Multifunctional regulation of NADPH oxidase in growth, microsclerotia formation and virulence in Metarhizium rileyi.

OBJECTIVES: Microsclerotia (MS), anti-stress structures produced by many filamentous fungi, have been proven to be a great substitute for conidia in the production of insecticides within entomogenous fungi. NADPH oxidase (Nox) is a highly conserved ROS-response protein family that is widespread in eukaryotes and plays distinct roles in environmental fitness among various filamentous fungi. However, it is not clear whether the formation of MS and pathogenicity in entomogenous fungi is regulated by the Nox inside. In this study, we reported the presence of NADPH oxidase homologs in a great potential biocontrol fungus, Metarhizium rileyi, and further showed multiple biological functions. RESULTS: Three Nox homologous genes in M. rileyi showed high expression throughout the entire process of MS formation. Targeted deletion of MrNoxA, MrNoxB and MrNoxR all led to a decrease in MS yield and impaired morphology. Moreover, the anti-adversity assay showed that they are indispensable for growth, osmotic pressure and oxidative stress regulation in Metarhizium rileyi. Most importantly, △MrNoxR and △MrNoxA but not △MrNoxB showed a dramatic reduction in virulence via inoculation. The normality of appressoria might be unaffected in mutants since there are no striking differences in virulence compared with WT by topical injections. CONCLUSION: Our results revealed that NADPH oxidase plays important roles in growth regulation, MS formation and pathogenicity in M. rileyi, perhaps in the ROS response and hyphal polarity.

Comparative transcriptome analysis to unveil genes affecting the host cuticle destruction in Metarhizium rileyi.

Insect pathogenic fungi, also known as entomopathogenic fungi, are one of the largest insect pathogenic microorganism communities, represented by Beauveria spp. and Metarhizium spp. Entomopathogenic fungi have been proved to be a great substitute for chemical pesticide in agriculture. In fact, a lot of functional genes were also already characterized in entomopathogenic fungi, but more depth of exploration is still needed to reveal their complicated pathogenic mechanism to insects. Metarhizium rileyi (Nomuraea rileyi) is a great potential biocontrol fungus that can parasitize more than 40 distinct species (mainly Lepidoptera: Noctuidae) to cause large-scale infectious diseases within insect population. In this study, a comparative analysis of transcriptome profile was performed with topical inoculation and hemolymph injection to character the infectious pattern of M. rileyi. Appressorium and multiple hydrolases are indispensable constituents to break the insect host primary cuticle defense in entomopathogenic fungi. Within our transcriptome data, numerous transcripts related to destruction of insect cuticle rather growth regulations were obtained. Most importantly, some unreported ribosomal protein genes and novel unannotated protein (hypothetical protein) genes were proved to participate in the course of pathogenic regulation. Our current data provide a higher efficiency gene library for virulence factors screen in M. rileyi, and this library may be also useful for furnishing valuable information on entomopathogenic fungal pathogenic mechanisms to host.

RFWD3 Participates in the Occurrence and Development of Colorectal Cancer via E2F1 Transcriptional Regulation of BIRC5.

Objectives: Colorectal cancer (CRC) is one of the most common human malignancies. It was reported that the alterations in the DNA damage response (DDR) pathways are emerging as novel targets for treatment across different cancer types including CRC. RFWD3 plays a critical role in replication protein A (RPA)-mediated DNA damage in cancer cells. More importantly, RFWD3 can response to DNA damage by positively regulating p53 stability when the G1 cell cycle checkpoint is activated. However, the functional significance of RFWD3 in CRC has not been reported in the existing documents. Materials and Methods: Here, we revealed high expression of RFWD3 in CRC tissues by IHC analysis and The Cancer Genome Atlas (TCGA) database. Besides, overexpression of RFWD3 in CRC cell lines was also confirmed by qRT-PCR and western blot assay. The Celigo cell counting method and wound-healing/transwell migration assay were applied to evaluate CRC cell proliferation and migration. The tumor growth indicators were quantified in nude mice xenografted with shRFWD3 and shCtrl RKO cells. Results: The results indicated that RFWD3 knockdown restricted CRC development in vitro and in vivo. In exploring the downstream mechanism of RFWD3's action, we found that RFWD3 could transcriptionally activate BIRC5 by interacting with E2F transcription factor 1 (E2F1). Accordingly, we identified BIRC5 as a downstream gene of RFWD3 regulating CRC. Subsequent loss- and gain- of function experiments demonstrated that upon overexpressing BIRC5 in RKO cells with down-regulated RFWD3, the inhibitory effects of cell proliferation, migration and colony formation could be reversed, while the capacity of cell apoptosis was ameliorated, suggesting that the effects of RFWD3 depletion was mainly due to BIRC5 suppression. Conclusion: Taken together, this study revealed that RFWD3 participates in the occurrence and development of colorectal cancer via E2F1 transcriptional regulation of BIRC5.

