# Dry ice/CO2 against wax moths



## Pondulinus

If your going to try this I would assume that its a good idea to leave a small hole at the top (if your placing dry ice on bottom) so the air gets displaced by the heavier/colder co2 more easily. The excess gas needs to exit somewhere anyway. If not i would place the dry ice on top. My only concern would be that the dry ice might sublimate to quickly to suffocate the larvae as you cant make the stack 100%gas-thight and the internal volume of the stack is large. 
How much would you use pr. Stack? And how large stacks?


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## shinbone

The stacks are about 9 medium supers tall. Dry ice goes in an empty super at the top. The seals are just heavy duty adhesive tape, so the stack is not air tight and the CO2 slowly leaks out everywhere. If the seals are good enough, and you put enough cakes of dry ice in the stack, enough CO2 will be produced quick enough and for long enough that the interior atmosphere will be saturated with CO2 for a time sufficient to kill the wax moths. That's the theory, at least, and I am not sure how many pounds of dry ice would be required.


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## Nabber86

I am wondering if the CO2 will actually kill the eggs.


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## Rader Sidetrack

Per this study, wax moth _eggs_ can be killed by appropriate levels of CO[SUB]2[/SUB], but it may take a number of _days _of CO[SUB]2[/SUB] exposure to do the job.

http://spiru.cgahr.ksu.edu/proj/iwcspp/pdf2/7/692.pdf


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## LeifLiberty

It will be interesting to see your results. 

I plan to use Acetic Acid. I think it will be less expensive and it has an odor so it can be another reminder that the stack has been treated. Would not want to break down a stack and get asphyxiated.


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## Huck

If the honey supers have never had brood in them, from my experience, the wax moths will not bother them. I have stacked mine for years without a problem.
Huck


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## GoodyFarms

shinbone said:


> The stacks are about 9 medium supers tall. That's the theory, at least, and I am not sure how many pounds of dry ice would be required.



If you want to get all scientific....

9 medium supers = 30 x 9 = 270L

Say you want 5x the volume to account for leaks and inefficiencies = 5 x 270 = 1350L CO2

I know it's not, but assume it's an ideal gas...

PV=nRT

P= 1atm
V=1350L
R= constant
T= 75F

Solve for n....

55.4 moles of CO2, which has a molecular weight of 44 g/mol.

*So you'll need ~2.4 kg (5.4 pounds) of dry ice.*

If you only want 1x, divide by 5 = ~1.1 lbs.


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## Phoebee

I have been playing around with CO2 as a wax moth fumigation method.

A group in Israel has a paper out on the lethality of CO2 to wax moths. Their results were similar to others who have used CO2 for insect pest fumigation. The basic numbers are, 4 hours at 100% is lethal to all stages of wax moth, eggs to adults, and 80% for 4 days is also lethal.

I can tell you from my trials that 95% for a few hours is NOT lethal to these tough little bugs. Somewhere around 50% they start crawling out of cells and dropping to the bottom of the container. By maybe 90% they are not moving. But come back a week later, and if the concentration did not stay up long enough, the rascals are still alive and moving. It is turning out to be a bear to make a container that will hold 100% for long enough to kill them unless a slow purge is operating. Dry ice ought to allow that slow purge if you use enough. Sealing up really, really well is needed. Expect a sloppy job to lose concentration fast.

I have been using bottled CO2 and I bought a CO2 meter development kit module from COZIR in order to verify concentration. My goal is to come up with methods that assure a kill without requiring the expensive meter. I have not tried it with dry ice yet, but that's on the short list of jobs to do.

Look up the "room purge equation". That predicts how concentration will build up over time as you inject a known volume of a gas to a closed space with a small exhaust vent, knowing that mixing is occurring. Any mathematician will tell you that you can never actually get to 100% concentration. Any engineer knows that after about 5 volumes of the closed space, you're close enough to make a good engineering approximation. There are off-color jokes illustrating this.

I believe that using dry ice, you should put in enough, by weight, to fill the treated volume 4-5 times (calculations already done above, I see). I would put in some of the dry ice broken up into gravel-sized pieces but at least one big chunk that will last for hours. That should give a faster initial purge but then keep the concentration up for long enough to assure a kill.

My thread on this topic: http://www.beesource.com/forums/sho...-Wax-Moth-Fumigation&highlight=CO2+fumigation

Paper: https://www.researchgate.net/public..._carbon_dioxide_enriched_modified_atmospheres


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## sakhoney

I gas my 20 foot containers 1 time a month with a bottle of CO2 - it works - but you need to check monthly - not just once.
NO2 will also work.


