Clinical outcomes and treatment necessity in patients with toxin-negative Clostridioides difficile stool samples.

PURPOSE: The clinical significance of negative toxin enzyme immunoassays (EIA) for Clostridioides difficile infections (CDIs) is unclear. Our study aimed to investigate the significance of toxin EIA-negative in the diagnosis and prognosis of CDI. METHODS: All stool specimens submitted for C. difficile toxin EIA testing were cultured to isolate C. difficile. In-house PCR for tcdA, tcdB, cdtA, and cdtB genes were performed using C. difficile isolates. Stool specimens were tested with C. difficile toxins A and B using EIA kit (RIDASCREEN Clostridium difficile toxin A/B, R-Biopharm AG, Darmstadt, Germany). Characteristics and subsequent CDI episodes of toxin EIA-negative and -positive patients were compared. RESULTS: Among 190 C. difficile PCR-positive patients, 83 (43.7%) were toxin EIA-negative. Multivariate analysis revealed independent associations toxin EIA-negative results and shorter hospital stays (OR = 0.98, 95% CI 0.96-0.99, p = 0.013) and less high-risk antibiotic exposure in the preceding month (OR = 0.38, 95% CI 0.16-0.94, p = 0.035). Toxin EIA-negative patients displayed a significantly lower white blood cell count rate (11.0 vs. 35.4%, p < 0.001). Among the 54 patients who were toxin EIA-negative and did not receive CDI treatment, three (5.6%) were diagnosed with CDI after 7-21 days without complication. CONCLUSION: Our study demonstrates that toxin EIA-negative patients had milder laboratory findings and no complications, despite not receiving treatment. Prolonged hospitalisation and exposure to high-risk antibiotics could potentially serve as markers for the development of toxin EIA-positive CDI.

Ineffectiveness of colistin monotherapy in treating carbapenem-resistant Acinetobacter baumannii Pneumonia: A retrospective single-center cohort study.

BACKGROUND: Acinetobacter baumannii, a common carbapenem-resistant gram-negative bacillus, usually causes nosocomial infections. Colistin has been used for carbapenem-resistant A. baumannii (CRAB) infections; however, only a few studies have evaluated colistin as a treatment option compared to appropriate controls. We investigated the effectiveness of colistin monotherapy in treating CRAB pneumonia compared to those treated without an active drug. METHODS: Adult patients (≥ 18 years) with CRAB isolated from respiratory specimens were screened from September 2017 to August 2022. Only patients with pneumonia treated with colistin monotherapy (colistin group) were included and compared to those without any active antibiotics (no active antibiotics [NAA] group). The primary and secondary outcomes were 30-day all-cause mortality and acute kidney injury within 30 days. The inverse probability of the treatment-weighted Cox proportional hazard model was used to compare mortality between groups. RESULTS: Among the 826 adult patients with CRAB in their respiratory specimens, 45 and 123 patients were included in the colistin and NAA groups, respectively. Most of the CRAB pneumonia (91.1%) cases were hospital-acquired pneumonia. The 30-day all-cause mortality rates in the colistin and NAA groups were 58.3% and 56.1%, respectively, and no difference was observed after adjustments (adjusted hazard ratio, 0.74; 95% CI, 0.47-1.17). The incidence of acute kidney injury was higher in the colistin group (65.3%) compared to the NAA group (39.0%) (P = 0.143). CONCLUSIONS: Colistin monotherapy did not significantly improve treatment outcomes for CRAB pneumonia. The development and evaluation of new antimicrobials for CRAB pneumonia should be advocated in clinical practice.

Bacterium secretes chemical inhibitor that sensitizes competitor to bacteriophage infection.

