Because I was so surprised at how quickly my sand dollar larvae (Dendraster excentricus) were developing, I checked my notebook from the invertebrate embryology course I took while in grad school to see if what I’m observing now is normal for these animals. It turns out that yes, Dendraster does develop at a much quicker rate than its cousin the sea urchin. And now that I think of it, when I took that 5-week embryology course the sand dollars were the only echinoids that we followed all the way to competence; we spawned and observed urchins as well, but none of them were as far along as the sand dollars by the time the class ended and we “graduated” our larvae off the dock.
Yesterday my Dendraster larvae were five days old. They already had two well-developed pairs of arms and were working on the third pair.
These larvae are big, too–500 µm long. Of course, they started from eggs that were over twice the size of urchin eggs, but they’ve still grown a lot in only five days. The fourth pair of arms will be the preoral arms. At the rate these larvae are developing, I wouldn’t be surprised if these arms show up in the next few days.
As beautiful as those long arms are, they may be a little too long. The larvae swim and gather food using a band of cilia that runs up and down all the arms; the entire body is ciliated, but the ciliated band is the primary locomotory system. I remember the instructor of my embryology course telling us that echinoid plutei will respond to lack of food by growing longer arms, which increases the length of the ciliated band and thus (presumably) the animal’s ability to capture the food that is available. There are two pieces of circumstantial evidence that my larvae may be a little food-deprived: (1) the really long arms; and (2) the lack of visible food cells in the stomachs. In urchin plutei that are feeding well I can see food cells churning away in the stomachs. These Dendraster plutei have beautifully transparent bodies, but I don’t see food in the guts. On they other hand, they are growing, so obviously they are eating. Just in case they are short of food, though, I’ll increase their food ration for the next few days and see how the animals respond.
In the meantime, I continue to be fascinated by the intricacy of the larval skeleton and the complexity of the skeletal rods themselves. Next time I’ll try to take photos of these.
My sand dollar larvae are developing very quickly! When I checked on them Thursday afternoon about 24 hours post-fertilization, I anticipated seeing them up in the water column because that’s how long it takes urchins to hatch. Remember, sea urchins and sand dollars are in the same taxonomic class (Echinoidea) and share a larval form called the echinopluteus. I’ve watched urchin development often enough that I have a sort of intuitive feel for how it goes, and am subconsciously comparing these sand dollars to the urchins’ time table. I need to stop doing that.
Anyway, on Thursday the sand dollars had indeed hatched. The big surprise was that when I examined them under the microscope I saw that they were much further along than urchin embryos would be at the same age. I expected to see the embryos swimming around as blastulae (hollow balls of ciliated cells); however, these sand dollar embryos had almost completed the process of gastrulation to form their archenteron, or first gut.
In echinoids the archenteron develops from an invagination into the blastula. Imagine a balloon. Now imagine poking your finger into the balloon–you’ve just made an invagination. If you continue the invagination across the entire balloon until your fingertip pops out on the other side, you’ve created a tube that penetrates through the balloon. This tube is the archenteron. Interesting tangent: You know that any one-way gut has two openings, right? One is the mouth and the other is the anus. The point in the blastula where gastrulation begins is called the blastopore. The fate of the blastopore is to be either the mouth or the anus of the archenteron. Echinoderms are deuterostomes (“second mouth”), a term that means the blastopore becomes the anus and the mouth is the second opening that forms when the invagination punches through to the other side of the blastula. Yes, sea urchins and sea stars and sand dollars all have an anus before they have a mouth. And guess what? So did you. The chordates, including us, are also deuterostomes.
These sand dollar embryos went from zygote almost to feeding larva in only 24 hours. In fact, some of them may have had mouths when I looked at them on Thursday. I had to start feeding them, so that food would be available as soon as they were able to eat.
Another 24 hours later, on Friday afternoon, I checked on the larvae again and they were bona fide plutei already. They had the cup-shaped body body of the pluteus larva and two pairs of arms, with complete guts. The stomachs in these larvae are huge, occupying almost the entire volume of the main body of the animal. Some of the larvae are also developing red pigment spots.
And how could I have forgotten that the plutei of Dendraster have fenestrated arm rods??? They are so beautiful! This is the same animal as in the photo above, but I focused in on the skeletal rods in the postoral arms. See the fenestrations in the rods? The larva is about 300 µm long.