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## GoodyFarms

sakhoney said:


> NO2 will also work.



Holy cow I hope you mean N2O not NO2! NO2 is definitely not for the novice unless you like acid eating your lungs.


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## sakhoney

N2 - the O is for old age


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## beepro

How does acetic acid work? I mean how do you apply them directly on
the drawn comb?


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## Bob J

beepro said:


> How does acetic acid work? I mean how do you apply them directly on
> the drawn comb?


Here is an excellent summary of treatments including acetic acid....

wax_moth


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## Nabber86

Rader Sidetrack said:


> Per this study, wax moth _eggs_ can be killed by appropriate levels of CO[SUB]2[/SUB], but it may take a number of _days _of CO[SUB]2[/SUB] exposure to do the job.
> 
> http://spiru.cgahr.ksu.edu/proj/iwcspp/pdf2/7/692.pdf




Keeping the necessary CO2 concentration for several days would be problematic, if not infeasible for most people.


Edit: Unless you have a setup like Sakhoney.


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## Phoebee

The papers I've seen, and my own results, say 100% for 4 hours will kill the eggs. I'd say to assume it is really 98% and hold for more hours. Maintain the concentration with a slow purge, which sublimating dry ice may do naturally.

80% for 4 days, without a constant purge, is the one that is problematic. I'm still trying to come up with a sealing method that does it without breaking the bank.

You will please notice that the OP is in Colorado. I think it is worth a caution that 100% "concentration" of CO2 at a 1 mile elevation is not the same amount of gas as it is at sea level. My CO2 meter, if not recalibrated for the lower air density, would say the concentration is significantly lower than 100%. CO2 does not work entirely by asphyxiation (displacing oxygen) but also by chemical changes such as turning water mildly acid, and upsetting the O2 control feedback in living things. In humans, CO2 is considered toxic at remarkably low concentrations. If you get a very short wiff of the 100% gas (I'd not recommend more), you instantly taste the sourness. 

So I'd recommend anyone doing this at high altitude to stretch the fumigation times beyond the recommendations of sea level studies. Either that, or watch the treated material and be ready to do a second and longer shot.


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## beepro

Why not just stack the frames inside a chest freezer for a
few days? Not sure how expensive is the CO2 here. But a freezer is
reusable after the first batch got packed away.


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## sakhoney

A large 244 cylinder of CO2 - $30 bucks - A chest freezer to hold what a 20 foot sea can will ???? Guess I could look into a refrigerated sea can


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## Phoebee

I addressed CO2 versus ice chests in my original thread. http://www.beesource.com/forums/showthread.php?331130-Dry-ice-CO2-against-wax-moths

For what I'm doing, I would be well ahead investing $150 in a small chest freezer. I've got that much in the CO2 tank and regulator, more than that in the meter. But a freezer of that cost is about 5 cubic feet. I know because we own one. A freezer gives a quick kill, no chemicals, and may well be most cost effective.

Downside: most of us with a chest freezer have it chock full of frozen food, with no room for frames, particularly frames that are going to drop larvae. The fact that our freezer is 130 miles from the apiary and full of food is what prompted me to try CO2. Commercial operators may be needing a huge walk-in freezer that ain't cheap.

But for people with a ready source of CO2, either dry ice or bottled CO2 for another purpose, there may be next to no equipment to buy. This assumes I, or some other tinkerer, can come up with a method that kills these things dependably without a $225 CO2 meter (and that's the cheap option). Among the people with CO2 on hand, one guy had a huge tank for refilling paint ball guns. Others who were interested could pick up dry ice cheap on short notice.

Other options include a specific subspecies and brand of Bacillus thuringensis, and paradichlorobenzene. Both have their limitations but they have been used for the purpose.


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## sakhoney

Oh contraire Phoebee - On my sea cans - I just hook up to the valve on the CO2 tank - with the proper nipple that fits the tank - and with a small length of copper tubing - I have a 3/8 hole drilled into the side of the sea can at the back. Crack the valve and let the gas flow - don't need a regulator. Going to dump in the whole tank anyway


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## Phoebee

sakhoney said:


> Oh contraire Phoebee - On my sea cans - I just hook up to the valve on the CO2 tank - with the proper nipple that fits the tank - and with a small length of copper tubing - I have a 3/8 hole drilled into the side of the sea can at the back. Crack the valve and let the gas flow - don't need a regulator. Going to dump in the whole tank anyway


Oughta work then. I would recommend a quick initial purge but save enough to keep a slow purge going for a few hours. I'm still hoping to get around that slow purge but I have not managed to seal a container well enough to quite get away with it. That includes the Ziploc sealed storage containers I'm using now ... the gasket is open cell foam and leaks out the initial purge too fast to effect the quick kill unless I keep up a little flow.

