MethMarkerDB: a comprehensive cancer DNA methylation biomarker database.

DNA methylation plays a crucial role in tumorigenesis and tumor progression, sparking substantial interest in the clinical applications of cancer DNA methylation biomarkers. Cancer-related whole-genome bisulfite sequencing (WGBS) data offers a promising approach to precisely identify these biomarkers with differentially methylated regions (DMRs). However, currently there is no dedicated resource for cancer DNA methylation biomarkers with WGBS data. Here, we developed a comprehensive cancer DNA methylation biomarker database (MethMarkerDB, https://methmarkerdb.hzau.edu.cn/), which integrated 658 WGBS datasets, incorporating 724 curated DNA methylation biomarker genes from 1425 PubMed published articles. Based on WGBS data, we documented 5.4 million DMRs from 13 common types of cancer as candidate DNA methylation biomarkers. We provided search and annotation functions for these DMRs with different resources, such as enhancers and SNPs, and developed diagnostic and prognostic models for further biomarker evaluation. With the database, we not only identified known DNA methylation biomarkers, but also identified 781 hypermethylated and 5245 hypomethylated pan-cancer DMRs, corresponding to 693 and 2172 genes, respectively. These novel potential pan-cancer DNA methylation biomarkers hold significant clinical translational value. We hope that MethMarkerDB will help identify novel cancer DNA methylation biomarkers and propel the clinical application of these biomarkers.

Hepatic GDP-fucose transporter SLC35C1 attenuates cholestatic liver injury and inflammation by inducing CEACAM1 N153 fucosylation.

BACKGROUND AND AIMS: Inflammatory response is crucial for bile acid (BA)-induced cholestatic liver injury, but molecular mechanisms remain to be elucidated. Solute Carrier Family 35 Member C1 (SLC35C1) can transport GDP-fucose into the Golgi to facilitate protein glycosylation. Its mutation leads to the deficiency of leukocyte adhesion and enhances inflammation in humans. However, little is known about its role in liver diseases. APPROACH AND RESULTS: Hepatic SLC35C1 mRNA transcripts and protein expression were significantly increased in patients with obstructive cholestasis (OC) and mouse models of cholestasis. Immunofluorescence revealed that the upregulated SLC35C1 expression mainly occurred in hepatocytes. Liver-specific ablation of Slc35c1 (Slc35c1 cKO) significantly aggravated liver injury in mouse models of cholestasis induced by bile duct ligation and 1% cholic acid-feeding, evidenced by increased liver necrosis, inflammation, fibrosis, and bile ductular proliferation. The Slc35c1 cKO increased hepatic chemokine Ccl2 and Cxcl2 expression and T-cell, neutrophil and F4/80 macrophage infiltration, but did not affect the levels of serum and liver BA in mouse models of cholestasis. LC-MS/MS analysis revealed that hepatic Slc35c1 deficiency substantially reduced the fucosylation of cell-cell adhesion protein CEACAM1 at N153. Mechanistically, cholestatic levels of conjugated BAs stimulated SLC35C1 expression by activating the STAT3 signaling to facilitate CEACAM1 fucosylation at N153, and deficiency in the fucosylation of CEACAM1 at N135 enhanced the BA-stimulated CCL2 and CXCL2 mRNA expression in primary mouse hepatocytes and PLC/PRF/5-ASBT cells. CONCLUSIONS: Elevated hepatic SLC35C1 expression attenuates cholestatic liver injury by enhancing CEACAM1 fucosylation to suppress CCL2 and CXCL2 expression and liver inflammation.

ChromLoops: a comprehensive database for specific protein-mediated chromatin loops in diverse organisms.

Chromatin loops (or chromatin interactions) are important elements of chromatin structures. Disruption of chromatin loops is associated with many diseases, such as cancer and polydactyly. A few methods, including ChIA-PET, HiChIP and PLAC-Seq, have been proposed to detect high-resolution, specific protein-mediated chromatin loops. With rapid progress in 3D genomic research, ChIA-PET, HiChIP and PLAC-Seq datasets continue to accumulate, and effective collection and processing for these datasets are urgently needed. Here, we developed a comprehensive, multispecies and specific protein-mediated chromatin loop database (ChromLoops, https://3dgenomics.hzau.edu.cn/chromloops), which integrated 1030 ChIA-PET, HiChIP and PLAC-Seq datasets from 13 species, and documented 1 491 416 813 high-quality chromatin loops. We annotated genes and regions overlapping with chromatin loop anchors with rich functional annotations, such as regulatory elements (enhancers, super-enhancers and silencers), variations (common SNPs, somatic SNPs and eQTLs), and transcription factor binding sites. Moreover, we identified genes with high-frequency chromatin interactions in the collected species. In particular, we identified genes with high-frequency interactions in cancer samples. We hope that ChromLoops will provide a new platform for studying chromatin interaction regulation in relation to biological processes and disease.

