Methyl jasmonate-induced bHLH42 mediates tissue-specific accumulation of anthocyanins via regulating flavonoid metabolism-related pathways in Caitai.

Anthocyanin is a type of plant secondary metabolite beneficial to human health. The anthocyanin content of vegetable and fruit crops signifies their nutritional quality. However, the molecular mechanism of anthocyanin accumulation, especially tissue-specific accumulation, in Caitai, as well as in other Brassica rapa varieties, remains elusive. In the present study, taking advantage of three kinds of Caitai cultivars with diverse colour traits between leaves and stems, we conducted a comparative transcriptome analysis and identified the molecular pathway of anthocyanin biosynthesis in Caitai leaves and stems, respectively. Our further investigations demonstrate that bHLH42, which is robustly induced by MeJA, closely correlates with tissue-specific accumulation of anthocyanins in Caitai; bHLH42 upregulates the expression of flavonoid/anthocyanin biosynthetic pathway genes to activate anthocyanin biosynthesis pathway, importantly, overexpression of bHLH42 significantly improves the anthocyanin content of Caitai. Our analysis convincingly suggests that bHLH42 induced by jasmonic acid signalling plays a crucial role in tissue-specific accumulation of anthocyanins in Caitai.

CsWRKY11 cooperates with CsNPR1 to regulate SA-triggered leaf de-greening and reactive oxygen species burst in cucumber.

Salicylic acid (SA) is a multi-functional phytohormone, regulating diverse processes of plant growth and development, especially triggering plant immune responses and initiating leaf senescence. However, the early SA signaling events remain elusive in most plant species apart from Arabidopsis, and even less is known about the multi-facet mechanism underlying SA-regulated processes. Here, we report the identification of a novel regulatory module in cucumber, CsNPR1-CsWRKY11, which mediates the regulation of SA-promoted leaf senescence and ROS burst. Our analyses demonstrate that under SA treatment, CsNPR1 recruits CsWRKY11 to bind to the promoter of CsWRKY11 to activate its expression, thus amplifying the primary SA signal. Then, CsWRKY11 cooperates with CsNPR1 to directly regulate the expression of both chlorophyll degradation and ROS biosynthesis related genes, thereby inducing leaf de-greening and ROS burst. Our study provides a solid line of evidence that CsNPR1 and CsWRKY11 constitute a key module in SA signaling pathway in cucumber, and gains an insight into the interconnected regulation of SA-triggered processes.

Changes in Diversity and Composition of Rhizosphere Bacterial and Fungal Community between Resistant and Susceptible Pakchoi under Plasmodiophora brassicae.

Plasmodiophora brassicae (P. brassicae) is a soil-born pathogen worldwide and can infect most cruciferous plants, which causes great yield decline and economic losses. It is not well known how microbial diversity and community composition change during P. brassicae infecting plant roots. Here, we employed a resistant and a susceptible pakchoi cultivar with and without inoculation with P. brassicae to analyze bacterial and fungal diversity using 16S rRNA V3-V4 and ITS_V1 regions, respectively. 16S rRNA V3-V4 and ITS_V1 regions were amplified and sequenced separately. Results revealed that both fungal and bacterial diversity increased, and composition was changed in the rhizosphere soil of the susceptible pakchoi compared with the resistant cultivar. In the four groups of R_mock, S_mock, R_10d, and S_10d, the most relatively abundant bacterium and fungus was Proteobacteria, accounting for 61.92%, 58.17%, 48.64%, and 50.00%, respectively, and Ascomycota, accounting for 75.11%, 63.69%, 72.10%, and 90.31%, respectively. A total of 9488 and 11,914 bacteria were observed uniquely in the rhizosphere soil of resistant and susceptible pakchoi, respectively, while only 80 and 103 fungi were observed uniquely in the correlated soil. LefSe analysis showed that 107 and 49 differentially abundant taxa were observed in bacteria and fungi. Overall, we concluded that different pakchoi cultivars affect microbial diversity and community composition, and microorganisms prefer to gather around the rhizosphere of susceptible pakchoi. These findings provide a new insight into plant-microorganism interactions.

Comparative Transcriptome Analysis between Resistant and Susceptible Pakchoi Cultivars in Response to Downy Mildew.

