Introduction

Botrytis bunch rot caused by Botrytis cinerea Pers.:Fr. (teleomorph: Botryotinia fuckeliana (de Bary) Whetzel) is a major fungal disease of grapevine causing severe economic damage worldwide (Kassemeyer and Berkelmann-Löhnertz, 2009; Wilcox et al., 2015). Common bunch rot control strategies were traditionally based on the application of fungicides with known activity against B. cinerea (botryticides). Substantial evidence for a loss of fungicide efficacy due to resistance in B. cinerea was reported on several occasions [see for instance Leroch et al. (2011) or Walker et al. (2013)], illustrating the need for better fungicide resistance management and alternative control strategies beyond fungicides. In recent years, the high efficacy of non-chemical crop cultural measures such as pre-flowering or early post-flowering leaf removal in the cluster-zone (Molitor et al., 2011a; Poni et al., 2006; Poni et al., 2008; Sternad-Lemut et al., 2015), cluster division (Molitor et al., 2012), late first shoot topping (Molitor et al., 2015a), artificial shading (Basile et al., 2015), leaf anti-transpirants (Intrieri et al., 2013) and flower debris removal (Jaspers et al., 2016; Molitor et al., 2015b) has been demonstrated in different regions. The success of individual disease control measures is usually measured as first described by Abbott, 1925).

Being aware of the significant impact of annual meteorological conditions on bunch rot epidemics (González-Domínguez et al., 2015; Molitor et al., 2016), an estimation of the overall degree of control efficacy of complex strategies built on a set of modules is of interest for the annual bunch rot control strategy. Furthermore, it is crucial to avoid combining control methods that may act in an antagonistic manner with regard to the overall efficacy.

Ostensibly, a straightforward approach for estimating the cumulative efficacy (E_ab…x) of combined measures would be the accumulation (additive consideration) of the efficacies (E) of each single measure. This approach might deliver an acceptable estimation of the real efficacy at low efficacy levels and/or low numbers of measures combined. However, there is an obvious limitation to this approach at high efficacy levels and/or in case of high numbers of measures combined: the overall efficacy cannot, by definition, exceed 100%. However, theoretical efficacies above 100% might be reached when accumulating efficacies of several single measures. Furthermore, combining several control measures in other pathosystems indicated multiplicative rather than additive effects of combining control measures (Blandino et al., 2012; Edwards, 2004).

Consequently, we hypothesize that the efficacy of bunch rot control strategies combining two or more measures could be more correctly estimated based on the multiplicative consideration of the efficacies of single measures than based on additive consideration.

To test this hypothesis, three field examinations on the efficacy of three single non-chemical and/or chemical measures to control bunch rot as well as of all possible combinations of these measures were conducted and analyzed in the white Vitis vinifera L. cultivars Pinot gris and Riesling in the years 2009 and 2015 in Luxembourg.

Materials and Methods

Results and discussion

As shown in B. cinerea disease severities in the untreated control ranged from 14.6 (Pinot gris 2009) to 51.9% (Riesling 2015). Efficacies of single measures ranged from 9.9% (bioregulator; Pinot gris 2009) to 68.4% (leaf removal; P. gris 2015). Combining three crop cultural measures [namely, bioregulator + leaf removal + botryticide (trial A) or cluster division + leaf removal + late first shoot topping (trials B + C)] resulted in the numerically highest efficacies [range from 66.7% (Riesling 2015) to 96.9% (Pinot gris 2015)] (). Generally, efficacies of the measures in 2015 tended to be higher in Pinot gris than in Riesling. This might be explained by the dense cluster structure of Pinot gris grapes in 2015. Here, control measures might have been more efficient than in Riesling, which showed less compact clusters in 2015.

Assuming additive effects, average deviations per trial between estimated and observed efficacies ranged from -1.0% to -21.3% with average absolute deviations between 6.7% and 21.3%. Here, the global average deviation was -10.6% and global average absolute deviation 14.7% (). The negative average deviations in all three trials suggest that additive considerations tend to overestimate the overall efficacies. This effect is, as expected, most pronounced in case of combining measures with high efficacies, as this was the case particularly in trial B. Here, assuming additive effects leads to estimated efficacies above 100%, confirming the limitations of this approach.

