Yellow banded possum wrasse breeding attempts

[DaJMasta]

More an announcement of an attempt, but I got my Parvocalanus an Tisochrysis lutea on Friday of this last week and both appear to have taken to my culturing setup well. The density seems good for early growing and the culture volume is almost half of what a full jug would be, so the plan so far is to try to collect eggs on Wednesday for the next attempt. If the culture isn't there yet or I just can't get them that day, I think Saturday and Sunday would both be viable for collection attempts.

Looking forward to getting them going again, but I've got a decent number of fire cleaner shrimplets and a handful of both sexy shrimp and mandarin dragonets started in the in-between.

Also got some 80 micron mesh in the mail, so I've got some gluing to do.

 

[DaJMasta]

Well some odd developments, but I've got an attempt. I tried to collect eggs on Wednesday night, getting started a little late, and from looking at them it looked like they had spawned earlier and wouldn't be, but after 20 mins of waiting, they came out again and seemed to be courting, so I set up the collector around 8:30 pm. I then fell asleep accidentally for half an hour or so, and by a little after 9 they were hiding again. While I didn't catch the spawn, I did find more than a hundred eggs in the collector later in the evening, so they probably spawned as late as I have ever seen 8:50 pm or later.

I moved them to the other system in a basket for them to hatch and checked the next day. Trying to get the timing narrowed down further, I checked early at like 1:30 pm, then 2, 3, 4, 5.... and they still hadn't hatched. I left home for a rehearsal around 6pm without seeing any spawn out thinking that it was my first spawn that wasn't fertilized. I came home around 10:30 pm to a bunch hatched! So there is a run going now, and while the spawning was late (maybe half an hour later) and the air temperature is 1.5C lower or so (not really water temperature...), they took at least 4 extra hours to hatch than I had come to expect. No idea why.

On the food side, I was growing out the parvocalanus culture and at around half volume and several days of promising growth in... I discovered brine shrimp in it! There were maybe 20 or so young adults, no nauplii obvious, and I have no idea how they got in there. I do raise brine shrimp, but on the other side of the room, not really sharing equipment and not tending to them before I set the culture up initially. I ran the whole culture through a 250 micron sieve, and while it definitely caught some parvocalanus adults, it got all but one of the brine shrimp (one I got with a baster after discovering it later), and in the following week I haven't seen them again.

The culture is not quite as dense as I would like, but they are multiplying and there are no signs of issues. My evening tomorrow is sort of packed, so I think I'm going to harvest them and do a first feeding in the afternoon, which given their hatch time may be before they are fully developed. The goal is to feed 45-100 micron parvocalanus and see what sort of density I can get (probably better than all four other strains at that size). I want to check for eating as well, but I don't know if I'll be able to before Saturday morning, but I'll check density of the culture water (sort of passively), feeding response of the larvae to what's in there, and whether I can spot anything in their gut. I should also be able to collect eggs on Saturday and Sunday for further attempts.

 

[DaJMasta]

Late this morning I got one of them under the microscope, and the result is still inconclusive, but I got the camera going at a decent framerate, so I've recorded some footage.

To my eyes, it's showing some hunting behavior, the gut is there, but the yolk sack and air bubble still seem physically connected to the gut, and I can't really determine if that portion looks different than it has or currently contains food. From previous ~62 hour images, it looks like the head area appears slightly longer before tapering down towards the spine, and this area does have the earlier parts of the gut, but also contains other organs and could just be a slight difference in development speed or coloration. I also see that the parvocalanus culture even in its current state produces a lot more nauplii that fit through the 100 micron sieve, but that those largest size are still definitely too large for one to eat. I will move to an 80 micron sieve for the size the food must pass through, but am considering ways to try and not rely on a 45 micron screen to catch it.... maybe first I check the water that comes through the 45 micron sieve for nauplii to see if it would even benefit.

Here's the view under the microscope, though, through the 2.5x, 5x, and 10x objectives and with some shots of both movement and the larva at least partly on its side to show those gut structures. Maybe someone else can make heads or tails of it.


I caught some more eggs this evening and I've still got some larvae in this run, so the project continues.

 

[JoJosReef]

Following. Great work, really amazing effort.

 

[DaJMasta]

Good news and bad news. Bad news first: The number of larvae in the attempt started on the 16th has dropped a lot - I saw one or two during the day, but it actually took me minutes to find one this evening. The eggs collected yesterday also hatched pretty late - around 7:20 pm, which is like 22.5 hours post spawn.

But there's good news too. The one I caught and looked at today looks different - bumps in the gut and those little round 'plates' with two dots on them behind the head are pushed out noticeably, and the little guy seemed eager to try and eat things.