ASMdb: a comprehensive database for allele-specific DNA methylation in diverse organisms.

DNA methylation is known to be the most stable epigenetic modification and has been extensively studied in relation to cell differentiation, development, X chromosome inactivation and disease. Allele-specific DNA methylation (ASM) is a well-established mechanism for genomic imprinting and regulates imprinted gene expression. Previous studies have confirmed that certain special regions with ASM are susceptible and closely related to human carcinogenesis and plant development. In addition, recent studies have proven ASM to be an effective tumour marker. However, research on the functions of ASM in diseases and development is still extremely scarce. Here, we collected 4400 BS-Seq datasets and 1598 corresponding RNA-Seq datasets from 47 species, including human and mouse, to establish a comprehensive ASM database. We obtained the data on DNA methylation level, ASM and allele-specific expressed genes (ASEGs) and further analysed the ASM/ASEG distribution patterns of these species. In-depth ASM distribution analysis and differential methylation analysis conducted in nine cancer types showed results consistent with the reported changes in ASM in key tumour genes and revealed several potential ASM tumour-related genes. Finally, integrating these results, we constructed the first well-resourced and comprehensive ASM database for 47 species (ASMdb, www.dna-asmdb.com).

Phylogeny and Fungal Community Structures of Helotiales Associated with Sclerotial Disease of Mulberry Fruits in China.

Mulberry fruit sclerotiniose is a prevalent disease caused by the fungal species Ciboria shiraiana, C. carunculoides, and Scleromitrula shiraiana of the order Helotiales, and severely affects the production of mulberry. However, these species have only been identified using morphological and rDNA-ITS sequence analyses, and their genetic variation is unclear. To address this, morphological and two-locus (ITS and RPB2) phylogenetic analyses were conducted using culture-dependent and independent methods for 49 samples from 31 orchards across four provinces in China. Illumina MiSeq sequencing was used to assess the fungal communities obtained from fruits varying in disease severity and color from an orchard in Wuhan. Conidial suspensions of C. shiraiana and C. carunculoides isolated from diseased fruits, diseased fruits affected with hypertrophy and pellet sorosis sclerotiniose, and mycelia of Sclerotinia sclerotiorum were determined to be pathogenic to the mulberry cultivar YSD10. However, fruits inoculated with S. sclerotiorum mycelia exhibited nontypical disease symptoms, and mycelia and conidia obtained from C. carunculoides and S. shiraiana strains were not pathogenic. Maximum parsimony and Bayesian analyses using the sequences of the assessed loci indicated species variability with no evidence of geographic specialization. Metagenomic analysis revealed that the diversity of fungal communities was reduced with disease progression. Furthermore, within a single fruit, the presence of two Ciboria spp. was detected. These results provide novel insights into Ciboria spp., revealing the secondary infections caused by conidia in diseased fruits, genetic variations of the pathogens, and the occurrence of coinfection. This improved understanding of fungal pathogens will aid in developing effective disease control strategies.

First Report of Neocosmospora mori Causing Root Rot and Stem Blight of Mulberry in Nanzhang, Hubei, China.

