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Notch signaling can have either an oncogenic or tumor suppressive function in cancer depending on the cancer type and cellular context. While Notch can be oncogenic in early prostate cancer, we identified significant downregulation of the Notch pathway during prostate cancer progression from adenocarcinoma to neuroendocrine prostate cancer where it functions as a tumor suppressor. Activation of Notch in neuroendocrine and Rb1/Trp53-deficient prostate cancer models led to phenotypic conversion towards a more indolent non-neuroendocrine state with glandular features and expression of luminal lineage markers. This was accompanied by up-regulation of MHC and type I interferon and immune cell infiltration. Overall, these data support Notch signaling as a suppressor of neuroendocrine differentiation in advanced prostate cancer and provides insights into how Notch signaling influences lineage plasticity and the tumor microenvironment.
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Forsythiaside A (FA) is an active constituent isolated from Forsythia suspensa, a beneficial herb used in traditional medicine known for its antioxidant and anti-inflammatory properties. Although various studies have suggested that FA has the protective effects, its impacts on arachidonic acid (AA) plus iron in vitro models and carbon tetrachloride (CCl4)-induced mouse liver damage in vivo have not been explored. In this study, HepG2 cells were subjected to AA + iron treatment to induce apoptosis and mitochondrial impairment and determine the molecular mechanisms. FA exhibited protective effects by inhibiting cell damage and reactive oxygen species (ROS) production induced by AA + iron, as assessed via immunoblot and flow cytometry analyses. Further molecular investigations revealed that FA resulted in the activation of extracellular-signal-related protein kinase (ERK), which subsequently triggered the activation of AMP-activated protein kinase (AMPK), a critical regulator of cellular oxidative stress. Additionally, FA modulated the nuclear factor erythroid 2-related factor 2 (Nrf2) signaling pathway, which is a significant antioxidant transcription factor regulated by the AMPK pathway. For in vivo studies, mice were orally administered FA and then subjected to induction of CCl4-based hepatotoxicity. The protective effect of FA was confirmed via blood biochemistry and immunohistochemical analyses. In conclusion, our findings demonstrated the protective effects of FA against oxidative stress both in vitro and in vivo, thus indicating that FA is a potential candidate for liver protection. Our study sheds light on the mechanistic pathways involved in the antioxidant effects of FA, highlighting the hepatoprotective potential of naturally occurring compounds in traditional herbs, such as FA.
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Proteínas Quinasas Activadas por AMP , Antioxidantes , Animales , Ratones , Proteínas Quinasas Activadas por AMP/metabolismo , Antioxidantes/farmacología , Antioxidantes/metabolismo , Factor 2 Relacionado con NF-E2/metabolismo , Estrés Oxidativo , Transducción de Señal , Especies Reactivas de Oxígeno/metabolismo , Quinasas MAP Reguladas por Señal Extracelular/metabolismo , Hierro/farmacologíaRESUMEN
Single-target rapid antigen tests (RATs) are commonly used to detect highly transmissible respiratory viruses (RVs), such as severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and influenza viruses. The simultaneous detection of RVs presenting overlapping symptoms is vital in making appropriate decisions about treatment, isolation, and resource utilization; however, few studies have evaluated multiplex RATs for SARS-CoV-2 and other RVs. We assessed the diagnostic performance of multiplex RATs targeting both the SARS-CoV-2 and influenza A/B viruses with the GenBody Influenza/COVID-19 Ag Triple, InstaView COVID-19/Flu Ag Combo (InstaView), STANDARDTM Q COVID-19 Ag Test, and STANDARDTM Q Influenza A/B Test kits using 974 nasopharyngeal swab samples. The cycle threshold values obtained from the real-time reverse transcription polymerase chain reaction results showed higher sensitivity (72.7-100%) when the values were below, rather than above, the cut-off values. The InstaView kit exhibited significantly higher positivity rates (80.21% for SARS-CoV-2, 61.75% for influenza A, and 46.15% for influenza B) and cut-off values (25.57 for SARS-CoV-2, 21.19 for influenza A, and 22.35 for influenza B) than the other two kits, and was able to detect SARS-CoV-2 Omicron subvariants. Therefore, the InstaView kit is the best choice for routine screening for both SARS-CoV-2 and influenza A/B in local communities.
