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Gomphus matijun, a new edible species, is described from southwestern China based on phylogenetic and morphological evidence. Phylogenetic analyses of the nrLSU and ITS datasets indicated that G. matijun is related to G. crassipes and G. ludovicianus with weak statistical support in maximum likelihood but strong statistical support in Bayesian analyses. Gomphus matijun is characterized by its gray-blue to blue or blue-purple pileus with a round or irregular outline, relatively smaller basidiospores [9-11(-13) × 6-7(-8) µm], mostly 2-spored basidia, and occurs in subtropical fagaceous forests. A key to the known species of Gomphus is provided. [ABSTRACT FROM AUTHOR]

The systematic position of the enigmatically mycoparasitic genus Squamanita (Agaricales, Basidiomycota) together with Cystoderma, Phaeolepiota, Floccularia, and Leucopholiota is largely unknown. Recently they were recognized as Squamanitaceae, but previous studies used few DNA markers from a restricted sample of taxa from the family and lacked a formal taxonomic treatment. In this study, with newly generated sequences of the type of the genus Squamanita, S. schreieri, and several additional species of the family, the phylogeny is reinvestigated with a concatenated (18S-5.8S-nrLSU-RPB2-TEF1-α) dataset. This study reveals that Cystoderma, Phaeolepiota, Squamanita, Floccularia, and Leucopholiota are a monophyletic clade with strong statistical support in Bayesian analysis and form Squamanitaceae. Phaeolepiota nested within Cystoderma; Squamanita, Leucopholiota, and Floccularia clustered together as two monophyletic subclades; and Squamanita was present as a monophyletic clade with strong statistical support in both Maximum Likelihood and Bayesian analyses. The family name Squamanitaceae is formally emended and a detailed taxonomic treatment is presented to accommodate the five genera. Meanwhile, another concatenated (18S-ITS-nrLSU-RPB2-TEF1-α) dataset is used to investigate phylogenetic relationships and species delimitation in Squamanita. Our data indicates that "S. umbonata" from the Northern hemisphere forms two species complexes, one complex includes six specimens from North America, Europe, and East Asia, the other includes two specimens from Central America and North America respectively. Futhermore, species of Squamanita can parasitize species of Amanita, besides other fungal species. Squamanita mira parasitizes A. kitamagotake (A. sect. Caesareae), while S. orientalis and S. sororcula are parasites of species belonging to the A. sepiacea complex (A. sect. Validae). "Squamanita umbonata" from Italy occurs on A. excelsa (A. sect. Validae). Three new species of Squamanita from East Asia, viz. S. mira, S. orientalis and S. sororcula are documented with morphological, multi-gene phylogenetic, and ecological data, along with line drawings and photographs, and compared with similar species. A key for identification of the global Squamanita species is provided. [ABSTRACT FROM AUTHOR]

In nature, foliar variegation has varied origins, and can be ascribed to two major mechanisms: pigment-related variegation and structural variegation caused by the optical properties of leaf structure. However, understanding of these mechanisms is still lacking, and structural mechanisms are often misinterpreted. In this study, six variegated plants native to Taiwan and two ornamental plants, with novel and unusual foliar variegation patterns, are studied to reveal their mechanisms of variegation. Two newly understood variegation patterns, the silvery white leaf surface of Begonia aptera and the varnish on basal leaflets of Oxalis corymbosa, are reported here. White to light green patches on leaf surfaces characterize the foliar variegation in the other six study species, Nervilia nipponica, O. acetosella subsp. griffithii var. formosana, Paphiopedilum concolor, Selaginella picta, Smilax bracteata subsp. verruculosa and Valeriana hsuii. All six Taiwan native plants exhibit structural variegation, five of which have air space type variegation. Surprisingly the silvery white leaf surface of B. aptera results from numerous sand-like white spots caused by intercellular space. The varnish on leaves of O. corymbosa is the epidermis type of variegation comprised of two newly identified subtypes, larger epidermal cells and thicker outer cell walls. The two ornamental plants have variegation caused by pigments: the chloroplast type from fewer chloroplasts in P. concolor and the chlorophyll type, from absence of chloroplasts in S. picta. This study extends understanding of the mechanisms of natural foliar variegation, illustrating the diversity of mechanisms by which plants may change the appearance of their leaves. [ABSTRACT FROM AUTHOR]

