Genetic study of resistance to strobilurins in isolates of Pseudocercospora fijensis from banana plantations in Costa Rica

Black Sigatoka, caused by the fungus Pseudocercospora fijiensis Morelet, is considered the most limiting foliar disease for banana and plantain production worldwide and with the greatest economic impact for the export banana industry. Chemical combat has been the main tool for managing this disease, however, due to the strong selection pressure exerted on the pathogen populations and the frequent use of the same active ingredients, the fungus has developed resistance to the main fungicides. specific sites released to the market, among them the external quinone inhibitors (QoI's), also known as strobilurins. As a consequence of the development of resistance, the number of applications of multi-site action fungicides has increased, with the consequent increase in the cost of combating the disease and the risk to the environment and health. With the aim of knowing in more detail the genetic bases of the resistance of P. fijiensis to strobilurins, the degree of in vitro sensitivity of sixty-five isolates of the pathogen (55 from commercial farms and 10 from a wild population) was determined. to azoxystrobin and pyraclostrobin. Subsequently, by sequencing, the presence or absence of mutations G143A, F129L and G137R associated with the loss of sensitivity of different phytopathogenic fungi to strobilurins was identified in P. fijiensis isolates. In addition, by reverse transcription and real-time PCR (polymerase chain reaction), the relative expression of the gene was quantified in 2 sensitive isolates and 8 isolates resistant to this group of fungicides. Finally, under greenhouse conditions, the pathogenicity and virulence of five isolates of P. fijiensis sensitive (G143) and five resistant (G143) to strobilurins were evaluated. When determining the degree of in vitro sensitivity to strobilurins, the loss of sensitivity to azoxystrobin and pyraclostrobin was confirmed in isolates of P. fijiensis from banana plantations in Costa Rica. Likewise, it was possible to identify the cross-resistance between them. Through the molecular evaluation of the cyt b gene, it was possible to identify the G143A mutation in 100% of the isolates from commercial farms that showed a loss of in vitro sensitivity to strobilurins, which reveals a clear relationship between this and resistance to this. group of fungicides in P. fijiensis. On the other hand, mutations G137R and F129L were not found in any of the isolates, suggesting their non-existence for P. fijiensis and even less their possible relationship with resistance to strobilurins in this pathogen. In addition, it was possible to quantify an overexpression of the cyt b gene in P. fijiensis isolates with loss of in vitro sensitivity to strobilurins, which reveals that this molecular mechanism may also be involved in the resistance of P. fijiensis to this family of fungicides. Finally, under greenhouse conditions, all P. fijiensis isolates were found to be pathogenic. However, the resistant isolates showed equal or greater virulence than the sensitive isolates. The information generated in this research provides valuable information in the knowledge of the genetics of P. fijiensis resistance to strobilurins. Information that could be used to molecularly monitor resistance to strobilurins in pathogen populations and contribute to the design of more efficient disease control programs