Culture Journal, Species: Brachionus plicatilis

General
Species:  Brachionus plicatilis
Species description:  Rotifers, L strain
Culture source (link if possible): Reed Mariculture 
If algae, CCMP # (Optional): 
http://ccmp.bigelow.edu/
Culture Establishment Date:  1/3/2013 
Continuation Date: 

Culturing Vessel Details
Salinity:  1.020
Temperature:  ambient room temp. ~79 F 
pH:  7.7

Vessel description:  5 gallon orange plastic buckets (from Home Depot)
Lighting description:  ambient room light, but culture buckets are covered with a lid to keep out cats
Lighting cycle:  mostly natural day cycle from ambient room lighting
Aeration description: slow boil   

Methodologies
Split methodology:

Culture medium description: 
new saltwater, mixed at least 24 hours prior to 1.025, and then diluted down to 1.020 

Cell count:
At culture start, it was ~1 million rotifers. 

Reference links:  

Additional Information
(No Pictures or Videos in the Section Please)
Notes: 
This is the third culture journal attempt.  The first culture never really got started. The second ran successfully from 9/11 - 12/11 and crashed from neglect while I had the flu.


You will be required to provide photographic evidence and as much detail as possible about your project in this thread.
If your thread does not contain detailed enough photos  and information the MBI Council will not be able to approve your reports.

- David Gibson
Denton, TX

It's been over 24 hours now, and I am still not able to confirm there are any live rotifers in the culture.
 
The RotiGrow+ does not seem to be clearing, or it's too slow to notice.  Also, I have filled a 5mL glass vial again and backlit it with a flashlight.  I was not able to make out any movement from anything to the naked eye.
 
I suppose the only way I'm going to know for sure is to get a microscope, but in the meantime, I am going to let the culture do what it's going to do and see if a few made it.

After a week, I've now split the culture into two buckets.  Something in it is eating the RG+, but I can't tell if it's rotifers yet.  I've got a microscope coming on loan next week, so hopefully I'll figure it out.
 
Here's the culture after sitting overnight before adding RG+

A good magnifying glass is all you really need to tell rotifers (even for my poor old eyes).  Just put a sample in a test tube, hold it up to a bright background, and use a moderately-powered magnifying glass.  You'll be able to see these U shaped things moving slowly under their own power.  Depending on how the light hits them, they may look white.  If they're carrying eggs, they will be U shaped things with little dark bumps on their bottoms!

My daily routine has been to add a few drops of RG+ if the cultures were growing lighter, and then an equal number of drops of Seachem Prime (no Chlor-Am-X locally, and I'm holding off buying a tub online with Reed's new Chloram-X products so close to release).  I haven't sieved out anything since establishing the culture, and there has been little detritus settling to the bottom. 
 
As of about a week ago, I hadn't been able to find anything that resembled a rotifer.  So, I took my cultures and ran them through a 120micron sieve into new buckets with 50% new water. 
 
The muck at the bottom of the buckets I put into some 450mL bottles and let sit until they settled.  I could see some white swimmers jumping around in that, which I believed to be some type of copepod nauplii.  I poured off most of the water, then put the remaining muck in a 1g bucket with new saltwater and some RG to see if I had any rotifer cysts settle out.
 
It's now a week later, and there isn't even signs of the copepods now.
 
I took a test tube full of culture water from each bucket and let them sit for a minute or so, and then backlit it with an LED flashlight.  I couldn't make out anything free swimming at all.  What few particles I saw slowly just settled down to the bottom.  No magnifying glass, but I do have good eyesight, so I thought I'd at least be able to see dust specs swimming around if I had anything.
 
More likely than not, I think I have something microscopic clearing the RG, but I don't think I've got any rotifers left.  I'm afraid I'm going to throw the towel in with this batch.  I can get a new starter culture locally and see if I can start over.
 
Frustrating, but all part of learning.

Don't feel bad.  My first attempt at rotifers went very much like what you are describing.  Yes, it is "all a part of learning."  Give it another go.  You'll get the hang of it!

Thanks for the encouragement, Jim.

I got another starter culture for round 2 today, this time instead of getting a local source I got some from a fellow club member that ordered from Reeds.
Do you think I've got the tint too light? 

