Novel Environmentally Friendly RNAi Biopesticides: Targeting V-ATPase in Holotrichia parallela Larvae Using Layered Double Hydroxide Nanocomplexes.

RNA interference (RNAi)-based biopesticides offer an attractive avenue for pest control. Previous studies revealed high RNAi sensitivity in Holotrichia parallela larvae, showcasing its potential for grub control. In this study, we aimed to develop an environmentally friendly RNAi method for H. parallela larvae. The double-stranded RNA (dsRNA) of the V-ATPase-a gene (HpVAA) was loaded onto layered double hydroxide (LDH). The dsRNA/LDH nanocomplex exhibited increased environmental stability, and we investigated the absorption rate and permeability of dsRNA-nanoparticle complexes and explored the RNAi controlling effect. Silencing the HpVAA gene was found to darken the epidermis of H. parallela larvae, with growth cessation or death or mortality, disrupting the epidermis and midgut structure. Quantitative reverse transcription-polymerase chain reaction and confocal microscopy confirmed the effective absorption of the dsRNA/LDH nanocomplex by peanut plants, with distribution in roots, stems, and leaves. Nanomaterial-mediated RNAi silenced the target genes, leading to the death of pests. Therefore, these findings indicate the successful application of the nanomaterial-mediated RNAi system for underground pests, thus establishing a theoretical foundation for developing a green, safe, and efficient pest control strategy.

Modeling knot features using branch scars from Mongolian oak (Quercus mongolica).

Wood quality is an important indicator for modern sawmills. Internal wood characteristics can be derived from their correlations with external appearances. In this study, we developed linear regression models to predict knot size from surface features of Mongolian oak (Quercus mongolica) using data collected from 53 trees. For this, manual measurements and X-ray computed tomography scanning technology was respectively used to obtain internal and external features of 1,297 knots. Our results showed that Mongolian oak knots were generally concentrated in the middle part of oak stems, with fewer knots observed at the top and base. The parameters of knot and scar showed significant correlations (P < 0.01), where length and diameter of the corresponding external scar increase with increasing the length and diameter of a knot. The corresponding external scar can be used as an effective indicator to predict the internal value of oak logs. The accuracy of our constructed model is more than 95% when assessed against independent test samples. These models thus can be applied to improve the practical production of oak timber and reduce commercial loss caused by knots. These additional data can improve the estimation of the influence of knots on wood quality and provide a theoretical foundation for investigating the characteristics of hardwood knots.

Group V Chitin Deacetylases Influence the Structure and Composition of the Midgut of Beet Armyworm, Spodoptera exigua.

Chitin deacetylase (CDA) can accelerate the conversion of chitin to chitosan, influencing the mechanical properties and permeability of the cuticle structures and the peritrophic membrane (PM) in insects. Putative Group V CDAs SeCDA6/7/8/9 (SeCDAs) were identified and characterized from beet armyworm Spodoptera exigua larvae. The cDNAs of SeCDAs contained open reading frames of 1164 bp, 1137 bp, 1158 bp and 1152 bp, respectively. The deduced protein sequences showed that SeCDAs are synthesized as preproteins of 387, 378, 385 and 383 amino acid residues, respectively. It was revealed via spatiotemporal expression analysis that SeCDAs were more abundant in the anterior region of the midgut. The SeCDAs were down-regulated after treatment with 20-hydroxyecdysone (20E). After treatment with a juvenile hormone analog (JHA), the expression of SeCDA6 and SeCDA8 was down-regulated; in contrast, the expression of SeCDA7 and SeCDA9 was up-regulated. After silencing SeCDAV (the conserved sequences of Group V CDAs) via RNA interference (RNAi), the layer of intestinal wall cells in the midgut became more compact and more evenly distributed. The vesicles in the midgut were small and more fragmented or disappeared after SeCDAs were silenced. Additionally, the PM structure was scarce, and the chitin microfilament structure was loose and chaotic. It was indicated in all of the above results that Group V CDAs are essential for the growth and structuring of the intestinal wall cell layer in the midgut of S. exigua. Additionally, the midgut tissue and the PM structure and composition were affected by Group V CDAs.

