Project description:Candidatus Liberibacter asiaticus (Las), a non-culturable phloem-limited bacterium, is the suspected causal agent of Huanglongbing (HLB) in Florida. HLB is one of the most devastating diseases of citrus and no resistant cultivars have been identified to date, though tolerance has been suggested in the genus Poncirus. A recent study conducted in our laboratory demonstrated tolerance of US-897, a hybrid of Poncirus trifoliata (L. Raf.) and the susceptible ‘Cleopatra’ mandarin (Citrus reticulata Blanco), to Ca. L. asiaticus, the presumed causal agent of HLB in Florida (Albrecht & Bowman, HortScience 46 (2011) 16-22). This study compares transcriptional changes in tolerant US-897 and susceptible ‘Cleopatra’ mandarin seedlings in response to infection with Las using the Affymetrix GeneChip citrus array with the main objective of identifying genes associated with tolerance to HLB. Such genes may be suitable as potential targets for biotechnology approaches, providing one strategy to possibly control this destructive disease of citrus.
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.
Project description:Candidatus Liberibacter asiaticus (Las), a non-culturable phloem-limited bacterium, is the suspected causal agent of Huanglongbing (HLB) in Florida. HLB is one of the most devastating diseases of citrus and no resistant cultivars have been identified to date, though tolerance has been suggested in the genus Poncirus. A recent study conducted in our laboratory demonstrated tolerance of US-897, a hybrid of Poncirus trifoliata (L. Raf.) and the susceptible ‘Cleopatra’ mandarin (Citrus reticulata Blanco), to Ca. L. asiaticus, the presumed causal agent of HLB in Florida (Albrecht & Bowman, HortScience 46 (2011) 16-22). This study compares transcriptional changes in tolerant US-897 and susceptible ‘Cleopatra’ mandarin seedlings in response to infection with Las using the Affymetrix GeneChip citrus array with the main objective of identifying genes associated with tolerance to HLB. Such genes may be suitable as potential targets for biotechnology approaches, providing one strategy to possibly control this destructive disease of citrus. Fifteen months-old seedlings of the genotypes 'Cleopatra' mandarin and US-897 were graft-inoculated with non-infected (control) or Las-infected tissue from greenhouse-grown 'Valencia' (C. sinensis L.) plants. Six non-infected control plants each from 'Cleopatra' (MC) and US-897 (TC) and six infected plants each from Cleopatra (MI) and US-897 (TI) were used. Four to six leaves per plant were excised at 32 weeks after inoculation (wai) and immediately frozen in liquid nitrogen. Total RNA was extracted from all 24 plants. Equal amounts of RNA from two samples were pooled to obtain three biological replicates per genotype and treatment and used for hybridization on Affymetrix citrus microarrays.
Project description:N-glycosylation is one of the most abundant and conserved protein modifications in eukaryotes. This modification serves various important functions, such as protein folding and cellular attachment, but also modulation of a protein’s function. Recently, it has been shown that N-glycosylation of proteins plays a vital role in insect development and survival, which makes it an interesting target for pest control. Despite the importance of protein N-glycosylation in insects, not much is known about insect N-glycoproteomes. Here, we report on the N-glycoproteomes of three major pest insects spanning different insect orders; Drosophila melanogaster (Diptera), Tribolium castaneum (Coleoptera) and Acyrthosiphon pisum (Hemiptera). The number of identified N-glycosylation sites ranged from 889 in T. castaneum, to 941 in D. melanogaster and 1,338 in A. pisum. Comparison between the different insect species revealed both conserved and species-specific glycoproteins. The functionality of the insect glycoproteins together with the conservation of the N-glycosites throughout evolution are discussed. This information can help in the elaboration of novel pest insect control strategies based on interference in insect glycosylation.
