Real-time quantitative reverse transcription PCR assay for the detection of Nuomin virus - An emerging tick-borne virus.

Nuomin virus (NOMV), an emerging tick-borne virus (TBVs) identified in 2020, has been associated with fever, headache, and potential liver dysfunction in infected individuals. This study presents a novel TaqMan real-time quantitative PCR method designed for the rapid, sensitive, and specific detection of NOMV, facilitating early diagnosis. Utilizing Beacon Designer software 8.0, we optimized the PCR assay including the development of primers and probes to precisely target the conserved region of the NOMV genome, followed by optimization of primer and probe concentrations and annealing temperature. The resulting assay demonstrated robust performance, with standard curve represented by the equation y=-3.29x+39.42, a high correlation coefficient (R2 = 0.995) and an efficiency 99.53 %. Importantly, the method exhibited exceptional specificity, which did not yield cross-reacting signals from other TBVs, including Songling virus (SGLV), Beiji virus (BJNV), tick-borne encephalitis virus (TBEV), Yezo virus (YEZV), Alongshan virus (ALSV), and severe fever with thrombocytopenia syndrome bunyavirus (SFTSV). The assay's detection limit was remarkably low, reaching 10 copies/µL, representing a 100-fold increase compared to semi-nested RT-PCR. Additionally, it demonstrated excellent repeatability, with coefficients of variation for intra- and inter-group tests consistently below 3 %. Clinical evaluations confirmed the assay's superior performance, highlighting its high specificity, sensitivity, and reproducibility for NOMV detection. In conclusion, the method developed in this study provides a valuable tool to support timely management of NOMV infections, with significant implications for clinical practice.

Identification of SNHG11 as a Therapeutic Target in Pulmonary Hypertension.

Pulmonary hypertension (PH) is a life-threatening condition characterized by pulmonary vascular remodeling and endothelial dysfunction. Current therapies primarily target vasoactive imbalances but often fail to address adverse vascular remodeling. Long non-coding RNA (lncRNA), which are key regulators of various cellular processes, remain underexplored in the context of PH. To investigate the role of lncRNA in PH, we performed a comprehensive analysis using Weighted Gene Co-expression Network Analysis (WGCNA) on the GSE113439 dataset, comprising human lung tissue samples from different PH subtypes. Our analysis identified the lncRNA SNHG11 as consistently downregulated in PH. Functional assays in human pulmonary artery endothelial cells (HPAECs) demonstrated that SNHG11 plays a critical role in modulating inflammation, cell proliferation, apoptosis, and the JAK/STAT and MAPK signaling pathways. Mechanistically, SNHG11 influences the stability of PRPF8, a crucial mRNA spliceosome component, thereby affecting multiple cellular functions beyond splicing. In vivo experiments using a hypoxic rat model showed that knockdown of SNHG11 alleviates PH development and improves right ventricular function. These findings highlight SNHG11 as a key regulator in PH pathogenesis and suggest it as a potential therapeutic target.

[Retracted] SOX2 knockdown inhibits the migration and invasion of basal cell carcinoma cells by targeting the SRPK1­mediated PI3K/AKT signaling pathway.

[This retracts the article DOI: 10.3892/ol.2018.9810.].

Calcium enrichment activity initiates extracellular calcium influx-dependent inflammatory response of biologically-derived hydroxyapatite.

Biologically-derived hydroxyapatite is a widely used biomaterial in various clinical applications including bone augmentation. However, the osteogenic application of biological hydroxyapatite is limited by inflammatory responses, and the underlying mechanism remains unknown. The current study aimed to elucidate the molecular mechanisms underlying the inflammatory response to biological hydroxyapatite. Porcine-derived hydroxyapatite (PHA) with two sintering temperatures (800 and 1600 °C), PHA800 and PHA1600, respectively, were prepared. A PHA/macrophage co-culture model was established. Transcriptome, polymerase chain reaction (PCR), and enzyme-linked immunosorbent assay (ELISA) analyses were used to determine the inflammatory effects and the main pathways activated by PHA800 and PHA1600. Intracellular calcium level, PHA-induced calcium enrichment, and related biological effects were used to determine the molecular mechanism at the PHA-cell interface. PHA800 significantly upregulated a TLR4 mediated inflammatory pathway in a calcium influx-dependent manner, and the calcium enrichment activity on the surface of PHA800 promoted calcium influx. In contrast, the calcium enrichment activity on the surfaces of the PHA1600 and PHA800 pretreated groups was attenuated, resulting in decreased calcium influx and mild inflammatory effects. Our results provide a fundamental basis for the development of novel bone substitutes that elicit low levels of inflammation response.

