Comparative transcriptome and DNA methylation analysis in temperature-sensitive genic male sterile wheat BS366.

BACKGROUND: Known as the prerequisite component for the heterosis breeding system, the male sterile line determines the hybrid yield and seed purity. Therefore, a deep understanding of the mechanism and gene network that leads to male sterility is crucial. BS366, a temperature-sensitive genic male sterile (TGMS) line, is male sterile under cold conditions (12 °C with 12 h of daylight) but fertile under normal temperature (20 °C with 12 h of daylight). RESULTS: During meiosis, BS366 was defective in forming tetrads and dyads due to the abnormal cell plate. During pollen development, unusual vacuolated pollen that could not accumulate starch grains at the binucleate stage was also observed. Transcriptome analysis revealed that genes involved in the meiotic process, such as sister chromatid segregation and microtubule-based movement, were repressed, while genes involved in DNA and histone methylation were induced in BS366 under cold conditions. MethylRAD was used for reduced DNA methylation sequencing of BS366 spikes under both cold and control conditions. The differentially methylated sites (DMSs) located in the gene region were mainly involved in carbohydrate and fatty acid metabolism, lipid metabolism, and transport. Differentially expressed and methylated genes were mainly involved in cell division. CONCLUSIONS: These results indicated that the methylation of genes involved in carbon metabolism or fatty acid metabolism might contribute to male sterility in BS366 spikes, providing novel insight into the molecular mechanism of wheat male sterility.

Genome-wide identification and transcriptional characterization of DNA methyltransferases conferring temperature-sensitive male sterility in wheat.

BACKGROUND: DNA methyltransferase (DMT) genes contribute to plant stress responses and development by de novo establishment and subsequent maintenance of DNA methylation during replication. The photoperiod and/or temperature-sensitive genic male sterile (P/TGMS) lines play an important role in hybrid seed production of wheat. However, only a few studies have reported on the effect of DMT genes on temperature-sensitive male sterility of wheat. Although DMT genes have been investigated in some plant species, the identification and analysis of DMT genes in wheat (Triticum aestivum L.) based on genome-wide levels have not been reported. RESULTS: In this study, a detailed overview of phylogeny of 52 wheat DMT (TaDMT) genes was presented. Homoeolog retention for TaDMT genes was significantly above the average retention rate for whole-wheat genes, indicating the functional importance of many DMT homoeologs. We found that the strikingly high number of TaDMT genes resulted mainly from the significant expansion of the TaDRM subfamily. Intriguingly, all 5 paralogs belonged to the wheat DRM subfamily, and we speculated that tandem duplications might play a crucial role in the TaDRM subfamily expansion. Through the transcriptional analysis of TaDMT genes in a TGMS line BS366 and its hybrids with the other six fertile lines under sterile and fertile conditions, we concluded that TaCMT-D2, TaMET1-B1, and TaDRM-U6 might be involved in male sterility in BS366. Furthermore, a correlation analysis showed that TaMET1-B1 might negatively regulate the expression of TaRAFTIN1A, an important gene for pollen development, so we speculated regarding an epigenetic regulatory mechanism underlying the male sterility of BS366 via the interaction between TaMET1-B1 and TaRAFTIN1A. CONCLUSIONS: Our findings presented a detailed phylogenic overview of the DMT genes and could provide novel insights into the effects of DMT genes on TGMS wheat.

Transcriptome analysis of wheat seedling and spike tissues in the hybrid Jingmai 8 uncovered genes involved in heterosis.