I also did a little testing with the sieves last night - I put the 100 to 45 micron sieves in the 30 and 25 micron sieve, and then examined what was getting through. There are definitely nauplii that the 30 micron sieve catches that the 45 doesn't, their characteristic jumps were visible even though they're tiny specks by eye. I neither saw any significant number in the 25 micron sieve or in the water that came through all of them, so for the run that starts getting food tomorrow morning, I'm using 80 microns as the upper limit and 30 microns as a lower limit. This probably won't last super long, since the 30 micron sieve takes so long to drain, but it seems like it will be very important in the first week, at least.

Some promising developments on the new food!

 

[F i s h y]

Good news and bad news. Bad news first: The number of larvae in the attempt started on the 16th has dropped a lot - I saw one or two during the day, but it actually took me minutes to find one this evening. The eggs collected yesterday also hatched pretty late - around 7:20 pm, which is like 22.5 hours post spawn.

But there's good news too. The one I caught and looked at today looks different - bumps in the gut and those little round 'plates' with two dots on them behind the head are pushed out noticeably, and the little guy seemed eager to try and eat things.


I also did a little testing with the sieves last night - I put the 100 to 45 micron sieves in the 30 and 25 micron sieve, and then examined what was getting through. There are definitely nauplii that the 30 micron sieve catches that the 45 doesn't, their characteristic jumps were visible even though they're tiny specks by eye. I neither saw any significant number in the 25 micron sieve or in the water that came through all of them, so for the run that starts getting food tomorrow morning, I'm using 80 microns as the upper limit and 30 microns as a lower limit. This probably won't last super long, since the 30 micron sieve takes so long to drain, but it seems like it will be very important in the first week, at least.

Some promising developments on the new food!

That's great news! Keep us updated.

 

[DaJMasta]

Not a whole lot to report today, I still have at least one from that run from eggs caught on the 16th, and I got it under the microscope. It does look subtly thicker and more developed, but it's not all that different from yesterday in overall form. I did see a gut contraction (a pinching near the base of the head that did not flex the spine) shortly before turning on the video capture, but it rolled over for me to give a good 3d effect.


Got one of the larvae from the second run under the microscope too, but those early larvae are really bulbous (hard to really get a good idea of what's going on), and since they don't stay horizontal and don't reposition as often, it's quite tough to get a good shot of them all in focus. I can't say there's obvious food in the gut, but the yolk is still quite large (larger than I remembered, maybe this is an indirect sign), but it should be clearer if they are eating tomorrow. I still have at least a few, so I'm hoping I get more to survive than the first batch with the smaller feed size.

 

[kdx7214]

Very cool to see! I'd love more info on your hatching/rearing setup. Someday I'd like to get back into breeding. I've bred clowns but would like to try something pelagic.

 

[DaJMasta]

I can show you some info, but it's probably not what you're expecting at this stage. Right now, the most success and the easiest checking on them by far has happened in a plastic basket that's about a 100mm cube hanging on the side of a mature aquarium. This is what the more recent run looks like today:

Not at all glamorous, not all that clean looking, and to my eyes I can't actually make out a larva even though I know there are a bunch in there. After the initial day or so of development, they seem to start drifting down in the water column - I think the same behavior that's been making them hard to image under the microscope. This may point to it being useful for the vessel to be deeper, because I think it's generally bad if they actually contact the bottom regularly, but I will have to see what options there are there. There is a little bit of skin on the surface of the water because I've only cleaned it once and there is virtually no surface agitation, and I wish the basket was hanging deeper in the tank because the ~25mm gap to the top of the basket makes it so that pouring the food in has a high minimum momentum and pushes the larvae inside around a lot.

That said, this image also shows something important - food availability. These parvocalanus nauplii seem to be pretty phototactic, and the little yellowish dots in a cloud in the middle are them. The important thing is that this is the basket that only started getting food yesterday, and only smaller than 80 micron sieved food, so the new parvocalanus culture is producing what appears to be an adequate number of nauplii of the selected size.

But the setup is this, the longest runs I've had have the larvae living entirely in these little boxes, except for brief stints under the microscope. I've gotten ok at handling them, there's a good bit of finesse/technique catching them (using a tiny beaker, trying to surface skim them up or slowly come from behind fully submerged) and keeping them from being injured in the process (after transferring to the well slide you need to check it for the larva, and it's a good habit to get a little more water in the beaker to make sure they aren't stuck to the side, then flush them out with a pipette and some extra tank water with the slide as close to the water line as possible when done), and there's a good bit of technique in just being able to see them - they are tiny, clear, and don't move a lot unless disturbed. The plan is to transfer to a bucket with a central spot for aeration, but it's also harder to monitor them at this size in there, so at least until I think they are reliably living a week or two, it's unlikely that I'll start many runs in the bucket - the extra volume also means it's harder to supply enough food and you have to do water changes (which are a nightmare with larvae this fragile).