For whatever reason, plutei of the local species of sea urchins don’t have fenestrated arm rods. This difference in larval morphology between the two most common echinoid species in the area should make it easy to identify plutei collected in plankton tows. We’re at the beginning of the spring bloom now, and I hope to keep an eye on how the plankton community develops through the spring and summer.
The next day I examine the larvae is Monday. I’ll see if they’re still on the fast track to metamorphosis.
This afternoon I met up with Joanna and Amy, who had come to the marine lab with some sand dollars (Dendraster excentricus) to try to spawn. Since sand dollars are in the same taxonomic group (the Echinoidea) as sea urchins, I’d try the same techniques on these animals I’d never spawned before. I did have to modify some things a bit, mostly to account for the difference in body shape between sand dollars and urchins. Urchins are globular, with quite a large internal body volume, while sand dollars are flat. There’s much less space inside a sand dollar for gonads and guts.
Gravid echinoids such as urchins and sand dollars can be pretty easily induced to spawn by injecting their internal body cavity with a solution of KCl. We shot up all eight sand dollars and five of them spawned, two males and three females. One of the males didn’t give enough sperm to be collected, so we didn’t use his gametes. The other male, though, gave us lots of sperm. And they were good sperm, too.
If you’ve never had a chance to see swimming sperm under a microscope, today is your lucky day!
And the eggs. Wow, sand dollar eggs are freakin’ cool! For one thing, they’re big, ~130 µm in diameter, compared to the 80 µm eggs of the purple urchin Strongylocentrotus purpuratus. Plus, they have a really thick jelly coat that contains red pigment cells; urchin eggs don’t have the pigment cells, either.
The eggs themselves were a little lumpy, not as perfectly round as I’m used to seeing with the urchins, but they fertilized just fine. In all three of the crosses, the fertilization rate was 90-95%. Apparently the sperm have no problem digging through the jelly coat to get to the egg surface.
In this photo you can see the familiar fertilization envelope raised off the surface of the egg, as well as the red pigment cells in the jelly coat. This may very well be the most beautiful zygote I’ve ever seen. How many people can say things like that?
After an hour and 20 minutes sitting on my desk at room temperature the zygotes started to cleave:
The blastomeres are still a little wrinkled and lumpy, but I think they’ll be okay. I’ve poured them into 1000-mL beakers and they’re sitting in one of my seawater tables. Tomorrow afternoon I hope to see them swimming up in the water column. Fingers crossed!
So. I have a batch of larvae from a spontaneous spawning of the leather star, Dermasterias imbricata, that occurred four weeks ago tonight. Until now I’ve never had an opportunity to work with this species, even though we have quite a few of them at the marine lab. I had my own for several years, until they became casualties of the plague about a year into the current sea star wasting syndrome event. In any case, this is the first time I’ve been able to spend time with larvae of this species. At the very least I wanted to see how big they would get and how quickly they would develop, compared to the species I’m more familiar with, Patiria miniata (bat star) and Pisaster ochraceus (ochre star).
When the Dermasterias spawned, the first thing I noticed was that the eggs are huge. I measured them at 220 µm in diameter, which is big even compared to what I’ve seen in other stars. Hatch rates were pretty good, and four days later the larvae were already in the 400-430 µm range. Since I have no experience culturing this species, I thought I’d divvy up my larvae and put them into three feeding treatments to see which larval diet resulted in the best overall success. According to the literature, Dermasterias larvae can be raised on a mixture of the unicellular algae Dunaliella tertiolecta (green) and Isochrysis galbana (golden). My three feeding treatments are: Dun only, a Dun/Iso mix, and Iso only.
A week into the experiment there was a clear difference between the larvae eating only the green food, and those eating either a mixture of green and golden or only the golden. Larvae from all food treatments were about the same size, but the ones eating only Dunaliella had noticeably green guts.
Fast forward two weeks, and the larvae were 20 days old. By this time they had progressed from the bipinnaria stage to the brachiolaria stage. The interesting thing was the absence of green pigment in any of the guts, even those that were eating only green food. The D. tertiolecta larvae looked good, actually. They were a little smaller than the other larvae but were perfectly formed.
