Take home messages

• Of the three principal fungicide modes of action used regularly for broadacre disease control:
• Group 11 quinone-outside inhibitors (QoIs) (strobilurins) are at the highest risk of pathogen resistance development, particularly the pathogens responsible for Septoria tritici blotch (STB) in wheat, and spot form of net blotch (SFNB) and powdery mildew in barley.
• Group 7 succinate dehydrogenase inhibitors (SDHIs) are at moderate to high risk of resistance development in the pathogen with evidence in New Zealand (NZ) and Europe of pathogen shifts in sensitivity to Ramularia leaf spot in barley and net form of net blotch (NFNB) and STB in Europe.
• Group 3 demethylase inhibitors (DMIs – triazoles) are generally considered at low to moderate risk, however recent developments in WA have challenged this view.
• Recent evidence from WA in the net blotch pathogens has revealed sinister mutations in the DMI target site governed by the Cyp51A gene that result in the over production of the target leading to highly resistant strains of net blotch.
• Use integrated disease management (IDM) measures (rotations, seed hygiene, resistant varieties, cultural control) to minimise the number of fungicide applications in a season. Ideally no more than one foliar application of the QoIs and SDHIs per season and no more than two DMIs.
• Never repeat the use of the same fungicide product or active ingredient in a growing season.
• Optimise foliar fungicide timings by considering the importance of the key yield bearing leaves of the crop — top four leaves in barley and top three in wheat.
• Continued monitoring of disease pathogens by the Centre for Crop and Disease Management (CCDM) is essential for early detection of pathogen resistance.

New fungicide active ingredients for better disease control

Over the past decade, there has been an unprecedented change in the fungicide armoury available to Australian growers and their advisers. At the start of the millennium, available fungicides in Australia were primarily from the older classes of DMI chemistry. Fungicide availability was limited and product introductions were a decade behind Europe. That has all changed and Australian broadacre croppers have much earlier access to newer chemistry at prices that allow far greater use. In part this availability is thanks to the manufacturers and GRDC who not only recognised the need to benchmark the sensitivity of common pathogens against available fungicide products, but also the need to encourage the registration of new active ingredients for the Australian market. The GRDC New Fungicide Actives project led by Curtin University (Project CUR 00019 and the new bilateral between Curtin/GRDC Program 9) in collaboration with the Foundation of Arable Research (FAR) Australia has worked with different target diseases in cereals and other major broadacre crops. This research has generated efficacy data that combined with manufacturers’ data has led to the registration of new fungicides with new modes of action. Now that there is access to newer fungicides, such as Group 7 (SDHIs) and Group 11 (QoIs), it comes with a responsibility to look after these chemistries as although they are very effective, they are also at high risk of the pathogen’s developing resistance and reduced sensitivity.

What is the current status of fungicide resistance and reduced sensitivity in Australia?

Over the past six years, the CCDM has been working with industry and other researchers to establish a fast, cost-effective monitoring system for the common diseases of broadacre grains crops. Current cases of fungicide resistance and reduced sensitivity in Australian broadacre crops are outlined in Table 1.

Table 1. Fungicide resistance and reduced sensitivity cases identified in Australian broadacre grains crops.

DMI fungicide resistance in NFNB (Pyrenophora teres f. teres) in WA

Since 2013, isolates of the pathogen causing NFNB have been detected with reduced levels of sensitivity to several DMI fungicides. Strains were classified as either sensitive, moderately resistant (MR), or highly resistant (HR) based on 50% effective concentrations (EC₅₀) to the Group 3 compounds — prochloraz, difenoconazole, propiconazole, prothioconazole, tebuconazole and epoxiconazole (Table 1). MR strains were first found in the Great Southern (Kojonup) region in 2013 and subsequently detected throughout the WA wheatbelt and Great Southern regions. Highly resistant isolates were found in the Esperance (Scaddan), southern wheatbelt (West Arthur) and central wheatbelt (Dandaragan) regions from 2017 onwards.

