The Lightning Project

The ongoing saga of the PNG Lightning Maroon Clownfish Breeding Project

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As promised, I finally got time to look at the offspring that Mike Doty had managed to rear from my F1 Lightning X Lighting Maroon Clownfish pair.  While Mike had initially stated he felt the split was 50/50, today’s visit showed something different.

All told, I only managed to count 3 white stripe offspring in the BRT. Meanwhile, there appeared to be 6 distinctive Lightning Maroon-type offspring. What I didn’t see were any fish that, at this point, looked atypical from either known phenotype.

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With only 9 (approximate) juveniles, the sample is far from conclusive. The 3/6 split could be representative of anything from a 50/50 split to a 25/75 split. A new phenotype could be missing because none simply survived in this first successful run.

That said, the 3/6 split, if a valid sample, would represent something we perhaps don’t want to  see.  It would imply that Lightning is straight dominant, but it would also imply that a double-dose of Lightning is in fact fatal to the offspring and they fail to develop.  This is one of the current working hypotheses for the Snowflake gene in Ocellaris, and anecdotal reports continue to bolster that line of thinking (reports are anecdotal in so much as a breeder saying “It’s about a 60-70% snowflake result” is not the same as a breeder saying “I got 140 Snowflakes and 65 wild types in this clutch”).

It is fair to say that there is still hope for these fish – they are QUITE YOUNG and all we can truly discern at this point is that they outwardly either are white stripes or aberrant.  My tune could change as these babies develop, or if subsequent results are different. It is not surprising to me though, to see similarities emerge in various mutation types.  We already have Picasso/Platinum and the sister mutation of DaVinci/Wyoming White. Much as I wonder if these twin mutations could represent the same gene in different species (or simply a similar genetic mutation in sibling species), would we come to find out that Lightning is in fact not unlike Snowflake? After all, genetic analysis has revealed that the Maroon Clownfish are in fact very closely related to the Percula/Ocellaris complex, so it would not be surprising to find similar or the same genes present in all these species based on their common shared ancestor.

It is also an interesting footnote to observe that these tiny juvenile clownfishes DO represent a the first F2 generation of Lightning Maroons.

Here’s a quick rundown.

The 5th spawn of the F1 Lightning X Lightning Maroon Clownfish pairing was collected and hatched by Mike Doty while I was away.  In short, he scraped off the eggs, hatched them in a 1 gallon jar in a water bath with simple aeration, 75% clean new water.  Come November 28th, Mike relayed that settlement had started.  The moment we’ve been waiting for was here – is there something new?

Well, Mike’s first words were “About 90% sure we have some normal striped fish.”

This, of course, does the following:

  • likely rules out Lightning as a simple recessive gene. If it WAS recessive, then both parents would be “double dose” aka. homozygous, represented as l/l, which means that each parent could only contribute a recessive lightning gene, and thus, each offspring would also get one copy each, one from mom, one from dad, and thus, could only be l/l as well. For the moment, while another couple test matings will bolster the data, the fact that there are white-stripe offspring pretty much precludes this being a standard single allele, single locus, recessive trait.
  • does not rule out straight dominance. If it was straight dominance, and each parent is “single dose” aka. heterozygous, represented as L/+, then 25% of the offspring would not get a gene from either parent, and thus, 25% would be white stripe maroon clownfish.
  • nor does it rule out partial dominance. This of course, would work the same way as dominance, except that 25% of the offspring would get a lightning gene from EACH parent, and would be homozygous for Lightning, represented as L/L. This is the scenario that most people are hoping for, because with the new homozygous state, there comes the potential for a new phenotype that could be different from the Lightning that we know.

As of today, 12-8-2014, I spoke with Mike briefly and have to relay this news – while he doesn’t have many babies left, he believes that the phenotype split is roughly 50/50.  That is to say, half white stripes, half lightnings.  So far, he also has not seen anything unique or new in this F2 generation.  I have yet to see the babies for myself, and have yet to take pictures or do a headcount, but these cursory, informal results, mirror another clownfish mutation that seems to not fit the mold as we’d expect – SNOWFLAKE in Ocellaris.  It’s my hope to get over to Mike’s today yet to see for myself.

