Phylogenomic insights into the origin and evolutionary history of evergreen broadleaved forests in East Asia under Cenozoic climate change.

The evergreen versus deciduous leaf habit is an important functional trait for adaptation of forest trees and has been hypothesized to be related to the evolutionary processes of the component species under paleoclimatic change, and potentially reflected in the dynamic history of evergreen broadleaved forests (EBLFs) in East Asia. However, knowledge about the shift of evergreen versus deciduous leaf with the impact of paleoclimatic change using genomic data remains rare. Here, we focus on the Litsea complex (Lauraceae), a key lineage with dominant species of EBLFs, to gain insights into how evergreen versus deciduous trait shifted, providing insights into the origin and historical dynamics of EBLFs in East Asia under Cenozoic climate change. We reconstructed a robust phylogeny of the Litsea complex using genome-wide single-nucleotide variants (SNVs) with eight clades resolved. Fossil-calibrated analyses, diversification rate shifts, ancestral habit, ecological niche modelling and climate niche reconstruction were employed to estimate its origin and diversification pattern. Taking into account studies on other plant lineages dominating EBLFs of East Asia, it was revealed that the prototype of EBLFs in East Asia probably emerged in the Early Eocene (55-50 million years ago [Ma]), facilitated by the greenhouse warming. As a response to the cooling and drying climate in the Middle to Late Eocene (48-38 Ma), deciduous habits were evolved in the dominant lineages of the EBLFs in East Asia. Up to the Early Miocene (23 Ma), the prevailing of East Asian monsoon increased the extreme seasonal precipitation and accelerated the emergence of evergreen habits of the dominant lineages, and ultimately shaped the vegetation resembling that of today.

Assessing candidate DNA barcodes for Chinese and internationally traded timber species.

Accurate identification of species from timber is an essential step to help control illegal logging and forest loss. However, current approaches to timber identification based on morphological and anatomical characteristics have limited species resolution. DNA barcoding is a proven tool for plant species identification, but there is a need to build reliable reference data across broad taxonomic and spatial scales. Here, we construct a species barcoding library consisting of 1550 taxonomically diverse timber species from 656 genera and 124 families, representing a comprehensive genetic reference data set for Chinese timber species and international commercial traded timber species, using four barcodes (rbcL, matK, trnH-psbA, and ITS2). The ITS2 fragment was found to be the most efficient locus for Chinese timber species identification among the four barcodes tested, both at the species and genus level, despite its low recovery rate. Nevertheless, the barcode combination matK+trnH-psbA+ITS2 was required as a complementary barcode to distinguish closely related species in complex data sets involving internationally traded timber species. Comparative analyses of family-level discrimination and species/genus ratios indicated that the inclusion of closely related species is an important factor affecting the resolution ability of barcodes for timber species verification. Our study indicates that although nuclear ITS2 is the most efficient single barcode for timber species authentication in China, complementary combinations like matK+trnH-psbA+ITS2 are required to provide broader discrimination power. These newly-generated sequences enrich the existing publicly available databases, especially for tropical and subtropical evergreen timber trees and this current timber species barcode reference library can serve as an important genetic resource for forestry monitoring, illegal logging prosecution and biodiversity projects.

Specialised metabolites as chemotaxonomic markers of Coptosapelta diffusa, supporting its delimitation as sisterhood phylogenetic relationships with Rubioideae.

From the aerial extracts of Coptosapelta diffusa (Champ. ex Benth.) Steenis, twenty-one compounds were isolated and identified by means of column chromatography and NMR and MS techniques, respectively. Amongst, ten ones were determined to be undescribed compounds including six seco-iridoid glucosides (1-6), 2-(hydroxymethyl)-1,2,3,4-tetrahydroanthracene-9,10-dione (7) and three guaiane-type sesquiterpenes (15-17). Compounds 7, 8 and 9 exhibited inhibitory activities against Staphylococcus aureus ATCC25923 with MIC of 8, 4 and 8 µg/mL. The use of 1-6 (iridoids), 7-14 (anthraquinones) and 15-17 (sesquiterpenes) as chemotaxonomic markers for this species was evidenced. Structurally, 7-14 are similar to those anthraquinones isolated from other species of the family Rubiaceae, confirming their close phylogenetic relationship. Whereas, these iridoids and sesquiterpenes with unique structures provided chemotaxonomic evidence to support the genus Coptosapelta (the tribe Coptosapelteae) as a sister of the subfamily Rubioideae. These results contrast with the general producing tendency of indole alkaloids by the species of the subfamily Cinchonoideae, and merit chemotaxonomic significance for the delimitation of Coptosapelta.

