Zhicheng Ding ¹ Yang Liu ² Shaoran Zhang ³ Fangkui Wang ³ Zong Qi ³ Anna Du ² Yiajie Zheng ² Jian Zhu ² Ling Jiang ^4*

• ¹ Department of Horticulture and Forestry, Huazhong Agricultural University National Key Laboratory of Germplasm Innovation and Utilization of Horticultural Crops,, Huazhong Agricultural University, Wuhan, China
• ² Department of Horticulture and Forestry, Huazhong Agricultural University National Key Laboratory of Germplasm Innovation and Utilization of Horticultural Crops, Huazhong Agricultural University, Wuhan, China
• ³ State Key Laboratory of Agricultural Microbiology, Huazhong Agricultural University, Wuhan, China
• ⁴ Department of Horticulture and Forestry, Huazhong Agricultural University National Key Laboratory of Germplasm Innovation and Utilization of Horticultural Crops, National Fruit Free-virus Germplasm Resource Indoor Conservation Center, Huazhong Agricultural University, Wuhan, China

Global citrus production has been severely affected by citrus Huanglongbing (HLB) disease and the development of effective control methods are crucial. This study used antimicrobial lipopeptide and phytohormone complex powder (L1) prepared from the fermentation broth of the endophytic plant growth-promoting bacterium (PGPB) of Bacillus amyloliquefaciens strain MG-2 to treat Candidatus Liberibacter asiaticus (CLas)-infected 'Citrus reticulata 'Chun Jian' plants. Real-time fluorescence quantitative polymerase chain reaction (qPCR) and PCR were employed for detection. The results revealed that after 15 spray-drench treatments with L1 solution, the HLB infection rate decreased from 100% to 50%, the bacterial titer decreased by 51.9% after L1 treatment compared with a 27.9% decrease in the control group, after 18 spray-drench treatments with L1 solution, the bacterial titer decreased by 57.2% after L1 treatment compared with 31.9% decrease in the control group. L1 treatment triggered the production of reactive oxygen species, increased lignin content, and increased defense enzyme activities (p<0.05). Defense-related gene expression significantly increased within 12 h of treatment. In addition, L1 application also promoted plant growth, as evidenced by higher transpiration rates and net photosynthetic rates as well as increased leave or root density. Root flora analysis after L1 treatment revealed that the abundances of Burkholderia_thailandensis, unclassified_g_Burkholderia-Caballeronia-Paraburkholderia, unclassified_g__Allorhizobium-Neorhizobium-Pararhizobium-Rhizobium, and Pseudomonas_mosselii were 1.64, 1.46, 5.84, and 6.93 times greater, respectively, than those in the control group. The levels of phenylpropanoids, polyketides, lipids, lipid-like molecules, organic acids, and derivatives, significantly increased following L1 treatment (FC>2, P<0.05). Additionally, salicylic acid, dihydrojasmonic acid, and isopentenyl adenosine levels in leaves markedly increased. High-performance liquid chromatography (HPLC) confirmed that L1 contained surfactin, iturin and fengycin cyclic-lipopeptides (CLPs) as well as indole-3-acetic acid (IAA), 3-indolebutyric acid (IBA), indole-3-carboxylic acid and indole-3-carboxaldehyde auxins, N6-entopentenyladenine and t-zeatin-riboside cytokinins, abscisic acid, 1-aminocyclicpanecarboxylic acid, salicylic acid, and gibberellin A1, A3 and A4 phytohormones. These findings provide insight into multiple mechanisms by which endophytic Bacillus PGPB L1 is able to combat HLB disease, to promote citrus plant growth, and to optimize the root flora for soil health which offering an innovative strategy for sustainable management of this severe disease and improving citrus plant growth and productivity.