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How does freezing kill parasites?


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#1 paulraphael

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Posted 16 October 2008 - 08:58 AM

From New York Health Dept:

"The most common fish parasite is the anisakis round worm that causes the illness anisakiasis. Fish eaten raw, marinated or partially cooked can be made safe by being frozen in one of two way (1) Frozen and stored in a freezer at -4°F or lower for 7 days or secondly, frozen at -31°F or below until solid and stored at that temperature for 15 hours."

How does this work? Especially the flash freezing that's used on sashimi-grade fish ... why wouldn't the minute size of the ice crystals that preserve the cell structure of the fish also preserve the cell structure of the creepy crawlies? I remember a kid at science fair flash freezing a goldfish and bringing it back to life.

Thoughts?

#2 Gifted Gourmet

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Posted 16 October 2008 - 10:14 AM

When thinking about my ceviche, gravlax, sushi prep after reading this topic, I located some rather interesting information on the topic:

freezing: does it kill parasites?

What freezing doesn't kill, I understand, is bacteria .... :hmmm:
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#3 Gabriel Lewis

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Posted 16 October 2008 - 10:43 AM

I believe the critical thing here is time. Yes you can flash freeze things and bring them back to an extent, but enough time spent at a low temperature will eventually lead to death. Freezing doesn't "stop" time (unless maybe you reach 0K), and all organisms are constantly doing internal things like replacing cells etc; their basic homeostatic mechanisms need to remain active in order for them to stay alive.

#4 Special K

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Posted 16 October 2008 - 10:46 AM

Here you go: http://www.cfsan.fda...mm/haccp4e.html

#5 paulraphael

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Posted 16 October 2008 - 10:50 AM

Freezing doesn't "stop" time (unless maybe you reach 0K), and all organisms are constantly doing internal things like replacing cells etc; their basic homeostatic mechanisms need to remain active in order for them to stay alive.


Well, the point isn't stopping time; it's stopping the mobility of water. Biological processes can't take place if water can't move. The temperature at which water is completely immobilized depends on what's disolved in it; in general by the time something is at -20F you can assume that all biological activity is stopped. Not just slowed.

Physical processes, like dehydration (freezerburn), and chemical processes, like oxidation (rancidification) can continue.

I still don't understand why a specific amount of freezer time kills parasites.

#6 Chris Hennes

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Posted 16 October 2008 - 11:14 AM

It's really a very interesting question: have you seen

Thomas L. Deardorff and Richard Throm, "Commercial Blast-Freezing of Third-Stage Anisakis simplex Larvae Encapsulated in Salmon and Rockfish." The Journal of Parasitology, Vol. 74, No. 4 (Aug., 1988), pp. 600-603

From that article's discussion section:

Further, the severe damage to the internal morphological structures of these worms may have been caused by the fast cooling and subsequent formation of intracellular ice.


This article seems to indicate that the difference between freezing for one hour and for 24 hours is merely one of ensuring that the interior regions of muscles are completely frozen. Clearly, this would occur well before the extremely long hold times recommended in food safety literature.

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#7 Gabriel Lewis

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Posted 16 October 2008 - 11:20 AM

You right, I guess I didn't really think about it enough. Checking Mcgee, the only tidbit he has is that parasites are susceptible due to their "more complex biological organization" (pg 186). I would assume that this refers primarily to the added complexity of a multicellular organism, but given that some parasites can be single celled protozoa, I have to wonder if freezing is effective for only some parasites rather than all of them. Certainly different parasites show different susceptibilities to freezing than others, and glancing through an article on pubmed, I see one case where a parasite was able to reproduce in horse meat that had been frozen for four weeks.

As to what actually leads to the death of the parasite in terms of how freezing affects a specific critical function(s) I can't even really guess, and I'm guessing that we probably just don't know in most cases. You would have to do a bunch of experiments comparing the histology of live/frozen killed parasites. As susceptibility varies between parasites, I'd guess that how freezing actually kills different parasites does too.

Myself, I'm content to believe that enough time spent at a low enough temperature will affect some critical biological function of a multicellular organism.