Obviously all of the larvae are assimilating enough of their food to grow and develop normally. I looked at them today but didn’t have time to take pictures. Qualitatively there is no difference between the Dun larvae and the Dun/Iso larvae. In the Iso jars, however, there are many larvae at earlier stages; some are still at the “jellybean” stage. I don’t know if this is because these larvae are developing more slowly, or because of some nonrandom distribution of earlier stages into those jars when I was setting up the feeding treatments.
Next week I’ll measure the larvae again, and will have three data points to track growth trajectories.
After pretty much neglecting us in February, El Niño has returned with a bang in March. Late yesterday and last night a weather station near me, more or less at sea level, recorded 4.67 inches of rain and wind speeds of 15 mph. Stations in the Santa Cruz mountains recorded close to 6 inches of rain yesterday, and there were patchy power outages throughout the county. This morning I woke to sunny, clear skies. Beautifully clear, with white puffy clouds. The forecast calls for another storm to head in this evening, giving me a window of opportunity to run up the coast and grab some mussels.
I have to say, El Niño’s timing could be better. We have alternating weeks of spring and neap tides, and this winter the storms seem to be arriving during the spring tides. More than one tide series has been washed out because of storm surge and majorly big swell. I had figured that this would be the case today, so I didn’t expect to get very far down in the intertidal. However the only thing I absolutely had to collect was mussels and I don’t need a very low tide for those. It was very unlikely that I’d be unable to collect them, and at the very least I’d be able to take some photos.
Walking across the beach to the rocks, I noticed my first Velella of the season. As usual for these strangely wonderful animals they were gathered into windrows at the high tide level. Many of them were very small, less than 1 cm long, and the largest I saw was about the length of my thumb.
While it is not at all unusual to find Velella washed up on the beach, I did find some in a place that I didn’t expect. More on that in a bit.
Conditions in and on the water were pretty rough. There were no surfable waves, therefore no surfers. They’d have been beat up by the waves crashing in all directions.
On a calmer day, the water at this beach can be glassy smooth with very gently breaking waves. Not so today:
Easily accessible beaches such as this one are typically crowded for these afternoon low tides. Most of the people there are just hanging out with their friends, family, and dogs. Every once in a while, however, I run into people who might not be entirely on the up and up. Much of the coast in California is designated as a marine protected area (MPA), and while allowed activities vary from MPA to MPA, in general I don’t have permission to collect at any of them with my current state-issued scientific collecting permit. This means that collecting, both scientific and recreational, is concentrated into the few places where it is allowed.
Today I arrived at the parking area at the same time as a family of three adults and about five kids. The men were wearing wellies and carrying 5-gallon buckets. It was clear that they were going to be collecting something. I can’t really say that I looked any different, in my hip boots and with my own bucket, so I just smiled a greeting to them and headed out on my way. Given that there was so little exposed rock, we were bound to keep running onto each other. At one such meeting I asked what they were doing, and they said they were collecting mussels to eat. I said I was, too, to use as food for animals at the marine lab. They asked what the limit was. I told them that I didn’t know what the limit was for taking with a marine fishing license (assuming that they had one), but the limit for my collecting permit is 35. We nodded and went our separate ways.
Now, I’m not a game warden and it’s not my job to enforce the state’s rules about collecting, or even to see if other citizens have the appropriate permits or licenses. I generally feel that the better part of valor is to mind my own business. These guys today were friendly enough and completely non-threatening, but my gut instinct tells me that they didn’t have a fishing license. Is that any of my business? I don’t think so; yet as a citizen of this state I have a vested interest in protecting our wildlife from unlawful take. I know there aren’t enough wardens to patrol all beaches all the time, and now that I think about it I don’t know that I’ve ever been stopped by a warden on an afternoon low tide. The enforcement strategy seems to be to let citizens patrol each other, in the sense that skullduggery is less likely on a crowded beach in the broad daylight of afternoon than at the crack of dawn on a morning low tide.
Anyway, on to the matter at hand. I’ve noticed that recently my eye has been drawn to patterns that occur among whatever objects happen to be around. Scrambling down a little cliff and continuing up the coast I noticed these smears of algae growing on the vertical sandstone face. It’s not that I hadn’t seen them before, but because of the recent rain there was water running down the cliff face, which added a sheen to the green algae that they don’t have when they’re dry.
