QT and Biofilm

[Humblefish]

@Gweeds1980 Maybe use lipase or H2O2 to wipe down the inside glass of the QT before setting it back up? Ever test vinegar against biofilms?

 

[Gweeds1980]

@Gweeds1980 Maybe use lipase or H2O2 to wipe down the inside glass of the QT before setting it back up? Ever test vinegar against biofilms?

We did, but bare in mind that our experiment involved biological destruction, not mechanical, compounds were added to Petri dishes without any scrubbing. From memory vinegar and alcohol (and anything else with a carbon source) caused growth not destruction. The hydrophobic layer created is very effective as repelling most substances, however it's worth noting that after the experiment, we used good old fashioned elbow grease to clean the dishes. Not a single biofilm stood up to hot water and a scrubbing brush!

 

[Gweeds1980]

@Humblefish any more news on this? Particularly interested in what's in these meds that biofilms are taking out to cause the breakdown. If we know that, we can narrow down the bacterial strains that are likely causing it and then focus treatments towards those strains. Key here is the idea of the biofilms being a bad thing... it really isn't. It's the strains of bacteria that are protected within the biofilms which are the issue.

In a normally functioning DT, biofilms are excellent news most of the time, our use of bacteria to break down waste is reliant on biofilm creation, to the extent that it isn't coincidence that biofilm creation rate (2-3 months) almost mirrors exactly the amount of time it takes to properly cycle an aquarium.

Without biofilms the bacteria we harness would fall off our rocks and filter media.

My knowledge on biofilm creation and destruction is pretty decent, but my knowledge of fish med ingredients and chemical equations is sadly lacking!

 

[Humblefish]

@Gweeds1980 I've been reading up on how stevia might be capable of killing Lyme disease, including its biofilm form. See links below:

https://draxe.com/stevia-kills-lyme-disease/

https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4681354/

 

[Gweeds1980]

Interesting... it's been known for a while that Stevia and other sweeteners have a negative impact on bacteria... studies have shown that it is effective at killing lactobacilli in vitro and in the gastrointestinal tract.

That it breaks down the bacteria within biofilm is certainly a surprise to me. I can't see the process in which RebA kills the bacteria, although I assume it must interrupt the bacterias ability to synthesise amino acids? If that is the case, breaking down the biofilm, which is mostly exopolysaccharides, proteins and nucleic acids, seems to make sense.

It would be relatively simple to carry out a basic, uncontrolled experiment to test Stevia in a marine setting... My concern would be that RebA is non specific in its ability to destroy bacteria and therefore you would expect those microbes involved in the nitrogen cycle to succumb also.

 

[Gweeds1980]

If be interested to see also if N-acetyl cysteine could be somehow incorporated into meds to prevent biofilm build up during treatment. If that could also include RebA to destroy what biofilm may already exist, you have the perfect anti-bacterial, anti-biofilm med and not have concerns over biodegradation. This is where I hand over to someone who knows more about the interactivity of these compounds with those already in various meds...

 

[Humblefish]

Wanted to add this great information provided by @Gweeds1980

I was just reading more on biofilms this morning. Apparently they can form on "virtually every non-shedding surface". So, I can only assume this would include sponges, ceramic rings, etc. Perhaps we can get @Gweeds1980's take on this as he has studied biofilms.

But if the above is true, it might be safer to use a bacteria in a bottle product to seed your QT or avoid using a sponge that has been sitting in the DT sump for months. @Gweeds1980 How quickly can a biofilm form on a new surface?

From a sterile surface to formation of a robust biofilm without the addition of known bacteria (ie just leaving it to self populate) biofilm will normally occur within 2 to 3 months. This depends upon the species of bacteria available as some will form a biofilm more readily than others.

The speed at which biofilms are created is also largely down to environmental conditions, biofilm creation is essentially a defence response.

It needs to be made clear that a biofilm will not always be the end result of bacterial colonisation. If environmental conditions are favourable then it's unlikely one will be formed... instead bacteria will compete against one another.

