MGRox
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This originally started to be written up as a PM, but I thought maybe it would be better to talk about this on the open forum. The intention of the thread is to investigate a more reliable way to deal with problems that "can" commonly arise with various aero style cloners and bare root cuttings. (turbo kloner, aero cloner, certain bubblers, etc).
First, if you look over all of the reported problems with these systems you tend to find one aspect mentioned more common than the rest; Slime and often gray slime. Now we all know that "Slimes" are typically related to one of the Cyano's (red, brown, black, gray etc) or possibly a couple fungus, right? In which case Quat's or Chlorine should work perfectly right? But many have not found this to be the case, or been successful with any treatment; even after sterilization.
Enter: "Slime Producing Bacteria". There is a "group" of bacteria that primarily function to produce a slime with little other direct effects. Further, it has been found that 67% of Pathogenic and 34% of non-pathogenic bacteria; produce a slime as well. It is not commonly known yet, but there are labs that are beginning to test specifically for slime producing bacteria (for pools, water treatment, process industry, food, medical etc).
Okay, so there's bacteria that produces slime; no big deal right? Heck we all know that bacteria and some level of gooey go hand in hand, so why am I talking about this?
Here's a little bit of info from a place that does testing for slime producing bacteria (both gram positive and negative! slimes):
"The slime forming bacteria is the name given to a group of bacteria that are capable of producing a variety of extracellular polysaccharide polymers. It is these long chain molecules which act as the foundation and cement for the formation of biofilm. The slime-like growth coating the inside of pipes and fixtures is called the biofilm. The purpose of this slime layer appears to be protective. Under harsh environments (e.g., temperature changes, chemicals, shortage of nutrients) slime layers can get thicker. As the biofilm matures, aerobic bacteria creating the biomass produce metabolic by-products all the while consuming oxygen. This facilitates micro-environments underneath the biofilm which can then support the growth of anaerobic bacteria. "
From other reading on slime producing bacteria, they tend to grow on an ORP front / highest oxidative section of an aqueous system. (I.e. the lid, stems, manifold and pump in these cloners). These slime bacteria Prefer areas that are higher in; organic matter, carbohydrates, nutrients, pH fluctuation and chemicals. As well ; Chlorine is actually inactivated AND incorporated into the slime exacerbating the thickness for protection. I believe that it would be similar with alcohol but didn't look for that specifically.
Further reading with a paper attempting to develop standards for slime producing bacterial test, found that not every bacterial cell will necessarily produce this slime. A parent might produce the slime but the "offspring" cell from that may not. Also, from the perspective of sample purity and accuracy; they found that 1 slime producing cell per 16,000, could cover the entire test area with slime (negating accuracy). To me this tends to suggest that any cleaning method that would have less than 99.994% of all surfaces clean.......would have a 100% chance for the slime to re-occur [(1/16,000)-1].
I.e. The development of slime forming bacteria is an inevitability rather than chance (which makes sense why slime is often a word used with these cloners). One could even argue that it is still an inevitability regardless of cleaning method; given enough time.
So, in order to expound the investigation into a wider range we should now switch to a broader (applicable) term; Biofilm. All of these biological films are the polysaccharide polymers and these coatings make bacteria pretty much IMPERVIOUS to any "normal" treatment method. Problems related to treatment of bacterial colonization as a result of this film, cover areas of industry such as; wells, plumbing, pools, foods, medical devices, papermilling, waste treatment and even inside our own bodies.
The investigations / researches into "biofilm" actually have been increasing quite dramatically in the last 15 years or so and it's being looked at more intently. Various industries have been unable to properly deal with these slimes and have had to make alterations accordingly. In the medical field; they are actually starting to quantify biofilms as a completely separate item from the bacteria itself. As unless the slime itself is treated; the underlying bacterial pathogen cannot be.
Here are actually a couple good video's that talk a bit about "biofilms" (though more relating to impacts in our bodies / medical).
VIDEO 1
VIDEO 2
At this point, besides knowing better why bacteria is everywhere; it should be agreeable to most that the slime itself is the issue to treat, not the bacteria. It is important to note that the films provide a safe haven for numerous other possible pathogens to develop. While the slime does also cut off oxygen, typically other pathogens that develop under the film are the final cause of death.
