Revisiting Meyer et al. 1990

In a paper published in Nature in 1990, Axel Meyer, Thomas Kocher, Pereti Basasibwaki, and Allan Wilson, using mitochondrial DNA sequences, provided evidence in support of a monophyletic origin of the cichlid diversity in Lake Victoria. Whats more, they found that the genetic variation among these cichlids was less than within the human species suggesting that the founding event for this cichlid radiation was fairly recent. Twenty-six years after the paper was published, I spoke to Axel Meyer about the origins of his interest in cichlid evolution, his memories of this study, and what we’ve learnt since about the evolutionary history of Lake Victoria’s cichlids.

Citation: Meyer, A., Kocher, T. D., Basasibwaki, P., & Wilson, A. C. (1990). Monophyletic origin of Lake Victoria cichlid fishes suggested by mitochondrial DNA sequences. Nature, 347(6293), 550-553.

Date of interview: 1 December 2016 (via Skype)

Hari Sridhar: I’d like to start by asking you about your motivation to do the work presented in this paper. This came just after you finished your PhD. And you had worked on cichlid fishes even for your Master’s dissertation, although on a different aspect.  How did you decide to do this piece of work?

AM: Well, the history is that I started to work as a PhD student in the Zoology Department in Berkeley on these interesting cichlids that live in crater lakes in Nicaragua that were polymorphic for coloration and, seemingly, for tooth morphology, this morphological aspect I discovered. At least we thought that they were polymorphic at that time, because there were only three species described. Now, almost 40 years later, there are 13 species described, some of which we, and others, described in the last years. These fish have a polymorphism in their color, which is why they’re called Midas cichlids. They start life as black and striped fish, but then some of them lose melanophores, and when they do then they turn gold – like King Midas had turned everything into gold that he touched. So, they have a colour polymorphism and a polymorphism in their pharyngeal jaws, which are very important for their feeding ecology and natural selection. I was interested to ask whether or not I would find genetic differences, or more pronounced genetic differences, between the colour morphs or the pharyngeal jaw morphs, because I was interested to ask whether natural or sexual selection is more important in driving diversification in this radiation. When I was a grad student at Berkeley, I went to Harvard to work for a year in the lab of a functional morphologist, who had tried to understand how fish heads work, and once I came back from Harvard – that was in the summer of 1987 – I started working in the lab of Allan Wilson in the biochemistry department for the last year of my PhD. I finished my PhD in 1988. And in that year, I began to address this question, initially with old methods of course. I isolated mitochondrial DNA and did restriction digests. DNA sequencing was very difficult and cumbersome at the time, but I was just lucky to be in Allan Wilson’s lab at the time when PCR was invented. PCR, of course, changed the world as we know it, and, for me, it was just a lucky coincidence to be there while the group of us postdocs (that included very talented and successful scientists such as Svante Paabo, Tom Kocher, Kelly Thomas and others) that I was then part of had worked on developing universal PCR primers (published in Kocher et al. 1989, PNAS) that were, then later, used in many, many studies (this paper has been cited more than 5000 times). And there were connections with the Wilson lab; a former postdoc had worked at Cetus Corporation where Kary Mullis worked and PCR was invented. And so, we were the first lab to use PCR for molecular evolutionary kinds of questions. Once PCR was being established in the lab, I could then begin to work on all kinds of questions. It was like a kid in the candy store. PCR changed my life, but it also changed science, arguably. My background was more in evolutionary biology and systematics so I knew what the open, important, pertinent questions were in vertebrate phylogenetics. So, I could work on lungfish and coelacanths, I did all kinds of studies on birds and whales, all kinds of things that came immediately after PCR was invented. And for my postdoc, I had a Sloan Foundation fellowship in molecular evolution. The Sloan Foundation always tried to sort of establish new fields, and I was one of the postdocs that they funded to work on molecular evolution. I was on a postdoc for two years -1988-1990 – in Allan’s lab, before I moved for my first job in Stony Brook. This project on the Lake Victoria cichlids was the project that was funded by the Sloan Foundation for my postdoc. And it also had a bit of a history. Allan Wilson had an ability to bring together people that knew their organisms – ‘ologists’ – like Scott Edwards, who was a ornithologist, me as an ichthyologist, or other people from the Museum of Vertebrate Zoology in Berkeley, and pair them with people that, you know, wouldn’t know a fish if it hit them on the head – more gene jocks -who were smart enough, interactive enough, so that the atmosphere in the lab was just fantastically exciting. I mean, in that time, there was a Nature paper almost every month. The journalist from New York Times would call us quite regularly to ask what we had in the pipeline. The Wilson lab really was at the forefront. Several other people besides Svante such as Russ Higuchi, Kelly Thomas, and Allan Cooper were there, who worked on ancient DNA; I too had extracted DNA from museum specimens.  Crichton, influenced by the work from Berkeley, had just written Jurassic Park. There were interactions there, because he got the idea for Jurassic Park, of course, from PCR, and getting dinosaurs from mosquito blood to work, to bring them back to life and so on. It was a super exciting time, and you could really feel that science history was made in that lab at that time. And, Allan Wilson was very interested in cichlids, because the last practical work he had done himself was on cichlids of Lake Victoria. He had done a sabbatical a few years before, in Nairobi in Kenya, at a museum. And he had used starch electrophoresis to look for genetic differences in these Lake Victoria cichlids. So, he knew about them and he was particularly keen to have somebody like me who wanted to work on African cichlids, and so this came together very nicely. Obviously, this paper was very important for my career, but it was, also, sort of, one of the main papers at that time using these methods for evolutionary questions.

