An endolysin gene from Liberibacter asiaticus confers dual resistance to huanglongbing and citrus canker.

The most damaging citrus diseases are Huanglongbing (HLB) and citrus canker, which are caused by Liberibacter asiaticus (Las) and pv. (), respectively. Endolysins from bacteriophages are a possible option for disease resistance in plant breeding. Here, we report improvement of citrus resistance to HLB and citrus canker using the LasLYS1 and LasLYS2 endolysins from Las. LasLYS2 demonstrated bactericidal efficacy against several Rhizobiaceae bacteria and , according to inhibition zone analyses. The two genes, driven by a strong promoter from , 35S, were integrated into Carrizo citrange via -mediated transformation. More than 2 years of greenhouse testing indicated that provided substantial and long-lasting resistance to HLB, allowing transgenic plants to retain low Las titers and no obvious symptoms while also clearing Las from infected plants in the long term. transgenic plants with improved HLB resistance also showed resistance to , indicating that LasLYS2 had dual resistance to HLB and citrus canker. A microbiome study of transgenic plants revealed that the endolysins repressed Xanthomonadaceae and Rhizobiaceae populations in roots while increasing Burkholderiaceae and Rhodanobacteraceae populations, which might boost the citrus defense response, according to transcriptome analysis. We also found that Lyz domain 2 is the key bactericidal motif of LasLYS1 and LasLYS2. Four endolysins with potential resistance to HLB and citrus canker were found based on the structures of LasLYS1 and LasLYS2. Overall, the work shed light on the mechanisms of resistance of Las-derived endolysins, providing insights for designing endolysins to develop broad-spectrum disease resistance in citrus.