Downy mildew caused by the obligate parasite Hyaloperonospora brassicae is a devastating disease for Brassica species. Infection of Hyaloperonospora brassicae often leads to yellow spots on leaves, which significantly impacts quality and yield of pakchoi. In the present study, we conducted a comparative transcriptome between the resistant and susceptible pakchoi cultivars in response to Hyaloperonospora brassicae infection. A total of 1073 disease-resistance-related differentially expressed genes were identified using a Venn diagram. The Gene Ontology and Kyoto Encyclopedia of Genes and Genomes pathway analyses revealed that these genes were mainly involved in plant-pathogen interaction, plant hormone signal transduction, and other photosynthesis-related metabolic processes. Analysis of the phytohormone content revealed that salicylic acid increased significantly in the resistant material after inoculation with Hyaloperonospora brassicae, whereas the contents of jasmonic acid, abscisic acid, and 1-aminocyclopropane-1-carboxylic acid decreased. Exogenous salicylic acid treatment also significantly upregulated Hyaloperonospora brassicae-induced genes, which further confirmed a crucial role of salicylic acid during pakchoi defense against Hyaloperonospora brassicae. Based on these findings, we suggest that the salicylic-acid-mediated signal transduction contributes to the resistance of pakchoi to downy mildew, and PAL1, ICS1, NPR1, PR1, PR5, WRKY70, WRKY33, CML43, CNGC9, and CDPK15 were involved in this responsive process. Our findings evidently contribute to revealing the molecular mechanism of pakchoi defense against Hyaloperonospora brassicae.

Integrated Transcriptome and Proteome Analysis Revealed the Regulatory Mechanism of Hypocotyl Elongation in Pakchoi.

Hypocotyl length is a critical determinant for the efficiency of mechanical harvesting in pakchoi production, but the knowledge on the molecular regulation of hypocotyl growth is very limited. Here, we report a spontaneous mutant of pakchoi, lhy7.1, and identified its characteristics. We found that it has an elongated hypocotyl phenotype compared to the wild type caused by the longitudinal growth of hypocotyl cells. Different light quality treatments, transcriptome, and proteomic analyses were performed to reveal the molecular mechanisms of hypocotyl elongation. The data showed that the hypocotyl length of lhy7.1 was significantly longer than that of WT under red, blue, and white lights but there was no significant difference under dark conditions. Furthermore, we used transcriptome and label-free proteome analyses to investigate differences in gene and protein expression levels between lhy7.1 and WT. At the transcript level, 4568 differentially expressed genes (DEGs) were identified, which were mainly enriched in "plant hormone signal transduction", "photosynthesis", "photosynthesis-antenna proteins", and "carbon fixation in photosynthetic organisms" pathways. At the protein level, 1007 differentially expressed proteins (DEPs) were identified and were mainly enriched in photosynthesis-related pathways. The comprehensive transcriptome and proteome analyses revealed a regulatory network of hypocotyl elongation involving plant hormone signal transduction and photosynthesis-related pathways. The findings of this study help elucidate the regulatory mechanisms of hypocotyl elongation in lhy7.1.

Transcriptome Analyses Reveal the Role of Light in Releasing the Morphological Dormancy of Celery Seed by Integrating Plant Hormones, Sugar Metabolism and Endosperm Weakening.

Celery seed is known to be difficult to germinate due to its morphological dormancy. Light is the key signal to release morphological dormancy and promote seed germination. However, this mechanism has rarely been studied. We performed physiological, transcriptome analyses on celery seed exposed to light and dark to decipher the mechanism by which light promotes germination of celery seed. The results showed that light significantly enhanced the expression of gibberellin synthesis genes and abscisic acid degradation genes and inhibited the expression of abscisic acid synthesis genes and gibberellin degradation genes. Moreover, gibberellin synthesis inhibitor could completely inhibit the germination capacity of celery seed, indicating that gibberellin is indispensable in the process of celery seed germination. Compared with dark, light also increased the activity of α-amylase and ß-amylase and the expression of related coding genes and promoted the degradation of starch and the increase of soluble sugar content, suggesting that light enhanced the sugar metabolism of celery seed. In addition, transcriptome analysis revealed that many genes related to endosperm weakening (cell wall remodeling enzymes, extension proteins) were up-regulated under light. It was also found that light promoted the accumulation of hydrogen peroxide in the radicle, which promoted the endosperm weakening process of celery seed. Our results thus indicated that light signal may promote the release of morphological dormancy through the simultaneous action of multiple factors.