In the accompanying paper, we described evolving a lipase to the point where variants were soluble, stable and capable of degrading C8 TAG and C8 esters. These variants were tested for their ability to survive in an environment that might be encountered in a washing machine. Unfortunately, they were inactivated both by treatment with a protease used in laundry detergents and by very low concentrations of sodium dodecyl sulfate (SDS). In addition, all the variants had very low levels of activity with triglycerides with long aliphatic chains and with naturally occurring oils, like olive oil. Directed evolution was used to select variants with enhanced properties. In the first 10 rounds of evolution, the primary screen was selected for variants capable of hydrolyzing olive oil whereas the secondary screen was selected for enhanced tolerance towards a protease and SDS. In the final six rounds of evolution, the primary and secondary screens identified variants that retained activity after treatment with SDS. Sixteen cycles of evolution gave variants with greatly enhanced lipolytic activity on substrates that had both long (C16 and C18) as well as short (C3 and C8) chains. We found variants that were stable for more than 3 hours in protease concentrations that rapidly degrade the wild-type enzyme. Enhanced tolerance towards SDS was found in variants that could break down naturally occurring lipid and resist protease attack. The amino acid changes that gave enhanced properties were concentrated in the cap domain responsible for substrate binding. [ABSTRACT FROM AUTHOR]

We identified Candidatus Borrelia fainii, a human pathogenic bacterium causing New World relapsing fever in a Myotis bat in eastern China. This finding expands knowledge about the geographic distribution of Borrelia spp. and the potential for infection with New World relapsing fever in China. [ABSTRACT FROM AUTHOR]

Cyanuric acid is a metabolic intermediate of s-triazines, such as atrazine (a common herbicide) and melamine (used in resins and plastics). Cyanuric acid is mineralized to ammonia and carbon dioxide by the soil bacterium Pseudomonas sp. strain ADP via three hydrolytic enzymes (AtzD, AtzE, and AtzF). Here, we report the purification and biochemical and structural characterization of AtzE. Contrary to previous reports, we found that AtzE is not a biuret amidohydrolase, but instead it catalyzes the hydrolytic deamination of 1-carboxybiuret. X-ray crystal structures of apo AtzE and AtzE bound with the suicide inhibitor phenyl phosphorodiamidate revealed that the AtzE enzyme complex consists of two independent molecules in the asymmetric unit. We also show that AtzE forms an α2β2 heterotetramer with a previously unidentified 68-amino acid-long protein (AtzG) encoded in the cyanuric acid mineralization operon from Pseudomonas sp. strain ADP. Moreover, we observed that AtzG is essential for the production of soluble, active AtzE and that this obligate interaction is a vestige of their shared evolutionary origin. We propose that AtzEG was likely recruited into the cyanuric acid-mineralizing pathway from an ancestral glutamine transamidosome that required protein-protein interactions to enforce the exclusion of solvent from the transamidation reaction. [ABSTRACT FROM AUTHOR]

Purpose: To investigate the anti-inflammatory effects of nobiletin on human osteoarthritic chondrocytes and also to explore possible related molecular events. Methods: Isolated human osteoarthritic chondrocytes were stimulated with IL-1β. The effect of nobiletin (75, 150 or 300 µg) on chondrocyte viability was assessed. Furthermore, the effect on NO production was determined using Griess reagent while the levels of IL-6 and PGE2 were assessed by enzyme linked immunosorbent assay (ELISA). The influence of nobiletin on the expression of COX-2, iNOS and proteins of PI3/Akt, NF-κB and MAPK cascades were also assessed. Results: Nobiletin (75, 150 and 300 μg) significantly (p < 0.05) improved the viability of chondrocytes, and remarkably reduced the levels of NO, IL-6 and PGE2. The expression levels of COX-2 and iNOS both at mRNA and protein levels were strikingly reduced by nobiletin, in a dose-dependent way. In addition, nobiletin caused marked (p < 0.05) down-regulation of the NF-κB signalling pathway. IL-1β- induced activation of PI3/Akt, and JNK, ERK and p38 MAPK cascades were significantly (p < 0.05) inhibited by nobiletin with 300 µg dose exhibiting maximum effects. Conclusion: Inflammatory cytokines are critically involved in the pathogenesis of OA. Significant suppression of cytokines and modulation of PI3/Akt and MAPK signalling cascades by nobiletin suggests its potent anti-inflammatory and anti-osteoarthritic effects. [ABSTRACT FROM AUTHOR]

