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Bletilla striata, a perennial herbaceous plant belonging to the family Orchidaceae, is native to China and is widely distributed in the Yangtze River basin. In China, B. striata is a popular medicinal plant that is typically used to reduce wound bleeding and inflammation. In September 2021, distinct leaf spot symptoms were observed in more than 50% of B. striata plants in a traditional Chinese medicine plantation (ca. 10 ha) in Xianju City, Zhejiang Province, China. Small, round, pale brown, necrotic spots were first observed on the leaves. Subsequently, these lesions became grayish brown in the center and dark brown with slight protuberances at the margins and eventually enlarged to 5–8 mm on the leaves. Over time, the small spots enlarged and coalesced into necrotic streaks (1–2 cm). Leaves with symptoms of disease were cut, surface-sterilized, and plated on potato dextrose agar (PDA). Fungal colonies (28×28 mm) with grayish-black mycelia from all tissues were produced after 3 days of incubation at 26 °C. The mature colonies eventually turned black in the center, with obvious rings appearing after 10 days of culture. Basal conidia ranged from pale to dark brown, whereas apical ones were pale brown, with central cells being larger and darker than basal cells. Conidia were smooth and either fusiform, cylindrical, or slightly curved with rounded tips. They ranged in length from 22.34 to 36.82 (mean = 28.63) μm with 2–4 septations and slight septal constrictions. Monospore isolation was performed to obtain a pure culture. Strain BJ2Y5 was subsequently stored in the strain Preservation Center of Wuhan University (Wuhan, China) and the strain preservation number CCTCC M 2023123 was obtained. Fresh mycelia and conidia that grew at 26 ℃ for 7 days were collected from PDA plates. DNA was extracted using the Ezup Column Fungi Genomic DNA Purification Kit (Sangon Biotech Co., Shanghai, China). The phylogenetic position of isolate BJ2-Y5 was clarified based on DNA sequence analysis of three loci, namely glyceraldehyde 3-phosphate dehydrogenase (GAPDH; Berbee et al. 1999), the internal transcribed spacer (ITS) region (White et al. 1990), and partial sequences of the second largest subunit of RNA polymerase II (RPB2; O’Donnell et al. 2007). A BLAST search (GenBank accession nos. OP913168, OP743380, and OP913171) showed 99% homology to the reference isolate CBS 220.52. Based on the morphological and molecular information presented in this study these isolates were identified as C. geniculata (Hosokawa et al. 2003). Furthermore, we evaluated the pathogenicity of B. striata leaves by smearing a conidial suspension (106 conidia/mL) on both sides of leaves with and without wounds. Five inoculated and three non-inoculated leaves (smeared with sterile distilled water as a negative control) were kept in a greenhouse at 26 °C under natural sunlight and covered with plastic bags for 72 h to maintain humidity. After 7 days, small round spots appeared on the wounds. Fifteen days later, the symptoms of disease on the wounded inoculated leaves were similar to those observed in the original sample, whereas the control plants remained healthy. No symptoms of infection were observed in unwounded inoculated leaves. C. geniculata was successfully re-isolated from all five inoculated leaves and was confirmed based on Koch’s postulates. To the best of our knowledge, C. geniculata infection has not been previously reported in B. striata.

Major depression disorder (MDD) is a neuropsychiatric disorder characterized by abnormal neuronal activity in specific brain regions. A factor that is crucial in maintaining normal neuronal functioning is intracellular pH (pHi) homeostasis. In this study, we show that chronic stress, which induces depression-like behaviors in animal models, down-regulates the expression of the hippocampal Na

Curcumin is a polyphenol substance considered to be effective in the treatment of a number of neurodegenerative diseases. However, details regarding the exact mechanisms for the protective effects of curcumin in neuropsychiatric disorders, like depression, remain unknown. In the pathogenesis of major depressive disorder (MDD) it appears that dysregulation of oxidative stress and immune systems, particularly within the hippocampal region, may play a critical role. Here, we show that pre-treatment with curcumin (40 mg kg−1) alleviates depression-like behaviors in a LPS-induced rat model of depression, effects which were accompanied with suppression of oxidative stress and inflammation and an inhibition of neuronal apoptosis in the hippocampal CA1 region, and results from ultramicrostructure electrophysiological experiments revealed that the curcumin pre-treatment significantly prevented excessive synaptic loss and enhanced synaptic functioning in this LPS-induced rat model. In addition, curcumin attenuated the increases in levels of miR-146a-5p and decreases in the expression of p-ERK signaling that would normally occur within CA1 regions of these depressed rats. Taken together, these results demonstrated that curcumin exerts neuroprotective and antidepressant activities by suppressing oxidative stress, neural inflammation and their related effects upon synaptic dysregulation. One of the mechanisms for these beneficial effects of curcumin appears to involve the miR-146a-5p/ERK signaling pathway within the hippocampal CA1 region. These findings not only elucidated some of the mechanisms underlying the neuroprotective/antidepressant effects of curcumin, but also suggested a role of curcumin as a potential therapeutic strategy for depression.