#8 Gabriel Lewis

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Posted 16 October 2008 - 11:44 AM

I can't stop thinking about this now, and I've come up with what I think is a plausible theory:

Even when things are "frozen" at -20C, at the molecular level its not 100% of the water that has turned into ice. To a very very small degree, there are continuous miniature freeze/thaw cycles of all the water in different parts of the frozen tissue. It is probably these miniature freeze/thaw cycles that do damage to the parasites, and this damage builds up with the length of time spent in the freezer. If you freeze them initially for 24 or 48 hours or whatever, it will be enough to freeze their bodies rock solid, but given their relatively simple biological organization it won't kill them. However, if you leave them in the freezer long enough for ice crystals to melt/reform all over their bodies, and do damage in each new affected area, then eventually the damage will surpass a threshold such that the parasites can no longer "come back to life".

#9 Peter the eater

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Posted 16 October 2008 - 01:14 PM

I see often those coiled up little roundworms in fish, usually not until I get home. I never worry too much about them since they're harmless once cooked. If I cure seafood or make ceviche I would avoid the species prone to anisakis such as cod, halibut, monkfish, herring, etc. If you're serving uncooked fish just make sure the pieces are too small to hide a bunch of worms.

My main seafood market Clearwater does their best to remove the parasites using visual inspection while processing and packaging, often with a light table. Another vendor sells sashimi grade tuna that is very fresh but has never been cold-treated for worms. They tell me that it is impossible to find an adult wild fish in the Atlantic ocean that doesn't have at least one parasite.
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#10 Adam Balic

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Posted 16 October 2008 - 04:59 PM

They tell me that it is impossible to find an adult wild fish in the Atlantic ocean that doesn't have at least one parasite.

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From my experience of parasitologist and fish eater I would say that was correct. My fishmonger in Scotland refused to eat cod on the grounds that it was always filthy with worms.

#11 savvysearch

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Posted 17 October 2008 - 05:43 AM

First, I have to question your premise. Are you sure the goldfish was truly frozen? Fish are cool-blooded animals so their bodies can survive at colder temperatures, and their internal processes slow down. Most likely the fish wasn't completely frozen. Maybe the exterior was truly frozen, but the fish's metabolism just slowed to give the appearance that it was completely frozen. As far as I know, fish can't come back to life after being put on ice or flash frozen.


Most multicellular organisms, especially animals, can't survive freezing. Parasites are no exception. Freezing doesn't necessarily preserve the cell structure. It'll prevent the cells from deterioration, but it tends to physically damage the cell first. When freezing, jagged ice crystals form, which can tears cells and tissues. Also, water outside of the cells form a highly organized crystal structure and therefore expands when frozen, which results in "squeezing" the cells. In addition, as water freezes, it pushes solutes out of the crystallized ice structure that is forming and against the outside of the cell walls. Osmosis is a physical process of water moving to areas of higher solute concentrations. So as solutes build up outside of the cell, water moves out of the cell to equalize the concentration. Thus the cell is also damaged by shrinking as water moves out of it.




I'd bet the thawing process has much to do with it as well. If freezing doesn't kill the parasite, then coming out of the frozen state, I suspect, inevitably will. Multicellular organisms have to be in a constant state of homeostasis where their internal and cellular mechanisms are constantly regulated and working "in sync". And as far as I know, there's no such thing as "flash thawing." Inevitably, thawing defrosts some parts faster than other parts. So some mechanisms or internal processes would begin working while other processes which they depend upon, won't. Let's say that one process that's still frozen would prevents oxygen and nutrients from arriving to already defrosted areas, and causing the cells in that area to die. You can see how this would work for organisms with some level of internal organization.

Edited by savvysearch, 17 October 2008 - 05:47 AM.


#12 Chris Hennes

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Posted 17 October 2008 - 07:08 AM

Most multicellular organisms, especially animals, can't survive freezing. Parasites are no exception. Freezing doesn't necessarily preserve the cell structure. It'll prevent the cells from deterioration, but it tends to physically damage the cell first. When freezing, jagged ice crystals form, which can tears cells and tissues. Also, water outside of the cells form a highly organized crystal structure and therefore expands when frozen, which results in "squeezing" the cells. In addition, as water freezes, it pushes solutes out of the crystallized ice structure that is forming and against the outside of the cell walls. Osmosis is a physical process of water moving to areas of higher solute concentrations. So as solutes build up outside of the cell, water moves out of the cell to equalize the concentration. Thus the cell is also damaged by shrinking as water moves out of it.