In case of the multiplicative consideration, the average deviations per trial between estimated and observed efficacies of combined measures ranged from 5.1% to 6.3% with average absolute deviations between 6.4% and 9.0%. Here, the global average deviation was 5.5% and global average absolute deviation 7.9% ().

Estimated and observed efficacies were in both approaches significantly correlated. Coefficients of determination of linear regressions between estimated and observed efficacies were higher in case of the multiplicative consideration (R² = 0.8574; p < 0.0001) than in case of the additive consideration (R² = 0.8280; p < 0.0001) (). The multiplicative approach assumes that each additional measure is affecting (in case of efficient measures: reducing) the disease severity level as the result of the previous/additional treatments rather than compared to the disease severity level in the untreated control.

Generally, the high goodness of fit as well as the observed low deviations between the estimated and the observed efficacies demonstrated the suitability of the approach assuming multiplicative effects to estimate the efficacy of combined viticultural measures. Ratios between observed and estimated efficacies (E_obs./E_est.) > 1 mean that the overall efficacy of the combination of two or more measures is above the expected efficacy according to equation (3). Such ratios are indicating that besides multiplicative effects slight synergistic effects might exist, while, on the other hand, E_obs./E_est. ratios < 1 are indicating slight antagonistic effects. In the present investigations, both slightly synergistic as well as slightly antagonistic effects were observed in the different trials as well as in the different combinations. Generally, the fact that the global average ratio between observed and estimated efficacies of 1.10 was close to 1 demonstrates the usefulness of equation (3) with a slight tendency towards synergistic effects in some combinations: e.g., in treatments that combined leaf removal in the cluster-zone with other measures that lead to a reduction of the cluster compactness (such as cluster division or the application of a bioregulator), slightly synergistic effects were observed in all three trials, while in other combinations (e.g., cluster division + late first shoot topping), slight antagonistic effects (E_obs./E_est. < 1) were recorded (). In the case of combining leaf removal with a late first shoot topping in 2015, synergistic efficacies were observed in Pinot gris (E_obs./E_est. = 1.09), while the efficacies in Riesling (0.98) were slightly antagonistic. The question of which combinations of measures under which conditions in which cultivar tend to show (besides generally multiplicative effects) synergistic or antagonistic effects and the underlying principles would merit further investigations based on a broader data set. Potentially, combining different measures affecting the complex pathosystem grapevine/bunch rot at distant loci, in different ways or at distant time points (e.g., bioregulator (effect on cluster compactness) + botryticide (direct effect on pathogen); ) might tend to slightly synergistic effects while the combination of measures inhibiting the pathogen at similar positions (e.g., cluster division (effect on cluster structure) + late first shoot topping (effect on cluster structure); ) might exhibit slight antagonistic effects (efficacy lower than expected based on the multiplicative consideration of single efficacies).

Under practical conditions, a broad spectrum of crop cultural measures is available to optimize the grape health status and hence to enable a prolongation of the maturation period (Molitor et al., 2012). Present () as well as previous studies demonstrate that crop cultural measures (non-chemical control) are often of higher efficacy than the application of botryticides (chemical control) (Evers et al., 2010; Molitor et al., 2011b) and might save costs as well as energy (Sternad-Lemut et al., 2015). Consequently, such non-chemical crop cultural measures represent efficient tools for reducing or partially replacing the pesticide input in viticulture as intended in Integrated Pest Management. For instance, a chemical treatment (such as a botryticide application) can be replaced by an efficient crop cultural measure, without jeopardizing the efficacy of the bunch rot control regime. As an efficient risk management strategy, the combination of several measures is recommended in practical bunch rot control programs. According to the present results, the efficacy of the overall control strategy can be estimated based on multiplicative consideration of the expected effects of the single measures. Which and how many viticultural measures are integrated in the strategy is determined by the specific local conditions, the specific varietal degree of bunch rot susceptibility as well as the production target.

Acknowledgements: The authors would like to thank M. Schultz L. Vesque M. Pallez M. Behr D. Evers and B. Untereiner for their technical and scientific support, the Institut Viti-vinicole (IVV) and the Domaine Duhr Frères for providing the experimental vineyards and Mélanie Gollier (LIST) for helpful comments on an early version of the present manuscript. This work was carried out in the framework of the research projects “ProVino” and “BioViM” financially supported by the IVV in Remich/Luxembourg.