Mulberry (Morus alba L.) has been cultivated for thousands of years in many temperate regions in East Asia and is commonly used to feed silkworms. In May 2021, 5 to 8% incidence of stem blight on 4-year-old mulberry 'Nongsang 14' was observed in several orchards in Nanzhang County, Hubei Province, China. The roots and stems showed symptoms of vascular discoloration, and the tender new shoots, surrounded by white hyphae, were detached easily. Symptomatic stem tissues (5 mm × 5 mm) were excised from the border between diseased and healthy tissues, surface sterilized in a 75% ethanol solution for 30 s and 2.5% sodium hypochlorite for 1.5 min, washed three times in sterile distilled water, then placed on potato dextrose agar (PDA, 250 g potatoes, 2% dextrose, 1.6% agar), and incubated at 25°C in darkness. Two isolates (Bq2 and Bq3) were subcultured using the single-spore method. On PDA, colonies were cottony, with whitish aerial mycelium and the daily growth rate was 4.25 to 5.50 mm/day at 25°C in darkness. On carnation leaf agar, macroconidia were fusiform with slightly curved apical cells and foot-shaped basal cells, three to five septate, measuring 47.5 to 80.3 × 3.6 to 5.6 µm (average 68.7 × 4.7 µm, n = 30). On spezieller nährstoffarmer agar, microconidia were produced in false heads on monophialides, mostly 0-septate, oval, obovoid, or reniform in shape, measuring 5.1 to 10.7 × 2.7 to 5.3 µm (average 8.5 × 3.3 µm, n = 30). Chlamydospores were 4.9 to 11.0 µm in diameter (average 6.8 µm, n = 30), round shaped, thick-walled, and produced individually or in pairs or in chains. For molecular identification, the ribosomal internal transcribed spacers (ITS), translation elongation factor 1α (EF-1α), 28S large subunit nrDNA (LSU), and calmodulin (CAM) genes were amplified and sequenced with primers ITS1/ITS4 (White et al. 1990), EF1H/EF2T (O'Donnell et al. 1998), LR0R/LR5 ( Vilgalys and Hester 1990; Vilgalys and Sun 1994), and CL1/CL2A (Geiser et al. 2021; Wang et al. 2011), respectively. The sequences were deposited in GenBank (OQ711943-OQ711944 for ITS, OQ722438- Q722439 for EF-1α, OQ722441-OQ722442 for CAM, and OR116152-OR116153 for LSU). A maximum-likelihood phylogenetic analysis based on multilocus sequences was conducted using MEGA7, which showed that the two isolates grouped into a clade with Neocosmospora mori (previously Fusarium solani species complex) supported by a high bootstrap value (85%), and hence, they were identified as N. mori based on morphological and molecular analyses (Brooks et al. 2022; Crous et al. 2021; Lombard et al. 2015; Zeng and Zhuang 2023). To complete Koch's postulates, three healthy 2-month-old seedlings grown in sterile peat mix were removed from pots and the roots were washed in sterile water. Each plant was inoculated by dipping wounded and unwounded roots in a spore suspension (1 × 107 conidia/ml) for 20 min, and then 10 mL of the spore suspension was poured over the roots of each seedling after transplanting. Three plants were treated with sterilized water as a control. The tested plants were then kept in a plastic box containing sterile water and incubated at 25°C in a 12 h/12 h light/dark cycle. The pathogenicity assay was repeated three times for each isolate. Root and stem blight was observed 10 days after inoculation, while the control plants were asymptomatic. Furthermore, fungi with morphological characteristics of N. mori were only reisolated from the symptomatic stems and sequences of LSU matched those of isolates Bq2 and Bq3. This pathogen has been reported previously causing stem blight on mulberry trees in Japan and South Korea (Sandoval-Denis et al. 2019), but to our knowledge, this is the first report of N. mori causing root rot and stem blight of mulberry in China. This report will facilitate the development of effective control strategies for the disease.

A Novel Acoustic Evaluation Method for the Diagnosis of Adductor Spasmodic Dysphonia.

INTRODUCTION: Adductor spasmodic dysphonia (ADSD) is characterized by involuntary laryngeal muscle spasms. Due to the lack of a quantitative evaluation method, most measurements have demonstrated difficulty in validity and reliability for diagnosing ADSD. This study aimed to establish a novel indicator for ADSD and determine its diagnostic effects. METHODS: We investigated 98 voice samples from 49 patients with ADSD and 49 healthy subjects. A sustained vowel was recorded by a high-definition audio recorder. Voice samples underwent regular acoustic evaluation and a novel global dimension method. Global dimension (GD), Jitter, Shimmer, HNR, Frequency shift, and CPPS were measured for both groups. RESULTS: Statistical analysis revealed that the global dimension method effectively differentiated ADSD patients from healthy subjects (P<0.001, D'>0.8). Subsequent multiclass receiver operating characteristic analysis demonstrated that GD possessed the most significant classification accuracy (AUC = 0.988) compared with other acoustic parameters. CONCLUSION: GD was an effective metric for objective differentiation between ADSD patients and healthy subjects. This metric could assist clinicians in the diagnosis of ADSD patients.