Deafness enhances perceptual span size in Chinese reading: Evidence from a gaze-contingent moving-window paradigm.

Using a gaze-contingent moving-window paradigm, we investigated whether/how deafness affects perceptual processing in Chinese reading. Besides the manipulation of window size, word length of sentences used in the experiment was also manipulated to check whether deafness enhanced the word length effect on perceptual span. Significant interactions of window constraints and deafness and a three-way interaction were observed on reading rate. Smaller effects of window constraints for deaf Chinese readers and nonreliable three-way interactions were observed on forward saccade length. This suggests that deaf Chinese readers exhibit a larger perceptual span, and word length affected the span from which information was acquired for comprehension whereas both deafness and word length might have little impact on the span from which information is acquired for oculomotor targeting during natural reading of Chinese.

Characterizing the Role of TaWRKY13 in Salt Tolerance.

The WRKY transcription factor superfamily is known to participate in plant growth and stress response. However, the role of this family in wheat (Triticum aestivum L.) is largely unknown. Here, a salt-induced gene TaWRKY13 was identified in an RNA-Seq data set from salt-treated wheat. The results of RT-qPCR analysis showed that TaWRKY13 was significantly induced in NaCl-treated wheat and reached an expression level of about 22-fold of the untreated wheat. Then, a further functional identification was performed in both Arabidopsis thaliana and Oryza sativa L. Subcellular localization analysis indicated that TaWRKY13 is a nuclear-localized protein. Moreover, various stress-related regulatory elements were predicted in the promoter. Expression pattern analysis revealed that TaWRKY13 can also be induced by polyethylene glycol (PEG), exogenous abscisic acid (ABA), and cold stress. After NaCl treatment, overexpressed Arabidopsis lines of TaWRKY13 have a longer root and a larger root surface area than the control (Columbia-0). Furthermore, TaWRKY13 overexpression rice lines exhibited salt tolerance compared with the control, as evidenced by increased proline (Pro) and decreased malondialdehyde (MDA) contents under salt treatment. The roots of overexpression lines were also more developed. These results demonstrate that TaWRKY13 plays a positive role in salt stress.

Interspecies introgressive hybridization in spiny frogs Quasipaa (Family Dicroglossidae) revealed by analyses on multiple mitochondrial and nuclear genes.

Introgression may lead to discordant patterns of variation among loci and traits. For example, previous phylogeographic studies on the genus Quasipaa detected signs of genetic introgression from genetically and morphologically divergent Quasipaa shini or Quasipaa spinosa. In this study, we used mitochondrial and nuclear DNA sequence data to verify the widespread introgressive hybridization in the closely related species of the genus Quasipaa, evaluate the level of genetic diversity, and reveal the formation mechanism of introgressive hybridization. In Longsheng, Guangxi Province, signs of asymmetrical nuclear introgression were detected between Quasipaa boulengeri and Q. shini. Unidirectional mitochondrial introgression was revealed from Q. spinosa to Q. shini. By contrast, bidirectional mitochondrial gene introgression was detected between Q. spinosa and Q. shini in Lushan, Jiangxi Province. Our study also detected ancient hybridizations between a female Q. spinosa and a male Q. jiulongensis in Zhejiang Province. Analyses on mitochondrial and nuclear genes verified three candidate cryptic species in Q. spinosa, and a cryptic species may also exist in Q. boulengeri. However, no evidence of introgressive hybridization was found between Q. spinosa and Q. boulengeri. Quasipaa exilispinosa from all the sampling localities appeared to be deeply divergent from other communities. Our results suggest widespread introgressive hybridization in closely related species of Quasipaa and provide a fundamental basis for illumination of the forming mechanism of introgressive hybridization, classification of species, and biodiversity assessment in Quasipaa.

DNA barcoding evaluation and implications for phylogenetic relationships in Lauraceae from China.