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Everything you say is true, though the commercial freezing process is specifically designed to prevent the formation of those sharp ice crystals. But even assuming that the ice crystals are to blame, it still does not explain why it sometimes takes weeks to kill all the parasites. In particular, to rid pork of Trichiniosis the FDA says you have to freeze at temperature X for time Y, where sometimes Y is measured in weeks. Can it really take weeks to fully freeze a piece of pork? The guidelines say nothing about muscle thickness, which would govern time-to-freeze. It seems we are still missing a piece of the puzzle. Why does time matter? I am interested in Gabriel Lewis's hypothesis about the continuous formation and melting of these microscopic ice crystals, but I have not read anywhere that this has been experimentally verified.

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#13 paulraphael

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Posted 17 October 2008 - 07:41 AM

First, I have to question your premise. Are you sure the goldfish was truly frozen?  Fish are cool-blooded animals so their bodies can survive at colder temperatures, and their internal processes slow down. Most likely the fish wasn't completely frozen. Maybe the exterior was truly frozen, but the fish's metabolism just slowed to give the appearance that it was completely frozen. As far as I know, fish can't come back to life after being put on ice or flash frozen.


You could be right. It was a science fair experiment from a long time ago. After "thawing," the fish lived about two minutes (during which he seemed completely drunk).

I've been doing some searches on cryonics, and most suggest that goldfish can't, in fact survive freezing. But according to many sources, including this one, some fish and small animals can:

"Viruses, bacteria, sperm/eggs, embryos at early stages of development, insects, and even small animals (small frogs, some fish) can be cryogenically frozen, preserved for an indefinite time (as long as low temperature is maintained) and then thawed and returned to a living state. Large animals or organs (a few centimeters and larger) can not be safely frozen because removing heat via thick tissue by natural thermoconductivity becomes so slow that ice microcrystals grow big enough to damage cell membranes."

At any rate, the issue seems like a complicated one. There are different mechanisms at work causing cell damage, not just in the freezing process, but in the storing and also the thawing process. And these processes seem to be dependent on many variables. It's still seems curious that freezing reliably kills parasites, but not some larger organisms like insects and embryos.

#14 Hawthorne

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Posted 17 October 2008 - 04:37 PM

First, I have to question your premise. Are you sure the goldfish was truly frozen?   Fish are cool-blooded animals so their bodies can survive at colder temperatures, and their internal processes slow down. Most likely the fish wasn't completely frozen. Maybe the exterior was truly frozen, but the fish's metabolism just slowed to give the appearance that it was completely frozen. As far as I know, fish can't come back to life after being put on ice or flash frozen.


You could be right. It was a science fair experiment from a long time ago. After "thawing," the fish lived about two minutes (during which he seemed completely drunk).

I've been doing some searches on cryonics, and most suggest that goldfish can't, in fact survive freezing. But according to many sources, including this one, some fish and small animals can:

"Viruses, bacteria, sperm/eggs, embryos at early stages of development, insects, and even small animals (small frogs, some fish) can be cryogenically frozen, preserved for an indefinite time (as long as low temperature is maintained) and then thawed and returned to a living state. Large animals or organs (a few centimeters and larger) can not be safely frozen because removing heat via thick tissue by natural thermoconductivity becomes so slow that ice microcrystals grow big enough to damage cell membranes."

At any rate, the issue seems like a complicated one. There are different mechanisms at work causing cell damage, not just in the freezing process, but in the storing and also the thawing process. And these processes seem to be dependent on many variables. It's still seems curious that freezing reliably kills parasites, but not some larger organisms like insects and embryos.

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I don't know how or why it happened, but I saw this done on a science program years ago - the gold fish was dunked into liquid nitrogen, which certainly froze it, and when it was returned to its bowl it was apparently restored. I don't recall it being apparently drunk, but I also don't know how long it might have lived beyond the experiment.

Feather mites are apparently killed by freezing, (a little off topic, but not entirely) but I always thought it was the thawing process which did the job, since I understand the thawing process to rupture cell membranes.

