Spotted Salamander Pond
Gaian Symbiosis in Miniature
Sacred Site of the Epic of Evolution
report by Connie Barlow, December 2011
JANUARY 1, 2012 - What better way to welcome in the New Year than to finally assemble my photos and notes from this past spring on Lookout Mountain, Alabama. This report will become yet another rumination on my personal experience with a "Sacred Site of the Epic of Evolution"! This one journeys back to our amphibian ancestry. It charts my multi-week participation in the life cycle of the world's second largest land-based salamander and by doing so, to viscerally sense the time when our vertebrate lineage was still dependent on water for its earliest stages of life.
As you will read, when I went online to learn more about the particular species I had come upon, even more marvels began to emerge. Thanks to a scientific discovery published only one year earlier, I would now have the opportunity to journey back even farther long before vertebrates evolved, long before eukaryotes evolved. I would experience, in microcosm, the primordial time when the bacterial realm was assembling the chemical cycles essential for life. Spotted Salamander egg masses enabled me to journey back in my imagination to that Archaean time because they so visibly display, what I like to call, Gaian symbiosis in miniature.
I'll begin this report with an entry I made in my journal. There you will see my excitement begin to build for, what would turn into, an opportunity to relive two profound transitions in our evolutionary heritage. As background, know that my husband and I were on a writing retreat in a cabin alongside the very rural home of our friend Tobey Miller in the northeastern corner of Alabama, from February 23 to April 24, 2011.
MARCH 1 journal entry:"Last night here in Tobey's forest on Lookout Mtn, AL, we heard Spring Peepers galore (it rained hard that afternoon). Michael at dusk was tossing branches and logs into Tobey's spring pond to support eggs of salamanders when they lay, and lo! he discovered a spotted salamander under one! He came back, got me, and we found 3 more when gently looking under logs near the ponds!!! First time I have seen the second-biggest land salamander in the world since Bets and I used to rescue them from Auntie Jo's basement window cellar in Michigan as kids. Wow!!!! Plus, tornado warning yesterday that drove us into Tobey's basement.
Later on March 1: "This afternoon I discovered 5 blobs of eggs in Salamander Pond new because three were attached to a large stick I tossed in just a day or two earlier."
ABOVE LEFT: March 2 at "Sallie" the Salamander Pond. (Notice this is very early spring, before anything greens up.) Salamanders lay their eggs only in ephemeral forest ponds such as this one. In a wet year, the pond will remain until the dry season of August. This is enough time for Spotted Salamander nymphs to grow lungs, become 2-inch-long juveniles, and venture out onto land.
ABOVE RIGHT: March 2, I temporarily pulled out to examine a glob of salamander eggs that had probably been laid the night before during the rain. The glob is attached to a leaf that was resting on the bottom, so it may have been laid a day or so before I tossed some hefty sticks into the pond in hopes of improving habitat for salamanders. (I knew big sticks are valued because, while living in the Pacific Northwest two decades earlier, I had come upon an ephemeral forest pond and noticed masses of newt eggs attached to floating or partially buried sticks.)
ABOVE: early March - Nearly every day I would visit "Sallie." I would wear water sandals so I could venture into the pond and monitor changes in any of three of the nearest egg masses.
ABOVE LEFT: March 14 - Notice the embryos are starting to take shape. Toward the right in that photo you can see two with their white bellies visible. One reason why I monitored the egg masses every day was that, on March 2 when I went online to learn how long it would take this species to hatch into swimming nymphs, I learned of a remarkable scientific discovery made in the previous year. I wanted to be able to see this amazing transformation myself!
ABOVE RIGHT: March 17 - This was the first day I detected the transformation. Notice how within the jelly mass, each embryo has its own protected sphere to develop within. Now notice how those spheres have turned bright green! Just the spheres are turning green not the enveloping protective jelly. What's going on?