Insect carboxylesterases from the αEsterase gene cluster, such as αE7 (also known as E3) from the Australian sheep blowfly Lucilia cuprina (LcαE7), play an important physiological role in lipid metabolism and are implicated in the detoxification of organophosphate (OP) insecticides. Despite the importance of OPs to agriculture and the spread of insect-borne diseases, the molecular basis for the ability of α-carboxylesterases to confer OP resistance to insects is poorly understood. In this work, we used laboratory evolution to increase the thermal stability of LcαE7, allowing its overexpression in Escherichia coil and structure determination. The crystal structure reveals a canonical α/β-hydrolase fold that is very similar to the primary target of OPs (acetylcholinesterase) and a unique N-terminal α-helix that serves as a membrane anchor. Soaking of LcαE7 crystals in OPs led to the capture of a crystallographic snapshot of LcαE7 in its phosphorylated state, which allowed comparison with acetylcholinesterase and rationalization of its ability to protect insects against the effects of OPs. Finally, inspection of the active site of LcαE7 reveals an asymmetric and hydrophobic substrate binding cavity that is well-suited to fatty acid methyl esters, which are hydrolyzed by the enzyme with specificity constants (∼106 M-1 S-1) indicative of a natural substrate. [ABSTRACT FROM AUTHOR]

There have been many approaches to solving problems associated with protein solubility. This article describes the application of directed evolution to improving the solubility of the C-terminal metal-binding domain of aminopeptidase P from Escherichia coli. During the course of experiments, the domain boundary and sequence were allowed to vary. It was found that extending the domain boundary resulted in aggregation with little improvement in solubility, whereas two changes to the sequence of the domain resulted in dramatic improvements in solubility. These latter changes occurred in the active site and abolished the ability of the protein to bind metals and hence catalyze its physiological reaction. The evidence presented here has led to the proposal that metals bind to the intact protein after it has folded and that the N-terminal domain is necessary to stabilize the structure of the protein so that it is capable of binding metals. The acid residues responsible for binding metals tend to repel one another − in the absence of the N-terminal domain, the C-terminal domain does not fold properly and forms inclusion bodies. Evolution of the C-terminal domain has removed the destabilizing effects of the metal ligands, but in so doing it has reduced the capacity of the domain to bind metals. In this case, directed evolution has identified active site residues that destabilize the domain structure. [ABSTRACT FROM AUTHOR]

Directed evolution is a useful tool in the study of enzymes. It is used in this study to investigate the means by which resistance to the antibiotic trimethoprim develops in dihyrofolate reductase from Escherichia coli. Mutants with clinical levels of resistance were obtained after only three generations. After four generations of directed evolution, several mutants were characterized, along with some point mutants made to investigate amino acid changes of interest. Several mutations were found to grant resistance to trimethoprim, both by reducing the binding affinity of the enzyme for the drug, and by increasing the activity of the enzyme. [ABSTRACT FROM AUTHOR]