In other news, the 6th spawn of the F1 Lightning X Lightning pair was put down on 11/29/2014. It appears I finally won the battle of the tiles:

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LxL Spawn #6

Meanwhile, I brought some new clownfish into the fishroom earlier in November (the 19th and 22nd) and was trying out the new Ick-Shield food from New Life Spectrum.  This is basically a Chloroquin-laced pellet food that is meant primarily to prevent disease such as Crytopcaryon, Amyloodium, Brooklynella etc…pretty much the things which are sensitive to the active ingredient. I decided to not simply feed this fish to the new arrivals, but also to feed it to one of my holding systems AND the wild Lightning Maroon and her mate as preventative medication, just in case.

Well, it turns out that there is an unfortunate side effect to this feed; it seems to shut down breeding activity.  All my routine pairs stopped spawning. The Lightning and her mate did finally put down a spawn on December 1st, 2014, #46.

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Unfortunately, it appears as though the spawn was not fertilized…the eggs didn’t develop, and after 48 hours they were gone. A few days after that, I read, anecdotally, that Chloroquin can cause male sterility??? Not permanent according to the rumor, but certainly a potential setback. As far as the efficacy of the food, my jury is out. The larger fish which were feeding well on it by and large remained disease free, but not all did.  I still had a Brooklynella outbreak, although not in the fish I would have necessarily expected. Once that outbreak started, it then affected other fish as well despite their feeding on the pellets.  I’m also seeing either Cryptocaryon or Amyloodium on fish which were visually “clean” upon arrival, which were in dedicated QT systems, feeding on this food from day one.  So the question here is were they simply getting ENOUGH feed as they were small fish which cannot readily eat the small pellet size.

So of course, one is left with questions, not answers.  There is no way to say the food didn’t work, nor is there any way to prove that it does work. Absence of disease is not proof of prevention, that much I know for certain. Lack of a cure, or lack of prevention, which IS documented, only raises questions about why it didn’t work as suggested and certainly requires investigation (eg. would a smaller pellet size be better accepted…could these failures stem from simply lack of feeding, or lack of sufficient feeding, thus insufficient dose to the fish?).

Circling back to LxL Spawn #6, as the week progressed an interesting change in behavior occurred starting around December 4th, 5 days post spawn.  The larger female F1 Lightning became belligerent towards the male, and over the day drove him from nest tending duties.  December 5th, a Friday, would have been 6 days post spawn, and the night of the first hatching.  I was simply swamped with preparations for sending our dog to live with my brother, and failed to pull the tile.  By morning, Saturday, December 6th, 2014, it appeared that I had not missed much…most if not all the eggs were still there. The pair remained at odds.  We left for the weekend to ship our family dog, and upon Sunday, December 7th, there were still a few dozen eggs remaining, although they appeared potentially dead and disappeared throughout the day.  By nightfall, the pair was starting to be less antagonistic, but I am still keeping a close eye on them.  Hopefully, we’ll get another spawn soon – this was the only mature pair in the house that didn’t receive Chloroquin-laced foods (as they don’t reside in the fishroom with the rest of the fish).

Just jotting down notes…Spawn #45 of the wild Lightning Maroon pair happened on the evening of 11-19-2014.  Meanwhile, LxL (the F1 Lightning Pair) Spawn #5 was gone by the same day….they obviously didn’t like me tiling their entire tank. I need to go by more terra cotta flowerpots…maybe they’ll like that.

So much going on and I’m NOT doing a proper job of tracking things.

I should start with the unrelated F0 PNG White Stripe Maroon Clownfish pair…they spawned sometime last week, possibly Wednesday the 12th. I didn’t make much of a fuss about documentation of it because I knew I couldn’t deal with them at any rate; I can’t remember if it was very late Sunday night, or more likely Monday afternoon, when I found the last few eggs just sitting around in the tank.  This pair, overall, is spawning quite infrequently.

The Lightning Maroon Spawn, #44, never made it to hatching.  Something happened, and the pair consumed the eggs.  Could the Lightning foundation pair be winding down or did they just have a bad spawn?  Their prior spawn produced a good amount of juveniles, which were undergoing settlement and on TDO A by last Thursday, which is when I drained them down to about a gallon, filled with clean new aged saltwater, and then turned them on to the larviculture system.

