Development in competitive immunoassay of a point-of-care testing for cotinine (COT) detection in urine.

We present a sensitive and selective lateral flow immunoassay (LFIA) for cotinine (COT), the primary metabolite of nicotine. COT is widely recognized as a superior biomarker to evaluate tobacco smoke exposure. The LFIA uses a competitive assay format where the COT-BSA capture competes with the target COT in urine samples for binding to the monoclonal antibody against COT (mAb-COT) conjugated with gold nanoparticles (mAb-COT-AuNPs). To improve the sensitivity and selectivity of the LFIA-COT, we focused on optimizing the diameter of AuNPs, the conjugation of mAb-COT, and the concentration of the COT-BSA capture. Our findings reveal that the utilization of 40 nm AuNPs in conjugation with a concentration of 4 mg mL-1 of mAb-COT demonstrated significantly greater efficacy compared to LFAs utilizing 20 nm AuNPs. Under the optimal conditions, the LFIA-COT demonstrated sensitive detection of COT at a level of 150 ng mL-1 within 15 min, as observed by the naked eye. It possesses a linear range of 25 to 200 ng mL-1 of COT, with the limit of detection (LOD) of 11.94 ng mL-1 in human urine samples when the color intensity is analyzed using ImageJ software. Our LFIA described here is simple and requires less time for COT detection. It can be used for the rapid and quantitative detection of COT in urine samples in clinical settings.

The Evaluation of a Lateral Flow Strip Based on the Covalently Fixed "End-On" Orientation of an Antibody for Listeria monocytogenes Detection.

In this study, the covalently fixed "end-on" orientation of a monoclonal Listeria monocytogenes antibody (mAb-Lis) to amino terminated oligo (ethylene glycol)-capped gold nanoparticles (NH2-TEG-AuNPs) was used to fabricate an in-house lateral flow strip (LFS), namely, the fixed "end-on" Lis-mAb-NH-TEG-AuNPs LFS. The aim was to evaluate the performance of the fixed "end-on" Lis-mAb-NH-TEG-AuNPs LFS in detecting L. monocytogenes. The proposed LFS enabled the sensitive detection of L. monocytogenes in 15 min with a visual limit of detection of 102 CFU/mL. Quantitative analysis indicated an LOD at 10 CFU/mL. The fixed "end-on" Lis-mAb-NH-TEG-AuNPs LFS showed no cross-reactivity with other pathogenic bacteria and practical performance across different food matrices, including human blood, milk, and mushroom samples. Furthermore, the clinical performance of the fixed "end-on" Lis-mAb-NH-TEG-AuNPs LFS for detecting L. monocytogenes was evaluated by using 12 clinical samples validated by the hemoculture method. It demonstrated excellent concordance with the reference methods, with no false-positive or false-negative results observed. Therefore, the fixed "end-on" Lis-mAb-NH-TEG-AuNPs LFS serves as a promising candidate for a point-of-care test (POCT), enabling the rapid, precise, and highly sensitive detection of L. monocytogenes in clinical samples and contaminated food.

Serum CCDC25 Levels as a Potential Marker for Metabolic Syndrome.

BACKGROUND/AIM: Metabolic syndrome (MetS) stands as a significant risk for developing various severe health problems. Therefore, the discovery of biomarkers capable of predicting the progression of metabolic conditions is crucial for improving overall health outcomes. Recently, we reported that coiled-coil domain containing 25 (CCDC25) might be associated with key proteins involved in metabolic pathways, by bioinformatics analysis. Thus, we assumed that serum CCDC25 levels might have an association with MetS status. PATIENTS AND METHODS: In this study, based on the modified National Cholesterol Education Program-Adult Treatment Panel III (modified NCEP-ATP III) criteria, the participants who had three or more of abnormal criteria were defined as MetS, and those who had 1 or 2 abnormal criteria as pre-MetS groups; those who had no abnormal criteria were classified as the healthy control (HC) group. Serum CCDC25 levels were measured using the dot blot assay. RESULTS: The results showed that serum CCDC25 levels of the MetS group (0.072±0.026 ng/µl) were significantly higher (p<0.001) than that of pre-MetS (0.031±0.011 ng/µl) or HC groups (0.018±0.007 ng/µl). We can discern a consistent trend indicating that serum CCDC25 level is well correlated with the number of abnormal criteria of MetS of each participant. Although serum CCDC25 levels correlated with the distribution of all 5 MetS criteria, the highest correlation was seen in serum CCDC25 levels and triglyceride (TG) levels, with r=0.563, followed by systolic blood pressure (SBP) levels (r=0.557) and high-density lipoprotein-cholesterol (HDL-C) levels (r=-0.545). CONCLUSION: CCDC25 showed correlations with all MetS parameters, particularly with TG, SBP, and HDL-C. This prompts speculation that heightened CCDC25 levels may indicate the development and/or progression of those MetS-associated diseases.

