Antibiotic Combination to Effectively Postpone or Inhibit the In Vitro Induction and Selection of Levofloxacin-Resistant Mutants in Elizabethkingia anophelis.

Fluoroquinolones are potentially active against Elizabethkingia anophelis. Rapidly increased minimum inhibitory concentrations (MICs) and emerging point mutations in the quinolone resistance-determining regions (QRDRs) following exposure to fluoroquinolones have been reported in E. anophelis. We aimed to investigate point mutations in QRDRs through exposure to levofloxacin (1 × MIC) combinations with different concentrations (0.5× and 1 × MIC) of minocycline, rifampin, cefoperazone/sulbactam, or sulfamethoxazole/trimethoprim in comparison with exposure to levofloxacin alone. Of the four E. anophelis isolates that were clinically collected, lower MICs of levofloxacin were disclosed in cycle 2 and 3 of induction and selection in all levofloxacin combination groups other than levofloxacin alone (all p = 0.04). Overall, no mutations were discovered in parC and parE throughout the multicycles inducted by levofloxacin and all its combinations. Regarding the vastly increased MICs, the second point mutations in gyrA and/or gyrB in one isolate (strain no. 1) occurred in cycle 2 following exposure to levofloxacin plus 0.5 × MIC minocycline, but they were delayed appearing in cycle 5 following exposure to levofloxacin plus 1 × MIC minocycline. Similarly, the second point mutation in gyrA and/or gyrB occurred in another isolate (strain no. 3) in cycle 4 following exposure to levofloxacin plus 0.5 × MIC sulfamethoxazole/trimethoprim, but no mutation following exposure to levofloxacin plus 1 × MIC sulfamethoxazole/trimethoprim was disclosed. In conclusion, the rapid selection of E. anophelis mutants with high MICs after levofloxacin exposure could be effectively delayed or postponed by antimicrobial combination with other in vitro active antibiotics.

Dictamnine Ameliorates DNFB-Induced Atopic Dermatitis Like Skin Lesions in Mice by Inhibiting M1 Macrophage Polarization and Promoting Autophagy.

Autophagy and M1 macrophage polarization play important roles in the regulation of inflammation in atopic dermatitis (AD). Dictamnine is one of the main ingredients in Cortex Dictamni, a widely used traditional Chinese medicine for the treatment of dermatitis. In the present study, we investigated the anti-inflammatory effects of dictamnine on AD like skin lesions and M1 macrophage polarization. A 2,4-dinitrofluorobenzene (DNFB) triggered AD like skin lesions models in mice was established to identify the ameliorative effects of dictamnine on AD in vivo. In addition, an M1 macrophage polarization model was co-stimulated by lipopolysaccharide (LPS) and interferon-γ (IFN-γ) using phorbol myristate acetate (PMA) differentiated THP-1 cells, to investigate the effect of dictamnine on promoting autophagy and inhibiting inflammatory factor release. Dictamnine suppressed DNFB-induced skin inflammation by inhibiting M1 macrophage polarization, up-regulating the expression of microtubule-associated protein 1A/1B-light chain 3 (LC3) expression, and promoting macrophage autophagy at inflammatory sites. Dictamnine also could reduce the release of interleukin-1ß (IL-1ß), tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), monocyte chemotactic protein-1 (MCP-1), and interleukin-8 (IL-8), and down-regulate the mRNA expression of these genes in LPS-IFN-γ triggered M1 polarized macrophages. Dictamnine ameliorates AD like skin lesions by inhibiting M1 macrophage polarization and promoting autophagy. Hence, dictamnine is expected to be a potential therapeutic candidate for AD.

In Vitro and In Vivo Antimicrobial Activities of Vancomycin and Rifampin against Elizabethkingia anophelis.