LxL spawn #4 was one I was going to let go because I wasn’t in town (I was off speaking at the Milwaukee Aquarium Society’s “Fish Bowl” event), but in short, Mike Doty decided to play with harvesting the eggs.  He took approximately 25% of the next out, using my little egg siphon, on Friday night (Nov. 14th).  I got a text while in Milwaukee…there was a hatch..and a big one.  Surprisingly, he simply did it in a 1 gallon olive jar with an air feed, sitting in a water bath with probably 75% new water.  If I had done that, every last egg would have been dead.  All credit where it’s due – Mike may well in fact rear Lightning X Lightning offspring before I do (they’re at HIS fishroom ;) )  Mike further relayed that this was a split hatch, with more eggs present Saturday the 15th, but all gone by 4 AM Sunday morning.

Yesterday, I caught wind that the F1 Lightning X Lightning pair was going to spawn again…the female was roving the tank with her ovipositor down.  I did my best to FURTHER add tile to their tank, blocking their prior spawning spot off..which caused them to move to another side of the tank’s back wall, which I then blocked off with tile. They were still going through the motions of finding a spawning site when I turned the lights off after Midnight; technically they spawned in the overnight on the wee-early morning of 11-18-2014, but if I was to be thinking about these eggs properly, then normally they’d be a 11-17 spawn.  Ultimately, they selected the ONLY section of black plastic background that I cannot get tile on (near the tank’s filter return).  Either the nest was small, or they ate most of it, and I am considering foregoing working with it.  I’m thinking that what this pair may really need is a clay pot…that might be the ticket here.  So LxL Spawn #5 has been put down, but for now, probably will be left alone.

So despite two nights of attempted hatching of harvested eggs, I ultimately only got ONE viable larvae out of LxL Spawn #3.  It is in a BRT all by itself.

On 11/7/2014 – the F1 Lightning X Lightning pair threw down another nest…and once again, not on a tile.  I won’t be in town to try trapping or harvesting this nest, so I’ll have to wait for the next spawn. Will update with some photos as I’m able.

Not a lot of time tonight, so I get to be real quick.

Spawn #44 was laid by the F0 pair on late evening of 11/3/2014.  Spawn #43 ended up no hatching much when it was finally set up, but seems to have hatched apparently during the morning.

I opted to harvest 50% of the nest from the F1 Lightning X Lightning pair tonight, the 7th night post hatch, the overnight from 11/3 into 11/4. I used a very simple harvesting technique, using a section of air tubing, inserting the tip section from a 2 ml pipette, which was cut at an angle on the harvesting end.  The eggs were scraped from the back wall while the tubing was siphoning into a specimen cup.  The harvested eggs were added to a brine shrimp hatchery resting in a water bath in a BRT, with a bubble rate of a few bubbles per second.  3 larvae were dislodged from their eggs during harvest…one was viable and started swimming, while the other two appeared lifeless.

It’s the 7th night for Spawn #43 which was spawned on 10-22; we’re on the night of 10-29 going into 10-30 now; there wasn’t any noticeable hatch on the tile over the 6th night (I left it with the parents yesterady)….it will get pulled.  This is what Spawn #43 looked like when it was laid fresh on 10-22…I should also point out this is the first time I’ve taken pictures of the F0 original wild pair of Lightning Maroon X White Stripe PNG Maroon Clownfish.

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The larvae from spawn #42 that are still alive have largely settled out…it was a small group, but I’ll find a way to work with them.  More importantly, it looks like LM X LM Spawn #2 might be over…I have not found any larvae in the BRT, so one more look, I repurposed  it for Spawn #43.  The eggs got the usual dip in H2O2, but started hatching during it, so they were moved straight into the BRT with 8 gallons of broodstock water and a few ML of RotiGrow Plus (only thing I had thawed…it’ll be fine for the first 24 hours and by that point I’ll have RotiGreen Omega up and running).

The most frustrating news is arguably LM X LM Spawn #3.  So..to recap.  Here was the first spawn:

F1 (Lightning X Lightning) first spawn for the Lightning Maroon Clownfish

 

So…I took some proactive steps and tiled the back wall of the tank.

This was LM X LM (or LxL if you prefer) Spawn #2 on the first attempted hatch night:

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Yeah..that gray patch on the side..those are the eggs…

So that didn’t turn out obviously…so I tiled the side wall as well.