To overtake competitors, microbes produce and secrete secondary metabolites that kill neighboring cells and sequester nutrients. This natural product-mediated competition likely evolved in complex microbial communities that included viral pathogens. From this ecological context, we hypothesized that microbes secrete metabolites that "weaponize" natural pathogens (i.e., bacteriophages) to lyse their competitors. Indeed, we discovered a bacterial secondary metabolite that sensitizes other bacteria to phage infection. We found that this metabolite provides the producer (a Streptomyces sp.) with a fitness advantage over its competitor (Bacillus subtilis) by promoting phage infection. The phage-promoting metabolite, coelichelin, sensitized B. subtilis to a wide panel of lytic phages, and it did so by preventing the early stages of sporulation through iron sequestration. Beyond coelichelin, other natural products may provide phage-mediated competitive advantages to their producers-either by inhibiting sporulation or through yet-unknown mechanisms.

Neurotrophic phenolic glycosides from the roots of Armoracia rusticana.

Armoracia rusticana P. G. Gaertner. belongs to the Brassicaceae family and has aroused scientific interest for its anti-inflammatory and anticancer activities. In a continuing investigation to discover bioactive constituents from A. rusticana, we isolated 19 phenolic glycosides including three undescribed flavonol glycosides and one undescribed neolignan glycoside from MeOH extract of this plant. Their structures were elucidated based on NMR spectroscopic analysis (1H, 13C, 1H-1H COSY, HSQC, and HMBC), HRESIMS, and chemical methods. The determination of their absolute configuration was accomplished by ECD and LC-MS analysis. All the compounds were assessed for their potential neurotrophic activity through induction of nerve growth factor in C6 glioma cell lines and for their anti-neuroinflammatory activity based on the measurement of inhibition levels of nitric oxide production and pro-inflammatory cytokines (i.e., IL-1ß, IL-6, and TNF-α) in lipopolysaccharide-activated microglia BV-2 cells.

Evaluation of Liftover Tools for the Conversion of Genome Reference Consortium Human Build 37 to Build 38 Using ClinVar Variants.

Although Genome Reference Consortium Human Build 38 (GRCh38) was released with improvement over GRCh37, it has not been widely adopted. Several liftover tools have been developed as a convenient approach for GRCh38 implementation. This study aimed to investigate the accuracy of liftover tools for genome conversion. Two Variant Call Format (VCF) files aligned to GRCh37 and GRCh38 were downloaded from ClinVar (clinvar_20221217.vcf.gz). Liftover tools such as CrossMap, NCBI Remap, and UCSC liftOver were used to convert genome coordinates from GRCh37 to GRCh38. The accuracy of CrossMap, NCBI Remap, and UCSC liftOver were 99.81% (1,567,838/1,570,748), 99.69% (1,565,953/1,570,748), and 99.99% (1,570,550/1,570,748), respectively. Variants that failed conversion via all three liftover tools were all indels/duplications: a pathogenic/likely pathogenic variant (n = 1) and benign/likely benign variants (n = 7). The eight variants that failed conversion were identified in the ALMS, TTN, CFTR, SLCO, LDLR, PCNT, MID1, and GRIA3 genes, and all the variants were not in the VCF files aligned to GRCh37. This study demonstrated that three liftover tools could successfully convert reference genomes from GRCh37 to GRCh38 in more than 99% of ClinVar variants. This study takes the first step to clinically implement GRCh38 using liftover tools. Further clinical studies are warranted to compare the performance of liftover tools and to validate re-alignment approaches in routine clinical settings.

Ravidomycin Analogs from Streptomyces sp. Exhibit Altered Antimicrobial and Cytotoxic Selectivity.

Six new ravidomycin analogs (1-4, 6, and 7) were isolated from Streptomyces sp. Am59 using UV- and LCMS-guided separation based on Global Natural Products Social (GNPS) molecular networking analysis. Furthermore, we isolated fucomycin V (9), which possesses the same chromophore as ravidomycin but features a d-fucopyranose instead of d-ravidosamine. This is the first report of 9 as a natural product. Four new analogs (10-13) of 9 were also isolated. The structures were elucidated by combined spectroscopic and computational methods. We also found an inconsistency with the published [α]D25 of deacetylravidomycin, which is reported to have a (-) sign. Instead, we observed a (+) specific rotation for the reported absolute configuration of deacetylravidomycin (containing d-ravidosamine). We confirmed the positive sign by reisolating deacetylravidomycin from S. ravidus and by deacetylating ravidomycin. Finally, antibacterial, antifungal, and cytotoxicity activities were determined for the compounds. Compared to deacetylravidomycin, the compounds 4-6, 9, 11, and 12 exhibited greater antibacterial selectivity.