Liquid CO2 in the tank gets wicked cold when you're letting it out. You may be able to see the level by a frost line on the tank.

For that matter, you may manage to freeze the contents of the can with that liberal a dose of CO2, delivered that fast. Which would work, too.


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## sakhoney

I'm talking a 20 foot sea can - a freight container


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## Phoebee

sakhoney said:


> I'm talking a 20 foot sea can - a freight container


Yeah, I got that. Looked it up to be sure it is what I call a shipping container. What are those, about 2000 cubic ft?

Looks like you have enough of an operation that the 5 cubic foot freezer is a joke, and a very good scale to make CO2 an interesting option.


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## sakhoney

20 foot 1280 CF
40 foot 2560 CF
A 20 will hold 790 mediums - and close to 400 deeps
a 40 - double the numbers


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## Phoebee

I could stick our entire apiary in one corner and still park my truck in there.


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## beepro

At this moment I'm thinking to turn my little outside porch 7x8x14' into
a sealed space to hold the frames inside. Seal the outside with the 2" thick
insulation house foam boards and then the LP or OSB boards added. Inside line it up with
the 6 or 10 mil plastic sheets and duct tape them up. Now you have a roomy area for 
your CO2 method.
How about the mylar line inside weed growing tent on ebay? You can order your dimension
too. Different size for you to choose. The bigger the more expensive.


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## Phoebee

I would *NOT* do this with a porch attached to my home. CO2 is toxic. A few percent will kill you.

You might use a shed, well away from any living spaces.

I deny any knowledge of weed growing tents. That was an unfounded rumor. Never touched the stuff. I do know that some people use elevated CO2 levels to enhance plant growth.


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## FlowerPlanter

Does CO2 kills beetles just the same?


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## Phoebee

FlowerPlanter said:


> Does CO2 kills beetles just the same?


At the concentrations needed to take out wax moth eggs and larvae, CO2 kills everything that uses oxygen. 

Maybe not Tardigrades. Those are something of a special case, but they're not bee pests. https://en.wikipedia.org/wiki/Tardigrade


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## LeifLiberty

beepro said:


> How does acetic acid work? I mean how do you apply them directly on
> the drawn comb?


http://www.blackburnbeekeepers.com/Acetic_Acid_Fumigation.pdf


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## Phoebee

FlowerPlanter said:


> Does CO2 kills beetles just the same?


I just gassed the frames from a robbed out and abandoned hive this morning. About a dozen small hive beetles fell out of the frames.


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## beepro

O2 does the same as CO2.
How come nobody use the O2?
Is it more combustible that is why?


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## sakhoney

ever see the results of using any oil product on a O2 regulator. It will self ignite and blow up - I would not want to use it on all that wax/frame products just looking for a spark


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## Phoebee

beepro said:


> O2 does the same as CO2.
> How come nobody use the O2?
> Is it more combustible that is why?


O2 is the oxidizer in welding torches and liquid fueled rocket.

CO2 is used in fire extinguishers. 

A few minor differences.


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## beepro

Yep, more danger with the O2 I guess.
Both will come out cold though.


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## Phoebee

Liquid nitrogen is colder than either and cheaper than liquid O2.

But CO2 is the only one you can bring home in an ice chest, weigh out a chunk on a postal scale, and toss into a box with frames.


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## EastSideBuzz

I got a trailer full of Moth issues. Any ideas on how to kill them in mass without having to freeze the whole trailer? It is not going to be cold enough for a month or two. Way to many boxes to put in a small freezer. Ideas? Will the Liquid 02 idea work?


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## Phoebee

EastSideBuzz said:


> I got a trailer full of Moth issues. Any ideas on how to kill them in mass without having to freeze the whole trailer? It is not going to be cold enough for a month or two. Way to many boxes to put in a small freezer. Ideas? Will the Liquid 02 idea work?