Here is Spawn #3, laid on 10/27/2014:

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The internet shorthand FML seems somehow appropriate. Clearly the pair is flipping me the middle fin.

 

It’s been a few late late nights waiting for results on the Lightning X Lightning pair’s second spawn. Given that they had either spawned on Thursday, Oct 16th, or Friday, Oct. 17th, and going off of their parents 6 day hatch window, I set up their tank on the evening of Wednesday, Oct 22nd, to prepare for a possible hatch.  This made sense given that it would either fit into the 6 day window if the eggs had been spawned on the 16th. No hatch was observed. On Thursday night, Oct 23rd, I encountered a very small hatch around 3 AM, a solid hour after lights out.  But, nothing furhter happened, so the 10 or so larvae I had snagged simply sat in the snagger for the next 24 hours.  And now, Friday night, the 24th, going into Saturday AM, it’s now just shy of 4:00 AM and I just finished “putting away” around 50-100 larvae from the F1 Lightning X Lightning pairing.  I snagged them with the Vossen Larval Snagger…ultimately most totally bypassed the snagger, so I had to set up a LED crank-charge flashlight and manually attract the larvae to it. I also had to really crank up the flow rate faster than I was comfortable with, but if I hadn’t done that, I watched as most every larvae would swim back out of the trap once it realized what was happening.

I gave LxL Spawn #2 a very weak dosage of RotiGrow Omega (yes, I know this is not ideal, but I need to thaw out some new RotiGreen) and placed a half gallon’s worth of SS-strain rotifers (because I have those going now as well, thanks Mike) along with roughly 4.5 gallons of broodstock water in a BRT downstairs. Now we wait. Knowing how these fish tend to color up quickly, as long as I don’t massively screw up, we should get an idea of what’s going on very soon…eg. within 30 days.

The larvae from Lightning Maroon Spawn #42 did not do well despite what appeared to be a solid hatch on their second night.  I’m guessing no more than 6 are still viable.

Back on Wednesday, Oct 22nd, 2014, the Lightning Maroon and her mate threw down Spawn #43.  A beautiful nest.  I have a lot of photos in the camera to post, but I’m still recovering from two back-to-back laptop rebuilds, so pictures are just going to have to wait for now.

Lots of little updates

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Spawn #41 was left to go for itself because the Lightning X Lighting Spawn #1 took priority.  Unfortunately, by the time the eggs reached the anticipated hatch night, Wed, Oct 8th, there were only 7 eyed up eggs left.  I attempted to harvest them by using a rigid air tube attached to a flexible tube to scrape and siphon them out…this broke one egg but the rest went through intact.  By the next morning in the incubation cup (a half gallon specimen cup with broodstock water), the eggs were dead.

So back to the F0 Lightning X White Stripe pair; Oct. 11 they laid Spawn #42, and tonight, Friday, October 17th, 2014, I pulled the eggs and gave them the typical H202 Bath..about 11 minutes in, we (Paul Whitby of HighDefCorals is in town to speak at our club) noticed that larvae had begun to hatch, so the dip was ended and I placed the tile in the cleaned, readied 6 gallon BRT which had 5 gallons broodstock water and 1 gallon new water.

In other news, the Lightning X Lightning pair put down their second spawn somewhere between 5 PM on Thursday, Oct 16th and 4:30 PM Friday, Oct. 17th. It’s a much larger nest, which was expected. Despite having basically tiled the entire backside of the aquarium, the pair thwarted my efforts by spawning around the corner in a tight area of black plastic background. I don’t know when the proper “harvest” date will be for these fish; I’ll like try my Vossen Larval Snagger this time around. But I also know that I’ll be putting even MORE tile into their tank now ;)

So yeah…October 2nd, I’m just walking by the tank and I see this…

F1 (Lightning X Lightning) first spawn for the Lightning Maroon Clownfish

We are about to embark on the next exciting chapter of the Lightning Maroon Clownfish and genes.  My hypothesis is that the Lightning gene is partially dominant or dominant. The offspring from this spawn, if successfully raised, will yield important answers that may shed new light on the genetics at play. Will we see something NEW come out of this? Well..I certainly hope so and it looks like I’ll get to name it!

Let’s give that a closer look:

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