Evaluation of Housekeeping Genes for Quantitative Real-Time PCR Analysis of Bradysia odoriphaga (Diptera: Sciaridae)

The soil insect Bradysia odoriphaga (Diptera: Sciaridae) causes substantial damage to Chinese chive. Suitable reference genes in B. odoriphaga (Bradysia odoriphaga) have yet to be identified for normalizing target gene expression among samples by quantitative real-time PCR (qRT-PCR). This study was focused on identifying the expression stability of 12 candidate housekeeping genes in B. odoriphaga under various experiment conditions. The final stability ranking of 12 housekeeping genes was obtained with RefFinder, and the most suitable number of reference genes was analyzed by GeNorm. The results revealed that the most appropriate sets of internal controls were RPS15, RPL18, and RPS18 across developmental phases; RPS15, RPL28, and GAPDH across temperatures; RPS15 and RPL18 across pesticide treatments; RSP5, RPS18, and SDHA across photoperiods; ACTb, RPS18, and RPS15 across diets; RPS13 and RPL28 across populations; and RPS15, ACTb, and RPS18 across all samples. The use of the most suitable reference genes versus an arbitrarily selected reference gene resulted in significant differences in the analysis of a target gene expression. HSP23 in B. odoriphaga was found to be up-regulated under low temperatures. These results will contribute to the standardization of qRT-PCR and will also be valuable for further research on gene function in B. odoriphaga

Study on Strength Variation of Permeable Concrete Based on Differential Calorimetry Method and Multi-Index Test

With the implementation of the Xiongan New Area and the urbanization construction plan surrounding the Beijing-Tianjin satellite, it is urgent to study and apply the sponge-permeable paving materials to solve the “urban waterlogging and heat island effect.” In this paper, the hydration microscopic characteristics of cementitious materials are analyzed by means of the differential scanning calorimetry (DSC) test. The test of water-cement ratio, porosity, and gradation structure on the strength and permeability coefficient is emphasized, and the strength change rule of porous permeable concrete is obtained. The research results are shown as follows: (1) The DSC test shows that the effect of temperature on the hydration process of cement is obvious. With the increase of temperature, the two exothermic peaks of cement hydration increase significantly and tend to overlap and the exothermic process is shortened. At 85°C, only one exothermic peak appeared, indicating that C3S hydration and ettringite formation process were completed at the same time in a relatively short time. (2) The optimal water-binder ratio of pervious concrete ranges from 0.24 to 0.30, and the optimal porosity ranges should be controlled within the range of 15%–25%. Moreover, the open gradation of pervious concrete mix ratio design is selected, and the cement content should be within the range of 20%–25%. (3) The mathematical model of permeability coefficient and porosity ratio of permeable concrete is established: k=−0.0003n2+0.0057n−0.0044; the mathematical model of permeability coefficient and compressive strength of open-graded pervious concrete: k=0.99006−0.05389c+8.02×10−4c2. The research results can provide theoretical support and technical guidance for the design and construction of permeable materials of sponge cities in Xiongan New Area and surrounding ring new area, which are of great engineering value

The gene expression profile of peripheral blood mononuclear cells from EV71-infected rhesus infants and the significance in viral pathogenesis.

Enterovirus 71 (EV71) is the major pathogen responsible for fatal hand, foot and mouth disease (HFMD). Our previous work reported on an EV71-infected rhesus monkey infant model that presented with histo-pathologic changes of the central nervous system (CNS) and lungs. This study is focused on the correlated modulation of gene expression in the peripheral blood mononuclear cells (PBMCs) from EV71-infected rhesus monkey infants. The expression of more than 500 functional genes associated with multiple pathways was modulated. The expression of genes associated with immune inflammatory responses was up-regulated during the period from days 4 to 10 post-infection. The expression of two genes (TAC1 and IL17A), which play major roles in inflammatory reactions, was remarkably up-regulated during the infection period. Furthermore, a higher expression level of the TAC1 gene was identified in the CNS compared to the lungs, but a high expression level of the IL-17A gene was observed in the lungs and not in the CNS. The results of this study suggest at least two facts about EV71 infection, which are that: the TAC1 gene that encodes substance P and neurokinin-A is present in both PBMCs and the hypothalamus; and the up-regulation of IL-17A is sustained in the peripheral blood

Study of the Integrated Immune Response Induced by an Inactivated EV71 Vaccine

<div><p>Enterovirus 71 (EV71), a major causative agent of hand-foot-and-mouth disease (HFMD), causes outbreaks among children in the Asia-Pacific region. A vaccine is urgently needed. Based on successful pre-clinical work, phase I and II clinical trials of an inactivated EV71 vaccine, which included the participants of 288 and 660 respectively, have been conducted. In the present study, the immune response and the correlated modulation of gene expression in the peripheral blood mononuclear cells (PBMCs) of 30 infants (6 to 11 months) immunized with this vaccine or placebo and consented to join this study in the phase II clinical trial were analyzed. The results showed significantly greater neutralizing antibody and specific T cell responses in vaccine group after two inoculations on days 0 and 28. Additionally, more than 600 functional genes that were up- or down-regulated in PBMCs were identified by the microarray assay, and these genes included 68 genes associated with the immune response in vaccine group. These results emphasize the gene expression profile of the immune system in response to an inactivated EV71 vaccine in humans and confirmed that such an immune response was generated as the result of the positive mobilization of the immune system. Furthermore, the immune response was not accompanied by the development of a remarkable inflammatory response.</p> <p> <em>Clinical Trial Registration: NCT01391494 and NCT01512706.</em></p> </div

The characterization of the gene expression profiles of the PBMCs from EV71-infected rhesus infants.

<p>(a) The expression heat map of important genes associated with transcriptional regulation during EV71 infection. (b) The up-regulated expression of important genes associated with cellular metabolism, structure and signal transmission during EV71 infection. (c) The down-regulated expression of important genes associated with cellular metabolism, structure and signal transmission during EV71 infection. The color scale indicates the levels of gene expression from low (green, blue and purple) to high (red, pink and yellow). The values are shown on a log₂ scale.</p

Vaccination with the inactivated EV71 vaccine modulated gene expression.

<p>A global view of gene modulation in the PBMCs of vaccinated infants (Vac, n = 20), 28 days after the vaccine booster, compared to the placebo cohort (Pla, n = 10) was determined by microarray analysis. The genes were classified according to their function. A heat map representation (a) and principal component analysis (PCA) (b) generated using the significantly modulated genes from at least one comparison versus day 0 are shown. (a) The color scale shows the significantly modulated genes from up- (dark red) to down-regulated (dark blue) to the right of each image. The values are shown on a log₂ scale. (b) The Y value is the log₂-fold-changes in gene expression of the vaccinated group vs. the placebo group. The up-regulated changes are above the baseline (Y = 0, the same with placebo group), and the down-regulated changes are below the baseline. The bars represent the maximum and minimum. The mean of the fold-changes in gene expression with a 95% CI is shown as the rectangle. The line in the rectangle represents the mean of these fold changes in gene expression. The number of genes that changed after immunization is shown beside the rectangles on the image.</p