Lauraceae are an important component of tropical and subtropical forests and have major ecological and economic significance. Owing to lack of clear-cut morphological differences between genera and species, this family is an ideal case for testing the efficacy of DNA barcoding in the identification and discrimination of species and genera. In this study, we evaluated five widely recommended plant DNA barcode loci matK, rbcL, trnH-psbA, ITS2 and the entire ITS region for 409 individuals representing 133 species, 12 genera from China. We tested the ability of DNA barcoding to distinguish species and as an alternative tool for correcting species misidentification. We also used the rbcL+matK+trnH-psbA+ITS loci to investigate the phylogenetic relationships of the species examined. Among the gene regions and their combinations, ITS was the most efficient for identifying species (57.5%) and genera (70%). DNA barcoding also had a positive role for correcting species misidentification (10.8%). Furthermore, based on the results of the phylogenetic analyses, Chinese Lauraceae species formed three supported monophyletic clades, with the Cryptocarya group strongly supported (PP = 1.00, BS = 100%) and the clade including the Persea group, Laureae and Cinnamomum also receiving strong support (PP = 1.00, BS = 98%), whereas the Caryodaphnopsis-Neocinnamomum received only moderate support (PP = 1.00 and BS = 85%). This study indicates that molecular barcoding can assist in screening difficult to identify families like Lauraceae, detecting errors of species identification, as well as helping to reconstruct phylogenetic relationships. DNA barcoding can thus help with large-scale biodiversity inventories and rare species conservation by improving accuracy, as well as reducing time and costs associated with species identification.

Molecular epidemiology and evolution of human enterovirus 71 and hand, foot and mouth disease.

Human enterovirus 71(EV71), one of the major pathogens of the hand, foot and mouth disease (HFMD), causes skin rashes in palms, feet and mouth ulcers and complication in the central nervous system such as aseptic meningitis and acute flaccid paralysis that may lead to death. EV71 infection has been reported to be associated with many outbreaks of HFMD worldwide, especially the great outbreaks that occurred in the Asia-Pacific region and caused numerous death since 1997. The studies of molecular epidemiology and evolution of EV71 are important for the prevention and control of HFMD since no vaccines and antiviral drugs have been developed except symptomatic treatment for HFMD. In this review, we summarize genotype classification, temporal and spatial distribution, evolutionary characteristics and modes of EV71 as well as typical EV71 epidemics. Further studies on EV71 and HFMD may lead to better understanding of pathological mechanisms of EV71, development of antiviral drugs and prevention and control of HFMD.

Effects of awareness on numerosity adaptation.

Numerosity perception is a process involving several stages of visual processing. This study investigated whether distinct mechanisms exist in numerosity adaptation under different awareness conditions to characterize how numerosity perception occurs at each stage. The status of awareness was controlled by masking conditions, in which monoptic and dichoptic masking were proposed to influence different levels of processing. Numerosity adaptation showed significant aftereffects when the participants were aware (monoptic masking) and unaware (dichoptic masking) of adaptors. The interocular transfer for numerosity adaptation was distinct under the different awareness conditions. Adaptation was primarily binocular when participants were aware of stimuli and was purely monocular when participants were unaware of adaptors. Moreover, numerosity adaptation was significantly reduced when the adaptor dots were clustered into chunks with awareness, whereas clustering had no effect on unaware adaptation. These results show that distinct mechanisms exist in numerosity processing under different awareness conditions. It is suggested that awareness is crucial to numerosity cognition. With awareness, grouping (by clustering) influences numerosity coding through altered object representations, which involves higher-level cognitive processing.

[Advances on gene-based association analysis].

Genome-wide association studies (GWAS) have been widely used for hunting the susceptibility genes for common diseases in the past years; however, the abundant information for the disease mechanism based on the GWAS data has not been fully mined. Recently, some researchers focused on the biological network and pathway analysis for the GWAS data to explore the potential disease mechanism. Since genes are the basic units for the biological network and pathway, the genetic effects from all or part of single nucleotide polymorphisms (SNPs) in the genes should be integrated into genetic scores, which are so-called "gene-based association analysis". Gene-based association analysis takes into account some important factors such as genetic effects of the SNPs, the number of the SNPs in the genes and the linkage disequilibrium structure of the SNPs. In this review, we will focus on the progress, principle and application of gene-based association analysis.

