Development and verification of a novel immunogenic cell death-related signature for predicting the prognosis and immune infiltration in triple-negative breast cancer.

BACKGROUND: Insufficient understanding of the pathogenesis and tumor immunology of triple-negative breast cancer (TNBC) has limited the development of immunotherapy. The importance of tumor microenvironment (TME) in immunotyping, prognostic assessment and immunotherapy efficacy of cancer has been emphasized, however, potential immunogenic cell death (ICD) related genes function in TME of TNBC has been rarely investigated. AIMS: To initially explore the role and related mechanisms of ICD in TNBC, especially the role played in the TME of TNBC, and to identify different relevant subtypes based on ICD, and then develop an ICD-related risk score to predict each TNBC patient TME status, prognosis and immunotherapy response. METHODS AND RESULTS: In this study, we identified distinct ICD-related modification patterns based on 158 TNBC cases in the TCGA-TNBC cohort. We then investigated the possible correlation between ICD-related modification patterns and TME cell infiltration characteristics in TNBC. By using univariate Cox and least absolute shrinkage and selection operator (LASSO) regression analysis, we created a risk scoring system (ICD score) to quantifiably evaluate the impact of ICD-related modification patterns in individual TNBC patient. Two different ICD-related modification patterns were found with significant differences in immune infiltration. Lower ICD score was correlated with higher immune infiltration, tumor mutational burden and significantly enriched in immune-related pathways, indicating a strong ability to activate immune response, which might account for relatively favorable prognosis of TNBC patients and could serve as a predictor to select suitable candidates for immunotherapy. We used two independent cohorts, GSE58812 cohort and Metabric cohort to validate prognosis and immunohistochemistry for preliminary in vitro validation. CONCLUSION: This study evidenced that the ICD-related modification patterns might exert pivotal roles in the immune infiltration landscape of TNBC and ICD score might act as potential predictors of prognostic assessment and immunotherapy response. This research provides unique insights for individualize immune treatment strategies and promising immunotherapy candidates screening.

Bilateral Pulmonary Enteric Adenocarcinoma on FDG PET/CT.

ABSTRACT: Pulmonary enteric adenocarcinoma(PEAC) is a rare variant of pulmonary adenocarcinoma. We herein report FDG PET/CT findings of PEAC in a 76-year-old woman. The images showed widespread opacities in both lungs with intense FDG uptake. The pathology revealed pulmonary enteric adenocarcinoma.

Sphingosine 1-phosphate receptor 1 regulates blood-brain barrier permeability in epileptic mice.

Destruction of the blood-brain barrier is a critical component of epilepsy pathology. Several studies have demonstrated that sphingosine 1-phosphate receptor 1 contributes to the modulation of vascular integrity. However, its effect on blood-brain barrier permeability in epileptic mice remains unclear. In this study, we prepared pilocarpine-induced status epilepticus models and pentylenetetrazol-induced epilepsy models in C57BL/6 mice. S1P1 expression was increased in the hippocampus after status epilepticus, whereas tight junction protein expression was decreased in epileptic mice compared with controls. Intraperitoneal injection of SEW2871, a specific agonist of sphingosine-1-phosphate receptor 1, decreased the level of tight junction protein in the hippocampus of epileptic mice, increased blood-brain barrier leakage, and aggravated the severity of seizures compared with the control. W146, a specific antagonist of sphingosine-1-phosphate receptor 1, increased the level of tight junction protein, attenuated blood-brain barrier disruption, and reduced seizure severity compared with the control. Furthermore, sphingosine 1-phosphate receptor 1 promoted the generation of interleukin-1ß and tumor necrosis factor-α and caused astrocytosis. Disruption of tight junction protein and blood-brain barrier integrity by sphingosine 1-phosphate receptor 1 was reversed by minocycline, a neuroinflammation inhibitor. Behavioral tests revealed that sphingosine 1-phosphate receptor 1 exacerbated epilepsy-associated depression-like behaviors. Additionally, specific knockdown of astrocytic S1P1 inhibited neuroinflammatory responses and attenuated blood-brain barrier leakage, seizure severity, and epilepsy-associated depression-like behaviors. Taken together, our results suggest that astrocytic sphingosine 1-phosphate receptor 1 exacerbates blood-brain barrier disruption in the epileptic brain by promoting neuroinflammation.

