacho, C., Diop, M. S., Marchant, T. A. & Edelaar, P. (2021). Selection on individuals of introduced species starts before the actual introduction. Evolutionary Applications14(3), 781-793.

Featured image: Black-headed Weaver (Ploceus melanocephalus) © Francesco Veronesi | Wikimedia Commons

How convincing is the evidence to split the Gentoo Penguin into four species?

A critical look at the genetic and morphological data supporting this taxonomic proposal.

Taxonomy is often in flux. As new data are collected or novel methods are being developed, the classification of certain sections on the Tree of Life might change. For example, a recent study in the journal Ecology and Evolution presented morphometric and genetic evidence to split the Gentoo Penguin (Pygoscelis papua) into four distinct species. Last year, this study attracted some media attention (including the BBC and the Oceanographic Magazine), but I remained somewhat skeptical about this taxonomic revision. I added the paper to my writing-list where it gathered dust from several months (there were so many other interesting papers to cover on the blog). Now, it has finally resurfaced and we can assess the evidence. How strong is the case for four species of Gentoo Penguin?

Genetic Lineages

In recent years, taxonomy has become more pluralistic, combining several lines of evidence to support taxonomic decisions (see for example this blog post on larks). The researchers in the penguin study also took an integrative approach and collected genetic and morphological data. The genetic analyses – based on more than 10,000 markers – pointed to four clearly distinct lineages, corresponding to populations from several islands (i.e. Falklands, South Georgia Island, South Shetland Islands + Western Antarctic Peninsula, and Kerguelen). In addition, species delimitation analyses supported a model that considers these four lineages as distinct species. However, as I have discussed in other blog posts (see here and here), genetic population structure does not necessarily coincide with species boundaries. Other data types are needed to validate the delimited species.

Genetic analyses revealed four distinct lineages. But do they represent different species? From: Tyler et al. (2020) Ecology and Evolution.

Morphological Overlap

Next, the researchers performed several statistical analyses on a set of morphological traits. A MANOVA test indicated that “all genetically distinct populations are significantly distinct from each other overall.” However, a closer look at the results of this MANOVA test shows that the p-values are just below the significance threshold of 0.05. For example, the p-value separating the Falklands from Kerguelen is 0.0446. Statistically significant, yes. But is this difference biologically relevant?

My skepticism towards the morphological patterns was reinforced by the output of the linear discriminant analysis where the authors reported that “a small number of specimens occupying positions closer to other lineages.” Moreover, assigning individuals to the genetic lineages using the morphological data resulted in an error rate of 10% (i.e. one individual out of ten was assigned to the wrong lineage). Clearly, the morphological differences are not absolute.

Linear Discriminant Analysis of the morphological data. Circles represent individual specimens with triangles showing the lineage mean. Notice the overlap between SGI (pink) and FALK (blue). Tyler et al. (2020) Ecology and Evolution.


So, how convincing is the evidence to recognize four species of Gentoo Penguin? There appears to be some conflict between the genetic and morphometric results. We can recognize four distinct genetic lineages, but they overlap morphologically. And that is a logical finding when you keep in mind that speciation is a gradual process in which different traits evolve at different rates (more details in this blog post). In this case, the Gentoo Penguin populations have become genetically distinct, but the morphological separation might still be under way (or it might stabilize in the current situation). In my opinion, these penguins are in a “taxonomic grey zone”. One could make a case to treat them as subspecies. Or one could argue that the morphological differences are large enough to classify them as distinct species.

An additional argument to recognize four distinct species concerns their conservation status. In the press release, the researchers say that “regarding the four populations as separate species, gives conservationists a better chance of protecting their diversity because if there’s a decline in one of them it will change the threat status as defined by the IUCN Red List”. This is not a biological reason for a taxonomic revision, but a political one. And that is no problem. In light of the current biodiversity crisis it is important to protect as many species as possible. We could have endless academic discussions whether to classify these penguins as species or subspecies, but that won’t safeguard their future. Let’s focus on what matters.


Tyler, J., Bonfitto, M. T., Clucas, G. V., Reddy, S., & Younger, J. L. (2020). Morphometric and genetic evidence for four species of gentoo penguin. Ecology and Evolution10(24), 13836-13846.

Featured image: Gentoo Penguin (Pygoscelis papua) © Ben Tubby | Wikimedia Commons

Hybrids between Zebra Finch subspecies provide evidence for a weak meiotic driver

Subtle deviations from Mendelian expectations point to a meiotic driver in the Timor subspecies.