MAIN CONCLUSION: Transcriptome analysis was carried out for wheat seedlings and spikes from hybrid Jingmai 8 and both inbred lines to unravel mechanisms underlying heterosis. Heterosis, known as one of the most successful strategies for increasing crop yield, has been widely exploited in plant breeding systems. Despite its great importance, the molecular mechanism underlying heterosis remains elusive. In the present study, RNA sequencing (RNA-seq) was performed on the seedling and spike tissues of the wheat (Triticum aestivum) hybrid Jingmai 8 (JM8) and its homozygous parents to unravel the underlying mechanisms of wheat heterosis. In total, 1686 and 2334 genes were identified as differentially expressed genes (DEGs) between the hybrid and the two inbred lines in seedling and spike tissues, respectively. Gene Ontology analysis revealed that DEGs from seedling tissues were significantly enriched in processes involved in photosynthesis and carbon fixation, and the majority of these DEGs expressed at a higher level in JM8 compared to both inbred lines. In addition, cell wall biogenesis and protein biosynthesis-related pathways were also significantly represented. These results confirmed that a combination of different pathways could contribute to heterosis. The DEGs between the hybrid and the two inbred progenitors from the spike tissues were significantly enriched in biological processes related to transcription, RNA biosynthesis and molecular function categories related to transcription factor activities. Furthermore, transcription factors such as NAC, ERF, and TIF-IIA were highly expressed in the hybrid JM8. These results may provide valuable insights into the molecular mechanisms underlying wheat heterosis.

Biogeography and community assembly of soil fungi from alpine meadows in southwestern China show the importance of climatic selection.

Soil fungi are pivotal in alpine and arctic ecosystems that are vulnerable to climate changes. Previous studies have shown broad connections between soil fungi in the arctic and alpine regions, but most of these studies are mainly from Europe and North America, with more sporadic studies from East Asia. Currently, little is known about the biogeographic relationships between soil fungi in alpine meadows of southwestern China (AMSC) and other regions of the world. In addition, the regional-scale spatial patterns of fungal communities in the AMSC, as well as their driving factors and ecological processes, are also poorly understood. In this study, we collected roots and surrounding soils of two dominant ectomycorrhizal plants, Bistorta vivipara and B. macrophylla from the AMSC, and performed bioinformatic and statistical analyses based on high-throughput sequencing of ITS2 amplicons. We found that: (1) fungi from the AMSC were closely related with those from boreal forests and tundra, and saprotrophic fungi had higher dispersal potential than ectomycorrhizal fungi; (2) community compositions exhibited clear divergences among geographic regions and between root and soil samples; (3) climate was the predominant factor driving regional-scale spatial patterns but had less explanatory power for saprotrophic and total fungi from roots than those from soils; (4) homogeneous selection and drift were the key ecological processes governing community assembly, but in communities of saprotrophic and total fungi from soil samples, drift contributed less and its role was partially replaced by dispersal limitation. This study highlights the importance of climatic selection and stochastic processes on fungal community assembly in alpine regions, and emphasizes the significance of simultaneously investigating fungi with different trophic modes and from both roots and soils.

Complete Genome Sequence of Aeromonas hydrophila Phage CC2.

Aeromonas hydrophila is one of the major pathogenic bacteria for fish and people. To develop an effective antimicrobial agent, we isolated a bacteriophage from sewage, named CC2, and sequenced its genome. Comparative genome analysis of phage CC2 with its relatives revealed that phage CC2 has higher sequence homology to A. salmonicida phage 65 than to A. hydrophila phage Aeh1. Here, we announce the complete genome sequence of CC2 and report major findings from the genomic analysis.

Complete genome sequence of an H3N2 canine influenza virus from dogs in Jiangsu, China.

A canine influenza virus (CIV) strain of avian origin designated A/Canine/Jiangsu/06/2010 (H3N2) was isolated from dogs exhibiting severe respiratory disease in Jiangsu, China. We announce the complete genome sequence of this viral strain and report major findings from the genomic analysis. This sequence will help us understand the molecular characteristics and evolutionary of H3N2 CIV in China.

A fungal insecticide engineered for fast per os killing of caterpillars has high field efficacy and safety in full-season control of cabbage insect pests.

