Biology subjects

Hunter, W. B.

Publications and source records attributed to Hunter, W. B..

17 recordsLinked to original sources

bioRxiv2021

Manual curation and phylogenetic analysis of chitinase family genes in the Asian citrus psyllid, Diaphorina citri

Chitinases are enzymes that digest the polysaccharide polymer chitin. Chitin is a major component of insect exoskeletons, which must be replaced multiple times during growth and development in a process known as molting. Insect genomes usually have multiple chitinase genes, some of which are required for molting and some whose functions are still unknown. Since knockdown of the chitinases required for molting causes high levels of lethality, chitinase genes have drawn interest as targets for RNAi-based pest control methods. The Asian citrus psyllid, Diaphorina citri, carries the bacterium that causes Huanglongbing, also known as citrus greening disease, which is devastating the citrus industry worldwide. We have identified and annotated 12 chitinase family genes from D. citri as part of a community effort to create high quality gene models to facilitate the design of interdictory molecules for pest control. Using predicted protein domain content and phylogenetic analysis, we categorized the D. citri chitinases according to a previously established classification scheme and re-evaluated the classification of chitinases in other hemipterans. In addition to chitinases belonging to known groups, we identified a novel class of chitinases present in D. citri and several related hemipterans that appears to be the result of horizontal gene transfer.

bioinformatics↗

bioRxiv2021

Genomic identification, annotation, and comparative analysis of Vacuolar-type ATP synthase subunits in Diaphorina citri

Detailed annotation and comparative analysis were performed on the Asian citrus psyllid (ACP), Diaphorina citri, vacuolar-type ATP synthase (V-ATPase) to support the biological understanding and development of novel therapeutics to manage psyllid vectors. D. citri is a hemipteran insect that vectors the causative agent, the bacteria Candidatus Liberibacter asiaticus (CLas), of the citrus greening disease, Huanglongbing (HLB). Millions of citrus trees have been destroyed by citrus greening and every grove in Florida has been directly impacted. In eukaryotic organisms, V-ATPase is an abundant heterodimeric enzyme that serves the cell with essential compartment acidification through the active processes that transport protons across the membrane. Manual curation was completed on 15 putative V-ATPase genes in the D. citri genome. Comparative genomic analysis reveals that the D. citri V-ATPase subunits share domains and motifs with other insects, including the V-ATPase-A superfamily domain from the V-ATPase catalytic subunit A, which shares a 92% identity with Acyrthosiphon pisum. Phylogenetic analysis separates D. citri V-ATPase subunits into expected clades with orthologous sequences. Based on the results of annotation and comparative genomic analysis, RNAi therapies targeting D. citri V-ATPase genes, which have been successfully utilized in related hemipterans, are being pursued. Annotation of the D. citri genome is a critical step towards the development of directed-pest management that will lead to the reduced spread of the pathogens causing HLB throughout the citrus industry.

bioinformatics↗

bioRxiv2021

Ubiquitin Proteasome Pathway annotation in Diaphorina citri can reveal potential targets for RNAi based pest management

Ubiquitination is an ATP-dependent process that targets proteins for degradation by the proteasome. In this study, we annotated 15 genes from the ubiquitin-proteasome pathway in the Asian citrus psyllid, Diaphorina citri. This psyllid vector has come to prominence in the last decade due to its role in the transmission of the devastating bacterial pathogen, Candidatus Liberibacter asiaticus (CLas). Infection of citrus crops by this pathogen causes Huanglongbing (HLB or citrus greening disease) and results in the eventual death of citrus trees. The identification and correct annotation of these genes in D. citri will be useful for functional genomic studies that aid in the development of RNAi-based management strategies aimed at reducing the spread of HLB. Investigating the effects of CLas infection on the expression of ubiquitin-proteasome pathway genes may provide new information regarding the role that these genes play in the acquisition and transmission of CLas by D. citri.

bioinformatics↗

bioRxiv2021

Annotation of glycolysis, gluconeogenesis, and trehaloneogenesis pathways provide insight into carbohydrate metabolism in the Asian citrus psyllid

Citrus greening disease is caused by the pathogen Candidatus Liberibacter asiaticus, which is transmitted by the Asian citrus psyllid, Diaphorina citri. There is no curative treatment or significant prevention mechanism for this detrimental disease that causes continued economic losses from reduced citrus production. A high quality genome of D. citri is being manually annotated to provide accurate gene models required to identify novel control targets and increase understanding of this pest. Here, we annotated genes involved in glycolysis, gluconeogenesis, and trehaloneogenesis in the D. citri genome, as these are core metabolic pathways and suppression could reduce this pest. Specifically, twenty-five genes were identified and annotated in the glycolysis and gluconeogenesis pathways and seven genes for the trehaloneogenesis pathway. Comparative analysis showed that the glycolysis genes in D. citri are highly conserved compared to orthologs in other insect systems, but copy numbers vary in D. citri. Expression levels of the annotated gene models were analyzed and several enzymes in the glycolysis pathway showed high expression in the thorax. This is consistent with the primary use of glucose by flight muscles located in the thorax. A few of the genes annotated in D. citri have been targeted for gene knockdown as a proof of concept, for RNAi therapeutics. Thus, manual annotation of these core metabolic pathways provides accurate genomic foundations for developing gene-targeting therapeutics to reduce D. citri.

