Pennogenin 3-O-ß-Chacotrioside Attenuates Hypertrophied Lipid Accumulation by Enhancing Mitochondrial Oxidative Capacity.

Excessive lipid accumulation in adipocytes is a primary contributor to the development of metabolic disorders, including obesity. The consumption of bioactive compounds derived from natural sources has been recognized as being safe and effective in preventing and alleviating obesity. Therefore, we aimed to explore the antilipidemic effects of pennogenin 3-O-ß-chacotrioside (P3C), a steroid glycoside, on hypertrophied 3T3-L1 adipocytes. Oil Red O and Nile red staining demonstrated a P3C-induced reduction in lipid droplet accumulation. Additionally, the increased expression of adipogenic and lipogenic factors, including PPARγ and C/EBPα, during the differentiation process was significantly decreased by P3C treatment at both the protein and mRNA levels. Furthermore, P3C treatment upregulated the expression of fatty acid oxidation-related genes such as PGC1α and CPT1a. Moreover, mitochondrial respiration and ATP generation increased following P3C treatment, as determined using the Seahorse XF analyzer. P3C treatment also increased the protein expression of mitochondrial oxidative phosphorylation in hypertrophied adipocytes. Our findings suggest that P3C could serve as a natural lipid-lowering agent, reducing lipogenesis and enhancing mitochondrial oxidative capacity. Therefore, P3C may be a promising candidate as a therapeutic agent for obesity-related diseases.

Antioxidant and anti-hepatitis A virus activities of Ecklonia cava Kjellman extracts.

Ecklonia cava is a nutrient-rich algae species that contains abundant physiological phytochemicals, including peptides, carotenoids, fucoidans, and phlorotannins. However, elucidation of the antiviral effects of this algae and identification of new functional ingredients warrant further investigation. This study was aimed at investigating the potential anti-hepatitis A virus activities of extracts of E. cava prepared in different solvents. E. cava extracts were prepared using hot water and 70 % ethanol. The antioxidant activities of the extracts were confirmed by analyzing the total phenolic content, as well as 2,2-diphenyl-1-picrylhydrazyl and 2,2'-azino-bis-3-ethylbenzothiazoline-6-sulfonic acid radical scavenging activities. The inhibitory effects of the extracts against hepatitis A virus were analyzed using real-time polymerase chain reaction. The E. cava extract yield was 22.5-27.2 % depending on the extraction solvent. The 2,2-diphenyl-1-picrylhydrazyl radical scavenging activity was 70.44 % and 91.05 % for hot water and ethanol extracts at a concentration of 1000 ppm. The 2,2'-azino-bis-3-ethylbenzothiazoline-6-sulfonic acid radical scavenging activity of the ethanol extract was the highest (93.57 %) at 1000 µg/mL. Fourier-transform infrared was used to identify the functional groups (phlorotannin and alginate) in the extraction solvents. Ultra-high performance liquid chromatography with quadrupole time-of-flight tandem mass spectrometry analysis revealed a potential bioactive compound previously unidentified in E. cava. Finally, we identified the antiviral activity of E. cava extracts against hepatitis A virus replication. These findings demonstrate that E. cava could be used as an anti-hepatitis A virus functional food and biological material.

Utilization of coffee waste as a sustainable feedstock for high-yield lactic acid production through microbial fermentation.

Lactic acid is an important industrial precursor; however, high substrate costs are a major challenge in microbial fermentation-based lactic acid production. Coffee waste is a sustainable feedstock alternative for lactic acid production via microbial fermentation. Herein, the feasibility of coffee waste as a feedstock was explored by employing appropriate pretreatment methods and optimizing enzyme combinations. Coffee waste pretreatment with hydrogen peroxide and acetic acid along with a combination of Viscozyme L, Celluclast 1.5 L, and Pectinex Ultra SP-L achieved the 78.9 % sugar conversion rate at a substrate concentration of 4 % (w/v). Lactiplantibacillus plantarum WiKim0126-induced fermentation with a 4 % solid loading yielded a lactic acid concentration of 22.8 g/L (99.6 % of the theoretical maximum yield) and productivity of 0.95 g/L/h within 24 h. These findings highlight the viability of coffee waste as an eco-friendly resource for sustainable lactic acid production.