Characterization and Localization of Sol g 2.1 Protein from Solenopsis geminata Fire Ant Venom in the Central Nervous System of Injected Crickets (Acheta domestica).

Solenopsis geminata is recognized for containing the allergenic proteins Sol g 1, 2, 3, and 4 in its venom. Remarkably, Sol g 2.1 exhibits hydrophobic binding and has a high sequence identity (83.05%) with Sol i 2 from S. invicta. Notably, Sol g 2.1 acts as a mediator, causing paralysis in crickets. Given its structural resemblance and biological function, Sol g 2.1 may play a key role in transporting hydrophobic potent compounds, which induce paralysis by releasing the compounds through the insect's nervous system. To investigate this further, we constructed and characterized the recombinant Sol g 2.1 protein (rSol g 2.1), identified with LC-MS/MS. Circular dichroism spectroscopy was performed to reveal the structural features of the rSol g 2.1 protein. Furthermore, after treating crickets with S. geminata venom, immunofluorescence and immunoblotting results revealed that the Sol g 2.1 protein primarily localizes to the neuronal cell membrane of the brain and thoracic ganglia, with distribution areas related to octopaminergic neuron cell patterns. Based on protein-protein interaction predictions, we found that the Sol g 2.1 protein can interact with octopamine receptors (OctRs) in neuronal cell membranes, potentially mediating Sol g 2.1's localization within cricket central nervous systems. Here, we suggest that Sol g 2.1 may enhance paralysis in crickets by acting as carriers of active molecules and releasing them onto target cells through pH gradients. Future research should explore the binding properties of Sol g 2.1 with ligands, considering its potential as a transporter for active molecules targeting pest nervous systems, offering innovative pest control prospects.

The effect of edible bird's nests on the expression of MHC-II and costimulatory molecules of C57BL/6 mouse splenocytes.

The glutinous nest that builds by the saliva secretion of swiftlet is recognizable as an edible bird's nest (EBN). It enriched a medicinal value and was regarded as supplementary food that exerts various beneficial health effects, especially immune boosters. This study's objective was to determine the impact of EBN on the expression of MHC-II and costimulatory molecules (CD86 and CD80) related to the initiation of T-cell activation. Both rEBN and pEBN samples were prepared with simulated gastrointestinal digestion for enhancing the bioaccessibility of bioactive compounds. Our result showed that digested EBN samples slightly influence the upregulation of MHC-II, CD86, and CD80 in gene expression of LPS-stimulated Raw 264.7 cells. The concern of endotoxin contamination in EBN samples, which may cause a false-positive result, was measured by quantitative PCR. We found that the inflammatory genes (IL-1ß and TNF-α) were not induced by EBN treatments. Moreover, cell surface protein expression in splenocytes treated with EBN was assessed using flow cytometric analysis. Digested EBN samples demonstrated their capacity to promote the elevation of MHC-II, CD86, and CD80 cell surface protein expression. Finally, the digested-EBN-treated splenocytes only exhibited a specific response in the T-cells population. Thus, EBN is a source of the bioactive compound that has been proposed to exert a role in the stimulation of both MHC-II and costimulatory molecules for TCR/pMHC-II interaction leading to T-cell activation.