The question for reliability with these cloners, now becomes how to eliminate existing slime and / or to prevent biofilm formation. Here's where I'm at / what I've found / what may work:
-Prophylactically - with bacteria only in planktonic state and assuming a 100% sterile unit; Chlorine can work, assuming there are not surfaces that are wet; without exposure to the Cl. If a Cl level of 2ppm is maintained permanently, or better ORP controlled to 780+; this "theoretically" would prevent bacteria from living. (this is also relates to why "city tap" is often recommended over R/o).
However, I hesitate to consider this "assurance" in any way; considering the virility / survivability of bacteria in general. Other areas that perform regular or prophylactic sterilization, (medical devices, papermilling and wastewater treatment plants) ALL still have biofilm formation despite continual treatments and regular sterilizations. Most specifically with waste plants and papermills as they use Cl already and at higher ppms that we can have with plants.
-Treatment of slime - Chlorine Dioxide WILL remove biofilms as well as being an antibacterial. There are a couple ways to remove biofilms, though it is more tough with plants and this is seemingly the best. Under continual application; dosages required to remove and prevent biofilms range 0.1 - 1.0 ppm. For intermittent application, effective dosages range 2 - 5 ppm.
Chlorine dioxide has recently been widely adopted in papermilling processes where chlorine had many disadvantages. Also, the wastewater industry is beginning to switch over to using this as opposed to Chlorine. It is more effective than Cl, does not react with organic matter, is not sensitive to pH, can be used in lower quantities and does not produce the same reacted byproducts.
Continual application as a preventative would, of course, be the most optimal situation and would virtually guarantee no future issues, ever. Though, the only current methods of continual application are via Chlorine dioxide generators that are pretty expensive (however, a large UC or NFT may well be worth it). As any concentration of chlorine dioxide at or above 3% needs to be generated on site. It is also fairly reactive and a soluble gas, so it does not store well in solution (some 2% sol. exist).
For treatment applications - Quite a few of the "water purification tablets" are, in fact, chlorine dioxide (typically instruct to 4ppm concentration). Also, there are a "couple" pool companies that sell this same tablet in a larger form; though most pools, with issues, install a generator. For our applications though, the "water purification tablets" (or drops) provide a way to treat and remove the biofilms.
The EPA maximum allowable level of chlorine dioxide for drinking is 0.8ppm and the purification tablets recommend a 4 hour wait, starting @ 4ppm. So in a "Still" environment and after 4 hours; the ppm of chlorine dioxide would be below 0.8ppm. The longevity of the Chlorine Dioxide in a cloner with the pump running would be, in the order of, less than 30 minutes.
---------------------
This is basically where I'm at atm. I have not yet tested the chlorine dioxide but intend to, upon seeing this issue. I have some cuts currently and the setup DID have the biofilm; though I had taken measures prior to researching all this. I fully expect that even with all sterilization I did, since old cuts went back in; that I will see this issue shortly.
My intention is to dissolve a tablet with pump off to 2 ppm. Once the tablet is dissolved, I will run the system for 30 minutes. After the 30 minutes, the system will be torn down and water dumped into a tray or container (large enough to hold cuts in neoprenes). All cuts will be transferred to sit in the water with neoprenes (from cloner) while rest of unit is cleaned with soap and hot water (not Cl). New R/O water will be added and cloner set back up. Cuts will be quickly washed across R/O before putting back into the unit. New Chlorine dioxide tablets will be added with pump off to 1 ppm. Finally system is ran as normal (timed or not).
Last, some changes / thoughts I have after learning all of this. First, even though some NPK may be debatably better (from certain perspectives) than none; I feel since the biofilm develops more in higher nutrient levels, it may be the lesser evil to run R/O (including comparing city or well). Also, since orgainic matter also increases biofilm producing bacteria; I'm going to avoid using ANY rooting accelerator (rapid start, etc) or humics / fulvics. As well keeping all of this as close to pure as possible, will help to keep the ORP lower; which again increases the biofilm. Finally, I noted that any stems which had been "nicked" (more technically basal wounding); had a dramatically higher level of biofilm than stems which were not. Here again I wonder if the advantage to greater root sites may be outweighed by the risk of biofilm; so I will not be doing basal wounding for this system.
I'm indecisive about whether or not to run a prophylactic Cl (would only do with a controller) as eventually the biofilm would occur and come on quick when it did. However, the chlorine dioxide treatment could still be performed; provided some time was left for Cl to dissipate first.