A Feature Selection Approach Guided an Early Prediction of Anthocyanin Accumulation Using Massive Untargeted Metabolomics Data in Mulberry.

Identifying the early predictive biomarkers or compounds represents a pivotal task for guiding a targeted agricultural practice. Despite the various available tools, it remains challenging to define the ideal compound combination and thereby elaborate an effective predictive model fitting that. Hence, we employed a stepwise feature selection approach followed by a maximum relevance and minimum redundancy (MRMR) on the untargeted metabolism in four mulberry genotypes at different fruit developmental stages (FDSs). Thus, we revealed that 7 out of 226 differentially abundant metabolites (DAMs) explained up to 80% variance of anthocyanin based on linear regression model and stepwise feature selection approach accompanied by an MRMR across the genotypes over the FDSs. Among them, the phosphoenolpyruvate, d-mannose and shikimate show the top 3 attribution indexes to the accumulation of anthocyanin in the fruits of these genotypes across the four FDSs. The obtained results were further validated by assessing the regulatory genes expression levels and the targeted metabolism approach. Taken together, our findings provide valuable evidences on the fact that the anthocyanin biosynthesis is somehow involved in the coordination between the carbon metabolism and secondary metabolic pathway. Our report highlights as well the importance of using the feature selection approach for the predictive biomarker identification issued from the untargeted metabolomics data.

Controllable Size-Independent Three-Dimensional Inertial Focusing in High-Aspect-Ratio Asymmetric Serpentine Microchannels.

High-throughput three-dimensional (3D) focusing of cells is the key prerequisite for enabling accurate microfluidic cell detection and analysis. In this work, we develop a high-aspect-ratio asymmetric serpentine (HARAS) microchannel for single-line inertial focusing of particles and cells at the 3D center of the channel. The mechanism of 3D focusing is explored by numerical simulation, and the focusing performances of differently sized particles are characterized experimentally at different flow rates. The results demonstrate the outstanding 3D single-line focusing capability of our HARAS microchannel. In addition, the phenomena of size-independent and position-controllable focusing over wide flow rates are observed. Finally, the applicability of our HARAS microchannel for processing real biological cells is validated by the 3D single-line focusing of A549 cells and MCF-7 cells. Our work overcomes the issue of off-centered focusing of most previous works and provides new insights into the 3D focusing in inertial microfluidics. The proposed HARAS microchannel is extremely easy for mass production and may provide a stable, high-throughput, and position-controllable scheme for subsequent single-cell detection and analysis.

First Report of Colletotrichum aenigma Causing Anthracnose on Mulberry Leaves in China.

