My last research paper. Part 2: Results

January 28, 2020 • 10:30 am

A few days ago I began a two-part summary (it’s now become three parts!) of what will probably be my final “research” paper: the last paper in which I pushed Drosophila flies with my own hands to gather data. (This doesn’t mean it’ll be my final science paper.)

That post discussed the aims and the methods of our paper in Genetics, which you can see by clicking on the screenshot below, or by downloading the pdf here (the full reference is at the bottom). If you can’t see the paper, and don’t have the legal Unpaywall app, a judicious inquiry will produce a pdf for you.  After having written this post, I think that I will divide the discussion into three parts, as this has gotten a bit long. Today we’ll have the results, and within a few days I’ll write about what I see as the significance of the results (i.e., what’s called the “discussion” section of the paper).

The rationale of the experiment is described in detail in the previous post, and I won’t repeat it here. In short, we took two pairs of species, and for each pair, made a hybrid swarm consisting of individuals having half of the genome from each parental species, and half of the organelles and cytoplasm of each parental species (the contents of the cells). Each pair comprised one widespread species and one island species (D. simulans/D. mauritiana in one experiment and D. yakuba/D. santomea in the other). The object was to see if the hybrid swarm would revert back to one or the other parent species, remain as a group of “mongrels” that had both species’ genes segregating, or, perhaps, might evolve into a new lab species that was reproductively isolated from both parental species.

We made eight replicate “swarms” for each pair of species to see how repeatable the changes were. And we measured four sets of characters, each of which characterized and differentiated the two parental species:

1.) Morphological traits that differentiated each pair of species. There were five of these in the D. simulans/D. mauritiana pair and three in the D. santomea/D. yakuba pair. These were described in the previous post, and the graphs below also name them. Characters in the swarm (and pure species) were measured every five generations up to generation 20.

2.) Mating behavior. As described before, there is mating discrimination (and shortened copulation) between the members of each pair. We took males and females from the hybrid swarm and mated them to individuals of the pure parental species to see if the hybrids behaved like one pure species or another (we did this at Generation 21; we kept the swarms going for 24 generations).

For example, copulations between D. simulans males and D. mauritiana females are very short compared to copulations in the pure species. If hybrid-swarm males also show short copulation duration when mated to pure D. mauritiana females, we can conclude their mating traits have reverted to those of D. simulans. (They’d also be expected to show no mating discrimination against D. simulans but substantial mating discrimination against D. mauritiana.) We did this with both pair of species.

3.) Sterility in species crosses. When you cross members of both pairs of species, you get hybrid males that are sterile and females that are fertile. (This pattern is called “Haldane’s Rule”.) Sterility also persists in the hybrid swarms for a while as genes producing the malfunction get weeded out of the population by selection. We measured the sterility of males in the population at generation 20 compared to that of the pure species (in which males are perfectly fertile). More important, we looked at the sterility of hybrids produced in matings between swarm individuals and the two pure species (all at Generation 20). For example, if you cross individuals from the D. mauritiana/D. simulans swarm to the parental species and get fertile offspring with D. simulans but sterile offspring with D. mauritiana, then you know that the genotypic constitution for fertility in the swarm has reverted to that of D. simulans.

4.) DNA composition of hybrid swarm. As I wrote in the previous post, both pair of species are differentiated at many sites on their DNA. We could thus sequence samples of the hybrid swarm and see what proportion of each species’ genome remained in that swarm after 20 generations. We did this survey in all eight replicates for each of the two swarms, but only once, as this endeavor involved sequencing about 20 million bases. The analysis is a bit complicated and I needn’t go into it here.

The paper:

I’ll give the results of four tests below. They are all consistent and unequivocal: in every analysis, either the morphology, behavior, or DNA sequence of the swarm reverted to that of the “dominant” (mainland) species: D. simulans in one swarm in D. yakuba in the other. And this was true of all eight replicates of each swarm. Also, by generation 20, the DNA sequences of all replicates had reverted almost completely to that of the dominant species, though some DNA of the island species remained. That “relict” DNA was pretty consistent across replicates.

