Simultaneous editing of three homoeologues of TaCIPK14 confers broad-spectrum resistance to stripe rust in wheat.

Wheat stripe rust caused by the fungus Puccinia striiformis f. sp. tritici (Pst) is one of the most destructive wheat diseases resulting in significant losses to wheat production worldwide. The development of disease-resistant varieties is the most economical and effective measure to control diseases. Altering the susceptibility genes that promote pathogen compatibility via CRISPR/Cas9-mediated gene editing technology has become a new strategy for developing disease-resistant wheat varieties. Calcineurin B-like protein (CBL)-interacting protein kinases (CIPKs) has been demonstrated to be involved in defence responses during plant-pathogen interactions. However, whether wheat CIPK functions as susceptibility factor is still unclear. Here, we isolated a CIPK homoeologue gene TaCIPK14 from wheat. Knockdown of TaCIPK14 significantly increased wheat resistance to Pst, whereas overexpression of TaCIPK14 resulted in enhanced wheat susceptibility to Pst by decreasing different aspects of the defence response, including accumulation of ROS and expression of pathogenesis-relative genes. We generated wheat Tacipk14 mutant plants by simultaneous modification of the three homoeologues of wheat TaCIPK14 via CRISPR/Cas9 technology. The Tacipk14 mutant lines expressed race-nonspecific (RNS) broad-spectrum resistance (BSR) to Pst. Moreover, no significant difference was found in agronomic yield traits between Tacipk14 mutant plants and Fielder control plants under greenhouse and field conditions. These results demonstrate that TaCIPK14 acts as an important susceptibility factor in wheat response to Pst, and knockout of TaCIPK14 represents a powerful strategy for generating new disease-resistant wheat varieties with BSR to Pst.

Resistance assessment of pyraoxystrobin in Magnaporthe oryzae and the detection of a point mutation in cyt b that confers resistance.

Pyraoxystrobin is a new QoI fungicide developed in China. The present study was aimed at determining the baseline sensitivity of M. oryzae to pyraoxystrobin and investigating the potential resistance risk and resistance mechanism of pyraoxystrobin in M. oryzae. The results showed that the mean EC50 of 109 M. oryzae isolates to pyraoxystrobin was 0.0094 µg/mL and the sensitivity exhibited a unimodal distribution. The established baseline sensitivity could provide critical data for monitoring sensitivity changes of M. oryzae to pyraoxystrobin in rice fields. The potential resistance risk was assessed by investigating the biological characteristics of the resistant mutants obtained by fungicide adaptation. The results indicated that the resistance risk of pyraoxystrobin in M. oryzae was medium to high with positive cross-resistance between pyraoxystrobin and azoxystrobin, but without cross resistance between pyraoxystrobin and carbendazim, isoprothiolane, and prochloraz. Further investigation revealed that the pyraoxystrobin-resistant mutants had a G143S mutation in the cyt b protein. Molecular docking confirmed that the G143S substitution conferred high resistance to pyraoxystrobin in M. oryzae. Collectively, the results of this study provided essential data for monitoring the emergence of resistance and developing resistance management strategies for pyraoxystrobin.

Three point-mutations in cytochrome b confer resistance to trifloxystrobin in Magnaporthe oryzae.

BACKGROUND: Rice blast, caused by Magnaporthe oryzae, is the most devastating disease in rice. Recently, trifloxystrobin was registered for the control of M. oryzae in China. The resistance profile and mechanism of M. oryzae to trifloxystrobin were investigated in the present study, providing important data for the recommended use of trifloxystrobin. RESULTS: The baseline sensitivity was established at a half maximal effective concentration (EC50 ) of 0.024 µg mL-1 . Nine stable trifloxystrobin-resistant mutants were generated with EC50 values ranging from 12.75 to 171.49 µg mL-1 . The mutants exhibited strong adaptive traits in sporulation, conidial germination, and pathogenicity. Positive cross-resistance was only observed between trifloxystrobin and azoxystrobin, but not between trifloxystrobin and carbendazim, isoprothiolane, prochloraz, or chlorothalonil. The point mutation G143S in cytochrome b (cyt b) protein was found in eight high-resistance mutants with resistant factor ranging from 2295.16 to 13 200.00; and the double mutation G137R/M296V only occurred in Mg117-1 with resistance factor ≈ 900. The G143S mutation weakened hydrogen bond interactions, and G137R/M296V changed the conformation of trifloxystrobin in the cyt b binding pocket. A molecular detection method was established for the rapid detection of G143S mutants in M. oryzae. CONCLUSION: The resistance risk of M. oryzae to trifloxystrobin could be moderate to high. Two genotypes with three point-mutations G143S, G137R, and M296V conferred resistance to trifloxystrobin in M. oryzae. © 2020 Society of Chemical Industry.