The influences of citrus apoplast pH on Xanthomonas citri subsp. citri invasion and canker formation

The pH plays a key role in the growth and colonization of plant pathogens as well as the onset and progression of the symptoms they cause within the host. Plants may quickly alter their apoplastic pH (pH_apo) to protect themselves against infection. However, pathogens can also alter the pH of their ambient environment to promote their own growth. Citrus canker is a serious plant disease caused by Xanthomonas citri subsp. citri (Xcc). This Gram-negative aerobic rod is usually cultured in Luria–Bertani (LB) medium at pH 7. However, little is known about the changes in pH both in this medium as Xcc grows and in the leaf apoplast in response to Xcc infection and colonization. Moreover, the differences in leaf apoplast pH between Xcc-resistant and Xcc-susceptible citrus genotypes are also unknown. Here, Xcc grew well in liquid LB medium at initial pH 6–8 and the pathogen altered the medium pH to 6.8 ± 0.4. Xcc growth declined at pH 5 and was zero at pH 3, 4, 9, and 10. In susceptible sweet orange infected with Xcc inoculum, canker symptoms were inhibited at pH 3, 4, and 10 but did not differ in the range of pH 5–9. As expected, canker symptoms were absent at all inoculum pH in the resistant Citron C-05. For both genotypes, Xcc only grew well in the leaves exposed to pH 5–8 inoculums. At four days post-inoculation (4 dpi), the foliar pH_apo of resistant Citron C-05 had rapidly declined from 5.6 to 4.4. At 2 dpi, the pH_apo of susceptible sweet orange had rapidly increased from 5.6 to 6.7, Xcc grew quickly, and canker symptoms appeared. Plasma membrane (PM) H⁺-ATPase activation with fusicoccin (FC) acidified the apoplast and upregulated the pathogenesis-related genes (PRs) in the sweet orange leaves. Hence, Xcc colonization and canker development were inhibited. The results of this study revealed that apoplastic acidification is implicated in the resistance of Citron C-05 to Xcc infection and provided insight into the association between pH_apo regulation and resistance to bacterial pathogen invasion in plants.