Evolutionary Ecology of Plant-Aphid Interactions

Genetically based color variation in four green peach aphid clones

Bringing hundreds of paper bags stuffed with aphid-infested plants inside our lab was a very bad idea. Thus began the aphid infestation. All the lab benches, pipettes, centrifuges, computers, everything, were crawling with aphids seeking a new host. Winged aphids swarmed around the lights and windows, and all the molecular-based work my lab mates were doing came to a halt.

Green peach aphid (Myzus persicae)

This was the aftermath of a field experiment I (well, actually Marc Johnson and I) conducted, testing the relative ecological importance of herbivore genetic variation and contemporary evolution on plants. In most systems we still don’t know much about the ecological importance of ongoing evolutionary processes. My goal was to directly compare ecological impacts of genetic variation and contemporary evolution to the presence/absence and abundance of a species, two ecological factors we clearly expect to be important.

I studied interactions between green peach aphids (Myzus persicae) and two host plant species (Brassica napus and Solanum nigrum). Aphids are insect herbivores that stab their pointy mouthparts into plants to feed on phloem. In large numbers they can be devastating to plants, and rapidly growing to large numbers is their specialty. During growing season most aphids reproduce asexually, giving live birth to dozens of adorable little menaces that can grow to give birth themselves in as little as four days. You can watch exponential growth in action over the course of several weeks.

Momma aphid and her kids

This natural history makes aphids a great system in which to study the ecological effects of genetic variation. I collected populations of green peach aphids from tobacco fields all across eastern North Carolina and established colonies starting from just one aphid. Because they reproduce clonally each colony contains no genetic variation. For my experiments I choose four aphid clones that represented the full range of population growth rate on plants in the lab. These four clones could be distinguished using molecular markers, a fact that will become important below.

Colonies of 30 different aphid clones collected in North Carolina 

To test the ecological effects of aphid genetic variation on plants I started populations of each clone on caged plants in the field. I also had no-aphid controls. Then the aphid counting began: kneeling in the field, flipping over every leaf, and clicking away (with a tally counter) for several days straight. Some of the plants had up to 3000 aphids after only 11 days. Once I had estimates of population size I harvested the plants and dried and weighed them – these were the plants that I foolishly brought into the lab.

Marc Johnson and I glad to be done transferring hundreds of plants into the field

I found, not surprisingly, that plants with thousands of aphids had greatly reduced biomass. And while aphid abundance was the most important predictor of plant biomass, aphid genotype and aphid presence/absence had similar effects (i.e. they explained similar amounts of variation). This suggests that which genotype of aphid lands on a plant may be of similar ecological importance to whether the plant does or does not have aphids on it at all.

Field of ghosts experiment. Each plant was caged in polyester bags to prevent movement of aphids between plants.

Aphids’ asexual reproduction and rapid population growth rates also make it possible to track evolutionary change over short timescales (a few weeks). To do this I started genetically diverse aphid populations with equal numbers of all four of my genotypes.  This provided standing genetic variation allowing populations to evolve as they grew. At the end of the experiment I collected a random subsample of aphids from each plant, brought them back to the lab, and identified the genotype of each aphid using microsatellite molecular markers. With this I inferred genotype frequencies on each plant, and by comparing a population’s starting genotype frequency with the final genotype frequency I quantified change in genotype frequency over time – AKA evolution.

I tested whether the host plant influenced the evolutionary rate or trajectory, and whether evolution feeds back to influence the host plant’s growth. I found that aphid populations did evolve over approximately 5 generations, but at the same rate and direction on both host plants. Basically, faster-growing clones increased in genotype frequency whereas slower-growing genotypes decreased.

Aphids collected from the field and ready for genotyping

I also found some evidence for an ecological impact of contemporary evolution on the plants. On one of the host plants, faster-evolving populations had a larger negative impact on the plants. The magnitude of this effect was comparable to the impacts of aphid presence/absence and abundance that I mentioned earlier. 

In the world of plant-herbivore interactions there are quite a few studies showing how plant genetic variation influences herbivores, but there are very few studies testing the impacts of herbivore genetic variation on plants. So I’m pretty excited to provide a clear example showing that herbivore genetic variation matters too.