Fungal insecticides developed from filamentous pathogens of insects are notorious for their slow killing action through cuticle penetration, depressing commercial interest and practical application. Genetic engineering may accelerate their killing action but cause ecological risk. Here we show that a Beauveria bassiana formulation, HV8 (BbHV8), engineered for fast per os killing of caterpillars by an insect midgut-acting toxin (Vip3Aa1) overexpressed in conidia has both high field efficacy and safety in full-season protection of cabbage from the damage of an insect pest complex dominated by Pieris rapae larvae, followed by Plutella xylostella larvae and aphids. In two fields repeatedly sprayed during summer, BbHV8 resulted in overall mean efficacies of killing of 71% and 75%, which were similar or close to the 70% and 83% efficacies achieved by commercially recommended emamectin benzoate but much higher than the 31% and 48% efficacies achieved by the same formulation of the parental wild-type strain (WT). Both BbHV8 and WT sprays exerted no adverse effect on a nontarget spider community during the trials, and the sprays did not influence saprophytic fungi in soil samples taken from the field plots during 4 months after the last spray. Strikingly, BbHV8 and the WT showed low fitness when they were released into the environment because both were decreasingly recovered from the field lacking native B. bassiana strains (undetectable 5 months after the spray), and the recovered isolates became much less tolerant to high temperature and UV-B irradiation. Our results highlight for the first time that a rationally engineered fungal insecticide can compete with a chemical counterpart to combat insect pests at an affordable cost and with low ecological risk.

Research review on methods of grassland health assessment.

Grassland health refers to the degree to which the integrity of soil and ecological processes is maintained, which primarily reflects the health status and productivity of grasslands. Evaluating the degree of grassland health is vital for the sustainable develop of grasslands. There are many methods for evaluating grassland health, with advantages and disadvantages for each one. However, there is still a lack of systematic literature offering an overview of methods of grassland health assessment and their applicability. We summarized 10 methods of grassland health assessment, including vigor-organization-resilience (VOR) index evaluation model, condition-vigor-organization-resilience (CVOR) index evaluation model, principal component analysis method, analytic hierarchy process, cluster analysis method, grey relational analysis, pressure-state-response evaluation model, fuzzy comprehensive evaluation method, comprehensive evaluation model of grassland health, and evaluation model using remote sensing technology. The advantages, disadvantages, and applicability of these methods were discussed, aiming to provide scientific basis for selecting more suitable methods of grassland health assessment for different scenarios in the future.

Bemisia tabaci (Hemiptera: Aleyrodidae) Insecticide Resistance in Shandong Province, China.

The Bemisia tabaci (Gennadius) (Hemiptera: Aleyrodidae) complex comprises important pests and virus vectors in agricultural crops worldwide. In China, B. tabaci has spread to more than 20 provinces and caused severe losses of vegetables, fruits, and ornamental plants. However, B. tabaci has developed resistance to many insecticidal classes in Shandong Province, eastern China. In this study, we investigated the cryptic species, insecticide resistance and detoxifying enzymes of B. tabaci from six representative locations exhibiting severe damage in Shandong. At four of the six locations, B. tabaci Mediterranean (MED) comprised 100% of the samples collected. In a further two locations, species composition was predominantly (>94%) MED with B. tabaci Middle East-Asia Minor 1 (MEAM1), comprising a low proportion (<6%) of the samples collected. For all field populations, avermectin was the most effective insecticide against adult B. tabaci, pyriproxyfen had a significant effect on B. tabaci eggs and field populations were susceptible to pymetrozine. Six field populations of B. tabaci have developed low-to-moderate resistance to neonicotinoids. The detoxifying enzyme activity of carboxylesterase, glutathione S-transferase, and multifunctional oxidase were quantified. Multifunctional oxidase and glutathione S-transferase activity were positively correlated with insecticide resistance in several B. tabaci populations.

Organocatalytic Asymmetric Decarboxylative Mannich Reaction of ß-Keto Acids with Cyclic α-Ketiminophosphonates: Access to Quaternary α-Aminophosphonates.