Table 2. Mean 50% effective concentrations (EC₅₀) and resistance factors of Group 3 MR isolates, HR isolates, and eight sensitive NFNB isolates collected 1996–2012, to the fungicides tebuconazole, epoxiconazole and prothioconazole.

Analysis of thegene for the DMI target, called Cyp51A, in resistant isolates revealed the presence of a point mutation, F489L, in the coding sequence found in all resistant isolates and not in any sensitive isolates. In MR isolates, a single copy of the F489L mutated allele was present. In HR isolates, up to 10 additional copies of the mutated allele were detected, indicating over production (or expression) of the Cyp51A gene with the F489L mutation.

DMI fungicide resistance in SFNB (Pyrenophora teres f. maculata)

Isolates of the pathogen causing SFNB with reduced levels of sensitivity to several DMI fungicides have been detected in WA since 2016. In vitro testing sorted strains into sensitive, MR, and HR groups based on 50% effective concentrations (EC₅₀) to the Group 3 compounds prochloraz, difenoconazole, propiconazole, prothioconazole and tebuconazole.The MR and HR groups showed a similar level of reduced sensitivity to epoxiconazole (Table 2). MR strains were detected from the Esperance region (Gibson and Munglinup) from 2016 onwards, and HR isolates were found in the Great Southern (South Stirling and Wellstead) and Esperance (Dalyup) regions from 2017 onwards.

Table 3. Mean effective concentration 50 (EC₅₀) in µg/ml of Group 3 MR isolates, HR isolates, and a reference population of sensitive SFNB isolates collected between 1996 and 2013. Resistance factors (fold number difference between EC₅₀ values from resistant isolates and average of sensitive isolates) are shown in brackets. Cultures were grown at different concentration ranges of the fungicides tebuconazole, epoxiconazole, prothioconazole and propiconazole.

^aMean of 20 sensitive SFNB isolates; ^bmean of a subset of three sensitive SFNB isolates.

Analysis of the target gene for the Group 3 (DMI) fungicide, Cyp51A, revealed the presence in MR and HR isolates of two different mutations that were not observed in sensitive isolates. In MR isolates, the mutation was a small fragment of DNA that was inserted in the fungicide target gene. This small fragment of new DNA was found at three different positions and its effect was over-production of the fungicide target. The presence of more fungicide target requires an increase in the amount of fungicide necessary to kill the MR isolates.

The small DNA fragment was also found in HR isolates, but at a different position, together with another mutation, F489L. This latter mutation has been previously observed in the closely related pathogen P. teres f. teres, the causative agent of NFNB, where it has been correlated with reduced sensitivity to a range of DMI fungicides (Mair et al. 2016).

MR strains were detected from the Esperance region (Gibson and Munglinup) from 2016 onwards, and HR isolates were found in the Great Southern (South Stirling and Wellstead) and Esperance (Dalyup) regions from 2017 onwards.

These new mutations in the net blotch pathogen in WA populations are sinister as they represent only the second example in Australia of gene overproduction (expression), the first being in NFNB pathogen in 2016 (Mair et al. 2016).

Anti-resistance measures when using fungicides

Clearly the best way to avoid fungicide resistance is not to use fungicides. However, in an IDM approach, when a variety’s genetic resistance breaks down or is incomplete, it is imperative that growers and advisers have access to a diverse range of fungicides (in terms of mode of action) for controlling the disease. The main anti-resistance measures in cereal crops that growers can adopt when using fungicides are:

• To minimise the number of fungicide applications using the same mode of action using other IDM measures to assist in controlling disease (rotations, seed hygiene, resistant varieties, cultural control and grazing used in conjunction with less fungicides).
• Avoid repeat applications of the same product and mode of action in the same crop and/or year after year, particularly when using QoI (strobilurins) and SDHIs alone, such as the seed treatment fluxapyroxad (Systiva®).
• Never apply the same DMI (triazole) Group 3 fungicide twice in a row.
• Avoid using tebuconazole as a stand-alone product in barley for any disease to avoid indirect fungicide resistance selection.
• Triazoles used alone are best reserved for less important spray timings targeted at less important plant structures that have historically given poorer economic returns and in situations where disease pressure is low.
• Ideally, use DMI-based mixtures (e.g. Prosaro® containing prothioconazole and tebuconazole) only once, followed by mixtures containing other actives (preferably from Groups 7 or 11).
• Ideally, apply no more than one QoI (strobilurin) or one SDHI in the course of a growing season (the current limit for registered foliar products is two applications of each mode of action in a season with seed treatments effective on foliar disease counting as one of those applications).
• For most scenarios in Australia, with the exception of Tasmania and some parts of the high rainfall zone (HRZ), two broad spectrum fungicide inputs should be regarded as a maximum in order to control the vast majority of disease outbreaks in susceptible varieties, with most crops requiring none or one application.
• With SDHI seed treatments that have activity on foliar diseases later in the spring, such as fluxapyroxad (Systiva®), consider foliar fungicide follow ups, which have a different mode of action and avoid repeat usage of Systiva® alone in successive years.

Influence of fungicide rate

The reality is that using the most appropriate rate for effective disease control (lower label rates for lower disease pressure and higher label rates for higher disease pressure) is the best strategy for managing resistance. Label rates have been developed to provide robust and reliable control of the target disease.

In many cases, the full label rate is the most appropriate rate for control. However, for diseases, the lower rate from the label range of a fungicide can be used in conjunction with a crop variety that has a good disease resistance rating, because disease pressure will be lower. There is evidence that using a higher rate than necessary increases the risk of resistance as removing all of the sensitive pathogen isolates provide more opportunity for resistant isolates to dominate the population and hence be the strain colonising the plant.

Conclusion

In summary, there are some exciting new fungicides available to us as an industry. Whilst we should not be afraid to use these agrichemicals when necessary, their longevity needs to be protected by considering the number of times they are used in a growing season. The evidence in Australia is increasing to suggest that whilst there is more rapid access to new fungicides than 15 years ago, there are also more issues with fungicide resistance. With the sound principles of IDM applied and a good monitoring system, we can at least slow down the development of resistance and increase the longevity of the fungicides being used.

References

FRAG-UK, 2018. Fungicide Resistance Management in Cereals.

https://cereals.ahdb.org.uk/media/175998/frag-18-fungicide-resistance-management-in-cereals.pdf

GRDC fact sheet, 2012. Barley Powdery Mildew.

https://grdc.com.au/__data/assets/pdf_file/0028/24958/grdc-fs-barley-powdery-mildew.pdf.pdf

GRDC Media Release, 2018. Fungicide resistance found in barley spot form of net blotch.

https://grdc.com.au/news-and-media/news-and-media-releases/west/2018/02/fungicide-resistance-found-in-barley-spot-form-of-net-blotch

GRDC Media Release, 2018. Research uncovers new changes in fungicide resistance in WA barley.

https://grdc.com.au/news-and-media/news-and-media-releases/west/2018/07/research-uncovers-new-changes-in-fungicide-resistance-in-wa-barley

Mair WJ, Deng W, Mullins JGL, West S, Wang P, Besharat N, Ellwood SR, Oliver RP & Lopez-Ruiz FJ, 2016. Demethylase Inhibitor Fungicide Resistance in Pyrenophora teres f. sp. teres associated with Target Site Modification and Inducible Overexpression of Cyp51. Front. Microbiol. 7:1279. doi: 10.3389/fmicb.2016.01279.

Acknowledgements

The research undertaken as part of this project is made possible by the significant contributions of growers through both trial cooperation and the support of the GRDC — the authors would like to thank them for their continued support.

Contact details

Nick Poole and Kat Fuhrmann
FAR Australia
23 High St, Inverleigh, Victoria 3221
03 5265 1290
nick.poole@faraustralia.com.au

Fran Lopez-Ruiz
Curtin University, Centre for Crop and Disease Management
School of Molecular and Life Sciences
Perth WA 6845
08 9266 3061
fran.lopezruiz@curtin.edu.au