I would be very interested to hear others thoughts and to see if others would like to do experimentation too. If there's a way to have consistent reliability with these units across a wide range of environments; it may well be one of the top overall methods. However IMHO, currently, it does seem that "in total" there is a lower overall success rate with this method (considering all users) vs virtually any other clone method.
First, if you look over all of the reported problems with these systems you tend to find one aspect mentioned more common than the rest; Slime and often gray slime. Now we all know that "Slimes" are typically related to one of the Cyano's (red, brown, black, gray etc) or possibly a couple fungus, right? In which case Quat's or Chlorine should work perfectly right? But many have not found this to be the case, or been successful with any treatment; even after sterilization.
Enter: "Slime Producing Bacteria". There is a "group" of bacteria that primarily function to produce a slime with little other direct effects. Further, it has been found that 67% of Pathogenic and 34% of non-pathogenic bacteria; produce a slime as well. It is not commonly known yet, but there are labs that are beginning to test specifically for slime producing bacteria (for pools, water treatment, process industry, food, medical etc).
Okay, so there's bacteria that produces slime; no big deal right? Heck we all know that bacteria and some level of gooey go hand in hand, so why am I talking about this?
Here's a little bit of info from a place that does testing for slime producing bacteria (both gram positive and negative! slimes):
"The slime forming bacteria is the name given to a group of bacteria that are capable of producing a variety of extracellular polysaccharide polymers. It is these long chain molecules which act as the foundation and cement for the formation of biofilm. The slime-like growth coating the inside of pipes and fixtures is called the biofilm. The purpose of this slime layer appears to be protective. Under harsh environments (e.g., temperature changes, chemicals, shortage of nutrients) slime layers can get thicker. As the biofilm matures, aerobic bacteria creating the biomass produce metabolic by-products all the while consuming oxygen. This facilitates micro-environments underneath the biofilm which can then support the growth of anaerobic bacteria. "
From other reading on slime producing bacteria, they tend to grow on an ORP front / highest oxidative section of an aqueous system. (I.e. the lid, stems, manifold and pump in these cloners). These slime bacteria Prefer areas that are higher in; organic matter, carbohydrates, nutrients, pH fluctuation and chemicals. As well ; Chlorine is actually inactivated AND incorporated into the slime exacerbating the thickness for protection. I believe that it would be similar with alcohol but didn't look for that specifically.
Further reading with a paper attempting to develop standards for slime producing bacterial test, found that not every bacterial cell will necessarily produce this slime. A parent might produce the slime but the "offspring" cell from that may not. Also, from the perspective of sample purity and accuracy; they found that 1 slime producing cell per 16,000, could cover the entire test area with slime (negating accuracy). To me this tends to suggest that any cleaning method that would have less than 99.994% of all surfaces clean.......would have a 100% chance for the slime to re-occur [(1/16,000)-1].
I.e. The development of slime forming bacteria is an inevitability rather than chance (which makes sense why slime is often a word used with these cloners). One could even argue that it is still an inevitability regardless of cleaning method; given enough time.
So, in order to expound the investigation into a wider range we should now switch to a broader (applicable) term; Biofilm. All of these biological films are the polysaccharide polymers and these coatings make bacteria pretty much IMPERVIOUS to any "normal" treatment method. Problems related to treatment of bacterial colonization as a result of this film, cover areas of industry such as; wells, plumbing, pools, foods, medical devices, papermilling, waste treatment and even inside our own bodies.
The investigations / researches into "biofilm" actually have been increasing quite dramatically in the last 15 years or so and it's being looked at more intently. Various industries have been unable to properly deal with these slimes and have had to make alterations accordingly. In the medical field; they are actually starting to quantify biofilms as a completely separate item from the bacteria itself. As unless the slime itself is treated; the underlying bacterial pathogen cannot be.
Here are actually a couple good video's that talk a bit about "biofilms" (though more relating to impacts in our bodies / medical).
VIDEO 1
VIDEO 2
At this point, besides knowing better why bacteria is everywhere; it should be agreeable to most that the slime itself is the issue to treat, not the bacteria. It is important to note that the films provide a safe haven for numerous other possible pathogens to develop. While the slime does also cut off oxygen, typically other pathogens that develop under the film are the final cause of death.
The question for reliability with these cloners, now becomes how to eliminate existing slime and / or to prevent biofilm formation. Here's where I'm at / what I've found / what may work:
-Prophylactically - with bacteria only in planktonic state and assuming a 100% sterile unit; Chlorine can work, assuming there are not surfaces that are wet; without exposure to the Cl. If a Cl level of 2ppm is maintained permanently, or better ORP controlled to 780+; this "theoretically" would prevent bacteria from living. (this is also relates to why "city tap" is often recommended over R/o).
