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ARS Home » Midwest Area » East Lansing, Michigan » Sugarbeet and Bean Research » Research » Publications at this Location » Publication #302341
Title: Use of PCR-RFLP analysis to monitor fungicide resistance in Cercospora beticola populations from sugarbeet (Beta vulgaris) in Michigan, United States

Author
item ROSENZWEIG, N - Michigan State University
item Hanson, Linda
item FRANC, G - University Of Wyoming
item STUMP, W - University Of Wyoming
item JIANG, Q - Michigan State University
item CLARK, G - Michigan Sugar Company
item STEWART, J - Michigan Sugar Company
item KIRK, W - Michigan State University

Submitted to: Plant Disease
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 9/4/2014
Publication Date: 3/1/2015
Citation: Rosenzweig, N., Hanson, L.E., Franc, G.D., Stump, W.L., Jiang, Q.W., Clark, G., Stewart, J., Kirk, W.W. 2015. Use of PCR-RFLP analysis to monitor fungicide resistance in Cercospora beticola populations from sugarbeet (Beta vulgaris) in Michigan, United States. Plant Disease. 99:355-362.

Interpretive Summary: Genetic resistance to two classes of fungicides may be responsible for recent declines in efficacy of chemical control management strategies for Cercospora leaf spot (CLS) caused by Cercospora beticola (Sacc.) in Michigan sugarbeet fields. The target genes and fungicide resistance mutations are known for these two fungicides. Based on this, two PCR-restriction fragment length polymorphism (RFLP) assays were developed to detect the point mutations in the fungal genes responsible for resistance. These mutations confer a high level of resistance to either quinone outside inhibitor (QoI) or benzimidazole fungicides. The presence of the mutations was monitored within C. beticola populations recovered from Michigan sugarbeet production fields in 2012. Both resistant alleles were detected directly from leaf tissue using the PCR-RFLP assay. The QoI- and benzimidazole-resistance mutations were detected in over 90% of the 118 field samples originating from Michigan sugarbeet production. While some of these fields had a mixture of sensitive and resistant isolates, over 60% of the fields yielded only isolates with the QoI resistance mutation and 80% of the fields yielded only isolates with the benzimidazole resistance mutation. Monitoring of the mutations in fields located in 9 counties and 58 townships indicated that the mutations were widespread in Michigan sugarbeet production areas. Results indicated a high level of resistance to both fungicides in the population. The PCR-RFLP assays developed and used in this study were effective in detecting the presence of the specific mutations in C. beticola field populations from Michigan. Both resistance mutations are present at high levels in the Michigan population. No evidence was found for the presence of other mutations in the C. beticola population to these two fungicide classes.

Technical Abstract: Genetic resistance to Quinone outside inhibitor (QoI) and benzimidazole fungicides may be responsible for a recent decline in efficacy of chemical control management strategies for Cercospora leaf spot (CLS) caused by Cercospora beticola (Sacc.) in Michigan sugarbeet (Beta vulgaris L.) fields. The target genes and fungicide resistance mutations are known for these two fungicides. Based on this, two standard PCR-restriction fragment length polymorphism (RFLP) assays were developed to detect the G143A and E198A point mutations in the fungal mitochondrial cytochrome b and the ß-tubulin genes respectively. These mutations confer a high level of resistance to either QoI or benzimidazole fungicides. The presence of the G143A and E198A mutations was monitored within C. beticola populations recovered from Michigan sugarbeet production fields collected in 2012. Both the Quinone outside inhibitor (QoI)-resistant cytochrome b allele and the benzimidazole-resistant ß-tubulin allele were detected directly from leaf tissue following a polymerase chain reaction restriction fragment length polymorphism (PCR-RFLP) assay. Using either detection assay, the G143A and E198A mutations were detected in over 90% of the 118 field samples originating from Michigan sugarbeet production under fungicide management programs for CLS control. Monitoring of the G143A and E198A mutations in fields located in 9 counties and 58 townships indicated that the mutations were widespread in Michigan sugarbeet production areas. Results indicated a high level of resistance to both fungicides in the population. The PCR-based assays used and developed in this study were effective in detecting the presence of the G143A and E198A mutations in C. beticola field populations from Michigan. Both resistance mutations are present at high levels in the Michigan population.