What am I doing wrong? (phyto culture)

[Zante]

I have a phyto culture that is going real well.
4 bottles, 3 litres each. each bottle has a different strain (nannochloropsis, tetraselmis, isochrisys and chlorella).

I split it weekly and it is going, as I said, real well.

My usage of phyto has recently gone up because I added a rotifer culture and because I trade left-over phyto with the reef club members, so I decided to take some of the phyto from the split (already mixed) and reculture it once in 4 bottles, 2 litres each.

I know that if phyto is cultured mixed eventually the dominant strain will take over, but in this case I am reculturing the mix only once, and every time I use a fresh mix from the separate cultures.

It has been going well for three or four times, but now every time the reculture bottles just crash. I haven't had a single bottle of recultured mix in four weeks. I do see them getting darker for a short while, then, one after the other they just crash.
The other cultures, the separate ones, are doing just fine.

What am I doing wrong?

Now, here are some details:
all of the water I use is sterilised with sodium hypocloride 15% solution 1ml per litre and neutralised with a solution of sodium thiosulfate (25g in 500ml of water) 3ml per litre.
The air going into the bottles is filtered by going through an airstone and bubbling in 4 inches of water.

The main cultures are near the window, but not on the windowsill and they are lit by 4 x T5 tubes put horizontally. It is basically a 2ft aquarium fixture propped up.

The recultures are on the windowsill and lit by the sun only. They are also close to a window that is always cracked open.

On all eight bottles the caps are airtight and so are the holes for the air to come into the bottle. The hole for the exit of the air is small (diameter of an airline) so there is a pressure of air going out keeping pollutants or contaminants from falling in

In all cultures I use F/2 but I use a double dose in the main cultures and the recommended dose for the recultures.

If you need any other details please do ask.

 

[billsreef]

Considering the time of the year, those cultures in the windowsill might be seeing temperature swings that they can't handle.

For the air filtering, you'd be better off with a length of PVC pipe stuffed with floss that gets changed periodically. The water trap your using now could get contaminated too easily.

 

[Zante]

Considering the time of the year, those cultures in the windowsill might be seeing temperature swings that they can't handle.

Tomorrow (split day) I was planning to keep them at the same level as the other bottles, to eliminate one difference between the cultures. We'll see if that was the case. It also means I will have to get out the bigger light fixture <grumble>.

For the air filtering, you'd be better off with a length of PVC pipe stuffed with floss that gets changed periodically. The water trap your using now could get contaminated too easily.

Unfortunately that would be impractical for me. What I do though is lace the water with the sodium hypocloride once a week so that any live stuff will be regularly sterilized. Also it is freshwater, which is likely to help with some of possible contaminants.

I am aware it's not a perfect solution, it's the best one I can go for, and I accept the fact that I will have to restart the cultures once a year, and that is good enough for my personal use.

 

[Aqualund]

I've had the same issues as you for the first 6 months of trying to perfect the art of stable and regular cultures. I'm not sure where you are going wrong, but what I can do is speak to what I am doing which is very similar to you except for maybe a couple minor things.

But Bill brings up a good point...the temp might be the only issue.

so here's what i do:

1. Each "simply orange juice" container is cleaned and sterilized in 15/15/70 calcium hypochlorite/Muratic acid/water for 24 hours prior to culture.

2. I empty the acid/bleach solution and rinse with saltwater.

3. Fill the bottle 60% with 1.021 RO saltwater (sterile).

4. Add 5 ml of a 50/50 calcium hypochlorate/water mix to each bottle. Test solution to ensure a good reading of killing levels.

5. let sit overnight.

6. add sodium thiosulfate (like 1 or 2 granules) to solution and mix.

7. Add the previous culture inoculate to the water.

8. All of my air tubes connect to a glass tube for inside the culture container. when not in use, these glass tubes sit in a 10/10/80 solution of bleach/muratic acid/water.

8. add the f/2 fertilizer. cap the bottle and place in the light.


My airline tubes go through a carbon filter...just basically a tube with 1/4" end on both sides stuffed with cotton balls and pelleted activated carbon.

This method has provided me with about 10 gallons of plankton a week for the better part of 8 months now without a crash. All of my errors in the beginning were attributed to improper sterilization methods, and assuming the water was ok when it wasnt (creeping bleach levels...too much sodium thiosulfate etc)

Good luck and feel free to msg me if I can help you.

 

[billsreef]

You don't want to mix and acid and a base together. They'll neutralize each other Usual routine is to just bleach, rinse well, and follow with a final 10% HCL rinse. Clean and sterilized vessels can be stored dry with some parafilm or saran wrap to keep them clean

 

[Aqualund]

"Maximum disinfecting efficacy is achieved at pH 4-5, because essentially all the chlorine is present as HOCl which is two orders of magnitude more effective than OCl–."

http://oregonstate.edu/dept/larc/sites/default/files/pdf/chlorine-fact-sheet.pdf

just a random source. But I originally got my info from the plankton culture manual:

http://www.amazon.com/Plankton-Culture-Manual-Sixth-Edition/dp/0966296044

Basically, at above 7.4 PH, the chlorine is present as OCL- and HOCL...and as you go higher you get more proportions of OCL- which is less effective than HOCL.

True, mixing too much of these will create chlorine gas...but you want a lower PH in the solution to increase the efficacy of the chlorine. So, I just make sure I am aware and wear a mask, and there is nothing that lives.

I can imagine a situation for Zante where the % of bleach he's adding just isnt as effective as necessary because of a high PH in his water. Dropping that PH with acid will definitely ensure there is nothing getting by...but that's not really what I was suggesting anyways...but just a thought.

I've also seen my saltwater solutions that were one day "good" as far as chlorine levels go, and then the next day being completely neutralized and undetectable without the addition of anything else to the system. I've had to then re-add chlorine to get the neutralizing going again.

**Also, not adding the solution within the vessel...sterilizing and then neutralizing within the same vessel allows for too many opportunities of contamination from outside sources. Doing it all in the same container eliminates virtually all possibility for outward contamination.

 

[Aqualund]

Actually even further the stoichiometry shows not a neutralization but rather just a production of chlorine gas which is where the sterilization comes in.

CaClO + 2 HCl → H2O + CaCl + Cl2

 

[Zante]

I've had a complete crash, so I will have to start over.

I will order new culture starters when I get back from my holidays, in a week's time. In the meantime I'll move the futures from the window and only use artificial light. I am now convinced it was the unstable environment and thermal excursions that crashed the cultures, as they've been running for a few months without any problem.

I have managed to get my hands on some panels of LED grow lights which I'm itching to try.

 

[Zante]

Would cigarette filters work to filter the air?