Identification and characterization of Jingmen tick virus from Rhipicephalus microplus in Hunan, China.

Jingmen tick virus (JMTV) is a tick-borne pathogen known to affect human beings, characterized by a segmented genome structure that defies the conventional understanding of the Flaviviridae family. In the present study, we employed metagenomic analysis to screen for tick-borne viruses in Hunan Province, China, and identified five JMTV variants with complete genomes from Rhipicephalus microplus ticks sampled from cattle. These viral strains exhibited the highest sequence similarity to JMTV isolates previously reported in Hubei Province, China. However, evidence of genomic reassortment was detected, particularly with the S2 segment showing greater similarity to the strains from Japan. Phylogenetic analysis demonstrated that JMTV strains cluster predominantly based on their geographic origin. In agreement with the homology data, the S1, S3, and S4 segments of the strains identified in this study grouped with those from Hubei Province, while the S2 segment displayed a distinct topological structure. Moreover, JMTV displayed limited replication in mammal-derived cells, but thrived in tick-derived cell lines. In addition to the commonly used R. microplus-derived BME/CTVM23 cells, we found that JMTV also proliferated robustly in both Ixodes scapularis-derived ISE6 and Ixodes ricinus-derived IRE/CTVM19 cells, offering new avenues for in vitro production of the virus. In summary, this study expands the known geographic distribution and genetic diversity of JMTV, providing valuable insights into its epidemiology and potential for in vitro cultivation.

An imine-functionalized silicone-based epoxy coating with stable adhesion and controllable degradation for enhanced marine antifouling and anticorrosion properties.

Biofouling and corrosion of submerged equipment caused by marine organisms severely restrict the rapid development of the marine industry. Traditional antifouling or anticorrosion coatings typically serve a sole purpose and exhibit limited degradability upon failure, rendering them inadequate for current demands. Herein, a novel imine-functionalized command-degradable bio-based epoxy coating (SAHPEP-DDM) with enhanced integrated antifouling and anticorrosion performances was synthesized utilizing 1,3-bis (3-aminopropyl)-1,1,3,3-tetramethyldisiloxane and syringaldehyde. Compared with commercial epoxy resins (E51-DDM) and polydimethylsiloxanes (PDMS), the SAHPEP-DDM coating exhibits superior antifouling and anticorrosion properties due to the existence of -C=N- and Si-O-Si chain segments in the cross-linking network. The coating shows promising resistance against bacteria, algae and proteins, as well as excellent corrosion resistance in artificial seawater. The coating also exhibits excellent chemical resistance in organic solvents as well as neutral and alkaline environments. Moreover, its controlled degradation after failure can be achieved in acid aqueous solutions through temperature and acidity adjustments, facilitated by the presence of -C=N-. This work presents a novel degradable coating successfully coupled the dual functions of antifouling and anticorrosion coatings, avoiding the employment of intermediate coat, indicating vast potential for application in marine engineering fields.

α-(N-Alkyl-N-heteroarenium)-α-diazoacetates: synthesis and reactivity of a novel class of 'onium' diazo compounds.

Treatment of alkyl α-(N-heteroaryl)-α-diazoacetates with alkylating reagents affords diazoacetate N-heteroarenium salts. These novel 'onium' diazo compounds are mostly yellow solids, displaying increased thermal and acid stability. Their tetrafluoroborates undergo rhodium catalyzed [2 + 1] and Doyle-Kirmse reactions under mild conditions, suggesting the N-quaternization an effective means of elimination of N-coordination caused catalyst toxicity.