The only raw fish I eat is Nova Scotia salmon, but I assume that that is less raw than is apparent. It's raw enough for me :)
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#15 Temple

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Posted 17 October 2008 - 10:21 PM

Freezing does kill vegetative bacteria quite effectively. One of the first things you learn as a microbiologist is that you don't store your bacterial samples in water alone. You have to store it along with a cryoprotectant, such as glycerol or DMSO, or else the concentration of viable cells will be dramatically reduced during freezing. Also, for lab use of commercial freezers, one of the primary requirements is that it doesn't have a de-icing feature, since the freezing-thawing of the de-icing cycle can wipe out all your samples.

The issue in regards to food safely is that freezing doesn't kill all bacteria, and viable unicellular organisms that do survive can rapidly replicate under the right conditions and become harmful once again. For multicellular organisms, like parasites, the freezing process will cause cellular damage and should be fatal; this is especially true for uncontrolled slow-rate freezing environments, like a normal freezer, that are being frozen primarily in its own fluids without cryoprotectants.

Now, in regards to cryobiology, small multicelluar organisms can be frozen if they are frozen fast enough and cold enough. Ultra-rapid vitrification methodology for cryopreservation is widely used to freeze things like embryos, however, even using this methodology 50% of embryos don't survive.

On an interesting somewhat off-topic note, its been reported that painted turtle hatchlings can survive freezing. Its thought to be because of high-concentration of glycerol in its blood that acts as a cryoprotectant and other physical changes that promote vitrification. A technique that is similarly by tardigrades (water bears); which can also survive slow-rate freezing.

#16 Patapsco Mike

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Posted 18 October 2008 - 11:40 AM

I think it's being made too complicated. It has nothing to do with sharp crystals. Freezing kills organic life forms that don't have some kind of anti-freeze for the same reason that a jug of water filled to the top and frozen will break. Water expands quite a lot when frozen, and cell membranes to not have room for the amount of expansion that takes place. The cells and even the structures inside the cells burst as the water expands, and this is fatal.

Quite a few organisms can live at below freezing temperatures (ice worms, some amphibians, etc). These organisms have one thing in common- they live in an environment where they have had evolutionary pressure that forces them to adapt to being at below freezing temperatures. They will still die if frozen solid, but they have various types of chemicals that allow the water in them to stay liquid at below freezing temperatures.

Fishes don't freeze, so the parasites that are adapted to live in them have never developed this ability. So freezing very reliably kills them.
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#17 Chris Hennes

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Posted 19 October 2008 - 11:14 AM

I think it's being made too complicated.  It has nothing to do with sharp crystals.  Freezing kills organic life forms that don't have some kind of anti-freeze for the same reason that a jug of water filled to the top and frozen will break. 

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OK, but WHY does TIME matter? Once something is frozen solid, I would have thought that was it. But it does not seem to be. For some reason, holding something at freezing temperatures for what to me seems like far longer than it would take to reach thermal equilibrium kills more critters. What is the mechanism through which this occurs? Or is this simply a matter of the FDA over-simplifying the science, and basically assuming that any time we freeze we are freezing a whole animal, and thus at high temperatures it really does take 2-3 weeks to actually freeze through?

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#18 Peter Green

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Posted 19 October 2008 - 01:47 PM

The Food Sci wife supports the "freeze and kill" thesis (which may be why I avoid Northern climes). These parasites are (relatively) complicated life forms, and won't stand up to the travails of an extended Canadian winter.

Mind you, neither would I.

P.S. - why does this thread have me fixated on Walt Disney?

#19 paulraphael

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Posted 19 October 2008 - 02:24 PM

Now, in regards to cryobiology, small multicelluar organisms can be frozen if they are frozen fast enough and cold enough.


This is what I would have assumed, but the bulk of fish sold for raw consumption is cryogenically flash frozen. It's required by law for all but one or two species of fish, in order to kill parasites ... but it's done fast enough that cellular damage is minimal, and the texture of the fish doesn't suffer.

I've read that some sushi chefs actually prefer flash frozen fish, because in many cases the texture penalty is minimal if it's there at all, and the improvement to freshness is noticeable.

I'd think these freezing conditions would be the exact ones that encourage parasites to survive, but apparently that's not the case.

#20 Patapsco Mike

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Posted 19 October 2008 - 03:16 PM

OK, but WHY does TIME matter?