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Recent studies have highlighted the positive effects of Kaempferol (KP), including its anti-inflammatory and antioxidant properties. However, its impact on oxidative damage induced by heavy metals and pro-inflammatory mediators, such as arachidonic acid (AA), has not yet been identified. Our objective was to specifically evaluate liver damage due to AA + iron-induced oxidative stress, both in vitro and in vivo. In HepG2 cells, KP activated the AMP-activated protein kinase (AMPK), suggesting a hepatoprotective effect through AMPK inhibition, as assessed by immunoblot and FACS analysis (EC50 = 10 µM). KP also stimulated autophagy, a degradation process that eliminates aged, damaged, and unnecessary components, via mTOR inhibition and ULK1 phosphorylation. This activation was further validated by the upregulation of autophagy-related genes (ATG5) and Beclin-1, along with the conversion of LC3BI to LC3BII. Ferroptosis, a non-apoptotic type of cell death characterized by oxidative stress from the production of reactive oxygen species (ROS) and excessive iron accumulation, was linked to the activation of autophagy, as confirmed through the protein expression of deferoxamine (DFO) and ferrostatin-1 (Fer-1), the representative ferroptosis inhibitors (positive controls). In mice, oral administration of KP demonstrated protective effects against CCl4-induced hepatotoxicity. In conclusion, KP provides hepatoprotective effects against oxidative stress induced by AA + iron treatment in vitro and CCl4 treatment in vivo.
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Proteínas Quinasas Activadas por AMP , Ferroptosis , Ratones , Animales , Proteínas Quinasas Activadas por AMP/genética , Proteínas Quinasas Activadas por AMP/metabolismo , Quempferoles/farmacología , Estrés Oxidativo , Especies Reactivas de Oxígeno/metabolismo , Ácido Araquidónico/farmacología , Autofagia , HierroRESUMEN
Ten new norterpene alkaloids, coscinoderines A-J (1-10), were isolated from the marine sponge Coscinoderma bakusi. Each coscinoderine contains a 1,2,5-trisubstituted pyridinium moiety bearing a terpene unit at the C-2 position. Their structures were elucidated by analysis of NMR and HRMS data, and the absolute stereochemistry of 4 with a 2-methylbutyl group attached to the nitrogen was determined from a comparison of the calculated and measured ECD spectra. The isolation of coscinoderines expands the repertoire of pyridinium alkaloids isolated from marine sponges.
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Alcaloides , Poríferos , Animales , Poríferos/química , Alcaloides/farmacología , Alcaloides/química , Espectroscopía de Resonancia Magnética , Terpenos , Estructura MolecularRESUMEN
Although coronavirus disease 2019 (COVID-19) is no longer a Public Health Emergency of International Concern (PHEIC), severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection has had a vast impact to date. Hence, continuous management is required, given the uncertainty caused by the potential evolution of SARS-CoV-2. Reverse transcription-quantitative PCR (RT-qPCR) diagnosis has been fundamental in overcoming this issue. In this study, the performances of two rapid RT-qPCR assays (Real-Q Direct SARS-CoV-2 Detection Kit and Allplex™ SARS-CoV-2 fast PCR Assay) with short PCR times were comparatively evaluated using a STANDARD M nCoV Real-Time Detection Kit (STANDARD M, conventional RT-qPCR assay). All kits showed a limit of detection values (102-103 copies/reaction). The evaluation showed that the two rapid assay tests had ≥97.89% sensitivity and ≥99.51% specificity (κ = 0.98) for individual samples and ≥97.32% sensitivity and ≥97.67% specificity for pooled samples compared to STANDARD M. These results indicate that the two rapid RT-qPCR kits, which showed significant time reduction in performance, are as effective as a conventional RT-qPCR assay. They are likely to increase not only the number of tests that can be performed but also the efficiency of sustainable management of COVID-19 in the long term.
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Dielectric layers are widely used in ferroelectric applications such as memory and negative capacitance devices. The wake-up and the split-up phenomena in the ferroelectric hafnia are well-known challenges in early-stage device reliability. We found that the phenomena even occur in the bilayer, which is composed of the hafnia and the dielectrics. The phenomena are known to be affected mainly by oxygen vacancies of hafnia. Dielectric layers, which are often metal oxides, are also prone to be affected by oxygen vacancies. To study the effect of the dielectric layer on the wake-up and the split-up phenomena, we fabricated ferroelectric thin-film capacitors with dielectric layers of various thicknesses and measured their field-cycling behaviors. We found that the movement of oxygen vacancies in the dielectric layer was predominantly affected by the polarization state of the ferroelectric layer. In addition, the mechanism of the field-cycling behavior in the bilayer is similar to that in ferroelectric thin films. Our results can be applied in ferroelectric applications that use dielectric layers.