Ca2+/Calmodulin-dependent Protein Kinase II Inhibitor KN-93 Enhances Chondrogenesis of Bone Marrow Mesenchymal Stem Cells and Delays Chondrogenic Hypertrophy.

BACKGROUND/AIM: Cartilage tissue engineering has been popularly applied in the treatment of articular cartilage defect because it is more effective in generating functional engineered cartilage than traditional methods. Although the chondrogenic differentiation of human bone marrow-derived mesenchymal stem cells (BM-MSCs) is well established, it is often accompanied by undesired hypertrophy. Ca2+/calmodulin-dependent protein kinase II (CaMKII) is a crucial mediator in the ion channel pathway which is known to be involved in chondrogenic hypertrophy. Therefore, this study aimed to reduce the hypertrophy of BM-MSCs by inhibiting CaMKII activation. MATERIALS AND METHODS: BM-MSCs were cultured in three-dimensional (3D) scaffold under chondrogenic induction with and without CaMKII inhibitor, KN-93. After cultivation, markers of chondrogenesis and hypertrophy were investigated. RESULTS: KN-93 at a concentration of 2.0 µM had no effect on the viability of BM-MSCs, while the activation of CaMKII was suppressed. A long period of KN-93 treatment significantly up-regulated the expression of SRY-box transcription factor 9 and aggrecan on day 28 compared to untreated BM-MSCs. Furthermore, KN-93 treatment significantly down-regulated the expression of RUNX family transcription factor 2 and collagen type X alpha 1 chain on days 21 and 28. Immunohistochemistry showed increased expression of aggrecan and type II collagen while the expression of type X collagen was reduced. CONCLUSION: A CaMKII inhibitor, KN-93 is able to enhance chondrogenesis of BM-MSCs and suppress chondrogenic hypertrophy, suggesting its potential applicability in cartilage tissue engineering.

Three-dimensional silk fibroin-gelatin/chondroitin sulfate/hyaluronic acid-aloe vera scaffold supports in vitro chondrogenesis of bone marrow mesenchymal stem cells and reduces inflammatory effect.

A limited self-healing ability of injured articular cartilage results in osteoarthritis and a joint dysfunction afterward. Cartilage tissue engineering is a promising approach to increase the treatment efficiency. Moreover, host response to implanted biomaterial has been increasingly concerned. Thus, this study aimed to establish three-dimensional (3D) scaffold that could support cartilage tissue engineering and reduce inflammatory. The various ratios of silk fibroin (SF), gelatin (G), chondroitin sulfate (C), hyaluronic acid (H), and aloe vera (A) were used to fabricate 3D scaffolds by lyophilization, designated as SF, SF-A, SF-gelatin/chondroitin sulfate/hyaluronic acid (GCH)-A-411, and SF-GCH-A-111. The physical and biological characteristics of the scaffolds were investigated. All scaffolds possessed interconnected porous structures, which the highest pore size of 209 µm was found in SF and SF-GCH-A-411 scaffolds. Moreover, high porosity, high water uptake, and good mechanical strength were observed in the SF-GCH-A-411 scaffold. The SF, SF-A, and SF-GCH-A-411 scaffolds could retain their structures up to 21 days, while SF-GCH-A-111 was rapidly degraded. The proliferation of human bone marrow mesenchymal stem cells (BM-MSCs) was significantly higher in SF-A and SF-GCH-A-411 than in the SF scaffold. Besides, the SF-A and SF-GCH-A-411 revealed significantly lower expression of pro-inflammatory cytokine, interleukin-1 beta than the SF scaffold, suggesting the beneficial role of aloe vera in anti-inflammatory effect. Furthermore, the SF-GCH-A-411 scaffold could support chondrogenic differentiation of BM-MSCs. In conclusion, based on its superior physical and biological characteristics that support chondrogenesis of BM-MSCs, the SF-GCH-A-411 scaffold is recommended for cartilage tissue engineering.

Unveiling the Potent Antiviral and Antioxidant Activities of an Aqueous Extract from Caesalpinia mimosoides Lamk: Cheminformatics and Molecular Docking Approaches.

