ABSTRACT: The green rice leafhopper (GRH), Nephotettix cincticeps Uhler, is a major insect pest of cultivated rice in temperate Asia. GRH2-near-isogenic line (NIL) TGRH11, GRH4-NIL (TGRH16) and GRH2/GRH4-pyramided line (PYL) TGRH29 were developed by introducing the GRH2 and GRH4 from indica rice (DV85). We identified GRH-inducible genes in respective rice lines. Furthermore, we compared the gene expression levels between NILs/PYL and control plants (T65). The gene expression changes in respective rice lines were detected by comparison between GRH-infested and pre-infested plants. Seedlings at the second-leaf stage were infested with 10 to 15 first- or second-instar nymphs in test tubes and shoots were collected at 30 h after GRH infestation. Detached-leaf blades at heading stage were infested with 10 to 15 first- or second-instar nymphs in test tubes and samples were collected at 30 h after GRH infestation. For each treatment, two biological replicates were performed.
Project description:KMD is genetically engenered to be highly resistant to lepidopteran pests through expressing a synthetic cry1Ab gene and its parent non-transgenic rice is Xiushui 11.The developmental duration of BPH feeding on KMD2 was significantly delayed. And moreover, the fecundity of BPH was significantly lower when fed on Bt rice than on the non-Bt parental plants.To investigate unintended effects in KMD2 that causes changes in BPH performance, we performed microarray (GeneChip) analysis to compare the gene expression profiles between Bt rice and non-transgenic parental plants in response to BPH infestation. We used microarrays to detect Bt-independent variation, which might render Bt rice more defensive or less nutritious to BPH. For BPH treatment, 10 second-instar nymphs were infested onto each 30-day-old seedling. After 72 h, the BPH nymphs were carefully removed and rice shoots of both BPH-infested and non-infested plants were sampled for microarray analysis. There were four treatments: Xiushui 11-non infested, Xiushui 11-BPH infested, KMD2-non infested, KMD2-BPH infested, three biological replications.
Project description:In the present study molecular interactions between potato plants, Colorado potato beetle (CPB) larvae and Potato virus YNTN (PVYNTN) were investigated by analyzing gene expression in potato leaves. mRNA samples of secondary PVYNTN-infected (CPB_PVY) and healthy potato plants (CPB_H) cultivar Igor and of RNAi coi1-silenced (CPB_coi1) and non-transformed (CPB_NT) potato plants cultivar Desiree collected 24 h post CPB infestation and respective control non-infested samples (CONT_PVY, CONT_H, CONT_coi1, CONT_NT).
Project description:Introduction: Ixodes scapularis ticks are hematophagous arthropods capable of transmitting many infectious agents to humans. The process of blood feeding is an extended and continuous interplay between tick and host responses. While this process has been studied extensively in vitro, no global understanding of the host response to ticks has emerged. To address this issue, we measured skin-specific expression of 233 discrete genes at 8 time points during primary and secondary infestations of mice with pathogen-free I. scapularis nymphs. Selected results were then validated at the mRNA and protein levels. Results: Primary infestation was characterized by the late induction of an innate immune response. Lectin pattern recognition receptors, cytokines, and chemokines were upregulated consistent with increased neutrophil and macrophage migration. Gene ontology and pathway analyses of downregulated genes suggested inhibition of gene transcription and Th17 immunity. During the secondary infestation, additional genes were modulated suggesting a broader involvement of immune cells including CD8 and CD4 positive T lymphocytes. The cytokine response showed a mixed Th1/Th2 profile with a potential for T regulatory cell activity. Key gene ontology clusters observed during the secondary infestation were cell migration and activation. Matrix metalloproteinases were upregulated, apoptosis-related genes were differentially modulated, and immunoreceptor signaling molecules were upregulated. In contrast, transcripts related to mitogenic, WNT, Hedgehog, and stress pathways were downregulated. Conclusions: Our results support a model of tick feeding where lectin pattern recognition receptors orchestrate an innate inflammatory response during primary infestation that primes a mixed Th1/Th2 response upon secondary exposure. Tick feeding inhibits gene transcription and Th17 immunity. Salivary molecules may also inhibit upregulation of mitogenic, WNT, Hedgehog, and stress pathways and enhance the activity of T regulatory cells, production of IL-10, and suppressors of cytokine signaling molecules (SOCS). This study provides the first comprehensive transcriptional analysis of the host response at the tick bite site and suggests both a potential model of the host cutaneous response and candidate genes for further description and investigation. Ear biopsies from BALB/cJ mice infested with Ixodes scapularis nymphs were assayed at 12, 48, 72, and 96 hours after infestation during a primary and secondary exposure. 3 mice were measured at each time point. Controls were 3 similarly housed but tick-free mice.