More generally, we know very little about the ecological importance of ongoing evolutionary change, in no small part because it is difficult to study in the field. But, similar to bringing thousands of live insects into a clean lab environment, assuming contemporary evolution is not important is probably not a good idea.

My car full of aphid infested plant collections

Reference: Turley NE, Johnson MTJ. 2015. Ecological effects of aphid abundance, genotypic variation, and contemporary evolution on plants. Oecologia. Link to the paper here.

All photos © Nash Turley, used with permission. 

Chance and direction in research

[This is post by Jessica Abbott.]

Since Andrew Hendry was kind enough to write a guest post about his career path to date, I was invited to return the favour. As with most researchers I know, my career path has been considerably influenced by chance events. In fact, now that I think about it, you can see this effect pretty much as far back as you want to go. Andrew started his story with his MSc work, but I’ve decided to put a bit more focus on the things that got me started on the road to research. I regularly give lectures for high school students, and one of the things they’re often interested in is how I decided to become an evolutionary biologist. Besides, all you have to do is look at my CV to get an idea of the things I’ve done during and after my PhD.

Some people you meet in science ended up there despite the fact that it was never their childhood dream. Others always wanted to be researchers. I fall into the second category. Ever since I was a kid I was interested in science, especially biology and astronomy. I first became interested in evolution when I read a book about it in 6^th grade. At that time I didn’t really realize that you could be a professional evolutionary biologist, though, so I never really considered it as a possible career.

By the end of high school I had settled on marine biology as an interesting field. But I didn’t want to work with dolphins! At some point I’d seen a lecture by a local researcher from Trent University, who talked about the development of new cancer treatments from naturally-occurring chemicals (for example taxol, which is derived from yew trees and can be used to treat ovarian cancer). She also mentioned marine sponges, and how they might be a promising subject for similar research since they have effective but relatively non-specific immune function. This sparked my interest as a way to combine research in marine biology with some practical applications. I therefore decided to study marine biology at the University of Guelph during my undergraduate degree.

Suberites domuncula, by Guido Picchetti. Charismatic, no?

It was my first-year introductory zoology class that really made me start thinking about evolutionary biology. Ron Brooks taught the class and basically seemed to completely ignore the material that was supposed to be covered in the course, at least judging by the information we covered in the labs. Instead he talked a lot about evolution and told everyone to read The Selfish Gene. I was a good student, so of course I read it. And it made me realize that this was the sort of thing that I really wanted to work with.

I also wanted to broaden my horizons on a personal level, so I applied to go on an international exchange for my third year. My destination, Lund University in Sweden, was pretty random. I had originally applied to go to Aberdeen or Sydney, because they were the only two places that had marine biology programs (at least among the universities that Guelph had a reciprocal exchange agreement with). But because both these locations were highly popular (meaning only one semester abroad was allowed) and I wanted to go for a whole year, the exchange office suggested some other options. Lund seemed to have the most interesting selection of courses, so that’s where I decided to go, despite knowing basically nothing about the country or the university.

Lund is lovely in the spring.

Once I got to Lund, I really liked it. The classes were small and the material was interesting. Swedes were hard to get to know, but nice once you knew them. It was fun learning a new language. And of course I met my future husband. So rather than go back to Guelph I registered as a student in Sweden for the next year. And near the end of my second academic year in Lund I started a master’s project with Erik Svensson. My choice of project was also somewhat random. Because I was interested in evolutionary questions in general, I wasn’t so picky about the type of study organism. I asked around to find out who had a project that needed a student, and just went with the one that sounded most interesting. That’s how I ended up working on Ischnura elegans. When the opportunity arose to continue working with Erik in the same system, I took it.

As I neared the end of my PhD I started thinking about what to do next. I was never especially enamoured with field work, so I thought it would be fun to try working with a lab-based system. I was interested in the evolution of sexual dimorphism (I’d done a bit of work on sexual dimorphism during my PhD), but also in genetic conflicts. I’d run across Bill Rice’s work on intralocus sexual conflict (then often called ontogenetic sexual conflict) which combined both of these things, but at that point there weren’t so many people working in that area, so it wasn’t really on my radar. Then I went to ESEB in 2005 and saw a talk by Russell Bonduriansky about intralocus sexual conflict. It made me realize that this could be a viable option after all. I therefore got in touch with Adam Chippindale to see about doing a postdoc with him.