Our group previously demonstrated that Caesalpinia mimosoides Lamk exhibits many profound biological properties, including anticancer, antibacterial, and antioxidant activities. However, its antiviral activity has not yet been investigated. Here, the aqueous extract of C. mimosoides was prepared from the aerial parts (leaves, stalks, and trunks) to see whether it exerts anti-influenza (H1N1) effects and to reduce the organic solvents consumed during extraction, making it a desirable approach for the large-scale production for medical uses. Our plant extract was quantified to contain 7 g of gallic acid (GA) per 100 g of a dry sample, as determined using HPLC analysis. It also exerts potent antioxidant activities comparable to those of authentic GA. According to untargeted metabolomics (UPLC-ESI(-)-QTOF-MS/MS) with the aid of cheminformatics tools (MetFrag (version 2.1), SIRIUS (version 5.8.3), CSI:FingerID (version 4.8), and CANOPUS), the major metabolite was best annotated as "gallic acid", phenolics (e.g., quinic acid, shikimic acid, and protocatechuic acid), sugar derivatives, and dicarboxylic acids were deduced from this plant species for the first time. The aqueous plant extract efficiently inhibited an influenza A (H1N1) virus infection of MDCK cells with an IC50 of 5.14 µg/mL. Of equal importance, hemolytic activity was absent for this plant extract, signifying its applicability as a safe antiviral agent. Molecular docking suggested that GA interacts with conserved residues (e.g., Arg152 and Asp151) located in the catalytic inner shell of the viral neuraminidase (NA), sharing the same pocket as those of anti-neuraminidase drugs, such as laninamivir and oseltamivir. Additionally, other metabolites were also found to potentially interact with the active site and the hydrophobic 430-cavity of the viral surface protein, suggesting a possibly synergistic effect of various phytochemicals. Therefore, the C. mimosoides aqueous extract may be a good candidate for coping with increasing influenza virus resistance to existing antivirals.

Polydopamine Nanoparticles Functionalized Electrochemical DNA Aptasensor for Serum Glycated Albumin Detection.

Polydopamine (PDA) has now been widely applied to electrochemical biosensing because of its excellent biocompatibility, abundant functional groups, and facile preparation. In this study, polydopamine nanoparticles (PDA-NPs)-functionalized electrochemical aptasensor was developed for the rapid, sensitive, and cost-effective detection of glycated albumin (GA), a promising biomarker for glycemic control in diabetic patients. PDA-NPs were synthesized at various pH conditions in Tris buffer. Cyclic voltammetry (CV) of PDA-NPs-coated screen-printed carbon electrodes (SPCEs) revealed that the materials were more conductive when PDA-NPs were synthesized at pH 9.5 and 10.5 than that at pH 8.5. At pH 10.5, the prepared PDA and PDA-aptamer NPs were monodispersed spherical morphology with an average size of 118.0 ± 1.9 and 127.8 ± 2.0 nm, respectively. When CV and electrochemical impedance spectrometry (EIS) were used for the characterization and detection of the electrochemical aptasensor under optimal conditions, the proposed aptasensor exhibited a broad linearity for detection of GA at a clinically relevant range of (1-10,000 µg mL-1), provided a low detection limit of 0.40 µg mL-1, appreciable reproducibility (less than 10%), and practicality (recoveries 90-104%). In addition, our developed aptasensor presented a great selectivity towards GA, compared to interfering substances commonly present in human serum, such as human serum albumin, urea, glucose, and bilirubin. Furthermore, the evaluation of the aptasensor performance against GA-spiked serum samples showed its probable applicability for clinical use. The developed PDA aptasensor demonstrated excellent sensitivity and selectivity towards GA detection with a simple and facile fabrication process. This proposed technique shows its potential application in GA measurement for improving the screening and management of diabetic patients in the future.

Recombinase Polymerase Amplification Combined with Lateral Flow Strip for Rapid Detection of OXA-48-like Carbapenemase Genes in Enterobacterales.