Regulation of conidiation, polarity growth, and pathogenicity by MrSte12 transcription factor in entomopathogenic fungus, Metarhizium rileyi.

Metarhizium rileyi, a well-known filamentous biocontrol fungus, is the main pathogen of numerous field pests, especially noctuid pests. To explore the potential factors involved in the fungal pathogenicity, MrSte12, an important and conserved functional transcription factor in mitogen-activated protein kinase pathway was carried out by functional analysis. Homologous recombination was used to disrupt the MrSte12 gene in M. rileyi. The deletant fungal strain exhibited malformed hyphae and impaired conidiogenesis, and conidia could not be collected from △MrSte12 in vitro towards SMAY medium. Although conidia could be collected again supplemented with KCl within SMAY medium, the conidial germination, growth and stress tolerance were much weaker compared with that in WT. Additionally, △MrSte12 showed a dramatic reduction in virulence in intra-hemolymph injections and no pathogenicity in topical inoculations against noctuid pests, which is due to the failure of appressorium formation. Moreover, the content of chitin and ß-1, 3-glucan in cell wall significantly reduced in mutant conidia. These results indicate that the MrSte12 gene markedly contributes to invasive growth and conidiation, as well as the major pathogenicity in M. rileyi.

New pathway ameliorating ulcerative colitis: focus on Roseburia intestinalis and the gut-brain axis.

BACKGROUND: The community of gut microbes is a key factor controlling the intestinal barrier that communicates with the nervous system through the gut-brain axis. Based on our clinical data showing that populations of Roseburia intestinalis are dramatically decreased in the gut of patients with ulcerative colitis, we studied the efficacy of a strain belonging to this species in the context of colitis and stress using animal models. METHODS: Dextran sulfate sodium was used to induce colitis in rats, which then underwent an enema with R. intestinalis as a treatment. The disease activity index, fecal changes and body weight of rats were recorded to evaluate colitis, while histological and immunohistochemical analyses were carried out to examine colon function, and 16S rRNA sequencing was performed to evaluate the gut microbiota change. Behavioral assays and immunohistochemical staining of brain were performed to assess the effect of R. intestinalis on the gut-brain axis. RESULTS: Colitis-related symptoms in rats were significantly relieved after R. intestinalis enema, and the stool traits and colon length of rats were significantly recovered after treatment. The gut epithelial integrity and intestinal barrier were restored in treated rats, as evidenced by the higher expression of Zo-1 in colon tissues, accompanied by the restoration of gut microbiota. Meanwhile, depressive-like behaviors of rats were reduced after treatment, and laboratory experiments on neuronal cells also showed that IL-6, IL-7 and 5-HT were downregulated by R. intestinalis treatment in both serum and brain tissue, while Iba-1 expression was reduced in treated rats. CONCLUSIONS: The administration of R. intestinalis contributes to restoration of the gut microbiota, promoting colon repair and the recovery of gastrointestinal function. These alterations are accompanied by the relief of depressive-like behaviors through a process modulated by the neuronal network and the regulation of inflammation by the gut-brain axis.

Induced Effect of Environmental Regulation on Green Innovation: Evidence from the Increasing-Block Pricing Scheme.

With increasing constraints on resources and the environment, it is of great practical importance to discover and utilize the induced effect of green technology through market-based tools, in order to simultaneously realize economic development and ecological sustainability. Based on unique patent data from 1999 to 2013, this paper examines the induced effect of China's increasing-block electricity pricing scheme (IBP) on energy-efficient patents and checks whether the effect is neutral or biased. Furthermore, the quality of the induced patents is identified. The results reveal that increased green innovation is strongly related to the IBP scheme. In addition, the induced effect is biased towards green technology such that, apart from autonomous technological advances, the biased effect of IBP induced two more energy-efficient patents per hundred technological patents. However, the quality of the induced innovation is relatively low: compared to high-quality inventions, low-quality utility models showed greater and more significant growth due to the IBP. Our paper provides quantitative insight into the impact of the IBP and indicates that a reasonable pricing scheme can benefit both the environment and the economy.

Spatial Disparity of Meteorological Impacts on Carbon Monoxide Pollution in China during the COVID-19 Lockdown Period.

