The non-negligible non-specific adsorption of oligonucleotides in target-immobilized Mag-SELEX.

Target-immobilized magnetic beads-based Systematic Evolution of Ligands by Exponential Enrichment (target-immobilized Mag-SELEX) has emerged as a powerful tool for aptamer selection owing to its convenience, efficiency, and versatility. However, in this study we systematically investigated non-specific adsorption in target-immobilized Mag-SELEX and found that the non-specific adsorption of the oligonucleotides to target-labeled magnetic beads was comparable to that of the screening libraries, indicating a substantial portion of captured sequences likely stem from non-specific adsorption. Longer nucleic acid sequences (80 nt and above, such as polyA80 and yeast tRNA) were found to attenuate this non-specific adsorption, with more complex higher-order structures demonstrating greater efficacy, while dNTP and short sequences such as primer sequences (20 nt), polyT(59), or polyA(59), did not possess this capability. Various evidence suggested that hydrophobic interactions and other weak interactions may be the primary underlying cause of non-specific adsorption. Additionally, surface modification of magnetic beads with polar molecule polyethylene glycol (PEG) also yielded a significant reduction in non-specific adsorption. In conclusion, our research underscores the critical importance of closely monitoring non-specific adsorption in target-immobilized Mag-SELEX.

Heat-dependent fecundity enhancement observed in Nilaparvata lugens (Hemiptera: Delphacidae) after treatment with triazophos.

The brown planthopper Nilaparvata lugens (Stål) (Hemiptera: Delphacidae) is a serious pest of rice crops in the temperate and tropical regions of Asia and Australia, and it is also a classic example of an insecticide-induced resurgent pest. Brown planthopper outbreaks have been reported to be closely associated with pesticide application. Previous studies have shown that the insecticide triazophos enhances thermal tolerance and fecundity in brown planthopper. However, the effects of triazophos and high temperature on reproductive capacity have not been studied in high temperature-conditioned reciprocal pairs of adult males and females. The present experiments showed that triazophos enhanced the reproductive capacity of brown planthopper under high temperature (34°C). The number of eggs laid by females treated with 40 ppm triazophos at 34°C approximately doubled compared with untreated insects. Furthermore, the triazophos-induced fecundity enhancement at 34°C was significantly greater than that at 26°C, and the number of eggs laid for mating pair of adult males at 34°C with adult females under 34°C (♂(34) × â™€(34)) were significantly greater than that of adult males at 26°C (♂(26) × â™€3(4)), suggesting that this insecticide enhances the resistance of brown planthopper to high-temperature stress. Insecticide-induced enhancement of reproductive capacity in brown planthopper under high temperatures should be of great concern, and it has important implications for forecasting future brown planthopper outbreaks as well as the pesticide-selection process.

Molecular basis for insecticide-enhanced thermotolerance in the brown planthopper Nilaparvata lugens Stål (Hemiptera:Delphacidae).

Climate change is likely to have marked ecological effects on terrestrial ecosystems, including the activities of insect pests. Most attention has focused on the increasing geographical ranges of pests; however, if extrinsic factors enhance their thermotolerance, populations may express increased voltinism and longer daily and annual activity periods. These changes in pest populations have the potential for severe consequences, including increased crop losses and decreased food security at the global level. The brown planthopper (BPH) Nilaparvata lugens Stål (Hemiptera: Delphacidae) is a serious pest of rice crops in temperate and tropical regions of Asia. It is often present in rice microclimates at temperatures close to its maximum thermotolerance. Recent BPH outbreaks in tropical Asia are considered to be associated with excess use of pesticides and increasing temperature. This study tested whether exposure to sublethal concentrations of triazophos (tzp), an insecticide widely used in Asian rice production, enhances thermotolerance of BPH. Tzp exposure (40 ppm at 40 °C) significantly decreased mortality (from 94% in controls to 50% at 48 h post-treatment) and increased lethal mean time (LT50 ) of adults by 17.2 h. To investigate the underlying molecular mechanism of this tzp-enhanced thermotolerance, we selected Hsp70 and Arginine kinase (Argk) for detailed study. Transcripts encoding both proteins in third-instar nymphs and brachypterous adult females were up-regulated, compared with controls, after exposure to tzp. RNAi silencing of both genes demonstrated that Hsp70 and Argk are essential for survival and tzp-increased thermotolerance. We propose that tzp induces thermotolerance in BPHs by increasing the expression of genes that act in cell protection mechanisms. The significance of our proposal relates to the importance of understanding the influence of sublethal concentrations of insecticides on pest biology. In addition to its influence on thermotolerance, tzp also enhances BPH reproduction. We infer that exposure to a pesticide stressor can produce cross-tolerance, that is, increased tolerance to one stressor also increases tolerance to other stressors, including temperature. Aside from needing a better understanding of these effects in nature and in other pest/cropping systems, we suggest that pest management programmes can be improved with better understanding of the influences of stressors, including increased environmental temperatures and sublethal concentrations of insecticides, on pest biology.

The effects of triazophos on the trehalose content, trehalase activity and their gene expression in the brown planthopper Nilaparvata lugens (Stål) (Hemiptera: Delphacidae).

A previous study demonstrated that the flight capacity of Nilaparvata lugens adults treated with triazophos was enhanced significantly. However, the physiological and regulative mechanisms of the flight enhancement are not well understood. Trehalose is a primary blood sugar in insects, and the enzyme trehalase is involved in energy metabolism. The present study investigated the effects of triazophos on the trehalose content, trehalase activity (soluble trehalase and membrane-bound trehalase) and the mRNA transcript levels of their corresponding genes (NlTre-1 and NlTre-2) in fifth instar nymphs, as well as in the brachypterous and macropterous N. lugens adult females. Our findings showed that the trehalose content in fifth instar nymphs as well as in the brachypterous and the macropterous adults significantly decreased following triazophos treatment. However, the glucose content, soluble trehalase activity and expression level of NlTre-1 mRNA increased significantly compared to the controls. No significant enhancement of NlTre-2 expression was found, indicating that regulation of energy metabolism of triazophos-induced flight capacity in N. lugens was not associated with NlTre-2 expression. In addition, soluble trehalase activity and the expression level of NlTre-1 mRNA in the macropterous females was significantly higher than that in the brachypterous females. The present findings provide valuable information on the molecular and regulative mechanisms of the increased flight capacity found in adult N. lugens after treatment with triazophos.