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This study evaluated the effects of Rorippa cantoniensis (Lour.) ohwi extract (RCE) on factors associated with inflammation-related skin lesions in RAW 264.7 and HaCaT cells. RCE inhibited the levels of proinflammatory mediators and cytokines such as nitric oxide (NO), prostaglandin E2 (PGE2), interleukin (IL)-6, and tumor necrosis factor (TNF)-α in lipopolysaccharide (LPS)-stimulated RAW 264.7 cells. In addition, RCE significantly inhibited the expression of chemokines and cytokines such as MDC/CCL22, TARC/CCL17, RANTES/CCL5, CTSS, IL-6, IL-1ß, and TNF-α in HaCaT cells costimulated by TNF-α and interferon (IFN)-γ in a concentration-dependent manner. These results suggest that RCE attenuated the TNF-α- and IFN-γ-induced release of proinflammatory chemokines and cytokines probably by suppressing the activation of MAPK (JNK and p38), NF-κB, and STAT1 signaling. Moreover, RCE significantly increased the expression of skin components such as hyaluronic acid and aquaporin, which play important roles in the physical and chemical barriers of the skin. These results suggest that RCE has significant anti-inflammatory and antiatopic activities, which may be beneficial for the topical treatment of inflammatory skin disorders.
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Células HaCaT , Rorippa , Animales , Ratones , Humanos , Rorippa/metabolismo , Factor de Necrosis Tumoral alfa/metabolismo , Queratinocitos , Línea Celular , Citocinas/metabolismo , Antiinflamatorios/farmacología , Antiinflamatorios/metabolismo , FN-kappa B/metabolismo , Quimiocinas/metabolismo , Células RAW 264.7RESUMEN
Nonalcoholic fatty liver disease (NAFLD) is a global health problem that is closely associated with obesity and metabolic syndrome. Spatholobi caulis (SC) is a herbal medicine with potential hepatoprotective effects; however, its active compounds and underlying mechanisms have not been fully explored. In this study, we combined a multiscale network-level approach with experimental validation to investigate SC's antioxidant properties and their impact on NAFLD. Data collection and network construction were performed, and active compounds and key mechanisms were identified through multi-scale network analysis. Validation was conducted using in vitro steatotic hepatocyte models and in vivo high-fat diet-induced NAFLD models. Our findings revealed that SC treatment improved NAFLD by modulating multiple proteins and signaling pathways, including AMPK signaling pathways. Subsequent experiments showed that SC treatment reduced lipid accumulation and oxidative stress. We also validated SC's effects on AMPK and its crosstalk pathways, emphasizing their role in hepatoprotection. We predicted procyanidin B2 to be an active compound of SC and validated it using a lipogenesis in vitro model. Histological and biochemical analyses confirmed that SC ameliorated liver steatosis and inflammation in mice. This study presents SC's potential use in NAFLD treatment and introduces a novel approach for identifying and validating active compounds in herbal medicine.
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Owing to the high transmissibility of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants, the capacity of testing systems based on the gold standard real-time reverse transcription-polymerase chain reaction (rRT-PCR) is limited. Rapid antigen tests (RATs) can substantially contribute to the prevention of community transmission, but their further assessment is required. Here, using 1503 nasopharyngeal swabs, we compared the diagnostic performance of four RAT kits (Abbott Panbio™ COVID-19 Ag Rapid Test, SD Biosensor Standard™ Q COVID-19 Ag Test, Humasis COVID-19 Ag Test, and SG Medical Acrosis COVID-19 Ag Test) to the cycle threshold (Ct) values obtained from rRT-PCR. The precision values, area under the curve values, SARS-CoV-2 variant detection ability, and non-SARS-CoV-2 specificity of all four kits were similar. An assay using the Acrosis kit had a significantly better positive detection rate with a higher recall value and cut-off value than that using the other three RAT kits. During the current COVID-19 pandemic, the Acrosis kit is an effective tool to prevent the spread of SARS-CoV-2 in communities.
