Bacterial Diversity Analysis of Huanglongbing Pathogen-Infected Citrus,
Using PhyloChip Arrays and 16S rRNA Gene Clone
Uma Shankar Sagaram,1‡ Kristen M. DeAngelis,2‡ Pankaj Trivedi,1 Gary L. Andersen,2 Shi-En Lu,3 and Nian Wang1*
Citrus Research and Education
Center, Department of Microbiology and Cell Science, University
of Florida/IFAS, 700 Experiment Station Road, Lake Alfred,
The bacterial diversity associated with citrus leaf midribs was characterized for citrus groves that contained
the Huanglongbing (HLB) pathogen, which has yet to be cultivated in vitro. We employed a combination of
high-density phylogenetic 16S rRNA gene microarrays and 16S rRNA gene clone library sequencing to
determine the microbial community composition for symptomatic and asymptomatic citrus midribs. Our
results revealed that citrus leaf midribs can support a diversity of microbes. PhyloChip analysis indicated that
47 orders of bacteria in 15 phyla were present in the citrus leaf midribs, while 20 orders in 8 phyla were
observed with the cloning and sequencing method. PhyloChip arrays indicated that nine taxa were significantly
more abundant in symptomatic midribs than in asymptomatic midribs. “Candidatus
was detected at a very low level in asymptomatic plants but was over 200 times more abundant in symptomatic
plants. The PhyloChip analysis results were further verified by sequencing 16S rRNA gene clone libraries,
which indicated the dominance of “Candidatus
Liberibacter asiaticus” in symptomatic leaves. These data
Liberibacter asiaticus” as the pathogen responsible for HLB disease.
Keywords:Candidatus Liberobacter spp.,;high-density phylogenetic 16S rRNA gene microarrays;Cuscuta campestris;Catharanthus roseus;16S rRNA gene clone libraries.
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