Workplace violence in primary hospitals and associated risk factors: A cross-sectional study.

AIM: To investigate the characteristics of workplace violence at primary hospitals in Southeast China and identify associated risk factors. DESIGN: A cross-sectional survey design was used for this work. METHODS: We distributed a workplace violence questionnaire among medical staff at primary hospitals in Southeast Zhejiang Province, China. The data were collected between December 2016 and December 2017. We analysed the categorical data by using the chi-square test and expressed it as frequencies. The risk factors were analysed by using multiple logistic regression analysis. RESULTS: Among the 2,560 questionnaires, 1,842 (71.9%) medical staff indicated that they had experienced workplace violence. Verbal assault was the most common type, followed by physical and sexual assault. Furthermore, gender, age, marital status, education, technical position and number of hospital beds' numbers were independent risk factors.

Impact of early postoperative activities on postoperative recovery in patients undergoing abdominal surgery: A protocol for systematic review and meta-analysis.

INTRODUCTION: A refined nursing process is utilized to formulate a detailed early postsurgical activity plan. The postsurgical activity aims to conduct focused and planned interventions to address the early postoperative activities of patients, enhance the awareness and compliance of the patients through the early postsurgical activities. Currently, in traditional clinical practice, there is no clear evidence showing the effect of initial postsurgical actions related to the rehabilitation of inpatients undertaking abdominal operations. The present study will systematically evaluate how initial postsurgical actions impact the rehabilitation of patients undertaking abdominal operation through the analysis of relevant domestic and foreign literature. OBJECTIVE: Analyze the how initial postsurgical actions impact the rehabilitation of abdominal surgery inpatients. METHODS AND ANALYSIS: The present systematic study will retrieve randomized controlled trials and case-control studies from online databases. The retrieved studies will describe the initial postsurgical activities in inpatients undergoing abdominal surgery. Accordingly, the following databases are searched for the aforementioned types of studies: Cochrane library, China National of Knowledge Infrastructure, Web of Science, PubMed database, WanFang database, and Embase database. Studies from inception to August 19, 2021 will be searched. The quality evaluation and data extraction for the studies that will satisfy the inclusion criteria will be conducted by 2 independent researchers. A meta-analysis on the postoperative indicators will be performed using RevMan 5.3.5 software.

Application of exogenous salicylic acid reduces disease severity of Plasmodiophora brassicae in pakchoi (Brassica campestris ssp. chinensis Makino).

Clubroot is one of the most serious diseases affecting Brassicaceae plants worldwide. However, there is no effective control method for clubroot. Salicylic acid (SA) is a plant hormone that plays a critical role in plant defense. In our study, we found the disease severity of a clubroot-sensitive cultivar of pakchoi, Xinxiaqing, was reduced with 0.6mM exogenous SA after the infection of P. brassicae. To investigate the mechanism of SA-reduced disease severity against clubroot, then we analyzed the plant growth, alteration of antioxidant enzyme system, and related gene expression of Xinxiaqing. Results showed that the clubroot incidence rate and disease index were decreased after being treated with 0.6 mM exogenous SA. Furthermore, plant growth, reactive oxygen species (ROS) contents, and membrane lipid peroxidation were changed. The activities of antioxidant enzymes, including superoxide dismutase (SOD), ascorbic acid-peroxidase (APX), catalase (CAT), and glutathione reductase (GR), were increased. Additionally, the production rates of malondialdehyde (MDA), hydrogen peroxide (H2O2), and superoxide anion (O2·-) were also inhibited. The expression levels of genes, encoding SOD, APX, CAT, and GR, were increased. By summering all results, we conclude that 0.6 mM SA contributes to the reduction of disease severity to clubroot by increasing the activities of antioxidant enzymes, abilities of osmotic regulation, and ROS scavenging to reduce the clubroot-induced damage in pakchoi.

Development and application of quality checklist for the prevention and control of COVID-19 in fever clinic and isolation ward of the general hospital.

