It is time to get the system up to full capacity. The last component, a half-height 10-gallon tank (yes, that’s really a 5-gallon tank for the mathematically inclined) is now in operation. Originally designed to be a nursery, it seemed better suited to use as a Bryopsis culturing tank. I have been pleased with growth rates under the Evergrow S2 hydropnics light, so I picked up another one from Exotic Reef Creations. So there are now two slug tanks and two algae tanks for the upcoming project. With the red light from the hydroponics lights and the green from the algae, one might be tempted to think it looks a little like Christmas around here.
Here’s a better view of the new algae tank, with eggcrate to acts as a substrate and a small Hydor propeller pump to provide current. Several of the original samples of Bryopsis had started growing in fellow aquarists coral propagation systems, and were provided conveniently mounted on “frag plugs” onto which fragments of coral colonies are normally glued to enable them to grow. In this case, the growth of the algae on the plugs allowed easy and stable placement in the eggcrate.
There have been a few other improvements as well. In November, I got the new Neptune Systems Apex controller hooked up. Although it is capable of much more, it is hooked up to monitor and log energy use, system pH, water temperature, air temperature (in case of failure of environmental controls in the building), and moisture on the floor. The unit controls all of the lights, pumps, chiller, and heater, and can, for example shut down the pumps automatically in case of a spill being detected. Further, the system is completely accessible through a cloud-based interface, which allows control while I am away from the office.
Along with keeping an eye on the system when I am away, the Apex provides a log of some parameters. For example, when all of the lights were on at the same time of day, pH fluctuated significantly. The upper plot in the graph show detail of pH rising from 8.1 in the early morning to 8.5 when the lights went out at the end of the day. Because CO2, which reduces pH through the formation of carbonic acid, is consumed during photosynthesis when the lights are on and builds up at night, it is not surprising that pH steadily rises during the day.
Although it did not seem to cause the plants or animals much stress, it seemed worth trying to reduce the fluctuation. Rather than having all lights on at the same time of day, I set the lights on the algae growth tank to be on during the night. In this way, photosynthesis would be relatively constant in the system over the course of the day, with the potential benefits of stabilizing pH and keeping consumption of nutrients relatively steady. I was pleasantly surprised to see the pH fluctuation reduced to between 8.25 and 8.35. Frankly, the livestock did not seem as excited as I was.
Finally, I added a small protein skimmer (a.k.a., foam fractionator) to remove microalgae and organic material from the water column. Some might argue that suspended material serves as a source of nutrients for the plants, but in my experience desirable algae and plants tend to outcompete nuisance algae when a skimmer is used. None of the the nutrients provided by the dosing system are removed by skimming, so the skimmer just removes the compounds that are not under my control. With the height restrictions of the sump, I went with a Reef Octopus BH50, which is designed for small systems. It has been easy to set up and adjust, and extracts significant amounts of smelly material every day.
For the next few weeks, we’ll be watching algae grow. After the holidays, it will be time to order up a batch of slugs for the next round of Slug Science!
As described in the last post, I was interested in improving the growth of the food plants. Among the many parameters to consider (spectrum and intensity of light, water flow, nutrients), I decided to systematically increase the nutrient levels by carefully dosing Guillard’s f/2 formula. I have been very pleased to see a significant improvement in color, growth rate and branchiness of the Bryopsis.
Previously, growth was unimpressive. The plants were pale and rangy, and I was becoming concerned that they would not ever grow enough to keep the slugs fed. In the photo below, the finer, fuzzier stuff is Derbesia, and the thicker strands are barely recognizable Bryopsis, which should be bushy and feathery. If you can’t figure out what Bryopsis should look like, it may help to read to the end of the post and then scroll back up here.
Within a few days, the Bryopsis started looking better. In the photo below, an astute viewer might be able to see that the tips of the branches are beginning to become a darker green. With the added nutrients, cyanobacteria (the red film coating some of the plants) have also increased. With a little adjustment of the dosing, I hope to see less red and more green.
In just over a week, growth is robust, color is a satisfying, deep green, and additional branches are starting to appear. At this point, I also changed the flow pattern a little to increase the current passing through this patch.
As of a few days ago, the plants are nice and green, and almost feathery. In addition, there are patches of new growth forming in areas of high flow, so the alga is spreading. At this point, NO3 levels are a little over 10 ppm, and PO4 is about 0.5. Over the next few weeks, I am going to try to bring the levels down a little bit to discourage cyanobacteria.
So what have we learned? First, a more balanced approach to feeding the algae results in faster growth and better color. Without systematic removals and subsitutions of components, it’s hard to know which of the ingredients was limiting for growth (N, P, Mg, Mn, Fe…..?), but the mix has quickly done the job. Second, based on the quality of the growth in areas of high flow, Bryopsis seems to like a lot of current. At present, I do not know which species I have (I hope that will change by the end of next semester), but some are found in intertidal zones with significant surf.
