An investigation for the efficacy of teaching model of combining virtual simulation and real experiment for clinical microbiology examination.

Background: As a convenient teaching tool, virtual simulation experiment technology had been widely utilized in the field of medical education. However, virtual learning could not fully replace the benefits of in-person instruction. Therefore, finding ways to integrate both methods was crucial for achieving optimal educational outcomes. The objective of this study was to compare the effectiveness of the self-built virtual simulation and design experiment combining teaching mode and the traditional experimental teaching mode in the clinical microbiology examination experiment teaching. Methods: This study was conducted at Shandong First Medical University in China. The experimental group consisted of 100 third-year students from the grade 2020 majoring in medical examination technology, who underwent an innovative teaching model combining virtual and real experiments. The control group comprised of 100 third-year students from the grade 2019 in the same major, who received traditional experimental teaching model. In this study, we referred to grade 2020 as cohort 2020 and grade 2019 cohort 2019. The performance of both groups was assessed via experimental and theoretical testing. Meanwhile, survey questionnaires were administered to evaluate the efficacy of the innovative experimental teaching model and students' level of satisfaction with it. Cohort 2020 conducted a survey for modules 1 to 4, while cohort 2019 only conducted a survey for module 4, as detailed in the Appendix. Results: The majority of students in the experimental group expressed satisfaction with the teaching model that combined virtual and real experiments, as evidenced by their superior performance on both experimental operational skills (87.54 ± 8.93 vs. 82.39 ± 10.55) and theoretical knowledge tests (83.65 ± 9.02 vs. 80.18 ± 8.24) compared to those in the control group. Conclusion: The combination of virtual simulation experiment and design experiment in the microbiological examination of clinical specimens represented an effective pedagogical approach. The instructional approach had the potential to incite a passion for learning, enhance proficiency in standardized experimental techniques, foster the ability to integrate theory with practice, and cultivate clinical reasoning skills.

The effects of ultrasonic processing on the antioxidant activity of Geotrichum candidum LG-8 and its cell wall extracts.

Introduction: Extraction techniques that influence cell wall polysaccharides (EPS) is crucial for maximizing their bioactivity. This study evaluates ultrasound technology for extracting antioxidant polysaccharides from Geotrichum candidum LG-8, assessing its impacton antioxidant activity. Methods: Ultrasound extraction of EPS from G. candidum LG-8 was optimized (18 min, pH 7.0, 40 W/cm2, 0.75 M NaCl). ABTS scavenging efficiency and monosaccharide composition of LG-EPS1 and LG-EPS3 were analyzed using Fourier-transform infrared spectroscopy (FTIR) and scanning electron microscopy (SEM). Results: The Results showed that ultrasonic treatment markedly increased the ABTS radical scavenging efficiency of LG-8 cells by 47%. At a concentration of 1 mg/mL, the ultrasonically extracted LG-EPS1 and LG-EPS3 polysaccharides exhibited significant ABTS radical scavenging efficiencies of 26% and 51%, respectively. Monosaccharide composition analysis identified mannose and glucose in LG-EPS1, while LG-EPS3 was primarily composed of mannose. FTIR spectra verified the polysaccharides' presence, and SEM provided visual confirmation of the nanoparticle structures characteristic of LG-EPS1 and LG-EPS3. Discussion: This research not only underscores the technological merits of ultrasound in polysaccharide extraction but also highlights the potential of G. candidum LG-8 derived polysaccharides as valuable bioactive compounds for antioxidant utilization.

The structure and in vitro starch digestibility of wheat starch-glycerol monopalmitin complexes: the effect of heating conditions and water content.

The effects of the heating conditions and water content on the structure and digestibility of wheat starch (WS)-glycerol monopalmitin (GMP) complexes were investigated. The results showed that the higher water content and the heating conditions of 90°C for 60 min after 100°C for 10 min favor the formation of more WS-GMP complexes with the greater short-range order, although the thermal transition temperatures of GWS-GMP-100 complexes are not significantly affected by the water content. Only the type I complexes were formed under the heating conditions of 90°C for 60 min. The heating conditions of 90°C for 60 min after 100°C for 10 min facilitates the formation of type II complexes, and the amounts of type II complexes increased with increasing water content. The digestion rates of WS-GMP complexes decreased slightly with increasing water content, and the extent of starch amylolysis of WS-GMP complexes significantly decreased after heating further at 90°C compared with that only heating at 100°C. The digestibility of complexes is mainly related to structural order rather than the number of complexes. This study is helpful to further understand starch-lipid complexes by showing that heating conditions and water content influence the formation of WS-GMP complexes.

