Dataset Information

Asian citrus psyllid proteome: whole insect samples, with or without Candidatus Liberibacter asiaticus

ABSTRACT: ‘Candidatus Liberibacter asiaticus’ (CLas), the bacterial pathogen associated with citrus greening disease, is transmitted by Diaphorina citri, the Asian citrus psyllid (ACP). We used quantitative mass spectrometry to compare the proteomes of CLas(+) and CLas(-) populations of D. citri. This experiment used adult mixed sex whole insect samples as starting material for protein extraction. DNA extracted from CLas(+) ACP was tested by qPCR to confirm presence of CLas.

INSTRUMENT(S): Orbitrap Fusion

ORGANISM(S): Diaphorina Citri (asian Citrus Psyllid)

TISSUE(S): Whole Body

SUBMITTER: John Ramsey

LAB HEAD: Michelle Cilia

PROVIDER: PXD003096 | Pride | 2018-10-25

REPOSITORIES: Pride

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Publications

Metabolic Interplay between the Asian Citrus Psyllid and Its Profftella Symbiont: An Achilles' Heel of the Citrus Greening Insect Vector.

Ramsey John S JS Johnson Richard S RS Hoki Jason S JS Kruse Angela A Mahoney Jaclyn J Hilf Mark E ME Hunter Wayne B WB Hall David G DG Schroeder Frank C FC MacCoss Michael J MJ Cilia Michelle M

PloS one 20151118 11

'Candidatus Liberibacter asiaticus' (CLas), the bacterial pathogen associated with citrus greening disease, is transmitted by Diaphorina citri, the Asian citrus psyllid. Interactions among D. citri and its microbial endosymbionts, including 'Candidatus Profftella armatura', are likely to impact transmission of CLas. We used quantitative mass spectrometry to compare the proteomes of CLas(+) and CLas(-) populations of D. citri, and found that proteins involved in polyketide biosynthesis by the end ...[more]

PMID: 26580079

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Project description:Root samples of ‘Sanhu’ red tangerine trees infected with and without Candidatus Liberibacter asiaticus (CLas) were collected at 50 days post inoculation and subjected to RNA-sequencingto profile the differentially expressed genes (DEGs) . Results showed that a total of 3956 genes were differentially regulated by HLB-infection. Comparison between our results and those of the previously reported showed that known HLB-modulated biological pathways including cell-wall modification, protease-involved protein degradation, carbohydrate metabolism, hormone synthesis and signaling, transcription activities, and stress responses were similarly regulated by HLB infection but different or root-specific changes did exist. The root unique changes included the down-regulation in genes of ubiquitin-dependent protein degradation pathway, secondary metabolisms, cytochrome P450, UDP-glucosyl transferase and pentatricopeptide repeat containing protein genes. Notably, nutrient absorption was impaired by HLB-infection as the expression of the genes involved in Fe, Zn, N and P adsorption and transportation were significantly changed. HLB-infection induced some cellular defense responses but simultaneously reduced the biosynthesis of the three major classes of secondary metabolites, many of which are known to have anti-pathogen activities. Genes involved in callose deposition were up-regulated whereas those involved in callose degradation were also up-regulated, indicating that the sieve tube elements in roots were hanging on the balance of life and death at this stage. In addition, signs of carbohydrate starvation were already eminent in roots at this stage. Other interesting genes and pathways that were changed by HLB-infection were also discussed based on our findings. Two-year-old seedlings of ‘Sanhu’ red tangerine were grafted with buds from CLas-infected or CLas-free ‘Gonggan’ mandarin trees. Mature leaves and roots were collected from the CLas-inoculated and the control trees every ten days to detect for CLas by PCR. DNA used for PCR was extracted with the use of the Plant DNA isolation Kit (Trans, Beijing, China) according to manufacturer’s instructions. Primers used for CLas detection were the same A2/J5. The fibrous roots of 3 CLas-positive and 3 control CLas-free trees were individually collected at 50 dpi (days post inoculation) when the HLB-inoculated trees became CLas-positive in both leaves and roots yet showed no visible chlorosis and other HLB symptoms. This should have allowed us not to miss too many early responsive genes but at the same time ensured that the trees were infected as expected. Total RNA was extracted from each sample using RNeasy plant mini kit (Qiagen, Valencia, CA) and further purified using the RQ1 Rnase Free Dnase Kit (Promega, Madison, USA). RNA quality and quantity were assessed by gel-electrophoresis and by absorbance at OD260/OD280, respectively. Aliquot RNA samples were stored at -80 ?. For RNA-seq analysis, RNA samples from the three trees were mixed in equal amount and used for cDNA library construction following the Illumina mRNA-sequencing sample preparation protocol (Illumina, San Diego, CA). The 90-bp raw paired end reads were generated by Illumina HiSeqTM 2000. Please note that the features (in the processed data, e.g.,clementine0.9_035093m.g) represent the gene IDs in the Citrus clementina genome at ftp://ftp.jgi-psf.org/pub/JGI_data/phytozome/v9.0/Cclementina/. However, the database has been updated (http://genome.jgi.doe.gov/pages/dynamicOrganismDownload.jsf?organism=PhytozomeV10) with new gene IDs, thus the gene IDs used for this study are not trackable anymore. Therefore, the gene annotation files downloadeded from the previous genome database (v0.9) and used for the data analysis are included in this record.

Project description:The Asian citrus psyllid (Diaphorina citri) is a pest of citrus and the primary insect vector of the bacterial pathogen, ‘Candidatus Liberibacter asiaticus’ (CLas), which is associated with citrus greening disease. Variability in CLas titer in insects collected from infected plants has been attributed in part to the host plant from which the insects were collected. CLas accumulates to high titers in infected Citrus macrophylla, and in D. citri feeding on the infected plants of this species. In contrast, in the citrus relative Murraya paniculata, CLas titers remain low in infected plants and in D. citri exposed to infected plants. In this study, top-down and bottom-up proteomics methods were used to investigate the impact of these different host plants on D. citri protein expression. Difference in gel electrophoresis (DIGE) was used to identify protein spots on two-dimensional gels that were larger in one of three insect sample classes compared to the other two: D. citri continuously reared on C. macrophylla, D. citri reared continuously on M. paniculata, and D. citri transferred to M. paniculata for five days feeding after continuous rearing on C. macrophylla. Peptide mass spectrometry was used to identify and quantify proteins in target spots upregulated in each sample class. Shotgun proteomics was used to identify and quantify proteins from analysis of tryptic peptide samples prepared from whole insects from four sample classes: the reciprocal host switch condition (D. citri transferred to C. macrophylla for five days feeding after continuous rearing on M. paniculata) in addition to the three sample classes used in DIGE analysis. Integration of the results of both analyses reveals proteins identified by separate experimental workflows to be upregulated in insects adapted to each host plant, and in insects adapting to a novel host plant. A peptidoglycan-degrading protein involved in the immune response to bacterial pathogens was found to be upregulated in M. paniculata-reared D. citri. In the absence of CLas infection, host plant factors specific to M. paniculata may prime the antibacterial immune response in D. citri. Understanding the insect proteins involved in the adaptation of D. citri to host plants with variation in their susceptibility to CLas will inform the development of control strategies aimed at stopping the spread of citrus greening disease.

Dataset Information

Asian citrus psyllid proteome: whole insect samples, with or without Candidatus Liberibacter asiaticus

Publications

Metabolic Interplay between the Asian Citrus Psyllid and Its Profftella Symbiont: An Achilles' Heel of the Citrus Greening Insect Vector.

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