Adam’s response was a pretty typical one – he’d love to have me as a postdoc but didn’t have the money to hire me himself. But he was happy to help me out in designing a project so that I could apply for my own funding to go to Queen’s University. I applied to both NSERC and the Swedish Research Council (VR), and was successful with VR. That’s how I got started working on experimental evolution, and Drosophila, a method and a system which I still use today.

When we moved to Kingston we had hoped to stay longer than the two years of my VR fellowship, but when I applied for an NSERC postdoc again (my last chance) I wasn’t successful. The choice was between returning to Sweden with a new repatriation fellowship from VR, or being unemployed and living in my parents’ basement. I think you can guess which was the more attractive choice. That’s how I ended up in Uppsala, working with Ted Morrow. I took my fly populations with me and continued the stuff that I’d done at Queen’s in Uppsala.

I liked the fact that there were a bunch of sexual conflict people in Uppsala, and I liked working with Ted. When my one-year repatriation grant was up, I was lucky enough to be offered a one-year postdoctoral stipend by Klaus Reinhardt, funded by the Volkswagen Foundation. During that period I continued to work in Uppsala, but on a collaborative project with Ted and Klaus. The stipend kept me going until I was successful in obtaining a Junior Researcher Project grant from VR.

Macrostomum lignano mating, by Lukas Schärer.

The Junior Researcher grant let me start up my own small group, and start work on a new study organism, Macrostomum lignano. (The story of how I decided to do a project on Macrostomum is also interesting and much influenced by chance events, but I won’t go into details here. This post is long enough already.) Although I considered staying in Uppsala, in the end I decided to move back to Lund, both for personal and professional reasons. I liked having a lot of people that shared my interest in sexual conflict in Uppsala, but the downside was that it meant that I was just one of many, and that I wouldn’t necessarily bring anything new to the department. Lund was also closer to old friends and my husband’s family. I’ve been working here since 2012.

Looking back, it’s clear that both chance and direction have played a role in my career path. In many ways, I’m exactly where I had hoped I would be at this stage, when I imagined my future as a teenager. I imagined myself working at a good research university (preferably abroad), in a good relationship (maybe kids – not essential), combining research, teaching, and popular science in an enjoyable mix. These things are all true (that’s where the direction part comes in). However exactly what I’m working on and where I am are different than what I expected (that’s where the chance part comes in).

It’s also been a lot harder than I had expected it to be. It’s not like I thought being a researcher would be easy. But being a postdoc with no option to plan long-term, no job security, and a family, was much harder than I had expected. A common theme when senior scientists talk about their career paths is “I just worked on whatever I thought was most interesting, I never tried to think strategically”. I know that PhD students and postdocs can find this a bit frustrating – even if this approach is perhaps a necessary condition for success, it’s probably not sufficient. There’s probably just as many people out there (or more!) who followed their hearts but didn’t get that tenure-track job or key big grant, as the ones who did. I can understand this frustration, because “just do what you think is fun” is not very helpful advice. However, one can also look at it another way. It’s good to have long-term goals in mind (direction), so that you can take the right opportunities as they come up (chance). But if you’re not really enjoying your work while you’re working on it, what’s the point? Don’t spend a lot of time doing things you don’t like just because you think they’re strategic. You might get hit by a bus tomorrow.

Are laboratory studies useful for understanding the “real world"?

“It is interesting to contemplate a tangled bank, clothed with many plants of many kinds, with birds singing on the bushes, with various insects flitting about, and with worms crawling through the damp earth, and to reflect that these elaborately constructed forms, so different from each other, and dependent upon each other in so complex a manner, have all been produced by laws acting around us.” (Darwin 1859)

Some of what we hope to understand.

I just returned from an “eco-evolutionary interactions” workshop at Yale organized by David Post, David Vasseur, and Paul Turner. In his opening introduction to the symposium, Paul referred to research from the “real world” when speaking about something of applied relevance to humans. He then hesitated for a second and took a tangent to complain about the term “real world” as all of the work was in the “real world.” (The Turner World?) A bit later I was talking with Alvaro Sanchez about the value of lab experiments for eco-evolutionary studies – a different sort of “real world” concern. Alvaro was asking my opinion partly because he had seen an advance copy of my book in which I state:

Eco-evolutionary studies with real organisms could proceed in the laboratory or in nature. Advantages of the laboratory are manifold: populations can be genetically manipulated, environments can be carefully controlled, replicates and controls can be numerous, and small organisms with very short generation times (e.g., microbes) allow the long-term tracking of dynamics (Bell 2008, Kassen 2014). These properties dictate that eco-evolutionary studies in the laboratory are elegant and informative, yet only in a limited sense. That is, such studies tell us what happens when we impose a particular artificial environment on a particular artificial population and, hence, they cannot tell us what will actually happen for real populations in nature. Understanding eco-evolutionary dynamics as they play out in the natural world instead requires the study of natural populations in natural environments. I will therefore focus to the extent possible on natural contexts, although I certainly refer to laboratory studies when necessary.

Is this place to do it?

This exchange reminded me of a debate Andy Gonzalez and I had undertaken for the amusement of our department, and it seemed an appropriate time to revisit my side of that debate in this blog. Specifically, I would like to ask “What – if anything – can laboratory studies tell us about real world?”

What we seek to understand in any eco-evolutionary work is the chain of effects shown below: the effect of ecological conditions on organisms and the evolutionary/ecological outcome. The way laboratory studies typically address this question is to create variation in some focal ecological condition, such as temperature or carbon dioxide (or both), expose some model organisms (often only just one or a few identical clones) to those different conditions, and then measure the evolutionary/ecological outcome. The real world is much more complicated, of course; populations or species of interest might be exposed to differences in a great diversity of biotic and abiotic factors and might have large amounts of genetic and non-genetic variation in a wide array of relevant traits. These differences lead to two main limitations of laboratory studies.

1. By ignoring variation in other natural factors, laboratory studies cleverly isolate particular effects, but those effects are likely to be very different in nature.

These differences can arise in at least three ways. First, correlations can exist between explanatory variables that fundamentally shape the effect of each. Second, indirect effects cascading through other organisms or environmental conditions can amplify or offset the direct effects usually measured in laboratory studies. Third, unmeasured contributors or random noise can swamp causal effects. Interestingly, these complications are all cited as supporting the great strength of laboratory studies: by getting rid of these correlations and effects, one can get rid of the noise and get closer to the true mechanism. Sure, this is all well and good – but such studies don’t then tell you anything about the importance and nature of the actual effects IN NATURE (which is what we really care about). Instead such experiments only provide proof of concept.

2. The use of model (as opposed to natural) populations is sure to yield a misleading expectation.

Evolutionary responses of a given suite of traits will be a function of selection acting directly on the genetic variation for each trait and selection (and constraints) acting indirectly through genetic covariation with other traits. Unfortunately, genetic (co)variances in nature never equal those in nature, meaning that the observed evolutionary response will not match the response that would emerge in nature. Moreover, many laboratory studies use only a single clone of organisms so that they can assess the effects of new mutations. Wonderful – but most immediate responses to environmental change in nature will be driven by standing genetic variation, which that approach eliminates.

Similarly, ecological responses to changes in a given suite of traits will be a function of the distribution of traits in the population and the effects of that variation on ecological variables. Sadly, the distribution of traits in the lab will (for the above reasons) never match the distribution in nature. In addition, the simplified laboratory environment will never capture how a given trait distribution will shape ecological responses in more realistic scenarios.

Having just argued from first principles that laboratory studies will not be predictive of evolutionary or ecological effects in nature, I will provide a few examples by way of illustration.  

Amphibians are a classic system for studying the effects of competition. Skelly and Kiesecker (2001 – Oikos) performed a meta-analysis of how the experimental “venue” (lab, mesocosm, field) influenced estimates of how competition influences amphibian growth. Based on 227 comparisons from 52 studies, the authors showed that the effects of intra-specific competition (white bars) were reasonably consistent across venues but the effects of inter-specific competition (black bars) were dramatically different between venues. Results from the laboratory from mesocosms and from experiments in natural populations all yielded very different results.

Many laboratory studies have shown that exposure to novel parasites causes the evolution of resistance to those parasites. Similarly, releasing populations from parasite pressures leads to the evolution of reduced resistance, presumably because resistance is costly. We tested the latter expectation in nature through assays of resistance to a common parasite (Gyrodactylus) in guppies from a highly parasitized population that was introduced into four replicate environments that lacked the parasite. We found – completely contrary to lab experiments – that all four of the introduced populations rapidly evolved INCREASED resistance to the parasite they no longer experienced. Presumably this result arose because other factors in nature caused selection on other traits that were pleiotropically coupled to resistance. The lead author (Felipe Dargent) wrote about this work in a previous post and the work has since been the subject of a comment-response that specifically contrasted the complexity of nature as a weakness or strength of experiments in nature.