Every biology student has worked his or her way through the pea-experiments of Gregor Mendel, creating Punnett squares with recessive and dominant alleles. One of the most important insights from these pivotal experiments was the observation that every allele at a genetic locus has an equal probability of being transmitted to the next generation (ultimately giving rise to the predictable ratios of dominant and recessive traits). However, some alleles show clear deviations from these expected patterns. These genetic elements are known as meiotic drivers, because they “drive” the meiotic cell division process in such as way that they have a higher chance of ending up in the gametes (i.e. eggs or sperm cells).

During meiosis, the chromosomes are sorted into four daughter cells. In birds, one daughter cell develops into the mature egg, while the other three develop into polar bodies. The spindle apparatus attaches to the centromeres of the chromosomes and drags them into the different daughter cells. Meiotic drivers are often found close to centromeres, because this allows them to influence the spindle fibers and ensure that they end up in the daughter cell that becomes the mature egg. Indeed, previous research in chickens reported a meiotic driver at the centromere of chromosome 1.

An schematic representation of the meiotic process. Meiotic drivers influence the sorting of chromosomes so that they are transmitted to the next generation. From: Wikipedia.


Meiotic drivers can be difficult to detect, because they tend to be transient phenomena. In some cases, the meiotic drivers lead to deleterious effects and are quickly suppressed by other genetic elements that restore proper Mendelian segregation. Alternatively, meiotic drivers are so successful that they rapidly spread through a population and become fixed (i.e. all individuals have the same genetic variant). One way to identify such cryptic meiotic drivers is to cross individuals from divergent populations. If a meiotic driver evolved in one population but not the other, it will become visible in the hybrids. A recent study in the journal Ecology and Evolution used this approach to look for meiotic drivers in the Zebra Finch (Taeniopygia guttata). The researchers crossed two subspecies – Australian (castanotis) and Timor (guttata) Zebra Finches – and traced the genetic ancestry of several molecular markers. Did some deviate from the expected Mendelian patterns?

An overview of the extensive breeding scheme in this study. From: Knief et al. (2020) Ecology and Evolution.


The experiments revealed “no clear evidence for any active meiotic driver in a cross between Australian and Timor Zebra Finches.” However, there was a significant deviation from Mendelian segregation in females of the first backcrossed generation. This finding suggests that there might be a weak meiotic driver which allows Timor centromeres to outcompete Australian variants in the race for the oocyte. It took numerous hybridization and backcrossing events to detect this subtle signal, indicating that weak meiotic drivers might be more common than we think. Another example of how avian hybrids can lead to exciting discoveries and new insights.

Segregation patterns in different hybrids and backcrosses between Australian an Timor Zebra Finches. Notice the slight deviation from the expected pattern (dotted line) in the female backcrosses (BC1). From: Knief et al. (2020) Ecology and Evolution.


Knief, U., Forstmeier, W., Pei, Y., Wolf, J., & Kempenaers, B. (2020). A test for meiotic drive in hybrids between Australian and Timor zebra finches. Ecology and evolution10(23), 13464-13475.

Featured image: Zebra Finch (Taeniopygia guttata) © Peripitus | Wikimedia Commons

Ancient DNA reveals low levels of past genetic diversity in the Andean Condor

Low genetic diversity might be the natural state for this vulture.

Since the first release of Andean Condors (Vultur gryphus) in Colombia in 1989, more than 200 individual birds have be re-introduced in the wild across South America. The goal of these introductions was to reinforce existing populations and re-establish extinct ones. One of the main arguments for such extensive management programs is the augmentation of genetic diversity. However, the relationship between genetic diversity and the risk of species extinction is not always straightforward (see for example this blog post). And in many cases, it is not clear how much genetic variation has been lost over time. So, what about the Andean Condor? A recent study in the journal Ecology and Evolution obtained DNA samples from museum collections to assess the historical levels of genetic diversity in this iconic vulture species.


Julian Padró and his colleagues sequenced several mitochondrial markers for 42 Andean Condors, covering a time period from 1884 to 2013. The historical and modern samples shared several haplotypes, but one haplotype from 1896 seems to have been lost. This genetic variant belonged to a now extinct population on the Patagonian coast. Demographic analyses indicated that this loss of genetic diversity coincided with the timing of European colonization in South America. Probably, the development of livestock production resulted in conflict between humans and Andean Condors.

