Phyto...

[sunny d polyp]

I felt that there is some very useful information from JRC concerning Phyto production. I asked his permission to re post some of his answers to my questions, he agreed.
Thx JRC and here are some.

your on track with the motile verses non motile and there are a few things to bear in mind
here. For one florida aqua farms is a very reputable aquaculture supplier the cross contamination you are seeing is quite normal for what we call "non axenic cultures".

Most cultures for aquaculture are non axenic meaning they arent fully isolated and purified to ensure against cross contamination. Axenic cultures are cultures that have been isolated cell by cell and treated with multiple procedures to ensure that you only get one type of cell. The procedures and time involved greatly increase the cost of these cultures when compared to non axenic types. Typically axenic cultures are more for the researcher and are wasted effort for most hobbyists as cross contamination will most likely occur at some stage of the culture process.

As far as FAF goes, typically they do a real good job given the low cost of their cultures and I've only run into a few instances where cross contamination (like you are seeing now) has occurred, in none of the instances was it a problem and your probably going to end up with a good viable culture in the end anyhow. I wouldnt worry about it too much unless you have a real yearn for single species isolation.

In any event FAF will provide axenic cultures upon request, processing time is typically rather long (20-40 day wait I beleive) but they will deliver. When doing research I prefer to order my cultures from an algae research bank such as the university of texas UTEX - The Culture Collection of Algae it's much cheaper than other sources but you've got to be pretty well in touch with exactly the strain you want (as they house algae for DNA studies and the like). Often when receiving cultures from UTEX there are long wait times as they have to prepare the culture and often from a cryogenic source.

hope that long winded spiel helps. in the long run your probably in great shape, eventually the lower species count will give way and while you may have a small amount of cells in each culture it wont be enough to even be considered a marginal contamination. It's zooplankton such as cilates and the like that we're most worried about for our cultures as they will often wipe entire species runs. If your truly concerned you can give the hoffs a call over there at FAF and they'll most likely ship you out a replacement culture.

[sunny d polyp]

she is correct, raising salinity will keep a lot of species from taking hold. Tet acclimates pretty easy to higher salinities so you should be fine if you do it in 2 to 3 steps. Be aware that you've got to balance the salinity of the cultures that are fed the algae!

[sunny d polyp]

she is correct, raising salinity will keep a lot of species from taking hold. Tet acclimates pretty easy to higher salinities so you should be fine if you do it in 2 to 3 steps. Be aware that you've got to balance the salinity of the cultures that are fed the algae!

I am assuming my rotifers will not tolerate consuming the phyto at a higher salinity? Or do you mean when adding the higher salinity culture water will raise the sg of the rotifer cultures themselves?
Also, I have 2.50/gals of rotifers, believe or not I am going through 1/gal of phyto feeding the rotifers during the 7 days before I split the phyto colonies (twice a day).
Another change in my routine is FAF's told me to add approx. 2/3 ml's of Algae Grow per 500ml's of culture water when beginning a new strain of phyto. I am now adding 16/18 ml's of AG (F-2) to my culture water I use to split them at 7 days.

[sunny d polyp]

he he the rotifers are most likely your source of crash on the other cultures! Bear in mind they will lay resting cysts that will survive pretty much anything (dessication, bleaching, etc). Best bet would be to replace the culture water vessel. I use old IO salt buckets, you can pick up a 1/4 inch plastic petcock with threading on one end and a john guest on the other and a small length of the plastic tubing for next to nothing at home depot and just drill a hole at the bottom front of the bucket and screw it in. Seal it with a touch of silicone then stack the bucket on another one (or a table or whatever) and you have a nice little dispenser for your culture water. You can then keep the sealed lid on nice and tight, refill from another bucket or carboy where you mix your salt and it works out pretty good. I typically toss the bucket and replace after a few months (as I'm going through the salt anyhow).