So, in the case of a sponge filter in a sump... there is a good environment to grow, nutrients are readily available and the bacteria aren't under constant attack. It may well be that a biofilm in the sense we are talking about would never develop.

However, remove that sponge and put it in a QT with antibiotics then a biofilm will rapidly begin to develop as the bacteria communally respond to the attack.

So in short, it's fine to seed a sponge in the DT, but once it's been in a QT with meds it will need to be discarded... which is likely what we do in any case in the course of sterilising the QT.

 

[Humblefish]

More good info from @Gweeds1980 on this subject:

Well, copper wouldn't be impacted by biofilm and prazi really only needs 2-3 hours to do it's job. (I recommend at least 24 hours exposure time out of an abundance of caution.) So, the two main medications people use in QT are probably safe from biofilm.

However, Chloroquine phosphate, antibiotics, metronidazole, etc. require long-term exposure in order to be effective and can be degraded by biofilm. I am concerned this could be the reason why so many are losing fish to bacterial infections i.e. sometimes it seems like dosing antibiotics has little to no effect on an infection.

Just to clarify here... whilst the likes of copper and prazi won't be degraded by biofilm, it should be noted that both would likely trigger a defence response from bacteria so could accelerate biofilm formation.

In addition, you mention that biofilms could be responsible for the apparent ineffectiveness of antibiotics. This is true, biofilms are able to degrade antibiotics but in practice the bigger issue is that the antibiotics (even if they are effective against the bacteria on the fish) won't impact the bacteria within the biofilm. Thus you will likely have a remaining population of whatever bacteria infected said fish, within the biofilm. This makes it so much more important to break down and sterilise any QT if the purpose of it is to remove the risk of any diseases making it into the DT.

It would be next to impossible to remove a biofilm adhered to live rock or sand (within an aquarium... heating it up to a couple of hundred degrees c is another matter)... another good reason to stick with non porous, smooth surfaces only in a QT.

I would strongly recommend the use of biological washing detergent and high % h2o2 in the sterilisation process along with good old fashioned elbow grease to remove biofilms.

A good analogy is to consider the most familiar of biofilms - plaque. It doesn't matter how much expensive mouthwash you use, none of it will affect plaque in any meaningful way... give your teeth a darned good scrub and rinse with h2o2 and hey presto...

 

[Humblefish]

More info from @Gweeds1980:

There's no way of knowing really. And as @Gweeds1980 pointed out a particular biofilm might target a specific medication but leave all others alone. I would say the only way to be truly safe is to break down your QT in-between batches of fish and only use bacteria in a bottle to seed your QT.

Remember it's not just the issue of biofilms breaking down medication, it's that it could also harbour harmful bacteria which will be protected from antibiotic treatments.

Breaking down the QT is a sensible precaution as long as it is sterilised sufficiently. The majority of biofilms are visible, its the nooks and crannies which can be problematic. Using biological washing detergent and elbow grease will help as the enzymes will break down the biofilm, follow up with a good rinse with strong h2o2 to kill off anything remaining.

I am keen to point out that biofilms in general are not a bad thing... as posted elsewhere we rely on them for our nitrogenous waste processing. The danger in a QT environment is that harmful bacteria could be protected within the biofilm and / or the biofilm could break down medications rendering them useless.

 

[Blue Carbon Reefing]

@Humblefish Thank you for starting this thread! Have you started a thread on how to effectively break down a QT after use? after reading through this thread, it sounds like using Hydrogen peroxide would be the most effective at breaking down the biofilms that built up after having the QT up for a 2 -3 months while medicating and observing.

 

[Humblefish]

@Humblefish Thank you for starting this thread! Have you started a thread on how to effectively break down a QT after use? after reading through this thread, it sounds like using Hydrogen peroxide would be the most effective at breaking down the biofilms that built up after having the QT up for a 2 -3 months while medicating and observing.

@melypr1985 wrote this article (and did a video) on sterilizing quarantine equipment: https://www.reef2reef.com/ams/video-how-to-sterilize-a-quarantine-tank.201/

Wiping all surfaces down with hydrogen peroxide would be an extra step but should eliminate any biofilms.