Elizabethkingia anophelis has emerged as a critical human pathogen, and a number of isolated reports have described the successful treatment of Elizabethkingia infections with vancomycin, a drug that is typically used to target Gram-positive bacteria. This study employed in vitro broth microdilution checkerboard and time-kill assays, as well as in vivo zebrafish animal models to evaluate the individual and combination antimicrobial effects of vancomycin and rifampin against E. anophelis. The minimum inhibitory concentration ranges of vancomycin and rifampin against 167 isolates of E. anophelis were 16-256 mg/L and 0.06-128 mg/L, respectively. The checkerboard assay results revealed a synergistic effect between vancomycin and rifampin in 16.8% (28/167) of the isolates. Time-kill assays were implemented for 66 isolates, and the two-drug combination had a synergistic interaction in 57 (86.4%) isolates. In vivo zebrafish studies revealed that treatment with vancomycin monotherapy, rifampin monotherapy, or vancomycin-rifampin combination therapy yielded a higher survival rate than the control group treatment with 0.9% saline. The results of this study support the use of vancomycin to treat E. anophelis infections.

Validation of 16S rRNA and Complete rpoB Gene Sequence Analysis for the Identification of Elizabethkingia Species.

Bacteria in the genus Elizabethkingia have emerged as a cause of life-threatening infections in humans. However, accurate species identification of these pathogens relies on molecular techniques. We aimed to evaluate the accuracy of 16S rRNA and complete RNA polymerase ß-subunit (rpoB) gene sequences in identifying Elizabethkingia species. A total of 173 Elizabethkingia strains with whole-genome sequences in GenBank were included. The 16S rRNA gene and rpoB gene sequences from the same Elizabethkingia strains were examined. Of the 41 E. meningoseptica strains, all exhibited >99.5% 16S rRNA similarity to its type strain. Only 83% of the 99 E. anophelis strains shared >99.5% 16S rRNA gene similarity with its type strain. All strains of E. meningoseptica and E. anophelis formed a cluster distinct from the other Elizabethkingia species in the 16S rRNA and rpoB gene phylogenetic trees. The polymorphisms of 16S rRNA gene sequences are not sufficient for constructing a phylogenetic tree to discriminate species in the E. miricola cluster (E. miricola, E. bruuniana, E. occulta, and E. ursingii). The complete rpoB gene phylogenetic tree clearly delineates all strains of Elizabethkingia species. The complete rpoB gene sequencing could be a useful complementary phylogenetic marker for the accurate identification of Elizabethkingia species.

Corrigendum to "Does the CFO serving as the secretary of the board affect the financial statement comparability?--Evidence from China" [Heliyon 9(3) (February 2023) e13609].

[This corrects the article DOI: 10.1016/j.heliyon.2023.e13609.].

Fisetin alleviates chronic urticaria by inhibiting mast cell activation via MRGPRX2.

OBJECTIVES: The activation of mast cell (MC) plays an important part in the pathogenesis of chronic urticaria (CU), and the expression of MRGPRX2 (Mas-related G-protein coupled receptor X2) and the circulating levels of SP (substance P) in skin MC of CU patients increased. Fisetin is a natural flavonoid with anti-inflammatory and antiallergic pharmacological effects. This study aimed to investigate the inhibitory effect of fisetin on CU via MRGPRX2 and its possible molecular mechanisms. METHODS: OVA/SP co-stimulated and SP-stimulated CU like murine models were used to evaluate the effect of fisetin on CU. MRGPRX2/HEK293 cells and LAD2 cells were used to perform the antagonism effect of fisetin on MC via MRGPRX2. KEY FINDINGS: The results indicated that fisetin prevented urticaria-like symptoms in murine CU models, and inhibited MCs activation by suppressing calcium mobilization and degranulation of cytokines and chemokines via binding to MRGPRX2. The bioinformatics analysis showed that fisetin might have an interaction relationship with Akt in CU. The western blotting experiments showed that fisetin downregulated the phosphorylation levels of Akt, P38, NF-κB, and PLCγ in C48/80 activated LAD2 cells. CONCLUSIONS: Fisetin alleviates CU progression by inhibiting mast cell activation via MRGPRX2, which may be a novel therapeutic candidate for CU.

Discovery of Potential Protein Markers Associated with Quality Characteristics of Antarctic Krill (Euphausia superba) Surimi Gel.