Low-Dose NIR-II Preclinical Bioimaging Using Liposome-Encapsulated Cyanine Dyes.

Fluorescence imaging in the second near-infrared window (NIR-II, 1000-1700 nm) provides a powerful tool for in vivo structural and functional imaging in deep tissue. However, the lack of biocompatible contrast agents with bright NIR-II emission has hindered its application in fundamental research and clinical trials. Herein, a liposome encapsulation strategy for generating ultrabright liposome-cyanine dyes by restricting dyes in the hydrophobic pockets of lipids and inhibiting the aggregation, as corroborated by computational modeling, is reported. Compared with free indocyanine green (ICG, an US Food and Drug Administration-approved cyanine dye), liposome-encapsulated ICG (S-Lipo-ICG) shows a 38.7-fold increase in NIR-II brightness and enables cerebrovascular imaging at only one-tenth dose over a long period (30 min). By adjusting the excitation wavelength, two liposome-encapsulated cyanine dyes (S-Lipo-ICG and S-Lipo-FD1080) enable NIR-II dual-color imaging. Moreover, small tumor nodules (2-5 mm) can be successfully distinguished and removed with S-Lipo-ICG image-guided tumor surgery in rabbit models. This liposome encapsulation maintains the metabolic pathway of ICG, promising for clinical implementation.

Surgical technique management of microvascular decompression for trigeminal neuralgia.

Trigeminal neuralgia is a severe neuropathic disorder, affecting the distribution area of the trigeminal nerve and often impairs the quality of life of patients. More and more scholars agree that one of the pathogenesis of trigeminal neuralgia is due to the demyelinating lesion caused by vascular compression or arachnoid bundle wrapping on the root exit zone of trigeminal nerve. In this regard, the most effective method is microvascular decompression, which can relieve the compression of the offending vessels and the thickened arachnoid on the trigeminal nerve. However, it still has some disadvantages, such as the possibility of fatal complications. In recent years, with the advancement of neurosurgical treatment technology, new progress has been made in microvascular decompression. This article mainly introduces the surgical techniques and new methods of the microvascular decompression.

Efficacy of Dialectical Comprehensive Treatment of Traditional Chinese Medicine in Patients with Chronic Stable Heart Failure: A Randomized Controlled Trial.

The treatment of chronic stable heart failure (CSHF) with integrated traditional Chinese and Western medicine has been of wide concern. We mainly discuss the clinical efficacy of TCM decoction combined with acupuncture and moxibustion (A&M) in CSHF treatment on the basis of syndrome differentiation and treatment (SDT). The control group was given conventional cardiac rehabilitation (CCR), and the treatment group was given TCM decoction combined with A&M treatment based on SDT on the basis of conventional cardiac rehabilitation. The clinical efficacy and cardiopulmonary exercise testing (CPET) indicators were evaluated. Left ventricular ejection fraction (LVEF), NT-proBNP, myocardial ischemia threshold (MIT), and 6-minute walking distance (6MWD) were measured by ultrasound, ELISA, electrocardiogram, and 6MWD test. After treatment, the clinical efficacy, LVEF, and 6MWD of the treatment group were better than in the control group. The NT-proBNP plasma level and MIT in the treatment group were lower than in the control group. The treatment group had enhanced AT, VO2 Peak, VO2 Peak/HR, and Peak power and decreased resting systolic pressure and peak systolic pressure, and the difference was statistically significant. Dialectical comprehensive treatment of TCM could effectively improve cardiac function and clinical treatment effect, which was worthy of clinical application.

Dietary Copper Intake and Risk of Stroke in Adults: A Case-Control Study Based on National Health and Nutrition Examination Survey 2013-2018.