Dual Antiplatelet Therapy and Outcomes in Acute Mild to Moderate Stroke With Versus Without Large-Artery Atherosclerosis Post Hoc Analysis of ATAMIS.

BACKGROUND: We conducted a post hoc analysis of the ATAMIS (Antiplatelet Therapy in Acute Mild to Moderate Ischemic Stroke) trial to investigate whether the priority of clopidogrel plus aspirin to aspirin alone was consistent between patients with and without stroke pathogenesis of large-artery atherosclerosis (LAA). METHODS AND RESULTS: Patients with stroke classification randomized to a clopidogrel-plus-aspirin group and aspirin-alone group in a modified intention-to-treat analysis set of ATAMIS were classified into LAA and non-LAA subtypes. The primary outcome was early neurologic deterioration at 7 days, defined as a >2-point increase in National Institutes of Health Stroke Scale score compared with baseline, and safety outcomes were bleeding events and intracranial hemorrhage. We compared treatment effects in each stroke subtype and investigated the interaction. Among 2910 patients, 225 were assigned into the LAA subtype (119 in the clopidogrel-plus-aspirin group and 106 in the aspirin-alone group) and 2685 into the non-LAA subtype (1380 in the clopidogrel-plus-aspirin group and 1305 in the aspirin-alone group). Median age was 66 years, and 35% were women. A lower proportion of early neurologic deterioration was found to be associated with dual antiplatelet therapy in the LAA subtype (adjusted risk difference, -10.4% [95% CI, -16.2% to -4.7%]; P=0.001) but not in the non-LAA subtype (adjusted risk difference, -1.4% [95% CI, -2.6% to 0.1%]; P=0.06). No significant interaction was found (P=0.11). CONCLUSIONS: Compared with the non-LAA subtype, patients with stroke of the LAA subtype may get more benefit from dual antiplatelet therapy with clopidogrel plus aspirin with respect to early neurologic deterioration at 7 days. REGISTRATION: URL: clinicaltrials.gov; UnIque identifier: NCT02869009.

Bioinspired Functional Composites for Enhanced Thermally Conductivity via Fractal-Growth CuNP Fillers.

Thermal conduction for electronic devices has attracted extensive attention in light of the development of 5G communication. Thermally conductive materials with high thermal conductivity and extensive mechanical flexibility are extremely desirable in practical applications. However, the construction of efficient interconnected conductive pathways and continuous conductive networks is inadequate for either processing or actual usage in existing technologies. In this work, spherical copper nanoparticles (S-CuNPs) and urchin-inspired fractal-growth CuNPs (U-CuNPs), thermally conductive metal fillers induced by ionic liquids, were fabricated successfully through the electrochemical deposition method. Compared to S-CuNPs, the U-CuNPs shows larger specific surface contact area, thus making it easier to build a continuous conductive pathway network in the corresponding U-CuNPs/liquid silicone rubber (LSR) thermally conductive composites. The optimal loading of CuNP fillers was determined by evaluating the rheological performance of the prepolymer and the mechanical properties and thermal conductivity performances of the composites. When the filler loading is 150 phr, the U-CuNPs/LSR produces optimal mechanical properties (e.g., tensile strength and modulus), thermal conductivity (above 1000% improvement compared to pure LSR), and heating/cooling efficiency. The enhanced thermal conductivity of U-CuNPs/LSR was also confirmed through the finite element analysis (FEA) overall temperature distribution, indicating that U-CuNPs with larger specific surface contact areas exhibit more advantages in forming a continuous network in composites than S-CuNPs, making U-CuNPs/LSR a promising and competitive alternative to traditional flexible thermally interface materials.

[Drafâ ¢ approach to unilateral frontal sinus for contralateral drainage].