An organocatalytic asymmetric decarboxylative Mannich reaction of ß-keto acids with cyclic α-ketiminophosphonates has been developed. By using saccharide-derived bifunctional amine-thiourea catalysts bearing an axial chiral binaphthyl scaffold, a wide range of quaternary α-amino-γ-oxophosphonates were obtained in up to 93% yield and >99% ee. Furthermore, two interesting α-aminophosphonate derivatives were synthesized from the corresponding decarboxylative Mannich product via simple transformations.

Chiral phosphoric acid-catalyzed direct asymmetric mannich reaction of cyclic C-acylimines with simple ketones: facile access to C2-quaternary indolin-3-ones.

A chiral Brønsted acid-catalyzed direct asymmetric Mannich reaction of simple ketones with cyclic C-acylimines has been established for the synthesis of C2-quaternary indolin-3-ones. In the presence of 5-10 mol% chiral phosphoric acid, a series of 2-(2-oxo-2-phenylethyl)-2-arylindolin-3-ones were obtained in good to high yield with up to 99% ee. The adducts obtained could be readily converted into indolines, tricyclic indolin-3-ones, and tetracyclic tetrahydro-indolo[1,2-a]quinolines by simple modifications.

Stress Ratio and Notch Effects on the Very High Cycle Fatigue Properties of a Near-Alpha Titanium Alloy.

Ultrasonic fatigue tests up to 1010 cycles were performed on a turbine engine titanium alloy (Ti-8Al-1Mo-1V) at the stress ratio (R) of -1 with smooth specimens and at R = -1, 0.1 and 0.5 with notched specimens. As a result, with increase of fatigue life, the source of reduced fatigue life caused by multi-point surface crack initiation changes from crack propagation stage to crack initiation stage in the high cycle fatigue regime. Notch effect further promotes the degeneration of high cycle and very high cycle fatigue strength at R > -1. The bilinear model, extended from the Goodman method, can better estimate the mean stress sensitivity of this titanium alloy. The fatigue mean stress sensitivity and fatigue-creep mean stress sensitivity of this material increased with the increase of fatigue life. The new model, based on the Murakami model, can provide more appropriate predictions for notch fatigue strength.

Generation of linear expression constructs by one-step PCR with vaccinia DNA topoisomerase I.

Linear expression constructs can facilitate gene function studies. We describe a method to generate linear expression constructs for mammalian cells by one-step polymerase chain reaction (PCR) with vaccinia DNA topoisomerase I (TOPO). Cytomegalovirus (CMV) 5\' promoter, the gene of interest, and V5 bovine growth hormone (BGH) polyA 3\' terminator elements were PCR-amplified with target-specific primers containing vaccinia DNA TOPO-specific sequence and complementary sequence to each other. We amplified specific and complementary sequences. These three elements were directionally joined with vaccinia TOPO. The joined products were then directly transfected into Chinese hamster ovary cells. Compared with the transfection of supercoiled plasmids, comparable expression signals were obtained for green fluorescent protein, chloramphenicol acetyltransferase, and beta-galactosidase proteins using Western blots. This is a quick and efficient method to generate linear expression constructs. Unlike Invitrogen TOPO Tools, our method avoided the secondary round of PCR and more rapidly yielded correct joining products. This method can be easily used in the function test of uncharacterized open reading frames.

Catalytic Asymmetric Mukaiyama-Mannich Reaction of Cyclic C-Acylimines with Difluoroenoxysilanes: Access to Difluoroalkylated Indolin-3-ones.

A catalytic enantioselective Mukaiyama-Mannich reaction of cyclic C-acylimines with difluoroenoxysilanes is reported. (S)-TRIP enables the enantioselective synthesis of a series of novel difluoroalkylated indolin-3-ones bearing a quaternary stereocenter in up to 97% yield and 98% ee. The synthetic utility of this protocol is highlighted by efficient conversion of the products to the corresponding indolin-3-one derivatives without any erosion of the enantiopurity.

