Diaphorina citri is a vector of Candidatus Liberibacter asiaticus (CLas), the gram-negative bacterial pathogen associated with citrus greening disease. Control measures rely on pesticides with negative impacts on the environment, natural ecosystems and human and animal health. In contrast, gene-targeting methods have the potential to specifically target the vector species and/or reduce pathogen transmission.
To improve the genomic resources needed for targeted pest control, we assembled a D. citri genome based on PacBio long reads followed by proximity ligation-based scaffolding. The 474 Mb genome has 13 chromosomal-length scaffolds. 1,036 genes were manually curated as part of a community annotation project, composed primarily of undergraduate students. We also computationally identified a total of 1,015 putative transcription factors (TFs) and were able to infer motifs for 337 TFs (33 %). In addition, we produced a genome-independent transcriptome and genomes for D. citri endosymbionts.
Manual annotation provided more accurate gene models for use by researchers and also provided an excellent training opportunity for students from multiple institutions. All resources are available on CitrusGreening.org and NCBI. The chromosomal-length D. citri genome assembly serves as a blueprint for the development of collaborative genomics projects for other medically and agriculturally significant insect vectors.

Citrus greening disease is caused by the pathogen Candidatus Liberibacter asiaticus and transmitted by the Asian citrus psyllid, Diaphorina citri. No curative treatment or significant prevention mechanism exists for this disease, which causes economic losses from reduced citrus production. A high-quality genome of D. citri is being manually annotated to provide accurate gene models to identify novel control targets and increase understanding of this pest. Here, we annotated 25 D. citri genes involved in glycolysis and gluconeogenesis, and seven in trehaloneogenesis. Comparative analysis showed that glycolysis genes in D. citri are highly conserved but copy numbers vary. Analysis of expression levels revealed upregulation of several enzymes in the glycolysis pathway in the thorax, consistent with the primary use of glucose by thoracic flight muscles. Manually annotating these core metabolic pathways provides accurate genomic foundation for developing gene-targeting therapeutics to control D. citri.

The circadian rhythm is a process involving multiple genes that generates an internal molecular clock, allowing organisms to anticipate environmental conditions produced by the earths rotation on its axis. This report presents the results of the manual curation of twenty-seven genes likely associated with circadian rhythm in the genome of Diaphorina citri, the Asian citrus psyllid. This insect acts as the vector of the bacterial pathogen Candidatus Liberibacter asiaticus (CLas), the causal agent of citrus greening disease (Huanglongbing). This disease is the most severe detriment to citrus industries and has drastically decreased crop yields worldwide. Based on the genes identified in the psyllid genome, namely cry1 and cry2, D. citri likely possesses a circadian model similar to that of the lepidopteran butterfly, Danaus plexippus. Manual annotation of these genes will allow future molecular therapeutics to be developed that can disrupt the psyllid biology.

The Asian citrus psyllid, Diaphorina citri, is an insect vector that transmits Candidatus Liberibacter asiaticus, the causal agent of the Huanglongbing (HLB) or citrus greening disease. This disease has devastated Floridas citrus industry and threatens Californias industry as well as other citrus producing regions around the world. To find novel solutions to the disease, a better understanding of the vector is needed. The D. citri genome has been used to identify and characterize genes involved in Wnt signaling pathways. Wnt signaling is utilized for many important biological processes in metazoans, such as patterning and tissue generation. Curation based on RNA sequencing data and sequence homology confirm twenty four Wnt signaling genes within the D. citri genome, including homologs for beta-catenin, Frizzled receptors, and seven Wnt-ligands. Through phylogenetic analysis, we classify D. citri Wnt-ligands as Wg/Wnt1, Wnt5, Wnt6, Wnt7, Wnt10, Wnt11, and WntA. The D. citri version 3.0 genome with chromosomal length scaffolds reveals a conserved Wnt1-Wnt6-Wnt10 gene cluster with gene configuration similar to that in Drosophila melanogaster. These findings provide a greater insight into the evolutionary history of D. citri and Wnt signaling in this important hemipteran vector. Manual annotation was essential for identifying high quality gene models. These gene models can be used to develop molecular systems, such as CRISPR and RNAi, that target and control psyllid populations, to manage the spread of HLB. Manual annotation of Wnt signaling pathways was done as part of a collaborative community annotation project.

Huanglongbing (HLB), also known as citrus greening disease, is caused by the bacterium Candidatus Liberibacter asiaticus (CLas) and represents a serious threat to global citrus production. This bacteria is transmitted by the Asian citrus psyllid, Diaphorina citri (Hemiptera) and there are no effective in planta treatments for CLas. Therefore, one strategy is to manage the psyllid population. Manual annotation of the D. citri genome can identify and characterize gene families that could serve as novel targets for psyllid control. The yellow gene family represents an excellent target as yellow genes are linked to development and immunity due to their roles in melanization. Combined analysis of the genome with RNA-seq datasets, sequence homology, and phylogenetic trees were used to identify and annotate nine yellow genes for the D. citri genome. Phylogenetic analysis shows a unique duplication of yellow-y in D. citri, with life stage specific expression for these two genes. Genomic analysis also indicated the loss of a gene vital to the process of melanization, yellow-f, and the gain of a gene which seems to be unique to hemipterans, yellow 9. We suggest that yellow 9 or the gene yellow 8 (c), which consistently groups closely to yellow-f, may take on this role. Manual curation of genes in D. citri has provided an in-depth analysis of the yellow family among hemipteran insects and provides new targets for molecular control of this psyllid pest. Manual annotation of the yellow gene family was done as part of a collaborative community annotation project.