Objective:Anatomical variation or scar atresia of the drainage channel of the frontal sinus on the affected side, and opening the frontal sinus through the drainage channel of the frontal sinus on the affected side may lead to surgical failure. The purpose of this study is to explore a modified Draf Ⅲ operation to complete the drainage of the affected frontal sinus by removing the floor wall and septum of the frontal sinus and connecting the bilateral frontal sinus through the healthy side of the frontal sinus. Methods:Through the anatomical study of 2 skull bone specimens and 2 fresh frozen specimens, the surgical landmark and surgical approach were explored. Four patients with frontal sinus atresia and frontal sinusitis after DrafⅡb surgery in Eye & ENT Hospital of Fudan University were retrospectively analyzed. Descriptive method was used to analyze the data. Results:The bottom wall of bilateral frontal sinus was removed, and the bilateral frontal sinus was enlarged above the nasal septum to form a large common cavity. The uncinate process and ethmoid bubble were retained, and the midline drainage of the affected frontal sinus in the healthy side of the nasal cavity was completed. From August 2022 to April 2023, 4 patients with frontal sinus atresia and frontal sinusitis after DrafⅡb surgery for unilateral frontal sinus papilloma in Eye & ENT Hospital of Fudan University were treated with surgery. The headache symptoms disappeared after surgery, and the drainage of frontal sinus was spacious, the mucosa healed well and the drainage was unobstructed under endoscopy. There were no other postoperative complications. Conclusion:DrafⅢ approach to unilateral frontal sinus for contralateral drainage can drain the affected frontal sinus adequately. The essence of this operation is to drain the bilateral frontal sinus in the unilateral nasal cavity, and this operation has short path, less trauma, and a broader prospect, which is suitable for promotion.

Effects of ankle Kinesio taping on knee and ankle joint biomechanics during unanticipated jumps in collegiate athletes.

OBJECTIVE: Most biomechanical research on the application of Kinesio taping (KT) to the ankle joint focused on testing anticipated movements. However, ankle sprains frequently occur in real life in unanticipated situations, where individuals are unprepared and face sudden external stimuli. This situation is completely different from the anticipated situation. The aim of the present study was to investigate the effects of ankle KT application on the kinematic and kinetic characteristics of the knee and ankle joints during unanticipated jump tasks in collegiate athletes. METHODS: Eighteen healthy collegiate athletes experienced three taping conditions in a randomized order: no taping (NT), placebo taping (PT), and KT, and performed unanticipated jump tasks. A 9-camera infrared high-speed motion capture system was employed to collect knee and ankle kinematic data, and a 3-dimensional force plate was utilized to collect knee and ankle kinetic data during the tasks. RESULTS: During the right jumps, KT significantly increased peak knee flexion angle (P = 0.031) compared to NT and significantly decreased peak vertical ground reaction force (P < 0.001, P = 0.001) compared to NT and PT. During the left jumps, KT significantly reduced peak ankle inversion angle (P = 0.022, P < 0.001) and peak ankle inversion moment (P = 0.002, P = 0.001) compared to NT and PT. CONCLUSION: During unanticipated jump maneuvers, KT reduced peak ankle inversion angle, peak vertical ground reaction force, and peak ankle inversion moment and increased peak knee flexion angle in collegiate athletes.

Circular RNA SNX27 Facilitates Gastric Cancer Progression By Sponging miR-638.

BACKGROUND/AIMS:  Accumulating evidences have shown an important role of circular RNAs (circRNAs) in the tumorigenesis of gastric cancer (GC). Nevertheless, whether circSNX27 plays a role in GC remains undetermined. MATERIALS AND METHODS:  Relative expression of circRNAs and related microRNAs (miRNAs) in GC tissues and cells were tested by quantitative reverse transcription polymerase chain reaction. Specific short hairpin RNAs were designed to knockdown the expression of circSNX27 in GC cells. CCK-8, colony formation, flow cytometry, wound healing, and transwell assays were used to access the function of circSNX27 silencing on GC cells. The target miRNAs of circSNX27 were predicted by 2 databases, circBank and Circinteractome. Dualluciferase reporter assay was used to verify the interaction between circSNX27 and miR-638. RESULTS:  circSNX27 was found to be upregulated in GC tissues and cell lines compared with normal controls. Silencing of circSNX27 repressed GC cell viability, proliferation, migration, and invasion. Moreover, circSNX27 silencing could accelerate GC cell apoptosis. Additionally, we found that circSXN27 decreased the expression of miR-638 by directly binding to it in GC cells. CONCLUSION:  Our results indicated that circSXN27 facilitated GC progression by acting as a sponge of miR-638.