ABOVE: April 22 (left) and April 23 (right) - On 4 August 2010, a leading science journal, Nature, published a report titled, "Salamander's Egg Surprise". Here is the article:Scientists have stumbled across the first example of a photosynthetic organism living inside a vertebrate's cells. The discovery is a surprise because the adaptive immune systems of vertebrates generally destroy foreign biological material. In this case, however, a symbiotic alga seems to be surviving unchallenged and might be giving its host a solar-powered metabolic boost.
The embryos of the Spotted Salamander Ambystoma maculatum have long been known to enjoy a mutualistic relationship with the single-celled alga Oophila amblystomatis. The salamanders' viridescent eggs are coloured by algae living in the jelly-like material that surrounds the embryo. The embryos produce nitrogen-rich waste that is useful to the algae, which, in turn, supply the developing embryos with extra oxygen. The algae clearly benefit their salamander hosts: Lynda Goff, a molecular marine biologist at the University of California, Santa Cruz, showed 30 years ago that salamander embryos lacking algae in their surrounding jelly are slower to hatch.
Ryan Kerney of Dalhousie University in Halifax, Nova Scotia, Canada, has now found that these algae also live inside the embryo's cells. Such a close coexistence with a photosynthetic organism has previously been found only in invertebrates, such as corals. Kerney took long-exposure fluorescent images of pre-hatchling salamander embryos, and saw scattered dots in the unstained tissue an indicator that it might contain chlorophyll. Transmission electron microscopy (TEM) images showed mitochondria in the salamander cells clustering close to the algae. Reporting the discovery on 28 July at the Ninth International Congress of Vertebrate Morphology in Punta del Este, Kerney suggested that the mitochondria might be taking advantage of both oxygen and carbohydrate generated by the alga's photosynthesis.
So when do the algae enter the embryos' cells? A time-lapse video made by Roger Hangarter at Indiana University in Bloomington, and presented by Kerney at the meeting, reveals a fluorescent green flash an algal bloom next to each embryo just as its nervous system begins to form. Most research on spotted salamander embryos has focused on earlier periods of development, which might explain why algae have not been seen inside the cells before.
One of Kerney's most curious discoveries suggests that the algae may be a maternal gift. He has found the same algae in the oviducts of adult female spotted salamanders, where the embryo-encompassing jelly sacs first form.
David Wake, an emeritus professor at the University of California, Berkeley, who watched Kerney's presentation, wonders whether algae could be getting into the reproductive cells. This would "really challenge the dogma" that vertebrates' immune systems ban such close relationships, he says. Both Wake and David Buckley, who studies salamander development at the National Museum of Natural Sciences in Madrid, agree that the work might tell us more about how vertebrate cells learn to identify intruders.
"It makes me wonder if other species of salamander that have known symbiotic relationships with algae also harbour algae inside their cells," adds Daniel Buchholz, a developmental biologist at the University of Cincinnati in Ohio. "I think that if people start looking we may see many more examples."
I choose to call Spotted Salamander embryos in their green sacs "Gaian symbiosis in miniature" because this image harks back to my very first scientific paper. Titled, "Open Systems Living in a Closed Biosphere: A New Paradox for the Gaia Debate," it was published in 1990 in the journal Biosystems.
To the left you will see the acceptance letter that I will always cherish, from one of the journal's scientific editors: Lynn Margulis, who died in 2011.
LYNN MARGULIS is best known for her success in advocating that some of the major transitions in evolution (notably, the birth of the eukaryotic cell) occurred through an intimate form of symbiosis called symbiogenesis. The scientific community now agrees that several of the components within eukaryotic cells were absorbed directly as bacteria from the surrounding environment, from which a mutual dependency emerged. Chlorophyl plastids within the cells of all photosynthesizing plants had spent hundreds of million of years entirely as free-living, sun-loving bacteria before the merger occurred. As well, the oxygen-consuming powerhouses in all plant and animal cells (the mitochondria) began as free-living bacteria, too.
So why do I call the alga-rich embryo sacs of Spotted Salamanders "Gaian symbiosis in miniature?"
I do so because the crux of my 1990 scientific paper is that it is a remarkable thing indeed that Planet Earth, while open to the sun's energy, must be entirely capable of keeping all of the matter essential for life circulating freely between the oceans, the atmosphere, the soils, the life forms themselves, and the far slower circulation of rocks propelled by plate tectonics. Technically, Earth and its biosphere is a "closed system".