Asian pear (Pyrus pyrifolia Nakai cv. cuiguan), is widely grown in Zhejiang province of China. In April 2019, symptoms consisting of small black, round leaf spots and blight flower petals were observed on over 30 % of 'Cuiguan' pear trees in an orchard (ca. 0.8 ha) near Cixi city, Zhejiang province, China. Initially, leaf spots were observed on leaf petioles, which, with time, enlarged and coalesced into necrotic streaks (1-2 cm) along the length of the petioles. Irregularly, reddish brown spots developed on flower petals, which hastened their senescence. Additional symptoms included round, black spots on leaves (2-3 mm in diameter) and necrosis of shoot tips. Symptomatic tissues from petals, petioles and leaves were plated onto potato dextrose agar (PDA). After five days of incubation at 26 °C, slimy fungal colonies (48×48 mm) with pinkish to orange-colored mycelia and with regular annulations were isolated from all tissues. After 10 days, cultured were shiny and dark brown in the center. The color of conidia ranged from hyaline to dark brown. Hyaline conidia were blastic, unicellular, ellipsoidal, smooth, with lengths that ranged from 11.03 to 27.14 (avg. 18.38) μm, and widths that ranged from 3.45 to 8.86 (avg. 6.04) μm (n = 50). Dark brown conidia were 1 to 2 celled, 10.89 to 26.03 (avg. 17.41) μm in length and 4.26 to 12.15 (avg. 6.94) μm in width (n = 50), and a slight septal constriction. Conidiogenous cells were clavate, hyaline, eseptate and top smoothly with 3-11 spores. Indistinct scars remained when the conidia dislodge from the conidiogenous cells. Single spore isolation was used to obtain pure cultures. Mycelia and conidia were scraped from cultures and DNA was extracted using Ezup Column Fungi Genomic DNA Purification Kit (Sangon Biotech). Amplified PCR products from the internal transcribed spacer (ITS) region ITS1F/ITS4 (White et al. 1990), the partial 28S rDNA (LSU) NL1/NL4 (Boekhout et al. 1995), the β-tubulin (TUB) gene Bt2a/Bt2b (GlassDonaldson 1995) and the partial elongase gene (ELO) ELO2-F/ELO2-R (Zalar et al. 2008) were sequenced (Tsingke Biotechnology Co., Hangzhou, Zhejiang). A blast search (GenBank Accession No. MT107050, OK485685, OK631951, OK631950) showed 99% Aureobasidium pullulans reference isolate CBS584.75 and EXF-150, which was consistent with the morphological data (Cene et al. 2014). Three-yr-old seedlings from 'Cuiguan' pears were spray with conidial suspension (106 conidia/ml) on the both sides of leaves without wounding. In a greenhouse (26 °C, natural light), six inoculated plants and three noninoculated plants (sprayed with sterile distilled water) enclosed in plastic bags to maintain humidity for 72 h. At 5 days after inoculation, shoot tips blackened and began to wilt. At 15 days after inoculation, symptoms similar to those on the original sample developed on inoculated petioles and leaves, whereas the control plants remained healthy. No symptoms were observed on leaves that were mature at the time of inoculation. Aureobasidium pullulans var. pullulans was reisolated from all inoculated plant. Overall, this disease shortened the life of pear flowers and reduced fruit set. To our knowledge, A. pullulans var. pullulans has not previously been reported as a pathogen of P. pyrifolia.

Background: Noise is unavoidable in the physiological signal measurement system. Poor quality signals can affect the results of analysis and disable the following clinical diagnosis. Thus, it is necessary to perform signal quality assessment before we interpreting the signal. Objective: In this work, we describe a method combing support vector machine (SVM) and multi-feature fusion for assessing the signal quality of pulsatile waveforms, concentrating on the photoplethysmogram (PPG). Methods: PPG signals from 53 healthy volunteers were recorded. Each had a 5 min length. Signal quality in each heart beat was manual annotated by clinical expert, and then the signal quality in 5 s episode was automatically calculated according to the results from each beat segments, resulting in a total of 13,294 5-s PPG segments. Then a SVM was trained to classify clean/noisy PPG recordings by inputting a set of twelve signal quality features. Further experiments were carried out to verify the proposed SVM based signal quality classifier method. Results: An average accuracy of 87.90%, a sensitivity of 88.10% and a specificity of 87.66% were found on the 10-fold cross validation. Conclusions: The signal quality of PPGs can be accurately classified by using the proposed method.

Aldehyde oxidases (AOXs) are a subfamily of cytosolic molybdo-flavoenzymes that play critical roles in the detoxification and degradation of chemicals. Active AOXs, such as AOX1 and AOX2, have been identified and functionally analyzed in insect antennae but are rarely reported in other tissues. This is the first study to isolate and characterize the cDNA that encodes aldehyde oxidase 5 (BmAOX5) in the pheromone gland (PG) of the silkworm, Bombyx mori. The size of BmAOX5 cDNA is 3,741 nucleotides and includes an open reading frame, which encodes a protein of 1,246 amino acid residues. The theoretical molecular weight and isoelectric point of BmAOX5 are approximately 138 kDa and 5.58, respectively. BmAOX5 shares a similar primary structure with BmAOX1 and BmAOX2, containing two [2Fe-2S] redox centers, a FAD-binding domain, and a molybdenum cofactor (MoCo)-binding domain. RT–PCR revealed BmAOX5 to be particularly highly expressed in the PG (including ovipositor) of the female silkworm moth, and the expression was further confirmed by in situ hybridization, AOX activity staining, and anti-BmAOX5 western blotting. Further, BmAOX5 was shown to metabolize aromatic aldehydes, such as benzaldehyde, salicylaldehyde, and vanillic aldehyde, and fatty aldehydes, such as heptaldehyde and propionaldehyde. The maximum reaction rate (Vmax) of benzaldehyde as substrate was 21 mU and Km was 1.745 mmol/liter. These results suggested that BmAOX5 in the PG could metabolize aldehydes in the cytoplasm for detoxification or participate in the degradation of aldehyde pheromone substances and odorant compounds to identify mating partners and locate suitable spawning sites.

As a population-based national surveillance region, Tianning District confronts with great challenges in birth defects (BDs) prevention. We aimed to describe the epidemiology of BDs in infants (including dead fetus, stillbirth, or live birth between 28 weeks of gestation and 42 days after birth) in Tianning District from 2014 to 2018. The data was collected from the national birth defect surveillance system. The prevalence rates of BDs were calculated by poisson distribution. Trends of incidence and the associations of regarding perinatal characteristics with BDs were analyzed by poisson regression. During the study period, the prevalence of BD was 155.49 per 10,000 infants. The ten leading BDs were congenital heart defects (CHD), polydactyly, Congenital malformation of kidney (CMK), syndactyly, cleft palate, hypospadias, Congenital hypothyroidism (CH), congenital atresia of rectum and anus, congenital talipes equinovarus (CTE), and microtia. A significant increase in the prevalence of CHD was observed with a prevalence rate ratio (PRR) of 1.191. Gravidity ≥ 3 (PRR = 1.38) and multiple births (PRR = 2.88) were risk factors for BDs. Premature delivery (PRR = 4.29), fetal death or stillbirth (PRR = 24.79), and infant death (PRR = 43.19) were adverse consequences of BDs. Strengthening publicity and education, improving the ability of diagnosis and monitoring, expanding surveillance time quantum of BDs system may be warranted.