At this site there are some little caves that you can get to at low tide. The tide wasn’t low enough to reach the caves that go back any appreciable distance, but I did get to a small one. It was more of deep fissure than a cave, really, large enough to duck into but only a couple of meters deep. The really cool thing about it was the waterfall cascading over the opening. Again, without the runoff from yesterday’s rain this little waterfall wouldn’t even exist.
Also, there is quite a bit of stuff living inside the cavelet. Not much in the way of algae, of course, with the exception of both encrusting and upright corallines, but in terms of animals there was more or less the same fauna that I’d expect in the high-mid intertidal.
The biggest surprise in this little cave was Velella! A bunch of them had apparently gotten washed up into the fissure by the last high tide. I found them stuck amongst barnacles and algae.
I can’t imagine there’s much nutrition in a Velella for a crab, but the animal is always right even (especially?) when it doesn’t make sense to us. The crab knows what it’s doing.
All told, it was a short but very satisfying little jaunt to the intertidal. The clouds had spent the afternoon talking about whether or not to build to anything, and by the time I left they’d come to consensus. The wind is picking up now, the rain should start soon, and the National Weather Service says we may be in for thunderstorms tonight. I’m tucked up at home, warm and dry. Have a good evening, everybody!
When serendipity strikes, I try to go with the flow and ride it as long as I can. The latest wave is my batch of Dermasterias larvae, which are developing nicely for the first four days of life. And now they look just like jellybeans!
They have complete guts now and have already grown a bit, measuring 400-430 µm long. It’s not always easy to catch these guys in the right orientation to take a photo, as they are spinning and swimming through three-dimensional space, but I got lucky:
Actually, it was a fortunately placed phone call from an aquarium curator that struck the other night. I was at home, having eaten dinner and reviewed my lecture for the following morning, when my phone rang. It was the curator, saying that he was making his last rounds of the evening and had noticed that some of his sea stars were spawning. Echinoderm sex–more specifically, the opportunity to collect gametes and observe larval development–always grabs my attention, so I told him I’d throw on some shoes and meet him at the marine lab in five minutes.
Lo and behold, there were leather stars (Dermasterias imbricata) spawning in several of the tanks and seawater tables. Many of the tables were cloudy with sperm, but I found only one female, which seems strange but isn’t so unusual. These spawning events occur in response to some environmental cue, such as day-length, a chemical of some sort, or the phase of the moon. When a sea star (or sea urchin) spawns it also releases chemicals that trigger spawning in nearby conspecifics, as to spawn by oneself is an enormous waste of energy. A single spawning animal can result in all the others of its kind spewing out huge numbers of gametes in an orgy of passive sex. However, an animal can be induced to spawn only if its gonads are ripe. Ripeness depends on the overall health of the animal and requires adequate food; animals that don’t receive enough food don’t have energy to allocate towards gamete production. As eggs are energetically expensive to produce, compared to sperm, it is not unusual for males of a species to mature earlier in the reproductive season than the females. In Washington the spawning season for D. imbricata is April-August. Here in California the reproductive season hasn’t been clearly defined, but I do remember a springtime spontaneous spawning event in the lab several years ago.
That creamy looking mass of goo on the star’s aboral surface is a pile of eggs. Sea star eggs are fairly large, compared to the urchin eggs I’m used to, and sticky. They tend to clump together in stringy globs until they are dispersed by water currents. The star whose arm is photobombing in the lower right corner is a male. He was also spawning copiously and is probably the individual who fertilized most of this female’s eggs.
Given the lateness of the hour and the fact that I had to get up early the next morning I didn’t take many pictures of the eggs, although I did look at them to make sure they were fertilized. They were, so I put them into a 1000-mL beaker of seawater and let them do their thing.
Fast forward to today, about a day and a half after fertilization. About two-thirds of the embryos had hatched and were swimming in the water column. Here’s what they look like under the dissecting scope:
I poured off the swimmers into jars and set them up on the paddle table. I gave them a little bit of food, in case their mouths break through before I can get back to the lab tomorrow afternoon. In the meantime, I took a sample of embryos and examined them under the microscope. They look really cool!
The embryos are almost spherical, measuring 290 µm long and 270 µm wide. They are ciliated all over and swim with the rounded end forward. The flattened end is where the process of gastrulation started. That visible invagination begins at a section of the embryo called the blastopore; the channel is the archenteron, the first gut of the larva. In echinoderms, as in chordates (including us humans), the blastopore will end up being the larva’s anus; the mouth breaks through later at the other end of the archenteron. This is why I don’t need to start feeding the larvae right away even though their gut has begun forming.