I can see lots of critters running around this time, and they are fairly active, so I'm already a step up from attempt #1.  Incidentally, the rotifer culture I got, they look just like what I had assumed were copepods in my earlier attempt.  I hadn't found any videos of rotifers that weren't magnified, so I didn't know what to look for.  I thought they would be ... shorter? fatter?  Definitely thought they'd be slower.  These guys have this lurching swim, darting forward then coming to a stop before darting again.  Sometimes spiraling around drunkenly.  Sound about right?
 

The new starter culture has been so much more successful, the results are like day and night.  I split the initial culture between the two buckets and kept the water tinted lightly (approx. 6 drops of RG+).  Within three days, I have had the water clear completely within six hours, so I have doubled the amount of food I am adding.
 
This weekend, I had to feed three times daily, about 12 drops of RG+ to each bucket, and it would still be just about clear by the next feeding.  I would also add the same number of drops of Prime with each feeding (I'm worrying that I could overdose that though).
 
I took a test tube with a taped off section of 1mL, and I had a hard time getting a count, but my estimate is about 50 per mL density.
 
I'm going to start my harvesting today.  Buckets are also getting a good bit of detritus on the walls of the bucket too.  I am going to try just stirring everything up and exporting as I harvest, but if that doesn't keep up, I'll switch over to doing bucket changes.

Both rotifer cultures have been very successful this time.  I'm having a hard time finding the right amount of food to add so I can go from feeding three times a day down to two times a day.  As it is, even at three times a day, the water is completely clearing each time.
 
I'm harvesting only every few days since I am going to be out of town for several days and my tank sitter won't be doing any harvesting.

My rotifer cultures continue to cruise along.  Due to breaking down my main tank and being in the middle of upgrading to a new 90g, my clowns have stopped laying, so I haven't been harvesting.
 
I did get a short video of a sample taken.  That's about 1.5mL worth visible there.
 

- David Gibson
Denton, TX

Ugh.  Got bronchial pneumonia and was bed ridden for almost a week.  I managed to drag myself to the living room to feed my fish every couple of days, but I couldn't keep up with food/ammonia control on all my cultures and they have all crashed.
 
I've kept the air going and the water tinted for a few days now, but haven't seen anything rise from the bottom.  If I can't get the culture restarted from eggs, then I'll be restarting from scratch.

- David Gibson
Denton, TX

Keep working on that culture, it should rebound.  Just don't over feed and try to keep the PH up.
 

Ha ha!  I was just about to give up on my rotifer cultures, but then I got a Celestron 44340 for Christmas.  Of course, the first thing I do is to take a sample and start looking for anything moving.
 

 
Success!  It's incredible the amount of neglect that these two buckets received, and yet have managed to start to recover.  

- David Gibson
Denton, TX

Hmm.  Well, I seemed to have crashed both cultures on the same day.  One day they are both dense and thriving, the next I can't see anything floating in the water.
 
I had done a water replacement a few days prior, so there hasn't been time to build up much crud.  My suspicion is that I've been feeding RG+ and using AmQuel Plus for ammonia control while waiting for my Reed's order to get in (RGComplete and a bucket of ChlorAm-X).  The bottle of AmQuel says to shake before using and I can't remember if I shook it up that day.  That's really the only thing I can currently imagine could have happened to impact both cultures so fast and at the same time.
 
I was trying to use up the AmQuel before I cracked open my RGComplete.  I think I'm just going to toss the AmQuel and switch now though.  Hopefully they had time to make some cysts and I can bring them back.

- David Gibson
Denton, TX

I have restarted my rotifer cultures as of 1/3/13 (http://www.mbisite.org/Forums/fb.ashx?m=78510).  I've updated the initial post of this journal specific to this new culture attempt.

- David Gibson
Denton, TX

I wish you best of luck in this attempt. Just keep going and results will show up!

Thanks, Peter.  My clownfish fry are pretty much done with meta now, so my demand for rotifers isn't a pressure anymore.  This recent attempt has kept up with being pretty dense (over 100 rotifer per ML) since establishment, so I'm pretty confident that it'll go the distance.  Just need to not lose this one like the last, so no getting the flu for me!

- David Gibson
Denton, TX

David, health is on your side this time, so I am optimistic about this attempt.
Looking forward to see your first newborn clowns.