Survival analysis of implants after surgical treatment of peri-implantitis based on bone loss severity and surgical technique: a retrospective study.

BACKGROUND: Few trials have compared the results of surgical treatment for peri-implantitis based on severity of peri-implantitis and surgical method. This study investigated the survival rate of implants based on type of surgical method used and initial severity of peri-implantitis. Classification of severity was determined based on bone loss rate relative to fixture length. METHODS: Medical records of patients who underwent peri-implantitis surgery from July 2003 to April 2021 were identified. Classification of peri-implantitis was divided into 3 groups (stage 1: bone loss < 25% (of fixture length), stage 2: 25% < bone loss < 50%, stage 3: bone loss > 50%) and performance of resective or regenerative surgery was investigated. Kaplan-Meier survival curves and Cox hazards proportional models were used to analyze the cumulative survival rate of implants. Median survival time, predicted mean survival time, hazard ratio (HR), and 95% confidence interval (CI) were calculated. RESULTS: Based on Kaplan-Meier analysis, 89 patients and 227 implants were included, and total median postoperative survival duration was 8.96 years. Cumulative survival rates for stage 1, 2, and 3 were 70.7%, 48.9%, and 21.3%, respectively. The mean survival time for implants in stage 1, 2, and 3 was 9.95 years, 7.96 years, and 5.67 years, respectively, with statistically significant difference (log-rank p-value < 0.001). HRs for stage 2 and stage 3 were 2.25 and 4.59, respectively, with stage 1 as reference. Significant difference was not found in survival time between resective and regenerative surgery groups in any peri-implantitis stage. CONCLUSIONS: The initial bone loss rate relative to the fixture length significantly correlated with the outcome after peri-implantitis surgery, demonstrating a notable difference in the long-term survival rate. Difference was not found between resective surgery and regenerative surgery in implant survival time. Bone loss rate could be utilized as a reliable diagnostic tool for evaluating prognosis after surgical treatment, regardless of surgical method used. TRIAL REGISTRATION: Retrospectively registered. (KCT0008225).

Supporting students with complex needs living in rural and regional New South Wales: is wraparound the answer?

Historically, students attending school in rural and regional New South Wales have experienced poorer outcomes than their peers attending metropolitan schools. The lack of coordinated support services for students with complex support needs compounds this issue. Wraparound models of support have been successful in improving outcomes for students with complex support needs, and the New South Wales government has prioritised the establishment of strong relationships between schools and communities to overcome the limitations of geographic isolation. The aim of the current study was to explore wraparound support for students with complex support needs attending schools in rural and regional New South Wales. A qualitative research approach was employed, and semi-structured interviews were conducted with key stakeholders to gain an in-depth understanding of current successes, barriers, and needs. The findings indicated that wraparound was most effective in rural and remote schools when school staff implemented bespoke approaches to wraparound, such as restorative practices. Resourcing was a barrier found to be central to all schools. Recommendations are provided to enhance the capacity of rural and regional NSW schools to provide wraparound support for students with complex support needs.

Determination of Toxic Elements and Arsenic Species in Salted Foods and Sea Salt by ICP-MS and HPLC-ICP-MS.

Toxic elements (Cd, Pb, and As) accumulate into the environment by industrialization and natural phenomena and then pass to organisms. Analysis of toxic elements in food must be accurately carried out on a regular basis so as to avoid any adverse impact. Salted foods are difficult samples and accurate analysis of As is not easy due to salt interference. In this study, analysis of As was carried without influence of salts in three types of salted foods via an analytical method, which was validated using spiking recovery experiments and by analyzing certified reference materials. As a result, toxic elements were detected in all samples but none of these exceeded the World Health Organization recommended limits. Among the As species, arsenobetaine (AsB) was the most abundant, while inorganic As was below the detection limit in all samples. All the analyzed salted food samples appeared to be safe for consumption. In addition, the analysis of sea shrimp, freshwater shrimp, and seawater verified As bioaccumulation in these organisms from the environment.

Associations between mandibular torus and types of temporomandibular disorders, and the clinical usefulness of temporary splint for checking bruxism.