HS: How did you get interested in fishes?

AM: I always say that I was one of those kids that always had a frog in the pocket.  I grew up on the outskirts of town, and I had a collection of beetles and newts and things like that and many different pets. When I was 10, I got my first aquarium, and then it went up exponentially. By the time I was in college, I had over 80 tanks that filled two walls. I was one of these kids that, you know, would breed fish and observe fish, and in the winter would collect water fleas to feed them; that kind of thing. I have always liked fish, or other aspects of natural history and evolution. I knew always that I wanted to be a biologist but was not quite clear what kind of biologist. And, as you said earlier in this conversation, my first papers were more on behavioral ecology, or animal behavior even, and then moved into functional morphology, and then into molecular systematics and molecular evolution and comparative genomics.

HS: When was the first time you visited Lake Victoria and Lake Malawi?

AM: That was in 1990. Actually, the fish that were used for this study came, partly, from Pereti Basasibwaki, who’s one of the middle authors on this paper. He was a fisheries officer from Jinja in Uganda who came to the Wilson lab and brought some of the samples. Others, I got from collaborators in Leiden and Holland and so on. I actually went to the lake for the first time after the publication of this paper.

HS: How did this group of authors come together and what did each bring to this piece of work?

AM: Tom Kocher was a little bit senior to me. The way this worked, in the Wilson lab, and the way I’m trying to emulate this now, in my lab, is if you come new to a lab, you’re sort of a shadow of somebody for six months to learn new tricks. Tom had been in the Wilson lab a year or so longer than me, and he knew molecular techniques better than me and trained me in that. So, his contribution there is more an acknowledgement of his training aspects, because all the sequences were from me, and Pereti was the one who brought the fish. That’s sort of how this came together. Tom, who was more interested in cichlid fish from Lake Malawi continued mostly to work on them after we both left the Wilson lab. And I continued to work on Lake Victoria and Lake Tanganyika cichlid and then, after I moved to Germany in 1997, also again on the cichlids of the crater lakes in Nicaragua.

HS: Could you give us a sense of how challenging it was to do this kind of work then, both in terms of the lab work and in terms of procuring the specimens? What did it involve?

AM: Well, obviously, all of this was done by manual sequencing, which meant pouring polyacrylamide gels and running radioactively-labeled nucleotides on these acrylamide gels. The lab was, as I said earlier, very, very exciting and was full of smart and very ambitious people. I mean, all of us ended up with very good jobs. But, for example, there were no PCR machines one could buy in 1987 or 88. So, this was done initially in 1.5 ml tubes rather than 0.5 ml tubes. It was done with NativeTaq and not with CloneTaq. So, it was super expensive and very cumbersome, because you had to move these tubes, following this stop clock, in different water baths, to do the polymerase chain reaction.