Carbapenem-resistant Enterobacterales (CRE) possessing various carbapenemases, particularly the OXA-48 group, are now rapidly spreading and becoming a major public health concern worldwide. Phenotypic detection of OXA-48-like carbapenemases is still suboptimal due to their weak carbapenemase activity, whereas highly sensitive and specific polymerase chain reaction (PCR)-based methods take at least 3-4 h. We, therefore, developed a recombinase polymerase amplification (RPA) combined with lateral flow (LF) strip assay for the rapid detection of blaOXA-48-like in Enterobacterales. A total of 131 clinical isolates including 61 blaOXA-48-like-carrying Enterobacterales isolates and 70 Gram-negative bacilli isolates containing other bla genes were subjected to the RPA method performed under isothermal conditions at 37 °C within 10 min and visually inspected by LF strip within 5 min. The RPA-LF assay provided 100% sensitivity (95% confidence interval, 92.6-100%) and 100% specificity (93.5-100%) for detecting blaOXA-48-like genes from bacterial colonies. Its detection limit was 100 times less than that of the PCR method. This assay is rapid, easy to perform, and provides excellent performance without any special equipment. It may be applied for directly identifying the blaOXA-48-like genes in Enterobacterales obtained from blood culture. Rapid identification of carbapenemase types is essential for selecting appropriate antimicrobial options, particularly the ß-lactams combined with novel ß-lactamase inhibitors.

Rapid detection of methicillin-resistant Staphylococcus aureus in positive blood-cultures by recombinase polymerase amplification combined with lateral flow strip.

Staphylococcus aureus, especially methicillin-resistant S. aureus (MRSA), is an important bacterium that causes community and healthcare-related infections throughout the world. However, the current conventional detection methods are time-consuming. We therefore developed and evaluated a recombinase polymerase amplification-lateral flow strip (RPA-LF) approach for detection of MRSA in positive blood-culture samples. Sixty positive blood-cultures from a hospital were tested directly without DNA extraction and purification before the amplification reaction. RPA primers and probes were designed for nuc (encoding thermonuclease) and mecA (encoding penicillin-binding protein 2a) genes to diagnose S. aureus and its methicillin-resistance status. The RPA reaction occurred under isothermal conditions (45°C) within 20 min and a result was provided by the LF strip in a further 5 min at room temperature. The evaluation of RPA-LF using blood-culture samples showed 93.3% (14/15) sensitivity for identifying S. aureus, and no cross-amplification was seen [100% (45/45) specificity]. For detection of methicillin resistance, the RPA-LF test provided 100% (16/16) sensitivity and 97.7% (43/44) specificity. The RPA-LF is rapid, highly sensitive, robust and easy to use. It can be used for direct detection of MRSA with no requirement for special equipment.

Multi Evaluation of a Modified GoldNano Carb Test for Carbapenemase Detection in Clinical Isolates of Gram-Negative Bacilli.

Carbapenemase-producing Gram-negative bacteria have been increasingly reported. Simple and sensitive methods for carbapenemase detection are still needed. In this study, a gold nanoparticle (AuNP) solution was modified by the addition of zinc sulfate (ZnSO4) for improving the conventional GoldNano Carb (cGoldC) test, and the modified GoldC (mGoldC) test was then evaluated for phenotypic detection of carbapenemase production in Gram-negative bacilli clinical isolates. ZnSO4 was added to give final concentrations of 0.25, 0.5, 0.75, and 1 mM. The performance of the mGoldC test was evaluated in Enterobacterales, Acinetobacter spp., and Pseudomonas aeruginosa isolates from six hospitals in different regions using polymerase chain reaction (PCR) as a gold standard. The AuNP solution with 0.25 mM ZnSO4 was used for the mGoldC test. Evaluation of the mGoldC test in 495 Enterobacterales, 212 Acinetobacter spp., and 125 P. aeruginosa isolates (including 444 carbapenemase producers and 388 non-carbapenemase producers) revealed sensitivity, specificity, a positive likelihood ratio, and a negative likelihood ratio of 98.6%, 98.2%, 54.7, and 0.01, respectively. This test is fast, easy to perform, cost-effective (~0.25 USD per test), and highly sensitive and specific for routine carbapenemase detection, thus leading to effective antimicrobial therapy and infection control measures.