Project description: Bacillus thuringiensis Cry toxin is used for insect pest management. When susceptible insect larvae ingest the Bt toxin it shows various physiological and cellular changes. We have employed Agilent One Color matrix based microarray to check the mid-gut gene expression upon Cry toxin exposure in Achaea janata (Castor semilooper). Microarray analysis shows significant variation in gene expression upon Cry toxin exposure.
Project description:The citrus red mite, Panonychus citri, is a major pest on citrus all around the world. Mitochondrial Electron Transport Inhibitors of complex I (METI-I) acaricides such as fenpyroximate have been used extensively to control P. citri populations, which resulted in multiple reports of METI-I resistant populations in the field. In this study, gene expression profile differences between METI-I resistant P. citri populations (Lahijan, Ramsar and Sari) and a susceptible P. citri population (Rasht) from Iran were compared using Illumina RNAseq.
Project description:A genomic insight into how an insect pest responds to the infection of a fungal insect pathogen, such as Beauveria bassiana, is critical for alternative strategy of insect pest contol based on fungal insecticides but has not been well probed. Here we constructed three pairs of digital expression libraries (transcriptomes) of Plutella xylostella (global lepidopteran pest) larvae 24, 36 and 48 hours post treatment of infection (hptI) and control (hptC) to reveal the host response to B. bassiana infection at genomic level. The paired libraries comprised 2144, 3200 and 2967 differentially expressed genes (DEGs) of P. xylostella at 24, 36 and 48 hptI/hptC, respectively. These DEGs were enriched in various immune pathways activated by the fungal infection, such as the pathways of complement and coagulation cascades, protein digestion and absorption, and drug metabolism - cytochrome P450. We found that 24 hptI was critical either for the cuticular penetration of B. bassiana or for the initial activation of the host defense system. The host immune response peaked at 36 hptI so that multiple defense mechanisms were activated against the fungal entry into the host hemocoel. At 48 hptI, many host genes involved in immunity and metabolism were downregulated, suggesting a success of fungal localization in the host hemocoel by overcoming the host defense reaction. Finally, we revealed that several fungal pathways could play important roles in the host-pathogen interaction, such as antioxidant activity, peroxidase activity and proteolysis. Up to 1636 fungal genes were co-expressed at the three time points, and 116 of them encode putative secretion proteins. Our results provide a novel insight into the pathogen-insect interaction and help to probe molecular mechanisms involved in the control of P. xylostella by B. bassiana. Here we constructed three pairs of digital expression libraries (transcriptomes) of Plutella xylostella (global lepidopteran pest) larvae 24, 36 and 48 hours post treatment of infection (hptI) and control (hptC) to reveal the host response to B. bassiana infection at genomic level
Project description:The resistance mechanisms evolved by insects to overcome host-plant allelochemicals is a key consideration in pest management. Camphor oil (EO) and its main component (i.e D-camphor) form a specific terpenoid-defensive system in camphor trees, Cinnamomum camphora. However, an emerging insect pest, Pagiophloeus tsushimanus has recently caused serious damage to this intractable plant species, which is largely elusive. Here, we used feeding bioassays and RNA-seq to investigate the mechanism underlying the resistance of the beetle to host-specific terpenoid defenses. Firstly, a hormetic response in both larval weight and developmental time was observed in terpenoid-feeding individuals, which is a highly generalized dose-response phenomenon in toxicology whereas occurs infrequently in the context of insect-plant interactions. Then, comparative transcriptome analysis between terpenoid-feeding and control groups indicated that both CYP450s-mediated metabolic resistance and CPs-mediated cuticular resistance were jointly employed to cope with terpenoid-induced stress. In addition, a small portion of genes involved in glucose transport pathway were up-regulated at the low D-camphor dose, suggesting an extra intake of glucose used for larval growth may contribute to a hormetic response. These findings probably implied that the dual terpenoid-resistance mechanisms existing in this specialist is an essential precondition for hormetic response in larval growth, ultimately contributing to successfully colonize host camphor trees with impunity. Overall, our study will open new avenues for understanding the insect-plant coevolutionary adaptation and developing durable pest control strategies.