In contrast to the southern populations, Andean Condors in the north of their range did lose any haplotypes, despite being driven to near-extinction in recent times. The similar genetic diversity in historical and present times, in combination with the relatively modest loss of one mitochondrial haplotype, suggest that the Andean Condor can cope with low levels genetic diversity. Similar patterns have been reported in other raptor species, such as the Spanish Imperial Eagle (Aquila adalberti) and the Cinereous Vulture (Aegypius monachus).

An overview of the sampling locations for the historical and contemporary samples. The haplotype shows the modest loss of genetic diversity over time. From: Padró et al. (2020) Ecology and Evolution.


The researchers concluded that “Low levels of genetic diversity found in the Andean condor represent a natural state of mtDNA, and thus are unlikely to be an immediate threat to long-term viability.” However, mtDNA represents only a tiny fraction of the total genetic diversity in a population. A genomic perspective is needed to assess the impact of population bottlenecks on the genetic make-up of the Andean Condor. These raptors might be able to withstand a decline in mitochondrial diversity, but what if other genomic regions have been eroded?


Padró, J., Lambertucci, S. A., Perrig, P. L., & Pauli, J. N. (2020). Andean and California condors possess dissimilar genetic composition but exhibit similar demographic histories. Ecology and Evolution, 10(23), 13011-13021.

Featured image: Andean Condor (Vultur gryphus) © Ltshears | Wikimedia Commons

The mystery of the white-faced Limestone Wren-babbler: a leucistic morph or a separate (sub)species?

Genomic and acoustic analyses of the species complex provide the first clues.

Somewhere in Myanmar, there is a white-faced population of the Limestone Wren-babbler (Napothera crispifrons). Ornithologists are not sure how to treat this population from a taxonomic point of view. Is it just a white morph or does it represent a distinct (sub)species? In addition to this mystery, the taxonomy of the Limestone Wren-babbler has been recently revised. Traditionally, this inconspicuous passerine was considered as a single species, comprised of three allopatric subspecies (crispifrons, annamensis and calcicola). However, analyses of plumage differences based on museum specimens suggested to split it into two species: the rufous-bellied N. calcicola and the grey-bellied N. crispifrons (containing two subspecies). An integrative approach is needed here, combining different data sources to support a taxonomic decision (see for example this blog post). And indeed, a recent study in the journal Molecular Ecology collected data on vocalizations and genomics to solve this puzzle.

Three Lineages

Chyi Yin Gwee, Qiao Le Lee and their colleagues generated genomic sequences for 15 individuals and uncovered three deeply divergent lineages. They could confidently discriminate between crispifrons from Myanmar and western Thailand, annamensis from Vietnam and calcicola from northeastern Thailand. More detailed analyses indicated that there has been no gene flow between these three lineages, suggesting that they have been reproductively isolated for some time. The genomic results contradict the plumage-based classification which combined the subspecies annamensis and crispifrons. It turns out that annamensis is more closely related to calcicola than to crispifrons. This finding nicely illustrates the dangers of solely relying on morphological data.

The acoustic data supported the genomic patterns. Analyses of 10 vocal parameters showed that annamensis produces sounds similar to calcicola. Because these taxa look quite different – annamensis is grey-bellied, while calcicola is rufous-bellied – vocal differences might be less important in species recognition. Based on these results, the researchers concluded that “the Limestone Wren-babbler complex consists of three mitochondrially and genomically diverged lineages, each supported by a combination of plumage and vocal characters that would allow them to be diagnosed as different species under many species concepts.”

Genomic analyses of the Limestone Wren-babbler species complex indicated three distinct lineages (see K=3 graph in figure a). Acoustic data separated crispifrons fro the other two taxa (figures b and c), which are morphologically distinct. From: Gwee et al. (2021) Molecular Ecology.

The White Mystery

And what about the white-faced population in Myanmar? The genomic data cluster it within the brown-plumaged populations of crispifrons. Comparing the genomes of white and brown individuals pointed to several outlier regions that contain a few candidate genes involved in pigmentation (including RAB3IP and SLC16A3). Research on other bird species has shown that a few genomic loci can drive drastic plumage differences (see for instance crows and warblers). At the moment, it is difficult to judge how stable the white-faced population is. The researchers might have captured the beginning of a diversification process between white and brown Limestone Wren-babblers, or the white-faced population might disappear in a few generations due to stochastic processes. Only time will tell.


