Comparative Analysis of Transcriptomics and Metabolomics Reveals Defense Mechanisms in Melon Cultivars against Pseudoperonospora cubensis Infection.

Melon (Cucumis melo L.) represents an agriculturally significant horticultural crop that is widely grown for its flavorful fruits. Downy mildew (DM), a pervasive foliar disease, poses a significant threat to global melon production. Although several quantitative trait loci related to DM resistance have been identified, the comprehensive genetic underpinnings of this resistance remain largely uncharted. In this study, we utilized integrative transcriptomics and metabolomics approaches to identify potential resistance-associated genes and delineate the strategies involved in the defense against DM in two melon cultivars: the resistant 'PI442177' ('K10-1') and the susceptible 'Huangdanzi' ('K10-9'), post-P. cubensis infection. Even in the absence of the pathogen, there were distinctive differentially expressed genes (DEGs) between 'K10-1' and 'K10-9'. When P. cubensis was infected, certain genes, including flavin-containing monooxygenase (FMO), receptor-like protein kinase FERONIA (FER), and the HD-ZIP transcription factor member, AtHB7, displayed pronounced expression differences between the cultivars. Notably, our data suggest that following P. cubensis infection, both cultivars suppressed flavonoid biosynthesis via the down-regulation of associated genes whilst concurrently promoting lignin production. The complex interplay of transcriptomic and metabolic responses elucidated by this study provides foundational insights into melon's defense mechanisms against DM. The robust resilience of 'K10-1' to DM is attributed to the synergistic interaction of its inherent transcriptomic and metabolic reactions.

Fine mapping of a novel QTL DM9.1 conferring downy mildew resistance in melon.

Downy mildew (DM) is a major foliar disease globally causing great economic loss in melon production. Utilizing disease-resistant cultivars is the most efficient approach for disease control, while discovery of disease-resistant genes is crucial for the success of DM-resistant breeding. To address this problem, two F2 populations were constructed using the DM-resistant accession PI 442177 in this study, and QTLs conferring DM resistance were mapped using linkage map and QTL-seq analysis, respectively. A high-density genetic map with the length of 1096.7 cM and density of 0.7 cM was generated by using the genotyping-by-sequencing data of a F2 population. A major QTL DM9.1 with the phenotypic variance explained proportion of 24.3-37.7% was consistently detected at the early, middle, and late growth stages using the genetic map. QTL-seq analyses on the two F2 populations also validated the presence of DM9.1. Kompetitive Allele-Specific PCR (KASP) assay was further carried out to fine map DM9.1 into 1.0 Mb interval. A KASP marker co-segregating with DM9.1 was successfully developed. These results not only provided valuable information for DM-resistant gene cloning, but also offered useful markers for melon DM-resistant breeding programs.

NAC Transcription Factor TwNAC01 Positively Regulates Drought Stress Responses in Arabidopsis and Triticale.

The NAC transcription factors play important roles in regulating plant growth, development, and senescence, and responding to biotic and abiotic stressors in plants. A novel coding sequence (1,059 bp) was cloned from hexaploid triticale in this study. The putative protein (352 amino acids) encoded by this sequence was over 95% similar to the amino acid sequence of a NAC protein from Aegilops tauschii (XP020161331), and it formed a clade with Ae. tauschii, durum wheat, and barley. The putative protein contained a conserved nature actomyosin (NAM) domain (129 consecutive amino acids) between the 20th and 148th amino acids at the N-terminus and three transcription activation regions at the C-terminus. The novel gene was identified as a triticale NAC gene localized in the nucleus and designated as TwNAC01 (GenBank accession MG736919). The expression levels of TwNAC01 were the highest in roots, followed by leaves and stems when triticale lines were exposed to drought, polyethylene glycol 6,000 (PEG6000), NaCl, cold, methyl jasmonate (MeJA), and abscisic acid (ABA). Transgenic Arabidopsis thaliana overexpressing TwNAC01 had significantly lower leaf water loss rates and longer roots than wild-type (WT) A. thaliana. Virus-induced silencing of the TwNAC01 gene in triticale delayed root development and decreased length of primary root. Under drought stress, leaves of TwNAC01-silenced triticale had higher levels of malondialdehyde (MDA) and hydrogen peroxide (H2O2), but lower relative water content (RWC), net photosynthetic rate, stomatal conductance, intercellular CO2 concentration, and transpiration rate than the leaves of the WT. Gene overexpression and silencing experiments suggested that TwNAC01 improves plant stress tolerance by increasing root length, regulating the water content of plant leaves by reducing MDA and H2O2 content, and adjusting respiration rate. The results suggest that TwNAC01 is a novel NAC transcription factor gene that can be exploited for triticale and cereal improvement.