The association between dietary copper intake and the risk of stroke is unknown. We included a total of 10,550 participants from the National Health and Nutrition Examination Survey (NHANES) 2013−2018. Two 24-h dietary recalls and a standard questionnaire were used to determine copper intake and stroke, respectively. We used logistic regression models to estimate the associations between dietary copper intake and the risk of stroke. The nearest-neighbor propensity score matching (PSM) with a ratio of 1:2 was used to reduce selection bias. The non-linear relationship was explored with restricted cubic splines (RCS). The correlation between copper intake and baseline characteristics was detected by the Pearson correlation coefficient. The median dietary copper intake was 1.072 mg/day (IQR = 1.42−0.799). Approximately 3.8% (399) of the participants had a history of stroke. A multivariate logistic regression analysis before and after matching showed that subjects in the higher quartile had significantly lower odds of stroke compared with subjects in the first quartile of copper intake. A stratified analysis showed that copper intake was a significant protective factor for women, individuals <65 years old, individuals with hypertension, individuals who smoke, and diabetic stroke patients. The RCS models showed an L-shaped nonlinear relationship (p for nonlinear < 0.001) between copper intake and stroke. Our results suggested that increased dietary copper intake was associated with a lower risk of stroke.

Structural and functional insights into a novel pre-clinical-stage antibody targeting IL-17A for treatment of autoimmune diseases.

The pro-inflammatory cytokine interleukin-17A (IL-17A) is a key driver of multiple inflammatory and immune disorders. Therapeutic antibodies targeting IL-17A have been proven effective in treating patients with these diseases; however, large variations in clinical outcomes have been observed with different antibodies. In this study, we developed HB0017, a novel monoclonal antibody that targets human IL-17A. HB0017 specifically and strongly bound to human, cynomolgus monkey, and mouse IL-17A at the physiological interface with the IL-17A receptor. In human and monkey cells, HB0017 potently antagonized the functions of IL-17A through competitive binding. HB0017 functioned equivalently to that of clinically approved antibodies in terms of therapeutic efficacy for inflammatory disorders and psoriasis in a mouse model. The results indicate that HB0017 may be an alternative biological therapy for treating patients with inflammation and autoimmune diseases.

Risk Factors for Epilepsy: A National Cross-Sectional Study from National Health and Nutrition Examination Survey 2013 to 2018.

OBJECTIVE: The purpose of this study is to investigate the risk factors of epilepsy based on the National Health and Nutrition Examination Survey (NHANES). METHODS: The data in this study was obtained from the NHANES database between 2013 and 2018. It included 14,290 participants aged between 20 and 80. We defined people with epilepsy (PWE) when they self-reported took at least one treatment medication for seizures or epilepsy. Analysis of risk factors for epilepsy mainly includes Student's t-test, chi-square test, univariate and multivariate logistic regression analysis. RESULTS: People aged 40-59 shared 1.8 times the risk of epilepsy than those who aged 20-39, P=0.034. People who never married had a 2.8-fold higher risk of epilepsy than those who married/living with partner, P<0.001. The risk of epilepsy in subjects with very good/good general health was 0.4 times than that of subjects with fair/poor general health, P<0.001. Moreover, subjects without sleep disorders had a 0.4-fold higher risk of epilepsy than those who had sleep disorders, P=0.042. CONCLUSION: People who are older, unmarried, and have sleep disorders are at higher risk of epilepsy. In addition, good/good general health condition is associated with a lower risk of epilepsy.

Perampanel, an AMPAR antagonist, alleviates experimental intracerebral hemorrhage­induced brain injury via necroptosis and neuroinflammation.