In the very early evolution of life, every time some essential element (notably, nitrogen) got stuck somewhere in the cycle, bacteria would evolve a new form of metabolism to "mine" that stuck resource and get it back into circulation.
SO WHEN I GAZE UPON THE GREEN SPHERES encompassing the developing salamander embryos, it reminds me of the profound achievements of Earth's early bacteria. They kept matter circling and cycling within a developing planetary system: Gaia.
In the case of salamanders, of course, the closure is not absolute. Obviously, the jelly is penetrable to oxygen and carbon-dioxide exchange if salamanders deprived of algae can still develop, albeit more slowly. But I do have my own adaptive theory:
From my observations in the spring of 2011 at Sallie Pond, I noticed a profound difference in the structural integrity of Spotted Salamander jelly blobs versus the jelly masses laid by various species of frogs and toads. As massive spring rains and winds jostled the denizens of the pond, all the jelly masses except those of the Spotted Salamander tended to disintegrate. As well, there was a much shorter time between egg laying and egg hatching for the smaller species, so they didn't need robust jelly for as long.
Hence my hypothesis: Because Spotted Salamander nymphs take so long to develop, a sturdier jelly would have been selected for by evolution. If a jelly mass can hold onto its stick or pond-bottom perch away from the danger of a diminishing and drying shoreline, then as the pond waters swirl, and waters rise and fall with the rains, the jelly mass will not become stranded ashore. The downside, of course, is that the sturdier the jelly, the less porous it is to air transfer. Thus, acquiring symbiotic algae would have given the species an opportunity to fortify its jelly without diminishing its metabolic rate.
Further speculation: Salamander embryos that did not enter into this form of algal symbiosis would have taken longer to develop, given the same robustness of jelly. They would have been "lost to the gene pool" because this trait would be a dangerous disadvantage. Certainly at the Sallie Pond I observed, the danger for slow hatching is that in some years, the ephemeral pools dry up before this large salamander has time to trade its gills for lungs. Stretching out the time for embryo development is thus a bad idea.
Why not simply switch from temporary forest ponds to perennial water bodies?
The answer is safety from predators. Perennial freshwaters of high enough quality to support salamanders would inevitably carry fish. Fish are dangerous predators for new hatchlings. But in an ephemeral forest pond, Spotted Salamander nymphs are the top aquatic predator!
ABOVE: April 1 I unintentionally speeded up the hatching of one Spotted Salamander egg mass that I put in a jar to take to Atlanta with me for a 2-day trip. I was scheduled to deliver an evolutionary children's program at a Unitarian Universalist district church gathering, so I figured I would first introduce the kids to a living form of, what looked to me like, green cat's-eye marbles. Here is my journal entry:
April 1 I collected the smallest Spotted Salamander jelly mass, plus extra pond water, and backpacked it to the cabin. Lo, two hatched by the time I got there, from the jostling. An hour later we left for gigs in Atlanta.After the program, I poured the contents of the bowl back into several jars (as I had added additional plain pond water to the bowl), and my husband and I made the return drive to Lookout Mountain. Upon arrival, Tobey and I immediately walked to the pond and released all the baby salamanders back into the cool waters of home. They had done such fabulous work!
The next morning, after arriving at the church, I emptied the jar into a large transparent plastic bowl, took it outdoors, and placed it on a bench so that the kids could gather around and look. Nobody wanted to touch the water or the jelly to begin with, but then I scooped one of the 3 hatched salamanders into my cupped palm for them to see, then picked up the jelly mass with the rest of the green spheres in it with embryos some were visibly hatching.
Then I left the kids to tend to them on their own, with an adult or two casually around. Girls were the oldest, so dominant, thank goodness. One 5-year old-boy comes into the room with his cupped hand dripping jelly, to show me a hatching salamader.
Adults told me the kids loved having them hatch in their hands. An adult told me that one hatchling got killed by dropping off a hand into the wood chips, but the rest seemed okay.