As far as the Book at FAF, it's become one of my bibles and has really lent itself to a lot of the knowledge I've learned now. Frank Hoff is known for making some really serious strides in rotifer culturing for commercial aquaculture and the book really is full of a ton of info. I still use it for reference constantly and am actually planning on buying a replacement sometime this year as the one I have is now completely beat and ragged! If you've got the money to spend, it's an amazing resource and it'll prove itself for not only bathroom reading but constant quick reference.

[THEJRC]

ah he he, you pushed it to the thread for all to see, awesome!

The trick with salnity changes is that rotifers will suffer from shock if a sudden salinity change (around 7 points) occurrs so you have to bear that in mind when feeding your cultures and using your cultures. obviously over time with splits and feeding your cultures will begin to rise in salinity. this can be offset by adding RO water when you perform your splits. I tend to keep almost all of my cultures at 1.022 as I have a large variance of species and it's easier to standardize on an acceptable salinity for all rather than have 20 different vats of culture media.

The changes in F strength are on par as well and will help the cultures start off. I'd highly recommend (for those seriously interested) taking a glance at the book by Thomas Kiorboe "A mechanistic Approach to Plankton Ecology" as it goes into great detail (mathematically no less) the mechanics of feeding cycles for microplankters. The theory is that there is a definate point where cells will become less active and slow growth when food is kept at elevated levels for too long. This would be the high end of the upswing (verses cells shutting down when feed levels are too low).

I havent delved into this one fully yet but after reading the theories I did cut back F strength on a few of my older cultures and received a growth spurt response, whether or not this is repeatable is another story and another experiment for another time.... Regardless, the book is one heck of a read!

For those interested ISBN 978-0-691-13422-2

[sunny d polyp]

Yes JRC I did bring your responses for anyone whom is interested to read, unfortunately it appears like it's my posts and I apologize for that.
I've made my culture water for the Tetraselmis @ 1.030, I do my splits on Thursday nights which is my 7 day routine. I believe I'll take your advice and keep all of my critters (roti's, tetraselmis, nanoc. and T.Iso) @ an even 1.022 which like you said will make it simpler in the long run.
I have also increased the f-2 formula for my splits to 16ml per/gal and have noticed they green up in just a few days and seem to stay at the same density until I split them on the 7th day. I'm hoping the last 4 days my cultures are consuming all of the extra ingredients. One observation is my chaeto in my 2 refuges seem to be taking off which I'm sure is an indicator that not all of the f-2 is being utilized. I will keep an eye on the tank parameters and post my results in a couple of weeks.
Right now I have 3/gals of phyto brewing, my roti's (2.5/gals) are using 1/gal per week, I remove around 4K to 5K ml (1/gal) of rotifers per day around 30/40% partials from the roti containers which seems to work fine so far. One thing I need to ask, is there a such thing as adding too many roti's? That may seem to be a silly question but I'm curious, I assume most end up in my fuges, the fish and corals consume alot of them, can you ever get an elevated population of roti's that live and mutiply inside of the aquarium?
Sorry for the long post but I am enjoying raising green water and little small specks as much as my tank itself. I may need to be committed to plankton anonymous (PA) for an undisclosed amount of time.
Again thanks JRC for all the help and insight you bring to TalkingReef.

[THEJRC]

he he, plankton is an addiction (at least thats how it started for me). I really got a kick out of the copepods when I started and it's been a downhill roll since....

As far as too many rotifers that might well be a question posed to someone more keen on the tank environments. Natural attrition will inevitably kick in and the available food sources will kick a population balance over time so short answer from me would be "too many is not likely". The downside here is that through attrition any die off or excess is converted to nutrients which would require some export.

As far as F/2 slipping into tanks, it's one of the reasons I chose the Guilliards mixture verses a lot of others. The algaes and such will continue to feed off the nutrients regardless of where they are at so long as they can photosynthesize and the main question of concern of course is elevated levels of trace and such. your refugium macro is doing it's job and keeping a balance for you so long as you keep it pruned (lest it die out) and keep that close eye on the nutrients you'll be in good shape.