However, I hesitate to consider this "assurance" in any way; considering the virility / survivability of bacteria in general. Other areas that perform regular or prophylactic sterilization, (medical devices, papermilling and wastewater treatment plants) ALL still have biofilm formation despite continual treatments and regular sterilizations. Most specifically with waste plants and papermills as they use Cl already and at higher ppms that we can have with plants.
-Treatment of slime - Chlorine Dioxide WILL remove biofilms as well as being an antibacterial. There are a couple ways to remove biofilms, though it is more tough with plants and this is seemingly the best. Under continual application; dosages required to remove and prevent biofilms range 0.1 - 1.0 ppm. For intermittent application, effective dosages range 2 - 5 ppm.
Chlorine dioxide has recently been widely adopted in papermilling processes where chlorine had many disadvantages. Also, the wastewater industry is beginning to switch over to using this as opposed to Chlorine. It is more effective than Cl, does not react with organic matter, is not sensitive to pH, can be used in lower quantities and does not produce the same reacted byproducts.
Continual application as a preventative would, of course, be the most optimal situation and would virtually guarantee no future issues, ever. Though, the only current methods of continual application are via Chlorine dioxide generators that are pretty expensive (however, a large UC or NFT may well be worth it). As any concentration of chlorine dioxide at or above 3% needs to be generated on site. It is also fairly reactive and a soluble gas, so it does not store well in solution (some 2% sol. exist).
For treatment applications - Quite a few of the "water purification tablets" are, in fact, chlorine dioxide (typically instruct to 4ppm concentration). Also, there are a "couple" pool companies that sell this same tablet in a larger form; though most pools, with issues, install a generator. For our applications though, the "water purification tablets" (or drops) provide a way to treat and remove the biofilms.
The EPA maximum allowable level of chlorine dioxide for drinking is 0.8ppm and the purification tablets recommend a 4 hour wait, starting @ 4ppm. So in a "Still" environment and after 4 hours; the ppm of chlorine dioxide would be below 0.8ppm. The longevity of the Chlorine Dioxide in a cloner with the pump running would be, in the order of, less than 30 minutes.
---------------------
This is basically where I'm at atm. I have not yet tested the chlorine dioxide but intend to, upon seeing this issue. I have some cuts currently and the setup DID have the biofilm; though I had taken measures prior to researching all this. I fully expect that even with all sterilization I did, since old cuts went back in; that I will see this issue shortly.
My intention is to dissolve a tablet with pump off to 2 ppm. Once the tablet is dissolved, I will run the system for 30 minutes. After the 30 minutes, the system will be torn down and water dumped into a tray or container (large enough to hold cuts in neoprenes). All cuts will be transferred to sit in the water with neoprenes (from cloner) while rest of unit is cleaned with soap and hot water (not Cl). New R/O water will be added and cloner set back up. Cuts will be quickly washed across R/O before putting back into the unit. New Chlorine dioxide tablets will be added with pump off to 1 ppm. Finally system is ran as normal (timed or not).
Last, some changes / thoughts I have after learning all of this. First, even though some NPK may be debatably better (from certain perspectives) than none; I feel since the biofilm develops more in higher nutrient levels, it may be the lesser evil to run R/O (including comparing city or well). Also, since orgainic matter also increases biofilm producing bacteria; I'm going to avoid using ANY rooting accelerator (rapid start, etc) or humics / fulvics. As well keeping all of this as close to pure as possible, will help to keep the ORP lower; which again increases the biofilm. Finally, I noted that any stems which had been "nicked" (more technically basal wounding); had a dramatically higher level of biofilm than stems which were not. Here again I wonder if the advantage to greater root sites may be outweighed by the risk of biofilm; so I will not be doing basal wounding for this system.
I'm indecisive about whether or not to run a prophylactic Cl (would only do with a controller) as eventually the biofilm would occur and come on quick when it did. However, the chlorine dioxide treatment could still be performed; provided some time was left for Cl to dissipate first.
I would be very interested to hear others thoughts and to see if others would like to do experimentation too. If there's a way to have consistent reliability with these units across a wide range of environments; it may well be one of the top overall methods. However IMHO, currently, it does seem that "in total" there is a lower overall success rate with this method (considering all users) vs virtually any other clone method.
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