BACKGROUND: Occlusal stress from oral parafunctional habits is one of the causes of temporomandibular disorders (TMD) and mandibular torus (MT). Although some studies have investigated the correlation between TMD and MT, understanding of the relationships between types of TMD and MT is insufficient. Therefore, we conducted this study to investigate the associations between presence of MT and TMD types. METHODS: This study included 77 patients diagnosed with TMD who first visited our clinic for TMD between March 2019 and July 2020. Among them, 30 (38.9%) had MT, and 54 (70.1%) had oral parafunction. Parafunctional activity during sleep was confirmed using a temporary splint for checking bruxism (TSCB). RESULTS: The relationship between prevalence of MT and oral parafunction in TMD patients was not statistically significant (P = 0.131), but the odds ratio was relatively high at 2.267. An analysis of TMD type revealed that Type I, which is classified as myalgia of the masticatory muscles, and MT had a significant association (P = 0.011). We fabricated a TSCB for 27 patients to wear during sleep and confirmed that 23 (85.2%) had nocturnal bruxism. The TSCB results and presence of MT showed a significant relationship (P = 0.047). CONCLUSION: Through the results of this study, clinicians may consider the hyperactivity of masticatory muscles in the presence of MT when treating TMD patients. In addition, TSCB has a great diagnostic value as it can be easily manufactured and be useful for discovering pre-existing oral parafunctions that patients are not aware of.

Proteomic evaluation of kimchi, a traditional Korean fermented vegetable, and comparison of kimchi manufactured in China and Korea.

Kimchi is a traditional Korean fermented vegetable, which is also widely consumed in Japan and China. However, little is known about the kimchi proteome. In this study, Korean and Chinese kimchi proteomes were evaluated by shotgun proteomics. A total of 250 proteins were annotated, and 29 of these were expressed at > 1% of the average relative abundance. Discrimination of the geographical origins of Korean and Chinese kimchi samples was possible using multivariate analysis of the proteomic data, and 23 proteins were expressed differently between the two types (p < 0.001), and represent possible markers to discriminate between Chinese and Korean kimchi. This study provides important insights into the kimchi proteome and illustrates the proteomic differences caused by geographical origin.

Interfacial behavior of surfactant-covered double emulsion in extensional flow.

We analyze the interface-interface interactions of a surfactant-covered double emulsion using the lattice Boltzmann method and study the interaction of the inner and outer interfaces and the local surfactant distribution under a uniaxial extensional flow. First, the capillary effects are analyzed. Upon surfactant application, the outer droplet deformation increases and the inner droplet deformation decreases. The concentrated surfactants on the outer interface increase deformation, and the inner droplet is affected by the inner flow. At a fixed Péclet number (Pe), the surfactant concentration at the outer interface increases with an increase in capillary number (Ca); however, such a tendency is difficult to identify at the inner interface. Next, the Pe effects are analyzed. With an increase in Pe, the deformation of the inner droplet decreases significantly. The local distribution of the surfactant considerably affects the double emulsion stabilization, which is analyzed in terms of internal flow. The interfacial tension gradient induced by the surfactant generates vortices internally, which is verified by applying the surfactant to each interface independently. The radius ratio affects droplet deformation and surfactant transport. The compression of the inner flow region increases the viscous force and decreases the interface velocity. Therefore, with an increase in radius ratio, the deformation increases, and the surfactant transport becomes slow.

Rapid and Simultaneous Quantification of Five Quinolizidine Alkaloids in Lupinus angustifolius L. and Its Processed Foods by UPLC-MS/MS.

Quinolizidine alkaloids (QAs) are toxic secondary metabolites of Lupinus plants. This study reports the simultaneous quantification of five alkaloids from Lupinus angustifolius L. and its processed foods by ultraperformance liquid chromatography with electrospray ionization mass spectrometric detection. After optimizing the extraction conditions, the analytical method was validated for the QAs in lupin beans and in three major processed foods through detection and quantification limits, linearity, precision, and accuracy. The detection and quantification limits of the QAs were 0.5-1.7 and 1.5-5.7 mg kg-1, respectively. The linearity was greater than 0.9992 and the precision was less than 3.1%. The recoveries of three different concentrations of each QA were 89.5-106.2%. For both raw lupin beans and processed foods, lupanine was the major QA, and 13-hydroxylupanine and angustifoline were detected in lower amounts. This method could be widely used for accurate risk assessment of QAs.

