Sunday, December 29, 2013

Homage to the Island of Misfit Toys, or why do wood frog populations seem locally maladapted to roads?

When I first began my dissertation research—bright-eyed, undaunted, and operating on full nights of sleep—I set my sights on roads, amphibians, and evolution. This was something of a transition from my previous incarnation, which is perhaps best described as a recovering liberal arts enthusiast with a mild interest in birds. Fortuitously, I think, I heeded my dissertation advisor’s suggestion by taking my “eye off the sky” in favor of what most might consider swamps.

A roadside pond primed for wood frog breeding. Due to its proximity, this pool has high concentrations of road salt. Yet, days after this photo was captured, hundreds of adult wood frogs began arriving to breed and lay their eggs.

It was in those swamps that I set about testing the predictions of evolutionary mediated responses to roads and runoff. With unabridged optimism, I set out to conduct a suite of experiments in my first field season. Seeing as how I was a complete newbie—having chosen to spend most of my undergraduate years embracing the youthful drive for renaissance over that of developing actual research skills—I had something to prove.

Renaissance, not research; me on the trombone in 2001.

Naturally, I chose to offset my lack of training with a healthy dose of enthusiasm and a ‘go-big’ approach. This amounted to doubling or tripling most sensible recommendations for sample size (and infrastructure). This conceal-inexperience-with-quantity logic seemed clear to me—if I could not impress my dissertation committee with a deep knowledge of ecology and evolution, I would impress them with my willingness to work hard. In reality, this overambitious plan required me to draggle friends and family aboard my sinking ship (of fool) to complete all the necessary sewing, dremeling, aquarium cleaning, surveying, and to of course keep me somewhat nourished if not adequately caffeinated.

My parents pitching in -- sewing tadpole cages late into the night.

Now before I discuss some actual biology—and hopefully make good on the title of this entry—let me backtrack a bit: why study roads? Well, there are an awful lot of them on the landscape. In the U.S. alone, we have enough paved roads to build a 25-lane highway from here to the moon, with a 12-lane soft shoulder to boot. Roads account for about 1% of the U.S. landscape, about the size of the state of Maine. And roads spur a host of consequences on habitats and organisms, from fragmentation to roadkill to runoff contamination. Among contaminants, road salt takes the cake (or the anti-cake if ferrocyanide is added..[1]). For example, in Connecticut, where I did most of this road-effects research, road salt is applied upwards of 30 tons per lane mile over the winter. That bears repeating: over the course of a single southern New England winter, each mile of road receives 30 tons of salt. That’s roughly the entire contexts of five dump trucks. By extension, over the course of the winter, the equivalent of a dump truck of salt is emptied every thousand feet. Bear in mind, this is per lane of travel.

The point is that that’s a lot of salt. Plus, salt use in deicing has been on the rise. Road salt is so widely and increasingly used that beginning in 2005, the deicing industry became, for the first time ever, the leading consumer of salts. This rather epic application of salts on the landscape has attracted a lot of research in environmental sciences (though for some reason, evolutionary perspectives have seldom been incorporated; I’ll save that for another blog entry). There is also interest stemming from public health as the increasing salinization of freshwater is now impacting private wells, which has raised concern for individuals with hypertension. While the effects of exposure to road salt varies, most organisms living near roads (which is to say, most organisms) do not fare especially well. Consequences can include arrested developmental and growth rates, behavioral modifications, and increased mortality. That is to say that road salt appears to be an agent of selection in roadside habitats. Not to mention, it’s rather tough on my 20-year-old Civic.

So, flashback to 2007 and my newfound interest in amphibians, roads, runoff, and the potential for contemporary evolution. I was investigating road effects in a system of pool-breeding amphibians in northeastern Connecticut, the so-called Quiet Corner of the state. Specifically, I was looking at road effects on the wood frog, a smallish frog that breeds in early spring, when adults abandon their subterranean winter hideouts in favor of ice-cold waters. They do this for one reason: to mate. Wood frogs are explosive breeders. While this term would undoubtedly yield interesting results on Urban Dictionary, here I am referring to the fact that a given pond can go from completely amphibian-free to overflowing with hundreds to thousands of adult wood frogs in a matter of days. It is quite the sight. And then, almost as quickly as it began, the breeding is done. The adults hop off to the woods, leaving behind the products of their reproductive endeavors; this amounts to about 800 externally fertilized eggs per female, glommed together in a single, gelatinous mass.

A wood frog on the move. Photo credit: Geoffrey Giller.