Neurotrophic neolignans of Pinus koraiensis twigs.

Four undescribed neolignan analogs, together with eight known compounds, were isolated from the twigs of Pinus koraiensis (Korean pine). The chemical structure of the isolated compounds was determined through extensive spectroscopic analysis and chemical method. Their relative and absolute configurations were assigned through a well-established empirical rule and electronic circular dichroism (ECD) analysis, respectively. Four compounds (3 and 9-11) at 20 µM concentration showed significant neurotrophic effect by inducing nerve growth factor (NGF) secretion in C6 cells with the stimulation levels a range of 140.82 ± 4.62% to 160.04 ± 11.04%. Additionally, the result indicated that the glycosylation of neolignan led to an improvement in neurotrophic activity compared to their aglycone form. A compound (7) inhibited nitric oxide production with an IC50 value of 31.74 µM in LPS-activated BV2 cells.

Structure Revision of Anti-Inflammatory Indole Alkaloids with a 1,2-Benzisoxazole Ring.

(R)- and (S)-2-(benzo[d]isoxazol-3-yl)-2-ethylindolin-3-one [(±)-1] were previously isolated from NIRAM, a natural blue dye from Polygonum tinctorium, and their structures were initially proposed to possess a 1,2-benzisoxazole ring. In this study, the structures of (±)-1 were revised to have an indole-anthranilic acid fused tetracyclic ring rather than the 1,2-benzisoxazole ring by reanalysis of one-dimensional (1D) and two-dimensional (2D) NMR followed by density functional theory (DFT) chemical shift calculation, DP4+ technique, and ECD simulation.

Associations between apolipoprotein B/A1 ratio, lipoprotein(a), and the risk of metabolic-associated fatty liver diseases in a Korean population.

OBJECTIVE: Metabolic-associated fatty liver disease (MAFLD) is new nomenclature for the fatty liver condition associated with metabolic dysfunction. This study aimed to investigate the association between apolipoprotein B/A1 (apo B/A1) ratio, lipoprotein(a), and MAFLD in a Korean population. METHODS: This study consisted of 14,419 subjects in the Korean population. Multivariate logistic regression was conducted to analyze the association between apo B/A1 ratio and MAFLD. RESULTS: The prevalence of MAFLD in the general Korean population was 34.5%. The apo B/A1 ratio (odds ratio: 3.913, P = .019) was independently associated with MAFLD. Lipoprotein(a) was significantly lower in patients with MAFLD with hepatic fibrosis (P < .0001). CONCLUSION: Apolipoprotein B/A1 ratio and lipoprotein(a) have opposite associations with MAFLD. This study suggests that lipoprotein(a) should be used with caution as a biomarker for MAFLD, especially in patients with hepatic fibrosis.

Anti-Inflammatory, Neurotrophic, and Cytotoxic Oxylipins Isolated from Chaenomeles sinensis Twigs.

Oxylipins are important biological molecules with diverse roles in human and plants such as pro-/anti-inflammatory, antimicrobial, and regulatory activity. Although there is an increasing number of plant-derived oxylipins, most of their physiological roles in humans remain unclear. Here, we describe the isolation, identification, and biological activities of four new oxylipins, chaenomesters A-D (1-4), along with a known compound (5), obtained from Chaenomeles sinensis twigs. Their chemical structures were determined by spectroscopic (i.e., NMR) and spectrometric (i.e., HRMS) data analysis including 1H NMR-based empirical rules and homonuclear-decoupled 1H NMR experiments. Chaenomester D (4), an omega-3 oxylipin, showed a potent inhibitory effect on nitric oxide (NO) production in lipopolysaccharide (LPS)-activated BV-2 cells (NO production, 8.46 ± 0.68 µM), neurotrophic activity in C6 cells through the induction of the secretion of nerve growth factor (NGF, 157.7 ± 2.4%), and cytotoxicity in A549 human cancer cell lines (IC50 = 27.4 µM).