Antarctic krill are a consumption resource with great exploitation potential. However, the poor gel properties of Antarctic krill meat seriously limit its high-value application. In the present study, the quality characteristics and proteome changes of the κ-/ι-carrageenan-Antarctic krill surimi gel were systematically analyzed and compared. In addition, the transcriptome sequencing of Antarctic krill was carried out, which filled the gap in the Antarctic krill database. Higher molecular forces (disulfide bond and hydrophobic interaction) and the degree of network cross-linking significantly promoted the formation of κ/ι-carrageenan-Antarctic krill surimi compared to that of Antarctic krill surimi. This is the first study to investigate and map potential protein markers for quality characteristics of Antarctic krill surimi based on mass spectrometry-based label-free quantitative proteomics. The results could provide a theoretical reference for the quality control of Antarctic krill during application.

Anti-inflammatory effect of dictamnine on allergic rhinitis via suppression of the LYN kinase-mediated molecular signaling pathway during mast cell activation.

Mast cells (MCs) are important therapeutic targets for allergic diseases. High-affinity immunoglobulin E (IgE) Fc receptors (FcεRI) trigger abnormal activation of MCs. Allergic rhinitis (AR) is an IgE-mediated antigen inhalation reaction that occurs in the nasal mucosa. MC aggravation and dysfunction were observed during the early stages of AR pathogenesis. Herb-derived dictamnine exhibits anti-inflammatory effects. Here, we investigated the pharmacological effects of herb-derived dictamnine on IgE-induced activation of MCs and an ovalbumin (OVA)-induced murine AR model. The results indicated that dictamnine attenuated OVA-induced local allergic reactions and reduced body temperature in OVA-challenged mice with active systemic anaphylaxis. Additionally, dictamnine decreased the frequency of nasal rubbing and sneezing in an OVA-induced murine AR model. Moreover, dictamnine inhibited FcεRI-activated MC activation in a dose-dependent manner without causing cytotoxicity, reduced the activation of the tyrosine kinase LYN in LAD2 cells, and downregulated the phosphorylation of PLCγ1, IP3R, PKC, Erk1/2, and Akt, which are downstream of LYN. In conclusion, dictamnine suppressed the OVA-stimulated murine model of AR and activated IgE-induced MCs via the LYN kinase-mediated molecular signaling pathway, suggesting that dictamnine may be a promising treatment for AR.

Alpha-linolenic acid improves nasal mucosa epithelial barrier function in allergic rhinitis by arresting CD4+ T cell differentiation via IL-4Rα-JAK2-STAT3 pathway.

BACKGROUND: Allergic rhinitis (AR) defined as inflammation and tissue remodeling of the nasal mucosa in atopic individuals after allergen exposure. Alpha-linolenic acid [cis-9, cis-12, cis-15-octadecatrienoic acid (18:3)] (ALA) as dietary supplementation can reduce inflammation and allergic symptoms. OBJECTIVE: To evaluate the potential therapeutic effect and mechanism of ALA in AR mouse model. METHODS: Ovalbumin sensitized AR mouse model were challenged with oral ALA administration. Nasal symptoms, tissue pathology, immune cell infiltration and goblet cell hyperplasia were investigated. Levels of IgE, TNF-ß, IFN-γ, IL-2, IL-4, IL-5, IL-12, IL-13 and IL-25 were determined by ELISA in serum and nasal fluid. Quantitative RT-PCR and immunofluorescence were performed for occludin and zonula occludens-1 expression. CD3+CD4+ T-cells from peripheral blood and splenic lymphocytes were isolated and Th1/Th2 ratio were determined. Mouse naive CD4+ T cell were isolated and Th1/Th2 ratio, IL-4Rα expression, and IL5/IL13 secretion were determined. IL-4Rα-JAK2-STAT3 pathway change in AR mice were performed by western blot. RESULTS: Ovalbumin induced AR, nasal symptoms, pathological performance, IgE, and cytokine production. ALA treated mice showed reduced nasal symptoms, nasal inflammation, nasal septum thickening, goblet cell hyperplasia, and eosinophil infiltration. In serum and nasal fluid of ovalbumin challenged mice, ALA decreased IgE, IL-4 levels, and the increase of Th2-cells. ALA prevented the disruption of the epithelial cell barrier in ovalbumin-challenged AR mice. Simultaneously, ALA prevents IL-4 induced barrier disruption. ALA treatment of AR by affecting the differentiation stage of CD4+T cells and block IL-4Rα-JAK2-STAT3 pathway. CONCLUSION: This study suggests that ALA has the potential therapeutic effect to ovalbumin-induced AR. ALA can affect the differentiation stage of CD4+T cells and improve epithelial barrier functions through IL-4Rα-JAK2-STAT3 pathways. CLINICAL IMPLICATION: ALA might be considered as drug candidate for improving epithelial barrier function through Th1/Th2 ratio recovery in AR.