Nematic superconducting state in iron pnictide superconductors.

Nematic order often breaks the tetragonal symmetry of iron-based superconductors. It arises from regular structural transition or electronic instability in the normal phase. Here, we report the observation of a nematic superconducting state, by measuring the angular dependence of the in-plane and out-of-plane magnetoresistivity of Ba0.5K0.5Fe2As2 single crystals. We find large twofold oscillations in the vicinity of the superconducting transition, when the direction of applied magnetic field is rotated within the basal plane. To avoid the influences from sample geometry or current flow direction, the sample was designed as Corbino-shape for in-plane and mesa-shape for out-of-plane measurements. Theoretical analysis shows that the nematic superconductivity arises from the weak mixture of the quasi-degenerate s-wave and d-wave components of the superconducting condensate, most probably induced by a weak anisotropy of stresses inherent to single crystals.

Fatigue Property of Open-Hole Steel Plates Influenced by Bolted Clamp-up and Hole Fabrication Methods.

Steel plates with open holes are commonly used in structural assemblies. The fatigue properties of such details are influenced by bolted clamp-up and hole fabrication methods. The fracture surface, stiffness degradation and fatigue life of test specimens are investigated in detail and compared with the contemporary test data. The analysis results show that the presence of draglines greatly influences the fatigue crack initiation at the open-hole cut by laser. The bolted clamp-up condition greatly enhances the stiffness and the fatigue life of the open-hole details. A discussion is also made from a comparison with the referred fatigue life of hole fabrication details, such as the influence of plate thickness and plasma cutting, drilling and oxy-fuel gas cutting, with the details studied herein. This work could enhance the understanding of the fatigue property and design of such details.

[Resistance risk, cross-resistance and biochemical resistance mechanism of Laodelphax striatellus to buprofezin].

In order to investigate the resistance development law and biochemical resistance mechanism of Laodelphax striatellus to buprofezin, spraying rice seedlings was used to continuously screen resistant strains of L. striatellus and dipping rice seedlings was applied to determine the toxicity and cross-resistance of L. striatellus to insecticides. After 32-generation screening with buprofezin, L. striatellus developed 168.49 folds resistance and its reality heritability (h2) was 0.11. If the killing rate was 80%-90%, L. striatellus was expected to develop 10-fold resistance to buprofezin only after 5 to 6 generations breeding. Because the actual reality heritability of field populations was usually lower than that of the resistant strains, the production of field populations increasing with 10-fold resistance would need much longer time. The results of cross-resistance showed that resistant strain had high level cross-resistance with thiamethoxam and imidacloprid, low level cross-resistance with acetamiprid, and no cross-resistance with pymetrozine and chlorpyrifos. The activity of detoxification enzymes of different strains and the syergism of synergist were measured. The results showed that cytochrome P450 monooxygenase played a major role in the resistance of L. striatellus to buprofezin, the esterase played a minor role and the GSH-S-transferase had no effect. Therefore, L. striatellus would have high risk to develop resistance to buprofezin when used in the field and might be delayed by using pymetrozine and chlorpyrifos.

Classification of a Hypervirulent Aeromonas hydrophila Pathotype Responsible for Epidemic Outbreaks in Warm-Water Fishes.