The first PCR machine we had in the lab was actually built by Tom Kocher. Tom is very good, technically. He just built himself an aeroplane! But at that time, he built it out of an old Apple II computer and washing machine valves that were controlled by this computer and a hot water bath where the temperature would circulate to bring about these PCR cycles. And so, we had this very clumsy, cumbersome looking thing with 20 tubes. And, of course, everybody wanted to use them. Later on, when Applied Biosystems – then it was still called PerkinElmer – sold the first PCR machine, it was booked out weeks in advance. And it was quite normal for people to come to the lab at two or three or four in the morning to take out the run of somebody else and to put your tubes in the PCR machine. For the 20 people or so in the lab, we only had one PCR machine that ran 24/7. So, it was very exciting, because everybody felt that this was, like I said, a historical period in evolutionary biology. And we all worked very, very hard to try to get those data. And, of course, the protocols, you know, you were happy to get 200 base pairs of sequence of these gels and you would read them by hand and enter them manually by hand in the computer and so on. It makes me sound old when I tell my students that I lived before PCR was invented, that I remember when GenBank would send us folders full of floppy disks. That’s how GenBank used to work. We used to send a floppy disk to them, if you had new sequences. Of course, it’s all very different now. Every base pair counted, and every gel one ran, you had to expose an X-ray film and then, again, read them off with a felt-tip pen or layer on tablet-like devices , by which you can do this more quickly. In any event, you’re really close to your data, let’s say and you could tell which gene one was looking at by the spatial patterns of Gs, As, Ts, and Cs!

HS: Was it challenging to procure samples?

AM: Yes and no. Lake Victoria cichlids are not as common, as hobby fishes, as Malawi cichlids, and I was well connected in that scene. But most of the samples that we use in this study were from Uganda, from Pereti, and also from Leiden. There’s a group in Leiden in Holland that used to work on Lake Victoria cichlids. And I had visited them before when I was still working on functional morphology and I knew the people there. So, some of the fish, I actually get from them.

HS: How did the idea to look at the origins of these fishes come about? Why was this interesting at that point in time?

AM: Well, that there’s something special about the cichlid fish was clear, in the sense that they’re so diverse. There are 500 or maybe more species endemic to Lake Victoria alone, 800 to 1000 in Lake Malawi and maybe 250 in Tanganyika. At the time when I started this, it was not clear though, how these three species flocks were related to each other. It had actually been suggested – I’m referring to a paper that I cite in this Nature paper – that the similarities that are found across the species flocks are not convergence but a sign of common descent. And, it was actually assumed that most of the species across lakes are more closely related to each other if they are more similar to each other. Only this study then showed that Lake Victoria had a single or maybe two ancestral lineages that contributed to it, and thereby, Malawi and Tanganyika evolved independent of what happened in Lake Victoria. And so it made it clear, already then, that similarities, across these variations, are parallel evolved phenotypes and not signs of common descent. In terms of molecular data to look at, there was very little done before. Only a couple of papers, one by Dick Sage, who was a former person in Wilson’s lab, who had done, with Allan Wilson, some work on allozymes of Lake Victoria cichlids. The sequences that I published in this 1990 Nature paper were the first DNA sequences of cichlid fish at all (except for those in the 1989 PNAS method paper), and, in many ways, the Lake Victoria radiation was known as an enigmatic, to be explained, situation that needed some looking at.

HS: Could you give us like a timeline for this work, from start to finish Do you remember in which year you started doing this work?

AM: That must have been 87-88, as soon as I got to the lab, funded as a Sloan-postdoc since the fall of 1988. I have to look at this. The time in Wilson’s lab, like I said, was very exciting. I can now understand this much better than then, that if you have a lab with 20 postdocs or 15 postdocs, you cannot be on top of the details of every project. And so, the way this usually worked is we tried to get Allan’s attention by faxing him a manuscript to a place where he was giving a talk. On the plane on the way back, he would read the manuscript, the fax, and mark up the manuscript. That’s the way we tried to get him to spend time with us. I remember one meeting that I had with him. The actual writing of this didn’t take very long at all. But there was one important conversation I had with Allan, where I remember going to his office and saying, look Allan, I don’t know what to make of this. All of these fish are almost identical. They’re all the same. Allan was a master in thinking outside the box. We sort of always joked in the lab that he can turn shit into gold. And he said, well, of course, this means that they are extremely young, and that they can be traced back to a single ancestral species. So I have to, sort of, give him credit for that insight, or that major message of the paper, where he said, well, usually, of course, you’re looking for genes that have phylogenetic signal. And these mitochondrial genes were so successful in that because, among closely related species, they would evolve quickly and you would still be able to find phylogenetic or population genetic signal (mutations). But here they were identical in cytochrome b and different only by two or three substitutions in the D-loop sequence, the fastest region, which, at the same time when I was at the Wilson lab, was used by Linda Vigilant to look at the ‘Mitochondrial Eve’ story, the migration of humans out of Africa. And she found much more variation using the same gene and primers as I, within human populations that are famously invariant, than I did in the Victoria cichlids. So you know, at first I thought, “Oh, my God, is this contamination? Why are they all identical?”  That took, really, Allan’s prodding or insight that this is the most likely explanation for the pattern that I found.

