Culture Journal, Species: Brachionus plicatilis

General
Species: Brachiionus plicatillis

Species description:  common L rotifer
Culture source (link if possible):  Washington University Zebrafish Facility
If algae, CCMP # (Optional): 
http://ccmp.bigelow.edu/
Culture Establishment Date:  7/29/2011 , renewed by removing ciliates 8/15/2011

Continuation Date:  forever if possible

Culturing Vessel Details
Salinity:  20 ppt
Temperature:    ambient, around 75-78F mostly
pH:  8ish

Vessel description:  3 five-gallon buckets
Lighting description:  room ambient, fluorescent
Lighting cycle:  14 day/10 night
Aeration description:  open rigid airline to bottoms of bucket.  Fairly rigorous "boil"

Methodologies
Split methodology: Buckets are filled to about 4 gallons max.  Bucket one is primary Culture. One gallon is dipped out daily from the Culture bucket and placed in the second, Enriching bucket, with some placed in the third Tiggerpods bucket.  Fresh saltwater, one gallon,  is dripped into the culture bucket during the day. 
 
Every week, rotifers and their culture water are decanted from the culture bucket into a clean bucket.  Residual sediment is suspended in the residual water, and the remaining contents are poured via funnel into a 2 liter soda bottle and allowed to settle further. The original Culture bucket is wiped out with a paper towel, and the large volume of decanted rotifers and their water are returned to the bucket.  This preserves any biofilm/filter that may have cultured on the bucket surfaces, while removing a majority of the decaying matter.  After settlement, the soda bottle rotifers and their water are also decanted back into the Culture bucket.  
 
After about a month of this, the dripping method was abandoned. Too much work. 
Now I use 1-2 ml of RGcomplete or a mix of RG+, 1 part, plus 6 parts Nrich and an equivalent volume of 16 teaspoons chloramX/1 liter water.  This is added twice a day.  Once a day a gallon is dipped out of each bucket and and a gallon of 20ppt saltwater is added, room temperature all.  I have one bucket of rotifers alone, mostly, and one bucket with every copepod in my basement included.  I probably feed and exchange water out of the mixed bucket less than the rotifer only bucket, and yet they have equivalent or even more rotifers in the mixed critter bucket.
Culture medium description: 
Chlorinated saltwater, obtained from spent water obtained from water changes of fish tanks, is diluted to about 20 ppt, and pH is adjusted. Bleach is added at 1 ml per gallon, and the water is allowed to sit overnight or longer.  Water is dispensed into 1 gallon milk or other type of cap-able jug. Each day, one gallon is dechlorinated with excess ChloramX, and 6 parts N-rich with 1 part Rotigrow+ is added as needed.     This medium is dripped into the culture bucket for several hours during the daytime.  If more food is needed at night, a bolus  of RotiGrow+ is added to the Culture bucket along with an = volume of ChloramX.    
 
Update: I've been bleaching and then the next day, dechlorinating (sodium thiosulfate) the whole 12 gallons or so at one time, and then using this water to do  water changes on the rotifers, the copepods, the daphnia, the ciliates, and the dinoflagellates. ( Immediately after bleaching, I set up bottles for culturing phytoplankton, as the bleach will sterilze the bottles and airline sticks as well.  I dechlorinate just before adding the f/2 and the innoculum.)

Cell count: varies. I am trying an experiment to see how dense I can get a 4 gallon bucket culture .  Once I started using the RGcomplete, my density increased visibly, though I have not done any serious counting.
   

Reference links:  

Additional Information
(No Pictures or Videos in the Section Please)
Notes:  If sediment is noticed in the Culture bucket during the week, a brine shrimp net is used to remove most of it.



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.

I need to keep better records of what I am doing.  And be more consistent. 
My goal with the record is to see how dense a culture I can obtain, given what I am willing to do to keep rotifers going.  There may be a better way, but I am only willing to spend so much time and money on rotifers.

Today I checked my colony of rotifers.  I doubt that I have even 50 rots per ml. However, I've been fairly consistent for the past couple of days with keeping the bucket tinted with food, and today I noticed for the first time that at least half of the rotifers are carying two eggs, instead of one or none.  Seemed like there is food inside them, and I can expect some growth in the next 24 hours.
 