Gwee, C. Y., Lee, Q. L., Mahood, S. P., Le Manh, H., Tizard, R., Eiamampai, K., Round, P. D. & Rheindt, F. E. (2021). The interplay of colour and bioacoustic traits in the differentiation of a Southeast Asian songbird complex. Molecular Ecology30(1), 297-309.

Featured image: Limestone Wren-babbler (Napothera crispifrons) © Francesco Veronesi | Wikimedia Commons

Does disrupted gene expression cause hybrid sterility in Flycatchers?

Taking a closer look at gene expression in the testis.

Every student of speciation should be familiar with the Bateson-Dobzhansky-Muller (BDM) model of genetic incompatibilities. Most evolutionary biologists can probably explain the rationale behind this model, but not everyone will know its interesting history (and why I chose to list these three names). The model was formulated by Dobzhansky (1934) and further developed by Muller (1942). However, Bateson (1909) already published an essentially identical model, apparently unknown to Dobzhansky and Muller, to explain the “secret of interracial sterility”. The BDM-model is very intuitive. Here is the short version from my PhD thesis:

Consider two allopatric populations diverging independently, with the same ancestral genotype AABB in both populations. In one population, a mutation (A -> a) appears and goes to fixation, resulting in aaBB, which is fertile and viable. In the other population, another mutation (B -> b) appears and goes to fixation, resulting in AAbb, which is also fertile and viable. When these populations meet and interbreed, this will result in the genotype AaBb. Alleles a and b have never “met” each other and it is possible that allele a has a deleterious effect that becomes apparent when allele b is present, or vice versa. Over evolutionary time, numerous of these incompatibilities may arise, each possibly contributing to hybrid sterility or unviability.

This model has been mostly applied to mutations in protein-coding genes, but could be extended to the regulation of gene expression. Regulatory regions come in two main types: cis-regulatory elements that are linked to nearby genes and trans-regulatory elements that affect distant genes (millions of DNA-letters apart). Interacting cis- and trans-regulatory elements often evolve in concert, and a mutation in one element can be compensated by a mutation in the other element. When species have experienced different compensatory mutations and interbreed, the gene expression in hybrids might be disturbed, leading to sterility or unviability.

The Bateson-Dobzhansky-Muller model of genetic incompatibilities. From: Wikipedia.

Sterile Males

A recent study in the journal Genome Research applied this reasoning to hybrids between Pied Flycatcher (Ficedula hypoleuca) and Collared Flycatcher (F. albicollis). These two species diverged about one million years ago and interbreed in several locations, including the Swedish island of Öland (where the group of Anna Qvarnström has been monitoring the breeding populations for numerous years). Previous work showed that male hybrids are infertile due to the production of abnormal sperm cells. Could male sterility be the result of disrupted gene expression due to mismatches between cis- and trans-regulatory elements? To answer this question, the researchers took a closer look at gene expression patterns in five Pied Flycatchers, five Collared Flycatchers and three natural hybrids.

The analyses focused on misexpression in hybrids, which can be detected by gene expression levels in hybrids that are either higher or lower than any of the parental species. The researchers reported “evidence for abundant hybrid misexpression in heart, kidney, and liver but not in brain or testis.” In addition, more detailed analyses of genes involved in spermatogenesis did not reveal misexpression in hybrids. All in all, this study could not provide evidence that disrupted gene expression in the testis causes sterility in hybrid males. However, the high levels of misexpression in other tissues could contribute to lower hybrid fitness in other ways.

Typical sperm from a collared flycatcher (a) and a pied flycatcher (b), compared to abnormal sperm from two hybrid flycatchers, indicated by arrows (c-f). From: Ålund et al. (2013) Biology Letters.

Evolution at Two Levels

Although the testis showed no clear signs of misexpression in hybrids, this tissue did experience the highest level of divergence in gene expression between Pied and Collared Flycatcher. More research will be needed to unravel the exact changes in gene expression and their contribution to male sterility, but it seems unlikely that mismatches between cis- and trans-regulatory elements play a major role.