Genome-wide association study for vitiligo identifies susceptibility loci at 6q27 and the MHC.

We conducted a genome-wide association study of generalized vitiligo in the Chinese Han population by genotyping 1,117 cases and 1,429 controls. The 34 most promising SNPs were carried forward for replication in samples from individuals of the Chinese Han (5,910 cases and 9,916 controls) and Chinese Uygur (713 cases and 824 controls) populations. We identified two independent association signals within the major histocompatibility complex (MHC) region (rs11966200, Pcombined=1.48x10(-48), OR=1.90; rs9468925, Pcombined=2.21x10(-33), OR=0.74). Further analyses suggested that the strong association at rs11966200 might reflect the reported association of the HLA-A*3001, HLA-B*1302, HLA-C*0602 and HLA-DRB1*0701 alleles and that the association at rs9468925 might represent a previously unknown HLA susceptibility allele. We also identified one previously undescribed risk locus at 6q27 (rs2236313, Pcombined=9.72x10(-17), OR=1.20), which contains three genes: RNASET2, FGFR1OP and CCR6. Our study provides new insights into the genetic basis of vitiligo.

[Oxaliplatin combined with capecitabine as first-line chemotherapy for patients with advanced gastric cancer].

BACKGROUND AND OBJECTIVE: Combination therapy of oxaliplatin and capecitabine has certain effects on advanced gastric cancer (AGC). This study was to investigate the efficacy and safety of oxaliplatin in combination with capecitabine as first-line chemotherapy for AGC patients. METHODS: Thirty-three chemotherapy-naive patients with AGC were entered into this study. They received 2 h intravenous infusion of oxaliplatin 130 mg/m2 on day 1 and oral administration of capecitabine 2000 mg/m2, given in two daily doses, on days 1-14 (XELOX regimen). The regimen was repeated every 21 days. A maximum of eight cycles were given. RESULTS: Thirty-three patients completed 159 cycles of chemotherapy with a median number of five cycles. Thirty-one patients were evaluable for efficacy. The response rate was 54.8% [95% confidence interval (CI): 37.3%-72.3%], with one complete response (3.2%), 16 partial responses (51.6%), eight stable diseases (25.8%), and six progressions (19.4%). At a mean follow-up of 10.5 months, the median time to progression and overall survival were 5.9 (95% CI: 4.7-7.1) and 10.4 months (95% CI: 7.9-12.9), respectively. The most common adverse events were myelosuppression, peripheral neuropathy, diarrhea, nausea/vomiting, and hand-foot syndrome. CONCLUSION: XELOX is an effective and well-tolerated first-line chemotherapy regimen for patients with AGC.

[Low dose-intensity docetaxel monotherapy for patients with anthracycline-resistant metastatic breast cancer in poor physical status].

OBJECTIVE: To evaluate the efficacy and safety of a weekly schedule of low dose-intensity docetaxel monochemotherapy for patients with anthracycline-resistant metastatic breast cancer (MBC) in poor physical status. METHODS: Thirty MBC patients who were previously exposed to anthracycline treatment received docetaxel alone at a dose of 30 mg/m2 on D1, D8 and D15, repeated every 4 weeks for a maximum of 6 cycles. RESULTS: Of the 30 evaluable patients, 2 (6.7%) achieved a complete response, and 9 (30.0%) a partial response, with an overall objective response rate of 36.7% (95% CI: 20.5%-53.9%). The most common adverse event was hematologic toxicity. After an average follow-up of 15.0 months, the median time to progression (TTP) was 8. 5 months and the median overall survival (OS) had not reached yet at the end of follow-up. CONCLUSION: The weekly low dose-intensity docetaxel monochemotherapy is effective and well-tolerated in patients with anthracycline-resistant metastatic breast cancer in poor physical status.

[Enhancing effect of antisense oligonucleotide targeting bFGF on apoptosis in hepatoma cells in vitro].