Lockdown due to the novel coronavirus disease 2019 (COVID-19) pandemic offers a unique opportunity to study the factors governing the variation in air pollution. A number of studies have investigated the cause underlying the occurrence of heavy haze pollution around the world during the lockdown period. However, information about spatiotemporal variations in gaseous pollutants and detailed quantifications of potential meteorological (METRO) impacts are limited. Ground measurements show that carbon monoxide (CO) pollution deteriorated in northern China despite strict control of human and industrial activities during the lockdown period in early 2020. In this study, a four-dimensional decomposition model was used to quantitatively extract the METRO impacts on the CO pollution over China. The results show that weakened winds elevated CO concentrations near Beijing and in northeastern China. Increased temperatures slightly elevated CO concentrations in northern and eastern China but reduced CO concentrations in northwestern China. Remarkable amounts of CO increases in northern China (e.g., by 0.21 mg/m3 within Beijing) were explained by anomalously high humidity, which could be associated with an enhanced interaction between aerosol and the boundary layer. After excluding the METRO impacts, the CO concentrations drastically declined across China (e.g., by 0.22 mg/m3 within Beijing), indicating that the lockdown indeed greatly lessened CO concentrations. However, the adverse METRO conditions counteracted the beneficial outcomes of emission reductions, leading to a deterioration of the CO pollution in northern China. These results indicate that the METRO factors can play a critical role in worsening air pollution despite a strict control of anthropogenic emissions.

Patterns and Prognostic Value of Lymph Node Metastasis on Distant Metastasis and Survival in Nasopharyngeal Carcinoma: A Surveillance, Epidemiology, and End Results Study, 2006-2015.

This study was conducted to identify factors associated with lymph node (LN) metastasis in nasopharyngeal carcinoma (NPC) patients, analyze node distribution patterns, and explore the prognostic value of the LN metastasis level for survival. We included 2994 patients with primary NPC diagnosed between 2006 and 2015 with information in the Surveillance, Epidemiology, and End Results (SEER) database. Patients' demographic and clinicopathologic features were compared according to LN status using chi-squared tests. The 5-year overall survival (OS) and cancer-specific survival (CSS) rates were calculated by the Kaplan-Meier method, and the differences were estimated by log-rank tests. Multivariate Cox proportional hazard models were used to evaluate independent risk factors for OS and CSS. Logistic regression was used to evaluate the risk of each LN metastasis category for distant metastasis. There were 695 patients in the N0 stage and 2299 with LN metastasis (classified as stage N1, N2, or N3). The overall incidence of LN metastasis was 76.8%. Sex and T stage were not associated with LN metastasis. Older patients had a significantly worse 5-year OS and CSS than younger patients. In terms of histologic type, keratinizing squamous cell carcinoma had the lowest 5-year OS and CSS at 48.2% and 53.8%, respectively. The most common nodal involvement level was II (65.9%), followed by III (29.1%), V (25.6%), I (17.6%), IV (15.7%), and retropharynx (13.5%). The skip metastasis rate was 5.7% (130/2299). Patients with only level II metastasis (classified as level 2) was the most common category, accounting for 30%. Compared to level 2, patients with only level I (classified as level 1) had an OR of 2.101 (95% CI: 1.090-4.047, P=0.027) for distant metastasis, patients with simultaneous levels II, III, IV, and V (classified as levels 2345) had the highest OR of 4.064 (95% CI: 2.155-7.666, P < 0.001) for distant metastasis, and level 24 had an OR of 3.003 (95% CI: 1.074-8.395, P=0.036) for distant metastasis. In survival analysis, levels 235 had a significant HR of 1.708 (95% CI: 1.089-2.678, P=0.020) for CSS compared to level 2 after adjustment for age, sex, race, histology, TNM (tumor, node, and metastasis) stage, and treatment.

A rhizobia strain isolated from root nodule of gymnosperm Podocarpus macrophyllus.

Symbiotic nitrogen fixation of rhizobia and leguminous plants is considered as the most important biologic nitrogen fixation system on earth. Symbiotic nodulation of gymnosperm Podocarpus macrophyllus and rhizobia has never been reported. In this study, 11 endophytic bacteria strains were isolated from root nodules of P. macrophyllus and its variation P. macrophyllus var. maki. The plant infection tests on these strains indicated that the isolated strains could be nodulated on P. macrophyllus plants, and weak nitrogenase activity of nodules was found in acetylene reduction method. According to the physiological and biochemical characteristics of the 11 strains, GXLO 02 was selected as the representative strain. 16S rDNA full-length sequence analysis of GXLO 02 confirmed that the representative strain GXLO 02 belongs to Rhizobium sp.