To develop a quality control checklist for the prevention and control of coronavirus disease 2019 （COVID-19） in fever clinic and isolation ward of the general hospital and to assess its application. Based on the relevant prevention and control plans and technical guidelines for COVID-19，Delphi method was used to identity items for evaluation，and a quality control checklist for the prevention and control of COVID-19 in the fever clinic and isolation ward was developed in Sir Run Run Shaw Hospital. The checklists included 8 dimensions and 32 items for fever clinic，7 dimensions and 27 items for the isolation ward. The appointed inspectors conducted daily quality control for each shift with this checklist. The expert authority coefficient was 0.88，the mean of the importance of each index in the quality control table was not less than 4.8，and the coefficient of variation was not more than 0.07. During the entire February 2020，8 problems were found and rectified on-the-spot with the application of the checklist. Quality inspection rate was 100% in both isolation wards and fever clinic. The compliance rate and accuracy rate of hand hygiene were 100%; the correct rate of wearing and removing protective equipment increased from 96% to 100%. During the same period，a total of 1915 patients were admitted to the fever clinic，including 191 suspected patients （all were isolated in the hospital，3 were confirmed）. There were no medical staff infected with COVID-19，no cross infection of patients and their families in the hospital. A quality control checklist for the prevention and control of COVID-19 has been developed and applied in the isolation wards and fever clinic，which plays an important role in preventing nosocomial infection.

ML264 inhibits osteosarcoma growth and metastasis via inhibition of JAK2/STAT3 and WNT/ß-catenin signalling pathways.

Osteosarcoma, the most common bone malignancy, has a high morbidity rate and poor prognosis. Krüppel-like factor 5 (KLF5) is a key transcriptional regulator of cellular proliferation whose overexpression is observed in osteosarcoma cell lines (U2OS, 143B, MG63 and SAOS2). ML264, a small-molecule inhibitor of KLF5, exerts antiproliferative effects in colorectal cancer; however, its function in osteosarcoma remains unknown. Here, we explored the possible antitumour effects of ML264 on 143B and U2OS cell lines and murine tumour xenograft model. ML264 suppressed proliferation and clonogenic ability of osteosarcoma cells in a dose-dependent manner. Moreover, ML264 induced G0/G1 cell cycle arrest, with no influence on apoptosis, and inhibited the migratory and invasive abilities of osteosarcoma cells, as demonstrated by wound-healing and Transwell assays. Exposure to ML264 reduced the mRNA and protein levels of molecules associated with epithelial-mesenchymal transition phenotype, including N-cadherin, vimentin, Snail, matrix metalloproteinase (MMP) 9 and MMP13. Inhibition of signal transducer and activator of transcription (STAT) 3 phosphorylation and Wnt signalling was also observed. In the murine model of osteosarcoma, tumour growth was efficiently suppressed following a 10-day treatment with ML264. Collectively, our findings demonstrate the potential value of ML264 as a novel anticancer drug for osteosarcoma.

MicroRNA-25-3p regulates osteoclasts through nuclear factor I X.

Osteoporosis is a bone metabolic disease, characterized by loss of bone density leading to fractures. Its incidence increases with age and affects patient quality of life. Although osteoclasts play a significant role in osteoporosis, their underlying regulatory mechanisms remain unclear. In this study, we found that microRNA (miR)-25-3p negatively regulates osteoclast function through nuclear factor I X (NFIX). Overexpression of NFIX promoted osteoclast proliferation and increased the expression of the osteoclast differentiation and activity markers tartrate-resistant acid phosphatase and cathepsin K. MiR-25-3p transfection inhibited NFIX expression, which in turn inhibited osteoclast proliferation. Collectively, our results suggest that miR-25-3p promotes osteoclast activity by regulating the expression of NFIX. Therefore, targeting miR-25-3p in osteoclasts could be a promising strategy for treating skeletal disorders involving reduced bone formation.

Integrating long noncoding RNAs and mRNAs expression profiles of response to Plasmodiophora brassicae infection in Pakchoi (Brassica campestris ssp. chinensis Makino).