Efficacy and safety of nanoparticle-albumin-bound paclitaxel compared with solvent-based taxanes for metastatic breast cancer: A meta-analysis.

The curative effects of nanoparticle albumin-bound (nab)-paclitaxel in the first-line treatment of metastatic breast cancer (MBC) are still controversial, with even more after the removal of marketing approval of indication of bevacizumab. Five electronic databases and the related resources were searched for eligible randomized clinical trials (RCTs) without year and language restrictions to perform a meta-analysis. The studies were comparing the efficacy and safety between nab-paclitaxel chemotherapy versus solvent-based (sb)-taxanes chemotherapy such as sb-paclitaxel and docetaxel. The primary end points were overall response rate (ORR) and disease control rate (DCR). Secondary end points were progression-free survival (PFS), overall survival (OS), adverse events (AEs), and dose discontinuation rate (DDR). Five RCTs (1,554 patients) were finally identified from 1,902 studies. When compared to sb-paclitaxel, nab-paclitaxel showed significant beneficial effects in terms of ORR (OR 2.39, 95% CI 1.69-3.37, p < 0.001), DCR (OR 1.89, 95% CI 1.07-3.35, p = 0.03), and PFS (HR 0.75, 95% CI 0.62-0.90, p = 0.002). Nab-paclitaxel also showed significantly longer OS (HR 0.73, 95% CI 0.54-0.99, p = 0.04) than docetaxel. AEs and DDR were comparable between the two arms. Using nab-paclitaxel could significantly improve efficacy with comparable toxicities in the treatment of MBC.

Dietary intake assessment of macro, trace, and toxic elements via consumption of kimchi in South Korea.

BACKGROUND: Kimchi, a Korean food, is now produced and consumed in many other countries. In this study, the macro, trace, and toxic element content in varieties of kimchi from Korea and China was evaluated. A health-risk assessment, involving the calculation of estimated dietary intake values, was undertaken. To discriminate among samples with different origins, linear discriminant analysis (LDA) and principal component analysis (PCA) were applied. RESULTS: The analytical methods used for analyses were successfully validated. In the quantification of elements, no significant differences were detected in the concentrations of the elements that were analyzed. Based on the fresh weight of kimchi, among the macro elements, the concentrations of Na and K were relatively high, with values of 4181.9-9919.8 mg kg-1 and 1661.8-6623.5 mg kg-1 , respectively. Among the trace elements that were analyzed, zinc in Korean samples, and strontium in Chinese samples showed the highest concentrations, at 2.67 and 2.98 mg kg-1 , respectively. Concentration levels of the toxic trace elements were within permissible limits. The amount of daily intake and provisional daily intake of each element calculated based on kimchi intake confirmed its safety for consumption. Linear discriminant analysis and PCA successfully discriminated among the samples of Chinese and Korean origin. CONCLUSION: The provisional daily intake of the toxic elements was within provisional tolerable daily intake values. The normal intake of kimchi could not pose a threat upon consumption. © 2019 Society of Chemical Industry.

Cone-beam computed tomography analysis of transverse dental compensation in patients with skeletal Class III malocclusion and facial asymmetry.

OBJECTIVE: The purpose of this study was to analyze the transverse dental compensation in reference to the maxillary and mandibular basal bones using cone-beam computed tomography (CBCT) and evaluate the correlations between transverse dental compensation and skeletal asymmetry variables in patients with skeletal Class III malocclusion and facial asymmetry. METHODS: Thirty patients with skeletal Class I (control group; 15 men, 15 women) and 30 patients with skeletal Class III with menton deviation (asymmetry group; 16 men, 14 women) were included. Skeletal and dental measurements were acquired from reconstructed CBCT images using OnDemand3D 1.0 software. All measurements were compared between groups and between the deviated and nondeviated sides of the asymmetry group. Correlation coefficients for the association between skeletal and dental measurements were calculated. RESULTS: Differences in the ramus inclination (p < 0.001), maxillary canine and first molar inclinations (p < 0.001), and distances from the canine and first molar cusp tips to the midmaxillary or midmandibular planes (p < 0.01) between the right and left sides were significantly greater in the asymmetry group than in the control group. In the asymmetry group, the ramus inclination difference (p < 0.05) and mandibular canting (p < 0.05) were correlated with the amount of menton deviation. In addition, dental measurements were positively correlated with the amount of menton deviation (p < 0.05). CONCLUSIONS: Transverse dental compensation was correlated with the maxillary and mandibular asymmetry patterns. These results would be helpful in understanding the pattern of transverse dental compensation and planning surgical procedure for patients with skeletal Class III malocclusion and facial asymmetry.