Spontaneous intracerebral hemorrhage (ICH) is a subtype of stroke with high mortality and morbidity due to the lack of effective therapies. The alpha­amino­3­hydroxy­5­methyl­4­isoxazolepropionic acid receptor antagonist perampanel has been reported to alleviate early brain injury following subarachnoid hemorrhage and traumatic brain injury by reducing reactive oxygen species, apoptosis, autophagy, and necroptosis. Necroptosis is a caspase­independent programmed cell death mechanism that serves a vital role in neuronal cell death following ICH. However, the precise role of necroptosis in perampanel­mediated neuroprotection following ICH has not been confirmed. The present study aimed to investigate the neuroprotective effects and potential molecular mechanisms of perampanel in ICH­induced early brain injury by regulating neural necroptosis in C57BL/6 mice and in a hemin­induced neuron damage cell culture model. Mortality, neurological score, brain water content, and neuronal death were evaluated. The results demonstrated that perampanel treatment increased the survival rate and neurological score, and increased neuron survival. In addition, perampanel treatment downregulated the protein expression levels of receptor interacting serine/threonine kinase (RIP) 1, RIP3, and mixed lineage kinase domain like pseudokinase, and of the cytokines IL­1ß, IL­6, TNF­α, and NF­κB. These results indicated that perampanel­mediated inhibition of necroptosis and neuroinflammation ameliorated neuronal death in vitro and in vivo following ICH. The neuroprotective capacity of perampanel was partly dependent on the PTEN pathway. Taken together, the results of the present study demonstrated that perampanel improved neurological outcomes in mice and reduced neuronal death by protecting against neural necroptosis and neuroinflammation.

Atorvastatin ameliorates early brain injury after subarachnoid hemorrhage via inhibition of pyroptosis and neuroinflammation.

Subarachnoid hemorrhage (SAH) is a subtype of stroke with high mortality and morbidity due to the lack of effective therapy. Atorvastatin has been reported to alleviate early brain injury (EBI) following subarachnoid hemorrhage (SAH) via reducing reactive oxygen species, antiapoptosis, regulated autophagy, and neuroinflammation. Which was the related to the pyroptosis? Pyroptosis can be defined as a highly specific inflammatory programmed cell death, distinct from classical apoptosis and necrosis. However, the precise role of pyroptosis in atorvastatin-mediated neuroprotection following SAH has not been confirmed. The present study aimed to investigate the neuroprotection and potential molecular mechanisms of atorvastatin in the SAH-induced EBI via regulating neural pyroptosis using the filament perforation model of SAH in male C57BL/6 mice, and the hemin-induced neuron damage model in HT-22. Atorvastatin or vehicle was administrated 2 h after SAH and hemin-induced neuron damage. The mortality, neurological score, brain water content, and neuronal death were evaluated. The results show that the atorvastatin treatment markedly increased survival rate, neurological score, greater survival of neurons, downregulated the protein expression of NLRP1, cleaved caspase-1, interleukin-1ß (IL-1ß), and IL-18, which indicated that atorvastatin-inhibited pyroptosis and neuroinflammation, ameliorated neuron death in vivo/vitro subjected to SAH. Taken together, this study demonstrates that atorvastatin improved the neurological outcome in rats and reduced the neuron death by against neural pyroptosis and neuroinflammation.

Development and Validation of Prognostic Survival Nomograms for Patients with Anal Canal Cancer: A SEER-Based Study.

OBJECTIVE: Anal canal cancer is a rare malignancy with increasing incidence in recent times. This study aimed to develop two nomograms to predict the overall survival (OS) and cancer-specific survival (CSS) of patients with anal canal cancer. METHODS: Information of patients with anal canal cancer from 2004 to 2015 was extracted from the surveillance, epidemiology, and end results (SEER) database. Cox analysis was used to select the risk factors for prognosis, and nomograms were constructed using the R software. The C-index, area under the curve (AUC) of time-dependent receiver operating characteristic (ROC) curves, calibration plot and decision curve analysis (DCA) were used to assess the clinical utility of the nomograms. RESULTS: A total of 2458 patients with malignant tumours of the anal canal were screened out. Sex, age, marital status, histological type, grade, tumour size, AJCC stage, SEER stage and chemotherapy were independent prognostic factors for OS, whereas sex, age, race, histological type, grade, tumour size, AJCC stage, SEER stage and radiotherapy were independent prognostic factors for CSS. In the training cohort, the C-index value for OS nomogram was 0.73 (95% CI, 0.69-0.77), and the AUC values that predicted the 1-, 3- and 5-year survival rates were 0.764, 0.758 and 0.760, respectively, whereas the C-index value for CSS nomogram model was 0.74 (95% CI, 0.69-0.79), and the AUC values were 0.763, 0.769 and 0.763, respectively. The calibration plot and DCA curves demonstrated good prediction performance of the model in both the training and validation cohorts. CONCLUSION: The established nomogram is a visualisation tool that can effectively predict the OS and CSS of patients with anal canal cancer.