Transcriptomic insights into the effects of abscisic acid on the germination of Magnolia sieboldii K. Koch seed.

Magnolia sieboldii K. Koch is a deciduous tree species. However, the wild resource of M. sieboldii has been declining due to excessive utilization and seed dormancy. In our previous research, M. sieboldii seeds have morphophysiological dormancy and low germination rates under natural conditions. The aim of the present study was to identify the genes involved in dormancy maintenance. In this study, the germination percentage of M. sieboldii seeds negatively correlated with the content of endogenous abscisic acid (ABA). The hydration of seeds for germination showed three distinct phases. Five key time points were identified: 0 h imbibition (dry seed, GZ), 0 day after imbibition (DAI), 16 DAI, 40 DAI, and 56 DAI. The comprehensive transcript profiles of M. sieboldii seeds treated with ABA and water at the five key germinating stages were obtained. A total of 9641 differentially expressed genes (DEGs) were identified, and 208 and 197 common DEGs were found throughout the ABA and water treatments, respectively. Compared with that in the GZ, 518, 696, 2133, and 1535 DEGs were identified in the SH group at 0, 16, 40 and 56 DAI, respectively. 666, 1725, 1560 and 1415 DEGs were identified in the ABA group at 0, 16, 40, and 56 DAI, respectively. Among the identified DEGs, 12 722 were annotated with GO terms, the top three enriched GO terms were different among the DEGs at 56 DAI in the ABA vs. SH treatments. KEGG pathway enrichment analysis for DEGs indicated that oxidative phosphorylation, protein processing in endoplasmic reticulum, starch and sucrose metabolism play an important role in seed response to ABA. 1926 TFs are obtained and classified into 72 families from the M. sieboldii transcriptome. Results of differential gene expression analysis together with qRT-PCR indicated that phase II is crucial for rapid and successful seed germination. This study is the first to present the global expression patterns of ABA-regulated transcripts in M. sieboldii seeds at different germinating phases.

[Responses of stand growth, regeneration, and understory species diversity in Quercus mongolica secondary forest to stand density.] / è’™å¤æ Žæ¬¡ç”Ÿæž—çš„ç”Ÿé•¿æ›´æ–°ä¸Žæž—ä¸‹æ¤è¢«å¤šæ ·æ€§å¯¹æž—åˆ†å¯†åº¦çš„å“åº”.

Six Quercus mongolica plots with an area of 0.1 hm2 were thinned in 2018. A field survey was carried out in 2020 to examine the effects of different stand densities (high: 900 trees·hm-2; medium: 720 trees·hm-2; low: 600 trees·hm-2) on growth and regeneration of stands and understory species diversity of secondary Q. mongolica forests in Qingyuan, Liaoning Province. Due to the short interval after thinning, there was no significant difference in tree height and diameter at breast height under different densities. However, the crown symmetry index under low stand density was significantly higher than that of high stand density, indicating that crown growth was more sensitive to stand density than trunk growth. The abundance of seedlings was the highest in the medium density, and the basal diameter of the seedlings with the same height was significantly higher, and the seedling regeneration and growth at the medium density were much better than the other two densities. A total of 70 species were recorded, belonging to 41 families and 67 genera. Quercus mongolica, Lespedeza bicolor, Melampyrum roseum, and Potentilla freyniana were the dominant species of trees and herbs, respectively. Simpson index, Pielou index and Shannon index of shrub layer and herb layer were the highest at the medium density. It indicated that the stand density of 720 trees·hm-2 could help maintain the sustainable development of Q. mongolica secondary forest in the mountainous area of eastern Liaoning.

Cloning and expression analyses of a Pyrabactin Resistance 1 (PYR1) gene from Magnolia sieboldii K. Koch.

