Botrytis cinerea testing | Vintessential Wine Laboratories

Introduction

The following is a written version of a presentation by the author to a meeting of the Interwinery Analysis Group in Adelaide on 26 July 2013.

Botrytis cinerea – What is it?

Botrytis cinerea is a common mould that occurs on many fruits. If you have ever seen strawberries with a grey fuzzy mould on them, that is Botrytis cinerea.
It is the most common bunch rot in grapes (1), and, as in strawberries, produces a grey fuzzy mould growth.
The conditions (unfortunately well known to viticulturists) that are required for Botrytis growth are primarily wet weather. Of course this does not occur in all seasons or all regions, but it can be devastating when it does, such as in 2011.

What does Botrytis do to grapes?

The grey fuzzy growth caused by Botrytis on grapes is known widely as “grey rot” and occurs with prolonged wet weather. In certain circumstances, particularly when wet weather is followed by dry days, Botrytis can form what is known as “noble rot”. This is the form that gives rise to the famous Botrytised sweet wines (2).

The growth of Botrytis on the skins of grapes can, in some instances, lead to infection by other species. Penicillium and Aspergillis niger are two relatively common organisms that can subsequently develop. This unfortunate condition is known as “vulgar rot” (2).

Worse still, the skins of the berries can split and microbes such as yeasts and bacteria can invade, growing on the sugars that are released. The subsequent fermentation that may occur can give rise to acids, particularly acetic acid. This condition is known as “sour rot” (2).

What effect can Botrytis have on wine?

The metabolite produced by Botrytis that has been most widely discussed is the oxidative enzyme laccase that can cause browning of red wines. However, there are many other problems caused by Botrytis that seem to be have been ignored or neglected by oenologists and winemakers, until now.

These other metabolites include:

Glucans
Glycerol (up to 25g/L)
Gluconic acid (up to 5 g/L)
Methanol
Off flavor compounds

As well as producing these compounds, Botrytis can cause fermentation problems as it uses up nutrients and oxygen (3) in the juice that would normally be utilized by the yeast for the primary fermentation.

Of the above metabolites, the long chain polysaccharide glucans can be very problematic as they can cause issues with filtration (2).

Glycerol, whilst being fairly innocuous, can also be a problem, not just due to increased viscosity but more so because of the sweetness perception – it is about 70% as sweet as glucose (4).

Gluconic acid is an oxidation product of glucose metabolism caused primarily by Botrytis. It does add to the overall acidity of the wine as importantly it is not further metabolized during fermentation or wine aging (2).

Methanol is, of course, a very nasty compound and its concentration is limited by law.

How is Botrytis commonly measured?

In Australia the laccase activity test (5) is widely used for the presence of Botrytis. This test has some serious drawbacks; in fact a study is underway by the AWRI to investigate issues with this analysis method. The laccase test is also very limited as it only looks at one parameter when in fact Botrytis can cause many issues in wine.

There is another simple qualitative test that is commonly used known to many as the white plate test. A sample of must or wine is left overnight (on a white plate) to see if browning occurs. This is obviously only qualitative and does not tell a complete picture of what is going on.

What other techniques are available?

Other tests that can be used include:

Gluconic acid by enzymatic analysis (now available from Vintessential)
% incidence Botrytis by ELISA test strip
Glucans test
Yeast Assimilable Nitrogen (YAN)
Glycerol
Methanol
Acetic acid
Ethanol

As can be seen from this list there are a number of very good tests that can be used to monitor the impact of Botrytis cinerea.

In particular, gluconic acid is an important metabolite and has been used for many years in other wine producing countries (such as Spain, France and USA) as a marker for Botrytis infection. One winery in Spain uses this test on all parcels of fruit and reduces payments to growers if the gluconic acid level is above 0.6 g/L and rejects fruit on the basis of higher results (6). Due to interest initially from our Test Kit distributor in the USA, a Gluconic Acid test kit is now available from Vintessential.

Conclusion and testing strategy

Below is a suggested testing strategy that we have developed to better monitor the impact of Botrytis:
If grey rot is present on grapes test the following:

Gluconic acid
Laccase
% Botrytis incidence by Quickstix

This will give a solid base for further decisions on the parcel of fruit concerned.

If the Gluconic acid is >0.5g/L or vineyard monitoring shows a high % Botrytis incidence also test for:

Glucans
YAN

This strategy is simple, gives a much better overall picture than laccase testing alone, and is easily achievable with most current wine lab setups.

References
1. Margalit, Y; Concepts in Wine Chemistry, The Wine Appreciation Guild San Francisco, 2004, p369
2. Zoecklein, B.W, et al; Wine Analysis and Production, Aspen Publishers New York, 1995, pp62 -67
3. Boulton, R.B, et al: Principles and Practices of Winemaking, Springer Science+Business Media, Inc., New York, 1996, p218
4. Margalit, Y; Concepts in Wine Chemistry, The Wine Appreciation Guild San Francisco, 2004, p184
5. Iland, P, et al; Monitoring the winemaking process from grapes to wine: techniques and concepts, Patrick Iland Wine Promotions Pty Ltd, Campbelltown, SA, Aus. 2004, p88
6. Santos, J; Enartis Vinquiry, Windsor, USA, personal communication