Development of a quantitative PCR-based method for the detection and monitoring azoxystrobin resistance in Pyricularia oryzae populations

Rice blast, caused by Pyricularia oryzae, is one of the most important rice diseases worldwide, causing economically important rice yield losses and representing a threat to global food security. Rice blast management relies on fungicides, especially in Europe, as susceptible rice varieties are intensively grown for their quality value. Azoxystrobin (a Quinone outside inhibitor fungicide, QoI) represents often the first – and sometimes the only –choice of chemical control of rice blast. However, it is a fungicide with a high risk of resistance development in pathogen populations. Indeed, P. oryzae azoxystrobin-resistant strains from rice have been identified in Japan, which poses concerns about the spread of QoI resistance also in other rice-growing areas. Despite this, no reliable and sensitive detection method of QoI resistance in P. oryzae populations exists at the moment. We developed two qPCRbased methods for the detection of QoI-resistant strains of P. oryzae. One is based on selective amplification of sensitive (S) or resistant (R) allele using S- and R-specific primers, while the other exploits high resolution melting analysis of the PCR product. Here, we evaluate and compare the sensitivity and specificity of the two methods using DNA mixtures of S and R strains at different ratios. This method is to be used where QoI application is the predominant means of rice blast management, to monitor the emergence of P. oryzae QoI-resistant strains.