The interesting one I noticed when I started down this road is that I actually had a sharp decline in nutrients with most my systems. Granted I tint one system green with various cultures and the others were receiving very healthy portions as well. The cells would continue splitting and consuming nutrients in the new envirionment untill either the cell died off through attrition (or some other means) or nutrients were no longer obtainable. Even more interesting is the fact that despite my constant addition of algal culture to the system directly, my skimmers were pulling less (so long as cultures were fed immediately from pulling off the shelf).

Caveat to this (noting the last line there). If I refrigerated or waited too long on a culture (past split date where cells begin to decline) I would see a nutrient spike and the skimmers would begin to pull more skimmate. Easily read as live nutrient consuming cell population verses dead decaying nutrient creating gunk.

Granted individual results may vary, my livestock bio load when examined in traditional senses is extremely low and I have a core focus on planktonic organisms and filter feeders. so long as there is a predator for the introduced prey and encounter rates are good balance will be acheived!

[sunny d polyp]

JRC I went to your www.copepodgeek.com Home, and I've believe you managed to ignite another proverbial spark. I'm culturing plenty of rotifers now for personal use (more than enough) and now I would like to try copepods, Tigriopus J. or Talitridae genus Orchetia, which would you recommend? I like the fact that they consume detritus from plants (algaes)and carrion, I could use them in my fuges not to mention feeding my fish. For now I am going to convert my 1/gal plastic jug that I use for roti's (that will leave me 1.5/gal.s for the roti's) and use the gal. for the pods.
Hopefully I won't get myself in trouble because I copied the 2 handbooks on your site and have them for quick reference in my cultivating room.
Just a tid-bit for anyone that has not tried it before, T Iso. (galbana) I purchased 2 cultures from FloridaAquaFarms and had a lot of trouble removing them from the culturing material. I ended up dumping most of the gel with the galbana still attached. I thought that the culture was bad or may have froze and was useless. So I strained off all of the gel (53 micron sieve) with most of the phyto attached to it and hoped to salvage as many of live cells as possible. I do have a live culture but it is extremely sparse and would probably take forever to get to a usable product. I called FAF and was told that T. Iso is notorious for binding to the substrate, it almost permeates itself. She said I should have left it that way and because the gel really does no harm to the culture. Anyway, live and learn I guess, I just hope my scenario will help someone else in the future.
If I keep finding new strains to cultivate (there is probably only 10,000 more) I'm going to need a bigger boat.
Thanks once again JRC and TalkingReef for allowing me to ramble.

[THEJRC]

no worries on the copy from the site, it's mostly random plotting of what I've found out over time slapped up there to hopefully help others. I do need to do some updating as I've learned so much more since I plopped up the handbooks. One of these days when I get a good clean block of time maybe.

As far as the copepod culturing, thats what got me into this whole thing anyhow. for some reason my ADHD brain pushed me from "what is this white thing on the glass" to counting algae cells with a hemocytometer and attempting to create a repeatable protocol for raising mythraculus sculptus (emerald crab) zoea. such is the way my head works.... In any case...

the garage hack of a biologist in me says go for T. Orchetia as I havent attempted the species (nor have I run into anyone that has on any scale to mention). Luis A M over at mofib has thrown up some good protocols on acartia tonsa, and despite the fact that T. californicus is super available it's beleived that the size is too large for most breeders. It'd be nice to see more people trying other species outside of the norm for sure!!

[sunny d polyp]

Have not had a tremendous amount of time to much research on A.tonsa but the requirements to cultivate them does not seem to be much different than tigriopus californicus, the size is though. T.orchetia was a different story, I could not find squat for referencing this copepod, no wonder not very many hobbyists are attempting it. As far as the tigriopus C. goes, you stated the size was a concern for larval fish. as of now I've not attempted breeding fish (at least not yet God help my wife) I just want another natural food source for my fish and inverts. There are a ton of them I've found, I am going small scale on everything like phyto and roti's, so will I with either tiger or A.tonsa. The scale of my phyto is a total of 4/gals soon to be 5/gals when the T.Iso gets cranking (optimism at it's best). You have been on track with all information in the past maybe if you could direct me as to where to find more info on T.orchetia I will research that strain further.
Your site although you said you have not up-dated it lately has a tremendous amount of information, thanks for not turning me in for copy right infringements. :-)

[THEJRC]

I'll see what I can find on the species, you may want to hit up the mofib and dibs folks as well! Theres a lot of resources out there such as FAF, Dr. Rhodes, and the Reeds that we can all draw on, it's one of the things I really love about the reef community!!