Magnolia sieboldii K. Koch is endemic to China and has high medicinal and ornamental values. However, its seed exhibits morphophysiological dormancy, and the molecular mechanisms of which are not clearly understood. To reveal the regulation mechanism of the ABA signal in seed dormancy, the M. sieboldii ABA receptor Pyrabactin Resistance 1 (PYR1) gene was cloned and analyzed. Analysis of the MsPYR1 sequence analysis showed that the full-length cDNA contained a complete open reading frame of 987 bp and encoded a predicted protein of 204 amino acid residues. The protein had a relative molecular weight of 22.661 kDa and theoretical isoelectric point of 5.01. The transcript levels of MsPYR1 were immediately upregulated at 16 DAI and then decreased at 40 DAI. The highest transcript level of MsPYR1 was found in the dry seeds, indicating that the MsPYR1 gene may play an important role in the regulation of dormancy. The MsPYR1 gene cDNA was successfully expressed in E. coli Rosetta (DE3), and the protein bands were consistent with the prediction. The Anti-MsPYR1antibody could detect the expression of MsPYR1 in M. sieboldii. The results provided a foundation for further study of the function of the MsPYR1 gene.ABBREVIATIONSABA: Abscisic acid; MPD: morphophysiological; PYR1: Pyrabactin Resistance1; PYL: Pyr1-Like; RCAR: Regulatory Components of Aba Receptors; PP2C: protein phosphatases 2C; SnRK2: sucrose non-fermenting1-related protein kinase2; DAI: day after imbibition; NCBI: National Center for Biotechnology Information; BCA: Bicinchoninic acid; CDD: Conserved Domains.

Integrated RNA and miRNA sequencing analysis reveals a complex regulatory network of Magnolia sieboldii seed germination.

Magnolia sieboldii K. Koch (M. sieboldii) is a deciduous Chinese tree species of the Magnoliaceae family with high ornamental, medicinal, and economic benefits. The germination of M. sieboldii seeds under natural conditions is extremely difficult, thereby hindering the cultivation and breeding of this important species. The molecular mechanisms underlying M. sieboldii seed germination remain unclear due to the lack of genomic and transcriptomic resources. Here, we integrated both mRNA and miRNA sequencing to identify the genes and pathways related to M. sieboldii germination. A comprehensive full-length transcriptome containing 158,083 high-quality unigenes was obtained by single-molecule real-time (SMRT) sequencing technology. We identified a total of 13,877 genes that were differentially expressed between non-germinated and germinated seeds. These genes were mainly involved in plant hormone signal transduction and diverse metabolic pathways such as those involving lipids, sugars, and amino acids. Our results also identified a complex regulatory network between miRNAs and their target genes. Taken together, we present the first transcriptome of M. sieboldii and provide key genes and pathways associated with seed germination for further characterization. Future studies of the molecular basis of seed germination will facilitate the genetic improvement M. sieboldii.

RNA interference-mediated functional characterization of Group I chitin deacetylases in Holotrichia parallela Motschulsky.

Chitin deacetylases (CDAs, EC 3.5.1.41) catalyze the N-deacetylation of chitin to produce chitosan, which is essential for insect survival. Hence, CDAs are promising targets for the development of novel insecticidal drugs. In this study, the putative Group I chitin deacetylase genes HpCDA1, HpCDA2-1 and HpCDA2-2 were identified from Holotrichia parallela. Conserved domain database search identified a chitin-binding peritrophin-A domain (ChBD), a low-density lipoprotein receptor class A domain (LDLa), and a putative CDA-like catalytic domain. RT-qPCR analysis showed that the Group I HpCDAs were expressed in various tissues and predominant in the integument. The developmental expression patterns from the first-instar to third-instar larvae showed that HpCDAs were highly expressed on the first day and gradually declined after molting. The functional characteristics of the Group I CDAs in cuticle organization were examined using RNA interference (RNAi) and transmission electron microscopy (TEM) methods. Administration of double-stranded HpCDA (dsHpCDA) through larval injection could suppress the expression levels of HpCDA1 and HpCDA2, thus resulting in abnormal or lethal phenotypes. TEM analysis revealed that RNAi of either HpCDA1 or HpCDA2 remarkably affected the cuticle integrity, as evidenced by cuticle disorganization and chitin laminae disruption, suggesting the crucial role of CDAs in chitin modification. These experimental results demonstrate the important contribution of putative key genes involved in chitin metabolism, and provide a foundation for developing new strategies to control H. parallela.