1.) Morphological traits. The two graphs below show the changes in morphology of the species-distinguishing traits occurring in all the swarm replicates over 20 generations (measurements were taken at generation 0, 5, 10, 15, and 20). The first plot shows the D. mauritiana/D. simulans swarm and the second the D. yakuba/D. santomea swarm. We also measured the morphology of both pure species as controls over time; these values are the straight and dotted lines at top and bottom (these values stayed pretty constant, as the traits are species-specific.) The hybrid replicates at each generation are shown as open circles, with the means among replicates plotted as best-fit lines, with each replicate being a different color.

In the first plot below, the values of pure D. mauritiana traits are the dotted lines; those of pure D. simulans traits are the solid lines. And you see that the hybrid swarm at generation 0 is largely intermediate between the lines, as it should be (forehead width, however, remains close to that of pure D. mauritiana for 10 generations before reverting to the D. simulans value).

The important result is that, over time, every replicate for every character reverted to the D. simulans value. The changes are especially marked between generation 10 and 15. By generation 20, the values of all traits are essentially those of pure D. simulans. For these traits the hybrids have, then, evolved to have all the trait values of D. simulans. The swarm looks like pure D. simulans individuals.


And this is the plot for traits in the D. yakuba/D. santomea swarm, with pure D. yakuba values being the solid line and those of D. santomea the dotted line. Again, the traits begin as intermediate in the first generation and then gradually take on the values of the dominant species (D. yakuba) after 20 generations. This is true in all eight replicates. In particular, the pigmentation of flies in the swarm winds up just as dark as that of pure D. yakuba individuals (a score of about 600 on a 1200-point maximum scale), rather than being almost unpigmented like D. santomea (that pure species has an average pigmentation score of 49). The characters change at different rates over time. For example, change in pigmentation is smooth over time but that of hypandrial bristles reverts within 5 generations to the value for D. yakuba.


2.) Mating behavior. I can summarize briefly: in all aspects of mating and copulation behavior, the hybrid swarms reverted to the “dominant” mainland species in all replicates by generation 21. In other words, in the D. yakuba/D. santoma swarm all the individuals behaved like pure D. yakuba flies, and in the D. simulans/D. mauritiana swarm all the individuals behaved like pure D. simulans individuals. This is true for both males and females. So we have reversion to the pure species in not just morphology, but in mating behavior.

3.) Sterility in species crosses. Again, in all the replicates of both hybrid swarms, individuals reverted to the fertility characteristics of the “dominant” mainland species by generation 20. For example, looking at the individuals in the 8 replicate D. simulans/D. mauritiana swarms, all swarm males produced fertile male offspring when crossed with D. simulans females but sterile male offspring when crossed with D. mauritiana females. The same held when we crossed hybrid-swarm females to either D. simulans or D. mauritiana males. And likewise again with the D.santomea/D. yakuba swarm: by the end of the experiment, both swarm males and swarm females behaved in their fertility relationships like pure D. yakuba individuals. Again, we observe ubiquitous and replicable reversion to the “dominant” mainland species.

4.) DNA composition of hybrid swarm. The assay at Generation 20 showed that in all the swarms of both species, the DNA of the island species had largely been eliminated, so that the genomes of the swarm were almost entirely that of the dominant species. However, some island-species genome remained in both replicates, as expected since some of it is “neutral” and wouldn’t be subject to selection one way or another. Here, for example, is the proportion of ancestry in each replicate (two bars for each replicate depending on which reference genome we used), with the dominant species’ DNA in yellow and island-species DNA in red. Very little of the island-species DNA remains, and what remains is present “segregating”, i.e., sites having one copy of island DNA and the other copy of mainland DNA at a given position (remember, there are two copies of every gene). Clearly, DNA from the island species is on the way out.