A Metabolite Produced by Gut Microbes Represses Phage Infections in Vibrio cholerae.

Vibrio cholerae is the causative agent of the severe diarrheal disease cholera. Bacteriophages that prey on V. cholerae may be employed as phage therapy against cholera. However, the influence of the chemical environment on the infectivity of vibriophages has been unexplored. Here, we discovered that a common metabolite produced by gut microbes─linear enterobactin (LinEnt), represses vibriophage proliferation. We found that the antiphage effect by LinEnt is due to iron sequestration and that multiple forms of iron sequestration can protect V. cholerae from phage predation. This discovery emphasizes the significance that the chemical environment can have on natural phage infectivity and phage-based interventions.

Variations in Nomenclature of Clinical Variants between Annotation Tools.

BACKGROUND: Accurate nomenclature of variants is an essential element for genetic diagnosis and patient care. OBJECTIVE: To investigate annotation differences of clinical variants between annotation tools. METHODS: We analyzed 218,156 clinical variants from the Human Gene Mutation Database. Multiple nomenclatures based on RefSeq transcripts were provided using ANNOVAR and snpEff. RESULTS: The concordance rate between ANNOVAR and snpEff was approximately 85%. Based on the Human Genome Variation Society (HGVS) nomenclature, snpEff was more accurate than ANNOVAR (coding variants, 99.3% vs 84.9%; protein variants, 94.3% vs 79.8%). When annotating each variant with ANNOVAR and snpEff, the accuracy of nomenclature was 99.5%. CONCLUSIONS: There were substantial differences between ANNOVAR and snpEff annotations. The findings of this study suggest that simultaneous use of multiple annotation tools could decrease nomenclature errors and contribute to providing standardized clinical reporting.

Clinical Interpretation Challenges of Germline-Shared Somatic Variants in Cancer.

OBJECTIVE: To investigate the interpretation differences of germline-shared somatic variants. METHODS: A total of 123,302 COSMIC variants associated with hematologic malignant neoplasms were used. The pathogenicity and actionability of shared variants were analyzed based on the standardized guidelines. RESULTS: The overall frequency of variants shared in ClinVar/HGMD and COSMIC was 10%. The pathogenicity of 54 shared variants was pathogenic/likely pathogenic (P/LP; n = 30), variants of unknown significance (n = 3), and benign/likely benign (n = 21). In total, 30 P/LP variants were reclassified to tier I/tier II (83%) and tier III (17%) variants. CONCLUSIONS: This is the first study about different clinical interpretations of shared variants based on the current standard guidelines. This study takes a meaningful step in bridging the interpretation gap between the somatic and germline variants.

Phenolic constituents from twigs of Aleurites fordii and their biological activities.

Three new neolignan glycosides (1-3), a new phenolic glycoside (15), and a new cyanoglycoside (16) were isolated and characterized from the twigs of Aleurites fordii together with 14 known analogues (4-14 and 17-19). The structural elucidation of the new compounds was performed through the analysis of their NMR, HRMS, and ECD spectra and by chemical methods. All isolated compounds were tested for their antineuroinflammatory and neuroprotective activities.

Inhibitory Effect of LGS and ODE Isolated from the Twigs of Syringa oblata subsp. dilatata on RANKL-Induced Osteoclastogenesis in Macrophage Cells.