FDA rules are the way they are for all sorts of reasons, food science being but one. It could be to make it reasonably likely that the food will be inspected while frozen- who knows...
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#21 Temple

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Posted 19 October 2008 - 08:01 PM

I think it's being made too complicated.  It has nothing to do with sharp crystals.  Freezing kills organic life forms that don't have some kind of anti-freeze for the same reason that a jug of water filled to the top and frozen will break.  Water expands quite a lot when frozen, and cell membranes to not have room for the amount of expansion that takes place.  The cells and even the structures inside the cells burst as the water expands, and this is fatal.

Quite a few organisms can live at below freezing temperatures (ice worms, some amphibians, etc).  These organisms have one thing in common- they live in an environment where they have had evolutionary pressure that forces them to adapt to being at below freezing temperatures.  They will still die if frozen solid, but they have various types of chemicals that allow the water in them to stay liquid at below freezing temperatures.

Fishes don't freeze, so the parasites that are adapted to live in them have never developed this ability.  So freezing very reliably kills them.

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The problem relative to cellular damage is the phase change between liquid to solid. But crystallization is part of it, as is the expansion of the medium as you have stated. I suppose it can be either complicated or simple depending on your perspective, however, any organism not adapted to live in cold temperatures will unlikely survive (as you have also stated).

Arctic animals do have 'anti-freeze', usually involving cold-induced mitochondrial changes that are correlated with increased glycerol-production. You are absolutely right in stating that most of these arctic creatures aren't completely frozen (there are very few animals that can survive those conditions like the painted turtle hatchling), and most parasites haven't adapted to those conditions and should not survive.

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#22 Temple

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Posted 19 October 2008 - 08:04 PM

I think it's being made too complicated.  It has nothing to do with sharp crystals.  Freezing kills organic life forms that don't have some kind of anti-freeze for the same reason that a jug of water filled to the top and frozen will break. 

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OK, but WHY does TIME matter? Once something is frozen solid, I would have thought that was it. But it does not seem to be. For some reason, holding something at freezing temperatures for what to me seems like far longer than it would take to reach thermal equilibrium kills more critters. What is the mechanism through which this occurs? Or is this simply a matter of the FDA over-simplifying the science, and basically assuming that any time we freeze we are freezing a whole animal, and thus at high temperatures it really does take 2-3 weeks to actually freeze through?

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Time doesn't stop because of freezing, there are still solid-solid phase transitions, and there are reactions going on after what we consider is frozen. In the case of supercooling the temperature continues to drop below the freezing point of the substance without any crystalline structures forming. Whether and to what extent a substance tends to supercool rather than undergo a phase change at its freezing point is often a function of the purity of the substance and the presence or absence of seed crystals or seed surfaces within the liquid. My assumption regarding FDA regulations is that these are the upper-time limits in which parasites cannot reliability survive.

#23 Temple

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Posted 19 October 2008 - 08:12 PM

Now, in regards to cryobiology, small multicelluar organisms can be frozen if they are frozen fast enough and cold enough.


This is what I would have assumed, but the bulk of fish sold for raw consumption is cryogenically flash frozen. It's required by law for all but one or two species of fish, in order to kill parasites ... but it's done fast enough that cellular damage is minimal, and the texture of the fish doesn't suffer.

I've read that some sushi chefs actually prefer flash frozen fish, because in many cases the texture penalty is minimal if it's there at all, and the improvement to freshness is noticeable.

I'd think these freezing conditions would be the exact ones that encourage parasites to survive, but apparently that's not the case.

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When we refer to 'flash frozen' from a food science perspective its dramatically different from the ultra-rapid vitirification that is used for cryogenic work. The requirements for keeping an organisms alive after frozen are incredibly strict from what we consider to be 'fresh' when eaten; as I'm sure you can imagine. For comparison, a standard liquid nitrogen freezer is -190 degrees Celsius.

The goal to make a multicelluar organism survive phase change between liquid to solid requires the liquid matter in the organism to change into an amorphous solid that is free from any crystalline structure. For many organisms DMSO is toxic (even at low concentrations), and glycerol isn't practical, hence cryoprotectants aren't an option in many circumstances. Even if those hurdles are met, the amount of cellular damage is fatal for most organisms, and under the most modern vitrification technologies there is still significant tissue damage. If we simplify our thinking; what percent cell damage do you think an organism can survive? 5%? 10%? 30%?