Efficacy of Combination Treatment with Sodium Metasilicate and Sodium Hypochlorite for Inactivation of Norovirus on Fresh Vegetables.

In recent years, fresh vegetables have frequently been associated with the foodborne transmission of enteric viruses, such as human norovirus (NoV). Therefore, several studies have focused on developing methods to inactivate foodborne viruses for preventing outbreaks of foodborne illnesses. Sodium hypochlorite (NaOCl) is commonly used as a disinfectant, but results in undesirable effects on the appearance and taste of foods and can generate toxic byproducts when it exceeds the allowable concentration. Here, we evaluated the efficacy of a range of NaOCl concentrations (50-1000 ppm) for reducing the amounts of human NoV (NoV GII.4) on lettuce (Lactuca sativa), celery (Apium graveolens L.), and white cabbage (Brassica oleracea ssp. capitata). In addition, the combination treatment of NaOCl and sodium metasilicate (SMS, 0.1-0.5%) pentahydrate was evaluated for its ability to decrease the populations of NoV GII.4 in the three food samples. An immunomagnetic separation procedure combined with reverse transcription quantitative polymerase chain reaction was used for virus detection. For lettuce, celery, and cabbage, the NoV GII.4 recovery rates were 57.3% ± 6.5%, 52.5% ± 1.7%, and 60.3% ± 3.9%, respectively, using a glycine/NaCl elution buffer (0.25 M glycine/0.14 M NaCl, pH 9.5). The reductions of NoV GII.4 were 3.17, 3.06, and 3.27 log10 genomic copies/µL for lettuce, celery, and cabbage, respectively, at 1000 ppm NaOCl, while a reduction of ∼3 log10 genomic copies/µL was obtained when the samples were treated with a combination of 100 ppm NaOCl and 0.4% SMS pentahydrate. Taken together, these results demonstrated that combined treatment with NaOCl and SMS pentahydrate was an efficient strategy to reduce the concentration of NaOCl for control of NoV GII.4 contamination in fresh vegetables.

Pretreatment with propidium monoazide/sodium lauroyl sarcosinate improves discrimination of infectious waterborne virus by RT-qPCR combined with magnetic separation.

RT-qPCR allows sensitive detection of viral particles of both infectious and noninfectious viruses in water environments, but cannot discriminate non-infectious from infectious viruses. In this study, we aimed to optimize RT-qPCR-based detection of chlorine-inactivated human norovirus (NoV) and pepper mild mottle virus (PMMoV) in suspension by pretreatment with an optimal combination of a monoazide and a detergent that can efficiently penetrate damaged viral capsids. Four methods were compared to determine the efficacy of chlorine disinfection (at 1, 3, and 5 min mg/L): (A) RT-qPCR alone, (B) RT-qPCR assay preceded by magnetic bead separation for enrichment of viral particles (MBS-RT-qPCR), (C) MBS-RT-qPCR assay with pretreatment with propidium monoazide (PMA-MBS-RT-qPCR), and (D) PMA-MBS-RT-qPCR assay with pretreatment with sodium lauroyl sarcosinate (INCI-PMA-MBS-RT-qPCR). On the basis of a PMA optimization assay, 200 and 300 µM PMA were used in subsequent experiments for NoV GII.4 and PMMoV, respectively. Optimal INCI concentrations, having minimal influence on NoV GII.4 and PMMoV, were found to be 0.5% and 0.2% INCI, respectively. For NoV GII.4, there were significant differences (P < 0.05) in log10 genome copies between the PMA-treated and the INCI + PMA-treated samples (log10 genome copies differed by 1.11 and 0.59 log10 for 3 and 5 min mg/L of chlorine, respectively). For PMMoV, INCI induced differences in log10 genome copies of 0.92, 1.18, and 1.86, for 1, 3, and 5 min mg/L of chlorine, respectively. Overall, the results of this study indicate that an optimal combination of PMA and INCI could be very useful for evaluating disinfection methods in water treatment strategies.