Osteoblasts and osteoclasts play a pivotal role in maintaining bone homeostasis, of which excessive bone resorption by osteoclasts can cause osteoporosis and various bone diseases. However, current osteoporosis treatments have many side effects, and research on new treatments that can replace these treatments is ongoing. Therefore, in this study, the roles of ligustroside (LGS) and oleoside dimethylester (ODE), a natural product-derived compound isolated from Syringa oblata subsp. dilatata as a novel, natural product-derived osteoporosis treatments were investigated. In the results of this study, LGS and ODE inhibited the differentiation of receptor activator of nuclear factor kappa-Β ligand (RANKL)-induced RAW264.7 cells into osteoclasts without cytotoxicity, and down-regulated the activity of TRAP, a specific biomarker of osteoclasts. In addition, it inhibited bone resorption and actin ring formation, which are important functions and features of osteoclasts. Also, the effects of LGS and ODE on the mitogen-activated protein kinase (MAPK) and nuclear factor kappa-light-chain-enhancer of activated B (NF-κB) and phosphoinositide 3-kinases (PI3K)/ protein kinase B (Akt)/mechanistic target of rapamycin (mTOR) signaling pathways that play important roles in osteoclast differentiation were evaluated. In the results, LGS and ODE downregulated the phosphorylation of RANKL-induced MAPK and PI3K/Akt/mTOR proteins in a concentration-dependent manner, translocation of NF-κB into the nucleus was inhibited. As a result, the compounds LGS and ODE isolated from S. oblate subsp. dilatata effectively regulated the differentiation of RANKL-induced osteoclasts and inhibited the phosphorylation of signaling pathways that play a pivotal role in osteoclast differentiation. Therefore, these results suggest the possibility of LGS and ODE as new natural product treatments for bone diseases caused by excessive osteoclasts.

Calcific tendinitis of the shoulder in the Korean population: demographics and its relation with coexisting rotator cuff tear.

BACKGROUND: To evaluate the demographics, clinical and radiographic features of calcific tendinitis of the shoulder in the Korean population, specifically focusing on the incidence of coexisting rotator cuff tear. METHODS: Between October 2014 and January 2015, we performed a prospective multicenter study with 506 patients from 11 training hospitals in Korea. We collected data of demographics and radiographic analysis based on simple radiographs, clinical assessments based on visual analog scale (VAS) and the American Shoulder Elbow Surgeons (ASES) score, and treatment modalities that are used currently. We also evaluated coexisting rotator cuff tear by ultrasonography (US) or magnetic resonance imaging (MRI) images. RESULTS: There were 402 female patients (79%) with mean age of 55 years (range, 31-87 years). Mean duration of symptoms was 16 months. Mean size of calcific materials was 11.4 mm (range, 0-35 mm). Mean value of VAS and ASES scores were 6.5 (range, 1-10) and 47 (range, 8-95), respectively. Of 383 patients (76%), 59 (15%) had rotator cuff tear including 15 full-thickness tears on US or MRI. Patients with rotator cuff tears were significantly associated with older age, recurrent symptoms, menstrual disorders in females, and having undergone calcification removal surgery and rotator cuff repair (all p<0.05). CONCLUSIONS: This study reported demographic, radiographic, and clinical features of calcific tendinitis of the shoulder in Korean population, which were not different from those of Western population. Coexisting rotator cuff tear was found with 15% incidence in this large series, suggesting that further radiographic study to evaluate rotator cuff tear might be needed in some calcific tendinitis patients of older age and presenting with recurrent symptoms.

The Frequency of Discordant Variant Classification in the Human Gene Mutation Database: A Comparison of the American College of Medical Genetics and Genomics Guidelines and ClinVar.

OBJECTIVE: Discordant variant classifications among public databases is one of the well-documented limitations when interpreting the pathogenicity of variants. The aim of this study is to investigate the level of germline variant misannotation from the Human Gene Mutation Database (HGMD) and the annotation concordance between databases. METHODS: We used a total of 188,106 classified variants (disease-causing mutations [n = 179,454] and polymorphisms [n = 8652]) in 6466 genes from the HGMD. All variants were reanalyzed based on the American College of Medical Genetics and Genomics (ACMG) guidelines and compared to ClinVar database variants. RESULTS: When variants were classified based on the ACMG guidelines, misclassification was observed in 3.47% (2289/65,896) of variants. The overall concordance between HGMD and ClinVar was 97.62% (52,499/53,780) of variants studied. CONCLUSION: Variants in databases must be used with caution when variant pathogenicity is interpreted. This study reveals the frequency of misannotation of the HGMD variants and annotation concordance between databases in depth.