OBJECTIVE: To investigate the cell cycle changes of hepatoma cells and the effect of antisense oligonucleotide targeting bFGF on apoptosis in the hepatoma cells. METHODS: The oligodeoxynucleotides were transfected with Lipofectin into hepatoma HepG2 cells. Inhibition of bFGF protein expression was assessed by confocal laser scanning microscopy and Western blot under the best condition of transfection of antisense oligonucleotide targeting bFGF, and the apoptosis in those cells was determined by flow cytometry. HepG2 cells were cultured in 24-well culture dish. The cultured cells were divided into 3 groups: group 1, the normal control group without any treatment; group 2, transfected with antisense oligonucleotide targeting bFGF; group 3, transfected with scrambled sequence targeting bFGF. RESULTS: The results from confocal microscopy and Western blot showed an inhibition of expression of bFGF at different levels under the best condition of transfection with antisense oligonucleotide targeting bFGF. The treatment with antisense oligonucleotide of bFGF not only reduced the expression of bFGF revealed by confocal microscopy and Western blotting, but also increased the apoptosis in HepG 2 cells (P < 0. 01). CONCLUSION: Treatment with antisense oligonucleotide of bFGF inhibits expression of bFGF protein and increase apoptosis. bFGF may take part in apoptosis regulation of hepatoma cells and may be used as a target in the treatment of hepatocellular carcinoma.

Biweekly oxaliplatin in combination with continuous infusional 5-fluorouracil and leucovorin (modified FOLFOX-4 regimen) as first-line chemotherapy for elderly patients with advanced gastric cancer.

OBJECTIVE: The aim of the study was to assess the toxicity and the clinical activity of biweekly oxaliplatin (OXA) in combination with continuous infusional 5-fluorouracil (5-FU) and leucovorin (LV) administered every 2 weeks (modified FOLFOX-4 regimen) in elderly patients with advanced gastric cancer (AGC). PATIENTS AND METHODS: A total of 44 previously untreated AGC patients aged 65 or older were treated with OXA 85 mg m-2 on day 1, LV 200 mg m-2 as a 2-hour infusion followed by a 22-hour infusion of 5-FU 1000 mg m using a multichannel programmable pump, repeated for 2 consecutive days every 2 weeks. RESULTS: All patients were assessable for toxicity and 40 patients for response. Median age was 69 years (65-83). The response rate according to Response Evaluation Criteria in Solid Tumors (RECIST) criteria was 52.5% (95% confidence interval: 44.6%-68.0%) with 3 complete responses, 18 partial responses, 11 stable diseases, and 8 progressions. Median time to progression was 6.5 months and median overall survival was 10.0 months. Toxicity was generally mild. Grade 3 hematologic toxicities of neutropenia, anemia, and thrombocytopenia were in 6.8%, 2.3%, and 4.5% of the patients, respectively. No grade 4 hematologic toxicities occurred. Grade 1 peripheral neuropathy was a common event (34.1%), whereas grade 3 peripheral neuropathy was recorded in only 1 (2.3%) patient. An acute hypersensitivity reaction was observed in 1 patient during administration. CONCLUSION: The modified FOLFOX-4 regimen is an active and well-tolerated chemotherapy for elderly patients aged > or =65 years with AGC. OXA may occasionally cause mild hypersensitivity.

[Antisense oligonucleotide targeting endostatin enhances hematopoiesis reconstitution in BMT mice].

OBJECTIVE: To explore the effect of antisense oligonucleotide targeting endostatin (endostatin-ASON) transfecting bone marrow stromal cells ( BMSC) on hematopoiesis reconstitution in BMT mice. METHODS: Inhibition of endostatin / VCAM-1 protein and mRNA expression was investigated by transfection of antisense oligonucleotide targeting endostatin with confocal microscopy, Western blot and RT-PCR. Bone marrow stromal cells were cultured and divided into 4 groups: group (1) without any treatment; group (2) BMT only; group (3) BMT + endostatin-ASON transfection; group (4) BMT + endostatin scrambled sequence transfection. RESULTS: (1) Endostatin-ASON was successfully introduced into BMSC in vitro, and the transfecting rate was 86% ;(2) After Endostatin-ASON transfected into BMSC, the expression of Endostatin mRNA and its protein on the BMSC was signficantly inhibited at different time point after BMT [the grey value of Endostatin was (0.09 +/- 0.03) - (1.44 +/- 1.19) and (0.02 + 0.02) - (0.14 +/- 0.05), respectively] (P < 0.01 and P < 0.05); (3) Transfecting with Endostatin-ASON effectively promoted the expression of VCAM-1 mRNA and its protein on the BMSC [the gray value of VCAM-1 was (1.60 +/- 0. 92) - (8.05 +/- 0.87) and (0.07 +/- 0.02) - (0.67 +/- 0.09) , respectively] (P <0.01 and P <0.05) ; (4) There was no effects of transfecting Endostatin scrambled sequence on the expression of Endostatin and VCAM-1 on the BMSC (P > 0.05). CONCLUSION: Endostatin-ASON could inhibit Endostatin expression and enhance VCAM-1 expression in BMSC after syngeneic-BMT in mice, which might be one of the mechanisms underlying the endostatin-ASON accelerating hematopoiesis reconstitution after allogeneic-BMT.