Development of a Reactive Force Field for Simulating Photoinitiated Acrylate Polymerization.

Light-driven and photocurable polymer-based additive manufacturing (AM) has enormous potential due to its excellent resolution and precision. Acrylated resins that undergo radical chain-growth polymerization are widely used in photopolymer AM due to their fast kinetics and often serve as a departure point for developing other resin materials for photopolymer-based AM technologies. For successful control of the photopolymer resins, the molecular basis of the acrylate free-radical polymerization has to be understood in detail. We present an optimized reactive force field (ReaxFF) for molecular dynamics (MD) simulations of acrylate polymer resins that captures radical polymerization thermodynamics and kinetics. The force field is trained against an extensive training set including density functional theory (DFT) calculations of reaction pathways along the radical polymerization from methyl acrylate to methyl butyrate, bond dissociation energies, and structures and partial charges of several molecules and radicals. We also found that it was critical to train the force field against an incorrect, nonphysical reaction pathway observed in simulations that used parameters not optimized for acrylate polymerization. The parameterization process utilizes a parallelized search algorithm, and the resulting model can describe polymer resin formation, crosslinking density, conversion rate, and residual monomers of the complex acrylate mixtures.

Development of a Pneumatic-Driven Fiber-Shaped Robot Scaffold for Use as a Complex 3D Dynamic Culture System.

Cells can sense and respond to different kinds of continuous mechanical strain in the human body. Mechanical stimulation needs to be included within the in vitro culture system to better mimic the existing complexity of in vivo biological systems. Existing commercial dynamic culture systems are generally two-dimensional (2D) which fail to mimic the three-dimensional (3D) native microenvironment. In this study, a pneumatically driven fiber robot has been developed as a platform for 3D dynamic cell culture. The fiber robot can generate tunable contractions upon stimulation. The surface of the fiber robot is formed by a braiding structure, which provides promising surface contact and adequate space for cell culture. An in-house dynamic stimulation using the fiber robot was set up to maintain NIH3T3 cells in a controlled environment. The biocompatibility of the developed dynamic culture systems was analyzed using LIVE/DEAD™ and alamarBlue™ assays. The results showed that the dynamic culture system was able to support cell proliferation with minimal cytotoxicity similar to static cultures. However, we observed a decrease in cell viability in the case of a high strain rate in dynamic cultures. Differences in cell arrangement and proliferation were observed between braided sleeves made of different materials (nylon and ultra-high molecular weight polyethylene). In summary, a simple and cost-effective 3D dynamic culture system has been proposed, which can be easily implemented to study complex biological phenomena in vitro.

Silibinin attenuated pseudo-allergic reactions and mast cell degranulation via PLCγ and PI3K/Akt signaling pathway.