Life is tough for any of these newly laid embryos—turtles might come along and nab them up; water levels can quickly drop, leaving eggs high and dry; or temperatures can fall, causing eggs to freeze. But things are even tougher in roadside ponds. Because of the small and shallow nature of these ponds, contaminants can accumulate at high concentrations. For example, I was finding that ponds located about 30 feet from the road were 10% as salty as the ocean. This is not normal; Northeastern CT is far from the ocean. Indeed, ponds a couple hundred feet from roads were virtually salt free. Despite this impressive increase in salt in these roadside ponds, wood frogs and other amphibians are still breeding there. In fact, as I continued my work, I found no difference in the number of adult wood frogs breeding in roadside versus nearby woodland ponds.

So given this strong signal of road salt entering these ponds, the first thing was to figure out whether roadside ponds had any negative effects on the wood frogs breeding there. And the next thing was to figure out whether adaptive outcomes were present. As to the first inquiry, roadside ponds are indeed difficult places to make a living. I found that wood frogs in roadside ponds survive, grow, and develop at lower rates. So, there was evidence that roads were imparting a selective force. Coupled with this, there was good reason to believe that wood frogs were particularly good at adapting. For one, they tend to occupy rather distinct populations at small spatial scales. For two, there was already evidence of contemporary adaptation among populations in this system, albeit not to roads. Specifically, my dissertation advisor had recently reported adaptive responses to dynamic forest canopy regimes that differ across the scale of tens of meters. So given that adaptive responses to natural perturbations were already detected in this system, I expected to find evidence for local adaptation to roads. Indeed, with my excited (read: naïve) sense of vision, I found myself dreaming up the title of the Nature paper that I would write following my experiments, which I imagined would reveal contemporary evolution in an amphibian (a vertebrate, no less!) adapting to roads and runoff… the phrase “Salt saltation” seemed like it would be perfect on the magazine’s cover. Well, that prediction turned out to be squarely wrong. After a host of reciprocal transplants, salt exposure experiments, and field surveys, the data suggested that adaptation to roads for these animals was about as likely as my envisioned Nature paper. Whereas I expected roadside populations to show increased ability to tolerate road adjacency and exposure to road salt, I found just the opposite. Regardless of where embryos from roadside populations were grown—whether in their natal ponds, in nearby uncontaminated woodland ponds, or in various experimental concentrations of road salt—they performed worse than those animals from woodland populations. This was not local adaptation – but what was it? Could it be local maladaptation? And given these negative effects, how did these populations persist at sizes equivalent to those inhabiting the clean, woodland ponds?

Survival (±1 SE) is shown here as the mean proportion of individuals surviving to feeding stage across all experimental units (N = 99). The woodland deme is represented by open circles while the roadside deme is represented by filled squares. The environment in which the animals were grown out is on the x-axis. From Brady SP. 2013. PeerJ. DOI: 10.7717/peerj.163/fig-2

Before we can answer these questions, we should try to define local maladaptation. From a research perspective, local maladaptation has not seen the attention that local adaptation has, so finding a clear definition is somewhat challenging. I think it is reasonable to use local adaptation as a corollary. In this case we would define local maladaptation as a genetically-based process leading a local population to have lower fitness in its local environment compared to that of a foreign population within that same environment. So far, the pattern I found is consistent with this process. But here’s a twist: even when roadside wood frogs were transferred to the woodland environments, they still survive at lower rates. So what would we call this outcome? Well, for lack of a better term, I have been referring to this as deme depression, drawing on the analogous pattern of reduced fitness that is caused by inbreeding depression. Certainly, my description of these outcomes and their significance is underdeveloped here and deserves more discussion at some point. But for now, let’s move on toward accounting for how these patterns might be persisting in this system.