HS: How long did the writing take?

AM: I think I can write pretty quickly, but I don’t recall. Again, of course, these were days when, you know, we were still using WordStar or WordPerfect. Allan, of course, had good connections to Nature -there were several people in his lab who had Nature papers – and we knew this was a big message. So, we sent it to Nature. I don’t remember the reviews, but they were, I think, quite favorable. There was another paper that I worked on, on lungfish and coelacanths, that was rejected from Nature, and ended up being published in Journal of Molecular Evolution, but I remember how Allan wrote to Nature to, sort of, scold them and say, how could you reject this paper? This one was done in a week or two; didn’t take long. And, of course, in those days, everything was slow faxing, and physically sending things and so on, and that’s why it was published in 1990 rather than while I was still a postdoc in 1989.

HS: Could you tell us how you and Allan worked together on the manuscript?  Was he involved a lot in the writing?

AM: Yes, I think I actually still have some of his comments on this manuscript (I kept the fax and keep lots of paper in my office), somewhere in some drawer. I don’t remember that they were so extensive. Like I said, Allan was – this sounds so bad – was very good in spinning a story. You know how it is; you have to have a message when you send a short three or four page paper to Nature. It has to be something sharp, clear and to the point. And Allan was a great teacher to all of us, teaching us to focus on one or two major findings of a study. I moved to Stony Brook in April of 1990. Looking back at the dates when we submitted this – “Received 30 July; accepted August 20” – so, it really went pretty quickly. And this second version was done, I guess, while I was already in Stony Brook, but the paper was written before that. But you know how it is with Nature. Sometimes they will say it was received on a certain day but that might have been Version 2. I was in contact with Allan while I had my first job in Stony brook, quite regularly, because we had also continued to work on other projects after I left his lab. Like I said, this was, at least in my recollection, not such a difficult paper to get into Nature, because the message was new and important and the method cutting-edge as well.

HS: Were the other two authors involved in the writing?

AM: Tom, initially by training me in 1987-88, but later less so, I don’t even know if he was still in Berkeley then. Allan was always very nice to have all kinds of minorities and people from foreign countries in the lab, and I’ve done the same over the years, always had students from Africa, from Uganda, Kenya. I have somebody right now from Botswana. It’s sort of a way, I guess, to give back a little bit, to train people from those countries where one catches the fish and so on. It’s the same with Nicaragua. I have a student from Nicaragua. I don’t want to belittle Pereti’s contribution, but his main contribution really was to bring the fish and the tissue, and identify the fish, maybe; I don’t remember this in detail. But I still have those tissues in my freezer. Over the decades, many, many tissues have been accumulated, here in Konstanz and in the seven years before when I was in New York.

HS: Can we go over the names in the Acknowledgments to get a sense of who these people were and how they helped?

AM: Sure.

HS: K. Barel

AM: He’s one of the people from Holland, from Leiden in the Netherlands, one of the functional ichthyologists that I knew quite well.  I had taken a course there as a student in 1984, and I had been in touch with them. And, like I said, some of the tissues came from Leiden. The same is true of Rob Hoogerhoud. Barel was a professor and Hoogerhoud was a student. Irv Kornfield, also a very dear friend, is a professor at the University of Maine. He had worked on cichlids before, also. He was a PhD student of Dick Koehn, who was a professor and friend of mine in Stony Brook. Dick had done some of the early classical work on LDH and Mytilus – highly important work on protein electrophoresis and protein variation and adaptation to temperature and so on. Irv Kornfield worked on cichlids. Peter Reinthal left science, but he used to be a Malawi cichlid expert, sort of like Irv. Melanie Stiassny is a curator of fishes at the American Museum of Natural History. Her specialty is cichlids. She and I, later on, wrote a review paper for Scientific American in 1999. Frans Witte is also from Leiden. So, Barel, Hoogerhoud and Witte are the main people that send me fish. Irv and Peter contributed, I think, the Malawi cichlids that are part of this paper. And, maybe, Melanie contributed Tanganyika cichlids; I don’t quite remember, 30 years later, off the top of my head.

HS: What about D. Irwin?

AM: Dave Irwin was also a postdoc at the Wilson lab. And he’s one of these people that I admire – super nice guy -that I would place more in the in the ‘gel jock’ area than in the “ologist” group in the Wilson lab. He was really good in the lab, and he helped me initially with the techniques.

