Cross-Resistance Between <i>Pseudomonas chlororaphis</i> Strain AFS009 Metabolites (Howler EVO) and Fludioxonil in <i>Botrytis cinerea</i>

Howler EVO is a biological fungicide based on metabolites of the bacterium <i>Pseudomonas chlororaphis</i> strain AFS009. One of the metabolites, pyrrolnitrin, is a chemical analogue of the phenylpyrrole fludioxonil used to manage gray mold of fruit crops caused by <i>Botrytis cinerea</i>. Resistance to fludioxonil in <i>B. cinerea</i> is well documented and linked to mutations in the transcription factor <i>mrr1</i>, leading to overexpression of the ATP-binding cassette transporter gene <i>BcatrB</i>. Moderately resistant isolates are designated MDR1 and MDR1h based on the specific variation of mutations in <i>mrr1</i> and the level of <i>BcatrB</i> expression. This study investigated concentration inhibiting 50% mycelium growth (EC₅₀) values of 54 <i>B. cinerea</i> isolates sensitive and with moderate resistance to fludioxonil for sensitivity to fludioxonil and Howler EVO. The Pearson correlation coefficient indicated a strong correlation between EC₅₀ values of fludioxonil and Howler EVO. Isolates that were moderately resistant to fludioxonil and classified as MDR strains were also moderately resistant to Howler EVO. The effect of Howler EVO and fludioxonil on <i>BcatrB</i> gene expression was studied by quantitative PCR. Both fungicides induced the <i>BcatrB</i> gene expression significantly up to 100-fold in sensitive <i>B. cinerea</i> isolates. Howler EVO significantly induced the <i>BcatrB</i> gene expression in all MDR1 isolates but not in the MDR1h isolate. In detached fruit assays on cherry, sensitive <i>B. cinerea</i> isolates were completely inhibited by formulated fludioxonil (Scholar) and significantly suppressed in growth by Howler EVO. However, MDR1 and MDR1h isolates produced disease in Scholar and Howler EVO treatments. Our results indicate cross-resistance between the synthetic fungicide fludioxonil and the biofungicide Howler EVO, indicating that, at least for some biofungicides, resistance management is necessary.