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We aimed to describe injury incidence and patterns at the 2019 World Taekwondo Championships (WTC), and to compare them with those of previous WTCs, based on new World Taekwondo (WT) competition rules, medical codes, and the Protector and Scoring System (PSS). This prospective cohort study utilized data obtained through the WT Injury Surveillance System. All athletes with injuries were evaluated by on-site sports medicine specialists, and ultrasonography was used to assess all musculoskeletal injuries. Of 936 athletes, 60 injuries were recorded (6.4 injuries/100 athletes, 95% confidence intervals [CI]: 4.8-8.0), and 4.5% (n = 42) sustained at least one injury. Males had a higher risk of sustaining injuries than females (incidence rate ratio: 1.57; 95% CI: 0.89-2.76). The most common sites, type, and mechanism were lower extremities (n = 26, 43.33%), contusion/hematoma/bruises (n = 33, 55.0%), and contact with another athlete (n = 50, 83.33%). Overall, the injury patterns associated with the mechanism of injury were similar in both the 2019 and 2017 WTCs. Refined WT competition rules and a re-established PSS at the 2019 WTC reduced the overall and severe injury incidence. Our findings can help revise Taekwondo competition rules, enhance protective equipment, optimize on-site venue medical systems, and develop injury prevention projects.
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Traumatismos en Atletas , Artes Marciales , Masculino , Femenino , Humanos , Estudios Prospectivos , Incidencia , Traumatismos en Atletas/epidemiología , AtletasRESUMEN
The World Health Organization recently highlighted the serious worldwide problem of the emergence of antibiotic-resistant or antibiotic multidrug-resistant bacteria. Carbapenem-resistant Enterobacterales, including carbapenemase-producing Enterobacterales (CPE), are major antibiotic-resistant bacteria that can be identified by various methods, including antibiotic susceptibility testing, PCR, and immunologic assays. However, there is a need for a faster, more accurate, low-cost, and easy method to detect CPE strains. We previously developed an osmotic shock matrix-assisted laser desorption/ionization mass spectrometry (OS-MALDI MS) method for directly detecting intact Klebsiella pneumoniae carbapenemase (KPC) using osmotic shock cell lysis. In this study, we evaluated the OS-MALDI MS method and compared it with two other methods (octyl-glucoside-aided direct KPC detection method [OG-MALDI MS] and Bruker's MBT subtyping module indirect method [MBT-SM MALDI MS]). We first completed an analytical performance evaluation of the OS-MALDI MS method according to Clinical and Laboratory Standards Institute guidelines. Clinical testing was performed with 437 clinical isolates, including 292 KPC-producing bacteria and 145 non-KPC-producing bacteria. The OS-MALDI MS method exhibited 95.9% sensitivity, 100.0% specificity, and 100.0% precision for detecting KPC. Accuracy of the OS-MALDI MS, OG-MALDI MS, and MBT-SM MALDI MS methods was 97.3%, 55.9%, and 50.2%, respectively. In conclusion, the OS-MALDI MS method clearly outperformed the other methods, exhibiting the highest accuracy and sensitivity of the three methods. We propose the OS-MALDI MS method as a practical, useful method for clinic environments, which may help guide appropriate antibiotic treatment and contribute to the prevention of the spread of CPE.
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Klebsiella pneumoniae , beta-Lactamasas , Humanos , Espectrometría de Masa por Láser de Matriz Asistida de Ionización Desorción/métodos , Presión Osmótica , Proteínas Bacterianas , Antibacterianos/farmacología , Pruebas de Sensibilidad MicrobianaRESUMEN
Eighteen scalarane sesterterpenoids (1-18), including eight new derivatives (1-8), were isolated from the sponge Hyrtios erectus (family Thorectidae), the extract of which showed cytotoxicity against the HeLa and MCF-7 cell lines. Of the new derivatives, six compounds (1-6) were found to contain a γ-hydroxybutenolide moiety capable of reversible stereoinversion at the hydroxylated carbon center. Under the influence of other adjacent functional groups, each derivative exhibited a different stereochemical behavior, which was fully deduced by ROESY experiments. All the isolated compounds were examined for their cytotoxicity by MTS assay using staurosporine as a positive control (IC50 0.18 and 0.13 µΜ against HeLa and MCF-7 cells, respectively), and they were found to show weak growth inhibitory activities against HeLa and MCF-7 cells, with a minimal IC50 value of 20.0 µΜ. The compounds containing a γ-hydroxybutenolide moiety (1-3, 10, 12) showed cytotoxicity, with IC50 values ranging from 24.3 to 29.9 µΜ, and the most potent derivative was heteronemin (16). Although the cytotoxicities of isolated compounds were insufficient to discuss the structure-activity relationship, this research could contribute to expanding the structural diversity of scalaranes and understanding the stereochemical behavior of γ-hydroxybutenolides.