Mulberry (Morus alba L.) has been grown worldwide as a crop for silkworm rearing for over five thousand years (Jiao et al. 2020). In July 2021, a leaf spot disease was observed on mulberry leaves in Wuhan city (114°33'E, 30°48'N), Hubei province, China, with approximately 40% of leaves (about 300 trees) affected. Early symptoms were light brown, with small lesions subsequently expanding to larger sometimes irregular dark brown or black spots surrounded by yellow-brown margins, with easily perforated necrotic lesions. Leaf tissues (5 mm×5 mm) were excised from the border between diseased and healthy tissues, surface sterilized with 75% ethanol solution for 30 s and 2.5% sodium hypochlorite for 2 min, washed thrice in sterile distilled water, and then placed on potato dextrose agar (PDA), and incubated at 25°C in darkness. Four isolates (C1, C9, CHS2, and CHS6) were subcultured using the single-spore method. On PDA, colonies were cottony, pale white from above, and white to grayish-green on the reverse side. Conidia were aseptate, hyaline, subcylindrical with broadly rounded ends, 8.4 to 18.3×4.1 to 7.7 µm (mean = 13.9×5.5 µm, n = 30). Appressoria were typically elliptic or irregular with a few lobes, dark brown, 5.9 to 9.6×4.2 to 8.1 µm (mean = 7.9 ×5.7 µm, n = 30). The morphological characteristics of the isolates matched the descriptions of Colletotrichum gloeosporioides species complex (Weir et al. 2012). The isolates were further identified by analysis of the ribosomal internal transcribed spacers (ITS), glyceraldehyde-3-phosphate dehydrogenase (GAPDH), calmodulin (CAL), actin (ACT), chitin synthase (CHS-1), glutamine synthetase (GS), and ß-tubulin 2 (TUB2) genes, amplified respectively with ITS1/ITS4, GDF/GDR, CL1C/CL2C, ACT-512F/ACT-783R, CHS-79F/CHS-345R, GSF1/GSR, and Bt2a/Bt2b (Glass and Donaldson 1995; Weir et al. 2012; White et al. 1990). The sequences were deposited in GenBank (ON492187-ON492214). Concatenated sequences of the seven genes in addition to Colletotrichum species sequences from GenBank were used to conduct a phylogenetic analysis using Maximum-Likelihood (ML) method in MEGA7. The four isolates were grouped into a clade with Colletotrichum aenigma supported by a high bootstrap value (89%), and hence, they were identified as C. aenigma based on the morphological and molecular analyses. To confirm Koch's postulates, wounded leaves of six healthy 2-month-old seedlings made by a sterile needle were inoculated with each isolate by spraying 10 ml of conidial suspensions (105 conidia/ml) on each plant, and the control plants were treated with sterile distilled water. All the treated plants were kept in a plastic box containing sterile water and incubated at 28°C in a 12 h/12 h light/dark cycle. The test was performed three times. After 7 days, typical anthracnose lesions appeared on all inoculated leaves, whereas control plants remained asymptotic. Furthermore, C. aenigma was only reisolated from the symptomatic leaves. Previous studies reported five Colletotrichum species (C. morifolium, C. fioriniae, C. brevisporum, C. karstii, and C. kahawae subsp. ciggaro) to cause this disease on mulberry in China (Tian, 1981; Xue et al. 2019). To our knowledge, this is the first report of C. aenigma causing anthracnose on mulberry in China. The finding will facilitate epidemiological studies and the development of effective control strategies for the disease.

Screening and Verification of Photosynthesis and Chloroplast-Related Genes in Mulberry by Comparative RNA-Seq and Virus-Induced Gene Silencing.

Photosynthesis is one of the most important factors in mulberry growth and production. To study the photosynthetic regulatory network of mulberry we sequenced the transcriptomes of two high-yielding (E1 and E2) and one low-yielding (H32) mulberry genotypes at two-time points (10:00 and 12:00). Re-annotation of the mulberry genome based on the transcriptome sequencing data identified 22,664 high-quality protein-coding genes with a BUSCO-assessed completeness of 93.4%. A total of 6587 differentially expressed genes (DEGs) were obtained in the transcriptome analysis. Functional annotation and enrichment revealed 142 out of 6587 genes involved in the photosynthetic pathway and chloroplast development. Moreover, 3 out of 142 genes were further examined using the VIGS technique; the leaves of MaCLA1- and MaTHIC-silenced plants were markedly yellowed or even white, and the leaves of MaPKP2-silenced plants showed a wrinkled appearance. The expression levels of the ensiled plants were reduced, and the levels of chlorophyll b and total chlorophyll were lower than those of the control plants. Co-expression analysis showed that MaCLA1 was co-expressed with CHUP1 and YSL3; MaTHIC was co-expressed with MaHSP70, MaFLN1, and MaEMB2794; MaPKP2 was mainly co-expressed with GH9B7, GH3.1, and EDA9. Protein interaction network prediction revealed that MaCLA1 was associated with RPE, TRA2, GPS1, and DXR proteins; MaTHIC was associated with TH1, PUR5, BIO2, and THI1; MaPKP2 was associated with ENOC, LOS2, and PGI1. This study offers a useful resource for further investigation of the molecular mechanisms involved in mulberry photosynthesis and preliminary insight into the regulatory network of photosynthesis.

The Ethylene Response Factor ERF5 Regulates Anthocyanin Biosynthesis in 'Zijin' Mulberry Fruits by Interacting with MYBA and F3H Genes.