2 ml of RG+ in the media jug this morning.  The drip is not perfect.  Dripping can start at 2 drops per second, but as the hours go by, it slows down.  I think the valve gets a little clogged.  I've tried a few different valves, but nothing is ideal. May just have to start fast to end at the right rate.  May just have to live with it.

great going!
I'm doing a bunch of nanno right now and in about a month I'll start my rotifers again. How do you like the RG+? Do you think it's more economic than nanno or just easier?

RLTW

180 Gallon Mixed Reef

Then I heard the voice of the Lord saying, "Whom shall I send? And who will go for us?" And I said, "Here am I. Send me!" Isaiah 6:8

easier and more effective.  For clowns, no further enrichment is needed.  I happen to have some N'Rich, that needs to get used, so I am enriching the rots further, but Roti-Grow + is really more than enough.  It's no more expensive than nano or the old rotifer food that Reed offered.

This evening Culture looked good with many two egg females.  Added 10 drop bolus Rotigrow+ and ChloramX.

Try bubbling your drip container with a little air to keep the RG+ in suspension.  You'll get best results with multiple feedings if RG+ over the day.  I'd try doing a drip in the AM and one late afternoon.

Thanks, I'm trying that today.

looks like I've got O marina in my rots….kinda messes with the experiment. I guess since my clownfish are past meta, I can just sieve the rots out, wash well, and start again………..It won't matter if it shocks the rot culture.
Well, I have to do that if I am to try this method with any kind of reliable, quantify-able results.  OK, starting fresh tomorrow.
 
 

Filtered the culture into a 53 micron.  rinsed well, put into a gallon of clean saltwater, 25ppt in a clean bucket.  Added 1 ml food: 6 parts N-rich to 1 part RG+.  Added 1 ml food to another gallon of clean saltwater , with amply chloramX and dripped it in all day. Came home to a cleared culture, added 6 more drops food, 6 drops chloramX.  Culture has had rigid airline with BOIL level of air.

August 17:
Dripped another gallon with 2 ml food.
August 18:
Morning: About 10% of rots had 2 eggs, most had one egg.
Removed one gallon to Enrichment bucket.
Dripped in another gallon with 2 ml food over the day.
came home to clear water, only half the gallon made it into the bucket.  The siphon clogged.
Loosened siphon, and water was tinted green by an hour later.  Added food to enrichment bucket and Tigger bucket.

Continued with above rhythm. Culture does not appear to be getting denser.  Next, increase food to 3 ml per , twice daily.
 
That O marina I  had before turns out to be ciliates. 
Here's pix;

 

 

I took a pudding cup's worth of culture, poured it thru a 53 micron screen, and back washed it onto a petrie dish( 6 cm diameter) .  Here's a picture.  Not many or any two egg rots, some one egg rots, and yet still lots o rots.

I'm afraid I'm not being very consistent.  I am still only on 2ml food twice a day, but I've switched back to just RotiGrow+ instead of the Andy recipe.  I've wiped out the bucket, and split the culture into two buckets, as I ramp up in anticipation of having orchid dottybacks to feed. Of course that will be after MACNA, and someone will have to take care of the rots while I'm gone, and my favorite fish sitter will be at MACNA as well, so that leaves it to my darling daughter.  I've given up on this drip idea as it is too fussy. Now I'm just shooting two ml RG+ in twice a day with chloramX.  I did get an API ammonia test kit, and my ammnonia is between .25 and 0.5 mg/l, so that can be fixed with 2 ml of 120g/l chloramx. Right now my stock solution is 60 g/l, so I just add twice as much.
 
I do remove a third of the culture and replace it with 20ppt clean saltwater daily.

A quick check under the scope showed a lot of one egg rots, and quite a few 2 egg rots, so I think the change has done them no harm.

Before macna i got down to one bucket, and since coming back I've increased feeding and now have a pretty dense looking population.  4 gallons, one gallon removed per day to either feed larvae or something else.  replaced with 20 ppm clean water daily. chloramX, and rotigrow +
 
...@ two ml twice a day. Culture seems to be thriving, and doing better than the slow drip.  Every few days, I refresh the bucket to get rid of detritus. As of 9/26/2011.  Now I'm sure I've jinxed it.