Morolic Acid 3-O-Caffeate Inhibits Adipogenesis by Regulating Epigenetic Gene Expression.

Obesity causes a wide range of metabolic diseases including diabetes, cardiovascular disease, and kidney disease. Thus, plenty of studies have attempted to discover naturally derived compounds displaying anti-obesity effects. In this study, we evaluated the inhibitory effects of morolic acid 3-O-caffeate (MAOC), extracted from Betula schmidtii, on adipogenesis. Treatment of 3T3-L1 cells with MAOC during adipogenesis significantly reduced lipid accumulation and decreased the expression of adiponectin, a marker of mature adipocytes. Moreover, the treatment with MAOC only during the early phase (day 0-2) sufficiently inhibited adipogenesis, comparable with the inhibitory effects observed following MAOC treatment during the whole processes of adipogenesis. In the early phase of adipogenesis, the expression level of Wnt6, which inhibits adipogenesis, increased by MAOC treatment in 3T3-L1 cells. To identify the gene regulatory mechanism, we assessed alterations in histone modifications upon MAOC treatment. Both global and local levels on the Wnt6 promoter region of histone H3 lysine 4 trimethylation, an active transcriptional histone marker, increased markedly by MAOC treatment in 3T3-L1 cells. Our findings identified an epigenetic event associated with inhibition of adipocyte generation by MAOC, suggesting its potential as an efficient therapeutic compound to cure obesity and metabolic diseases.

Antineurodegenerative Labdane Diterpenoid Glycosides from the Twigs of Pinus koraiensis.

Eleven new labdane-type diterpenoid glycosides, koraiensides A-K (1-11), together with two known analogues were isolated from the twigs of Pinus koraiensis. Their structures were elucidated via NMR, HRMS, and ECD data, DP4+ statistical analysis, and hydrolysis. The metabolites were tested for induction of nerve growth factor in C6 glioma cells to evaluate their potential neuroprotective activity. The compounds were measured for production of nitric oxide levels in lipopolysaccharide (LPS)-activated murine microglia BV2 cells to assess their antineuroinflammatory activity. Compounds 10 and 13 showed NGF secretion inducing effects from C6 glioma cells (162.3 ± 13.9% and 162.7 ± 6.9%, respectively). Compound 6 showed an IC50 value of 24.1 µM, implying significant inhibition of NO production.

Mutation analysis in Korean patients with T-cell acute lymphoblastic leukemia.

Genomic studies have illuminated the alterations in pathways underlying T-cell acute lymphoblastic leukemia (T-ALL) pathogenesis, but detailed mutation data by next-generation sequencing have not been reported in Korean patients. We aimed to investigate mutation frequency, spectrum, and pattern in the Korean patients with T-ALL. We designed a multigene panel targeting 101 genes and validated it using 10 reference materials. The mutation analysis was done in a total of 10 patients with T-ALL. Clinical data and laboratory tests including immunophenotyping, cytogenetics, and molecular genetic tests were also investigated. All of the 10 patients harbored at least one mutation (range 1-6 per patient). A total of 34 clinically significant mutations including 15 novel mutations were identified in 23 genes. The median of variant allelic frequencies (VAFs) and blasts were counted upto 33% (range 5-91%) and 79% (range 38-90%), respectively. Recurrent mutations were involved in epigenetic regulators (60%), NOTCH1 signaling (40%), PI3K-AKT (40%), JAK-STAT (30%), and transcription factors (30%). We found that both NOTCH signaling and JAK-STAT signaling were positively associated with epigenetic regulators, while showed mutually exclusive patterns with PI3K-AKT pathway. This study showed that the frequency of mutations in epigenetic regulators in Korean patients was significantly higher than expected. Distribution of VAF as well as mutation spectrum is considerably heterogeneous in Korean patients with T-ALL. Although from a limited number of patients, this study provides the first detailed mutational portrait of T-ALL of Korean patients, and gives additional insight into molecular pathogenesis of the disease.