Ethylene promotes ripening in fruits as well as the biosynthesis of anthocyanins in plants. However, the question of which ethylene response factors (ERFs) interact with the genes along the anthocyanin biosynthesis pathway is yet to be answered. Herein, we conduct an integrated analysis of transcriptomes and metabolome on fruits of two mulberry genotypes ('Zijin', ZJ, and 'Dashi', DS, with high and low anthocyanin abundance, respectively) at different post-flowering stages. In total, 1035 upregulated genes were identified in ZJ and DS, including MYBA in the MBW complex and anthocyanin related genes such as F3H. A KEGG analysis suggested that flavonoid biosynthesis and plant hormone signaling transduction pathways were significantly enriched in the upregulated gene list. In particular, among 103 ERF genes, the expression of ERF5 showed the most positive correlation with the anthocyanin change pattern across both genotypes and in the post-flowering stages, with a Pearson correlation coefficient (PCC) of 0.93. Electrophoresis mobility shift assay (EMSA) and luciferase assay suggested that ERF5 binds to the promoter regions of MYBA and F3H and transcriptionally activates their gene expression. We elucidated a potential mechanism by which ethylene enhances anthocyanin accumulation in mulberry fruits and highlighted the importance of the ERF5 gene in controlling the anthocyanin content in mulberry species. This knowledge could be used for engineering purposes in future mulberry breeding programs.

Effects of long-term straw retention on soil microorganisms under a rice-wheat cropping system.

The objective of this study was to investigate how straw-incorporating practices affect bacterial communities and carbon source utilization capacity under a rice-wheat rotational farming practice in central China. To clarify the effect of long-term straw incorporation in microbial abundance and carbon metabolism, a long-term field experiment was initiated in May 2005 (rice-planting season). Soil bacterial communities were revealed by high-throughput sequencing technology. After ten cycles of annual rice-wheat rotation (2005-2015), 2 M (straw incorporation) and 2 M + NPK (high straw incorporation + chemical fertilizer) treatments had significantly more bacterial phyla compared with CK (non-fertilization) and NPK (chemical fertilizer) treatments. Taxonomic analysis revealed that 2 M and NPK + 2 M treatments had a significantly greater abundance of microbial communities, especially the Gemmatimonadetes, Acidobacteria, Firmicutes, and Actinobacteria. In the NPK versus 2 M, 2 M treatment had a significantly greater abundance of Rozellomycota (P < 0.05). In the NPK + 2 M versus NPK, NPK + 2 M treatment also had significantly greater abundance of Ascomycota (P < 0.05). Principal component analysis (PCA) analysis showed that 2 M treatment was separate from other treatments. Using biolog-ECO method, the metabolic diversity and functional characteristics of microbial communities were used to indicate the ability of microorganisms to utilize carbon source. The carbon utilization ability of soil microorganisms in 2 M + NPK treatment was significantly higher than that of CK treatment (P < 0.05). The utilization ability of carboxylic acids, polymers, and other mixtures of carbon sources in 2 M treatment was higher than those of other treatments. These findings suggest that long-term straw incorporation affects the abundance and carbon utilization ability of soil microorganisms within 0-20 cm soil depths, among which, Gemmatimonadetes, Firmicutes, and Actinobacteria may play crucial roles in bacterial communities and carbon source utilization capacity.

Automated Microfluidic Instrument for Label-Free and High-Throughput Cell Separation.

Microfluidic technologies for cell separation were reported frequently in recent years. However, a compact microfluidic instrument enabling thoroughly automated cell separation is still rarely reported until today due to the difficult hybrid between the macrosized fluidic control system and the microsized microfluidic device. In this work, we propose a novel and automated microfluidic instrument to realize size-based separation of cancer cells in a label-free and high-throughput manner. Briefly, the instrument is equipped with a fully integrated microfluidic device and a set of robust fluid-driven and control units, and the instrument functions of precise fluid infusion and high-throughput cell separation are guaranteed by a flow regulatory chip and two cell separation chips which are the key components of the microfluidic device. With optimized control programs, the instrument is successfully applied to automatically sort human breast adenocarcinoma cell line MCF-7 from 5 mL of diluted human blood with a high recovery ratio of ∼85% within a rapid processing time of ∼23 min. We envision that our microfluidic instrument will be potentially useful in many biomedical applications, especially cell separation, enrichment, and concentration for the purpose of cell culture and analysis.

Inexpensive, rapid fabrication of polymer-film microfluidic autoregulatory valve for disposable microfluidics.