Tomorrow afternoon I’ll have a brief window of time when I can check on the larvae and see how they’re doing. I think they may have complete guts by then!
While much of America was glued to the television watching a football game, I went out to the intertidal at Davenport Landing to do some collecting and escape from Super Bowl mania. The Seymour Center and I have a standing agreement that some animals–small hermit crabs and certain turban snails, for example–are always welcome, which gave me an excuse to look for them. I also needed to pick up some algae for labs that I’m teaching later this week, so it was an easy decision to be alone in nature for a couple of hours.
As usual, I was easily distracted by the animals, especially the anemones. They are simply the most photogenic animals in the rocky intertidal. And we have an abundance of beautiful anemones in our region; I feel very lucky to photograph them where they live. I would like to share them with you.
Along the central California coast we have four species of anemones in the genus Anthopleura. Two of them, A. xanthogrammica and A. sola, are large and solitary; in other words, they do not clone. The geographic ranges of these two species overlap in central California. Anthopleura xanthogrammica has a more northern distribution, from Alaska down to southern California, while A. sola typically lives from central California into Mexico.
I’ve seen these congeneric anemones living side-by-side in tidepools at Natural Bridges and at Davenport. Here is a photograph from yesterday. The animals are almost exactly the same size, and are separated by about 30 cm. Can you tell which is which?
The pièce de résistance yesterday was a treasure trove of Anthopleura artemisia anemones. I’d seen and photographed them several times before, and always appreciated the variety of colors they come in. For some reason, though, yesterday they really caught my eye. I had a number of “Wow!” moments.
These stark white tentacles are new to me. The anemone measured about 4 cm across. In every other aspect it looks like A. artemisia, and I’m almost entirely certain that’s what it is. It does feel special to me. I will hopefully be able to keep an eye on this individual and see if its colorless tentacles are a temporary or long-term condition. And now that my eye has been primed to see the colors that A. artemisia comes in, I may notice more unusual color morphs.
My most recent batch of sea urchin larvae continues to do well, having gotten through the dreaded Day 24. I haven’t written about them lately because they’re not doing very differently from the group that I followed last winter/spring. However, I’ve been taking photos of the larvae twice a week and it seems a shame to let them go to waste, so I’ve put together a progression of larval development. As a reminder, the last time I wrote about these larvae they were six days old.
Age 9 days: The larvae had four arms and were growing their skeletal arm rods. Their stomachs, which we keep an eye on because their size can tell us whether or not we’re feeding them enough, were a bit small but not so much so that I worried.
Age 12 days: The larvae were growing their third pair of arms. Some had just begun growing the fourth pair of arms. Red pigment spots also start appearing all over the body. Some larvae develop lots of red spots, others have very few. Notice that the stomach is slightly pear-shaped; this is normal.
Age 17 days: This larva doesn’t look appreciably different from the previous one. This photograph, though, is a bit clearer. The stomach has taken on a pink tinge, due to the red color of the food the animal is eating, and the mouth is the large rounded triangular in the in-focus plane. The pair of skeletal arm rods that are in focus are protruding from the ends of the arms, which raises is something to be concerned about. Sometimes the first sign of imminent doom is the shriveling of the arms, so seeing the rods sticking out makes me think “Uh-oh. . .”
Age 24 days: This is about the time in larval development when things often start to go wonky. I’ve looked back at my notes from previous spawnings of S. purpuratus, and seven of the 20 cultures that crashed did so in the week between days 20-28 of development. Some of these cultures were doing well right up to the point that they all died. They were literally there one day and gone the next.
Nonetheless, the current batch of larvae continued to do well. The fourth pair of arms were slow to grow but otherwise the larvae look fine. The top larva in the picture below is lying on its back, so you are looking onto the ventral surface. On the left side of the stomach there’s a little upward-facing invagination; this is part of the initial water vascular system forming. Note also that the overall shape of the larvae is changing a bit. They are becoming less pointy and a bit rounder.