As far as copyright, nothing on the copepodgeek site is, and hopefully I'll never have to. Anything I do in this hobby deserves to be public domain for sure. I've gained a ton of knowledge from a ton of people out there who all have been more than gracious to inform me without fail. It's a beautiful thing!! It's also one of the reasons why I highly recommend people such as Reed Mariculture, Florida Aqua Farms, and Ocean Pods (read Adelaide Rhodes in my book) as they all despite having commercial stake will share information for the sake of the hobby and the sciences progression! I'll fire off a few emails to some copepodologists and see what I can come up with but the sad (but not so sad) thing is that with the extreme number of species out there direct species examination tends to be pointed for only certain reasons (such as pollutant examination with freshwater species, etc.). Sadly this makes it hard for us to find information, the great part is that it allows those of us who are slightly more than interested hobbyists and much less than marine biologists and copepodologists the chance to find out something new for mankind.

If you can easily obtain the species I personally would be greatly interested in getting a sample (at my cost of course). This is what makes me tick with the plankton, the easiest way to describe it is:

"we know more about space than our oceans and it's food webs, if a J***A** (read goofball) like me can find something someone else can use, I think it's amazing"

I love to see anyone interested for sure (I hope it shows) and I hope to see many more smarter folks than me get into it on a more accessible level. Much like any other field of open research funding and direction often stifle true findings and innovation, the same has proven for the oceanic and planktonic studies. I'd love to see more "garage mechanics" bounding around the traffic cones! If my information has helped you I'm happy for that, just do me a favor and let me know when you prove me wrong or find something new, that's what excites me!

[sunny d polyp]

I found their site last night but did not get a chance to research the forums much. I did call FAF and she couldn't tell me much about them either, so it's on to get some info on them.

I contacted Adelaide Rhodes last night and she sent me another link for purchasing tigriopus C., since I'm extremely impatient ( patience is not a virtue of mine) I may try out Tiger pods for now. I do notice when I'm researching species that many of the links for pods are from university studies, extreme cultures like phenominally high PH or salinity, to see just how much strictnine they can take before they meet their maker. That's why forums that are written from real people whom want to grow their cultures and not send them to thier makers are so valuable.

It seems to me that most of what we rear (roti's and bigger pods) are not very nutritious until we gut load them, with the exception of artemedia for the first 8 or so hrs. I don't look at them as a small package of what ever we feed them combined into one, I look on it as a challenge, like obviously you do. I have no doubt noticed a remarkable transition when it comes my tank, it always was healthy but now my polyps are extended for longer periods of time, my fish are more active and have never seen the colors so vibrant. It's just the opposite of what you would think, I've been adding alot of nutrients thru natural feedings and absolutely no nitrates, phosphates, nothing bad. I'm sold on culturing my own food. I had a friend ask me how much work went into my phyto, roti's and artemedia, I told him I guess if I ever thought of it as work I wouldn't be doing it.

As far as proving you wrong, that will be the day, you have probably forgot more than I know at this point. I could maybe teach you about technical diving (cave NSS/CDS) but you may already know that. I'm learning alot from you and the entire reefing community, i will never learn it all.

[THEJRC]

Generally I'm not one for quoting unless it's needed but this is a great example of my own struggles I've got to do it!!!