With these lessons in mind, I will be converting the nursery tank into a Bryopsis cultivation tank to provide food for the hungry Elysia that will arrive in a month or so for the student research project.
To possibly belabor a point, the limiting factor in slug husbandry seems to be food. So more, better food = more, better slugs?
Given how hard most aquarists work to eradicate nuisance algae, including the Bryopsis that Elysia clarki find irresistable, it would seem that growing algae should not be all that horribly challenging, right? One just needs good, strong light, plus some nutrients to support growth. The trick is to grow the species one wants, without having the cultures overtaken by undesirables. In my case, the “good” algae are Penicillus, Bryopsis, and, to a lesser extent, Derbesia, Halimeda, and Avrainvillea. The primary nuisance I am trying to avoid is “red slime” cyanobacteria, which can overtake and smother the other species.
Plants require nutrients such as carbon, nitrogen, potassium, and phosphorous in significant quantities, along with smaller amounts of calcium, magnesium and sulfur . Based on my measurements, the desirable algae consumed considerable amounts of nitrate, but adding phosphate did not appear to encourage growth target algae and increased the growth of cyanobacteria. So, along with calcium (Penicillus and Halimeda use it for structural support) and bicarbonate (a pH buffer and source of carbon) I have been using the dosing system to add potassium nitrate in order to maintain nitrate at about 5-10 ppm. For everthing else, I have been relying on the artificial seawater mix to provide an adequate balance. Perhaps not the most systematic approach to feeding the plants.
It has, however, worked reasonably well. In the photos above, there is a dense growth of Penicillus on the right side of the Broodstock tank, and a mat of Derbesia and Bryopsis on the left. The resident E. clarki seem quite happy to spend their days grazing without exhausting the supply. The other tanks are similarly productive. Nonetheless, it might be possible to increase algae productivity, thereby increasing slug capacity and the frequency of mating and egg laying.
There is, of course, an actual literature on the nutrient requirements of aquatic plants. Some commonly used formulas rely on extraction of nutrients from soil to provide the correct balance of elements. I might have tried that had I been able to buy the correct brand of English planting soil. However, based on a number of studies (Kumar et al., 2011 e.g.) Guillard’s f/2 medium (Guillard, 1975) yields excellent growth of macroalgae and does not require purchasing and soaking British soil.
Florida Aqua Farms supplies Guillard’s f/2 medium under the name “Plant Fuel,” providing pre-measured dry ingredients along with a suggested dosing regimen. Seemed worth a try to find out whether more complete supplementation will translate into faster growth of desirable algae. What’s the worst that can happen, aside from massive bloom of toxic, undesirable algae?
The components arrived, I mixed them up, and the dosing system is adding 1 ml of f/2 per 2.5 liters of system volume, distributed over the course of each week. We’ll see…
The project has had the feel of watching grass grow lately, mostly because I have been spending a lot of time collecting and establishing potential food algae for the slugs, and then watching them grow. Nonetheless, there is beginning to be some motion.
For example, starting Spring semester (late January), I will have a couple of students starting some simple molecular experiments. Although the diet of Elysia clarki is well-characterized, that of the species we find in Baja California, E. diomedea, is still not known for certain. On the next trip to Bahia de los Angeles, I plan to collect some E. diomedea, and identify their food plants based on the DNA of the chloroplasts that they have stolen from their food. This kind of work is straightforward in a comfortable lab where one has access to liquid nitrogen and other luxuries. However, we will need to develop protocols that will work in the heat and limited resources of the field station in Mexico. So, we will use the semester to develop a protocol for extracting and purifying DNA that utilizes the simplest methods possible. Meantime, it will be necessary to put together reading lists on the biology of Elysia and methods of DNA extraction so that we can hit the ground running.
The animal care system is also evolving. As I described a while ago, I have been lucky enough to get hefty samples of a few varieties of hair algae such as Bryopsis and Derbesia from local aquarists. The trick has been to get enough growth to maintain a self-sustaining supply of food. Although filling a tank with algae, sticking it into a window, and dumping hefty amounts of ammonia and phosphate into it produced decent results, it was not very stable.
Instead, I have added a 20 gallon tank devoted to growing algae to the slug culture system. The fancy hydroponics light gives a great spectrum for growth, but the strong red-pink quality of the light is not the most pleasing. There are a couple of species of hair algae, plus a tub of marine sand for macroalgae that need a substrate. To take advantage of the automated dosing of NO3, Ca and HCO3, it is plumbed into the rest of the system. Dosing of NO3 has already been increased a few times to keep up with the growth of the algae.
In other news, the controller that currently controls the lights, temperature and pH will soon be replaced by a newer, cloud-based model. This will allow remote monitoring of temperature, pH, and power use, as well as sensing moisture under the tanks in case of leaks. It can send email alerts in case of equipment failure, and should help prevent loss of animals or damage to property.