Despite the “negative” result, this study nicely highlights the potential involvement of regulatory changes in evolution and the formation of new species. In 1975, Mary-Claire King and A. C. Wilson already drew attention to the contrast between evolution at the sequence level and changes in patterns of gene expression. Focusing on human evolution, they noted that “a relatively small number of genetic changes in systems controlling the expression of genes may account for the major organismal differences between humans and chimpanzee.” At the time, we did not have the methods to explore how regulatory changes shape evolutionary trajectories. The development of new techniques, such as RNAseq, provide exciting opportunities to understand how changes in gene expression contribute to the origin of new species. What a wonderful time to be an evolutionary biologist.


Mugal, C.F., Wang, M., Backström, N., Wheatcroft, D., Ålund, M., Sémon, M., McFarlane, S.E., Dutoit, L., Qvarnström, A. & Ellegren, H. (2020). Tissue-specific patterns of regulatory changes underlying gene expression differences among Ficedula flycatchers and their naturally occurring F1 hybrids. Genome Research30(12), 1727-1739.

Featured image: Collared Flycatcher (Ficedula albicollis) © Andrej Chudy | Wikimedia Commons

Integrative taxonomy of the Lesser Short-toed Lark and Sand Lark species complex

The combination of genetic and non-genetic data points to four distinct species.

The advent of genetic – and later genomic – data turned out to be a double-edged sword for taxonomists. On the one hand, DNA sequences allow researchers to discriminate between morphologically similar species. On the other hand, the ability to detect ever more fine-scaled genetic differentiation between populations complicates the drawing of species boundaries. Several species delimitation programs using molecular data have been developed, but it remains a daunting task to translate the output of these programs into clear taxonomic arrangements. Jeet Sukumaran and Lacey Knowles nicely described this issue in a PNAS paper: “Until new methods are developed that can discriminate between structure due to population-level processes and that due to species boundaries, genomic-based results should only be considered a hypothesis that requires validation of delimited species with multiple data types, such as phenotypic and ecological information.” In other words, genetic patterns will need to be corroborated by non-genetic data.

Five Lark Lineages

Several months ago, I covered a molecular study on the Lesser Short-toed Lark (Alaudala rufescens) and Sand Lark (A. raytal) species complex (see this blog post). The genetic analyses pointed to five genetic lineages that could be classified into four distinct species. Indeed, the researchers wrote that “our results call for a taxonomic revision, and we tentatively suggest that at least four species should be recognized, although we stress the need for an approach integrating molecular, morphological and other data that are not yet available.” A follow-up study in the journal Molecular Phylogenetics and Evolution provides these missing data, using plumage patterns, biometrics, songs, geographical distributions and bioclimatic factors to evaluate the genetic patterns.

A slightly more elaborate genetic analysis uncovered the same five lineages as the previous study. In addition, the species delimitation program STACEY suggested that four lineages were sufficiently divergent to warrant a species status, namely the heinei clade, the raytal clade, the rufescens clade and the cheleensis + leucophaea clade (see figure below). But what about the non-genetic data? The researchers performed several detailed analyses and an exhaustive overview of the results is not feasible within the scope of a short blog post. So, I will try to summarize the main findings below:

  • The five lineages could not be separated by plumage. This is probably a consequence of the convergent evolution due to adaptation to a similar habitats.
  • Classification analyses based on the wing, tail and bill lengths discriminated between all clades, except for the cheleensis and leucophaea clades. Several individuals from these groups were misclassified.
  • Song characteristics were significantly different between the five clades, although the songs of cheleensis and leucophaea seemed to show a clinal pattern.
  • All five lineages occur in open habitats with scant vegetation, but there appear to be some differences in habitat preferences.
  • Bioclimatic parameters, such as rainfall and temperature, could discriminate between the different lineages.
Genetic analyses indicated five distinct clades. Do they all represent different species? From: Alström et al. (2021) Molecular Phylogenetics and Evolution.

Integrative Taxonomy

Based on these patterns, the researchers concluded that “the rufescens, heinei and raytal clades were unanimously supported as independent lineages by mtDNA, morphology and bioacoustics as well as by the STACEY multilocus analysis. The raytal clade was also supported by its unique habitat. The combined cheleensis and leucophaea clade was also supported by the same datasets.” The latter two clades could not be confidently separated, because they showed clinal variation in several traits. The proposed taxonomy thus includes four species:

  • Lesser Short-toed Lark (A. rufescens)
  • Heine’s Short-toed Lark (A. heinei)
  • Asian Short-toed Lark (A. cheleensis)
  • Sand Lark (A. raytal)

This study nicely illustrates the use of multiple data sources to inform taxonomic decisions (an approach known as integrative taxonomy). It is interesting to see how the genomic revolution has drawn attention to the taxonomic importance of non-genetic data, such as morphology, behavior and bioacoustics. Several authors have highlighted this perspective. I recently argued that genomics provides another line of evidence in the pluralistic approach to species classification (see this book chapter). Similarly, Carlos Daniel Cadena and Felipe Zapata called for the integration of genomic and phenotypic data in avian taxonomy (see this paper). The development of new genetic techniques is bringing us back to the basics.