Functions of microRNA in seed development, dormancy and germination processes.

In plants, microRNAs (miRNAs) are a class of non-coding small RNAs that play important roles in various biological processes, such as seed development, dormancy and germination. In these stages, seeds are regulated by various extrinsic environmental factors and intrinsic hormones. Different species of seeds have distinct development and dormancy characteristics. Recent investigations have identified more and more miRNAs to regulate seed morphogenesis, material metabolism and stress response capacities through hormonal signal transduction, antioxidant effects and transcription factors, etc. In this review, we summarize recent progresses on regulatory mechanisms of miRNAs, and also complex multilevel regulation of miRNAs in the process of seed development, dormancy and germination. We hope to provide insights into further research.

Proteomic analysis of Magnolia sieboldii K. Koch seed germination.

Magnolia sieboldii is a deciduous tree native to China. This species has a deep dormancy characteristic. To better understand seed germination, we used protein analysis of changes in seed protein at 0, 65, 110 and 150 d of stratification. Comparative 2DE analysis of M. sieboldii seed protein profiles at 0, 65, 110 and 150 d of stratification revealed 80 differentially abundance protein species. Comparative analysis showed that ADP-glucose pyrophosphorylase small subunit was degraded during germination. In particular, it was degraded almost completely at 110 d of germination. Starch granules in the microstructure decreased after 65 d of stratification. Starch granules provided a sufficient amount of substrates and ATPs for subsequent germination. Four storage protein species were identified, of which all were down accumulated. Spots 44 and 46 had different MW and pI values, spots 36 and 46 had nearly the same MW with pI shift in the 2-DE gels, suggesting that they might be present as different isoforms of the same protein family and the post translational modification. Our results suggested that degradation of starch granules and storage protein species prepared the seed embryo for growth, as well as regulated seed germination. The present proteomics analysis provides novel insights into the mobilisation of nutrient reserves during the germination of M. sieboldii seeds. BIOLOGICAL SIGNIFICANCE: To better understand seed germination, a complex developmental process, we developed a proteome analysis of M. sieboldii seed. We performed the first comprehensive proteomic and microstructure analysis during different seed stratification stages of M. sieboldii. Among the 80 protein species, 26 were identified, 7 and 14 protein species were up or down accumulated significantly. Many of the identified key proteins were involved in embryo development, starch biosynthesis and energy metabolism, Microstructure of stratification seed analysis revealed degradation of starch was used for preparing the seed embryo for growth. These data may help us to develop a comprehensive understanding of the physiological status and mobilisation mechanisms in M. sieboldii seed germination.

Proteomics profiling of ethylene-induced tomato flower pedicel abscission.