bioinformatics↗

bioRxiv2021

Annotation of Putative Circadian Rhythm-Associated Genes in Diaphorina citri (Hemiptera : Liviidae)

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.

bioinformatics↗

bioRxiv2021

Annotation of Hox cluster and Hox cofactor genes in the Asian citrus psyllid, Diaphorina citri, reveals novel features

Hox genes and their cofactors are essential developmental genes that specify regional identity in animals, including insects. A particularly interesting feature of Hox genes is their conserved arrangement in clusters in the same order in which they specify identity along the anterior-posterior axis. Among insects, breaks in the cluster have been reported in a few species, but these seem to be the exception rather than the rule. We have annotated the ten Hox genes of the Asian citrus psyllid, Diaphorina citri, and determined that there is a split in its Hox cluster between the Deformed and Sex combs reduced genes. This is the first time a break at this position has been observed in an insect Hox cluster. We have also annotated the D. citri orthologs of the Hox cofactor genes homothorax, PKNOX and extradenticle. Interestingly, we found an additional copy of extradenticle in D. citri that appears to be a retrogene. Expression data and sequence conservation suggest that the extradenticle retrogene may have retained the original extradenticle function and allowed the parental extradenticle gene to diverge.

bioinformatics↗

bioRxiv2021

Quantification of new and archived Diaphorina citri transcriptome data using a chromosomal length D. citri genome assembly reveals the vector's tissue-specific transcriptional response to citrus greening disease

Background Huanglongbing (HLB) is the most serious disease of citrus. HLB is caused by the obligate, intracellular bacterium "Candidatus Liberibacter asiaticus" (CLas). CLas is transmitted by Diaphorina citri, the Asian citrus psyllid. Development of transmission blocking strategies to manage HLB relies on knowledge of CLas-D. citri interactions at the molecular level. Prior transcriptome analyses of CLas-infected and un-infected D. citri point to changes in psyllid biology due to CLas-infection. These studies relied on incomplete versions of the D. citri genome, lacked proper host plant controls, and/or were analyzed using different statistical approaches. Therefore, we used standardized experimental and computational approaches to identify differentially expressed genes in both CLas (+) and CLas (-) D. citri. The comparative analysis utilized the newest chromosomal length D. citri genome assembly Diaci_v3. In this work, we present a quantitative transcriptome analysis of excised heads, salivary glands, midguts and bacteriomes from CLas (+) and CLas (-) insects. Results Each organ had unique transcriptome profiles and responses to CLas infection. Though most psyllids were infected with CLas, CLas-derived transcripts were not detected in all organs. By analyzing the midgut dataset using both the Diaci_v1.1 and v3.0 D. citri genomes, we showed that improved genome assembly led to significant and quantifiable differences in RNAseq data interpretation. Conclusions Our results support the hypothesis that future transcriptome studies on circulative, vector-borne pathogens should be conducted at the tissue specific level using complete, chromosomal-length genome assemblies for the most accurate understanding of pathogen-induced changes in vector gene expression.

genomics↗

bioRxiv2020

Segmentation pathway genes in the Asian citrus psyllid, Diaphorina citri

Insects have a segmented body plan that is established during embryogenesis when the anterior-posterior (A-P) axis is divided into repeated units by a cascade of gene expression. The cascade is initiated by protein gradients created by translation of maternally provided mRNAs, localized at the anterior and posterior poles of the embryo. Particular combinations of these proteins activate specific gap genes to divide the embryo into distinct regions along the A-P axis. Gap genes then activate pair-rule genes, which are usually expressed in part of every other segment. The pair-rule genes, in turn, activate expression of segment polarity genes in a portion of each segment. The segmentation genes are generally conserved among insects, although there is considerable variation in how they are deployed. We annotated 24 segmentation gene homologs in the Asian citrus psyllid, Diaphorina citri. We identified most of the genes that were expected to be present based on known phylogenetic distribution. Two exceptions were eagle and invected, which are present in at least some hemipterans, but were not identified in D. citri. Many of these genes are likely to be essential for D. citri development and thus may be useful targets for pest control methods.

genomics↗

bioRxiv2020

Annotation and analysis of yellow genes in Asian citrus psyllid, Diaphorina citri, vector for the Huanglongbing disease