Administration Timing and Efficacy of Tocilizumab in Patients With COVID-19 and Elevated IL-6.

BACKGROUND: Tocilizumab (TCZ), an interleukin-6 receptor antibody, has previously been used for treating patients with the coronavirus disease 2019 (COVID-19), but there is a lack of data regarding the administration timing of TCZ. OBJECTIVES: This study aimed to evaluate the timing and efficacy of TCZ in the treatment of patients with COVID-19. METHODS: Laboratory-confirmed patients with COVID-19 with an elevated interleukin-6 (IL-6) level (>10 pg/ml) were offered TCZ intravenously for compassionate use. Clinical characteristics, laboratory tests, and chest imaging before and after the administration of TCZ were retrospectively analyzed. RESULTS: A total of 58 consecutive patients who met the inclusion criteria and with no compliance to the exclusion criteria were included. Of these 58 patients, 39 patients received TCZ treatment, and 19 patients who declined TCZ treatment were used as the control cohort. In the TCZ-treatment group, 6 patients (15.4%) were in mild condition, 16 (41.0%) were in severe condition, and 17 (43.6%) were in critical condition. After TCZ treatment, the condition of 27 patients (69.2%) improved and 12 (30.8%) died. Compared with the improvement group, patients in the death group had higher baseline levels of IL-6 (P = 0.0191) and procalcitonin (PCT) (P = 0.0003) and lower lymphocyte percentage (LYM) (P = 0.0059). Patients receiving TCZ treatment had better prognoses than those without TCZ treatment (P = 0.0273). Furthermore, patients with a baseline IL-6 level of ≥100 pg/ml in the TCZ-treatment group had poorer clinical outcomes than those with an IL-6 level of <100 pg/ml (P = 0.0051). CONCLUSION: The administration of TCZ in an early stage of cytokine storm (IL-6 level < 100 pg/ml) may effectively improve the clinical prognosis of patients with COVID-19 by blocking the IL-6 signal pathway.

Analysis of viral load in different specimen types and serum antibody levels of COVID-19 patients.

BACKGROUND: COVID-19 has caused a global pandemic and the death toll is increasing. However, there is no definitive information regarding the type of clinical specimens that is the best for SARS-CoV-2 detection, the antibody levels in patients with different duration of disease, and the relationship between antibody level and viral load. METHODS: Nasopharyngeal swabs, anal swabs, saliva, blood, and urine specimens were collected from patients with a course of disease ranging from 7 to 69 days. Viral load in different specimen types was measured using droplet digital PCR (ddPCR). Meanwhile, anti-nucleocapsid protein (anti-N) IgM and IgG antibodies and anti-spike protein receptor-binding domain (anti-S-RBD) IgG antibody in all serum samples were tested using ELISA. RESULTS: The positive detection rate in nasopharyngeal swab was the highest (54.05%), followed by anal swab (24.32%), and the positive detection rate in saliva, blood, and urine was 16.22%, 10.81%, and 5.41%, respectively. However, some patients with negative nasopharyngeal swabs had other specimens tested positive. There was no significant correlation between antibody level and days after symptoms onset or viral load. CONCLUSIONS: Other specimens could be positive in patients with negative nasopharyngeal swabs, suggesting that for patients in the recovery period, specimens other than nasopharyngeal swabs should also be tested to avoid false negative results, and anal swabs are recommended. The antibody level had no correlation with days after symptoms onset or the viral load of nasopharyngeal swabs, suggesting that the antibody level may also be affected by other factors.

Comparative metabolomics shows the metabolic profiles fluctuate in multi-drug resistant Escherichia coli strains.