Gene-expression profiles in gastric epithelial cells stimulated with spiral and coccoid Helicobacter pylori.

Human gastric epithelial immortalized GES-1 cells were infected with spiral and coccoid Helicobacter pylori. Scanning electron microscopy was used to determine the ability of the two forms of H. pylori to adhere to GES-1 cells. GES-1 cell apoptosis induced by coccoid and spiral H. pylori was analysed using flow cytometry. A cDNA microarray for 22,000 human genes was used to identify the gene-expression differences in GES-1 cells infected with the two forms of H. pylori, and the gene expression identified by the cDNA microarray was confirmed by RT-PCR. Scanning electron microscope observation showed that both coccoid and spiral bacteria can adhere to GES-1 cells. After 4 h infection, apoptosis induction was 27.4% for spiral-form infection and 10.2% for coccoid-form infection. Of 268 differentially expressed genes identified by cDNA microarray, 166 showed higher expression with the spiral H. pylori infection than with the coccoid H. pylori infection. To the best of the authors' knowledge, this is the first report that GES-1 cells infected with spiral H. pylori have higher expression of cxcl10, ccl11, ccl5, groalpha, TLR5, ATF3, fos, fosl2, gadd45a and myc. The cells infected with coccoid H. pylori had higher expression of survivin. The global profile of gene expression in GES-1 cells infected with coccoid and spiral H. pylori is described for the first time.

[A PCR-RFLP analysis to HLA-B gene among Chinese northern Han populations].

OBJECTIVE: To set up the method for analyzing HLA-B gene polymorphism with PCR-RFLP, and to gain population data among northern Chinese Hans of HLA-B's restricted fragments after NlaIII digestion, and to achieve application in forensic medicine practice. METHODS: Sample DNA was extracted by the phenol/chloroform extraction method, 943 bp-long fragments containing HLA-B exon 2 and 3 were got by PCR. The endonuclease NlaIII was applied to cut the PCR products into polymorphic fragments shorter than 943bp, then PAGE and silver staining were used to detect the digestion results, finally the digestion sites were assured by DNA sequencing. RESULTS: Along 943bp-long PCR products, 14 length-different fragments, 20 kinds of fragment combinations were got and 6 cutting site were observed after NlaIII digestion. CONCLUSION: HLA-B gene was highly polymorphic among Chinese northern Hans. Even with only one endonuclease, 14 restricted fragments were got and the PIC was great. Such a HLA-B PCR-RFLP analysis will have values in forensic medicine applications.

Identification of androgen-responsive element ARE and Sp1 element in the maspin promoter.

Maspin is a serine protease inhibitor (serpin) with tumor-suppressing function in mammary gland. It is down-regulated in primary prostate cancer cells and lost in metastatic cells. To better understand the transcriptional regulation of maspin gene, the 860bp (-765 approximately +95) of its promoter sequence was amplified by PCR from the human genomic DNA. Then this 860bp sequence and a series of deletions from 5' and 3' ends were inserted into the upstream of luciferase reporter gene respectively. Results from dual luciferase reporter assay and electrophoretic mobility shift assay indicated that there were a negative androgen-responsive element (ARE) in the region of -277 to -262 and a positive Sp1 element in the region of +14 to +35, respectively. In addition, androgen receptor (AR) can recognize and bind to the ARE element, and then inhibit the activity of maspin promoter.