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Antineoplásicos , Poríferos , Animales , Humanos , Estaurosporina , Poríferos/química , Células MCF-7 , Relación Estructura-Actividad , Carbono , Estructura Molecular , Sesterterpenos/farmacología , Sesterterpenos/química , Antineoplásicos/farmacología , Antineoplásicos/química , Ensayos de Selección de Medicamentos AntitumoralesRESUMEN
Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the third highly pathogenic human coronavirus and is rapidly transmitted by infected individuals regardless of their symptoms. During the COVID-19 pandemic, owing to the dearth of skilled healthcare workers (HCWs) to collect samples for early diagnosis, self-collection emerged as a viable alternative. To evaluate the reliability of self-collection, we compared the virus detection rate using 3990 self-collected swabs and HCW-collected swabs, procured from the same individuals and collected immediately after the self-collection. The results of multiplex reverse-transcription quantitative polymerase chain reaction revealed that the viral load in the HCW-collected swabs was marginally (18.4-28.8 times) higher than that in self-collected swabs. Self-collection showed no significant difference in sensitivity and specificity from HCW-collection (κ = 0.87, McNemar's test; p = 0.19), indicating a comparable performance. These findings suggest that self-collected swabs are acceptable substitutes for HCW-collected swabs, and that their use improved the specimen screening efficiency and reduced the risk of SARS-CoV-2 infection among HCWs during and after the COVID-19 pandemic.
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Sports-related traumatic brain injuries are the most common injury in adolescents and young adults due to recurrent concussion experiences and head shock. Therefore, this study was designed to describe player characteristics and situational factors associated with concussions in the World Taekwondo Championships using systematic video analysis. Athlete injury data were collected using a web-based injury surveillance system at the World Taekwondo Championships organized by World Taekwondo from 2017 to 2019. Seven video footage were independently analyzed by four analysts using a modified Heads-Up Checklist. Descriptive statistical analysis was used. The incidence of concussion was 3.21 per 1000 games. Most players with concussions were shorter than their opponents, and most concussions were caused by a roundhouse kick on the front of the face. Regarding the acceleration direction of the head after the impact, transverse and multiplane directions were the most common. Most players with a concussion have used a closed stance and did not use blocking techniques during the defense. The rate of concussions caused by penalties was 42.9%. Based on our findings, no other injury mechanisms, except for direct blows to the head, were observed. Therefore, education on the risk and symptoms of concussion, the appropriate management and blocking techniques should be emphasized in TKD-S to reduce incidence of concussion.
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Traumatismos en Atletas , Conmoción Encefálica , Artes Marciales , Adolescente , Traumatismos en Atletas/etiología , Conmoción Encefálica/diagnóstico , Humanos , IncidenciaRESUMEN
BACKGROUND: The effectiveness of operative treatments other than neurectomy for Morton's neuroma remains debatable despite several reported studies. This review aimed to evaluate the effects of operative treatments for Morton's neuroma other than neurectomy using an algorithmic approach and a structured critical framework to assess the methodological quality of reported studies. METHODS: Several electronic databases were searched for articles published until August 2021 that evaluated the outcomes of operative treatments other than neurectomy in patients diagnosed with Morton's neuroma. Data searches, extraction, analysis, and quality assessments were performed according to the Preferred Reporting Items for Systematic Reviews and Meta-Analyses guidelines, and the clinical outcomes were evaluated using objective, subjective, and negative outcomes; complications; and reoperation rate and type. RESULTS: After reviewing 11,213 studies, 22 studies were finally included. Although a number of studies with high level of evidence are limited, we divided them according to four categories: (1) neurolysis with or without nerve transposition, (2) minimally invasive nerve decompression, (3) metatarsal osteotomy, and (4) additional procedures after nerve transection or neurectomy. All categories showed reliable outcomes except minimally invasive nerve decompression. The proportion of postoperative neurogenic symptoms was lower with neurolysis than with neurectomy. CONCLUSION: Whether the alternative procedures were superior to neurectomy remains unclear as the number of good quality studies was limited. The proportion of postoperative neurogenic symptoms was lower with neurolysis than with neurectomy. Furthermore, performing simultaneous dorsal transposition of the nerve along with neurolysis is more recommended than neurolysis alone. Surgeons should be more careful with minimally invasive deep transverse intermetatarsal ligament release and metatarsal shortening osteotomy as their effectiveness remains inconclusive. Finally, we strongly recommend performing intramuscular embedding or intermuscular transposition of the nerve cutting end if neurectomy or nerve transection is inevitable. LEVEL OF EVIDENCE: Level III, systematic review.