Changzhou has been confronted with great challenges in birth defects (BDs) prevention, as the prevalence rates of BDs in Changzhou increased rapidly. The aims of this study were to describe the epidemiology of BDs in perinatal infants (PIs, including dead fetus, stillbirth, or live birth between 28 weeks of gestation and 7 days after birth) in Changzhou during the period from 2014 to 2018.The BD surveillance data of PIs were collected from 56 hospitals of Changzhou. The prevalence rate of BDs with 95% confidence interval (CI) were calculated by Poisson distribution. Univariate and multivariate Poisson regression was performed to identify the changing trends of prevalence rates of BDs by year and the association of regarding BD characteristics including year, infant gender, maternal age, and season with BDs successively.From 2014 to 2018, there were a total of 238,712 PIs of which 1707 had BDs, with the average prevalence of 71.509 per 10,000 PIs, showing a remarkable uptrend (aPRR = 1.133, 95%CI: 1.094-1.173). The ten leading BDs were polydactyly, congenital heart defects (CHD), syndactyly, microtia, cleft lip and palate (CLP), hypospadias, cleft palate, other malformation of external ear (OMEE), congenital atresia of rectum and anus, and congenital talipes equinovarus (CTE). During the study period, the prevalence rates of polydactyly, CHD and syndactyly increased significantly (PRR = 1.195, 95%CI: 1.109-1.288, PRR = 1.194, 95%CI: 1.105-1.291, and PRR = 1.143, 95%CI: 1.007-1.297, respectively); the prevalence rates of congenital esophageal atresia decreased significantly (PRR = 0.571, 95%CI: 0.395-0.826). The risk of BDs was higher in male PIs versus female PIs (aPRR = 1.235, 95%CI: 1.123-1.358).A significant increase in the prevalence of BDs was detected from 2014 to 2018 in Changzhou. CHD, polydactyly, and syndactyly increased much and congenital esophageal atresia declined much. Male PIs was risk factor for occurrence of BDs. Collecting information on factors associated with BDs, setting the report time of BDs system at smaller gestational age so as to get an exact prevalence and make better prevention strategy, strengthening the publicity and education, improving the ability of monitoring, and wider use of new diagnosis technology are important to reduce the prevalence of BDs in PIs.

Background/Aims: Ginsenoside Rg1 has been demonstrated to exhibit neuroprotective effects in various studies. This study aimed to investigate the neuronal mechanisms underlying the neuroprotective and antidepressant-like effects of ginsenoside Rg1 in a rat model of depression. Methods: Chronic unpredictable mild stress was used to induce depression-like behaviors in rats. Transmission electron microscopy was used to observe neuronal synapses within the basolateral amygdala (BLA). The expression of microRNA (miR)-134 in the BLA was verified by real-time quantitative PCR. Finally, the synaptic plasticity-associated proteins CAMP-response element binding protein (CREB) and brain-derived neurotrophic factor (BDNF) were detected by immunoblotting. Results: Results showed that chronic stress effectively induced depression-like behaviors in rats, which were associated with significant ultrastructural changes within BLA neurons. Moreover, chronic stress decreased the expression of miR-134 in the BLA, which was accompanied by decreased phosphorylation of CREB and decreased expression of BDNF. Remarkably, chronic administration of ginsenoside Rg1 (40 mg/kg, i.p., 5 weeks) significantly ameliorated the neuronal structural abnormalities and biochemical changes induced by chronic stress, as well as preventing depression-like behaviors in these rats. Conclusion: Results suggested that ginsenoside Rg1 may exhibit neuroprotection and antidepressant-like effects by activating the CREB-BDNF system within the BLA in this rat model of depression. Amelioration of depression-like behaviors by ginsenoside Rg1 appears to involve modulation of the synapse-associated factor miR-134 within the BLA. Therefore, these findings demonstrate some of the neuronal mechanisms associated with depression and the therapeutic potential of ginsenoside Rg1 for use in the treatment of depression in clinical trials.

Objective The aim of this study was to explore health anxiety (HA) in a sample of hospital medical employees and to identify factors that influence HA. Methods A consecutively recruited sample of 1702 medical employees with or without HA was obtained from 13 hospitals across China. Participants’ demographic and clinical characteristics were collected using a standardized protocol and data collection procedure. Subjects were divided into a HA and non-HA group according to their scores on the Chinese version of the Short Health Anxiety Inventory. Comparisons between groups were conducted and binary logistic regression was used to identify risk factors of HA. Results Total HA prevalence was 30.14%. There were significant differences between the HA and non-HA groups in number of working years, hospital category, sex, marital status, family income, personality, physical disease and education degree. Working in a specialist hospital, being female, being married, low income, introversion, graduate education or above and presence of physical disease were risk factors of HA. Conclusions HA is common in medical employees. More investigation of the long-term impact of HA is warranted.