The boxes they're in are part of this: https://www.thingiverse.com/thing:6168731

The buckets I'm planning on eventually moving them to are this: https://www.thingiverse.com/thing:6436667

 

[armyvetheather]

This is really cool and I'm excited to follow along and see what happens. Best of luck!

 

[b1llvance]

I really need to print this snagger. I hate printing with supports so I’ve put it off for a year now but I need to do it.

 

[DaJMasta]

Some adjustments to the spacing of the interface layer, the gap between the top of the support and the bottom of the part, and making sure the temperature isn't too high, and it's not so bad for me. I go with the conventional ones rather then the organic supports, and while the holders for it definitely need them, they're all flat surfaces and none are super critical to the geometry of it, so a little ragged and they'll still fit and won't be too visible.

So long as you're using familiar materials, it's a bunch of testing to get a reasonable result, and then you just keep those settings indefinitely.

 

[kdx7214]

Some adjustments to the spacing of the interface layer, the gap between the top of the support and the bottom of the part, and making sure the temperature isn't too high, and it's not so bad for me. I go with the conventional ones rather then the organic supports, and while the holders for it definitely need them, they're all flat surfaces and none are super critical to the geometry of it, so a little ragged and they'll still fit and won't be too visible.

So long as you're using familiar materials, it's a bunch of testing to get a reasonable result, and then you just keep those settings indefinitely.

Also need to make sure you use PETG. PLA will soften in saltwater due to the ph.

 

[DaJMasta]

Definitely PETG, I think most brands/colors should work, but I've used mostly Overture branded black and clear PETG in my tanks for a few years now and have yet to see ill effects or degradation.

I think that run collected on the 16th really is down to one individual, but that means I saw the same fish under the microscope today, and if I can see it at all tomorrow, it's a record for longest survivor so far. It looks a little more developed/defined than yesterday and still shows good signs of eating, but interestingly, those occasional contractions I could see yesterday have gotten more pronounced. Right at the base of the head, and right ahead of the fins I can see muscle movement, not so much specifically of the gut, but that kind of moves the direction and protrusion of the head slightly - like a neck, but this is a fish. My assumption now is that the visible fins are probably the ventral fins, whereas the contracting muscles I'm seeing will eventually be associated with the pectoral fins... but it could just be some jaw muscles developing or something, I'm certainly not qualified to say one way or another. This isn't a behavior I've seen before in others, though, so I think this is developmental progress even if it doesn't have a significantly different looking form.

The run collected on the 19th is starting to look like a proper fish shape now, and you can see both the tiny stubs of fins as well as a slight pinching right around that area in the body shape now in early day 4. I can't say for certain that there's food in the gut, but it does appear that it's not wholly smooth and not super closed. The population is also about the same density as the last day - maybe a dozen or two remain - both of which I think are good signs.

They aren't hugely different in length, but there is more definition and more contrast in the older larva, and their movements are definitely more coordinated and measured. You can see the lens of the eye developing in the 4 day old larva, but it isn't until the back of the eyeball is opaque that it's going to be usable. In the start of day 7 larva, you can see more differentiated, fully darkened eyeballs, though I haven't seen them move yet.

 

[DaJMasta]

I saw it during day 7, but not at the end of it. The run that started on the 16th ended maybe 4 hours shy of a full week post spawn. That means it's my longest run by about 12 hours, but of course not the ending I hoped for. While it's a literally microscopic larvae, I did feel a little attached - it was the only one of the batch for 3-4 days, so I caught it and saw it under the microscope several times.

The batch that started on the 19th is still going, though, and with multiple individuals. Rather than a microscope picture today, how about my normal view:

See it? To the left about half way from the vortex.

Zoomed in a bit:

The focus of the phone is a fickle thing, and the bright side lighting distorts the look, but the ball on the end of the stick has a tiny bit of cleft - the two shining spots of eyes in the face, and by far the most reliable way to spot them even confined to these 100 mm cubes.

In any case, some notes from the now ended 3 day older run:
Twice I missed a feeding from my 3 500 mL of culture a day, I noticed losses both times, though they could be coincidental
The second loss was the last one, I fed once around lunch time, missed the afternoon one, and didn't see it in the evening
I don't think it's just that - but prey density may still be on the low side. I did not see the little cloud in the box with it in the final day, and I was feeding less because the single 500mL-of-culture feeding was split with the smaller portion to that box, the larger portion to the run with more individuals left.