Under no circumstances should you use liquid O2 (oxygen). The resulting intense fire or explosion probably would kill the moths, leaving a barren scorch mark where the trailer once stood, but the paperwork you'll have to do with the fire department, and the resulting tongue-lashing, probably would be enough to make you never try again.

The thought with CO2 (carbon dioxide) is that you don't have to freeze it (although if you use dry ice and it does get cold, that helps too).

How air tight can you make the trailer? If you can make it tight enough to hold the gas for a day or two, figure the volume of the trailer in cubic ft, multiply that by about 0.62, and you get the weight of CO2 in pounds needed to purge the trailer with 5x its volume in gas. That should produce almost 100% CO2. 

As an example, a trailer that is 12 ft long, 8' wide, and 8' high would be 768 cubic ft. 475 pounds of dry ice would purge produce a lethal level for the moths. And anything else in there, beekeepers included. So air it out before going back in. 

Yeah, 475 pounds is a lot of dry ice. Somebody check my math, but I think .62 pounds per cubic foot is close. That much bottled CO2 would also do it if you can get it cheaper.

The concentration needs to stay that high for hours, or better yet days. The effort is wasted if it is not really tight enough. 

I have not read the acetic acid trick above. It is probably worth the read.

A lot of people here would find the brand of Bta (a strain of Bacillus thurengensis) that is approved to kill wax moth larvae, Xen Tari.


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## EastSideBuzz

It is a 33 foot trailer. If it was road worthy I would find a company that had a big freezer and park it inside for a day or two.  I am looking to buy a freezer trailer so I can avoid this in the future. Almost bought one at auction a few months ago and gave up 100 bucks to soon in the bidding. Darn.

I dont think it is air tight enough for the 02 thing.


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## Phoebee

EastSideBuzz said:


> It is a 33 foot trailer. If it was road worthy I would find a company that had a big freezer and park it inside for a day or two.  I am looking to buy a freezer trailer so I can avoid this in the future. Almost bought one at auction a few months ago and gave up 100 bucks to soon in the bidding. Darn.
> 
> I dont think it is air tight enough for the 02 thing.


If you could get a good reefer trailer that is roadworthy and can get below freezing, possibly you could drive it around to other apiaries and freeze their frames? I wonder if there is a business in that. You probably would need other uses for it as well. What does it cost to rent one for a week?


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## EastSideBuzz

Phoebee said:


> If you could get a good reefer trailer that is roadworthy and can get below freezing, possibly you could drive it around to other apiaries and freeze their frames? I wonder if there is a business in that. You probably would need other uses for it as well. What does it cost to rent one for a week?


So my thought was to do it quarterly fill it with my boxes and then freeze them for a couple days quarterly. Letting others come in for a few days is a nice thing to do also.


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## LeifLiberty

I think for acetic acid you would need to remove the boxes from the trailer, else enough acetic acid to be effective would likely be enough to harm the trailer. Acetic acid would corrode the metal trailer body. . .


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## sakhoney

Eastside - your over thinking this method - Here's what I do for a 20 foot sea can
In the front of the sea can I put in a hole just big enough for a length of copper tubing or a 1/4 hose. (very small hole) Juts above the floor board. Connect this directly to a bottle of C02 I get from my local welding shop. I get a 251 cubic foot bottle (same size as a large oxygen bottle) While there - buy the nipple that fits the bottle of CO2. Connect the bottle to the hole you put in the covered trailer and just crack the valve releasing the CO2 into the trailer.
Your wanting to gas the trailer - not make it cold. People with smaller containers are getting dry ice - CO2 in solid form - to do smaller containers. Again it not about the cold - it about the gas.
Now remember before entering this trailer - you need to air it out as the CO2 will knock you on your butt as well.
Your displacing the oxygen in the trailer - something both you and the wax moth needs for breathing. 
I gas 1 time monthy


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## EastSideBuzz

sakhoney said:


> Eastside - your over thinking this method - Here's what I do for a 20 foot sea can
> ........ I get a 251 cubic foot bottle (same size as a large oxygen bottle) While there -


So if my trailer is 33ft I should 1/2 + your formula? 251.

So called a welding gas refilling place. They said 251 is called a 50lb tank and is 9 inches by 48 inches. Cost $35 bucks to refil/swap $14 a month to rent. The next size up is industrial so do I use 1.5 or do you think that the 50lb tank would do it.?


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## sakhoney

I would think that 50 would do


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## EastSideBuzz

So drill hole seal hose and empty tank. Wait <Fill in number here> then open air out and check for live worms.? Sounds too easy. How many days should I wait before checking?