Recent studies have indicated that physiological parameters change with different emotion states. This study aimed to quantify the changes of vascular function at different emotion and sub-emotion states. Twenty young subjects were studied with their finger photoplethysmographic (PPG) pulses recorded at three distinct emotion states: natural (1 minute), happiness and sadness (10 minutes for each). Within the period of happiness and sadness emotion states, two sub-emotion states (calmness and outburst) were identified with the synchronously recorded videos. Reflection index (RI) and stiffness index (SI), two widely used indices of vascular function, were derived from the PPG pulses to quantify their differences between three emotion states, as well as between two sub-emotion states. The results showed that, when compared with the natural emotion, RI and SI decreased in both happiness and sadness emotions. The decreases in RI were significant for both happiness and sadness emotions (both P< 0.01), but the decreases in SI was only significant for sadness emotion (P< 0.01). Moreover, for comparing happiness and sadness emotions, there was significant difference in RI (P< 0.01), but not in SI (P= 0.9). In addition, significant larger RI values were observed with the outburst sub-emotion in comparison with the calmness one for both happiness and sadness emotions (both P< 0.01) whereas significant larger SI values were observed with the outburst sub-emotion only in sadness emotion (P< 0.05). Moreover, gender factor hardly influence the RI and SI results for all three emotion measurements. This pilot study confirmed that vascular function changes with diffenrt emotion states could be quantified by the simple PPG measurement.

This pilot study investigated the differences of heart rate variability (HRV) indices between two opposite emotion states: happiness and sadness, to reveal the differences of autonomic nervous system activity under different emotional states. Forty-eight healthy volunteers were enrolled for this study. Electrocardiography (ECG) signals were recorded under both emotion states with a random measurement order (first happiness emotion measurement then sadness or reverse). RR interval (RRI) time series were extracted from ECGs and multiple HRV indices, including time-domain (MEAN, SDNN, RMSSD and PNN50), frequency-domain (LFn, HFn and LF/HF) and nonlinear indices (SampEn and FuzzyMEn) were calculated. In addition, the effects of heart rate (HR) and mean artery pressure (MAP) on the aforementioned HRV indices were analyzed for both emotion states. The results showed that experimental order had no significant effect on all HRV indices from both happiness and sadness emotions (all P > 0.05). The key result was that among all nine HRV indices, six indices were identified having significant differences between happiness and sadness emotion states: MEAN (P = 0.028), SDNN (P = 0.002), three frequency-domain indices (all P < 0.0001) and FuzzyMEn (P = 0.047), whereas RMSSD, PNN50 and SampEn had no significant differences between the two emotion states. All indices, except for SampEn, had significant positive correlations (all P < 0.01) for the two emotion states. Four time-domain indices decreased with the increase of HR (all P < 0.01), while frequency-domain and nonlinear indices demonstrated no HR-related changes for each emotional state. In addition, all indices (time-domain, frequency-domain and nonlinear) showed no MAP-related changes. It concluded that HRV indices showed significant differences between happiness and sadness emotion states and the findings could help to better understand the inherent differences of cardiovascular time series between different emotion states in clinical practice.

Peroxisomes are ubiquitous organelles in eukaryotic cells that fulfil a variety of biochemical functions. The biogenesis of peroxisomes requires a variety of proteins, named peroxins, which are encoded by PEX genes. Pex14/17 is a putative recently identified peroxin, specifically present in filamentous fungal species. Its function in peroxisomal biogenesis is still obscure and its roles in fungal pathogenicity have not yet been documented. Here, we demonstrate the contributions of Pex14/17 in the rice blast fungus Magnaporthe oryzae (Mopex14/17) to peroxisomal biogenesis and fungal pathogenicity by targeting gene replacement strategies. Mopex14/17 has properties of both Pex14 and Pex17 with regard to its protein sequence. Mopex14/17 is distributed at the peroxisomal membrane and is essential for efficient peroxisomal targeting of proteins containing peroxisomal targeting signal 1. MoPEX19 deletion leads to the cytoplasmic distribution of Mopex14/17, indicating that the peroxisomal import of Pex14/17 is dependent on Pex19. The knockout mutants of MoPEX14/17 show reduced fatty acid utilization, reactive oxygen species (ROS) degradation and cell wall integrity. Moreover, Δmopex14/17 mutants show delayed conidial generation and appressorial formation, and a reduction in appressorial turgor accumulation and penetration ability in host plants. These defects result in a significant reduction in the virulence of the mutant. These data indicate that MoPEX14/17 plays a crucial role in peroxisome biogenesis and contributes to fungal development and pathogenicity.

To develop and verify the Psychosomatic Symptom Scale (PSSS) among psychosomatic patients and the cut-off value of PSSS in distinguishing psychosomatic patients from health controls.The PSSS was drafted by an expert workgroup. 996 patients and 366 controls from 14 general hospitals in China were recruited to complete PSSS, Patient Health Questionnaire-15 (PHQ-15) and Symptom Checklist-90 (SCL-90). Student's t-test, Kruskal-Wallis test, Cronbach's α, Spearman's correlation, and confirmatory factor analysis (CFA) were used to verify the PSSS. Receiver operating characteristic (ROC) analyses were used to determine the cut-off value.Cronbach α of PSSS was 0.907. The PSSS was significantly correlated with SCL-90 somatization subscale (r = 0.682, P 0.001) and PHQ-15 (r = 0.724, P 0.001). CFA supported the theoretical two-factor structure of the PSSS, with comparative fit index (CFI) = 0.979, Tucker-Lewis index (TLI) = 0.977, root mean square error of approximation (RMSEA) = 0.039 (90% CI: 0.035-0.042), and standardized root mean residual (SRMR) = 0.054. As the sum score of PSSS was significantly higher in female, cut-off values were determined as 11 in females and 10 in males respectively.The PSSS is a reliable and valid instrument for measuring psychosomatic symptoms.