The control of abscission is an important agricultural concern because of its substantial effect on crop yield and quality. Changes in gene expression are correlated with the ethylene-mediated execution of abscission. However, only few large-scale proteomic studies focused on tomato pedicel abscission. Isobaric tag for relative and absolute quantification labeling was used to examine the protein and phosphoprotein changes in the tomato pedicel AZ (AZ) treated with ethylene or 1-methylcyclopropene. Among the 1429 quantified proteins, 383 unique peptides corresponding to 166 proteins showed higher than 1.5-fold change in abundance. A total of 450 phosphopeptides were detected, among which 85 phosphopeptides corresponding to 73 phosphoproteins were significantly regulated (>1.5-fold abundance change) in response to ethylene. Protein and phosphoprotein sets showed 26 similar proteins. Six phosphorylation motifs were extracted from the 138 phosphorylation sites. By analyzing translational and modification levels, we found that the modification level was not due to the translational changes. Comparison between the protein and phosphoprotein functions revealed that the proteins acted mainly in the metabolic process and showed catalytic activity, whereas most of the phosphoproteins showed signaling and transporting activities. Data revealed the unique features of the AZ phosphoproteomics, thereby suggesting the involvement of a complex network of kinase-substrate and phosphatase-substrate interactions in response to ethylene. Some phosphorylation sites from calcium-dependent protein kinase (CDPK5(S523)), CDPK5(S527), and SRL3(S329) were also found to perform protective functions for AZ and to be helpful in ethylene signal transduction. BIOLOGICAL SIGNIFICANCE: Organ abscission has both positive and negative roles. Abscission is conducive for the fall of ripe fruits and the release and dispersion of seeds, but abscission has been a major limiting factor for crop productivity. Hence, more details about the process may aid in the regulation of organ abscission. However, at present, the detailed mechanism of abscission is still unclear. In tomato, several transcriptome analyses were performed using pedicels as materials. Yet, no large-scale proteomics and phosphoproteomic studies of abscission zone have been reported so far. Hence, in this present study, we determined the ethylene-induced changes in the proteomics and phosphoproteomics of tomato flower AZ tissue using the isobaric tag for relative and absolute quantification (iTRAQ). Proteomics data from both data sets revealed the differentially expressed proteins that are associated with the translational and modification levels relevant to abscission mechanism. Two key proteins (CDPK (CDPK5(S523) and CDPK5(S527)) and SRL3(S329)) among ethylene signal transduction and defense-related proteins were obtained from the phosphoproteins. The set of tomato phosphorylation sites presented in this work is useful in at least two ways. First, as a database resource, the data would facilitate research on the identified phosphoproteins. Second, the identified sites of the related proteins could provide enough knowledge for further experiments. Hence, our results contribute to the understanding of the mechanism of abscission in plants.

[Variations of arsenic species in the solution of arsenic-contaminated paddy soil under flooding and at different temperatures].

Taking arsenic (As)-contaminated paddy soil as test object, and by using high performance liquid chromatography inductively coupled plasma-mass spectrometry (HPLC-ICP-MS), this paper studied the variations of As species in soil solution when the soil was sterilized or non-sterilized and incubated at different temperatures (5, 27, and 50 degrees C) under flooding. In the soil solution (pore water), only As(III) (arsenite), As(V) (arsenate), and DMA(V) ( dimethylarsinic acid) were detected, but no MMA(V)(mono methylarsinic acid) was found. With the increasing time of flooding and at the test temperatures, arsenite became the predominant species, averagely accounting for 64%, followed by As(V), with the proportion of 35%, and DMA(V), with the least proportion of 1%. Soil sterilization or non-sterilization had less effect on the concentrations of As(III) and DMA(V) in the soil solution, but remarkably affected the reduction of As(V) and the methylation of As(III). The promotion effect of soil sterilization decreased gradually with the increasing time of flooding and incubation. At 50 degrees C and after flooded for 23 days, the DMA(V) concentration in sterilized soil solution was the highest and up to 23.7 ng x mL(-1), indicating that some thermophilic microbes remained in sterilized soil became predominant species, and promoted the methylation of As(III) In sum, the total arsenic concentration in non-sterilized soil at incubation temperature 27 degrees C and flooded for 23 days was relatively low (501 ng x mL(-1)), and thus, in As-contaminated paddy rice planting areas, to adopt the water management mode of short cycle flooding-non-flooding could decrease the As level in soil solution as far as possible, and in the same time, save water resources and ensure yielding.

Arsenic speciation in Chinese Herbal Medicines and human health implication for inorganic arsenic.

Rice and drinking water are recognized as the dominant sources of arsenic (As) for human intake, while little is known about As accumulation and speciation in Chinese Herbal Medicines (CHMs), which have been available for many hundreds of years for the treatment of diseases in both eastern and western cultures. Inorganic arsenic was the predominant species in all of CHMs samples. The levels of inorganic arsenic in CHMs from fields and markets or pharmacies ranged from 63 to 550 ng/g with a mean of 208 ng/g and 94 to 8683 ng/g with a mean of 1092 ng/g, respectively. The highest concentration was found in the Chrysanthemum from pharmacies. It indicates that the risk of inorganic As in CHMs to human health is higher in medicines from markets or pharmacies than that collected directly from fields. Some CHMs may make a considerable contribution to the human intake of inorganic arsenic.