The intracellular pathogenesis-related (PR) proteins of common bean (Phaseolus vulgaris L.) are encoded by a highly polymorphic family of at least 20 genes. One member, the Ypr1O*c gene, has been isolated and characterised. The deduced amino acid sequence of the encoded protein, PR-10, exhibits similarities to tree-pollen allergens, to food allergens from celery and apple and to ginseng ribonuclease peptide sequences. We show by RNA blot analysis that the Ypr10 gene family, including Ypr10*c, is strongly expressed in bean roots. In leaves Ypr10 transcript levels are low in young and mature stages but are elevated during senescence and in diseased states. Dark treatment of leaves causes strong induction of Ypr10 transcripts, which is reversible by light, and diurnal rhythms of transcript accumulation during the night are observed. Ypr10 genes are responsive to external stimuli related to pathogen-defence such as glutathione or salicylic acid. Transcriptional activity of a Ypr10*c promoter-β-glucuronidase fusion gene in transgenic tobacco was observed in roots, in developing xylem and phloem of stems, and in the blade of senescent leaves, with highest levels at the onset of senescence. The most striking characteristic of developmental expression was the specific localisation of β-glucuronidase activity in the transmitting tract of styles in flowers at anthesis. Feeding of various pathogen-related and stress-related stimuli to young tobacco leaves led to accumulation of GUS activity in leaf blades. We identify considerable spatiotemporal similarities between reported expression patterns of Ypr10 genes and ribonuclease genes, which, together with the significant sequence similarity to the ginseng ribonuclease, support the hypothesis of a ribonuclease function for PR-10 proteins and allow the prediction of possible biological roles. [ABSTRACT FROM AUTHOR]

The article offers the author's view on the succesful strategies for the nanowire assemblies ordering in China.

PCR amplification of the rrs2 gene indicated that 50% (62/124) of insectivorous bats from eastern China were infected with Leptospira borgpetersenii, L. kirschneri, and several potentially new Leptospira species. Multilocus sequence typing defined 3 novel sequence types in L. kirschneri, suggesting that bats are major carriers of Leptospira. [ABSTRACT FROM AUTHOR]

We analyzed the effects of a traditional Chinese medicine, Qizhi Jiangtang Jiaonang (QJJ), on insulin resistance (IR) in vitro. After an in vitro model of IR was established by treating human liver cancer cells (HepG2 cells) with palmitic acid, the cells were then treated with various concentrations of QJJ. Treatment with 400 µM palmitic acid for 24 h induced IR in HepG2 cells. The survival rate for HepG2 cells in the IR group was significantly lower than that of the untreated control group (P < 0.001); however, QJJ restored HepG2 cell survival (P < 0.001). As compared with HepG2 cells in the IR group, QJJ at all doses analyzed significantly increased glucose consumption (all P < 0.05). Moreover, treatment with all the QJJ doses significantly reduced the mean intracellular reactive oxygen species levels as compared with the IR group (all P < 0.05). Furthermore, high-dose QJJ reduced both TNF-α and IL-6 levels as compared to the IR group (all P < 0.05). QJJ ameliorated the altered PI3K, GLUT4, and RAGE expression observed with IR. In conclusion, QJJ can improve IR in HepG2 cells, which may be mediated through the IRS-1/PI3K/GLUT4 signaling pathway as well as regulation of NF-κB-mediated inflammation and oxidative stress. [ABSTRACT FROM AUTHOR]

Phosphotriesterases catalyze the hydrolytic detoxification of phosphotriester pesticides and chemical warfare nerve agents with various efficiencies. The directed evolution of phosphotriesterases to enhance the breakdown of poor substrates is desirable for the purposes of bioremediation. A limiting factor in the identification of phosphotriesterase mutants with increased activity is the ability to effectively screen large mutant libraries. To this end, we have investigated the possibility of coupling phosphotriesterase activity to cell growth by using methyl paraoxon as the sole phosphorus source. The catabolism of paraoxon to phosphate would occur via the stepwise enzymatic hydrolysis of paraoxon to dimethyl phosphate, methyl phosphate, and then phosphate. The Escherichia coli strain DH10B expressing the phosphotriesterase from Agrobacterium radiobacter P230 (OpdA) is unable to grow when paraoxon is used as the sole phosphorus source. Enterobacter aerogenes is an organism capable of growing when dimethyl phosphate is the sole phosphorus source. The enzyme responsible for hydrolyzing dimethyl phosphate has been previously characterized as a nonspecific phosphohydrolase. We isolated and characterized the genes encoding the phosphohydrolase operon. The operon was identified from a shotgun clone that enabled E. coli to grow when dimethyl phosphate is the sole phosphorus source. E. coli coexpressing the phosphohydrolase and OpdA grew when paraoxon was the sole phosphorus source. By constructing a short degradative pathway, we have enabled E. coli to use phosphotriesters as a sole source of phosphorus. [ABSTRACT FROM AUTHOR]

