I have noticed recently that there has been a lot of concern about the possibility of mould developing resistance to fungicides.  A number of  tanners have begun alternating fungicides on a six month about regimen. Is this necessary?

There is a good reason NOT to alternate. One of the two fungicides will be less cost effective.  Using a more expensive treatment costs you money, and using a less effective treatment may add to the risk of failure.

We need to understand that there are two types of resistance.  Inherent and acquired.
Inherent resistance is that which the mould has built in.  For example; the mould Trichoderma viride which mainly grows on wood is quite resistant to TCMTB. It has not acquired this. This resistance has always been present. It is inherent.

However, the resistance of the skin infection “golden staph” to antibiotics is definitely acquired.  20 years ago, any antibiotic would easily kill these bacteria.  Today, the resistant strains are almost impossible to kill. This resistance is acquired.

Alternating fungicides on a six monthly basis will be effective only against acquired resistance - not inherent.

 What are the key requirements for the acquiring of resistance?

1.  The dose of  chemical to kill the mould must be too small - a sub lethal dose that kills most, but not all of the bugs.

2.  The repeated doses of sub lethal fungicide must be against the original population. Not against new infections from outside.

3. Enough generations must pass.  How many?  Laboratory research shows that  200 are needed for bacteria. Perhaps 20 for mould?

How do these principles apply to a tannery or pelt house?  For a mould that infects leather to acquire resistance, a single population must be exposed to a sub lethal fungicide dose over at least 20 generations.

A tanner would have to continue to use an ineffective treatment for 20 generations (over 3 years), with mould growing on all batches, and leaving the mouldy stock inside the plant to keep reinfecting the new production. You are all too competent to allow this to happen.  I think it is clear that the likelihood of resistance to fungicide being acquired is low.  To prevent it happening:

1.  Use an effective fungicide at an effective level
2.  Remove mouldy wet blues or pickled pelts so they cannot infect new batches.
3.  Six month alternations of fungicide?  Yes.  But only if we do not follow rules 1 & 2

So what are the real causes of failure by a fungicide, and the subsequent expensive mould growth?

1.  Process variables.  Sometimes we use a fungicide in the wrong way, or change a chemical process to make it incompatible with a fungicide.  Some examples.

a.  Adding TCMTB into high pH degrades this fungicide
b.  Recycling liquor and thus increasing grease levels. Some fungicides like octyl isothiazolone are grease loving and will be removed.
c.  Adding phenols into low pH conditions will reduce their effectiveness.

2.  A new mould species may appear that has inherent resistance to the fungicide. This can affect any fungicide at all.  Some examples.

a.  A tannery in years gone by was using Trinap 40.  A farmer built a hay shed upwind of the tannery.  Suddenly mould was all through the plant and growing over all wet blue production. A change to TCMTB fixed this problem.
b.  Several tanneries I am aware of which used TCMTB have imported Trichoderma viride (probably on mouldy wooden pallets) and been hit with subsequent mould outbreaks.
c.  Another tannery in New Zealand using a phenolic blend as fungicide had an outbreak of Amorphotheca resinae - inherently resistant to phenolics.

3.  Lousy housekeeping.  A tannery or pelthouse should be kept clean.  This has worker health and safety implications as well as affecting the mould risk. High spore counts in a plant lead to high levels of attack on wet blues or pelts.  Unclean plants have organic matter build up with mould growing on this rubbish, causing these dangerous high spore counts.

Note that the column labelled MAX is what I consider to be the maximum average spore count a bovine wet blue tannery should see before trouble strikes! The figure should be a bit higher for a pelt house, and a bit lower for ovine wet blue production.

You will observe the seasonal changes - rising in spring and falling in autumn.  There is also shown an anomalous reading for July - showing a temporary problem. Forget the theory!

How do we get practical and stop these outbreaks? What is the best fungicide treatment?

This will vary from plant to plant. The only way to find out is by trials with the leading fungicides. I will talk about how to run such a trial shortly.  Find the very best, and most cost effective fungicide.  Use this product all the time, since it gives you the best results.

However, it is important also to choose a secondary fungicide, in case one of these inherently resistant moulds turn up.  This product will be one that you have trialled and proven to work.  It will be one that can be brought in quickly. It will be held in stock in New Zealand.  It will be sufficiently different from the primary fungicide, so that it will not have the same weaknesses against specific moulds.