(From paper): Genetic ancestry rapidly and consistently regressed to that of one of the two parental species in all admixed populations. (A) The proportion of sites either fixing for D. simulans ancestry or still segregating for both parental species’ ancestry in each of the eight admixed D. mauritiana/simulans populations. (B) The proportion of sites either fixing for D. yakuba ancestry or still segregating for both parental species’ ancestry in each of the eight admixed D. santomea/yakuba populations. Sites were considered to still be segregating for both parental species’ ancestry if any of the ploidy = 8 genotypes 2 | 6 through 6 | 2 received a posterior probability >1/3. The left bar for each population summarizes results obtained when mapping to either the D. mauritiana (A) or the D. santomea reference genomes (B). Bars to the right, for each population, summarize results obtained when mapping to either the D. simulans (A) or D. yakuba (B) reference genomes.

Here is a broad genome scan, with each vertical line representing a 5000-base window, with its species composition indicated in yellow (dominant species DNA present) or red (island species DNA present, but not in very high frequency). I’ve put the caption from the paper here, too. As you read down each of the two figures, you see the DNA on that particular chromosome arm among the eight replicates (there are four chromosomes, with the X having only one arm, the second and third having two arms each, and the fourth being very small).


(From paper): Genome-wide distribution of ancestry in all admixed populations. Heatmaps showing ancestry estimates summarized in 5-kb genomic windows for each chromosome or chromosomal arm in the D. simulans (A) and D. yakuba (B) reference genomes. Each row is a different admixed population and colors reflect ancestry ranging from 0 (fixed for “minor” parent ancestry) to 1 (fixed for “major” parent ancestry). The bottom row summarizes the number of populations that showed evidence of a given genomic window still segregating for both parental species’ ancestry (i.e., ancestry estimate < 0.8).

As you see, the vast bulk of the genome is yellow, coming from the dominant species (the red bars stick out, but they are not nearly as frequent as they look against the pure yellow background).

One also sees that particular regions of the genome tend to remain “segregating” across all replicates: for example the tip of the third chromosome in the D. simulans/D. mauritiana swarm and the middle of the right arm of the second chromosome in the D. yakuba/ D. santomea swarm.

Finally, to show you the meager amount of foreign genome remaining in most parts of the genome, here is a plot showing, for each of the five chromosome arms, how much of the swarm genome was segregating: i.e., what proportion of the DNA had some sequence from both species (it was almost all “heterozygous”, with one copy of the mainland sequence and one of the island sequence). The eight replicates are given different symbols for each arm.

As you see, for almost all chromosome sites except for some replicates of chromosome arm 3L in the D. simulans/D. mauritiana swarm and arm 2R in the D. yakuba/D. santomea swarm, on average less than 5% of the island species DNA remained. In all arms but one in the D. yakuba/D. santomea swarm, almost no foreign DNA remained.

What’s also notable is that the amount of foreign DNA on the X (“sex”) chromosome was the lowest in both swarms. This may be because the X chromosome contains a number of genes producing hybrid male sterility, and thus “island” DNA was quickly eliminated by selection. (Also, since the X is present in only one copy in males, both recessive and dominant “sterility genes” are fully expressed, so they’re eliminated much more quickly.)

(Caption from paper): The proportion of genomic windows where both parental species’ ancestry still segregated varied across chromosomes. Each point represents the proportion of 5-kb genomic windows that have evidence for both parental ancestries still segregating after 20 generations following initial hybridization between the parental species. (A) D. simulans/D. mauritiana; (B) D. yakuba/D. santomea.


The Big Conclusion: In all replicates in both swarms, and for all traits measured—morphological, mating behavior, fertility relationships, and DNA sequence—the traits and the DNA of the swarm evolved (reverted) in the laboratory back to that of one pure species. And in all cases that pure species was the “dominant” mainland species: D. simulans in one swarm and D. yakuba in the other. We did not get a “hybrid” species, but rather got back a population whose DNA was largely that of a pure species. The results were markedly consistent among replicates, and the overall results very similar in both swarms.

In the next (and last) post, I’ll try to describe the significance of these results, and float some theories about why, in all cases, our swarms reverted to the dominant mainland species instead of the island species.