Capacity and kinetics of zearalenone adsorption by Geotrichum candidum LG-8 and its dried fragments in solution.

The application of LG-8 and its dry fragments as zearalenone (ZEN) adsorbents was investigated. The study showed that Geotrichum candidum LG-8 and its fragments dried at 55°C or through lyophilization are able to adsorb around 80% of ZEN. However, besides in water and 55°C-drying conditions, SEM indicated that higher 90% of ZEN binding tended to occur when cell walls of fragments were intact with less adhesion among themselves. Notably, ZEN/LG-8 fragments complexes were quite stable, as only 1.262% and 1.969% of ZEN were released after successive pH treatments for 4 h and 5 min. The kinetic data signified that adsorption of ZEN onto LG-8 fragments followed well the pseudo-first-order kinetic model. Isotherm calculations showed Langmuir model was favourable and monolayer adsorption of ZEN occurred at functional binding sites on fragments surface. Therefore, we conclude that it can be an alternative biosorbent to treat water contained with ZEN, since LG-8 is low-cost biomass and its fragments have a considerable high biosorption capacity avoiding impacting final product quality and immunodeficient patients.

Lentinus edodes Polysaccharides Alleviate Acute Lung Injury by Inhibiting Oxidative Stress and Inflammation.

Acute lung injury (ALI) is a kind of lung disease with acute dyspnea, pulmonary inflammation, respiratory distress, and non-cardiogenic pulmonary edema, accompanied by the mid- and end-stage characteristics of COVID-19, clinically. It is imperative to find non-toxic natural substances on preventing ALI and its complications. The animal experiments demonstrated that Lentinus edodes polysaccharides (PLE) had a potential role in alleviating ALI by inhibiting oxidative stress and inflammation, which was manifested by reducing the levels of serum lung injury indicators (C3, hs-CRP, and GGT), reducing the levels of inflammatory factors (TNF-α, IL-1ß, and IL-6), and increasing the activities of antioxidant enzymes (SOD and CAT) in the lung. Furthermore, PLE had the typical characteristics of pyran-type linked by ß-type glycosidic linkages. The conclusions indicated that PLE could be used as functional foods and natural drugs in preventing ALI.

HBV T1719G mutation reduced HBV replication through mutant Enh II and HBx protein in vitro.

It was repeatedly reported that the hepatitis B virus (HBV) T1719G mutation was very common and related to progression and malignancy of liver disease. However, its effect on viral replication efficiency remains unclear. In this study, we aimed to evaluate the function and mechanisms of the T1719G mutation on viral replication capacity. Wild-type and T1719G mutation-bearing HBV1.2× plasmids were transfected into Huh7 and HepG2 cells, respectively, and HBV total RNA, 3.5 kb RNA and supernatant HBV DNA were assessed using real-time PCR, hepatitis B surface antigen (HBsAg) and hepatitis B e antigen (HBeAg) levels were measured by time-resolved fluoroimmunoassay. In order to assess Enh II activity and the binding capacity of HNF3ß to Enh II sequence, dual-luciferase assay and Chromatin immunoprecipitation (ChIP)-PCR were employed, respectively. Simultaneously, the HBx or HBx-mut (T1719G) plasmid was co-transfected to evaluate the effect of HBx on viral replication. Our results showed that the T1719G mutation impaired viral replication efficacy compared with the wild type both by reducing Enh II activity and binding capacity of HNF3ß with Enh II. And such reduction caused by T1719G mutation could be rescued by HBx protein. Our results show that the T1719G mutation decreases HBV viral replication capacity possibly by mutant HBx protein and altered Enh II activity.

Complete genome sequencing and clinical analysis of intrahepatic hepatitis B virus cccDNA from HCC.

BACKGROUND: More than half of hepatocellular carcinomas (HCCs) are etiologically attributed to hepatitis B virus (HBV) infection, but it remains unclear whether HBV mutations are virological factors that contribute to formation of HCC or instead reflect accumulation during the progression of HBV-related disease. METHODS: Rolling-cycle amplification and PCR sequencing were used to characterize covalently closed circular DNA (cccDNA) mutations in tumor tissues. Paired non-tumor tissues were used as controls. RESULTS: High frequencies of C1653T, T1753V, and A1762T/G1764A cccDNA mutations were observed in both tumor and non-tumor tissues. T1719G, C1329A, and T3098C mutations were related to the overall survival of HCC patients. Patients with G1719 tended to be in the high Barcelona Clinic Liver Cancer stage and had lower levels of total DNA and cccDNA per cell than patients with T1719. Additionally, in vitro analysis revealed that T1719G mutation reduced viral replication efficacy. Finally, significantly higher levels of preoperative alpha-fetoprotein were observed in patients harboring the G1078T, C1653T, G1727A, C1913A, T1978C, or C3116T mutations at the cccDNA level. CONCLUSIONS: We speculated that HBV cccDNA mutations accumulated over the course of HBV-related disease development, and that some key mutations had prognostic value for patients with HBV-related HCC.