RGcomplete arrived Friday.  It's Monday. I've used it at 3 times the volume I was feeding RotiGrow + for my rotifers, as directed.  This morning, there are significantly more rots in the filter, and when checked under the scope there are quite a few 2 egg females, whereas it has been quite a while since I saw any 2 egg females under the old regimen. I was skeptical at first, but the conversion is happening.  This may be a great product.  Thanks to the Reeds and Gresham.
 

The rotifers and I are loving RGcomplete. The indoor polyglot culture returned a dense brown, critter concentrate from a gallon of culture, once filtered.  Daphnia, T. californicus, and rotifers are thriving.

RGcomplete... one word (just like the label)   I'll hound you all till you get it right.  I will not have another Tiger-pod deal on my hands
 

whatever we call it, surely you cannot miss the praise this product is receiving.  When will it be commercially available?

I am not missing the praise, every letter in every praise post has been passed onto the rest of the company
 
However as we're in the middle of branding this product line, it's imperative that first adapters get the name right when talking about it.  The internet is pretty permanent and Google doesn't know if you spelled it wrong, nor will others when looking it up.

Just for you, I've changed all my misspellings.  I understand the need.  Sorry I messed up initially.  Spelling has not been my thing lately, nor typing neither.

Rotifers continue to do well indoors.  RGcomplete is good stuff, but I'm almost out of it, and I want to buy some, as well as some TDO A, B1, and lots of C1, but I don't want to place an order until I can get some RGcomplete! I guess I'll call tomorrow to see if I can get some more.
 
Rotifers also continue to do well, if not better in the mixed critter bucket. I am less attentive to water changes and feeding, yet it seems that there are more rots in the the mixed population than in the single population bucket, that I am actively monitoring.

what all is in the mixed bucket?

RLTW

180 Gallon Mixed Reef

Then I heard the voice of the Lord saying, "Whom shall I send? And who will go for us?" And I said, "Here am I. Send me!" Isaiah 6:8

A little of everything.  Rotifers, M. salina, T. californicus, A. panamensis, ciliates, and hopefully some O marina.
 
I've continued to feed this bucket, taking out a daily gallon full to feed aiptasia right now.  It seems to thrive.

i did a rough count of the rotifers only bucket, and got about 50 rots per ml.  Not great, but not bad either.

And for my continuing culture, here are a picture and a video:

 
click to see video:

 
I seem to be struggling with focus today, so I hope you'll forgive an old woman.

Rotifers seem to be going strong, and contaminating everything!
 

awesome. So are they doing better since it's all mixed together?

RLTW

180 Gallon Mixed Reef

Then I heard the voice of the Lord saying, "Whom shall I send? And who will go for us?" And I said, "Here am I. Send me!" Isaiah 6:8

hard to say. I haven't been able to get a good count of either bucket. The mixed culture always has a lot of detritus in it as well as the lots of critters.  The rot bucket has a contaminating tigger pod population. Not sure that it improves the rotifers. I am sometimes inconsistent in my husbandry.

Last week I put the "rotifers only" culture through my "coffee" filter which is a plastic mesh, probably 300 microns or so. Also, I've changed my daily routine.  At least once a day, instead of dipping out a gallon and replacing that water, I prepare a clean bucket with aproximately 1 gallon of clean culture water, then decant the culture bucket into it, thus removing the detritus that sinks to the bottom on a daily basis. Some detritus pours into the new bucket, but the the majority is removed.
 
From the old culture bucket,  I then enrich the rotifers, filter out the gunk, and feed the rotifers to my baby fish and aiptasia.
 
Since I've started doing this, I've noticed a large number of 3 egg females and 2 egg females, however my density remains at about 50 rots /ml.  I would like to double that.
 
Yesterday and today, even though I've checked and have lots of 2 and 3 egg females, the water is not clearing as fast as before.  Still at 50 rots per ml.  Not sure what is happening.

 
Hmm, that sure sounds easier than bottom siphoning the 5 gallon tanks like I've been doing.  One point for bucket vs aquarium.  I think they are tied now.

If you still want to do tanks, try running a fine brine shrimp net across the bottom.  I replaced one net with polyester organza from the fabric store, and I never siphoned again. The net collects the gunk in one or two swoops, and I can rinse it out in the sink.