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## sakhoney

Should kill everything within 24/48 hours. But if you air it out they will return. I know it works so I don't check. I leave the gas in there. Then on the first of the month I hit it again until spring. When I start taking the supers out


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## Phoebee

sakhoney said:


> I would think that 50 would do


I'd be curious to see your math. What weight of CO2 per cubic foot are you using to get an effective kill? 50 pounds for a sea can the size you're using is way lower than my numbers, although possibly workable if you can maintain the concentration for a month. 

I have seen wax moth larvae fall from comb at around 50% concentration, although they're not dead at that level and will recover if the concentration falls in a few hours. But if you could hold 50-60% long term, that level takes far less gas to achieve than the near 100% levels needed for a quick kill.


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## sakhoney

While I never ran any math on this I know what works. I also know if you don't air out before entering you will pass out. I had to drag one of my hands out of a container a few years back


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## Phoebee

sakhoney said:


> While I never ran any math on this I know what works. I also know if you don't air out before entering you will pass out. I had to drag one of my hands out of a container a few years back


If you lived near me I'd ask if I could check out your containers and try my CO2 sensor in one. Apparently you're managing to seal them up very effectively. If so, a tip of the Stetson to you as that is a big problem with the CO2 fumigation method, at least per my experience. One project that treated stored grain in silos commented that leaks amounting to a square cm in a large silo were enough to render the fumigation ineffective.

I have a test running in which I've modified my sealing gaskets to reduce permeability, but previous tests on my 60-quart containers have dropped concentration down to 10% or less after 100 hours. But sea cans may be tighter than I would guess, to keep sea air out of commercial cargo.


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## sakhoney

They are sold as wind and water tight. Just duct tape the 4 little vents in thr corners.


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## EastSideBuzz

Phoebee said:


> If you lived near me I'd ask if I could check out your containers and try my CO2 sensor in one.


Now that is an interesting idea. What sensor do you recommend?


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## Phoebee

EastSideBuzz said:


> Now that is an interesting idea. What sensor do you recommend?


I am using a COZIR CM-0121, a development kit module ranged for 0-100% concentration. They have the sensor itself for less than the dev kit, but the dev kit is a plug and play USB gizmo with software to get it running. They want $229 for the dev kit, and the sensor itself I think may be about half that if you like to tinker with computer gadgets.

The dev kit will run on Windows 7 or Windows XP. I'm using a cranky old laptop to run it.

Low concentration CO2 sensors are pretty common. Sensors for 100% are not. This is the cheapest option I've found. Most are over $1k.

http://www.co2meter.com/collections/co2-sensors/products/c20-sensor-development-kit


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## GoodyFarms

EastSideBuzz said:


> So if my trailer is 33ft I should 1/2 + your formula? 251.
> 
> So called a welding gas refilling place. They said 251 is called a 50lb tank and is 9 inches by 48 inches. Cost $35 bucks to refil/swap $14 a month to rent. The next size up is industrial so do I use 1.5 or do you think that the 50lb tank would do it.?


I don't know. Your box is ~1800 customer ft so a 250 cu ft tank will result in a CO2 concentration of 14%. It will decrease the O2 concentration by a minimal amount (from 21 to 18%). 

While not the same species, some papers suggest a higher concentration (~20%) for 30 days is needed to kill moths. Another study on coddling moths showed 20-30 days at 14% co2 is effective. 

I guess the take home is seal it well and don't open it for at least a month. 24-48 hours is probably way to short for effective kill rate at the concentration your using. 

https://www.usaemergencysupply.com/...-storage/indian-meal-moths-and-carbon-dioxide

http://hortsci.ashspublications.org...9gpt_ki_EtKBede7A&sig2=MDnlQMubxI9E7QP1UoKfPw


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## Rader Sidetrack

Praxair says a 50 lb cylinder of CO[SUB]2 [/SUB]equates to 436 cu.ft. ...
http://www.praxairdirect.com/PXItem...Id=11530&langId=-1&storeId=10152&itemID=66714

... so in an 1800 cu.ft. container that would be 24% CO[SUB]2[/SUB].

One way to get a higher percentage without having to rent an industrial size CO[SUB]2 [/SUB]tank would be to empty the 50 lb tank into the container, then go back for a refill on the 50 lb CO[SUB]2 [/SUB]tank.