Survival Analysis and Prognostic Factors After Endonasal Resection of Advanced Olfactory Neuroblastomas: A Single Institution Experience.

OBJECTIVES: To explore the prognostic factors in patients with advanced olfactory neuroblastoma (ONB) underwent endoscopic surgery. MATERIALS AND METHODS: Retrospective medical records were reviewed of patients with pathologically proven ONB who underwent endoscopic surgical resection. Clinicopathological characteristics including patient demographics, treatment, complications, follow-up, and outcomes were analyzed. Kaplan-Meier overall survival (OS) and disease-free survival (DFS) curves were plotted. Univariate and multivariate Cox regression models were used to determine prognostic factors. RESULTS: Eighty-five patients with Kadish stage C ONB were examined. According to the various staging systems used, most patients harbored modified Kadish stage C (78.8%). Twenty-six patients (30.6%) underwent bony skull base resection, 11 (12.9%) underwent dura resection, and 24 (28.2%) underwent additional intracranial resection that included the olfactory bulb and duct. Median follow-up was 39 months. Five-year OS and DFS rates were 83.7% and 74.9%, respectively. Five-year OS was 100% in patients treated with bony skull base resection and 77.5% in those who were not (P = .052). Dura resection did not improve OS. Multivariate Cox regression analysis identified perioperative complications (P = .009), gross total resection (P = .004), orbital invasion (P = .014), postoperative radiotherapy (P = .030), and bony skull base resection (P = .019) as independent prognostic predictors. CONCLUSION: For patients with advanced ONB, endoscopic surgery in conjunction with radiotherapy and chemotherapy is effective and safe. Dura resection should be performed with caution in selected patients to balance survival and complications. Postoperative radiotherapy is important to improve OS and DFS.

Long COVID patients' brain activation is suppressed during walking and severer symptoms lead to stronger suppression.

This research aims to study the factors contributing to Long COVID and its effects on motor and cognitive brain regions using population surveys and brain imaging. The goal is to provide new insights into the neurological effects of the illness and establish a basis for addressing neuropsychiatric symptoms associated with Long COVID. Study 1 used a cross-sectional design to collect data on demographic characteristics and factors related to Long COVID symptoms in 551 participants. In Study 2, subjects with Long COVID and SARS-CoV-2 uninfected individuals underwent fNIRS monitoring while performing various tasks. Study 1 found that gender, age, BMI, Days since the first SARS-CoV-2 infection, and Symptoms at first onset influenced Long COVID performance. Study 2 demonstrated that individuals in the SARS-CoV-2 uninfected group exhibited greater activation of cognitive function-related brain regions than those in the Long COVID group while performing a level walking task. Furthermore, individuals in the Long COVID group without functional impairment displayed higher activation of brain regions associated with motor function during a weight-bearing walking task than those with functional impairment. Among individuals with Long COVID, those with mild symptoms at onset exhibited increased activation of brain regions linked to motor and cognitive function relative to those with moderate symptoms at onset. Individuals with Long COVID exhibited decreased activation in brain regions associated with cognitive and motor function compared to SARS-CoV-2 uninfected individuals. Moreover, those with more severe initial symptoms or functional impairment displayed heightened inhibition in these brain regions.

Advancing precision medicine in gliomas through single-cell sequencing: unveiling the complex tumor microenvironment.

