Abstract
The bacterial microbiomes of citrus plants were characterized in response to ‘Candidatus Liberibacter asiaticus’ (Las)-infection and treatments with ampicillin (Amp) and gentamicin (Gm) by Phylochip-based metagenomics. The results revealed that 7,407 of over 50,000 known Operational Taxonomic Units (OTUs) in 53 phyla were detected in citrus leaf midribs using the PhyloChip™ G3 array, of which five phyla were dominant, Proteobacteria (38.7%), Firmicutes (29.0%), Actinobacteria (16.1%), Bacteroidetes (6.2%) and Cyanobacteria (2.3%). The OTU62806, representing ‘Candidatus Liberibacter’, was present with a high titer in the plants graft-inoculated with Las-infected scions treated with Gm at 100 mg/L and in the water-treated control (CK1). However, the Las bacterium was not detected in the plants graft-inoculated with Las-infected scions treated with Amp at 1.0 g/L or in plants graft-inoculated with Las-free scions (CK2). The PhyloChip array demonstrated that more OTUs, at a higher abundance, were detected in the Gm-treated plants than in the other treatment and the controls. Pairwise comparisons indicated that 23 OTUs from the Achromobacter spp. and 12 OTUs from the Methylobacterium spp. were more abundant in CK2 and CK1, respectively. Ten abundant OTUs from the Stenotrophomonas spp. were detected only in the Amp-treatment. These results provide new insights into microbial communities that may be associated with the progression of citrus huanglongbing (HLB) and the potential effects of antibiotics on the disease and microbial ecology.
Highlights
Huanglongbing (HLB), the most devastating citrus disease worldwide, is vectored by phloem-feeding insects and caused by fastidious bacterial pathogens (Candidatus Liberibacter) [1,2,3]
This study provides a comprehensive survey of the richness and composition of microbial communities in the leaf midribs of HLB-affected citrus and healthy citrus plants as well as HLB-affected scions treated with antibiotics in greenhouse
These include an increase from 500,000 to 1.1 million probes, the inclusion of strain-specific probe sets, the ability to detect over 50,000 Operational Taxonomic Units (OTUs) instead of,9,000 OTUs [18], and the utilization of over 320,000 sequences in the reference database, which is over 10 times greater than that for the PhyloChipTM G2
Summary
Huanglongbing (HLB), the most devastating citrus disease worldwide, is vectored by phloem-feeding insects and caused by fastidious bacterial pathogens (Candidatus Liberibacter) [1,2,3]. Soon after a bacterium was reported to be associated with HLB, antibiotics were first used to control the pathogen. Different types of antibiotics, such as tetracycline and penicillin, were injected into infected citrus trees to temporarily relieve HLB symptoms and decrease Las bacterial titers [10]. Different kinds of antibiotics were tested for efficacy against the HLB bacterium while assessing their phytotoxicity to citrus. The microbial community structure in Las-infected field citrus plants treated with the above antibiotic combination has been analyzed. Bacterial cells in close proximity may be able to modify their microenvironment, making the composition of the microbial community an important factor in the ability of Las to cause HLB progression [14]. Some plant growth-promoting bacteria, such as Bacillus and Burkholderia, were detected in non-infected leaf samples [15], while bacteria such as Methylobacterium and Sphingobacterium were present in root samples from HLB-affected trees [16]
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