Anaphylaxis is a type of potentially fatal hypersensitivity reaction resulting from the activation of mast cells. Many endogenous or exogenous factors could cause this reaction. Silibinin is the main chemical component of silymarin and has been reported to have pharmacological activities. However, the anti-allergic reaction effect of silibinin has not yet been investigated. This study aimed to evaluate the effect of silibinin to attenuate pseudo-allergic reactions in vivo and to investigate the underlying mechanism in vitro. In this study, calcium imaging was used to assess Ca2+ mobilization. The levels of cytokines and chemokines, released by stimulated mast cells, were measured using enzyme immunoassay kits. The activity of silibinin was evaluated in a mouse model of passive cutaneous anaphylaxis (PCA). Western blotting was used to explore the related molecular signaling pathways. In results, silibinin markedly inhibited mast cell degranulation, calcium mobilization, and preventing the release of cytokines and chemokines in a dose-dependent manner via the PLCγ and PI3K/Akt signaling pathway. Silibinin also attenuated PCA in a dose-dependent manner. In summary, silibinin has an anti-pseudo-allergic pharmacological activity, which makes it a potential candidate for the development of a novel agent to arrest pseudo-allergic reactions.

Myricetin served as antagonist for negatively regulate MRGPRX2 mediated pseudo-allergic reactions through CD300f/SHP1/SHP2 phosphorylation.

BACKGROUND: Mas-related G protein-coupled receptor X2 (MRGPRX2) plays a vital role in mast cells (MCs) degranulation and pseudo-allergic reactions. Leukocyte mono-immunoglobulin-like receptor 3 (CD300f) can negatively regulate MCs degranulation. Identification of drug candidates which target CD300f represents a promising prospect in drug development. Myricetin is widely distributed in plants and has been reported to inhibit allergic reactions in OVA-induced murine models. OBJECTIVE: This study aims to determine whether myricetin can activate CD300f to arrest MCs degranulation mediated by MRGPRX2. RESULTS: Myricetin inhibited the allergic mediator and cytokine release triggered by MRGPRX2 in vivo and in vitro. Under C48/80 stimulation, the release of ß-hexosaminidase, TNF-α, IL-8 and MCP-1 in CD300f knockdown in LAD2 cells was significantly increased compared with NC-LAD2 cells. Myricetin displayed good structural affinity (KD = 7.21 × 10-5) with CD300f by SPR. Molecular docking results showed that hydrogen bonds were formed between myricetin and CD300f, indicating high binding ability (5.6653). Myricetin can upregulate the phosphorylation of SHP-1 and SHP-2 and dephosphorylation in the MRGPRX2 signaling pathway, involving PLCγ1, AKT, P38, and ERK1/2. CONCLUSION: In the present study, myricetin is identified as an exogenous ligand for CD300f, which negatively regulates MRGPRX2-mediated MCs activation via CD300f to inhibit MCs degranulation and pseudo-allergic reactions.

Does the CFO serving as the secretary of the board affect the financial statement comparability?-evidence from China.

One person serving as both CFO and board secretary is a unique institutional feature in China. Individuals in this senior executive role are responsible for not only the preparation of financial statements but also the coordination of information disclosure. We investigate the relation between a CFO serving as board secretary and financial statement comparability. We find that a CFO serving as board secretary improves the comparability of a firm's financial statements in additional analysis. This positive effect is more significant when the CFO is female or middle-aged, or has a Bachelor's degree or higher or a financial background. This paper enriches the theoretical research on CFO's employment characteristics and the quality of financial reports, and provides reference value for improving the quality of financial reports.

Gamma Knife Radiosurgery Irradiation of Surgical Cavity of Brain Metastases: Factor Analysis and Gene Mutations.

(1) Background: Surgical resection for the removal of brain metastases often fails to prevent tumor recurrence within the surgical cavity; hence, researchers are divided as to the benefits of radiation treatment following surgical resection. This retrospective study assessed the effects of post-operative stereotactic radiosurgery (SRS) on local tumor control and overall survival. (2) Methods: This study examined the demographics, original tumor characteristics, and surgical outcomes of 97 patients who underwent Gamma Knife Radiosurgery (GKRS) treatment (103 brain metastases). Kaplan-Meier plots and Cox regression were used to correlate clinical features to tumor control and overall survival. (3) Results: The overall tumor control rate was 75.0% and overall 12-month survival was 89.6%. Tumor control rates in the radiation group versus the non-radiation group were as follows: 12 months (83.1% vs. 57.7%) and 24 months (66.1% vs. 50.5%). During the 2-year follow-up period after SRS, the intracranial response rate was higher in the post-craniotomy radiation group than in the non-radiation group (p = 0.027). Cox regression multivariate analysis determined that post-craniotomy irradiation of the surgical cavity is predictive of tumor control (p = 0.035). However, EGFR mutation was not predictive of overall survival or tumor control. (4) Conclusions: Irradiating the surgical cavity after surgery can enhance local tumor control; however, it does not have a significant effect on overall survival.

