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Enhanced detoxification of exogenous toluene gas in transgenic Ardisia pusilla expressing AtNDPK2 gene
The Arabidopsis nucleoside diphosphate kinase 2 (AtNDPK2) gene is known to regulate the cellular redox state, and to enhance tolerance to multiple stressors in plants. In this study, we transferred AtNDPK2 under the stress-inducible promoter SWPA2 into Ardisia pusilla to enhance the plants’ ability to detoxify toluene gas. Thirty transgenic A. pusilla lines were confirmed by PCR analysis with AtNDPK2 and NPTII gene-specific primers. In addition, four transgenic A. pusilla lines were further confirmed by Southern blot analysis to verify the gene copy number. Three transgenic lines showed a single-copy transgene insertion, and one transgenic line had two transgene insertions. To test the gene expression of AtNDPK2 in the transgenic A. pusilla lines exposed to and not exposed to toluene treatment, qRT-PCR analysis was performed. The gene expression of AtNDPK2 in transgenic A. pusilla plants exposed to toluene treatment was significantly higher than that of transgenic plants not exposed to toluene treatment. Finally, we measured toluene removal efficiency of the transgenic and non-transgenic A. pusilla lines exposed to toluene-contaminated air. There was a statistically significant difference between the transgenic and non-transgenic A. pusilla lines at all time points (p < 0.001). The highest toluene removal efficiency (797.33 ± 59.41 µg m−3 cm−2 leaf area) was recorded in the transgenic A. pusilla line NDPK2-12-4 after 3 h of exposure to toluene, while the non-transgenic line showed little toluene removal efficiency (206.2 ± 31.19 µg m−3 cm−2 leaf area). These results suggest that the capacity for detoxifying toluene gas is related to the AtNDPK2 gene in A. pusilla. Therefore, this study provides useful results to reduce toluene pollution in indoor air.
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Survival strategy of Pseudomonas aeruginosa on the nanopillar topography of dragonfly (Pantala flavescens) wing
Discovery of nanopillars on the surface of the insect wings had led to the understanding of its bactericidal property. Nanopillar topography is deterrent to only those bacteria that are attached, or in close contact with the nanopillars. The present study investigated the variation in the viability of Pseudomonas aeruginosa strains PAO1 (virulent) and ATCC 9027 (avirulent) on the wing surface of dragonfly (Pantala flavescens). Viability study indicated that only 0.2% ATCC 9027 survived when incubated with wing for 48 h in Phosphate buffered saline, while under the same conditions 43.47% PAO1 survived. Enumeration of Pseudomonas attached to wing surface suggested that, the number of PAO1 attached on the wing surface was three times lesser than ATCC 9027. Propensity of attachment of P. aeruginosa strains PAO1 and ATCC 9027 on the wing surface investigated using scanning probe microscope indicated that P. aeruginosa ATCC 9027 showed adhesion to 88% of regions and, PAO1 showed adhesion to only 48% regions tested on wing surface. PAO1 survived the bactericidal effect of wing surface by evading attachment. Three clinical isolates tested which showed viability similar to PAO1 strain, also showed lower propensity to attach to wing surface. Transcriptional level analyses using RT-PCR suggested that flagellar genes (fliE and fleS) were downregulated and genes responsible for reversible to irreversible attachment (gcbA and rsmZ) were upregulated in ATCC 9027 than PAO1 on wing surface, indicating relatively higher attachment of ATCC 9027 on wing surface. The study suggests that virulent strains of P. aeruginosa may evade attachment on wing surface. The results gain significance as bioinspired surfaces are being created towards developing antibacterial medical implants and other antibacterial surface applications.
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Expression of the microfold cells in three-dimensional coculture system for in vitro cultivation of human norovirus
Optimization of Caco-2 cells monoculture in the alginate hydrogel beads showed optimum number of cells of 1 × 106 cells/ml, indicated by the intact structure of the beads. Result of SEM showed clear structure of monoculture in the alginate hydrogel beads indicated by the presence of smooth and regular apical surface while the coculture showed reduced apical surface of M cells. The result of WB showed downregulation of Ulex europaeus antibody expression.
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