Pathogen derived resistance mediated by toxin of toxin-antitoxin system from Xylella fastidiosa

Sweet orange is one of the most important cultures produced in Brazil given its economic importance for national agriculture. However, this sector suffers big losses due phytosanitary problems as diseases and pests. Among them, the bacterial diseases such as Citrus Variegated Chlorosis (CVC) caused by Xylella fastidiosa, and citrus canker, caused by Xanthomonas citri subsp. citri generate big losses to sweet orange production. Among the technologies that aim to obtain resistance to plant pathogens, transgenesis can be used as a biotechnological tool to obtain tolerant/resistant plants for different pathogens. Among strategies of transgenesis, pathogen derived resistance (PDR) has been showed promising to control citrus pathogens. In this way, a possible candidate gene to confer tolerance against CVC and citrus canker would be the mqsR gene, which encodes a toxin of a toxin-antitoxin system from X. fastidiosa. This toxin is an endoribonuclease and was already described to abolish the pathogenicity of X. fastidiosa when the bacterium overexpress the toxin. In this way MqsR has potential for genetic transformation of plants aiming tolerance to X. fastidiosa, being able to inhibit growth, consequentely, reduce bacterial colonization and symptoms. X. citri does not have an homologous of this TA system in its genome, thus, transgenic plants overexpresing MqsR toxin could also interfere in X. citri colonization, decreasing the symptoms of citrus canker. In this study, were obtained transgenic plants of tobacco and citrus overexpressing the MqsR toxin, which were confirmed by GUS, PCR, gene and protein expression. These plants were challenged with X. fastidiosa and showed a reduction of symptoms when compared with non-transformed plants. Also it was verified that the population of X. fastidiosa was significantly smaller in transgenic plants, showing that the toxin possibly interfered with bacterial multiplication and colonization and, consequently, decreased the disease symptoms. It was also verified that the MqsR toxin could penetrate in X. citri cells and inhibit their growth. In detached leaves of transgenic citrus plants infiltrated with X. citri was verified a reduction of bacterial colonization and population, and the toxin was detected inside the bacterial cells, showing that, as observed in X. fastidiosa, the MqsR toxin altered the behavior of X. citri. Supporting the observed phenotype, it was showed a repression of genes associated with bacterial multiplication and colonization. In greenhouse conditions, it was also verified a reduction of severity of symptoms in transgenic plants in a time course experiment. Therefore, the findings suggest that the toxin overexpressed by the plant can be interfering with the multiplication and colonization of X. fastidiosa and X. citri, leading to tolerance to both pathogens (AU)