Dengue Meteorological Determinants during Epidemic and Non-Epidemic Periods in Taiwan.

The identification of the key factors influencing dengue occurrence is critical for a successful response to the outbreak. It was interesting to consider possible differences in meteorological factors affecting dengue incidence during epidemic and non-epidemic periods. In this study, the overall correlation between weekly dengue incidence rates and meteorological variables were conducted in southern Taiwan (Tainan and Kaohsiung cities) from 2007 to 2017. The lagged-time Poisson regression analysis based on generalized estimating equation (GEE) was also performed. This study found that the best-fitting Poisson models with the smallest QICu values to characterize the relationships between dengue fever cases and meteorological factors in Tainan (QICu = −8.49 × 10−3) and Kaohsiung (−3116.30) for epidemic periods, respectively. During dengue epidemics, the maximum temperature with 2-month lag (ß = 0.8400, p < 0.001) and minimum temperature with 5-month lag (0.3832, p < 0.001). During non-epidemic periods, the minimum temperature with 3-month lag (0.1737, p < 0.001) and mean temperature with 2-month lag (2.6743, p < 0.001) had a positive effect on dengue incidence in Tainan and Kaohsiung, respectively.

Screening of ulcerative colitis biomarkers and potential pathways based on weighted gene co-expression network, machine learning and ceRNA hypothesis.

BACKGROUND: Ulcerative colitis (UC) refers to an intractable intestinal inflammatory disease. Its increasing incidence rate imposes a huge burden on patients and society. The UC etiology has not been determined, so screening potential biomarkers is critical to preventing disease progression and selecting optimal therapeutic strategies more effectively. METHODS: The microarray datasets of intestinal mucosal biopsy of UC patients were selected from the GEO database, and integrated with R language to screen differentially expressed genes and draw proteins interaction network diagrams. GO, KEGG, DO and GSEA enrichment analyses were performed to explore their biological functions. Through machine learning and WGCNA analysis, targets that can be used as UC potential biomarkers are screened out. ROC curves were drawn to verify the reliability of the results and predicted the mechanism of marker genes from the aspects of immune cell infiltration, co-expression analysis, and competitive endogenous network (ceRNA). RESULTS: Two datasets GSE75214 and GSE87466 were integrated for screening, and a total of 107 differentially expressed genes were obtained. They were mainly related to biological functions such as humoral immune response and inflammatory response. Further screened out five marker genes, and found that they were associated with M0 macrophages, quiescent mast cells, M2 macrophages, and activated NK cells in terms of immune cell infiltration. The co-expression network found significant co-expression relationships between 54 miRNAs and 5 marker genes. According to the ceRNA hypothesis, NEAT1-miR-342-3p/miR-650-SLC6A14, NEAT1-miR-650-IRAK3, and XIST-miR-342-3p-IRAK3 axes were found as potential regulatory pathways in UC. CONCLUSION: This study screened out five biomarkers that can be used for the diagnosis and treatment of UC, namely SLC6A14, TIMP1, IRAK3, HMGCS2, and APOBEC3B. Confirmed that they play a role in the occurrence and development of UC at the level of immune infiltration, and proposed a potential RNA regulatory pathway that controls the progression of UC.

2D Hexagonal Assemblies of Amphiphilic Double-Helical Poly(phenylacetylene) Homopolymers with Enhanced Circularly Polarized Luminescence and Chiral Self-Sorting.