If you’ve gotten this far, thanks for reading!


Matute, D. R., A. A. Comeault, E. Earley, A. Serrato-Capuchina, D. Peede, A. Monroy-Eklund, W. Huang, C. D. Jones, T. F. C. Mackay, and J. A. Coyne. 2020. Rapid and predictable evolution of admixed populations between two Drosophila species pairs. Genetics 214:211-230.

37 thoughts on “My last research paper. Part 2: Results

  1. I just wanted to say that, despite my not understanding all the technical aspects of the research and the content, your paper, and others like it, are extremely interesting and keep my brains challenged. Thanks!

      1. I found it fascinating and seemingly easy to understand which leads me to believe that I don’t really understand as much as I think.

    1. That’s me, though I’m nowhere near understanding much at all. It makes me realise the ignorance of creationists, who have no idea that this kind of work goes on, assuming that everything we know about evolution is just some sort of guess.

    2. Yes, I’m so dumb that I’m still waiting for the paper that backs up the assertion that “time flies like an arrow, but fruit flies like a banana”. (I know… but just because something is old doesn’t mean it isn’t good.)

  2. Waiting with bated breath for your discussion. Have my own thoughts on the matter! But need to read more carefully(?)also.

  3. Thank you Jerry for taking the time to write these articles explaining your research. Fascinating so far and I am looking forward to your 3rd article to see if any of my amateur hypotheses about how your results might shed light on some other puzzles have any hint of plausibility.

  4. This really got me thinking about resilience in evolution. I am almost always fixated on change and mutation, but rarely on resiliency of life to maintain a specific form. Even with shortcomings, some species find their niche and maintain that adaptation for as long as they can. I am not sure that entirely what was shown in this work, but that’s what it made me think about.

  5. “When you cross members of both pairs of species, you get hybrid males that are sterile and females that are fertile. (This pattern is called “Haldane’s Rule”.)”

    I believe male mules are sterile but females are fertile with a parent species. I’m not sure exactly or if this is the same rule.

    “…or, perhaps, might evolve into a new lab species that was reproductively isolated from both parental species.”

    I would have guessed that this would have happened. It’s interesting that it did not. I’m very curious about the why.

    This is fascinating.

    1. Each species had evolved to fill a specific niche. A doctor told me once that nature has a way of getting rid of junk. Any change into a new species would usually to occur over s long period of time based on changes that make the changes necessary. The organism the subordinate DNA as unnecessary material and weed it out.

      I took only three biology courses in college and most of the details are beyond my interest and understanding but am interested in the results and the theories.

      1. What was the proposed mechanism for getting rid of junk? A tendency for traits established over a longer period of time to be dominant?

        1. I had had sone surgery and had a big wad of blood swollen up in my body part. He was warning ne that the whole thing might bust open and be expelled by a natural process. That did not happen, it was all slowly absorbed ans expelled. That example gas nothing to do with DNA but seemed to me to apply. That same statement or rule could be applied in many different areas and applications. Ge was using the term in a broader meaning also but I understood his point. I know absolutely nothing about mechanisms for DNA changes over time.

  6. Thanks, PCC(e), for sharing this fascinating research study with us. I used to work in a
    completely different area, one which might be called phenetics. However, I have retained an interest in your subject ever since, long long ago as a grad student in biochem and genetics, I read Dobie’s “Genetics and the Origin of Species”. I still have that marked and dog-eared volume, as well as a few more
    contemporary relics from those days—long before it all turned into Gnome Sciences.

  7. You have made a complex topic understandable to a layperson.

    That speaks to both your acuity as a researcher and your ability as a teacher.

    Thank you for both your work and your very clear explanation


  8. I’ve been very interested in these posts and am looking forward to your thoughts about why the flies reverted. I can’t think of any plausible reason myself though I’d guess the two species being dominant to start with is a factor.

  9. What exactly is a hybrid species then? I didn’t think that fertile hybrids of two species had to have exactly 50/50 DNA mixtures.