Project description:Ixodes species ticks are competent vectors of tick-borne viruses including tick-borne encephalitis and Powassan encephalitis. Tick saliva has been shown to facilitate and enhance viral infection. This likely occurs by saliva-mediated modulation of host responses into patterns favorable for viral infection and dissemination. Because of the rapid kinetics of tick-borne viral transmission, this modulation must occur as early as tick attachment and initiation of feeding. In this study, the gene expression profile of cutaneous bite-site lesions created by uninfected ticks were analyzed at 1, 3, 6, and 12 hours after Ixodes scapularis nymphal tick attachment to discover host pathways or responses potentially important in tick-borne viral establishment. Four milimeter ear biopsies from BALB/cJ mice infested with Ixodes scapularis nymphs were assayed using Affymetrix genechip 430A 2.0 arrays at 1, 3, 6, and 12 hours after infestation during a primary exposure. 3 mice were measured at each time point. Controls were 3 similarly housed but tick-free mice.
Project description:Spider mites, including the two-spotted spider mite (Tetranychus urticae, TSSM) and the Banks grass mite (Oligonychus pratensis, BGM), are becoming increasingly important agricultural pests. The TSSM is an extreme generalist documented to feed on more than 1100 plant hosts. In contrast, the BGM is a grass specialist, with hosts including important cereal crops like maize, wheat, sorghum and barley. Historically, studies of plant-herbivore interactions have focused largely on insects. However, far less is known about plant responses to spider mite herbivores, especially in grasses, and whether responses differ between generalists and specialists. To identify plant defense pathways responding to spider mites, we collected time course RNA-seq data from barley (Hordeum vulgare L.) infested with TSSMs and BGMs. Additionally, and as a comparison to the physical damage caused by spider mite feeding, a wounding treatment was also included. The experiment was performed with four biological replicates across each of the following (28 samples in total): no infestation (C, control), 2hr after wounding (W2), 24hr after wounding (W24), 2hr after TSSM infestation (T2), 24hr after TSSM infestation (T24), 2hr after BGM infestation (B2), and 24hr after BGM infestation (B24).
Project description:The purpose of this study was to identify genes involved in the skin immune response that changed expression in response to scabies mite infestation. Transcriptomic analysis was undertaken using total RNA extracted from skin biopsies. Four biological replicates for non-infested control group and eight (four for crusted scabies disease phenotype and four for ordinary scabies diseases phenotype) biological replicates for the mite-infested group were used for gene expression analysis at baseline week 0, and at weeks 1, 2, 4 and 8 post-infestation using A-GEOD-16571 Agilent Porcine Gene Expression Microarray 4 x 44K.
Project description:Phloem-feeding pests cause extensive crop damage throughout the world yet little is understood about how plants perceive and defend themselves from these threats. The silverleaf whitefly (SLWF; Bemisia tabaci type B) is a good model for studying phloem-feeding insect-plant interactions as SLWF nymphs cause little wounding and have a long, continuous interaction with the plant. Using the Arabidopsis ATH1 GeneChip, the global responses to Silverleaf Whitefly 2nd instar feeding were examined. Experiment Overall Design: The silverleaf whitefly colony (Bemisia tabaci type B; Bemisia argentifolii Bellows and Perring) was maintained on Brassica napus cv. “Florida broad leaf” grown under fluorescent and incandescent lights (180 μE m-2 s-1) 27°C with 55% relative humidity under long-day (16 hr light: 8 hr dark) conditions in the Insectory and Quarantine Facility at the University of California, Riverside. Adult whiteflies are collected from infested plants by aspiration into 15-ml falcon tubes. Experiment Overall Design: Individual Arabidopsis thaliana ecotype Columbia plants were grown for 21 days in 4-inch diameter, round pots under fluorescent and incandescent lights (180 μE m-2 s-1) with 50% relative humidity, 23° C and an 8 hr-light/16-hr dark cycle. One hundred adult whiteflies were collected into each 15-ml falcon tube and the tube was placed upright in each pot. Plants were individually encased with 5- by 10-inch nylon bags that were secured to each pot with a rubber band. The whiteflies were released by unscrewing the falcon tube. After seven days, the adult whiteflies were removed from the plants by aspiration. The infested and non-infested plants were caged for the remainder of the experiment to ensure any adults that escaped aspiration could not reach the plants. Rosette tissue was collected after 28 days, when 2nd and 3rd instars were observed on wild-type Columbia plants. Developmentally matched leaves were harvested from uninfested plants. Infestations were performed in two growth