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## GoodyFarms

That sounds about right for 50 pounds. Pvnrt at 75f is 445 cu ft. 

Slowly let it in from the bottom with an exit at the top.


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## sakhoney

Is it just me or is anybody taking into account that why you may be right on the numbers for a empty container this container is full of boxes. This cuts down on the cubic footage alot.


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## Phoebee

sakhoney said:


> Is it just me or is anybody taking into account that why you may be right on the numbers for a empty container this container is full of boxes. This cuts down on the cubic footage alot.


Of course it does, although, remember, these are boxes loaded with frames drawn with comb that the bees make with the absolute minimum amount of wax they can manage. The boxes are still largely empty. Any benefit you get just acts as insurance the method will work.

Balancing that, the effects of wood and comb on the CO2 concentration are unknown. If moisture is present, this will absorb some of the CO2. In fact, wax moth larvae also absorb CO2 (which is what kills them). So I would expect some drop in concentration not due to leaks.

Unless the container is unusually strong, or has an external bladder to handle the volume changes, any change in temperature or atmospheric pressure is going to affect the concentration. Heating or lowering outside pressure will force some gas out. Cooling or higher outside pressure will force clean air in. That's why the sea cans were designed with vents, and the plastic storage containers I am using were designed to not seal totally. This could be fixed by adding an external bladder made of an impermeable material.

There are a lot of unknowns in this. My experience so far is that the largest unknown is unsuspected leaks in the container. Sakhoney has struck upon a very good container, but not everyone will have the same result. If you start with a damaged container or one with a small flaw in the seals, this may not work at all.

Hence, my investment in the CO2 meter. 

My hope is for me to use the meter to come up with methods reliable enough that others don't need one. But once you are on the scale of a large cargo container, your investment is high enough, and the cost of gas is high enough, that a few hundred bucks for the meter is a much less significant cost, with the potential to save way more than it costs. One bad treatment in a large container could damage more comb than the meter costs, by a large margin.


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## EastSideBuzz

Phoebee said:


> I am using a COZIR CM-0121, a development kit module ranged for 0-100% concentration.
> http://www.co2meter.com/collections/co2-sensors/products/c20-sensor-development-kit


Just ordered one. I called right before I ordered and asked questions and they were so helpful. Gave me really good advice on the use and spec's. I ordered it online and within an hour I got shipping info. Amazingly quick turn around time. Cant wait to get it and start treating them in the container.


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## Phoebee

EastSideBuzz said:


> Just ordered one. I called right before I ordered and asked questions and they were so helpful. Gave me really good advice on the use and spec's. I ordered it online and within an hour I got shipping info. Amazingly quick turn around time. Cant wait to get it and start treating them in the container.


They gave me some assistance when I had trouble getting my old XP to connect. I needed to update a piece of the operating system on that antique, but it works fine now.

Their basic software for that unit is one function only, but they do have some other sensors, and they have a white paper out on tying sensors together using a pet snake. Which makes no sense until you read the joke in this link.

https://image-store.slidesharecdn.com/24cbdfd4-2efe-4dde-b164-b5a4435d1725-original.jpeg

If the idea of integrating this with something else, like maybe a WeatherBug station, sounds useful, Google "Python, Unicorns & Beer Steins".


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## EastSideBuzz

Phoebee said:


> If the idea of integrating this with something else, like maybe a WeatherBug station, sounds useful, Google "Python, Unicorns & Beer Steins".


I am going to run it on a Minnowboard off a car battery. http://www.adiengineering.com/products/minnowboard-turbot/ and then run it remotely over the internet.


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## Phoebee

I got up to our apiary and found a perfect 200 hour recording on my latest modification. Success!

I am currently using 60 quart Ziploc plastic storage containers, sold by Walmart. These have a gasket and six latches around the lid. They should seal up well, but every run up to now, they've fallen off to less than 10% over 100 hours (4 days is 96 hours). That means they can't meet the 4-day at 80% lethality criteria. These containers also would not hold measureable pressure, i.e. I could not measure any water column pressure at all on the exhaust vent line I had rigged. The problem is they used open cell foam on the gasket, apparently to KEEP it from holding any pressure at all, and making it allow the contents to breathe.

This time I took a tube of Permatex windshield silicone sealant, the really runny stuff, and doped the gasket. I also worked the silicone down the edges of the gasket. The result was that after starting out with about a 1-hour purge that got close to 100%, at 100 hours the concentration was just below 80% (should still be lethal since it started out way higher), and at 200 hours the concentration was 54%. Back pressure was a remarkably steady 15 mm H2O regardless of flow, suggesting the pressure lifts the lid or gasket a little. This should mean the container can still breathe a little, but retains gas much better than it did.