So given the locally reduced survival inherent in roadside populations, we have evidence for local maladaptation. More accurately, we can say that roadside populations ‘perform maladaptively’, but we do not know whether it is maladaptation per se. This is because maladaptation sensu stricto refers to reduced fitness owing to genetics. In the context of roads and runoff (and other human-modified environments), we might expect maladaptive genetic changes to be more common than, say, in natural environments. I offer this suggestion cautiously, as it is based purely on the known diversity of mutagenic contaminants found in roadside ponds. These include heavy metals like platinum and copper from catalytic converters and brake pad wear, and aromatic hydrocarbons (think toluene, benzene, PAHs) from fuel, tires, and the road surface itself. Yet, because the animals in my experiments originated from wild-laid eggs, we cannot say whether there is a genetic component underlying this performance deficit. Instead, this pattern of reduced survival—so-called maladaptive performance—might well be the workings of an experimental artifact. Specifically, it could be that the wild-laid eggs I collected were compromised by runoff the minute they hit the water. In other words, early embryonic exposure might have caused performance detriment in later life history stages. It is certainly plausible to think this might be happening, and so, I tested it. And it turns out that this was not the case: early exposure to roadside water had no carryover effects on performance. Still, environmental effects can be transmitted directly from parent to offspring without the offspring experiencing the aquatic environment first-hand. These so-called maternal effects (or, more broadly, inherited-environmental effects) are widespread in nature and certainly might explain maladaptive performance here. While I found no difference in egg size (a typical maternal effect in amphibians), a colleague and I did find that eggs are pre-loaded with elevated concentrations of methylmercury. The magnitude of methylmercury found in eggs corresponded to that found in the mother. So now we know it is possible for wood frog eggs to be exposed to and accumulate contaminants through their mothers before they are laid into the external environment.

Between inherited environmental effects and these putative mutagenic effects, we have one plausible and one maybe possible mechanism to explain locally maladaptive performance. We could also invoke inbreeding depression and/or drift as explanations, but since these populations do not differ in size, it is difficult to conceive of these processes playing out in the roadside environment but not in the woodland environment. Regardless of the actual mechanisms at play, we still need to figure out how these maladaptively performing populations persist. Could it be that roadside habitat is a verifiable sink, with woodland populations serving as the source? Perhaps. A rescue effect of sorts—wherein woodland animals are leaving to further colonize roadside ponds—could certainly explain why these roadside populations don’t appear to be dwindling in size. But if that were the case, we would expect a more similar phenotype between roadside and woodland populations. In other words, if roadside populations are sustained through the arrival and reproductive contribution of woodland emigrants, wouldn’t the performance traits of the offspring be similar between the two population types? Actually, the answer to that question depends on whether the emigrating population is a random sample of the woodland population. If it is, then yes, we would expect comparable phenotypes. But what if the emigrating group was not composed of a random sample of the population? What if instead the emigrating group comprised some biased subset of the population that was already compromised in some way?

Enter the Island of Misfit Toys. If you’re unfamiliar with the classic stop motion animation version of Rudolph the Red-Nose Reindeer, or if you’ve overlooked some of the finer details of the movie, here’s short a recap: (cue Solsbury Hill background music) a young Rudolph, made to feel reindeer non grata for his glowing nose, runs away from home. In the odyssey that follows, Rudolph meets up with another pariah (Hermey, the elf whose dream is not that of making toys but of mending teeth) and an eccentric silver and gold prospector (Yukon Cornelius). During their adventure, this crew of misfits eventually lands upon the Island of Misfit Toys, where all mal-designed, unwanted toys live out their days.

So perhaps, just as the misfit toys sought solace and companionship on their island, misfit frogs are seeking as much in roadside ponds. Perhaps misfittery loves company? This misfit island hypothesis could be playing out in a variety of ways, all of which would hinge on habitat-oriented dispersal into roadside ponds. For example, in wood frogs, we know that younger adults are relatively poor provisioners of offspring. Thus, misfit frogs might just be younger, and more likely to disperse into inferior (roadside) habitat. Alternatively, perhaps the dispersing animals are weaker (regardless of age), and therefore less capable of securing mates among the healthier animals that breed in woodland ponds, thus seeking companionship among the similarly conditioned animals in roadside ponds.

There are several other processes that might account for the persistence of these maladaptively performing roadside populations. One is that there appears to exist a quantity over quality tradeoff in reproductive output. Namely, while roadside embryos show inherently reduced survivability compared to woodland embryos, roadside females lay more eggs per clutch. So while the chance of survival is lower for a given roadside embryo, each roadside female lays approximately 10.5% more eggs than her woodland counterpart. Therefore, it is possible that this numerical investment helps offset population decline. Another possible explanation for the persistence of this maladaptive pattern is that roadside wood frogs might be adapted to later life history stages. For example, it may be the case that adults are locally adapted to the terrestrial environment. It is further possible that such an adaptation is associated with poor offspring performance, but that adult survival is more important for population fitness.

In the end, Santa promised Rudolph that he would find a home for all the misfit toys. Here, I have no such promise to offer—the fate of the (putative) misfit frogs remains an active area of research. So for now, this story is to be continued.

[1] Ferrocyanide is sometimes added to road salt to prevent it from caking up; hence the unfortunate pun.

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