Correlation between Changes in Microbial/Physicochemical Properties and Persistence of Human Norovirus during Cabbage Kimchi Fermentation.

Recently, cabbage kimchi has occasionally been associated with the foodborne diseases of enteric viruses such as human norovirus (HuNoV). This study aimed to evaluate the correlation between microbial/physicochemical properties and persistence of HuNoV in experimentally contaminated cabbage kimchi fermented and stored at 4°C or 10°C for 28 days. Changes in organic acid content, lactic acid bacteria (LAB), acidity, pH, and salinity were analyzed. The recovery of structurally intact HuNoV was examined for up to 28 days post-inoculation, using a NoV GII.4 monoclonal antibody-conjugated immuno-magnetic separation method combined with quantitative real-time reverse transcription polymerase chain reaction. On day 0, LAB loads were 4.70 log10 colony forming units/g and HuNoV GII.4 titers were 2.57 log10 genomic copies/µl, at both temperatures. After 28 days, intact HuNoV titers decreased to 1.58 (4°C) and 1.04 (10°C) log10 genomic copies/µl, whereas the LAB density increased. This correlated with a gradual increase in lactic acid and acetic acid at both temperatures. Our findings support a statistical correlation between changes in physicochemical properties and the recovery of structurally intact HuNoV GII.4. Moreover, we determined that the production of organic acid and low pH could affect HuNoV GII.4 titers in cabbage kimchi during fermentation. However, HuNoV GII.4 was not completely eliminated by microbial/physicochemical factors during fermentation, although HuNoV GII.4 was reduced. Based on this, we speculate that the persistence of HuNoV GII.4 may be affected by the continually changing conditions during kimchi fermentation.

Exploration of the metal coordination region of concanavalin A for its interaction with human norovirus.

Rapid methods for the detection and clinical treatment of human norovirus (HuNoV) are needed to control foodborne disease outbreaks, but reliable techniques that are fast and sensitive enough to detect small amounts of HuNoV in food and aquatic environments are not yet available. We explore the interactions between HuNoV and concanavalin A (Con A), which could facilitate the development of a sensitive detection tool for HuNoV. Biophysical studies including hydrogen/deuterium exchange (HDX) mass spectrometry and surface plasmon resonance (SPR) revealed that when the metal coordinated region of Con A, which spans Asp16 to His24, is converted to nine alanine residues (mCon AMCR), the affinity for HuNoV (GII.4) diminishes, demonstrating that this Ca2+ and Mn2+ coordinated region is responsible for the observed virus-protein interaction. The mutated carbohydrate binding region of Con A (mCon ACBR) does not affect binding affinity significantly, indicating that MCR of Con A is a major region of interaction to HuNoV (GII.4). The results further contribute to the development of a HuNoV concentration tool, Con A-immobilized polyacrylate beads (Con A-PAB), for rapid detection of genotypes from genogroups I and II (GI and GII). This method offers many advantages over currently available methods, including a short concentration time. HuNov (GI and GII) can be detected in just 15 min with 90% recovery through Con A-PAB application. In addition, this method can be used over a wide range of pH values (pH 3.0 - 10.0). Overall, this rapid and sensitive detection of HuNoV (GI and GII) will aid in the prevention of virus transmission pathways, and the method developed here may have applicability for other foodborne viral infections.

Comparison of the bonding strengths of second- and third-generation light-emitting diode light-curing units.