HS: M. Nishida

AM: Mutsumi Nishida, a famous Japanese fish evolution person. I don’t quite recall, but he must have come to the Wilson lab to discuss the project. E. Prager was the main technician at Wilson’s lab. This is almost 30 years ago so I don’t remember the details, but I’m somebody who has a hard time throwing things out. I have boxes of correspondence, and I know that I have the fax of the manuscript that Allan wrote on, because I came across it the other day, while I was looking for something else. I must have discussed the results with Mutsumi and Ellen Prager, or even shown versions of the manuscript to them, but I don’t remember the details.

HS: How was the paper received when it was published? Did it attract a lot of attention?

AM: Yes. John Avise wrote a “News and Views” in Nature about this. Presumably, he was a reviewer; I don’t know. This happens sometimes, that the people that write the “News and Views” were reviewers. The paper was written up in New York Times and German newspapers also. It was picked up, I think, in quite a few media.

For my career, like I said, it was very important. I remember when I was still a student or first year postdoc, I think in 1988, I went to a meeting of the American Society of Ichthyologists and Herpetologists (ASIH) in Ann Arbor, Michigan. I was giving a talk on PCR and the first cichlid sequences, the ones I had sequenced that I had collected in Nicaragua. And it was such a, sort of, sensation, that I was invited to repeat my talk twice. I gave the same talk three times at this meeting (and two people offered me postdocs). People were so excited by this new method and the new results. It was game-changing that you can actually sequence DNA sequences. And that helped me quite a bit in my career. Several people wanted me to be a postdoc in their lab and stuff like that. It was difficult for me because, being a German student in the States, t was not clear if I could stay on in the States after my PhD, because I didn’t have a green card. And there were only a handful of fellowships I could apply for, because I didn’t have citizenship or green card. But it was a very exciting time, with a happy end. I was just in McGill a few weeks ago for a talk. And I remembered, at that time, I was asked to be a postdoc in that lab at McGill, and also in David Hillis’s lab in Texas. The methods were so new and I was one of the few people that had used them for molecular evolutionary work. And so, yes, it stirred up some interest and I had many interviews and job offers from several prestigious universities in the years after my first Nature paper in 1990 (but, I had a Nature paper pretty much every year after that for some years).

HS: What kind of an influence did it have on the future course of your research? Did this become the major focus of your research immediately afterwards?

AM: Yes. I was quite young. I was offered the job in Stony Brook as an assistant professor when I was 27. I moved there after I finished my postdoc, so I was still only 29 when I started my job. My first students were older than me; my first postdocs were older than me. So, it was very exciting to start my own lab. And, I had quite a few visitors, many of whom are still friends, who wanted to learn these methods from me. Just like in the Wilson lab in the 1980s, there were many people, who are quite famous now, who visited. Yeah, it changed my life, you know. I had a Nature paper every year, at the time when I was on the faculty of Stony Brook. Many job offers. It affected my life a lot. And like I said, it was like a kid in a candy store, to sort of say, oh, you know, I know that people are debating how turtles are related to other amniote lineages, so we’ll work on turtles; oh, the lungfish-coelacanth question; or the origin of whales. All of these papers ended up being, whatever, Science, Nature, PNAS, papers. And, it was very useful that we could use these methods faster and earlier than other people, in combination with being classically trained, I guess, already, in Germany in comparative morphology and phylogenetics.

HS: At the time when you’re doing this work, did you already have an inkling that this was going to be important? Was that something you thought about?

AM: I guess what happened to me at that ASIH meeting, that, as a student, I would be invited to repeat my talk twice, so that more people could come to hear it, was a hint that this is something interesting and exciting. Christian Sturmbauer was my first or second postdoc and we had a Nature paper in 92, with Michel Milinkovitch in 93 and another on swordtail fishes in 94. Not every paper ended up in Nature, but it was certainly a time where this method was at the forefront and could answer all kinds of important questions. Now, of course we do entire genomes. We are revisiting some of these questions, but the answers often remain the same. And so, yes, by today’s standards, of course the database then was ridiculously small, but still, again, for many questions the answers remain the same. I think this is quite typical. I tell this story to my students about Susumo Ohno who discovered that fish-specific genome duplication or, more generally, the importance of gene duplication in evolution. The database he had was laughable by today’s standards. I mean, he would take photographs of karyotypes, cut out the photo paper, and weight the paper to see how big a genome is. So primitive were the methods that he used. By today’s standards, of course, you know, also hair-raisingly inaccurate. But still, I guess, like I said, because of the manual work, I was closer to the data. I could, you know, recite by memory, good chunks of the cytochrome b sequence and knew many amino acid motifs of this or other genes that I’d sequenced many, many times. I knew where the variable regions were, and you could tell a transition from a transversion, immediately, and so on. You were really close to the data, and I think that’s worth a lot. Today, when you have next-gen sequencing and you get terabytes of data, you don’t look at it anymore. You can’t even really look at it. At least, you don’t have, sort of, an immediate sense of the data.