Lineages of hypervirulent Aeromonas hydrophila (vAh) are the cause of persistent outbreaks of motile Aeromonas septicemia in warm-water fishes worldwide. Over the last decade, this virulent lineage of A. hydrophila has resulted in annual losses of millions of tons of farmed carp and catfish in the People's Republic of China and the United States (US). Multiple lines of evidence indicate US catfish and Asian carp isolates of A. hydrophila affiliated with sequence type 251 (ST251) share a recent common ancestor. To address the genomic context for the putative intercontinental transfer and subsequent geographic spread of this pathogen, we conducted a core genome phylogenetic analysis on 61 Aeromonas spp. genomes, of which 40 were affiliated with A. hydrophila, with 26 identified as epidemic strains. Phylogenetic analyses indicate all ST251 strains form a coherent lineage affiliated with A. hydrophila. Within this lineage, conserved genetic loci unique to A. hydrophila were identified, with some genes present in consistently higher copy numbers than in non-epidemic A. hydrophila isolates. In addition, results from analyses of representative ST251 isolates support the conclusion that multiple lineages are present within US vAh isolated from Mississippi, whereas vAh isolated from Alabama appear clonal. This is the first report of genomic heterogeneity within US vAh isolates, with some Mississippi isolates showing closer affiliation with the Asian grass carp isolate ZC1 than other vAh isolated in the US. To evaluate the biological significance of the identified heterogeneity, comparative disease challenges were conducted with representatives of different vAh genotypes. These studies revealed that isolate ZC1 yielded significantly lower mortality in channel catfish, relative to Alabama and Mississippi vAh isolates. Like other Asian vAh isolates, the ZC1 lineage contains all core genes for a complete type VI secretion system (T6SS). In contrast, more virulent US isolates retain only remnants of the T6SS (clpB, hcp, vgrG, and vasH) which may have functional implications. Collectively, these results characterize a hypervirulent A. hydrophila pathotype that affects farmed fish on multiple continents.

[Construction and characterization of some virulence determinants-deficient strain in Aeromonas hydrophila].

Cell conjugation was carried out between the donor E. coli CG120 with pAM120 (Tc(r)/Tn916) and the recipient Aeromonas hydrophila J-1 by filter mating. 3800 positive clones were gained according to the ability of growth on LB medium with 10 g/mL tetracycline (Tc) and 100 g/mL cephazolin (cfz). Conjugation efficiency was 3 x 10(-5) per donor. When 16S rDNA PCR amplification was performed with special primers, positive results were gained in all the 38 conjugants. To demonstrate the insertion of Tn916 into the genomes of conjugants, tetracycline gene (tet) PCR amplification was performed. A special band could be gained in the conjugants with Tc(r). Compared with the parent J-1 strain, some genes of main virulence determinants, such as proteases, hemolysins, DNase and amylases, could not be expressed in the conjugants. The pathogenicity capability of conjugants was greatly decreased and the 50% lethal dose for mice was more than 10(9) CFU. By serially passaged for 10 times, the above virulent characters had not been regained despite the disappearance of Tc(r). The results showed that by the insertion of Tn916, stable and avirulent A. hydrophila mutants could be gained. The mechanisms, by which Tn916 induced the changes of virulence characters in A. hydrophila, are not definite. It is suspected that there might exist hot spots of Tn916 or pathogenicity island on the chronosome of A. hydrophila.

Evaluation of Fatigue Strength Improvement by CFRP Laminates and Shot Peening onto the Tension Flanges Joining Corrugated SteelWebs.

Corrugated steel web with inherent high out-of-plane stiffness has a promising application in configuring large span highway bridge girders. Due to the irregularity of the configuration details, the local stress concentration poses a major fatigue problem for the welded flange plates of high strength low alloy structural steels. In this work, the methods of applying CFRP laminate and shot peening onto the surfaces of the tension flanges were employed with the purpose of improving the fatigue strength of such configuration details. The effectiveness of this method in the improvement of fatigue strength has been examined experimentally. Test results show that the shot peening significantly increases hardness and roughness in contrast to these without treatment. Also, it has beneficial effects on the fatigue strength enhancement when compared against the test data of the joints with CFRP strengthening. The stiffness degradation during the loading progress is compared with each treatment. Incorporating the stress acting on the constituent parts of the CFRP laminates, a discussion is made regarding the mechanism of the retrofit and related influencing factors such as corrosion and economic cost. This work could enhance the understanding of the CFRP and shot peening in repairing such welded details and shed light on the reinforcement design of welded joints between corrugated steel webs and flange plates.