Discovery of Potential Flavonoid Inhibitors Against COVID-19 3CL Proteinase Based on Virtual Screening Strategy.

The outbreak of 2019 novel coronavirus (COVID-19) has caused serious threat to public health. Discovery of new anti-COVID-19 drugs is urgently needed. Fortunately, the crystal structure of COVID-19 3CL proteinase was recently resolved. The proteinase has been identified as a promising target for drug discovery in this crisis. Here, a dataset including 2030 natural compounds was screened and refined based on the machine learning and molecular docking. The performance of six machine learning (ML) methods of predicting active coronavirus inhibitors had achieved satisfactory accuracy, especially, the AUC (Area Under ROC Curve) scores with fivefold cross-validation of Logistic Regression (LR) reached up to 0.976. Comprehensive ML prediction and molecular docking results accounted for the compound Rutin, which was approved by NMPA (National Medical Products Administration), exhibited the best AUC and the most promising binding affinity compared to other compounds. Therefore, Rutin might be a promising agent in anti-COVID-19 drugs development.

The functional identification of glycine-rich TtASR from Tetragonia tetragonoides (Pall.) Kuntze involving in plant abiotic stress tolerance.

In this study, we reported on an ASR gene (TtASR) related to salt/drought tolerance from the edible halophyte Tetragonia tetragonoides (Pall.) Kuntze (Aizoaceae). A phylogenetic analysis revealed that TtASR was evolutionarily close to other two halophytic glycine-rich ASR members, SbASR-1 (from Salicornia brachiate) and SlASR (from Suaeda liaotungensis), with a typical abscisic acid (ABA)/water-deficit stress (WDS) domain at C-terminal. Quantitative RT-PCR analyses showed that TtASR was expressed in all tested different organs of the T. tetragonoides plant and that expression levels were apparently induced after salt, osmotic stress, and ABA treatments in T. tetragonoides seedlings. An induction of TtASR improved the growth performance of yeast and bacteria more than the control under high salinity, osmotic stress, and oxidative stress. TtASR was not a nuclear-specific protein in plant, and the transcriptional activation assay also demonstrated that TtASR could not activate reporter gene's expression in yeast. TtASR overexpressed Arabidopsis plants exhibited higher tolerance for salt/drought and oxidative stresses and lower ROS accumulation than wild type (WT) plants, accompanied by increased CAT, SOD activities, higher proline content, and lower MDA content in vivo. The results indicated that the TtASR was involved in plant responses to salt and drought, probably by mediating water homeostasis or by acting as ROS scavengers, and that it decreased the membrane damage and improved cellular osmotic adjustment that respond to abiotic stresses in microorganisms and plants.

Investigating the Mechanism of L-Valine in Improving the Stability of Gabapentin Combining Chemical Analysis Experiments with Computational Pharmacy.

The mechanism of L-Val on how to improve the stability of gabapentin (GBP) was described by the combination of chemical analysis experiments and computer simulations. Scanning electron microscope (SEM), powder X-ray diffraction (PXRD), and differential scanning calorimeter (DSC), coupled with attenuated total reflectance Fourier transform infrared spectroscopy (ATR-FTIR), were used to identify ß-GBP prepared by rapid solvent removal method. The reaction barriers on crystal planes, ß-GBP (100) and ß-GBP (10-1), are smaller than α-GBP and γ-GBP, reaching 276.65 kJ/mol and 299.57 kJ/mol, respectively. Thus, it was easier for ß-GBP to form lactam, and the occurrence of ß-GBP would lead the worse stability of α-GBP. The addition of neutral amino acids such as L-Val could improve the stability of α-GBP effectively. The adsorption energy of α-GBP (002) crystal plane with L-Val is larger than that of other crystal planes, reaching 42.17 kJ/mol. Hydrogen bond was the combination of L-Val and GBP main crystal planes, which could inhibit the crystal transformation of α-GBP. These results suggest that neutral amino acid protectants, such as L-Val, could improve the stability of α-GBP effectively, and inhibition of crystal transformation is one of the effective methods to improve the stability of polymorphic drugs.