I do notice when I'm researching species that many of the links for pods are from university studies, extreme cultures like phenominally high PH or salinity, to see just how much strictnine they can take before they meet their maker. That's why forums that are written from real people whom want to grow their cultures and not send them to thier makers are so valuable.

alright!!! you've found my problem number one!!!! problem number two is that most of the good papers out there are expensive and/or hard to get ahold of. You can spend time tracking down and writing to the great people who wrote the paper and obtain it for free... but time itself also must be considered into cost factors, especially for the hobbyist. It's frustrating for sure!! but when you do finally talk to a "ologist" with a good interest, it always pays off. Be persistent and dont lose faith!!

It seems to me that most of what we rear (roti's and bigger pods) are not very nutritious until we gut load them.

Ahyup, and if you pop over to the side of ornamental breeding there are people finding out that movement, size, and a whole manner of other factors come into play. Thats why it's important that those of us with a general interest in microplankters (read little itty bitty zooplankton) keep at it. The guys on the other side of the field studying the fish larvae are praying for us to find efficient and available ways to breed food sources that are more viable and useful!!

As far as proving you wrong, that will be the day, you have probably forgot more than I know at this point. I could maybe teach you about technical diving (cave NSS/CDS) but you may already know that. I'm learning alot from you and the entire reefing community, i will never learn it all.

Nah, you'll prove me wrong three times this week most likely (I hope so!). I've already done it myself a few times in the last few months. The beauty about this still being a serious hobby for me is that I'm in it for the joy of learning and tinkering. The day I cant find out something new is the day I need to quit and do something else. That day will never come here! I've learned so many new things from everyone I've talked to and each little tidbit sends me off spinning in a different direction. Things from Jake Adams and his flow examination, to people like Adelaide Rhodes in the trenches figuring out how to breed viable species in dixie cups... It's invigorating for me to be in the thick of it as the little guy watching and listening and playing around. To be brutally honest I can spell isaeidae all day long but despite the fact that I've been studying the amphipod's predation habits for months now I'm still not quite sure how to pronounce the name (perhaps someone will pick up on that and answer the question he he).

As I sit here now typing this post with two M. sculptus (emerald crabs) blowing spawn not more than a foot away from me I cant help but wonder how many people are out there that know a tidbit or three that the community can use but wont mention it out of fear of being proven wrong at one point or another. (FYI on that note, I'm actually in the process of trying to prove my own spawning induction theories surrounding this species from a few months ago wrong with another test...)

One of these days I'll stop tinkering with spawning induction and actually focus on rearing the zoea, but for now it's a serious attempt at spawning induction and only a few feeble attempts at larval rearing. With real life (read "work") I've got to pick my battles, just as those who do this for a living must balance interest and goodwill of themselves with interest and goodwill of those that fund them!

[sunny d polyp]

Quote away JRC, makes me know someone is listening to my ramblings hehe.

Hopefully she won't be upset with me but I'm am going to try and pick Dr. Adelaide brain as much as I can. I asked a few questions on her site about some of the harder to find species like A.Tonsia and Tisbe and she actually replied to my questions. I know she is probably extremely busy with all she does, but I'm going to try.

What I have been doing is taking 1/3 of my roti cultures (3/2 liter reactors) I purchased from FAF, they sell them as brine shrimp hatchers and work perfect for my needs. I remove 1/3 from each reactor daily which is about 660ml'sX3, this gives me plenty of copes to add to my 46/Bow reef tank. This not only culls my brood but also keeps the culture water relatively clean, they are fed every day (twice a day) with a mixture of T.Iso, Tet and Ncp) on the third day I (the second feeding for that day) put 1/2 ml of selcon to each reactor. This has worked well for me (has not been more that a couple of months) so far. I wonder if Brachionus Plicitilis may be the magic bullet? My analogy would be the preference for nutrient export, some like calipera and some use Chaeto and ect. The roti's in themselves don't really have very much nutrition right? The roti's are gut loaded with nutrition for export to what we are feeding. They are very easy to culture and multiply like like crazy, because of my anal retentiveness (not certifiable yet but close) I have to create a challenge and different species of critters seems to fill the void.