HS: Do you have a sense of what this paper’s been mostly cited for?

AM: I haven’t really checked this. I presume, both, for the rate of evolution, and, maybe, the primary sequences that were in there. It was just, historically, one of the first studies to use this technology to address an evolutionary question. And, I guess the cichlids are, next to the Darwin’s finches, maybe one of the main model systems for rates of speciation and biodiversity. And, you know how it is, sometimes paper takes on a life of their own too, so I don’t know what exactly the paper has been cited for, in every case. We had another paper in 2003 – so 13 years later – in Science where we revisited this initial 1990 Nature paper with much more data and we end up with the same result, but of course a much more complete geographic and phylogenetic history of the colonization, origin and diversification of Lake Victoria cichlids. But it was the first study to assess the origin of an adaptive radiation through DNA sequences, so it is of interest to all kinds of people because of that. I mean, it’s also a textbook example, of course.

HS: Would you say that the main conclusions of this paper still hold true, more or less? And, if you were to redo this piece of work today, what would you do differently?

AM: The basic message has remained the same, tested by us and other labs with more and more data species and individuals included. Like I said, in the paper in 2003, we ended up concluding that there may have been two ancestral lineages rather than one. But the basic message holds, that there was, sort of, a cichlid eve, if you will, that colonized Lake Victoria and then gave rise to all of those species. We are now, of course, sequencing complete genomes. We have sequenced over 500 genomes of this group of species that we’re interested in, in Nicaragua, for example. And we’re now looking for and found genes and mutations that are responsible for phenotypic differences, for repeated adaptations, for species differences. We can now ask much more precise and, to some extent, different questions. In evolutionary biology, and probably also ecology, we like to think that those fields are driven more by theory than by methods or techniques. But I think the advent of PCR is an example that clearly shows that that’s not the case. With new methods, we cannot only revisit old questions, we can definitively also ask new questions. Like the whole ancient DNA field; that was not possible before PCR. And I always tell my students – I mean, it’s now a few years ago – but I sort of tell them, look, in my generation, PCR was the thing that changed the world; in your generation, next-gen sequencing is changing the world. You better be on top of this, you better know how to deal with these data, bioinformatically, and so on, because you can ask new questions. For example, the molecular basis of phenotypic differences, or the molecular basis of species differences, or what happens during speciation, what loci are involved and what mutations are involved. These are questions that were not doable 25 years ago, at least not in non-model systems. One of the things that I think was so important about PCR is that the data that were created were there for eternity. Cytochrome-b gene sequences, or whatever gene sequence, are in GenBank, can be associated with a particular species, as long as the species and the place where it was caught is documented. This is always something to compare to, whereas the kinds of data that were collected before that, restriction digests of mitochondrial DNA or starch electrophoresis data, or, later on, maybe with microsatellites, those are data that were lab-specific and also had a short shelf life. My most cited paper, this PNAS paper – Kocher et al. 1989 – has been cited more than 5000 times. And the reason for that is the primer sequences, and the methods of how you get DNA sequences out of single-stranded PCR that we developed at that time in the lab. Wilson called it the democratization of the genetic code, because it became cheaper and faster, and almost anybody could do it, whereas, it used to be much more difficult, more expensive and more elitist, if you will, only the best or richest labs could do this before that. And so, I think, since this paper was timely in that regard, and maybe showed what can be done with a very small number of base pairs, maybe that’s part of the reason for why it’s been cited so much.

HS: In the paper, you say, “the flock contains less genetic variation than does the human species”. Does that still remain the case, based on new genetic information and techniques?

AM: Extremely low. We were part of the cichlid genome project that was published in 2014 in Nature and that involved sequencing of five complete genomes. On average, there is less than 0.1 or 0.2 percent sequence variation between these species. They’re extremely homogeneous. Depending on the data and the quality of the data, there’s maybe a SNP every thousand base pairs or so between species that belong to different genera, a similar level of variation as in the entire species Homo sapiens.