Burkholderia sp. strain SG-MS1 and Pseudomonas sp. strain SG-MS2 have previously been found to mineralize (+)-pinoresinol through a common catabolic pathway. Here, we used comparative genomics, proteomics, protein semipurification, and heterologous expression to identify a flavoprotein from the vanillyl alcohol oxidase/pcresol methyl hydroxylase (VAO/PCMH) enzyme family in SG-MS2 that carries out the initial hydroxylation of (+)-pinoresinol at the benzylic carbon. The cognate gene is translationally coupled with a downstream cytochrome gene, and the cytochrome is required for activity. The flavoprotein has a unique combination of cofactor binding and cytochrome requirements for the VAO/PCMH family. The heterologously expressed enzyme has a Km of 1.17µM for (+)-pinoresinol. The enzyme is overexpressed in strain SG-MS2 upon exposure to (+)-pinoresinol, along with 45 other proteins, 22 of which were found to be encoded by genes in an approximately 35.1-kb cluster also containing the flavoprotein and cytochrome genes. Homologs of 18 of these 22 genes, plus the flavoprotein and cytochrome genes, were also found in a 38.7-kb cluster in SG-MS1. The amino acid identities of four of the other proteins within the SG-MS2 cluster suggest they catalyze conversion of hydroxylated pinoresinol to protocatechuate and 2-methoxyhydroquinone. Nine other proteins upregulated in SG-MS2 on exposure to (+)-pinoresinol appear to be homologs of proteins known to comprise the protocatechuate and 2-methoxyhy droquinone catabolic pathways, but only three of the cognate genes lie within the cluster containing the flavoprotein and cytochrome genes. IMPORTANCE (+)-Pinoresinol is an important plant defense compound, a major food lignan for humans and some other animals, and the model compound used to study degradation of the γ-γ' linkages in lignin. We report a gene cluster, in one strain each of Pseudomonas and Burkholderia, that is involved in the oxidative catabolism of (+)-pinoresinol. The flavoprotein component of the α-hydroxylase which heads the pathway belongs to the 4-phenol oxidizing (4PO) subgroup of the vanillyl alcohol oxidase/p-cresol methyl hydroxylase (VAO/PCMH) enzyme family but constitutes a novel combination of cofactor and electron acceptor properties for the family. It is translationally coupled with a cytochrome gene whose product is also required for activity. The work casts new light on the biology of (+)-pinoresinol and its transformation to other bioactive molecules. Potential applications of the findings include new options for deconstructing lignin into useful chemicals and the generation of new phytoestrogenic enterolactones from lignans. [ABSTRACT FROM AUTHOR]

PCR amplification indicated the minimum infection rate of Rickettsia spp. was 0.66% in Haemaphysalis longicornis ticks collected from Shandong Province, China. Phylogenetic analysis based on the rrs, gltA, ompA, and ompB genes indicated that the ticks carried R. japonica, Candidatus Rickettsia longicornii, and a novel Rickettsia species related to R. canadensis. [ABSTRACT FROM AUTHOR]

Graphene-based composites have gained great attention in the field of gas sensor fabrication due to their higher surface area with additional functional groups. Decorating one-dimensional (1D) semiconductor nanomaterials on graphene also show potential benefits in gas sensing applications. Here we demonstrate the one-pot and low cost synthesis of W18O49 NWs/rGO composites with different amount of reduced graphene oxide (rGO) which show excellent gas-sensing properties towards toluene and strong dependence on their chemical composition. As compared to pureW18O49 NWs, an improved gas sensing response (2.8 times higher) was achieved in case of W18O49 NWs composite with 0.5 wt. % rGO. Promisingly, this strategy can be extended to prepare other nanowire based composites with excellent gas-sensing performance. [ABSTRACT FROM AUTHOR]