Combined OPCML and AXL Expression as a Prognostic Marker and OPCML Enhances AXL Inhibitor in Cholangiocarcinoma.

BACKGROUND/AIM: Cholangiocarcinoma (CCA) is a type of liver cancer originating from bile duct epithelium which has an unfavorable prognosis. Therefore, novel prognostic markers and effective therapeutic regimens are required. Opioid-binding protein/cell adhesion molecule-like (OPCML) is a tumor-suppressor protein that suppresses CCA cell proliferation via AXL receptor tyrosine kinase/signal transducer and activator of transcription 3 (AXL/STAT3) inactivation. However, this association in clinical samples remains unknown. We aimed to determine OPCML and AXL expression and investigate their association with clinicopathological features in patients with CCA. In addition, we also addressed whether OPCML enhanced the sensitivity of CCA cells to AXL inhibitor R428 in vitro. MATERIALS AND METHODS: The expression of OPCML and AXL was determined by immunohistochemistry in 90 CCA tissue samples. The study of CCA cell line sensitivity to R428 was performed by cell viability assay. RESULTS: The expression of OPCML was significantly lower while AXL expression was substantially higher in CCA than in adjacent normal tissue (p<0.001). Furthermore, high AXL expression was significantly associated with lymph node metastasis (p=0.035). Interestingly, patients with combined low OPCML/high AXL expression had significantly shorter overall survival (p=0.007). OPCML enhanced the effect of AXL inhibitor R428 in AXL-expressing CCA cell lines. CONCLUSION: Combined expression of OPCML and AXL shows potential value as a prognostic marker and OPCML as an agent enhancing the effect of R428 may contribute to better prognosis for patients with CCA.

3D Silk Fibroin-Gelatin/Hyaluronic Acid/Heparan Sulfate Scaffold Enhances Expression of Stemness and EMT Markers in Cholangiocarcinoma.

BACKGROUND/AIM: Cholangiocarcinoma (CCA) is a stem cell-based cancer. The in vivo tumor microenvironment is not present in two-dimensional (2D) cultures, which is one of the limitations in cancer stem cell (CSC) research. Thus, we aimed to establish three-dimensional (3D) culture mimicking extracellular matrix (ECM) that could serve as a niche for CSC enrichment in CCA. MATERIALS AND METHODS: Silk fibroin-gelatin/hyaluronic acid/heparan sulfate (SF-GHHs) scaffolds were fabricated by lyophilization in various ratios and compared to silk fibroin (SF) scaffold. The physical and biological characteristics of the scaffolds were investigated. RESULTS: The SF-GHHs 1:2 scaffold with pore size of 350±102 µm harbored optimal porosity, good water uptake, and stable beta-sheet that supported the increase in KKU-213A cell proliferation and aggregation. The CSC and the epithelial-mesenchymal transition (EMT) markers were significantly upregulated in this scaffold compared to 2D. Moreover, drug sensitivity against cisplatin and gemcitabine in 3D culture was significantly higher than that in 2D culture. CONCLUSION: The SF-GHHs 1:2 scaffold could simulate ECM that may serve as a CSC niche of CCA, and reinforce stemness and EMT properties, suggesting its suitability for 3D CCA model, which supports CSC and new targeting drug research in CCA.

Anti-Proteus Activity, Anti-Struvite Crystal, and Phytochemical Analysis of Sida acuta Burm. F. Ethanolic Leaf Extract.