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 was done as part of a collaborative community annotation project (https://citrusgreening.org/annotation/index).

genomics↗

bioRxiv2020

Characterization of chitin deacetylase genes in the Diaphorina citri genome

Chitin deacetylases (CDAs) are one of the least understood components of insect chitin metabolism. The partial deacetylation of chitin polymers appears to be important for the proper formation of higher order chitin structures, such as long fibers and bundles, that contribute to the integrity of the insect exoskeleton and other structures. Some CDAs may also play a role in bacterial defense. Here we report the characterization of four CDA genes from the Asian citrus psyllid, Diaphorina citri, laying the groundwork for future study of these genes. D. citri is the vector for Candidatus Liberibacter asiaticus (CLas), which is responsible for the global outbreak of Huanglongbing (citrus greening) disease. The manual annotation was done as part of a collaborative community annotation project (https://citrusgreening.org/annotation/index).

genomics↗

bioRxiv2020

Chitin Biosynthesis Genes in Diaphorina citri

The polysaccharide chitin is critical for the formation of many insect structures, including the exoskeleton, and is required for normal development. Here we report the annotation of three genes from the chitin synthesis pathway in the Asian citrus psyllid, Diaphorina citri (Hemiptera: Liviidae), the vector of Huanglongbing (citrus greening disease). Most insects have two chitin synthase (CHS) genes but, like other hemipterans, D. citri has only one. In contrast, D. citri is unusual among insects in having two UDP-N-acetylglucosamine pyrophosphorylase (UAP) genes. One of the D. citri UAP genes is broadly expressed, while the other is expressed predominantly in males. Our work helps pave the way for potential utilization of these genes as pest control targets to reduce the spread of Huanglongbing.

genomics↗

bioRxiv2020

Characterization of Wnt Signaling Genes in Diaphorina citri

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 further be used to develop molecular systems, such as CRISPR and RNAi, that target and control D. citri populations, to manage the spread of HLB. Manual annotation of Wnt signaling pathways was done as part of a collaborative community annotation project (https://citrusgreening.org/annotation/index).

genomics↗

bioRxiv2018

BAPC-assisted CRISPR/Cas9 System: Targeted Delivery into Adult Ovaries for Heritable Germline Gene Editing (Arthropoda: Hemiptera)

Innovative gene targeting strategies are often limited in application across arthropod species due to problems with successful delivery. In hemipterans, embryonic injections often used to deliver CRISPR components fail due to nearly complete embryo mortality. The Asian citrus psyllid, Diaphorina citri, Kuwayama, (Hemiptera: Liviidae), is the vector for a pathogenic bacterium, Candidatus Liberibacter asiaticus, CLas, which is devastating the U.S. citrus industries. The disease called, Huanglongbing, HLB, (aka. Citrus greening disease), is transmitted during psyllid feeding. Infection causes severe tree decline, loss of fruits, and eventually tree death. The citrus tree pathogen, CLas, is a fastidious alpha-proteobacterium, which has spread into all citrus growing regions worldwide. The economic losses are estimated in the billions of dollars, in U.S.A., Brazil, and China. Innovative technologies aimed at reducing psyllid populations using targeting RNA suppression, like RNAi, or gene-editing tools, like CRISPR/Cas9 have potential to reduce psyllid vectors and the pathogen in a highly specific manner. Breakthroughs that improve gene editing in psyllids, such as the BAPC-assisted-CRISPR/Cas9 System, enabled delivery by injection of CRISPR/Cas9 components directly into nymphs and adult females. Injection near ovaries produced heritable germline gene editing in subsequent generations. This method opens the world of gene editing across arthropods and bypasses the need for microinjection of eggs. Effective development of therapeutic treatments to reduce insect vectors, and stop pathogen transmission would provide sustainable citrus and grapevine industries.

genetics↗

bioRxiv2018

Leafhopper Viral Pathogens: Ultrastructure of salivary gland infection of Taastrup-like virus in Psammotettix alienus Dahlbom; and a novel Rhabdovirus in Homalodisca vitripennis (Germar) Hemiptera: Cicadellidae]