Pyricularia oryzae is the pathogen for rice blast disease, which is a devastating threat to rice production worldwide. Lysine succinylation, a newly identified post-translational modification, is associated with various cellular processes. Here, liquid chromatography tandem-mass spectrometry combined with a high-efficiency succinyl-lysine antibody was used to identify the succinylated peptides in P. oryzae. In total, 2109 lysine succinylation sites in 714 proteins were identified. Ten conserved succinylation sequence patterns were identified, among which, K*******Ksuc, and K**Ksuc, were two most preferred ones. The frequency of lysine succinylation sites, however, greatly varied among organisms, including plants, animals, and microbes. Interestingly, the numbers of succinylation site in each protein of P. oryzae were significantly greater than that of most previous published organisms. Gene ontology and KEGG analysis showed that these succinylated peptides are associated with a wide range of cellular functions, from metabolic processes to stimuli responses. Further analyses determined that lysine succinylation occurs on several key enzymes of the tricarboxylic acid cycle and glycolysis pathway, indicating that succinylation may play important roles in the regulation of basal metabolism in P. oryzae. Furthermore, more than 40 pathogenicity-related proteins were identified as succinylated proteins, suggesting an involvement of succinylation in pathogenicity. Our results provide the first comprehensive view of the P. oryzae succinylome and may aid to find potential pathogenicity-related proteins to control the rice blast disease. Significance Plant pathogens represent a great threat to world food security, and enormous reduction in the global yield of rice was caused by P. oryzae infection. Here, the succinylated proteins in P. oryzae were identified. Furthermore, comparison of succinylation sites among various species, indicating that different degrees of succinylation may be involved in the regulation of basal metabolism. This data facilitates our understanding of the metabolic pathways and proteins that are associated with pathogenicity.

Peroxisomes are ubiquitous organelles in eukaryotic cells that fulfil a variety of biochemical functions. The biogenesis of peroxisomes requires a variety of proteins, named peroxins, which are encoded by PEX genes. Pex14/17 is a putative recently identified peroxin, specifically present in filamentous fungal species. Its function in peroxisomal biogenesis is still obscure and its roles in fungal pathogenicity have not yet been documented. Here, we demonstrate the contributions of Pex14/17 in the rice blast fungus Magnaporthe oryzae (Mopex14/17) to peroxisomal biogenesis and fungal pathogenicity by targeting gene replacement strategies. Mopex14/17 has properties of both Pex14 and Pex17 with regard to its protein sequence. Mopex14/17 is distributed at the peroxisomal membrane and is essential for efficient peroxisomal targeting of proteins containing peroxisomal targeting signal 1. MoPEX19 deletion leads to the cytoplasmic distribution of Mopex14/17, indicating that the peroxisomal import of Pex14/17 is dependent on Pex19. The knockout mutants of MoPEX14/17 show reduced fatty acid utilization, reactive oxygen species (ROS) degradation and cell wall integrity. Moreover, Δmopex14/17 mutants show delayed conidial generation and appressorial formation, and a reduction in appressorial turgor accumulation and penetration ability in host plants. These defects result in a significant reduction in the virulence of the mutant. These data indicate that MoPEX14/17 plays a crucial role in peroxisome biogenesis and contributes to fungal development and pathogenicity.

The family members of PEX11 are key factors involved in regulation of peroxisome proliferation. Sixty-six PEX11p candidates of PEX11 gene family from 26 representative fungal species were obtained and analyzed by bioinformatic strategies. In most filamentous fungi, 2 or 3 potential PEX11ps were found, in contrast with 1 or 2 in yeast species. Compared with other fungal species, the Ascomycetes tend to have more PEX11ps, and even 5 in several individuals. The data of phylogenetic analysis and protein structure indicated that all of the PEX11ps were divided into 3 groups: I, II, and III. The members of group I and group III existed in most species, while those in group II were found only in Pezizomycotina. By MEME analysis, 5-6 conserved motifs were found in each PEX11ps. Among them,motif 8 in C-terminal had the most conservation, indicating that this motif probably plays a key role in maintaining the proper function of PEX11p.

Chickpea sprouts (CS) may become one of the natural products of high demand due to their nutritional values. The aim of this work was to study the enrichment of vanadium (V) during the germination of chickpea seeds in order to combine the high nutritive value of sprouts with a readily bioavailable source of essential trace element-V. Chickpea seeds were germinated in sodium metavanadate solutions (NaVO(3)) with different concentrations (50, 100, 150, 200, 400, 600, 800, 1,000, 1,200, 1,400, and 1,600 mg/L as NaVO(3)) for 6 days at 28 °C. It was found that the addition of sodium metavanadate produced slight stress effect, and the growth of CS was inhibited. The addition of sodium metavanadate significantly increased V content in CS in a dose-dependent manner. The highest V content (1.52 mg/g) in CS was observed when germination was carried out in the NaVO(3) solution with 1,200 mg/L NaVO(3) concentration. Chickpea seeds germinated in 50 ∼200 mg/L NaVO(3) solutions for 6 days at 28 °C showed a higher germination rate (>99%%) and a higher content of isoflavones including formononetin and biochanin A than those obtained in NaVO(3) solutions with other concentration in the same germination conditions. Therefore, a concentration of 50∼200 mg/L sodium metavanadate solutions should be used for the production of V-enriched chickpea sprouts (VCS).

To find new genes involved in fungal pathogenicity, a mutant (B11) exhibiting enhanced pathogenicity was isolated from an Agrobacterium -mediated transformed Magnaporthe oryzae mutant library. Southern blotting analysis showed that T-DNA insertion in the B11 genome was a single copy. TAIL-PCR and sequence alignment analyses revealed that a putative gene locus MG01679 was interrupted by the T-DNA fragment. By using the PCR-based method, the DNA and cDNA of the mutant gene MG01679 was cloned and sequenced. The open reading frame of MG01679 includes one intron and two exons, and the coding sequence is 696 bp in length and encodes a 231 amino acid peptide. Protein similarity analysis indicated that the gene belongs to the ThiJ/Pfp I protein family, and the gene was thus designated MgThiJ1 . MgThiJ1 showed 57% similarity to FOXG_09029 from Fusarium oxysporum and 54% similarity to FGSG_08979 from F. graminearum in protein sequence. MgThiJ1 gene might act as a negative regulator in vegetative growth and pathogenesis in filamentous fungi, and its specific mechanism needs to be studied further.