What a great experience for the kids! I walked them thru figuring out the green algae was on the inside of the spheres, and several times hoped to coax them to understand the mutualism of CO2 and oxygen exchange. I definitely told them that scientists had only discovered this the previous year.
I also told them about my love of nature and tending salamanders as a kid, and that I wrote science books.
"Does anybody here think they might want to be a scientist?" Of the 8 kids total, about 5 raised their hands.
I then pulled off a single-egg-sac chunk of one of the untouched egg masses in the pond and put it in my jar to carry back to the cabin. I wanted to have one unstressed nymph to care for in the remaining weeks before we were scheduled to head north.
ABOVE: April 7 - One week after I jostled and stressed about 20 nymphs into hatching (in my quest to give the church kids a profound nature experience), I carefully examined the untouched jelly masses I could easily wade out to in the pond. A few had hatched, but by far the majority were still developing. Meanwhile . . .
Meanwhile, on April 4 the single "green marble" I had put in a jar of pond water and carried back to the cabin (as calmly as I could) hatched. For the next 20 days I developed a whole new relationship with Sallie Pond: for I had to find the best places and a workable technique to capture denizens just barely visible to my eye: primarily daphnia ("water fleas") and tiny red roundworms.
This is because Spotted Salamader nymphs are total carnivores. In fact, they have been described as the T. rex of ponds. At first I named my hatchling T. rex, but I soon substituted the name Triceratops. It just looked like Triceratops! See the photo above left with the feathery external gills stretching outwards, like the bony fringe plate that protected Triceratops' neck.
In my quest to gather food for my temporary pet, I discovered a whole new world in the pond! And, I was amazed to occasionally see an impossibly tiny polliwog appear overnight in the extra jar of daphnia-and-algae-rich pondwater I kept for restocking Triceratops' home. Apparently, there are amphibian species that lay invisible eggs in the algal masses.
Looking down into the jar or through the side, I watched Triceratops a lot. But only twice did I see this lurking predator jut out in an instant and swallow something that had been passing directly overhead. Nonetheless, the daphnia and roundworms I kept adding to the mix tended to disappear.
On April 22 I wrote this email to a friend:"In 2 days we leave this paradise, so "Triceratops" will be carried back to its pond to join the other hatchlings. If we continue to get lots of rain, Triceratops will likely mature and maybe live another 20 years. If not, it will be salamander-eat-salamander, as I have read. I have no idea if my guy/gal is bigger for the early hatching, or smaller as maybe the size of the jar would limit its growth. Its front legs became visible about 5 days ago."
ABOVE LEFT: April 13 - Spring vegetation is apparent now at Sallie Pond, a full 6 weeks after the salamanders had laid their eggs.
ABOVE RIGHT: April 24 - Tobey watches Connie return "Triceratops" to its natal home.
All the remaining viable egg sacs in the pond had long since hatched. Despite best efforts, Connie could locate no "wild" nymphs in samples of nearshore detritus she searched through. These salamander nymphs are, after all, lurkers not swimmers. And perhaps they found deeper waters more to their liking.
MONTHS LATER: Tobey told Connie that the summer produced good rain; the drought that had swept the southeastern United States for several years was in hiatus. Surely, a good many Spotted Salamanders (like the one at the top of this page, which I found 40 feet from a pond in the Finger Lakes area of New York the previous August) had crawled out of Sallie Pond and found their way to abundant fallen logs and rocks. Deep below these surface features they would live the life of a cold-blooded mole venturing near surface only in the moistest of times.
I'd like to think that Triceratops was among them. If so, my little friend will eventually grow to 8 inches in length and it could, quite possibly, outlive me. This species is known to live as long as 32 years!
Good luck, Triceratops! May you and your descendants continue to make annual pilgrimages to precious forest pools. And may my own species learn the joys of making pilgrimages to Sacred Sites of the Epic of Evolution!
Here is a terrific 6:24 minute video to learn more about Spotted Salamanders, and to get a sense of how, except for seeing the egg masses, it is very unusual to actually see a spotted salamander nymph or adult during the daytime.
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