    If these populations were to maintain their current DNA proportions (of parental and island variant) as an isolated population in the wild, would they not perhaps be considered subspecies, more closely resembling the mainland variant but still different from both?

    Or do you think if this experiment were continued all of the ‘island’ DNA would eventually be eliminated entirely?

    1. A hybrid species (if it’s a diploid) is a new species that originated from hybrids between two existing species, and evolves reproductive isolation from both of the parental species. Diploid hybrid species are pretty rare.

      I these were an isolated population, I don’t know how they would be classified, as their behavior and morphology are the same as one parental species, though they (now) have DNA from the other species. Subspecies designation is subjective, but I doubt that a little bit of DNA from one species would be enough to do that. I don’t know if all the DNA would be eliminated; neutral DNA would likely remain variable, but, in nature, a hybridization even moved the entire mitochondrion, and the genome, from D. yakuba into D. simulans, so it wasn’t eliminated.

      1. Thank you for replying.

        Do you think what you observed here is typical of island/mainland species crosses in general? That the isolated species is simply less viable, and its alleles generally impart lower fitness?

        And was the loss of the island species character mediated entirely through increased sterility/decreased reproductive success, or were the individuals less viable in other ways?

        I apologize if this was covered already in the paper. I only read your summary here.

  10. Thi sis very interesting, and may shed some light on the orchid hybrid swarms that I am seeing in my study area. I look forward to the next installment!

  11. Seems to me this supports the idea that sympatric speciation is going to be exceedingly rare, if it exists at all, at least outside of plants (which have way more flexible genomes than animals).

  12. That ~5% relict value is about the level of Neandertal in our genomes. Is there any sense that our Neandertal segments remain because they are also neutral, or are they spread across the genome from one to another individual vs tending to cluster in certain spots as in 2R in santomena x yakuba above?

    1. One difference I noticed. In the final generations that had largely reverted to the dominant species but still retained a small percentage of the island species DNA, that DNA was largely the same among the entire population. But that doesn’t seem to be the case with Neanderthal genes in modern humans.

      From what I recall results vary from there being from 40% to 70% of the entire Neanderthal genome still existing scattered among various groups of modern humans. Individual humans average only about 2% – 4% Neanderthal genes but unlike with this fly experiment those genes are not largely the same among the population of modern humans.

        1. What kind of blows me away is the enormous time spans that the several species of flies have been separated. Still somewhat ‘interspecies’ hybrid possible. And what short time spans in comparison regarding Neanderthal/sapiens crosses. I would love an extended conversation on all these topics. How many generations did the lab.flies go thru? Around 50? Were they culled? How many since hybridisation of sapiens with neanderthal? With back crossing?
          Do crossovers occur at meiosis at around the same frequency in these very different species. Albeit very different numbers of chromosomes. Etc etc etc.
          Certainly rouses multiple questions.

  13. This is such an interesting result. Does this mean that the selective pressures on the islands must have been quite different than those in your lab? Or otherwise the island populations could never have become separate from the mainland populations they presumably did in fact evolve from?

  14. This paper is so cool! I’ve barely digested most of the technical details but taking from the highlight conclusions and thoroughness of the experimental design, what a cool scientific question to ask and tour de force paper summarizing so much work. And such an unequivocal and definitive result.

    It’s reminiscent of Lenski’s long-term evolution experiment in E. coli but in a much higher species. Bravo Prof. Coyne and congratulations! What a capstone to a distinguished career.

  15. So, when we find a species with a small amount of DNA from another species, is there a way of differentiating between the following two cases: (1) an initially large hybrid swarm that has (mostly) reverted to only on of the original species; (2) a single species into which a very small amount of hydridization has occurred? I am thinking of the fact the modern humans have a small amount of Neanderthal and Densisovian DNA. It would be nice to know if early non-African humans were once a large hybrid swarm that reverted to (mostly) African homo sapiens or if they were always dominated by African homo sapiens but with a small amount of interbreeding.

  16. After reading about the thought and care that went into your experiment, I can see why you got so angry about the Washington Post article about the new “species” of parrot!

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