Proteus mirabilis is a significant cause of urinary tract infection that may contribute to struvite stones. Anti-infection of this bacterium and anti-struvite formation must be considered. Sida acuta Burm. F. (SA) has been used for the treatment of diseases related to kidneys. Therefore, we investigated the effects of the SA leaf ethanolic extract (SAEE) on growth and on virulent factors (swarming motility and urease activity) of Proteusmirabilis isolated from kidney stone formers. We also evaluated anti-struvite crystal formation and phytochemical constituents of SAEE. The minimum inhibitory concentrations (MICs) of SAEE against three clinical P. mirabilis isolates were 8 mg/mL. Intriguingly, the 1/2MIC of SAEE had significant inhibitory effects on the swarming motility and urease activity of clinical P. mirabilis isolates when compared with the condition without SAEE. The SAEE at the various concentrations significantly inhibited the average weights of struvite crystals in a dose-dependent manner, compared with the control. The phytochemical analysis revealed that SAEE contained catechin, chlorogenic acid, rutin, and ferulic acid. This study indicated that SAEE has anti-P. mirabilis and anti-struvite crystal activities via its bioactive compounds. For this reason, SAEE may be developed as a new agent for the treatment of struvite stone induced by P. mirabilis.

Proteomic profiling reveals antitumor effects of RT2 peptide on a human colon carcinoma xenograft mouse model.

A comparative study of human colon HCT-116 xenograft in nude mice treated with and without peptide RT2 at high doses is performed along with a label-free proteomic analysis of the tissue in order to understand the potential mechanisms by which RT2 acts in vivo against colorectal tumors. RT2 displays no significant systematic toxicity, but reduces tumor growth after either intraperitoneal or intratumoral injection demonstrating it is a safe and efficacious antitumor agent in vivo. Of the 3196 proteins identified by label-free proteomics, 61 proteins appear only in response to RT2 and are involved in cellular processes largely localized in the cells and cell parts. Some of the proteins identified, including CFTR, Wnt7a, TIA1, PADI2, NRBP2, GADL1, LZIC, TLR6, and GPR37, have been reported to suppress tumor growth and are associated with cell proliferation, invasion, metastasis, angiogenesis, apoptosis, and immune evasion. Our work supports their role as tumor biomarkers and reveals RT2 has a complex mechanism of action in vivo.

Novel non-cytotoxic antimicrobial peptides WSKK11 and WSRR11 with potent activity against Cutibacterium acnes.

OBJECTIVES: Cutibacterium acnes is one of the common multifactorial causes that play an important role in the pathophysiology of acne vulgaris. We aimed to develop novel antimicrobial peptides for reduction of the hypercolonization. METHODS: Six cationic peptides were derived by de novo designation. The antimicrobial and cytotoxic activities of peptides were investigated. The peptide conformation was determined by circular dichroism spectrometry. The antimicrobial effects of peptides were evaluated using scanning electron microscopy (SEM), transmission electron microscopy (TEM) and DNA-binding ability assay. RESULTS: Among designed peptides, WSKK11 and WSRR11 were effective antimicrobials against C. acnes at MICs of 128 and 64 mg/L, respectively. The MICs of WSKK11 against Staphylococcus epidermidis, Staphylococcus aureus and Candida albicans were 8, 8 and 32 mg/L, while those of WSRR11 were 64, 32 and 32 mg/L, respectively. WSKK11 and WSRR11 were less toxic to human erythrocytes (<2%) and not toxic to macrophages, keratinocytes and fibroblasts up to 512 mg/L. WSKK11 and WSRR11 mostly revealed the conformation of the undefined or random coil structures under mimicked environmental conditions. The peptides affected cell surfaces and cell membranes of C. acnes as well as possibly translocating through the cell membrane, observed by a combination of SEM and TEM, respectively. WSKK11 and WSRR11 had the ability to bind bacterial DNA. CONCLUSIONS: The two novel antimicrobial peptides WSKK11 and WSRR11 are members of a new class of antimicrobial agents that could deal with acne problems. Therefore, the antimicrobial peptides may be promising novel active agents for dermatological, beauty and cosmeceutical applications.

Risk factors for canine magnesium ammonium phosphate urolithiasis associated with bacterial infection.