Two-dimensional (2D) chiral materials have been attracting immense attentions owing to their unique properties. Herein, we successfully developed a unique assembly strategy of amphiphilic homopolymers to construct stable free-standing 2D chiral nanosheets in solution. The amphiphilic poly(phenylacetylene) (PPA) homopolymers bearing the hydrophobic and hydrophilic dendritic side chains adopt a DNA-like double-helical conformation. The regular hexagonal nanosheets were formed in THF/EtOH through nucleation and epitaxial growth. The sizes of the nanosheets can be modulated from nanometers to submillimeters upon varying the ratio of binary solvents, while the thickness is linearly correlated with the molecular weights. The 2D architecture can significantly enhance the CPL of polymers with a high dissymmetry factor ≈0.1. Driven by a discrimination of helical conformation, the PPAs can self-sort into homochiral 2D nanosheets, as directly visualized by using fluorescent microscopy.

Effect of Intragenomic Sequence Heterogeneity among Multiple 16S rRNA Genes on Species Identification of Elizabethkingia.

Accurate identification of Elizabethkingia species mostly requires the use of molecular techniques, and 16S rRNA gene sequencing is generally considered the method of choice. In this study, we evaluated the effect of intraspecific diversity among the multiple copies of the 16S rRNA gene on the accuracy of species identification in the genus Elizabethkingia. Sequences of 16S rRNA genes obtained from the 32 complete whole-genome sequences of Elizabethkingia deposited in GenBank and from 218 clinical isolates collected from 5 hospitals in Taiwan were analyzed. Four or five copies of 16S rRNA were identified in the Elizabethkingia species with complete genome sequences. The dissimilarity among the copies of the16S rRNA gene was <1% in all Elizabethkingia strains. E. meningoseptica demonstrated a significantly higher rate of nucleotide variations in the 16S rRNA than did E. anophelis (P = 0.011). Nucleotide alterations occurred more frequently in regions V2 and V6 than in other hypervariable regions (P < 0.001). E. meningoseptica, E. anophelis, and E. argenteiflava strains were clustered distinctly in the phylogenetic tree inferred from 16S rRNA genes, and the intragenomic variation of gene sequences had no profound effect on the classification of taxa. However, E. miricola, E. bruuniana, E. ursingii, and E. occulta were grouped closely in the phylogenetic analysis, and the variation among the multiple copies of the 16S rRNA in one E. ursingii strain affected species classification. Other marker genes may be required to supplement the species classification of closely related taxa in the genus Elizabethkingia. IMPORTANCE Incorrect identification of bacterial species would influence the epidemiology and clinical analysis of patients infected with Elizabethkingia. The results of the present study suggest that 16S rRNA gene sequencing should not be considered the gold standard for the accurate identification of Elizabethkingia species.

Induced Pluripotent Stem Cell-Derived Corneal Cells: Current Status and Application.

Deficiency and dysfunction of corneal cells leads to the blindness observed in corneal diseases such as limbal stem cell deficiency (LSCD) and bullous keratopathy. Regenerative cell therapies and engineered corneal tissue are promising treatments for these diseases [1]. However, these treatments are not yet clinically feasible due to inadequate cell sources. The discovery of induced pluripotent stem cells (iPSCs) by Shinya Yamanaka has provided a multitude of opportunities in research because iPSCs can be generated from somatic cells, thus providing an autologous and unlimited source for corneal cells. Compared to other stem cell sources such as mesenchymal and embryonic, iPSCs have advantages in differentiation potential and ethical concerns, respectively. Efforts have been made to use iPSCs to model corneal disorders and diseases, drug testing [2], and regenerative medicine [1]. Autologous treatments based on iPSCs can be exorbitantly expensive and time-consuming, but development of stem cell banks with human leukocyte antigen (HLA)- homozygous cell lines can provide cost- and time-efficient allogeneic alternatives. In this review, we discuss the early development of the cornea because protocols differentiating iPSCs toward corneal lineages rely heavily upon recapitulating this development. Differentiation of iPSCs toward corneal cell phenotypes have been analyzed with an emphasis on feeder-free, xeno-free, and well-defined protocols, which have clinical relevance. The application, challenges, and potential of iPSCs in corneal research are also discussed with a focus on hurdles that prevent clinical translation.