HS: At one place in the paper, you talk about two ecological groups in Lake Malawi: sand dwellers and rock dwellers, and you say that these groups contain many more species, but you’ve sampled only a few, and so it might not be adequate to say anything about the association between phylogeny and ecology. Today, do we have a better idea about relationships within these ecological groups?

AM: Yes. This is a funny story. Tom Kocher and I, since we were the first to work on DNA sequences of cichlids, we, sort of, divided up the world among us. Tom had worked as an undergraduate in Lake Malawi, and he was more interested in Malawi, and sort of had, you know, the precedent there. And so, we agreed that I would stay away from Malawi for the rest of my career. He could work in Malawi, and I could do work on Victoria and Tanganyika. That’s, sort of, the agreement we had when we were both postdocs. My lab has revisited Malawi, to some extent. In subsequent papers, we sequenced a few more individuals, and the pattern has held up. There was a paper last year in Science by different groups of people from the Sanger Centre in England, and they had sequenced quite a few Malawi cichlids and they can, essentially, confirm what we had found before, based on mitochondrial DNA, that there is a group that lives over sand and a group that lives over rocks – they’re called ‘mbuna’ – and then there are three or four other lineages, like a big fish-eating group, and some others, that, also, we had already identified in papers in the early 1990s. In the early 1990s, we had a couple of papers and book chapters about this, and in the last 2-3 years I’ve had postdocs in my lab that were interested in this and have also confirmed these findings with complete genomes. The pattern has not held up 100%, but almost. The conclusions really have not changed that much. Maybe, we have much more detailed information now about how much sharing of genetic variation there is across lineages and so on, something that we couldn’t identify with the initial mitochondrial data sets, but the big picture was already established in that paper.

HS: With the new genetic information, has the systematics of the group changed a lot?

AM: No, not really. Lake Victoria is still a mess. Even with complete genomes. it is very difficult to decide what a genus is and the boundaries between species are fluid or nearly impossible to delineate objectively, with all the typical problems, including the philosophical issue of what a species is. So, systematically, Lake Victoria is still a mess. Malawi is a little bit more hopeful because it’s older and we have more genetic variation to work with. And actually, I have somebody in my lab that is working with me on that now. Tanganyika is the most hopeful. That’s where we have, over the last 20 years, my students and postdocs and I have a pretty good understanding of the phylogenetics. The entire cichlid DNA field, of course, has grown since Tom Kocher and I determined the first cichlid DNA sequences almost 35 years ago. It really started with Tom and me, and now our students, and our academic grandchildren, if you will – our students’ students – have established labs, and now there’s a community of, maybe 30-50 labs worldwide. When we have cichlid meetings, about one hundred people will come together and talk about cichlids. So, yes, this is somewhat beautiful growing.

HS: In the 1990 paper, you put the date of origin as 200,000 years for the Victoria flock and 700,000 years for the two groups in Malawi. Today, are those numbers still, more or less, the same?

AM: More or less. There’s, still some debate about this. We and others now think it’s even less in Lake Victoria, less than 100,000, maybe only 14,000, because there are some evidence to suggest that most, maybe all, of Victoria was dry only 14,000 years ago. For Lake Malawi, it is still in the same ballpark. The estimates range between million or two or three for Lake Malawi. Lake Tanganyika estimates are also around 10 million or so – that’s clearly the oldest. There are problems, of course, with the dating of these things, because the rate estimates, particularly among young radiations, especially if they involve hybridization or sharing of ancestor polymorphisms, make things very complicated. But the ballpark figures still are the same.

HS: In the very last sentence of the paper, you say,” we are losing the opportunity to study the cichlid fauna of Lake Victoria because much of it is going or has gone extinct as a result of the introduction of a non-endemic predatory fish.” What’s the situation today? Have more species gone extinct? Are there new problems?

AM: This is a tricky one. The introduction of Nile Perch by British fisheries officers- this is relying, mostly, on the work of the people in Leiden, who had been monitoring this from a fisheries perspective in the Mwanza Gulf in Tanzania, for three decades or so – initially, it seemed, had wiped out almost everything, or at least the abundance was very strongly reduced. And then there were some suggestions that certain ecological guilds – types that were in the open water more and those associated with rocks and so on – were affected by this. And there are still plenty of cichlids, and also diverse species of cichlids in Lake Victoria. But the fishery, of course, has shifted very strongly, now, towards Nile Perch. Introduction of the Nile Perch, and also the water hyacinth, ended up changing things. Also, because the Nile Perch is so large and the flesh is so oily, what people need to do in this area, since there’s no electricity or refrigeration possible, they cut down the trees to smoke the meat. And so, the cutting down of trees in the Lake Victoria region, leads to more erosion and therefore more water turbidity and so on. My perception is that it’s a very sad situation of a human-induced disaster, initially due to the Nile Perch, and then overfishing and so on and so on. When I was still in the US, I was part of a group of people that were trying to maintain aquarium stocks of many of these species. But that’s, of course, always difficult, because, you know, you still might get hybrids. Maybe, it’s not a hundred percent disaster, but it’s, certainly, a very sad of human-induced loss of biodiversity. And, unfortunately, the human population growth, pollution, and aquaculture have not only very much affected Lake Victoria, but pretty much all freshwaters in East Africa with consequent loss of, or at least threats to, much of the native fish fauna.