The peroxisomal matrix proteins involved in many important biological metabolism pathways in eukaryotic cells are encoded by nucleal genes, synthesized in the cytoplasm and then transported into the organelles. Targeting and import of these proteins depend on their two peroxisomal targeting signals (PTS1 and PTS2) in sequence as we have known so far. The vectors of the fluorescent fusions with PTS, i.e., green fluorescence protein (GFP)-PTS1, GFP-PTS2 and red fluorescence protein (RFP)-PTS1, were constructed and introduced into Magnaporthe oryzae Guy11 cells. Transformants containing these fusions emitted fluorescence in a punctate pattern, and the locations of the red and green fluorescence overlapped exactly in RFP-PTS1 and GFP-PTS2 co-transformed strains. These data indicated that both PTS1 and PTS2 fusions were imported into peroxisomes. A probable higher efficiency of PTS1 machinery was revealed by comparing the fluorescence backgrounds in GFP-PTS1 and GFP-PTS2 transformants. By introducing both RFP-PTS1 and GFP-PTS2 into Deltamgpex6 mutants, the involvement of MGPEX6 gene in both PTS1 and PTS2 pathways was proved. In addition, using these transformants, the inducement of peroxisomes and the dynamic of peroxisomal number during the pre-penetration processes were investigated as well. In summary, by the localization and co-localization of PTS1 and PTS2, we provided a useful tool to evaluate the biological roles of the peroxisomes and the related genes.

Vanadium compounds have been recognized for their hypoglycemic effects; however, potential short and long-term vanadium toxicity has slowed the acceptance for therapeutic use. In the present work, three batches of vanadium-enriched chickpea sprout (VCS) were prepared by incubating chickpea seeds in presence of 200, 100, and 50 microg/ml of sodium orthovanadate (SOV). The effects of oral administration of chickpea sprout (CS) and VCS food for 8 weeks on streptozotocin-induced (STZ) diabetic rats were investigated. Both CS and VCS food was found to ameliorate some hyperglycemic symptoms of the diabetic rats, i.e. improve lipid metabolism, decrease blood glucose level, prevent body weight loss, and reduce impairment of diabetic related spatial learning and memory. Serum insulin was substantially elevated in treated diabetic rats, which is probably one important reason for the hypoglycemic effect. Compared with CS alone, VCS100 food exhibited remarkably enhanced effectiveness in alleviating diabetes induced hyperglycemia and memory loss. Moreover, vanadium-enriched chickpeas appeared to abolish the vanadium induced toxicity associated with administration of this metal for diabetes during the 8-week study period. This study suggested further work of the vanadium speciation in CS and novel hypoglycemic mechanism for the antidiabetic activity of vanadium agents. Vanadium containing (VCS) food could be a dietary supplement for the diabetic status.

Peroxisomes play important roles in metabolisms of eukaryotes and infection of plant fungal pathogens. These organelles proliferate by de novo formation or division in response to environmental stimulation. Although the assembly of peroxisomes was documented in fungal pathogens, their division and its relationship to pathogenicity remain obscure. In present work, we analyzed the roles of three Pex11 family members in peroxisomal division and pathogenicity of the rice blast fungus Magnaporthe oryzae. Deletion of MoPEX11A led to fewer but enlarged peroxisomes, and impaired the separation of Woronin bodies from peroxisomes, while deletion of MoPEX11B or MoPEX11C put no evident impacts to peroxisomal profiles. MoPEX11A mutant exhibited typical peroxisome related defects, delayed conidial germination and appressoria formation, and decreased appressorial turgor and host penetration. As a result, the virulence of MoPEX11A mutant was greatly reduced. Deletion of MoPEX11B and MoPEX11C did not alter the virulence of the fungus. Further, double or triple deletions of the three genes were unable to enhance the virulence decrease in MoPEX11A mutant. Our data indicated that MoPEX11A is the main factor modulating peroxisomal division and is required for full virulence of the fungus.

A flow injection on-line coprecipitation system with diethyldithiocarbamate (DDTC) nickel (II) being used as a carrier was coupled to the flame atomic absorption spectrometry (FAAS) for the determination of trace copper, lead, cadmium and iron in environmental and biological samples. Metal ions were on-line coprecipitated with DDTC-Ni (II) in 0.3 mol·l−1 nitric acid, and the precipitate was collected in a knotted reactor. The precipitate was then dissolved by isobutyl methyl ketone (IBMK), and the concentrated zone was transported directly into the nebulizer-burner system of a FAAS. Enhancement factors of 60, 58, 65, and 59 were obtained for copper, lead, cadmium and iron, respectively, at the sampling frequency of 60 h−1. The detection limits (3[sgrave]) were 0.5 μg·l−1 for copper, 2.7 μg·l−1 for lead, 0.2 μg·l−1 for cadmium, and 2.5 μg·l−1 for iron respectively, and a relative standard deviations (n=10) were 3.0% for 40 μg·l−1 copper, 2.0% for 80 μg·l−1 lead, 3.5% for 10 μg·l−1 cadmium, and 2.8...

A flow injection wetting-film extraction system without segmentor and phase separator has been coupled to flame atomic absorption spectrometry for the determination of trace copper. Isobutyl methyl ketone (MIBK) was selected as coating solvent and 8-hydroxyquinoline (oxine) as the chelating reagent. By switching of a 8-channel valve and alternative initiation of two peristaltic pumps, MIBK, sample solution containing copper chelate of oxine, and air-segment sandwiched eluting solution (1.0 mol l −1 nitric acid) were sequentially aspirated into an extraction coil made of PTFE tubing of 360 cm length and 0.5 mm i.d. The formation of organic film in the wall of the extraction coil, extraction of the copper chelate into the organic film and back-extraction of the analyte into the eluting solution occurred consecutively when these zones aspirated into the extraction coil were propelled down the extraction coil by a carrier solution at a flow rate of 2 ml min −1 . After leaving the extraction coil, the concentrated zone was transported to the nebulizer at its free uptake rate for atomization. Under the optimized conditions, an enrichment factor of 43 and a detection limit of 0.2 μg l −1 copper were achieved at a sample throughput rate of 30 h −1 . Eleven determinations of a standard copper solution of 60 μg l −1 gave a relative standard deviation of 1.5%. Foreign ions possibly present in tap water and natural water did not interfere with the copper determination. The developed method has been successfully used to the determination of copper content of tap water and river water.

