Barley powdery mildew caused by the fungus Blumeria graminis f. sp. hordei (Bgh) has emerged as the most damaging disease of barley in Western Australia (WA). Many of the available cultivars display high levels of disease in the field when the climatic conditions are conducive. As a result, fungicides have become the main method of disease control in the last 10 years. Different types and sources of genetic disease resistance are available but to optimise their deployment it is necessary to evaluate the spectrum of pathotypes present in the pathogen population.Sixty isolates of Bgh were collected in the 2009 season from 9 locations, single spored and characterised by infection on reference barley lines and cultivars, 18 unique pathotypes were collected. Virulence against many of the R-genes in the reference lines was present in at least one pathotype. Isolates were virulent against 16 out of a total of 23 resistance gene combinations. Undefeated resistance genes included the major R-genes Mla-6, Mla-9, Ml-ra and the combinations of Mla-1 plus Mla-A12 and Mla-6 plus Mla-14 and Mla-13 plus Ml-Ru3 and the recessive resistance gene mlo-5. There was significant pathotype spatial differentiation suggesting limited gene flow between different regions with WA.On the basis of the results we recommend a number of strategies to manage powdery mildew disease levels within WA.

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The increased occurrence of triazole fungicide resistant strains of Blumeria graminis f. sp. hordei (Bgh) is an economic concern for the barley industry in Australia and elsewhere. High levels of resistance to triazoles in the field are caused by two separate point mutations in the Cyp51 gene, Y136F and S509T. Early detection of these mutations arising in pathogen field populations is important as this allows time for changes in fungicide practices to be adopted, thus mitigating potential yield losses due to fungicide failure and preventing the resistance from becoming dominant. A digital PCR (dPCR) assay has been developed for the detection and quantification of the Y136F and S509T mutations in the Bgh Cyp51 gene. Mutation levels were quantifiable as low as 0.2% in genomic DNA extractions and field samples. This assay was applied to the high throughput screening of Bgh field and bait trial samples from barley growing regions across Australia in the 2015 and 2016 growing seasons and identified the S509T mutation for the first time in the Eastern states of Australia. This is the first report on the use of digital PCR technology for fungicide resistance detection and monitoring in agriculture. Here we describe the potential application of dPCR for the screening of fungicide resistance mutations in a network of specifically designed bait trials. The combination of these two tools constitute an early warning system for the development of fungicide resistance that allows for the timely adjustment of management practices.

BACKGROUND Powdery mildew caused by Blumeria graminis f. sp. hordei (Bgh) is a constant threat to barley production but is generally well controlled through combinations of host genetics and fungicides. An epidemic of barley powdery mildew was observed from 2007 to 2013 in the West Australian grain belt. RESULTS We collected isolates across Australia, examined their sensitivity to demethylation inhibitor (DMI) fungicides and sequenced the Cyp51B target gene. Five amino acid substitutions were found, of which four were novel. The most resistant haplotypes increased in prevalence from 0% in 2009 to 16% in 2010 and 90% in 2011. Yeast strains expressing the Bgh Cyp51 haplotypes replicated the altered sensitivity to various DMIs and these results were complemented by in silico protein docking studies. CONCLUSIONS The planting of very susceptible cultivars and the use of a single fungicide mode of action was followed by the emergence of a major epidemic of barley powdery mildew. Widespread use of DMI fungicides led to the selection of Bgh isolates carrying both the Y137F and S524T mutations, which, as in Zymoseptoria tritici, account for resistance factors varying from 3.4 for propiconazole to 18 for tebuconazole, the major azoles used at that time in WA. © 2019 The Authors. Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

The barley powdery mildew pathogen, Blumeria graminis f. sp. hordei (Bgh), exists in numerous haplotypes and displays significant differences in fungicide sensitivity. It causes considerable yield losses throughout the world. Microsatellite SSRs are useful tools to study the population level and biogeographic aspects of intraspecific diversity, but so far none have been defined for Bgh. Here, eight polymorphic microsatellite loci were identified and characterized. Primer pairs amplifying the loci were then applied to 111 isolates of Bgh from Australia. The number of alleles per locus ranged from 4 to 13, and Nei's genetic diversity ranged from 0·25 to 0·76. The microsatellite primers detected several clones among the isolates and defined 97 unique haplotypes. There was little evidence for regional genotypic subdivision, suggesting that gene flow may not be restricted among geographic regions. All data was consistent with high levels of genetic diversity, potentially resulting from random mating and spread within each region.