Sclerotinia sclerotiorum is one of the most damaging and economically important necrotrophic plant pathogens, infecting more than 400 plant species globally. Although the phenylpyrrole fungicide
fludioxonil has high activity against S. sclerotiorum, reports indicate that there is also substantial potential for the development of fungicide resistance. However, the current study investigating five
fludioxonil-resistant laboratory mutants found a significant fitness cost associated with
fludioxonil resistance resulting in significantly (P < 0.05) reduced mycelial growth and sclerotia formation on potato
dextrose agar as well as significantly (P < 0.05) lower pathogenicity on detached tomato leaves, with one mutant, LK-1R, completely losing the capacity to cause
infection. In addition, all of the
fludioxonil-resistant mutants had significantly (P < 0.05) increased sensitivity to osmotic stress (0.5 M of
potassium chloride and 1.0 M of
glucose), which is consistent with the proposed
fludioxonil target sites within the high osmolarity
glycerol stress response
mitogen-activated protein kinase (HOG1-MAPK) signaling transduction pathway. Sequence analysis of six genes from this two-component pathway, including SsHk, SsYpd, SsSk1, SsSk2, SsPbs, and SsHog, revealed several mutations that may be associated with
fludioxonil resistance. For example, six separate point mutations were found in SsHk that led to changes in the predicted amino acid sequence, including A136G, F249V, G353A, E560K, M610K, and K727R. Similarly, SsPbs had three mutations (D34G, S46L, and L337E), SsSk1 and SsYpd had two (S53G and A795V for SsSk1, and E67G and Y141H for SsYpd), and SsHog and SsSk2 had one each (V220A and S763P, respectively). To our knowledge, these constitute the first reports of
amino acid changes in
proteins of the HOG1-MAPK pathway being associated with
fludioxonil resistance in S. sclerotiorum. This study also showed a positive cross-resistance between
fludioxonil and dimethachlone and
procymidone, but none with
tebuconazole or
carbendazim, indicating that the inclusion of
tebuconazole within an integrated pest management program could reduce the risk of
fludioxonil resistance developing in field populations of S. sclerotiorum and ensure the sustainable production of soybeans in China into the future.