Processing optimization and anti-oxidative activity of enzymatic extractable polysaccharides from Pleurotus djamor.

In present study, a quadratic regression model based on the response surface methodology (RSM) coupled with Box-Behnken design (BBD) was employed to optimize the enzyme-assisted extraction (EAE) process of enzymatic-extractable Pleurotus djamor polysaccharides (EnPPs). By solving the regression equations and analyzing the model graphs, the optimum conditions were at pH 7.36, water to material ratio 56.78, extraction time 44.77min and extraction temperature 35.36°C, respectively. Under these conditions, the experimental yields of EnPPs reached 3.61%, which were in good agreement with the validated values (3.57±0.51%). Basic physicochemical properties including molecular weights, structural identification by FT-IR and monosaccharide compositions were processed. In addition, the in vivo inhibiting activities against oxidative stress were investigated. The results provided an alternative bioresource for the exploitation of EnPPs under EAE from P. djamor, and the EnPPs had potential effects in prevention of oxidative stress.

[Construction of a recombinant eukaryotic expression vector containing HSV 2gD and its immune responses].

AIM: To construct and express pcDNA3.1-IL-18-HSV P6 recombinant DNA vaccine, and to observe the immune responses of pcDNA3.1-IL-18-HSV P6 genetic vaccine. METHODS: Genes of P6 and IL-18 were cloned into the eukaryotic expression vector pcDNA3.1(-) respectively, and confirmed by enzyme digestion and sequence analysis, the recombinant plasmid pcDNA3.1-IL-18-HSVP6 was transformed into CHO cells. The expressed protein was characterized by indirect immunofluorescence and Western blotting. The recombinant plasmid pcDNA3.1-IL-18-HSVP6 was used to inoculate BALB/c mice by intramuscular injection for three times, once a week. One week after the last vaccination, the levels of specific IgG antibody, IFN-γ and IL-18 were detected by ELISA; One month after the last vaccination, spleen cells of vaccinated mice were separated and proliferation of spleen lymphocytes were detected by MTT assay. RESULTS: Recombinant pcDNA3.1-IL-18-HSV P6 plasmid was confirmed by enzyme digestion and DNA sequencing. After inoculated by pcDNA3.1-IL-18-HSV P6 vaccine, the mice could produce higher level of splenocytes proliferation and secrected higher level of IFN-γ, IL-18 and specific antibody. CONCLUSION: DNA vaccine of pcDNA3.1-IL-18-HSV P6 has been successfully constructed, and it can effectively induce humoral and cellular immune responses, which provided a basis for constructing new type of DNA vaccine.

[Cloning and expression of human interleukin-32 and studies on its bioactivity].

AIM: To clone human interleukin-32(hIL-32) gene and express it in E.coli efficiently. METHODS: The gene of hIL-32 was amplified by RT-PCR from human peripheral blood mononuclear cells (PBMC) which stimulated with Con A for 60 h. The PCR product was inserted into the pMD18-T vector. The hIL-32 cDNA confirmed by sequencing was inserted into expression vector pET-30a(+) and expressed in E.coli BL21(DE3) strain. The hIL-32 protein expression was induced by IPTG and assayed by SDS-PAGE and coomassie blue stain. The recombination protein was identified by Western blot and its biological activity was analyzed. RESULTS: DNA sequencing confirmed that the cloned cDNA was identical to the published sequence of hIL-32 that the nucleotide sequence of this gene was 567 bp. The recombinant plasmid pET30a-hIL32 was transformed into E.coli BL21(DE3) strain for expression. An expected 28 kDa protein of hIL-32 was found mainly in the induced host strains by SDS-PAGE and coomassie blue stain. The 28 kDa protein was recognized by anti-IL-32 antibody in western blot. The purified recombination protein could induce the producing of IL-6 in the human PBMC. CONCLUSION: We have successfully cloned the gene and expressed the protein of hIL-32 and the expressed protein has specific bioactivity.