Well, despite daily bucket exchanges, and frequent feedings, I still ran out of rotifers for my clownfish, and I'm expanding right now, so there are a lot of babies to feed.  I tried the T device that Gresham is working on, comparing it to just an open bucket with an airline, and there was no great improvement.  I think it is still being tweaked.  I think that air is more difficult to deliver to this device, although it improves if the bucket is nearly full.
 
I'm trying something else right now with some success.  I'm using a 17 gallon storage container, right now filled to only 8 gallons. I acquired a supplement of rots from my friend at the university, and have been able to gradually feed more, and get a better concentration of rotifers.  I haven't counted yet, but I can tell when I harvest them that there are more on the filter.  3/4 gallon of the new culture is giving me more than 4 gallons did of the old culture, and there is no gunk in it.
 
My friend at the U gave me a rather large sheet of the polyfilter that Reed recommends as an aggregated algae attractant.  I've attached it to a needlepoint sheet, and this lives in the container, submerged, until I rinse it under the full force of the tap at the sink.  I let it drain, and scrub the sides and bottom of the container with a toilet brush reserved for this purpose. Back it goes in after the brushing.  The container has stayed relatively clean, compared to the bucket method.
 
The filter material does take out some rotifers, but one must do that daily anyway. The daily spraying doesn't completely clean it, so I wonder how long it will be before it will need to be replaced.
 
I am using more air than before, with an open ended rigid tube.  The T device had an airstone, but the larger container sees a lot of water movement with the central airline and a lot of flow.
 
I am probably using 10-12 ml of RGcomplete daily, and since i am home, I feed every few hours if I notice that it is clearing a bit.
 
I should count them.
 

WELL THAT WAS INTERESTING!
 I counted a ml of the culture and got 35 rots per ml. Hmm, seems low.  Counted again, same thing.  Counted a third time, but took the sample from the floor of the container.  There were too many rotifers to count, perhaps 1000 per ml.  
!!!!!
 
Took a sample from the filter, and there were, again, too many to count. These were dirtier, whereas the floor sample was clean. 
 
This makes me think that the rotifers appreciate a little substrate to hang around in. Perhaps we should try regular filter floss in addition to the filtering rotifer floss.

Also, in my old method of decanting the old bucket into a clean bucket each day, I was perhaps removing more rotifers than I thought, and that may be why I would see the numbers dwindle.  If they are more concentrated at the bottom of the containers we are growing them in, we should brush and stir BEFORE we take out the 25-33% of the culture each day.

This new method with the larger container and the rotifer floss and the frequent feeding is working well.  I have lots of rotifers, and the container stays pretty clean. I just broke down and ordered a peristaltic pump from Bulk Reef Supply, because I am now spending more time with the rotifers than I am with the fish, and this constant monitering and feeding will have to stop.  Someday, I hope to be employed again….

After several days of fiddling with the pump and the timer to regulate the pump, I am reasonably well trained to keep this thing going, and hopefully grow even more rotifers.  Due to fiddling and failing to get the pump to do my bidding, I think I lost a bunch of rots, but we live and learn, and all is well right now.  Once I finish the final touches, I'll post a picture or two.  I may be rebuilding the table I've set the container on...

Well, when one is measuring, one cannot help but notice that one is going through 40 ml of RGcomplete each day, and still the rotifers look red instead of green, a sure sign that they are not being fed enough.  
 
I had a blooming blooming culture of O. marina in with them.  They were eating the algae before the rotifers could get to it.  I had to filter out all my culture, and completely change the water, and I'll probably have to do it again in a few days.  In the course of changing everything, and remaking my rotifer stand, as it was getting wobbly, I alternately left the air out, the heater off, and other mess ups.  Many rotifers died.  However, this morning, they seem active and multiplying again!  They will recover.  I hope it's in time for the next big nest, tomorrow.
 
Also, while I was changing everything, I installed a valve at the bottom of the tub.  Now the procedure to remove rotifers is to brush and stir the contents of the tub, and drain off the rots into a plankton filter over the bucket.  Then replace the volume with clean water.  Currently I am keeping 8 gallons going.  Now that I'm not feeding 8 gallons of O. marina, I expect that the density of rotifers will increase, and I won't go broke feeding them.

You're feeding 40ml of RGC? What size cultures? I'm doing fine feeding ~3ml, twice a day, in each 4G culture.

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