The moth larvae looked pretty profoundly dead.


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## Fusion_power

I used to make a toxic atmosphere by sealing a candle into a container. Wonder if you could put a candle in and let it burn until it goes out thereby reducing oxygen content to under 5%. The best I recall, 1 cubic ft of air will keep a candle burning about 3 to 5 minutes depending on how high the flame is burning. There are 7.5 gallons in a cubic foot so a 60 quart container would keep a candle burning less than 10 minutes. Use obvious precautions dealing with flame. CO2 sinks so put the candle near the top of the container to take out most of the oxygen.


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## Phoebee

I recall a lab experiment in which we burned a candle under an inverted test tube in a pan of water. It drew water up into the tube. I'd expect it would cause a partial vacuum and either collapse the container or cause it to draw fresh air in.


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## CowDoc1

Please note that it is primarily the lack of oxygen that kills the moths, larva and eggs. Using a gas that is heavier than oxygen (like CO2) encourages that gas to settle to the bottom, displacing the oxygen upward, and hopefully out of the container. Of course, there is some mixing. So you should add several times more CO2 volume than the container holds.

Once the oxygen is flushed, it doesn't matter what happens to the CO2. Technically, the CO2 isn't killing them. It's the lack of oxygen. If the CO2 were replaced by nitrogen or helium or any gas except oxygen, all of the bugs would still die. A complete vacuum would work just as well. The main goal is to keep the oxygen out.


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## shinbone

I cracked open one of my stacks of supers that I had CO2'ed in September, and pulled two supers off the top to put on hives. The extracted combs inside were moth free, as far as I could tell.

Cowdoc1 - I am no biologist, but I thought in addition to displacing O2, the high concentration of CO2 is toxic, too.

I am not sure if I can drawn much of a conclusion from this one apparent success because it is honey comb that I CO2'ed, and as Huck points out in Post #7, wax moth tend to leave honey comb alone.




.


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## Phoebee

CowDoc1 said:


> Please note that it is primarily the lack of oxygen that kills the moths, larva and eggs. Using a gas that is heavier than oxygen (like CO2) encourages that gas to settle to the bottom, displacing the oxygen upward, and hopefully out of the container. Of course, there is some mixing. So you should add several times more CO2 volume than the container holds.
> 
> Once the oxygen is flushed, it doesn't matter what happens to the CO2. Technically, the CO2 isn't killing them. It's the lack of oxygen. If the CO2 were replaced by nitrogen or helium or any gas except oxygen, all of the bugs would still die. A complete vacuum would work just as well. The main goal is to keep the oxygen out.


Check the MSDS on CO2. Technically, it is listed as toxic. That's more true of mammals than it is for insect larvae, but it is true for most animals that use oxygen. The underlying reasons are that CO2 lowers pH (which nitrogen gas does not), and triggers regulatory mechanisms in resipiration systems. 

Relatively low levels of CO2 are harmful to humans. Wax moth larvae are remarkably resistant, but levels around 60%, or even lower, will eventually do them in if maintained.


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## CowDoc1

Phoebee said:


> Check the MSDS on CO2.


Here's a typical carbon dioxide MSDS, per OSHA guidelines. 

https://www.airgas.com/msds/001013.pdf

CO2 is classified as a simple asphixiant (Section 2, classification) "Simple" asphyxiants are inert but can be dangerous because they displace oxygen. "Cellular" asphixiants (like carbon monoxide or hydrogen sulfide) actually disrupt cellular function. CO2 is not classified as toxic. :no:

That being said, it's clear CO2 is a byproduct of cellular function and is used to regulate PH levels in human blood. Breathing air with a high CO2 concentration does cause neurologic effects and death in humans. Then again, breathing air with a high oxygen concentration can have the exact same effect (See Carbon Dioxide Narcosis). Obviously, we don't classify oxygen as toxic. :scratch:

Regardless, the kicker here is that insects don't have blood. They have copper based hemolymph, which doesn't transport oxygen or CO2. Most CO2 affects on humans don't apply directly to insects. Insect cells absorb oxygen straight from the surrounding air and they can be surprisingly resistant to low oxygen levels. Grain silos use burners that maintain an atmosphere of less than 1 percent oxygen to kill insects. Some flood systems use nitrogen and others use CO2.