WEE1 is a serine/threonine protein kinase that inactivates cell division cycle 2 and is therefore a critical cell cycle regulator. Increased WEE1 expression has been observed in numerous types of human malignancies, including hepatocellular carcinoma and melanoma. WEE1 inhibition also results in evident anti-tumor effects in several human tumor cells including colon cancer cells, suggesting WEE1 as a potential therapeutic target for the treatment of cancer. However, the expression pattern of WEE1 in colorectal cancer (CRC) remains unclear. In the present study, WEE1 mRNA expression in 43 cases of CRC tissues matched with adjacent normal tissues was determined by reverse-transcription quantitative polymerase chain reaction. The results demonstrated that WEE1 mRNA expression was significantly increased in CRC tissues and that this upregulation correlated significantly with hepatic metastasis, distant metastasis and high tumor node metastasis (TNM) stage of CRC. Additionally, WEE1 protein in 102 CRC tissue samples was detected by immunohistochemistry, and positive staining of WEE1 was identified in more than half of patients with CRC. WEE1 staining scores were also observed to be associated with distant metastasis and high TNM stage of CRC. In addition, patients with CRC with high WEE1 staining score (2+ or 3+) exhibited either poorer overall survival or poorer disease-free survival compared with those with low WEE1 staining score (0 or 1+). The multivariable Cox model also identified a high WEE1 staining score as well as high TNM stage to be independent prognostic factors for CRC. In conclusion, WEE1 upregulation is associated with a high degree of malignancy and poor prognosis of CRC, suggesting WEE1 as a potential prognostic biomarker for CRC. [ABSTRACT FROM AUTHOR]

Severe fever with thrombocytopenia syndrome (SFTS) is an emerging hemorrhagic fever in East Asia caused by SFTS virus (SFTSV), a newly discovered phlebovirus. The Haemaphysalis longicornis tick has been suspected to be the vector of SFTSV. To determine whether SFTSV can be transmitted among ticks, from ticks to animals, and from animals to ticks, we conducted transmission studies between developmental stages of H. longicornis ticks and between ticks and mice. Using reverse transcription PCR, we also analyzed the prevalence of SFTSV infection among H. longicornis ticks collected from vegetation in Shandong Province, China. Our results showed a low prevalence of SFTSV among collected ticks (0.2%, 8/3,300 ticks), and we showed that ticks fed on SFTSV-infected mice could acquire the virus and transstadially and transovarially transmit it to other developmental stages of ticks. Furthermore, SFTSV-infected ticks could transmit the virus to mice during feeding. Our findings indicate ticks could serve as a vector and reservoir of SFTSV. [ABSTRACT FROM AUTHOR]

The metallo-glycerophosphodiesterase from Enterobacter aerogenes (GpdQ) has been cloned, expressed in Escherichia coli and purified. Initial screening of crystallization conditions for this enzyme resulted in the identification of needles from one condition in a sodium malonate grid screen. Removal of the metals from the enzyme and subsequent optimization of these conditions led to crystals that diffracted to 2.9 Å and belonged to space group P213, with unit-cell parameter a = 164.1 Å. Self-rotation function analysis and VM calculations indicated that the asymmetric unit contains two copies of the monomeric enzyme, corresponding to a solvent content of 79%. It is intended to determine the structure of this protein utilizing SAD phasing from transition metals or molecular replacement. [ABSTRACT FROM AUTHOR]

The article presents a study on hemorrhagic fever with renal syndrome (HFRS) in Jiaonan County in Shandong Province, China from December 2012 to November 2013. It is caused by hantavirus from serotype-specific reservoir hosts like rodents and shrews with two serotypes, such as Hantaan virus (HTNV) and Seoul virus (SEOV). The mammals were amplified for RNA lung samples to determine the types of hantavirus using reverse transcription polymerase chain reaction (PCR) and serotype-specific primers.