Wang, Yuehu

AbstractBACKGROUNDAsian citrus psyllid (ACP), also known as Diaphorina citri, is the natural vector of Candidatus Liberibacter asiaticus (CLas), which is responsible for Huanglongbing (HLB), a devastating citrus disease. Previously, the pathogen was successfully excluded from diseased citrus plants by using the indigenous endophyte Bacillus subtilis L1‐21. However, the pathogen elimination and colonization potential of B. subtilis L1‐21 in the carrier vector ACP, as well as the recruitment of native microbial communities of psyllid in the presence of endophytes, are still unknown.RESULTSInitially, we suggested that endophyte L1‐21 reduced the CLas copies in ACP from 6.58 × 106 to 5.04 × 104 per insect after 48 h, however, the pathogen copies remained stable in the negative control. The endophyte was stable for 48 h after application. Among the bacterial genera those highlighted in ACP were Candidatus Liberibacter, Pseudomonas, Candidatus Profftella, Methylobacterium‐Methylorubrum, Pantoea, Curtobacterium, Wolbachia, Actinomycetospora, and Bacillus. Interestingly, B. subtilis L1‐21 easily colonizes the midgut of ACP but cannot be detected in eggs. When ACP with endophyte L1‐21 was allowed to feed on new citrus leaves, the highest colonization was observed. We also found that psyllids carrying endophyte L1‐21 after feeding on citrus leaves reduced the CLas copies in leaves on the 0, 3rd and 5th day from 8.18 × 10,4 2.6 × 10,3 and 0 pathogen copies/g fresh midvein, respectively.CONCLUSIONSWe propose that B. subtilis L1‐21 is a native endophyte in citrus and psyllid, which efficiently reduces the CLas pathogen in both citrus and psyllids, provides a more protective effect by increasing the number of cultivable endophytes, and successfully colonizes the midgut of ACP. © 2022 Society of Chemical Industry.

Huanglongbing (HLB) has turned into a devastating botanical pandemic of citrus crops, caused by Candidatus Liberibacter asiaticus (CLas). However, until now the disease has remained incurable with very limited control strategies available. Restoration of the affected microbiomes in the diseased host through the introduction of an indigenous endophyte Bacillus subtilis L1-21 isolated from healthy citrus may provide an innovative approach for disease management. A novel half-leaf method was developed in vitro to test the efficacy of the endophyte L1-21 against CLas. Application of B. subtilis L1-21 at 104 colony forming unit (cfu ml−1) resulted in a 1,000-fold reduction in the CLas copies per gram of leaf midrib (107 to 104) in 4 days. In HLB-affected citrus orchards over a period of 2 years, the CLas incidence was reduced to < 3%, and CLas copies declined from 109 to 104 g−1 of diseased leaf midribs in the endophyte L1-21 treated trees. Reduction in disease incidence may corroborate a direct or an indirect biocontrol effect of the endophytes as red fluorescent protein-labeled B. subtilis L1-21 colonized and shared niche (phloem) with CLas. This is the first large-scale study for establishing a sustainable HLB control strategy through citrus endophytic microbiome restructuring using an indigenous endophyte.