Might I suggest a simple experiment? Make two test runs using 80% CO2 and 20% oxygen versus 80% CO2 and 20% helium, nitrogen or argon... any inert gas other than oxygen. I suggest using a glass erlenmeyer flask. They seal well and you can see the moths/larva. I believe you'll find the second run more effective for controlling wax moths.

I'm not saying CO2 doesn't kill bugs. I'm just saying bugs need oxygen and large scale grain operations keep oxygen at less than 1% for a reason. I also think you've done some nice testing. :thumbsup:


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## Phoebee

For that matter, carbon monoxide also has little effect insects, probably less than CO2. Because there's no hemoglobin to bind to. Insects do have myoglobin, though.

CO2 knocks ants down in seconds at concentrations around 50%, and from the looks of the effect, it hurts. That's just something I've noticed while introducing CO2 to my wax moth test setup.

Deliberately taking a short puff into the mouth, the taste is very sour ... the pH effect is quite fast. That's clearly different from asphyxiants such as nitrogen.

All that said, EPA lists CO2 as "generally recognized as safe", while OSHA sets maximum permissible levels in the workplace, and FDA lets soda makers supersaturate soft drinks with the stuff.


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## CowDoc1

Phoebee said:


> the taste is very sour ... the pH effect is quite fast. That's clearly different from asphyxiants such as nitrogen.


I would add, "in humans". I don't think insects use CO2 to regulate PH as humans do. They don't have much of a circulatory system. Nitrogen and CO2 might "taste" similar to them. I don't believe we can equate CO2s effects on humans to insects. Very different physiology.

It would be nice to know what happens physiologically when insects are exposed to CO2. In people, CO2 crosses the blood / brain barrier because it's a non-polar molecule. That's why it causes neurologic effects. Most chemicals / drugs can't cross that barrier. I'm not sure insects have a blood / brain barrier to protect them. Yet, they're pretty resistant to chemicals. :kn:

I suspect someone even geeker than me has written an article about it in Entomology for Dumbies. Something like, "Physiologic Effects of Noble and Not so Noble Gases on Achroia grisella" :s


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## Phoebee

CowDoc1 said:


> I don't believe we can equate CO2s effects on humans to insects. Very different physiology.
> 
> It would be nice to know what happens physiologically when insects are exposed to CO2.


Insects contain water. When CO2 dissolves in water, it produces carbonic acid. Insects have a very different breathing apparatus, so no doubt it has some different regulatory mechanisms, but exposure to more CO2 than an insect can expel will lower its pH. It's just chemistry. The taste of CO2 in my mouth is sour because it almost instantly acidifies my spit. And it will instantly acidify an insect.

The remarkable thing is that wax moth larvae can withstand the enormous levels they do. And there are several peer-reviewed papers out on it. One or another of these CO2 fumigation threads call them out.


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## Rader Sidetrack

Randy Oliver has a page that is mostly discussing CCD, but has this interesting snippet on CO[SUB]2[/SUB] ...



> Even more remarkable, van Nerum & Buelens (1997) describe how the colony conserves energy and water by reducing ventilation with the tight winter cluster. The bees allow carbon dioxide levels to rise to as high as 6% (5% is dangerous to humans) inside the cluster. At the same time they create a condition of hypoxia (low oxygen), reducing the oxygen level from the 21% in the atmosphere to about 15%, which induces them to enter into an “ultra low metabolic rate”—similar to the diapause that other insects use to survive the winter. In this state, the colony uses minimum energy, and traps critical metabolic water within the cluster (the center experiences low relative humidity, and the cooler shell traps escaping moisture). (No mention was made of the effect that such an extraordinarily high CO2 level would have in increasing the acidity of the haemolymph—this may be an effect deserving a bit of research).
> 
> http://scientificbeekeeping.com/old-bees-cold-bees-no-bees-part-1/
> '


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## gww

Rader
Interesting find.
gww


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## Phoebee

OSHA CO2 exposure limits:

Carbon dioxide 5000 ppm (0.5%) for long-term (8-10 hr days depending on the jurisdiction)
30,000 ppm (3%) short term

Recommended levels to kill wax moths are typically above 60%, and it may take weeks at the lower levels. 80% kills in 4 days, 100% kills in 4 hours. The larvae appear to be the toughest stage to kill. Adults drop fast when the fumigation starts.


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