Even though eight to ten different products have been trialled at various times in New Zealand, right now there are only four in regular use.

TCMTB     [2(thiocyanomethylthio)benzothiazole as 30% emulsifiable concentrate]

OITZ [octyl isothiazolone as 10% solution]

MBT [sodium mercaptobenzothiazole as 50% solution]

Phenolic blend [para chloro meta cresol -PCMC- at 30% plus ortho phenyl phenol - OPP-at 15%]

Carbendazim [currently available in several forms] This is not currently used much, but I believe it has great potential.

1. TCMTB has various advantages.

• The “TT” factor (tried and true).  This is the most used product in New Zealand, and the most trusted. It has to be seen as the standard against which other products are measured.
• Superb performance in pelt houses.  Look no further than TCMTB. It works exceptionally well at the low pH levels of pickle solution.
• Very good performance at low doses.
• Very good broad spectrum of activity.
• Readily available at good prices.

2.  OITZ is also an excellent product, and is very similar to TCMTB in many ways (in spite of being chemically quite different).  It has the same advantages of high potency at low doses, and a broad spectrum (although my tests show a couple of worrying gaps).  Like TCMTB, it appears to have a weakness against Trichoderma. It is now available at a reasonable price. It is also fat loving and prone to problems from process variables.  I suspect it is also going to prove useful at about half of our tanneries.   It also causes skin irritation.

Because TCMTB and OITZ are so similar in performance, I would not recommend that they be chosen as primary and secondary fungicides.  This is because if one fails to perform, the other might not also.  Instead use one as primary, and a totally different product such as the phenolic blend or MBT as secondary.

3.  MBT has great potential and has proven itself on several sites.  I have personally been involved in four trials, and each succeeded.  It is inexpensive (half the TCMTB price). About double the TCMTB dose is required, so it works out about equally cost effective. I have yet to see any major problems from skin irritation.

MBT appears (so far) to be more forgiving in relation to process variables.  For example, it can be added at both high and low pH levels. It appears to be a lot less fat loving than either TCMTB or OITZ.  It has certainly solved some very difficult mould problems. It is also a good choice as primary fungicide.

4.  The phenolic blend has a high TT rating. It has been used for a long time, and with great success. It has solved a lot of problems for tanners where TCMTB has failed.

A unique disadvantage of phenolics is that it is not possible to check their performance in a tropical chamber. They all (including the old Trinap 40) seem to fail in the tropical chamber, while they succeed in the real world. It is only recently that we have found out why.  Phenolics do have the odd gap in their spectrum.  This includes several fungi that thrive in tropical chambers.

The PCMC/OPP blend has the added disadvantage that a high dose, and hence a high cost is involved. While having a good broad spectrum of activity, there are still several moulds that have high inherent resistance. Tim Allsop pointed this out in a paper several years ago, and it is still worth looking at again. This has led to failure on several occasions in recent history.

For these reasons, I would regard the phenolic blend to be an ideal choice as secondary fungicide, but only after your primary choice has failed.

5.  Carbendazim is the one product I added to the list which is not widely used in New Zealand. I believe this situation will change, however.  Carbendazim is an ideal fungicide additive in that it is cheap, potent, and adds nicely to the spectrum of activity of several fungicides. It is also very low in toxicity and non irritant to skin.  My own tests show an excellent spectrum of activity, and high potency at low doses.

Buckman Laboratories already has a product that is a blend of TCMTB and carbendazim.  I have tested it, and it works very well. It has an excellent spectrum of activity. To render carbendazim into liquid form in combination with TCMTB is a costly process, and unfortunately, this excellent product is very expensive to use..

I believe that there are much less expensive alternatives which will allow the benefit of carbendazim to be realised.  For example, it is possible to obtain an inexpensive 50% wettable powder for separate addition.  This has yet to be trialled. However, my own feeling is that a supplementary addition of cheap carbendazim will give a very big added insurance factor and solve some real existing problems.

What is the spectrum of activity of these products?  This is usually measured as the MIC.  This is a laboratory test that measures how much fungicide is needed to kill specific fungi. Suppliers all will readily provide tables of MIC results.