In this study, two susceptible strains and two multi-drug resistant clinical Escherichia coli strains were obtained by Kirby-Bauer method, and then a GC-MS-based metabolomics method was used to compare the differential expression of metabolites between two drug sensitive (CK1 and CK2) and two multidrug-resistant (MDR1 and MDR2) clinical strains of E. coli. We characterized a total of 273 metabolites, including 77 commonly altered metabolites, between MDR vs. antibiotic sensitive strains. Interestingly, the PCA score plot clearly discriminated drug sensitive and MDR strains. The following bioinformatics analysis showed that biosynthesis of amino acids, biosynthesis of phenylpropanoids and purine metabolism were commonly enriched in MDR strains. Moreover, microbial metabolism in diverse environments, carbon metabolism，and pyrimidine metabolism pathways were more likely to be enriched MDR1 strain, while ABC transporters, and cysteine and methionine metabolism pathways were enriched in MDR2 strains. The enzyme activities in several involved metabolic pathways were further measured and metabolite candidates were validated by GC-MS-SIM method. These results indicated that antibiotic resistance affects the metabolite profiles of bacteria. In general, our study provides evidence on the study and prediction of MDR characteristics and mechanisms in bacteria at the metabolite level. BIOLOGICAL SIGNIFICANCE: Overuse and abuse of antibiotics has led to the emergence of antibiotic-resistant strains of bacteria; however, relatively little is known about their resistance mechanisms. In this study, metabolomics method was used to compare the differential expression of metabolites between sensitive and multidrug-resistant clinical strains of E. coli. Results show that the PCA score plot clearly discriminated sensitive and MDR strains, indicating that they had different metabolic profiles. Further, bioinformatics analysis showed that biosynthesis of amino acids, biosynthesis of phenylpropanoids and purine metabolism may be related to resistance. Finally, the enzyme activities in several involved metabolic pathways were further measured and metabolite candidates were validated by GC-MS-SIM method. In general, our study provides evidence on the study and prediction of MDR characteristics and mechanisms in bacteria at the metabolite level.

ß-defensin 1 expression in HCV infected liver/liver cancer: an important role in protecting HCV progression and liver cancer development.

ß-defensin family plays a role in host defense against viral infection, however its role in HCV infection is still unknown. In this study, we demonstrated that ß-defensin 1 was significantly reduced in HCV-infected liver specimens. Treatment with interferon and ribavirin upregulated ß-defensin-1, but not other ß-defensin tested, with the extent and duration of upregulation associated with treatment response. We investigated ß-defensin family expression in liver cancer in publicly available datasets and found that among all the ß-defensins tested, only ß-defensin 1 was significantly downregulated, suggesting ß-defensin 1 plays a crucial role in liver cancer development. Further analysis identified E-cadherin as the top positive correlated gene, while hepatocyte growth factor-regulated tyrosine kinase substrate as the top negative correlated gene. Expression of two proteoglycans were also positively correlated with that of ß-defensin 1. We have also identified small molecules as potential therapeutic agents to reverse ß-defensin 1-associated gene signature. Furthermore, the downregulation of ß-defensin 1 and E-cadherin, and upregulation of hepatocyte growth factor-regulated tyrosine kinase substrate, were further confirmed in liver cancer and adjacent normal tissue collected from in-house Chinese liver cancer patients. Together, our results suggest ß-defensin 1 plays an important role in protecting HCV progression and liver cancer development.

Malignant angiomyolipoma in the liver: a case report with pathological and molecular analysis.

Malignant angiomyolipoma (AML) of the liver is rare. We report a case of AML with malignant transformation and metastases. A 30-year-old man had developed giant hepatic masses. Microscopically, the periphery of the tumor showed components of classic hepatic AML, but the central region contained atypical epithelioid components with extremely pleomorphic and hyperchromatic nuclei with frequent mitotic figures. Immunohistochemical analysis revealed that the epithelioid cells were positive for HMB-45 and smooth muscle actin. Furthermore, the atypical epithelioid cells displayed P53 immunoreactivity and mutation at exon 7 for p53. The tumor showed a typical monoclonal pattern but no loss of heterozygosity or microsatellite instability. Markedly atypical epithelioid cells with vascular invasion, distant metastasis, and fatal outcome were interpreted as malignant characteristics of hepatic AML. It is suggested that large tumor size, pleomorphic nuclei with high proliferation activity, and P53 immunoreactivity may predict the existence of malignant transformation of hepatic AML.