Peroxisomes participate in various important metabolisms and are required in pathogenicity of fungal plant pathogens. Peroxisomal matrix proteins are imported from cytoplasm into peroxisomes through peroxisomal targeting signal 1 (PTS1) or peroxisomal targeting signal 2 (PTS2) import pathway. PEX5 and PEX7 genes participate in the two pathways respectively. The involvement of PEX7 mediated PTS2 import pathway in fungal pathogenicity has been documented, while that of PTS1 remains unclear. Through null mutant analysis of MoPEX5, the PEX5 homolog in Magnaporthe oryzae, we report the crucial roles of PTS1 pathway in the development and host infection in the rice blast fungus, and compared with those of PTS2. We found that MoPEX5 disruption specifically blocked the PTS1 pathway. Δmopex5 was unable to use lipids as sole carbon source and lost pathogenicity completely. Similar as Δmopex7, Δmopex5 exhibited significant reduction in lipid utilization and mobilization, appressorial turgor genesis and H(2)O(2) resistance. Additionally, Δmopex5 presented some distinct defects which were undetected in Δmopex7 in vegetative growth, conidial morphogenesis, appressorial morphogenesis and melanization. The results indicated that the PTS1 peroxisomal import pathway, in addition to PTS2, is required for fungal development and pathogenicity of the rice blast fungus, and also, as a main peroxisomal import pathway, played a more predominant role than PTS2.

Because the end-point parameters of an electric arc furnace (EAF) are affected by both quantitative factors and non-quantitative factors, we combine Nonlinear Gray Bernoulli-Markov model with Support Vector Machine (SVM) to produce a Nonlinear Gray Bernoulli-Markov-SVM prediction model for estimating the end-point parameter values of an EAF. The effects from the non-quantitative factors on the prediction values of end-point parameters are reflected by Nonlinear Gray Bernoulli-Markov model; while the effects from the quantitative inputs are reflected by the SVM. The Nonlinear Gray Bernoulli model that reflects non-quantitative factors is established firstly, and then, its prediction values are revised by the Markov chain. Because the effect from the quantitative inputs can not be reflected by Nonlinear Gray Bernoulli-Markov model, the Nonlinear Gray Bernoulli-Markov model is certainly not free from prediction errors from the quantitative inputs. These prediction errors are compensated by the SVM model. Further, combining Differential Evolution (DE) with Particle Swarm Optimization (PSO), a Differential Evolution Particle Swarm Optimization (DEPSO) is proposed. The parameter of Nonlinear Gray Bernoulli Model is optimized by the PSO algorithm, while that of SVM are optimized by the DEPSO algorithm. The final prediction values of the end-point parameters in EAF are thus obtained. Simultaneously, a rolling forecasting mechanism is introduced for improving the prediction precision.

The paper works out an online self-learning control plant for multiple support vector machine inverse control. The multiple support vector machine model applies subtractive clustering algorithm by which the input space is divided into several small local spaces. By means of least squares support vector machine, the sub-models are established. The prediction output of each sub-model is connected by principal components regression method so that identification of the inverse dynamics model of the system is achieved. Combining inverse model of the system as a system controller with the controlled plant, a SVM direct inverse control system is constituted. In order to overcome the influence of inverse model identification error, a SVM direct inverse control system with the PID compensation is designed in the paper. The simulation research proves that the control strategy can provide the system with good tracking performance, resistance to interference and a better robustness.

A moving object detection method based on the dense velocity field detection followed by an unsupervised spatiotemporal clustering by means of the mean shift algorithm is proposed. The method firstly makes use of a novel imaging system and an algebraic solution to detect the extremely dense velocity vector distribution with a pixel-wise spatial resolution and a frame-wise temporal resolution. It is based on the complex sinusoidal-modulated imaging with a three-phase correlation image sensor (3PCIS) and an exact algebraic inversion method based on an equivalent weighted integral form of the optical flow partial differential equation. Since the inversion method is free from time derivatives, any limitations on the object velocity and inaccuracies due to approximated time derivatives are thoroughly avoided. Secondly, in order to segregate dominant velocity regions from noisy background and identify the exact moving object, the mean shift clustering method is employed to the spatially and temporarily dense velocity vector distribution. To avoid too many cluster centers or over clustering, the traditional mean shift method is improved by setting an emerging condition. An experimental system was constructed with a 320×256 pixel 3PCIS device and a standard PC for inversion operations and display. Several experimental results are shown including an application to dense motion capture of face and gesture and traffic scene, which including several independent moving objects.

In the view of characteristics for coke micrograph, a segmentation algorithm combining mean shift and edge confidence, is proposed. Firstly, the edge confidence of image pixels is calculated, and with the edge confidence the weighting function of mean shift algorithm is computed, the sampling points of feature space are weighted in order to improve the accuracy of detected modes. Secondly, coke image is segmented preliminarily by iterating the weighted mean shift vector. Because that the number of clusters in initial segmentation is larger than that of the actual clusters, which may result in over-segmentation, the combining conditions are set by the spatial distance and the average value of the edge confidence. The coke photomicrograph is finally segmented with the new combining conditions. Experimental results show that with the proposed algorithm the segmentation among different optical textures of coke is more reasonable and effective.