 

[ShaunRobinson]

Interesting stuff folks. So what would be an average lifespan for a qt before it needs sterilising?

 

[Humblefish]

Interesting stuff folks. So what would be an average lifespan for a qt before it needs sterilising?

2-3 months

 

[Cment]

Wanted to add this great information provided by @Gweeds1980

From a sterile surface to formation of a robust biofilm without the addition of known bacteria (ie just leaving it to self populate) biofilm will normally occur within 2 to 3 months. This depends upon the species of bacteria available as some will form a biofilm more readily than others.

The speed at which biofilms are created is also largely down to environmental conditions, biofilm creation is essentially a defence response.

It needs to be made clear that a biofilm will not always be the end result of bacterial colonisation. If environmental conditions are favourable then it's unlikely one will be formed... instead bacteria will compete against one another.

So, in the case of a sponge filter in a sump... there is a good environment to grow, nutrients are readily available and the bacteria aren't under constant attack. It may well be that a biofilm in the sense we are talking about would never develop.

However, remove that sponge and put it in a QT with antibiotics then a biofilm will rapidly begin to develop as the bacteria communally respond to the attack.

So in short, it's fine to seed a sponge in the DT, but once it's been in a QT with meds it will need to be discarded... which is likely what we do in any case in the course of sterilising the QT.

I’m about to break down my QT and have a new batch of fish coming...so for clarification, it’s ok to seed the NEW QT with matrix from my DT? This will not carry over any type of biofilm and lessen the effectiveness of meds in the QT?

 

[Gweeds1980]

I’m about to break down my QT and have a new batch of fish coming...so for clarification, it’s ok to seed the NEW QT with matrix from my DT? This will not carry over any type of biofilm and lessen the effectiveness of meds in the QT?

There will likely be a biofilm developed on the matrix, if it's been in your DT system for 3 months or more.

What will you be treating the QT with?

 

[Rattzreef]

I really want a spectrophotometer now.

In addition to being able to measure CP in the water; if one were to see a rapid decrease after dosing fresh CP, one could assume biodegradation was occurring and it was high time to sterilize one's QT.

Great Idea. Then we can send you samples from our QT for verification also. Like an ICP test.

 

[Cment]

There will likely be a biofilm developed on the matrix, if it's been in your DT system for 3 months or more.

What will you be treating the QT with?

CP and everything else other than copper. Although, I’m thinking of doing copper now since biofilms don’t break it down.

 

[Gweeds1980]

CP and everything else other than copper. Although, I’m thinking of doing copper now since biofilms don’t break it down.

Lol... ok, assuming the matrix is older than 3 months, it could affect meds. Best bet would be to either seed new matrix in the display now or manage ammonia using prime and WCs until 'quick' meds are done, the add the matrix once copper started. If using CP, I'd suggest seeding new matrix now. Unfortunately there's no way you'll be able to tell if either the CP is being broken down or if any biofilm is even capable of doing that in the tank.

 

[Rattzreef]

This was a good read. Thanks to all that contributed. A lot of great information here. Would having separate systems for copper and meds be better than trying to use one tank for both? I have a 10g QT setup for almost 2 months now and was considering using meds after I was done with copper treatment but after reading through this I question whether or not that is the best course to go. Would setting up a new 20g tank, dosing bacteria in a bottle, and using meds for new additions be better? Then, tearing that tank down to sterilize until next use? Or would it be better to tear down the current QT also and sterilize it if biofilms can shelter undesirable bacteria?
Thanks for any info or opinions.

 

[Humblefish]

Great Idea. Then we can send you samples from our QT for verification also. Like an ICP test.

Unfortunately, I've learned (thanks to @Christoph) that using a spectrophotometer isn't always the best way to test Chloroquine in the water.

The two posts below explain in greater detail:

https://www.reef2reef.com/threads/chloroquine-phosphate.192309/page-71#post-5147777

https://www.reef2reef.com/threads/chloroquine-phosphate.192309/page-71#post-5147817