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Enfermedades del Pie , Neuroma de Morton , Neuroma , Desnervación/efectos adversos , Pie , Enfermedades del Pie/cirugía , Humanos , Neuroma de Morton/cirugía , Neuroma/etiología , Neuroma/cirugía , Osteotomía/métodosRESUMEN
Background and Objectives: Nontuberculous mycobacteria (NTM) infections are increasing worldwide. We evaluated the annual trends of NTM isolation on acid-fast bacillus (AFB) culture, compared to that of Mycobacterium tuberculosis, and the distribution of NTM species nationwide in South Korea. Materials and Methods: The study was conducted in a diagnostic center that is a major referral laboratory for the diagnosis of tuberculosis and NTM in South Korea. All laboratory results of AFB culture from January 2014 to December 2019 were collected. All NTM identified were definitively identified to the species level. Results: A total of 345,871 tests were performed for the diagnosis of mycobacteria. The isolation rate of NTM and M. tuberculosis was 3.7% (12,969 cases) and 4.4% (15,081 cases), respectively. The annual isolation rate of NTM increased gradually from 2.7% in 2014 to 4.8% in 2019, whereas that of M. tuberculosis decreased from 6.2% to 3.3%. There were 4988 cases of NTM identified to the species level. M. avium complex (MAC) was the most common species isolated from pulmonary sites (59.8%), followed by M. gordonae (9.2%), M. abscessus (7.0%), and M. fortuitum (5.5%). Extrapulmonary NTM were identified in 29 cases (0.6%). MAC was also the most common NTM species isolated from extrapulmonary sites (65.5%), followed by M. kansasii (10.3%), M. abscessus (6.9%), and M. fortuitum (6.9%). Conclusions: The annual isolation rate of NTM has increased gradually, whereas that of M. tuberculosis has decreased. Follow-up studies of the increases in NTM identification and NTM infections in South Korea are needed.
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Infecciones por Mycobacterium no Tuberculosas , Micobacterias no Tuberculosas , Humanos , Laboratorios , Infecciones por Mycobacterium no Tuberculosas/diagnóstico , Infecciones por Mycobacterium no Tuberculosas/epidemiología , Derivación y Consulta , República de Corea/epidemiologíaRESUMEN
PURPOSE: Clinical mass spectrometry requires a simple step process for sample preparation. This study aims to optimize the method for isolating periplasmic protein from Gram-negative bacteria and apply to clinical mass spectrometry. EXPERIMENTAL DESIGN: The Klebsiella pneumoniae carbapenemase (KPC)-producing E. coli standard cells were used for optimizing the osmotic shock (OS) lysis method. The supernatant from OS lysis was analysed by LC-MS/MS and MALDI-TOF MS. The effectiveness of the OS lysis method for KPC-2-producing Enterobacteriaceae clinical isolates were then confirmed by MALDI-TOF MS. RESULTS: The optimized OS lysis using KPC-2 producing E. coli standard cells showed a high yield of KPC-2 protein and enriches periplasmic proteins. Compared with other lysis methods, the detection sensitivity of KPC-2 protein significantly increased in MALDI-TOF MS analysis. Nineteen clinical isolates were validated by MALDI-TOF MS using the OS method, which also showed higher detection sensitivity compared to other lysis method (e.g., 1.5% n-octyl-ß-D-glucopyranoside) (p < 0.001). CONCLUSIONS AND CLINICAL RELEVANCE: This study provides a straightforward, rapid, affordable, and detergent-free method for the analysis of periplasmic proteins from Enterobacteriaceae clinical isolates. This approach may contribute to MS-based clinical diagnostics.