HS: When was the last time you visited Lake Victoria?

AM: This is a long time ago. Let me think. Maybe it was ‘97. I shifted then to work more in Lake Tanganyika, where I’ve been much more regularly.

HS: When did you last visit Tanganyika?

AM: In 2013 and students of mine have gone regularly since.

HS: Yu said you visited Lake Victoria, for the first time, in 1990. Was that on a research trip?

AM: Yes. I actually visited directly Pereti Basasibwaki in Uganda in the fishery center there and I collected fish in different regions in Uganda.

HS: Have you ever read this paper after it was published?

AM: Yes, surely. We I give overview talks I regularly use figures from it or refer back to it. I’m still proud of that paper.

HS: Would you count this as one of your favorites?

AM: Definitely in the top 10. Like I said, it was important, scientifically, and also for my personal career. Thinking back, like I said, the data basis is small, by comparison to now. I had my first cover in Nature with this study, although the cover was, kind of, a blurry photograph. It’s kind of funny. The cover photograph actually shows Malawi cichlids that were not even the focus of that paper. I think I got the photo from Peter Reinthal, because at that time I had not visited Lake Malawi myself. We’ve had other Nature and Science covers since then with cichlid fish on them. One of the other studies that I like, because it has a long history with me, is my involvement in the genome of the coelacanth. We had a Nature paper and cover for that in 2013. That question on the lungfish and coelacanth is something that goes back since I was a boy and read a book on the discovery of the coelacanth. There’s a very famous book by JLB Smith, a South African ichthyologist, who talks about “old four legs”, and it, sort of, has this sort of African mystique of the dark continent and all these adventures catching fish and so on. I remember, I probably was, 12 or 14 when I read this book in the German translation (where it’s called “Vergangenheit steig taus dem Meer”). I think I do this out of personal satisfaction and fascination with biodiversity. I am not ashamed to admit that I like fish. I like watching them. I tell all my students, you should have an aquarium on your desk, to, sort of, get into the head of the fish, so that you can ask biologically more meaningful questions. You have to try to become one with your organism, to ask, what’s the world of fish like? Of course, I can take a step back and say, well, this was important for rates of speciation or molecular evolution or whatever, and those things are very exciting, but I still approach much of my research, intuitively, sort of, from my belly! You know what I mean? I just like nature and its breathtaking diversity. For me, cichlids are not primarily a model system- yes, of course they are a model system that we helped to get established, but – I’m just curious about the diversity out there, I just like watching nature and I like to swim with fish and dive and see them in their natural habitat. And, being in Konstanz and in Germany is a privileged situation where a society has enough money to afford people like me, who just love what they do, to ask basic questions in science that will not immediately result in the new products or something applicable or something that money can be made from. I’m always aware of this privilege, that I’m able to follow a whim, and to get paid to ask fundamental questions in evolutionary biology is not something I can expect every government to fund.

HS: What would you say to a student who’s about to read this paper? What should he or she take away from it? Would you would you add any caveats to their reading?

AM: I think it is very difficult to think back what it was like before we had smart phones, or before there was PCR. Nowadays, everybody lives life as if we’ve always had smart phones or email or internet. And I think it is difficult for them to see what science was like 30 years ago. I know, because these methods aren’t used anymore, it’s hard for them to imagine that you can sequence DNA with radioactivity. Why would you do that? You can do it with fluorescence now. So, I view this a little bit as a lesson of how –Avery or Watson or Crick or whoever – how they worked in a generation or two before me, to try to imagine what that was like and what the data were like and how cumbersome it was to get only a little bit of new information. I think this is maybe a lesson in that, that it’s difficult to understand, from a young student’s perspective, in particular, how these kinds of technical advances really contributed to progress in science generally and to better understanding the patterns and processes that describe and shaped biological diversity on our planet.

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