Glioblastoma (GBM) displays an infiltrative growth characteristic that recruits neighboring normal cells to facilitate tumor growth, maintenance, and invasion into the brain. While the blood-brain barrier serves as a critical natural defense mechanism for the central nervous system, GBM disrupts this barrier, resulting in the infiltration of macrophages from the peripheral bone marrow and the activation of resident microglia. Recent advancements in single-cell transcriptomics and spatial transcriptomics have refined the categorization of cells within the tumor microenvironment for precise identification. The intricate interactions and influences on cell growth within the tumor microenvironment under multi-omics conditions are succinctly outlined. The factors and mechanisms involving microglia, macrophages, endothelial cells, and T cells that impact the growth of GBM are individually examined. The collaborative mechanisms of tumor cell-immune cell interactions within the tumor microenvironment synergistically promote the growth, infiltration, and metastasis of gliomas, while also influencing the immune status and therapeutic response of the tumor microenvironment. As immunotherapy continues to progress, targeting the cells within the inter-tumor microenvironment emerges as a promising novel therapeutic approach for GBM. By comprehensively understanding and intervening in the intricate cellular interactions within the tumor microenvironment, novel therapeutic modalities may be developed to enhance treatment outcomes for patients with GBM.

Risk factors for mortality in patients receiving extracorporeal membrane oxygenation.

OBJECTIVE: Patients on extracorporeal membrane oxygenation (ECMO) are often complex and have a high mortality rate. Currently, risk assessment and treatment decisions for patients receiving ECMO are controversial. Therefore, we sought to identify risk factors for mortality in patients receiving ECMO and provide a reference for patient management. METHODS: We retrospectively analyzed the clinical data of 199 patients who received ECMO support from December 2013 to April 2023. Univariate and multivariable logistic regression analyses were used to identify risk factors. The cutoff value was determined by receiver operating characteristic (ROC) curve analysis. RESULTS: A total of 199 patients were selected for this study, and the mortality rate was 76.38%. More than half of the patients underwent surgery during hospitalization. Multivariable logistic regression analysis revealed that continuous renal replacement therapy (CRRT) implantation (OR = 2.994; 95% CI, 1.405-6.167; p = 0.004) and age (OR = 1.021; 95% CI, 1.002-1.040; p = 0.032) were the independent risk factors for mortality. In the ROC curve analysis, age had the best predictive effect (AUC 0.646, 95% CI 0.559-0.732, p = 0.003) for death when the cutoff value was 48.5 years. Furthermore, in patients receiving combined CRRT and ECMO, lack of congenital heart disease and previous surgical history were the independent risk factors for mortality. CONCLUSIONS: CRRT implantation and age were independent risk factors for patients with ECMO implantation in a predominantly surgical cohort. In patients receiving a combination of CRRT and ECMO, lack of congenital heart disease and previous surgical history were independent risk factors for mortality.

BET degrader exhibits lower antiproliferative activity than its inhibitor via EGR1 recruiting septins to promote E2F1-3 transcription in triple-negative breast cancer.

The bromodomain and extraterminal domain (BET) family proteins serve as primary readers of acetylated lysine residues and play crucial roles in cell proliferation and differentiation. Dysregulation of BET proteins has been implicated in tumorigenesis, making them important therapeutic targets. BET-bromodomain (BD) inhibitors and BET-targeting degraders have been developed to inhibit BET proteins. In this study, we found that the BET inhibitor MS645 exhibited superior antiproliferative activity than BET degraders including ARV771, AT1, MZ1 and dBET1 in triple-negative breast cancer (TNBC) cells. Treatment with MS645 led to the dissociation of BETs, MED1 and RNA polymerase II from the E2F1-3 promoter, resulting in the suppression of E2F1-3 transcription and subsequent inhibition of cell growth in TNBC. In contrast, while ARV771 displaced BET proteins from chromatin, it did not significantly alter E2F1-3 expression. Mechanistically, ARV771 induced BRD4 depletion at protein level, which markedly increased EGR1 expression. This elevation of EGR1 subsequently recruited septin 2 and septin 9 to E2F1-3 promoters, enhancing E2F1-3 transcription and promoting cell proliferation rate in vitro and in vivo. Our findings provide valuable insights into differential mechanisms of BET inhibition and highlight potential of developing BET-targeting molecules as therapeutic strategies for TNBC.

A colorimetric assay for Neospora caninum utilizing the loop-mediated isothermal amplification technique.