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Escherichia coli/metabolismo , Proteínas Periplasmáticas/análisis , Cromatografía Líquida de Alta Presión , Escherichia coli/enzimología , Escherichia coli/aislamiento & purificación , Presión Osmótica , Proteínas Periplasmáticas/aislamiento & purificación , Proteínas Periplasmáticas/metabolismo , Cloruro de Sodio/química , Espectrometría de Masa por Láser de Matriz Asistida de Ionización Desorción , beta-Lactamasas/metabolismoRESUMEN
Classification of clinical symptoms and diagnostic microbiology are essential to effectively employ antimicrobial therapy for lower respiratory tract infections (LRTIs) in a timely manner. Empirical antibiotic treatment without microbial identification hinders the selective use of narrow-spectrum antibiotics and effective patient treatment. Thus, the development of rapid and accurate diagnostic procedures that can be readily adopted by the clinic is necessary to minimize non-essential or excessive use of antibiotics and accelerate patient recovery from LRTI-induced damage. We developed and validated a multiplex real-time polymerase chain reaction (mRT-PCR) assay with good analytical performance and high specificity to simultaneously detect four bacterial pathogens causing pneumonia: Klebsiella pneumoniae, Pseudomonas aeruginosa, Staphylococcus aureus, and Moraxella catarrhalis. The analytical performance of mRT-PCR against target pathogens was evaluated by the limit of detection (LOD), specificity, and repeatability. Two hundred and ten clinical specimens from pneumonia patients were processed using an automatic nucleic acid extraction system for the "respiratory bacteria four" (RB4) mRT-PCR assay, and the results were directly compared to references from bacterial culture and/or Sanger sequencing. The RB4 mRT-PCR assay detected all target pathogens from sputum specimens with a coefficient of variation ranging from 0.29 to 1.71 and conservative LOD of DNA corresponding to 5 × 102 copies/reaction. The concordance of the assay with reference-positive specimens was 100%, and additional bacterial infections were detected from reference-negative specimens. Overall, the RB4 mRT-PCR assay showed a more rapid turnaround time and higher performance that those of reference assays. The RB4 mRT-PCR assay is a high-throughput and reliable tool that assists decision-making assessment and outperforms other standard methods. This tool supports patient management by considerably reducing the inappropriate use of antibiotics.
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Klebsiella pneumoniae/aislamiento & purificación , Moraxella catarrhalis/aislamiento & purificación , Neumonía/diagnóstico , Pseudomonas aeruginosa/aislamiento & purificación , Staphylococcus aureus/aislamiento & purificación , Humanos , Reacción en Cadena de la Polimerasa Multiplex/métodos , Neumonía/microbiología , Sensibilidad y EspecificidadRESUMEN
OBJECTIVE: To compare the shear bond strength (SBS) after thermocycling of four universal adhesives applied in self-etch mode on dentin and etch-and-rinse mode on enamel. STUDY DESIGN: Flat 144 buccal or lingual dentin and enamel surfaces from 72 non-carious primary molars were prepared. Samples were segregated into 12 groups (n=12): Adper Single Bond 2 etch-and-rinse (SB_T) and Clearfil SE Bond self-etch (SE_S) applied to enamel and dentin served as controls. Scotch Bond Universal Adhesive (SBU), Clearfil S3 Bond Universal Adhesive (SEU), Tetric N-Bond Universal Adhesive (TEN), and All Bond Universal (BIS) were applied in etch-and-rinse mode to enamel and in self-etch mode to dentin. They were thermocycled for 5000 cycles. SBS testing and the evaluation of fracture mode were performed. RESULTS: SB_T showed statistically higher SBS than other adhesive groups using etch-and-rinse mode on enamel. SE_S and BIS had statistically higher SBS than other adhesive groups using self-etch mode on dentin. Mixed failure was the most common failure mode in each group. CONCLUSION: The universal adhesives did not show higher SBS than SB_T when using etch-and-rinse on enamel. All universal adhesives showed higher SBS than SB_T and had SBS similar to SE_S, except SBU when using self-etch mode on dentin.