This work presents the fabrication of a microfluidic autoregulatory valve which is composed of several layers of thin polymer films (i.e., polyvinyl chloride (PVC), polyethylene terephthalate (PET) double-sided tape, and polydimethylsiloxane (PDMS)). Briefly, pulsed UV laser is employed to cut the microstructures of through grooves or holes in the thermoplastic polymer films, and then the polymer-film valves are precisely assembled through laminating the PDMS membranes to the thermoplastic polymer films through the roll-lamination method. The effective bonding between the PVC film and the PDMS membrane is realized using the planar seal method, and the valve is sandwiched and compressed by a home-made housing to achieve the good seal effect. Then, the flow performances of the prototype valve are examined, and constant flow autoregulation is realized under the static or dynamic test pressures. The long-term response of the valve is also studied and minimum flow-rate decrements are found over a long actuation time. The fabrication method proposed in this work is successful for the low-cost and fast prototyping of the polymer-film valve. We believe our method will also be broadly applicable for fabrication of other low-cost and disposable polymer-film microfluidic devices.

Identification and Characterization of Fusarium Species Associated with Wilt of Eleocharis dulcis (Chinese water chestnut) in China.

Fusarium wilt is one of the most important diseases of Eleocharis dulcis (Chinese water chestnut) in China. In order to characterize the pathogens responsible, 69 Fusarium isolates were collected from diseased plants in E. dulcis production areas of the Chinese provinces Anhui, Fujian, Hubei, Hunan, Jiangsu, and Zhejiang. These were then identified based on morphological and molecular characteristics. F. commune was the most common species (92.8%) and was widely distributed in the six provinces. A novel species within the Gibberella fujikuroi species complex (GFSC) was found in Hubei and Zhejiang provinces (5.8%), and an unidentified Fusarium sp. was found only in Hubei province (1.4%). Thirty F. commune isolates from different provinces and four GFSC isolates were selected for sequence analyses of the translation elongation factor 1-α (EF-1α), the mitochondrial small subunit (mtSSU) ribosomal DNA, and the nuclear ribosomal intergenic spacer region (IGS). Maximum parsimony and Bayesian analyses of the multilocus sequence data of these two species plus other taxa showed that the two species formed two distinct, well-supported clades among the three individual and combined gene genealogies. Isolates from different locations were scattered, with no evidence of geographic specialization. Pathogenicity assays showed that the two Fusarium spp., including the unidentified Fusarium sp., were pathogenic to E. dulcis 'Tuanfeng seven'. There was no relationship between the source of isolates and their pathogenicity. This is the first description of F. commune, a novel species within the GFSC, and an unidentified Fusarium sp. as causal agents of Fusarium wilt of E. dulcis in China.

Preliminary investigation of the influence of CREB1 gene polymorphisms on cognitive dysfunction in Chinese patients with major depression.

Research has increasingly focused on the role of the cyclic adenosine monophosphate (cAMP) response element binding (CREB) protein in learning and memory, particularly its role in cognitive disorders and neurodegeneration, such as Huntington's disease, Alzheimer's disease, Rubinstein-Taybi syndrome, and Coffin-Lowry syndrome. The cognitive dysfunction of patients with major depressive disorder (MDD), which is widely recognized, is not completely in accordance with depressive severity, and the dysfunction persists upon clinical remission in some patients. However, few studies have focused on the role of CREB on cognitive function in patients with MDD. This study aimed to investigate the influence of CREB1 polymorphism on cognitive function in patients with MDD. The current study comprised 113 patients with MDD. The severity of depression was measured using the 17-item Hamilton Depression Rating Scale, and cognitive function was assessed using the Stroop Neuropsychological Screening Test, verbal fluency test, and tests of immediate logical memory and visual reproduction. All subjects were genotyped with regard to CREB1 polymorphisms (rs10932201, rs2551645, rs2254137, rs6740584, and rs2551640). For the verbal fluency test, the results showed significant differences for all single-nucleotide polymorphism genotypic groups. For the Stroop color-word task, a significant difference was found only for rs6740584. No significant differences were found for the Stroop color task, the immediate logical memory test or the visual reproduction test. In conclusion, there was an effect of CREB1 polymorphism on selective attention and retrieval of long-term memory, but not on immediate memory.

High-throughput and simultaneous inertial separation of tumor cells and clusters from malignant effusions using spiral-contraction-expansion channels.