BACKGROUND: With limited information available, the association among urinary tract infections, urease-producing bacteria and the presence of magnesium ammonium phosphate (MAP) urolithiasis in canines in Thailand requires more study. OBJECTIVES: This study aimed to investigate the association between demographic characteristics of canines and the presence of MAP urolithiasis in canines, and to evaluate antimicrobial susceptibility patterns of bacteria isolated from canine uroliths. METHODS: A total of 56 canines admitted for treatment with surgical removal of uroliths were recruited. Demographic characteristics and clinical chemistry data were recorded. Bacteria isolated from the removed uroliths were identified. Chemical compositions of the uroliths were analyzed by Fourier transform infrared spectrometer. Potential risk factors were determined with univariable and multivariable logistic regression analyses. RESULTS: Of 56 canine urolithiasis, bacteria were isolated from uroliths of 38 canines (27 MAP and 11 non-MAP) but not from uroliths of 18 canines (5 MAP and 13 non-MAP). The most common bacteria found in nidus of MAP uroliths was Staphylococcus pseudintermedius (approximately 51%). An antimicrobial resistance was frequently found in Staphylococci isolates (42.86%). Multivariate logistic regression analysis showed that the predictors of MAP urolith in canine urolithiasis were being female (p = 0.044; adjusted odds ratio [OR], 10.22; 95% confidence interval [CI], 1.06-98.24) and the positive urolith culture (p = 0.012; adjusted OR, 8.60; 95% CI, 1.60-46.30). CONCLUSIONS: Our results indicate that S. pseudintermedius (a urease-producing bacterium) is the major causative bacteria of MAP uroliths. A positive urolith culture and being female are risk factors of MAP urolithiasis in canines.

Rapid detection of Enterococcus and vancomycin resistance using recombinase polymerase amplification.

Vancomycin-resistant enterococci (VRE), especially Enterococcus faecium, have been a global concern, often causing serious healthcare-associated infections. We established a rapid approach for detecting E. faecium and vancomycin-resistance genes (vanA and vanB) in clinical samples using isothermal recombinase polymerase amplification (RPA) combined with a lateral-flow (LF) strip. Specific RPA primer sets and probes for ddl (to identify the presence of E. faecium) vanA and vanB genes were designed. The RPA reaction was performed under isothermal condition at 37 °C within 20 min and read using the LF strip within a further 5 min. A total of 141 positive blood-cultures and 136 stool/rectal swab samples were tested using RPA-LF method compared to the conventional PCR method. The RPA-LF method exhibited 100% sensitivity in both blood-culture (60 E. faecium; 35 vanA type and two vanB type) and stool/rectal-swab samples (63 E. faecium and 36 vanA type) without cross-reaction (100% specificity). The lower detection limit of the RPA-LF was approximately 10 times better than that of the conventional PCR method. The RPA-LF method is an alternative rapid method with excellent sensitivity and specificity for detecting E. faecium, vanA, and vanB, and it has the potential to be used as a point-of-care device for VRE therapy and prevention.

A Disposable Electrochemical Biosensor Based on Screen-Printed Carbon Electrodes Modified with Silver Nanowires/HPMC/Chitosan/Urease for the Detection of Mercury (II) in Water.

This work describes the facile preparation of a disposable electrochemical biosensor for the detection of Hg(II) in water by modifying the surface of a screen-printed carbon electrode (SPCE). The surface modification consists of the immobilization of a composite layer of silver nanowires, hydroxymethyl propyl cellulose, chitosan, and urease (AgNWs/HPMC/CS/Urease). The presence of the composite was confirmed by scanning electron microscopy (SEM) and its excellent conductivity, due chiefly to the electrical properties of silver nanowires, enhanced the sensitivity of the biosensor. Under optimum conditions, the modified SPCE biosensor showed excellent performance for the detection of Hg(II) ions, with an incubation time of 10 min and a linear sensitivity range of 5-25 µM. The limit of detection (LOD) and limit of quantitation (LOQ) were observed to be 3.94 µM and 6.50 µM, respectively. In addition, the disposable and portable biosensor exhibited excellent recoveries for the detection of Hg(II) ions in commercial drinking water samples (101.62-105.26%). The results are correlated with those obtained from inductively coupled plasma optical emission spectrometry (ICP-OES), indicating that our developed sensor is a reliable method for detection of Hg(II) in real water samples. The developed sensor device is a simple, effective, portable, low cost, and user-friendly platform for real-time detection of heavy metal ions in field measurements with potential for other biomedical applications in the future.