The populations not exposed to parasites (LL, UL, T, C) are now MORE resistant to the parasite than is the ancestral population (S) that remains exposed to parasites.

The position I have taken in this post is: if we are to understand nature we must work in nature. Of course, this doesn’t mean that we shouldn’t also work in the lab. In particular, laboratory experiments provide critical proof of concept that a given ecological factor can have a particular evolutionary effect under a given set of conditions. Similarly, they provide proof of concept that a given evolutionary change can have a given ecological effect under a given set of conditions. The critical next step, however, is to then test key aspects of those outcomes in the messy natural world.

So – in the end – the study of a given eco-evolutionary effect would ideally couple laboratory and field (and theoretical) studies. Reassuringly, this was a major theme of the working group – designing questions and experiments that can be explored simultaneously in theory, in the lab, and in nature. I look forward to seeing what emerges.

The crew!

Jokes Go In The Acknowledgements - or Anywhere on April 1

Much ink has been spilled over the dry and boring nature of scientific writing. This style stems, of course, from the desire to communicate information in the fewest possible words with the greatest possible clarity. I generally agree with this goal and sentiment. At the same time, however, the quality of the writing itself can increase the appreciation of science and make you WANT to read that paper that is sitting there on your desktop. 

So how best to accomplish this balance between clarity/brevity and an appreciation of the writing itself. Two approaches are common. First, in what we might call the "subtle" (or even subversive) approach, authors outwardly adhere to the formal scientific style but sprinkle fun details into key parts, most commonly the title, the figures, or the acknowledgements. Second, in what we might call the "overt" approach, authors throw out the rules and find a good venue to just write a fun paper from start to finish. In the spirit of Steve Heard’s wonderful “On Whimsy, Jokes, and Beauty: Can Scientific Writing Be Enjoyed,” I wanted to here provide some examples of these approaches, both my own attempts and the successes of others.

In the subtle approach, jokes and whimsy often appear in titles or figures. The title option is most common, where one simply adds a colon between the serious and funny parts of the title. This approach is very common and sometimes works and it sometimes doesn’t. Here are a few examples and I will let you judge which work well and which would best have been avoided.

The good and the bad of colon-based title jokes.

The figure option is also somewhat common, and I have already posted a “Top Ten best scientific figures” to outline some great examples, with a few shown here. 

Probably by far the most common way to sneak some surprises into a paper is in the acknowledgements, as these are either not carefully read by editors or simply a place where they are willing to allow some lee-way. Posts have already been written about how acknowledgements are ways to insult or criticize reviewers or colleagues – but they are also good places to follow your whimsy muse. Here are some of my own attempts.

From a paper in Conservation Genetics in 2000, titled "Questioning Species Realities"

From a paper in Journal of Evolutionary Biology - the unnamed Deciding Editor with the great sense of humor was Rhonda Snook.

In the subtle approach, one has to hope that either the reviewers or editors don’t read carefully enough to catch your jokes – or that they catch the jokes and have enough of a sense of humor to allow them to continue regardless of you thumbing your nose at convention. Sometimes it works and sometimes it doesn’t - an instance is noted below for a "Whither Adaptation" paper I published.

In the overt approach, one really needs an editor who buys into the whole thing by either allowing you to produce a joke paper from start to finish or by allowing you throw out the norms of writing even if the paper and topic itself is not a joke. Even serious journals will sometimes allow joke papers to be published, with three examples given below.

Probably the best tongue-in-cheek paper ever writen.

For serious papers that are written in an unconventional way, whether tongue in cheek or self-mocking, you often have to get out of the serious scientific journals and into “lower-tier” journals or journals that allow some “philosophy.” Here are a couple of examples. 

Much of the paper was written in this style but we also wanted to add an asterisk to the author names with the indication "each author things to other contributed less." However, it was excised despite repeated requests.

Of course, another solution is to publish your paper on April Fools, such as this paper published today in Nature.

Happy writing!