Tumor cell clusters are regarded as critical factors in cancer pathophysiology, and increasing evidence of their higher treatment resistance and metastasis compared to single tumor cells has been obtained. However, existing cell separation methods that are designed for single tumor cells cannot be used to simultaneously purify tumor cell clusters. To address this problem, we demonstrated a microfluidic approach for the high-throughput, continuous-flow ternary separation of single tumor cells, tumor cell clusters, and WBCs from clinical pleural or abdominal effusions by coupling slanted spiral channels and periodic contraction-expansion arrays. We first systematically explored the influence of particle size and flow rate on particle focusing. The separation performance indicated that 94.0% of WBCs were removed and more than 97% of MDA-MB-231 tumor cells were recovered at a high flow rate of 3500 µL/min. Moreover, more than 90% of tumor cell clusters were effectively preserved after separation. Finally, we successfully applied our device for the ternary separation of single tumor cells, tumor cell clusters, and WBCs from different malignant effusions collected from patients with metastatic cancer. Thus, our spiral-contraction-expansion device has potential as a sample pretreatment tool for the cytological diagnosis of malignant effusions.

Introducing the ratio of cross diameters of the appendix for ultrasound diagnosis of acute appendicitis can significantly increase specificity.

PURPOSE: To validate whether the introduction of the ratio of the cross diameters on the transverse section of the appendix (RATIO) ≤ 1.18 is useful for improving the ultrasound diagnosis of acute appendicitis (AA). METHODS: Data from 220 patients with AA and 110 patients with a normal appendix were retrospectively studied. The RATIO ≤ 1.18, maximal outer diameter (MOD) > 6 mm, and a combination of RATIO ≤ 1.18 and MOD > 6 mm were used for predicting AA. The area under the receiver operating characteristic curve (AUC), sensitivity, specificity, positive and negative predictive values (PPV and NPV) were calculated. RESULTS: The RATIO ≤ 1.18, MOD > 6 mm, and a combination of RATIO ≤ 1.18 and MOD > 6 mm for predicting AA showed a sensitivity of 90, 100, and 90%; specificity of 79.1, 27.3, and 88.2%; and AUC of 0.845, 0.636, and 0.891, respectively. When comparing the outcomes between MOD > 6 mm and a combination of MOD > 6 mm and RATIO ≤ 1.18, the specificity and PPV increased from 27.3 to 88.2% and 73.3 to 93.8%, respectively (all P < 0.0001). The sensitivity and NPV decreased from 100 to 90% and 100 to 81.5%, respectively (all P < 0.0001). The AUC increased from 0.636 to 0.891 (P < 0.0001). When comparing the AUC of MOD > 6 mm, and a combination of RATIO ≤ 1.18 and MOD > 6 mm for predicting AA with the AUC in a previous study, there were no significant differences between each other (all P > 0.05). CONCLUSION: Introducing the RATIO ≤ 1.18 for the evaluation of AA can improve the diagnostic performance and significantly increase specificity.

Fluoxetin suppresses tau phosphorylation and modulates the interaction between tau and tubulin in the hippocampus of CUMS rats.

Depression is considered a crucial psychiatric disease correlated with neuronal-dysfunctions induced by stress-stimuli. This study aimed to investigate effect of Fluoxetine (FL) on chronic unpredictable mild stress (CUMS) and explore the associated mechanisms. CUMS rat model was established by treating with lots of stresses. CUMS rats were administered FL, SB216763 (SB), Wortmannin (WT) alone or in combination. CUMS rats were administered 1 % sugar water to conduct sugar water consumption experiment. Acet-Tub, Tyr-Tub, tau46, p-tau-Ser199/202, p-tau-Ser396, p-tau-Ser231, expression was examined using immunohistochemical assay and western blotassay. Interaction between tau and tubulin was evaluated with immunoprecipitation assay. Double immunohistochemical assay was used to identify interaction between Nestin and Tau. The results indicated that FL treatment only increased sugar consumption of CUMS rats (P < 0.05), but also strengthened effects of SB and WT. FL significantly treatment decreased tau phosphorylation (p-tau) in hippocampal tissues of rats compared to those of rats in CUMS group (P < 0.05). FL treatment markedly decreased Acet-Tub and increased Tyr-Tub expression in hippocampal tissues of rats compared to those of rats in CUMS group (P < 0.05). The effects of FL treatment on p-tau down-regulation and tubulin modulation in hippocampal tissues were independent from PI3K and GSK-3 signaling pathways. FL treatment could also enhance proliferation and total tau of newborn neurons of CUMS rats. FL treatment strengthened interaction between tau and botulin in hippocampal tissues of CUMS rats. In conclusion, Fluoxetin suppressed phosphorylation of tau and modulated the interaction between tau and tubulin in hippocampus of adult CUMS rats.