The biotrophic protist Plasmodiophora brassicae causes serious damage to Brassicaceae crops grown worldwide. However, the molecular mechanism of the Brassica rapa response remains has not been determined. Long noncoding RNA and mRNA expression profiles in response to Plasmodiophora brassicae infection were investigated using RNA-seq on the Chinese cabbage inbred line C22 infected with P. brassicae. Approximately 5,193 mRNAs were significantly differentially expressed, among which 1,345 were upregulated and 3,848 were downregulated. The GO enrichment analysis shows that most of these mRNAs are related to the defense response. Meanwhile, 114 significantly differentially expressed lncRNAs were identified, including 31 upregulated and 83 downregulated. Furthermore, a total of 2,344 interaction relationships were detected between 1,725 mRNAs and 103 lncRNAs with a correlation coefficient greater than 0.8. We also found 15 P. brassicaerelated mRNAs and 16 lncRNA interactions within the correlation network. The functional annotation showed that 15 mRNAs belong to defense response proteins (66.67%), protein phosphorylation (13.33%), root hair cell differentiation (13.33%) and regulation of salicylic acid biosynthetic process (6.67%). KEGG annotation showed that the vast majority of these genes are involved in the biosynthesis of secondary metabolism pathways and plant-pathogen interactions. These results provide a new perspective on lncRNA-mRNA network function and help to elucidate the molecular mechanism of P. brassicae infection.

Two QTLs controlling Clubroot resistance identified from Bulked Segregant Sequencing in Pakchoi (Brassica campestris ssp. chinensis Makino).

Clubroot, caused by Plasmodiophora Brassicae, is a serious soil-borne disease in worldwide. In recent years, progression of clubroot is rapid and serious in Shanghai, China. In this study, The inheritance of clubroot resistance (CR) were determined in pakchoi using F2 segregation population that were developed by crossing highly resistant line 'CR38' and susceptible line 'CS22'. Two novel QTLs, qBrCR38-1 and qBrCR38-2, was identified by BSA-seq (Bulked Segregant Sequencing) resistant to P. brassicae physiological race 7. Two significant peak qBrCR38-1 and qBrCR38-2 were observed by three statistical methods between interval of 19.7-20.6 Mb in chromosome A07 and 20.0-20.6 Mb in chromosome A08, respectively. In addition, Polymorphic SNPs identified within target regions were converted to kompetitive allele-specific PCR (KASP) assays. In target regions of qBrCR38-1 and qBrCR38-2, there were twenty SNP sites identified, eleven KASP markers of which are significantly associated to CR (P < 0.05). Seven candidate genes were identified and found to be involved in disease resistance (TIR-NBS-LRR proteins), defense responses of bacterium and fungi and biotic/abiotic stress response in the target regions harboring the two QTLs. Two novel QTLs and candidate genes identified from the present study provide insights into the genetic mechanism of CR in B.rapa, and the associated SNPs can be effectively used for marker-assisted breeding.

Effects of low light on photosynthetic properties, antioxidant enzyme activity, and anthocyanin accumulation in purple pak-choi (Brassica campestris ssp. Chinensis Makino).

Anthocyanins are secondary metabolites that contribute to red, blue, and purple colors in plants and are affected by light, but the effects of low light on the physiological responses of purple pak-choi plant leaves are still unclear. In this study, purple pak-choi seedlings were exposed to low light by shading with white gauze and black shading in a phytotron. The responses in terms of photosynthetic properties, carbohydrate metabolism, antioxidant enzyme activity, anthocyanin biosynthetic enzyme activity, and the relative chlorophyll and anthocyanin content of leaves were measured. The results showed that chlorophyll b, intracellular CO2 content, stomatal conductance and antioxidant activities of guaiacol peroxidase, catalase and superoxide dismutase transiently increased in the shade treatments at 5 d. The malondialdehyde content also increased under low light stress, which damages plant cells. With the extension of shading time (at 15 d), the relative chlorophyll a, anthocyanin and soluble protein contents, net photosynthetic rate, transpiration rate, stomata conductance, antioxidant enzyme activities, and activities of four anthocyanin biosynthetic enzymes decreased significantly. Thus, at the early stage of low light treatment, the chlorophyll b content increased to improve photosynthesis. When the low light treatment was extended, antioxidant enzyme activity and the activity of anthocyanin biosynthesis enzymes were inhibited, causing the purple pak-choi seedlings to fade from purple to green. This study provides valuable information for further deciphering genetic mechanisms and improving agronomic traits in purple pak-choi under optimal light requirements.