To improve the efficiency of targeted gene replacement (TGR), a dual screen (DS) system with gusA gene as negative selective marker (GUS-DS) was developed in Magnaporthe oryzae. First, we tested the endogenous beta-glucuronidase (GUS) activities of 78 fungal strains. All tested strains were GUS-, only with 3 exceptions. Whereas, after the gusA being introduced in, M. oryzae, Fusarium oxysporum and Colletotrichum lagenarium acquired high GUS activities. The gusA is thus usable as a selective maker in fungal species. With gusA as the negative marker, HPH gene as the positive marker, and the peroxisomal targeting signal receptor genes MGPEX5 and MGPEX7 as 2 instances of target genes, we established the GUS-DS system. After transformation, we collected the transformants from hygromycin B screen media and then tested the GUS activities of them. The GUS- ones were selected as potential mutants and checked in succession by PCR and Southern blotting to identify the true mutants and calculate the efficiency of GUS-DS. As a result, GUS-DS improved the screen efficiency for delta mgpex5 from 65.8% to 90.6%, and for delta mgpex7 from 31.2% to 82.8%. In addition, we established a multiple PCR (M-PCR) method for mutant confirmation. By amplifying the different regions at the targeted locus, M-PCR differentiated the wild type, the ectopic transformants and the mutants effectively and rapidly, and had the same reliability as Southern blotting. In conclusion, GUS-DS and M-PCR are useful tools to improve the efficiency of TGR and would be helpful for fungal genomics.

Cloning of a Homologous Gene of PMK1 Type Mitogen-Activated Protein Kinase (MAPK) from the rice false smut fungus Ustilaginoidea virens.According to the conserved amino acid sequence of several filamentous fungus MAPKs, which were homologous to Magnaporthe grisea PMKI, degenerate PCR primers were designed to amplify the MAPK internal DNA fragment from Ustilaginoidea grisea. The complete UVMK1 DNA and cDNA sequences were obtained using Thermal Asymmetric Interlaced-PCR (TAIL-PCR) and RT-PCR methods. Functional Identification was done by using the M. grisea APMKI mutant stain nn78, including appressoria differentiation assay and barley infection test.The total length of UVMKJ was 1435 bp. It contained 3 introns and encoded 355 amino acids. The induced amino acid sequence showed identical to Magnaporthe grisea PMKI, Fusarium oxysporum FMKJ, Fusarium solani FsMAPK, Colletotrichum lagenarium CMKI, Botrytis cinerea BMKI, Claviceps purpurea CMPKI. After transformation of the APMK1 mutant of M. grisea using a complement vector with the complete cDNA of UVMK1 (under the M. grisea MPG1 promoter), five transformants were obtained. Furthermore, the selected two transformants fully restored their ability to form appressoria and infect a barley leaf.In this study, we characterized the frst MAPK protein from U. virens, and that UVMK1 is a homologue of M. grisea PMK1.

Magnaporthe oryzae is a hemibiotrophic fungal pathogen that causes rice blast disease. A compatible interaction requires overcoming plant defense responses to initiate colonization during the early infection process. Nitric oxide (NO) plays important roles in defense responses during host-pathogen interactions. Microbes generally protect themselves against NO-induced damage by using enzymes. Here, we characterized an S-(hydroxymethyl)- glutathione dehydrogenase gene in M. oryzae, MoSFA1, the homologs of which are involved in NO metabolism by specifically catalyzing the reduction of S-nitrosoglutathione (GSNO) in yeasts and plants. As expected from the activities of S-(hydroxymethyl)glutathione dehydrogenase in formaldehyde detoxification and GSNO reduction, MoSFA1 deletion mutants were lethal in formaldehyde containing medium, sensitive to exogenous NO and exhibited a higher level of S-nitrosothiols (SNOs) than that of the wild type. Notably, the mutants showed severe reduction of conidiation and appressoria turgor pressure, as well as significantly attenuated the virulence on rice cultivar CO-39. However, the virulence of MoSFA1 deletion mutants on wounded rice leaf was not affected. An infection assay on barley leaf further revealed that MoSFA1 deletion mutants exhibited a lower infection rate, and growth of infectious hyphae of the mutants was retarded not only in primary infected cells but also in expansion from cell to cell. Furthermore, barley leaf cell infected by MoSFA1 deletion mutants exhibited a stronger accumulation of H2O2 at 24 and 36 hpi. MoSFA1 deletion mutants displayed hypersensitivity to different oxidants, reduced activities of superoxide dismutases and peroxidases, and lower glutathione content in cells, compared with the wild type. These results imply that MoSFA1-mediated NO metabolism is important in redox homeostasis in response to development and host infection of M. oryzae. Taken together, this work identifies that MoSFA1 is required for conidiation and contributes to virulence in the penetration and biotrophic phases in M. oryzae.

Peroxisomes are present ubiquitously and make important contributions to cellular metabolism in eukaryotes. They play crucial roles in pathogenicity of plant fungal pathogens. The peroxisomal matrix proteins and peroxisomal membrane proteins (PMPs) are synthesized in the cytosol and imported post-translationally. Although the peroxisomal import machineries are generally conserved, some species-specific features were found in different types of organisms. In phytopathogenic fungi, the pathways of the matrix proteins have been elucidated, while the import machinery of PMPs remains obscure. Here, we report that MoPEX19, an ortholog of ScPEX19, was required for PMPs import and peroxisomal maintenance, and played crucial roles in metabolism and pathogenicity of the rice blast fungus Magnaporthe oryzae. MoPEX19 was expressed in a low level and Mopex19p was distributed in the cytoplasm and newly formed peroxisomes. MoPEX19 deletion led to mislocalization of peroxisomal membrane proteins (PMPs), as well peroxisomal matrix proteins. Peroxisomal structures were totally absent in Δmopex19 mutants and woronin bodies also vanished. Δmopex19 exhibited metabolic deficiency typical in peroxisomal disorders and also abnormality in glyoxylate cycle which was undetected in the known mopex mutants. The Δmopex19 mutants performed multiple disorders in fungal development and pathogenicity-related morphogenesis, and lost completely the pathogenicity on its hosts. These data demonstrate that MoPEX19 plays crucial roles in maintenance of peroxisomal and peroxisome-derived structures and makes more contributions to fungal development and pathogenicity than the known MoPEX genes in the rice blast fungus.