I am finding it near impossible not to contaminate my three species of phyto, I'm now sieving (53micron) all of my cultures when I split them. I have been using the same 53 micro sieve with all three ( I wash the sieve carefully with vinegar after I split each species). I believe this is where most of my contamination has take place. I now purchased separate sieves for the three phytos, roti's and my culture water before I split (roti's get every where!) I shake the cultures of phyto twice a day and have motile and non-motile species mixed in. It's like you stated "it's hard to keep them separated", but I believe being as careful as possible you can at least prolong the inevitable.

Also whom ever might be interested, I have started filling plastic ice trays half way with phyto and letting them freeze for a few hours, then I remove them from the freezer and put live artemedia (about 10 to 15 thousand in each ice compartment) then 1/2 ml of selcon on top of the artemedia (each compartment) and top the tray off with phyto. In my 46/gal reef (which is heavily stocked) I thaw out 3 cubes every 3 days, turn off the sump returns and let the solution mingle for a couple of hours. I do this when just the actinics are on (morning) because I get a much better reaction from my corals. Please keep in mind this works for me, I run carbon 24/7 and partials (10/20%) every weekend. I also monitor the levels in my tank minimum once a week. So please use common sense and before you try this routinely, make sure this will work for your set-up also.

JRC, thank you once again for taking the time to write what you know and explain in such a way that we can easily understand the information.

[THEJRC]

I wonder if Brachionus Plicitilis may be the magic bullet? My analogy would be the preference for nutrient export, some like calipera and some use Chaeto and ect. The roti's in themselves don't really have very much nutrition right?

quite possibly, any new species examined is a good thing!!! As far as the roti's easiest way to think of them is tiny little bags. We fill those bags with food (phyto, selcon, etc) and use the bags as a transport to get the food into our target livestock.

I am finding it near impossible not to contaminate my three species of phyto, I'm now sieving (53micron) all of my cultures when I split them. I have been using the same 53 micro sieve with all three ( I wash the sieve carefully with vinegar after I split each species). I believe this is where most of my contamination has take place. I now purchased separate sieves for the three phytos, roti's and my culture water before I split (roti's get every where!) I shake the cultures of phyto twice a day and have motile and non-motile species mixed in. It's like you stated "it's hard to keep them separated", but I believe being as careful as possible you can at least prolong the inevitable.

I used to seive my splits but now tend to split a touch differently as sieving invites all kinds of trouble. I now use multiple funnels and during my splits I only utilize the first portion of each culture for split, the rest is then shaken up and sieved before being used for feeding this way you reduce contamination possibility on your seed cultures and the harvest doesnt really matter as much as I'm not doing any crazy axenic research, it all goes to the same place... the guts of zooplankters and polyps of corals.

As far as cleaning equipment, I spent the few dollars with FAF to get some alconox soap, the stuff is a godsend. I keep my rigid airline tubing in a 64 ounce container filled with water and about a teaspoon of the soap and I do the same with my other equipment (funnels, sieves, etc) keeping that in a shallow tub with water and the soap with lid on tight. Prior to use I run through a cleaning regimen where I dump the containers, get the water as hot as possible and rinse everything. I then refill the containers and put what I'm not going to use back in, dip what I am going to use and follow the dip with a cold water rinse, I then set it out and avoid use till it's dry. This is a hack of a protocol at best but it's worked pretty well. Given the cheap cost of litre bottles of water (which is what I use) I toss my culture vessels when they get too cruddy to clean within just a few minutes with a toilet brush, or I toss em after about 3-4 runs of use. The rigid tubing gets tossed every few months and replaced as well. If I start noticing a lot of cultures going south or growth impeded I'll typically do a "refresher run" where I strip out and toss all airline hosing, rigid tubing, vessels, and replace my air filters. I also use opaque bottles intended for liquid medicine for my F/2, this bottle has a cap that allows for removal of solution with a syringe only and I'll replace the syringe every few runs.