Effects of Hot Water and Plant Growth Regulator Treatments on Bud Germination and Pathogen Elimination in Citrus Scions

The global citrus industry is under severe threat from devastating diseases, including Huanglongbing (HLB), caused by Candidatus Liberibacter asiaticus (CLas), and Citrus tristeza disease, caused by Citrus tristeza virus (CTV). Establishing efficient pathogen elimination techniques is crucial for developing virus-free citrus nursery systems and helps facilitate the prevention and control of devastating diseases. This study systematically evaluated the multiple effects of hot water treatment and plant growth regulator application on bud germination, pathogen elimination, antioxidant enzyme activities, and CsWUS expression in citrus scion. The results showed that moderate hot-water treatment at 40 °C for 5 min promoted bud germination, while higher temperatures or extended durations were inhibitory. Both 6-benzylaminopurine (6-BA) and melatonin (MT) significantly enhanced germination, with 50 mg/L 6-BA and 100 μM MT being most effective. All treatments significantly enhanced pathogen elimination and increased the activities of antioxidant enzymes SOD, POD, and CAT, with melatonin concentrations of 100 μM and above reducing CLas and CTV titers by over 93.64% and 96.07%. Furthermore, hot-water treatment and MT, but not 6-BA, upregulated the expression of the virus-resistance-related gene CsWUS. A combined treatment of hot water and MT did not yield synergistic benefits. In conclusion, melatonin treatment, particularly at 100 μM, optimally balances high bud germination with effective pathogen elimination, likely through activating antioxidant defenses and CsWUS-mediated immunity. This study provides a practical and efficient strategy for citrus pathogen-free seedling production.