I have been a little suspicious of these tables.  New Zealand fungi are different to the fungi used overseas to make up these tables. For example, it used to be thought that there were six species of Trichoderma.  DNA testing has shown that the real number is over 200.  Thus, it is highly unlikely that New Zealand Trichoderma viride is the same as American Trichoderma viride, and the MIC tables are not accurate for New Zealand.

I have been playing with MIC tests and the results are below.  Fungi I have used are :

A.  Amorphotheca resinae
B.  Aspergillus flavus
C.  Aspergillus niger
D.  Fusarium culmorum
E.  Paecilomyces variottii
F.  Penicillium chrysogenum
G.  Penicillium glabrum
H.  Trichoderma viride

MBT has high potency at higher doses, and a spectrum very similar to the phenolic blend.

OITZ seems to be a bit more potent than MBT, and almost as good as TCMTB.

The phenolic blend, as stated before shows a need for higher doses, but has a good activity against Trichoderma.

So, looking at all the factors, what is the ideal treatment as primary fungicide?
There is still work to be done, and especially to look at carbendazim as a supplement.  Before anyone can determine their ideal treatment, they must run trials.

With these proviso’s, my suggested “ideal” treatment would be a combination of TCMTB, OITZ, or MBT with a supplementary dose of cheap carbendazim.  Please trial the system thoroughly first.

This raises the question of how to run an ideal fungicide trial.  This is not a new skill for too many of you, so I will not insult your intelligence by talking about this in detail.  A couple of points are worth making, however.

The actual trial, of course, consists mostly of simply replacing the usual fungicide with the new product at recommended amounts. The real problems comes with evaluating results. Let me make a couple of suggestions.

• You may, quite rightly, feel a bit nervous about trusting a whole batch of valuable wet blues or pelts to an untried fungicide treatment.  You can add extra insurance by removing samples for mould resistance testing, and then adding extra fungicide and running the batch for a while longer.  TCMTB works very well for this as it is absorbed very quickly.
• The number of samples removed for testing is crucial.  Two or three pieces will not give good results.  The raw material we work with is not homogeneous. One piece is different to another.  We need enough samples to get good averages.  Let me illustrate with a real set of results. (Numbers show the percentage of sample surface covered with mould after 29 days in a tropical chamber)

 

Fungicide X	Fungicide Y
0	0
1	0
2	0
3	2
3	2
10	2
10	5
20	5
40	5
50	5

Fungicide X  0,1,2
Fungicide Y 5,5,5

Please use at least 10 samples per trial batch.

We use accelerated laboratory tests to evaluate mould resistance, which will not reflect reality entirely accurately. However, they are the best tools we have and are generally quite reliable, as long as we understand their limitations.

1.  The LASRA test.  This involves placing a small test sample onto a sterile disc on agar in a petri dish, inoculating the agar with known fungal species, and incubating for 90 days.  If the sample remains free of mould for 90 days, it is said to pass the test.

This method certainly has a high TT factor, and is well trusted.  It has the big advantage that LASRA is seen to be unbiased, and very professional in their approach. This test is the standard by which others are measured. Certainly, something as important as a fungicide trial should use the LASRA test.

It has the disadvantage that it takes 90 days.  For a trial, this is OK, but is probably too long to be useful as a quality check on normal production. There is also the disadvantage of exposure to a limited number of species.

Beware of anyone running this test by placing the sample in direct contact with agar.  If the fungicide leaches, they will show a “zone of inhibition” which must be seen as product failure - not success! Fungicides must not leach.

Also be aware that the choice of mould species here is crucial.  An unscrupulous operator can ensure success for his product by selecting moulds that are always killed. Another good reason to use LASRA.
 

2. The tropical chamber.  This is a modified insulated cabinet (I use an old fridge) heated to 25 to 30 degrees Celsius; kept at very high humidity; with fungal innoculum added; and spores circulated with a low power fan.  Samples hung in the chamber grow mould very quickly.  After a while, such a chamber becomes a real unpredictable mixture of many mould species.

Advantages of the chamber include:

• the speed to get results.  My chamber will predict a failed treatment within two weeks.  This makes it excellent for quality assurance needs.
• It also has the advantage of exposing samples to many species of mould.

No-one expects a tanner or fellmonger to be an expert at fungicides.  Each of you should be prepared to ask for assistance from LASRA or from chemical suppliers. Your choice of fungicide treatment is vital, and you should expect those who help you to spend the time to make sure proper and complete trials are run.

I am now available for questions.

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