Seasonal variation and detection frequency of Candidatus Liberibacter asiaticus in Binchuan, Yunnan province China

Munir, Shahzad; Li, Yongmei; He, Ping; He, Pengjie; Cui, Wenyan; Wu, Yixin; Li, Xingyu; He, Yueqiu

doi:10.1016/j.pmpp.2019.01.004

Cited by 8 publications

(33 citation statements)

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“…Healthy trees had more endophytes compared to symptomatic and asymptomatic trees, which may have been because the www.nature.com/scientificreports www.nature.com/scientificreports/ healthy trees were free of CLas. Previous studies reported that microbial communities in citrus trees are negatively affected by CLas 37,41,42 . In addition, microbial colonization of the branches, stems, roots, and leaves are affected by various factors.…”

Section: Discussionmentioning

confidence: 98%

“…Samples were subsequently processed for PCR template preparation. DNA was extracted using the cetyl trimethylammonium bromide (CTAB) method with slight modifications 36,37 . Briefly, the midribs of leaves were separated and frozen in liquid nitrogen.…”

Section: Methodsmentioning

confidence: 99%

“…The recombinant plasmid (pUC18-382) solution (10 µl) was used to transform 50 µl competent Escherichia coli TG1 cells (Takara) for 8-10 h at 4 °C. The resulting mixture was spread onto Luria Bertani (LB) plates containing ampicillin and positive clones were confirmed by PCR using CLas-specific primers 37 . The positive recombinant plasmid was extracted using a HiBind DNA Mini Column (Omega), following the instructions for the Plasmid DNA Mini Kit (Omega).…”

Section: Construction Of Recombinant Plasmid and Standard Curvementioning

confidence: 99%

“…All collected citrus leaves were investigated for the presence of CLas with CLas-specific primers 37 . The number of CLas copies was different between symptomatic and asymptomatic plants, with symptomatic plants having lower cycle threshold (CT) values and more CLas copies.…”

Section: Detection Of Clasmentioning

confidence: 99%

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Core endophyte communities of different citrus varieties from citrus growing regions in China

Munir

Huang

et al. 2020

Sci Rep

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The native microbiomes of citrus trees play important roles in plant health, with good communication between the native microbiome and the host plant. Here, we report on the native endophytes in 24 citrus varieties in nine citrus growing regions in China; some of the trees were healthy and others had asymptomatic or symptomatic huanglongbing, which is caused by the pathogen Candidatus Liberibacter asiaticus (CLas). We used culture-dependent analysis and characterized the isolates by partial 16S rRNA gene sequencing. The endophytes were compared between different citrus varieties, regions, and disease states (healthy, asymptomatic, and symptomatic). The total number of endophytes isolated from most of the citrus varieties was 10 4-10 6 CFU/g of leaves, but it differed significantly by disease state, with the highest numbers in the healthy leaves and the lowest in the symptomatic leaves (p < 0.05). Among the citrus varieties, the Valencia variety had the maximum number of endophyte species (22). The most dominant endophytes were Bacillus subtilis, B. velezensis, Curtobacterium luteum, and Microbacterium testaceum. The higher frequency of B. subtilis in the healthy/asymptomatic plants compared to the symptomatic plants suggests that it has a role in huanglongbing resistance. Native endophyte communities in various citrus varieties could be used to improve citrus growth and combat CLas.

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Section: Discussionmentioning

confidence: 98%

Section: Methodsmentioning

confidence: 99%

Section: Construction Of Recombinant Plasmid and Standard Curvementioning

confidence: 99%

Section: Detection Of Clasmentioning

confidence: 99%

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Core endophyte communities of different citrus varieties from citrus growing regions in China

Munir

Huang

et al. 2020

Sci Rep

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“…Biocontrol of plant pathogens has yielded effective results using bacteria under laboratory conditions [18][19][20] and moderate effects under eld conditions [21], but the potential of indigenous bacteria as economically useful biocontrol agents of plant pathogens has not been investigated. This approach may be used to manage citrus disease caused by phloem-limited α-proteobacteria [2], because there is evidence for negative effects on pathogen titers of Clas of changes in microbe population dynamics during HLB disease progression [22][23][24].…”

Section: Introductionmentioning

confidence: 99%

Restructuring citrus endophytic diversity through potential indigenous endophytes could eliminate huanglongbing pathogen Candidatus Liberibacter asiaticus

Munir¹,

Li²,

He³

et al. 2020

Preprint

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Background Huanglongbing (HLB) is a major botanical pandemic of citrus crops caused by Candidatus Liberibacter asiaticus (Clas). It is important to understand the different mechanisms involved in interaction of pathogen with plants to develop novel management strategy against HLB. However, until now there has been no control strategy to manage this disease in vitro and on large scale in citrus grove. We found that, indigenous endophyte Bacillus subtilis L1-21, a patented strain isolated from healthy citrus tree, may have the potential to reduce the impact of pathogen through restructuring of core endophytes. Results A novel half-leaf method was developed to test the efficacy of B. subtilis L1-21 against Clas. Concentration of B. subtilis L1-21 at 104 cfu ml− 1 resulted in a 1000-fold reduction in Clas copy densities per gram of leaf midrib (107 to 104) by 4 d after treatment. With endophytes, where HLB incidence was reduced to < 3% and Clas copy density was reduced from 109 to 104 pathogen g− 1 of diseased leaf midrib. We found that 16 of 93 tree samples became Clas-free and functional pathways and pathogen resistance genes were regulated in diseased citrus trees after treatment. Conclusions This is the first large-scale study using an indigenous endophyte and shows its potential utility in sustainable disease management through strengthening the citrus microbiome.

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Potential of citrus endophyte Bacillus subtilisL1‐21 in the control of Candidatus Liberibacter asiaticus in Asian citrus psyllid, Diaphorina citri

et al. 2022

Pest Management Science

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BACKGROUND Asian 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. RESULTS Initially, 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. CONCLUSIONS We 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.

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Seasonal variation and detection frequency of Candidatus Liberibacter asiaticus in Binchuan, Yunnan province China

Cited by 8 publications

References 49 publications

Core endophyte communities of different citrus varieties from citrus growing regions in China

Core endophyte communities of different citrus varieties from citrus growing regions in China

Restructuring citrus endophytic diversity through potential indigenous endophytes could eliminate huanglongbing pathogen Candidatus Liberibacter asiaticus

Potential of citrus endophyte Bacillus subtilisL1‐21 in the control of Candidatus Liberibacter asiaticus in Asian citrus psyllid, Diaphorina citri

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