Age 30 days: At this stage the juvenile rudiment is clearly visible. You can see it as a rather nondescript blob of stuff to the left of the gut. The fourth pair of arms have also grown quite a bit but are still considerably shorter than the others. This individual has two bands of cilia, called epaulettes, that encircle the body. These epaulettes will become more conspicuous as the larva approaches competency.
Age 33 days: Today I got lucky! The larvae looked good when I changed their water this morning <knock on wood> and although I’m keeping my fingers crossed I have high hopes for these guys. They’re about as big as they’re going to get, measuring 760-800 µm in length. They will get heavier and more opaque as the juvenile rudiment continues to develop.
The really cool thing is that one of the larvae landed on the slide exactly as I wanted it to. It happened to fall onto its left side and stayed there, so I was able to focus up and down through the body to get the rudiment into focus.
Do you see five small roundish blobs that are evenly spaced around the larger golden circular blob? The large blob is the stomach, seen in side view. Those smaller blobs are tube feet! Don’t believe me? Then take a look at this close-up:
Now if those don’t look like tube feet, then I’ll eat my hat. What’s also noteworthy about this larva is that its epaulette bands are both visible, especially the posterior-most one.
So far, so good. I won’t know how successful larval development is for these guys until they either make it through metamorphosis, or not. In a very real sense, I won’t be able to draw any conclusions about the success of larval development until they either become established as juvenile urchins, or not. One of my graduate advisors inherited a couple of sayings that he passed on to me, as well as to a whole generation of aspiring invertebrate zoologists:
The animal is always right.
and
The life cycle is the organism.
The first is a given, right? The animal knows what it is and what it’s doing, even if we humans have no clue about what’s going on and can’t decide what its name should be.
The second saying might be a little less intuitive. What it means is that, for organisms with a multi-stage life cycle, you have to consider all of the stages if you want to understand them. This is a much more holistic view of biology, and it’s the one that appeals most strongly to me. When I’m thinking as a naturalist, I find my thought process constantly switching between “forest” and “trees” as I seek to understand even a teensy bit of the world around me. While it’s easy to get distracted by all the cool details of organisms, it’s important to step back and ask myself, “What does it all mean? What is the big picture here?” So yeah. Perhaps when (if!) these larvae turn into urchins and I’ve got them feeding on macroalgae in a few months, I’ll be able to say whether or not larval development was successful. If all goes well this larval phase, as all-consuming and fascinating as it is to me, will be only a small part of these animals’ lives.
About two and a half months ago, the ongoing disaster of sea star wasting syndrome raised its ugly head again when one of my bat stars (Patiria miniata) developed lesions on its aboral surface. Here’s what it looked like then:
See how the lesion is sort of fluffy? It looks as though tissue may be sloughing off the surface. Wanting to see how the syndrome would progress, I let it remain in its table and kept an eye on it. Every so often I took it out and examined it, and nothing really seemed to change. The animal continued to eat, retained its internal turgor pressure, and none of its table mates became sick. Eventually I sort of forgot about it.
Until two of my students last week asked if I had any pictures of sick sea stars that they could borrow for their end-of-the-semester project. This question jump-started my brain and I remembered this particular bat star, and told the students they could come to the lab and take their own pictures of it. . . that is, if it were still alive. They were able to visit me this past Monday and together we looked at the animal.
Lo and behold! it’s not dead, and actually looks pretty good.
The star has a few pale areas in addition to the original lesions, but overall doesn’t seem sick at all. It’s nice and firm, righted itself quickly when we placed it in the bowl with its oral surface up, and crawled around very actively.
Not only that, but take a closer look at the lesion itself:
The lesion appears to be somewhat sealed off, as if the epidermis has recovered. I gently poked the surface of the lesion with my forceps, and it feels a little firm and nothing squirted out of or peeled off the surface of it. I think it’s analogous to a scab that forms over a skinned knee. Of course, while a scrape on my knee would heal after a few days, sea stars have a much slower metabolism so I’m not really surprised that it would take over two months for this individual to show signs of a healing lesion.
Of course, I could be entirely wrong about what’s going on with this lesion. It’s the same size as it was back in September, so I’m not convinced that it’s healing. However, it seems that closure of the wound is better than a wide-open gaping sore that leaves the animal’s innards exposed to the external environment. If, over the next several weeks the edges of the wound begin to come together, then I’ll be more confident that this animal is on the road to recovery. In this season on Thanksgiving, this is something to be grateful for.