Alström, P., van Linschooten, J., Donald, P. F., Sundev, G., Mohammadi, Z., Ghorbani, F., Shafaeipour, A., van den Berg, A., Robb, M., Aliabadian, M., Wei, C., Lei, F., Oxelman, B. & Olsson, U. (2021). Multiple species delimitation approaches applied to the avian lark genus AlaudalaMolecular Phylogenetics and Evolution154, 106994.

Featured image: Lesser Short-toed Lark (A. rufescens) © Juan Emilio | Wikimedia Commons

More markers, more power? Genomic analyses uncover fine-scale population structure in Philippine Broadbills

Ornithologists reconstruct the evolutionary history of several island populations.

Scientists love a good acronym. A recent study counted more than one million unique acronyms in papers published between 1950 and 2019, but just over 2,000 (0.2%) were used regularly. You can find some clever ones here, including BIGASS (Bright Infrared Galaxy All Sky Survey) and Gandalf (Gas AND Absorption Line Fitting algorithm). Ornithologists working in Asia might be familiar with the abbreviation PAIC, which stands for Pleistocene Aggregate Islands Complexes. This acronym refers to groups of islands that were connected by land bridges during the Pleistocene when sea levels were lower. The avifauna of the Philippine Archipelago seems to adhere to the PAIC model with islands complexes from different PAICs showing clear genetic divergence (see for example this study by Peter Hosner and his colleagues). However, the evolutionary history of bird populations within a single PAIC remains largely unknown. In a recent paper in the Biological Journal of the Linnean Society, researchers addressed this knowledge gap and took a closer look at the Philippine broadbills (genus Sarcophanops) of the Greater Mindanao PAIC.

Genetic Patterns

Luke Campillo and his colleagues collected samples from the Wattled Broadbill (S. steerii) and the Visayan Broadbill (S. samarensis). Genetic analyses – based on thousands of genetic markers – revealed a deep split between both species. A similar pattern emerged in a previous study using mitochondrial DNA. The researchers attributed these findings to the habitat unsuitability of the Leyte Gulf during the Pleistocene, which prevented birds from the northern and southern islands from mixing.

Apart from this deep split, the genetic analyses pointed to fine-scale diversification within the Visayan Broadbills, separating the populations from the islands Samar/Leyte and Bohol. The authors speculate that “rising sea levels at the end of the Pleistocene would have isolated Bohol first, whereas prolonged connectivity between Samar and Leyte could have promoted gene flow, thus obscuring population genetic effects of inter-island diversification.”

The genome-wide markers clearly differentiated between the two species: Wattled Broadbill (in yellow) and Visayan Broadbill (in blue). Within the latter species, further diversification between islands became apparent. Adapted from: Campillo et al. (2020) Biological Journal of the Linnean Society.

Genomic Power

The population structure within the Visayan Broadbill was not apparent in the mitochondrial DNA, highlighting the power of genome-wide markers to detect subtle signatures of population diversification. A few years ago, I covered this recent genomic development in a book chapter with several colleagues, writing that “genomic data has increased the potential for fine-scale resolution of population structure and determination of population boundaries and population membership.” However, this increase in genomic power can complicate analyses because populations tend to fall on a continuum from isolation to panmixia. Delineating populations and drawing species limits with genomic can become a daunting task. It will be interesting to follow how the genetic patterns in this study will impact the taxonomy of Philippine broadbills. Is the fine-scale population structure in the Visayan Broadbill large enough to justify subspecies or not?


Campillo, L. C., Manthey, J. D., Thomson, R. C., Hosner, P. A., & Moyle, R. G. (2020). Genomic differentiation in an endemic Philippine genus (Aves: Sarcophanops) owing to geographical isolation on recently disassociated islands. Biological Journal of the Linnean Society131(4), 814-821.

Featured image: Wattled Broadbill (Sarcophanops steerii) © Bram Demeulemeester | Flickr