There's also the chance it was hydroids - I see a couple of fibrous things in the bottom of the vessel that have stuck to some detritus and it would not be the first time some hydroid spores/fragments had been collected when collecting the eggs. It could also still be something else - water quality in these boxes is probably not splendid, and I was handling it quite a bit (though it appeared to be acting normal before I lost it.)


In any case, the run from the 19th is starting to approach the record in a couple of days, and while I didn't collect any eggs this evening, I should be able to get some tomorrow night for another ongoing attempt.

 

[kdx7214]

I saw it during day 7, but not at the end of it. The run that started on the 16th ended maybe 4 hours shy of a full week post spawn. That means it's my longest run by about 12 hours, but of course not the ending I hoped for. While it's a literally microscopic larvae, I did feel a little attached - it was the only one of the batch for 3-4 days, so I caught it and saw it under the microscope several times.

The batch that started on the 19th is still going, though, and with multiple individuals. Rather than a microscope picture today, how about my normal view:

See it? To the left about half way from the vortex.

Zoomed in a bit:

The focus of the phone is a fickle thing, and the bright side lighting distorts the look, but the ball on the end of the stick has a tiny bit of cleft - the two shining spots of eyes in the face, and by far the most reliable way to spot them even confined to these 100 mm cubes.

In any case, some notes from the now ended 3 day older run:
Twice I missed a feeding from my 3 500 mL of culture a day, I noticed losses both times, though they could be coincidental
The second loss was the last one, I fed once around lunch time, missed the afternoon one, and didn't see it in the evening
I don't think it's just that - but prey density may still be on the low side. I did not see the little cloud in the box with it in the final day, and I was feeding less because the single 500mL-of-culture feeding was split with the smaller portion to that box, the larger portion to the run with more individuals left.

There's also the chance it was hydroids - I see a couple of fibrous things in the bottom of the vessel that have stuck to some detritus and it would not be the first time some hydroid spores/fragments had been collected when collecting the eggs. It could also still be something else - water quality in these boxes is probably not splendid, and I was handling it quite a bit (though it appeared to be acting normal before I lost it.)


In any case, the run from the 19th is starting to approach the record in a couple of days, and while I didn't collect any eggs this evening, I should be able to get some tomorrow night for another ongoing attempt.

I've seen other posts where people have included some phyto in the culture container to help improve water quality. I realize that it would make visibility tougher, but might help control water quality a little bit. Just a thought

 

[DaJMasta]

That's certainly been a thought, but I'm in the process of working it out. I have added phyto to cultures before and seen mostly negative results - but I think I know why. I believe my proportions of f/2 added were too high, and in the last few weeks I've been trying to decrease the dose and monitored culture density as well as nitrate in the mature culture (and also centrifuged the culture to make sure the phyto cells don't interfere with the nitrate test), but I haven't yet tested a mature culture with the current fertilizer dosage, so until I can verify that it won't add any nitrates to the culture, I won't be adding it. Once I can confirm it's just phyto, I will probably add some for trials - but I also have to be careful with when. The prey items are very small but grow quickly, so feeding them in the first may week may actually make them grow to outpace the size of the larvaes' mouths, so while clean phyto should help prey density and nutritional value, it also probably can't be introduced too early.

While the tank the basket holding the larvae is in has decent water quality, it isn't the pristine water I sometimes I hear has a positive impact on larval development. The few ppm of nitrate in the system combined with the very low exchange with the small volume in the basket may be a negative, which is probably another reason to get into a bucket as soon as I can verify they're making it through the early stages and I no longer need to keep such close watch.

 

[kdx7214]

That's certainly been a thought, but I'm in the process of working it out. I have added phyto to cultures before and seen mostly negative results - but I think I know why. I believe my proportions of f/2 added were too high, and in the last few weeks I've been trying to decrease the dose and monitored culture density as well as nitrate in the mature culture (and also centrifuged the culture to make sure the phyto cells don't interfere with the nitrate test), but I haven't yet tested a mature culture with the current fertilizer dosage, so until I can verify that it won't add any nitrates to the culture, I won't be adding it. Once I can confirm it's just phyto, I will probably add some for trials - but I also have to be careful with when. The prey items are very small but grow quickly, so feeding them in the first may week may actually make them grow to outpace the size of the larvaes' mouths, so while clean phyto should help prey density and nutritional value, it also probably can't be introduced too early.

While the tank the basket holding the larvae is in has decent water quality, it isn't the pristine water I sometimes I hear has a positive impact on larval development. The few ppm of nitrate in the system combined with the very low exchange with the small volume in the basket may be a negative, which is probably another reason to get into a bucket as soon as I can verify they're making it through the early stages and I no longer need to keep such close watch.

Thanks for keeping this updated. I've bred benthic fish, but pelagic is something completely new to me. It's amazing to watch you figure it out for this species.