OBJECTIVE: With the introduction of third-generation light-emitting diodes (LEDs) in dental practice, it is necessary to compare their bracket-bonding effects, safety, and efficacy with those of the second-generation units. METHODS: In this study, 80 extracted human premolars were randomly divided into eight groups of 10 samples each. Metal or polycrystalline ceramic brackets were bonded on the teeth using second- or third-generation LED light-curing units (LCUs), according to the manufacturers' instructions. The shear bond strengths were measured using the universal testing machine, and the adhesive remnant index (ARI) was scored by assessing the residual resin on the surfaces of debonded teeth using a scanning electron microscope. In addition, curing times were also measured. RESULTS: The shear bond strengths in all experimental groups were higher than the acceptable clinical shear bond strengths, regardless of the curing unit used. In both LED LCU groups, all ceramic bracket groups showed significantly higher shear bond strengths than did the metal bracket groups except the plasma emulation group which showed no significant difference. When comparing units within the same bracket type, no differences in shear bond strength were observed between the second- and third-generation unit groups. Additionally, no significant differences were observed among the groups for the ARI. CONCLUSIONS: The bracket-bonding effects and ARIs of second- and third-generation LED LCUs showed few differences, and most were without statistical significance; however, the curing time was shorter for the second-generation unit.

Detection of hepatitis A virus in seeded oyster digestive tissue by ricin A-linked magnetic separation combined with reverse transcription PCR.

Outbreaks of hepatitis A virus (HAV) infections are most frequently associated with the consumption of contaminated oysters. A rapid and selective concentration method is necessary for the recovery of HAV from contaminated oysters prior to detection using PCR. In this study, ricin extracted from castor beans (Ricinus communis) was tested as an alternative to antibody used in immunomagnetic separation while concentrating HAV prior to its detection using reverse transcription PCR. Initially, the extracted proteins from castor beans were fractionated into 13 fractions by gel filtration chromatography. Pretreatment of different protein fractions showed a variation in binding of HAV viral protein (VP) 1 to oyster digestive tissue in the range of 25.9 to 63.9%. The protein fraction, which caused the highest reduction in binding of VP1 to the tissue, was identified as ricin A by quadrupole time-of-flight mass spectrometry. Ricin A could significantly inhibit binding of VP1 to the tissue with a 50% inhibitory concentration of 4.5 µg/ml and a maximal inhibitory concentration of 105.2%. The result showed that the rate of inhibition of HAV binding to tissue was higher compared to the rate of ricin itself binding to HAV (slope: 0.0029 versus 0.00059). However, ricin A concentration showed a higher correlation to the relative binding of ricin itself to HAV than the inhibition of binding of HAV to the tissue (coefficient of determination, R(2): 0.9739 versus 0.6804). In conclusion, ricin A-linked magnetic bead separation combined with reverse transcription PCR can successfully detect HAV in artificially seeded oyster digestive tissue up to a 10(-4) dilution of the virus stock (titer: 10(4) 50% tissue culture infective dose per ml).

Development of lectin-linked immunomagnetic separation for the detection of hepatitis a virus.

The accuracy and sensitivity of PCR-based methods for detection of hepatitis A virus (HAV) are dependent on the methods used to separate and concentrate the HAV from the infected cells. The pH and ionic strength affect the binding affinity of the virus to cells. In this study, we initially investigated the effects of pH (4.0-10.0) and metal ions (Fe²âº, Co²âº, Cu²âº, Mg²âº, K⁺, and Ca²âº) on the binding of HAV to oyster digestive cells. The lowest relative binding (RB) of HAV to the cells was found at pH 4.0 and in FeSO4 solution (64.6% and 68.1%, respectively). To develop an alternative to antibody-dependent immunomagnetic separation prior to detection of HAV using RT-PCR, the binding of HAV to five lectins, peanut agglutinin (PNA), Dolichos biflorus agglutinin (DBA), Helix pomatia agglutinin (HPA), Ulex europaeus agglutinin (UEA-1) and soybean agglutinin (SBA), was evaluated using ELISAs. SBA showed significantly higher RB to HAV than the other lectins tested. In addition, HAV could be concentrated within 30 min using SBA-linked magnetic bead separation (SMS) prior to the RT-PCR assay. Our findings demonstrate the feasibility of using SMS combined with RT-PCR to detect HAV at dilutions ranging from 10⁻¹-10⁻4 of a HAV stock (titer: 104 TCID50/mL).