chambers; each chamber contained one replicate experiment, which included 10 control and 10 infested plants. This experiment was repeated for a total of 8 biological replicate experiments. Experiment Overall Design: Total RNA from the eight biological replicates was isolated using the RNAwiz protocol (Ambion Inc., Austin, TX) and purified using a RNAeasy column (Qiagen, Valencia, CA). RNA from the two biological replicates performed in each growth chamber were pooled to eliminate variance due to different environmental factors. This yielded the infested and control RNA pools used in the microarrays (Exp1 and Exp2) and RT-PCRs (Exp3 and Exp4). The quality of the RNA was determined by A260/A280 absorbance readings. RNA integrity (1 µg) was verified by fractionation on a 1% formaldehyde gel. Experiment Overall Design: Hybridization Experiment Overall Design: Biotin-labeled cRNAs were synthesized from infested and control RNAs for Exp1 and Exp2 at the UC Irvine Microarray Facility using the Affymetrix Eukaryotic One-Cycle Target Labeling Assay protocol (Affymetrix GeneChip Expression, Analysis Technical Manual, Affymetrix, Santa Clara, CA). The labeled cRNA was hybridized to Affymetrix Arabidopsis genome ATH1 Chip arrays, washed, and scanned using a Hewlett Packard Genearray scanner (Hewlett-Packard, Palo Alto, CA). Experiment Overall Design: MAS 5.0 was performed using the standard parameters (Affymetrix GeneChip Expression, Analysis Technical Manual, Affymetrix, Santa Clara, CA.) Genes with “absent” calls in replicate experiments were removed from further analysis.
Project description:Infestation with white-backed planthopper (WBPH) to rice caused induced resistance to rice pathogens but brown planthopper (BPH) infestation induce weaker resistance to rice pathogens. We compared changes in gene expression in rice plants infested with WBPH and BPH to gain some insight into the WBPH-induced resistance to rice pathogens. An analysis, using microarrays, of gene expression in rice plants infested with these planthoppers revealed that WBPH infestation caused high induction of many defense-related genes including pathogenesis-related (PR) genes than BPH infestation. Furthermore, hydroperoxide lyase 2 (OsHPL2) which is an enzyme to produce C6 volatiles was induced by WBPH infestation, but not by BPH infestation. Experiment Overall Design: Agilent rice oligo microarray was used to investigate the gene expression profiling in rice plants infested with WBPH or BPH. Total RNA was extracted from pooled leaf blades infested with WBPH or BPH for 24 h and from mock-treated pooled leaf blades. Total RNA (200 ng) was labeled with Cy-3 or Cy-5 using an Agilent low RNA input linear amplification kit. Fluorescently labeled targets were hybridized to Agilent rice oligo microarrays. Hybridization and wash processes were performed according to the manufacturer’s instructions, and hybridized microarrays were scanned using an Agilent DNA microarray scanner. Agilent Feature Extraction software was employed for the image analysis and data extraction processes. Fold changes in expression level in each treatment were compared with those of the respective mock-treated controls. In each treatment, the experiment was performed independently three times.
Project description:The differentiation of specialized feeding sites in Zea mays root cells in response to nematode infestation involves substantial cellular reprogramming of host cells that is not well characterized at the molecular level. Expression data was generated from Zea mays root cells undergoing giant cell formation due to nematode infestation and from non-infested control root cells. Cells were laser captured 14 and 21 days after infestation. Each time point (14 day and 21 day) consisted of three biological replicates per treatment (control root cells or giant cells). Control cells were captured from an area ~13,000,000 um2 in size and giant cells were captured from an area ~5,000,000 um2 in size. RNA samples were isolated using the PicoPure RNA Isolation Kit (Arcturus, Mountain View, USA). RNA amplifications were carried out with the NuGEN WT-Ovation Pico kit.
Project description:The differentiation of specialized feeding sites in Arabidopsis root cells in response to nematode infestation involves substantial cellular reprogramming of host cells that is not well characterized at the molecular level. Expression data was generated from Arabidopsis root cells undergoing giant cell formation due to nematode infestation and from non-infested control root cells. Cells were laser captured 14 and 21 days after infestation. Samples, collected 14 days post infestation, consisted of three biological replicates per treatment (control root cells or giant cells). RNA samples were isolated from ~150 control cells or from ~80 giant cells using the PicoPure RNA Isolation Kit (Arcturus, Mountain View, USA). RNA amplifications were carried out with the NuGEN WT-Ovation Pico kit. GSM546568-GSM546577: Control and giant cells collected 21 days post infestation.