Neospora caninum (N. caninum) is a protozoan parasite that poses a serious risk to livestock by infecting various domestic and wild animals. Loop-Mediated Isothermal Amplification (LAMP) offers a cost-effective, highly sensitive, and specific method for detecting protozoan parasites. This study aims to develop a precise, rapid, and visually assessable colorimetric LAMP method, improving on traditional techniques. We employed a rigorous screening process to identify the optimal primer set for this experiment. Subsequently, we fine-tuned the LAMP reaction at 65 °C for 40 min with 270 µmol/L neutral red. We then confirmed the specificity of primers for N. caninum through experimental validation. The LAMP method demonstrated a lower detection limit compared to traditional Polymerase Chain Reaction (PCR) techniques. While LAMP offers clear advantages, the prevalence of DNA detected in 89 sheep serum and 59 bovine serum samples using the nested PCR method was 3.37 % (3/89) and 1.69 % (1/59), respectively. In contrast, when the LAMP method was employed, the prevalence of detected DNA rose to 5.61 % (5/89) for sheep and 3.38 % (2 /59) for bovine. A comparison of two molecular assays using the intragroup correlation coefficient (ICC) resulted in a value of 0.999 (95 % CI: 0.993-0.996, p < 0.001), indicating the LAMP method is in the "better" range according to James Lee's categorization. The LAMP technique, optimized with specific primers of N. caninum and neutral red dye, not only exhibited higher sensitivity but also provided convenience over conventional PCR methods, highlighting its potential for on-site applications and cost-effective field detection.

Paospora carinifang n. gen., n. sp. (Microsporidia: Spragueidae), a parasite of the ridgetail white prawn, Palaemon carinicauda.

A new microsporidian disease of the pond-reared ridgetail white prawn, Palaemon carinicauda, was found in China. Light microscopy, pathology, and scanning electron microscopy showed that the parasite infected the host's skeletal muscle tissue and formed spherical sporophorous vesicles (SPOVs). Electron microscopy revealed that its merogonic life stages developed in direct contact with the host cytoplasm. The sporogonic life stages underwent octosporoblastic sporogony with the formation of eight uninucleate spores in each SPOV. Fresh SPOVs were 5.4 ± 0.55 µm in diameter. The octospores were oval and measured 2.3 × 1.5 µm (fresh) and 1.96 × 1.17 µm (fixed). The isofilar polar filament was coiled with 9-10 turns and arranged in two rows. Phylogenetic analysis based on the SSU rRNA gene suggests that this microsporidium has close affinities with members of the genera Potaspora and Apotaspora, but represents an independent generic taxon. We therefore propose the establishment of a new genus and species (Paospora carinifang n. gen., n. sp.) within the family Spragueidae. We also propose a taxonomic revision to transfer Potaspora macrobrachium to this new genus and reclassify it as Paospora macrobrachium comb. nov.

Rapid identification of antibody impurities in size-based electrophoresis via CZE-MS generated spectral library.

Methods for the reliable and effective detection and identification of impurities are crucial to ensure the quality and safety of biopharmaceutical products. Technical limitations constrain the accurate identification of individual impurity peaks by size-based electrophoresis separations followed by mass spectrometry. This study presents a size-based electrophoretic method for detecting and identifying impurity peaks in antibody production. A hydrogen sulfide-accelerated degradation method was employed to generate known degradation products observed in bioreactors that forms the basis for size calibration. LabChip GXII channel electrophoresis enabled the rapid (< 1 min) detection of impurity peaks based on size, while capillary zone electrophoresis-mass spectrometry (CZE-MS) facilitated their accurate identification. We combine these techniques to examine impurities resulting from cell culture harvest conditions and forced degradation to assess antibody stability. To mimic cell culture harvest conditions and the impact of forced degradation, we subjected samples to cathepsin at different pH buffers or exposed them to high pH and temperature. Our method demonstrated the feasibility and broad applicability of using a CZE-MS generated spectral library to unambiguously assign peaks in high throughput size-based electrophoresis (i.e., LabChip GXII) with identifications or likely mass of the antibody impurity. Overall, this strategy combines the utility of CZE-MS as a high-resolution separation and detection method for impurities with size-based electrophoresis methods that are typically used to detect (not identify) impurities during the discovery and development of antibody therapeutics.