Investigation of Fused Alumina Based-Mold Facecoats for DZ22B Directionally Solidified Blades.

The interaction between alloy melt and mold facecoats is the main factor affecting the surface quality of investment casting nickel-based superalloys. An investigation was undertaken to develop suitable refractories as facecoat materials for the directionally solidified blades of DZ22B nickel-based superalloys in order to avoid a sand-burning defect. The wettability and interface reactions between alloy melt and various facecoats were studied by using a sessile drop experiment and the real investment casting method, respectively. The results show that by adding Cr2O3 powder with the amounts of 2 wt.%, 5 wt.% and 10 wt.% in the fused alumina-based facecoats, the wetting angles between the alloy melt and facecoats decreased from 105.40° to 100.37°, 99.96° and 98.11°, respectively, while the sand-burning defect on the casting blade surfaces still formed during the process of directional solidification. However, by adding h-BN powder in the fused alumina-based facecoats, the wetting angles between the alloy melt and facecoats dramatically increased, the sand-burning defect on the casting blade surfaces was effectively inhibited and a metallic luster on the directionally solidified blades could be obviously observed. In this study, the suitable composition of mold facecoats for the investment casting of blades is 2 wt.% h-BN added fused alumina.

Expression signatures of long non-coding RNA and mRNA in human traumatic brain injury.

Long non-coding RNAs (lncRNAs) play a key role in craniocerebral disease, although their expression profiles in human traumatic brain injury are still unclear. In this regard, in this study, we examined brain injury tissue from three patients of the 101st Hospital of the People's Liberation Army, China (specifically, a 36-year-old male, a 52-year-old female, and a 49-year-old female), who were diagnosed with traumatic brain injury and underwent brain contusion removal surgery. Tissue surrounding the brain contusion in the three patients was used as control tissue to observe expression characteristics of lncRNAs and mRNAs in human traumatic brain injury tissue. Volcano plot filtering identified 99 lncRNAs and 63 mRNAs differentially expressed in frontotemporal tissue of the two groups (P < 0.05, fold change > 1.2). Microarray analysis showed that 43 lncRNAs were up-regulated and 56 lncRNAs were down-regulated. Meanwhile, 59 mRNAs were up-regulated and 4 mRNAs were down-regulated. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses revealed 27 signaling pathways associated with target genes and, in particular, legionellosis and influenza A signaling pathways. Subsequently, a lncRNA-gene network was generated, which showed an absolute correlation coefficient value > 0.99 for 12 lncRNA-mRNA pairs. Finally, quantitative real-time polymerase chain reaction confirmed different expression of the five most up-regulated mRNAs within the two groups, which was consistent with the microarray results. In summary, our results show that expression profiles of mRNAs and lncRNAs are significantly different between human traumatic brain injury tissue and surrounding tissue, providing novel insight regarding lncRNAs' involvement in human traumatic brain injury. All participants provided informed consent. This research was registered in the Chinese Clinical Trial Registry (registration number: ChiCTR-TCC-13004002) and the protocol version number is 1.0.

Karyopherinß1 regulates proliferation of human glioma cells via Wnt/ß-catenin pathway.