The eggs hatched last Monday (10/5/15), but I am finally getting around to posting. Assuming you have a little imagination, you should be able to see some little veligers whizzing around among the very wiggly embryos. The hatchery was not ready for them (they only took about 5 days to hatch), so I just added the veligers directly to the growout tank and hoped for the best. Whatever this species is, it could be very useful in the lab to have embryonic development done in less than a week.
Meantime, I have added some fresh Bryopsis from a local reef tank, so maybe the E. clarki will be in the mood to lay eggs soo.
Two days later, the embryos have started to spin in their eggs. The video below has been filtered somewhat to reduce noise, so they appear to be moving a little more slowly than in the raw data. Have a look in full screen mode for the complete Elysia-in-motion experience.
A little over a month down the road, the little slugs that rode in with the last batch of macroalgae are still pumping out eggs. At some point, I will really need to figure out the actual species, rather than referring to them (probably erroneously) as E. papillosa. Always more to do than time to do it.
Here is the young lady laying eggs this morning. No scale, but she is probably about 4 cm at this point. Note that the egg mass is different from the tidy coil seen in a previous clutch, seen here.
If past experience holds true, the eggs will hatch within about 5 days, which is a lot faster than the larger E. clarki. In order to compare the developmental sequences of the two species, I have started taking photos of the little guys’ development.
Below is a shot at about 20X, showing a couple of strands.
At higher magnification.
Given their rapid development, it will be interesting to see what they look like tomorrow. For a sense of scale, try to compare the eggs in the micrographs to the barely resolvable dots in the egg cluster as the eggs are being deposited.
Although they will eat a number of algal species, Elysia clarki strongly prefer Bryopsis, which is a feathery filamentous alga considered by most aquarists to be a pestilent nuisance. For the past year, I have been benefiting from the struggles of a local reefkeeper who has had a plague of the stuff in his 500 gallon system. Alas, through rigorous reduction of phosphate levels, his battle with Bryopsis seems to be coming to an end.
That sent me into a panic. Well, more like mild anxiety. Even though it has been possible to maintain my own stock of Bryopsis, it had started to run low. Hard to believe, but I am having trouble growing enough of a nuisance alga. In my defense, it is a bit of a trick to grow one form of plague without another taking over. I was starting to be concerned that the slugs would not be ready to lay eggs if they ran out of their favorite food and had to make do with other algae.
Fortunately (for me), another WAMAS member is in the midst of a battle with Bryopsis. It required a trip to Virginia, but I returned with a bucket full of soft, feathery algae. It seems finer than the variety I was getting from the previous source.
Nonetheless, the slugs are lapping the stuff up. Despite their reputation for slow movement, slugs are pretty good at moving around when they have appropriate motivation.
The adult above dug right in. As did the juvenile, below. Up to her rhinophores in Bryopsis.
Below is a shot of her cruising around, before having found the food pile.
Now that they have been fattening up on algae, they have taken on a much deeper green color. Will have to post some before and after photos next.
It has been quite some time since I have posted anything about Box of Slugs 2, the tank at home that is connected to a coral reef aquarium. There has been a lot of excitement, such as the appearance of several species of Sacoglossans that had ridden in with live plants. However, without large grazers such as urchins or snails, it has also been difficult to control the growth of algae, such that much of the tank was quite furry and subject to blooms of cyanobacteria (blue green algae, a.k.a. red slime).
The mass of hair algae may have had some benefits as food for growing Elysia, but it also created dead spots with poor circulation, and overwhelmed some of the desirable species of macroalgae.
After a few tweaks, and a little patience, the mass of filamentous algae has died back, and the macroalgae and seagrasses are thriving. I got rid of the large rock that impeded flow, started dosing vinegar as a carbon source for the plants, and assembled a filter to remove debris and increase circulation.
The tank is looking pretty good at this point. There is a variety of macroalgae (Penicillus, Udotea, Avrainvillea, Halimeda), along with some Bryopsis and turtle grass. A few gorgonians add a little variety. All the plant species seem to be growing well, and there is little sign of the plague algae coming back.
Probably the most important change was the addition of this filter to increase circulation and remove some of the debris. It consists of a sponge sleeve over a perforated 1″ PVC pipe, connected to an Eheim 1048 pump.
The pump is stuck to the wall of the tank with a magnet. The arrangement allows the pump to be partway out of the water to minimize heat transfer. The output goes through a Hydor Flo rotating deflector, which provides circulation throughout the tank.
The resident slugs are three E. crispata. I have decided to separate the larger Elysia species, with clarki in the hatchery at USG and crispata in BoS2. Each system also has a few smaller hitchhiking species. BoS2 has a few of the frilly presumed Cyerce, while BoS1 has some presumed E. papillosa.
The E. crispata seem quite happy, but are largely ignoring all of the macroalgae species in favor of a few small patches of Bryopsis.