Viruses that are pathogenic to insect pests can be exploited as biological control agents. Viruses that are pathogenic to beneficial insects and other arthropods, as in honey bees, silk worms, and shrimp, cause millions of dollars of losses to those industries. Current advances in next generation sequencing technologies along with molecular and cellular biology have produced a wealth of information about insect viruses and their potential applications. Leafhoppers cause economic losses as vectors of plant pathogens which significantly reduce the worlds food crops. Each year more viruses are discovered primarily through the use of next generation sequencing of the leafhopper hosts. The diversity of viruses from leafhoppers demonstrates a wide range of taxonomic members that includes genomes of DNA or RNA from families like: Reoviridae, Iridoviridae, Dicistroviridae, Iflaviridae, and others yet to be classified. Discussed is a recent viral pathogen isolated from the leafhopper Psammotettix alienus, name Taastrup Virus. Taastrup virus (TV) is a novel virus with a RNA genome, a Filovirus-like morphology, being tentatively placed within the Mononegavirales. Adult Psammotettix alienus infected with TV, showed the highest concentration of virions in salivary glands, consisting of a principal gland (type I-VI-cells) and an accessory gland. Examination of thin sections revealed enveloped particles, about 1300 nm long and 62 nm in diameter, located singly or in paracrystalline arrays in canaliculi of type III- and IV-cells. In gland cells with TV particles in canaliculi, granular masses up to 15 m in diameter were present in the cytoplasm. These masses are believed to be viroplasms, the sites for viral replication. TV particles were observed at the connection between a canaliculus and the salivary duct system. A TV-like virus with strongly similar morphology was discovered in the ornamental plant, Liriope, near Fort Pierce, Florida, USA. When the virus was inoculated to a leafhopper cell culture, HvWH, made from the glassy-winged sharpshooter, Homalodisca vitripennis (Germar), the cells rapidly degraded with 100% mortality in 48 hours. These two instances are the only reported cases of this newly discovered viral pathogen of leafhoppers.

pathology↗

bioRxiv2018

Rapid, Affordable, Collection and Analysis of Bioaerosol Viral Pathogens (Begomoviruses and Whitefly Vectors)

Detection of pathogens is critical to monitoring their distribution and spread, and is a key component in the prediction and management of disease epidemiology. Monitoring for pathogens as bioaerosols requires developing techniques which are sensitive, affordable, and time saving before they will have widespread impact. This approach also overcomes private property issues, which are a major pitfall in monitoring diseases in complex agricultural and urban settings. In this study, we have applied an emerging technology of electrostatic sampling to the detection of an insect-transmitted plant pathogen as a bioaerosol. Where insects aggregate in large numbers, as with whiteflies, leafhoppers, psyllids and honey bees, the pathogen (ie. virus or bacteria) becoming aerosolized as thousands of excreta droplets fall from the plants during feeding. Agricultural systems have not fully measured the impact of bioaerosols on disease epidemiology. Electrostatic sampling provides a valuable, affordable, method for monitoring for diseases as bioaerosols, which includes plant, animal and human pathogens. This study shows results which successfully used an electrostatic sampling device to collect an aerosolized begomovirus from the air near whiteflies feeding on virus-infected tomato plants.

pathology↗

bioRxiv2018

Evaluation of North American Asian citrus psyllid (Hemiptera: Liviidae) population genetics by cytochrome P450 melt curve analysis

The Asian Citrus Psyllid (ACP) (Diaphorina citri) is a Hemipteran which feeds on Citrus and is the principal vector of the pathogen Candidatus Liberibacter asiaticus (Las); the primary causal agent of Huanglongbing (HLB) or citrus greening disease. Currently HLB is the single greatest threat to the American citrus industry. Previous work has shown that that Cytochrome P450 (CYP4) genes are associated with insecticidal resistance to neonicotinoid insecticides. Infection by Las has also been shown to affect the expression of CYP4 genes. In this study the genetic relationships of ACP populations was evaluated using melting curve analysis of CYP4 genes. Principal component analysis shows the presence of two potential haplotypes. The groups identified by principal component analysis are significantly different from each other (P < 0.05) for all CYP4 genes tested. All populations are most likely within 2-3 mutations of each other. The existence of these mutations sheds light on the potential evolution of ACP in North America. Indicating either coevolution with Las or difference in imidacloprid basal selective pressure in the United States and Mexico.

evolutionary biology↗

bioRxiv2018

The Genomic Basis of Arthropod Diversity

BackgroundArthropods comprise the largest and most diverse phylum on Earth and play vital roles in nearly every ecosystem. Their diversity stems in part from variations on a conserved body plan, resulting from and recorded in adaptive changes in the genome. Dissection of the genomic record of sequence change enables broad questions regarding genome evolution to be addressed, even across hyper-diverse taxa within arthropods.\n\nResultsUsing 76 whole genome sequences representing 21 orders spanning more than 500 million years of arthropod evolution, we document changes in gene and protein domain content and provide temporal and phylogenetic context for interpreting these innovations. We identify many novel gene families that arose early in the evolution of arthropods and during the diversification of insects into modern orders. We reveal unexpected variation in patterns of DNA methylation across arthropods and examples of gene family and protein domain evolution coincident with the appearance of notable phenotypic and physiological adaptations such as flight, metamorphosis, sociality and chemoperception.\n\nConclusionsThese analyses demonstrate how large-scale comparative genomics can provide broad new insights into the genotype to phenotype map and generate testable hypotheses about the evolution of animal diversity.

genomics↗