Karyopherinß1 (KPNB1), one of the cytosolic factors involved in the selective protein transport across nucleus, docked at nuclear pore complex and transported through nuclear envelope in an ATP-dependent style, assisting proteins to be recognized as import substrates. It has been reported to be bound up with the origination and progress of lung cancer, cervical cancer, head and neck cancer and hepatocellular carcinoma. In current study, we demonstrated for the first time that the role of KPNB1 in human glioma. KPNB1 was over-expressed as the well-known trend of Ki-67(p < 0.01) and tightly closed to poor prognosis, as an independent prognostic factor. In vitro, up-regulation of KPNB1 was accompanied by certain rising levels of proliferation markers, employing U251 and U87MG cells as serum-starve models. Silencing KPNB1 in U251 and U87MG led to G1 phase arrested directly via flow cytometry analysis. In the nucleus of KPNB1-depletion cell models, the decreasing expression of KPNB1 and ß-catenin was detected respectively, which indicated that KPNB1 functioned via ß-catenin signal. Besides, the interaction between KPNB1 and ß-catenin was proved clearly by immunoprecipitation. Taken together, it showed that KPNB1 might enhance human glioma proliferation via Wnt/ß-Catenin Pathway.

Biochemical characterization of a thermostable HNH endonuclease from deep-sea thermophilic bacteriophage GVE2.

His-Asn-His (HNH) proteins are a very common family of small nucleic acid-binding proteins that are generally associated with endonuclease activity and are found in all kingdoms of life. Although HNH endonucleases from mesophiles have been widely investigated, the biochemical functions of HNH endonucleases from thermophilic bacteriophages remain unknown. Here, we characterized the biochemical properties of a thermostable HNH endonuclease from deep-sea thermophilic bacteriophage GVE2. The recombinant GVE2 HNH endonuclease exhibited non-specific cleavage activity at high temperature. The optimal temperature of the GVE2 HNH endonuclease for cleaving DNA was 60-65 °C, and the enzyme retained its DNA cleavage activity even after heating at 100 °C for 30 min, suggesting the enzyme is a thermostable endonuclease. The GVE2 HNH endonuclease cleaved DNA over a wide pH spectrum, ranging from 5.5 to 9.0, and the optimal pH for the enzyme activity was 8.0-9.0. Furthermore, the GVE2 HNH endonuclease activity was dependent on a divalent metal ion. While the enzyme is inactive in the presence of Cu(2+), the GVE2 HNH endonuclease displayed cleavage activity of varied efficiency with Mn(2+), Mg(2+), Ca(2+), Fe(2+), Co(2+), Zn(2+), and Ni(2+). The GVE2 HNH endonuclease activity was inhibited by NaCl. This study provides the basis for determining the role of this endonuclease in life cycle of the bacteriophage GVE2 and suggests the potential application of the enzyme in molecular biology and biotechnology.

Fungal communities from the calcareous deep-sea sediments in the Southwest India Ridge revealed by Illumina sequencing technology.

The diversity and ecological significance of bacteria and archaea in deep-sea environments have been thoroughly investigated, but eukaryotic microorganisms in these areas, such as fungi, are poorly understood. To elucidate fungal diversity in calcareous deep-sea sediments in the Southwest India Ridge (SWIR), the internal transcribed spacer (ITS) regions of rRNA genes from two sediment metagenomic DNA samples were amplified and sequenced using the Illumina sequencing platform. The results revealed that 58-63 % and 36-42 % of the ITS sequences (97 % similarity) belonged to Basidiomycota and Ascomycota, respectively. These findings suggest that Basidiomycota and Ascomycota are the predominant fungal phyla in the two samples. We also found that Agaricomycetes, Leotiomycetes, and Pezizomycetes were the major fungal classes in the two samples. At the species level, Thelephoraceae sp. and Phialocephala fortinii were major fungal species in the two samples. Despite the low relative abundance, unidentified fungal sequences were also observed in the two samples. Furthermore, we found that there were slight differences in fungal diversity between the two sediment samples, although both were collected from the SWIR. Thus, our results demonstrate that calcareous deep-sea sediments in the SWIR harbor diverse fungi, which augment the fungal groups in deep-sea sediments. This is the first report of fungal communities in calcareous deep-sea sediments in the SWIR revealed by Illumina sequencing.