Objective To evaluate the performance of microfluidic chips in the identification and genotyping of Malassezia species.Methods This study included 6 reference Malassezia strains and clinical Malassezia isolates from the scrapings of patients with pityriasis versicolor and follicular contents of patients with Malassezia folliculitis.These isolates were identified by DNA sequencing,random amplified polymorphic DNA (RAPD)-PCR and microfluidic chips.Cluster analysis was carried out and tree diagrams were generated.Results A total of 83 Malassezia isolates were obtained from 72 patients with pityriasis versicolor and 11 patients with Malassezia folliculitis.Genomic DNA of most strains was successfully amplified by PCR with two primers S22 and S24,and PCR with S22 primer produced more stable and clear amplification bands than that with S24.Positive bands of different sizes were repetitively obtained by using microfluidic chips,with interspecies and intraspecies polymorphisms observed in all the strains.On the basis of DNA sequencing,microfluidic chips and RAPD-PCR could be used to successfully distinguish the following eight species,i.e.,M.furfur,M.sympodialis,M.globosa,M.pacbydermatis,M.slooffiae,M.Japonica,M.yamatoensis and M.dermatis.Conclusions As a rapid,high-throughput and high-sensitivity method,microfluidic chips combined with RAPD-PCR shows an advantage for analyzing interspecies genetic diversity,genetic relationship of Malassezia species,as well as for identifying new Malassezia species.

Methanotrophs could degrade methane and various chlorinated hydrocarbons. The analysis on methane monooxygenase gene cluster sequence would help to understand its catalytic mechanism and enhance the application in pollutants biodegradation. The methanotrophs was enriched and isolated with methane as the sole carbon source in the nitrate mineral salt medium. Then, five chlorinated hydrocarbons were selected as cometabolic substrates to study the biodegradation. The phylogenetic tree of 16S rDNA using MEGE5.05 software was constructed to identify the methanotroph strain. The pmoCAB gene cluster encoding particulate methane monooxygenase (pMMO) was amplified by semi-nested PCR in segments. ExPASy was performed to analyze theoretical molecular weight of the three pMMO subunits. As a result, a strain of methanotroph was isolated. The phylogenetic analysis indicated that the strain belongs to a species of Methylocystis, and it was named as Methylocystis sp. JTC3. The degradation rate of trichloroethylene (TCE) reached 93.79% when its initial concentration was 15.64 μmol/L after 5 days. We obtained the pmoCAB gene cluster of 3 227 bp including pmoC gene of 771 bp, pmoA gene of 759 bp, pmoB gene of 1 260 bp and two noncoding sequences in the middle by semi-nested PCR, T-A cloning and sequencing. The theoretical molecular weight of their corresponding gamma, beta and alpha subunit were 29.1 kDa, 28.6 kDa and 45.6 kDa respectively analyzed using ExPASy tool. The pmoCAB gene cluster of JTC3 was highly identical with that of Methylocystis sp. strain M analyzed by Blast, and pmoA sequences is more conservative than pmoC and pmoB. Finally, Methylocystis sp. JTC3 could degrade TCE efficiently. And the detailed analysis of pmoCAB from Methylocystis sp. JTC3 laid a solid foundation to further study its active sites features and its selectivity to chlorinated hydrocarbon.
Methylocystaceae ; classification ; metabolism ; Multigene Family ; Oxygenases ; genetics ; Phylogeny ; Polymerase Chain Reaction ; RNA, Ribosomal, 16S ; genetics ; Sequence Analysis, DNA ; Trichloroethylene ; metabolism

OBJECTIVE:To study the preventive effect of emodin and astragalus polysaccharides on experimental hepato-carcinoma in rats.METHODS:The experimental rats were randomly divided into the normal group,model group,emodin group,and emodin combined with astragalus polysaccharides group.hepatocarcinogenesis in rat models were induced by di-ethylnitrosamine,while at the same time all the groups were administered with pre-set drugs by stomach irrigation.Rat's we_ ight,serum indices of ALT,ALP,?-GT,?-L-fucoxidase were determined,and pathological examination was made before and after administration.RESULTS:The group administered with emodin and astragalus polysaccharides improved more than any other groups in terms of rat's weight,serum indices,onset time of hepatocarcinogenesis,and pathological grade.CONCLU_ SION:Emodin combined with astragalus polysaccharides have certain preventive effect on experimental hepatocarcinoma in rats.

Objective To explore the clinical characteristics of metabolic disorder and the incidence rate of metabolic syndrome (MS) in the patients with posthepatitic cirrhosis.Methods Four hundred and fifty-five patients with posthepatitic cirrhosis were included in the study and divided into hepatitis B related cirrhosis group (LCB group,351 cases) and hepatitis C related cirrhosis group (LCC group,104cases).The prevalence of metabolic disorder was recorded and the characteristics of posthepatitic cirrhosis with MS and without MS were compared and analysed.Results The prevalence of hyperglycemia,hypertension,hyperlipemia,obesity and MS in the posthepatitic cirrhosis patients was 46.59% (212/455),15.16% (69/455),15.38% (70/455),22.64% ( 103/455),12.53% (57/455) respectively.The prevalence of MS in LCB and LCC was 8.26% (29/351) and 26.92% (28/104).The levels of body mass index (BMI),fasting blood glucose,hypertension,cholesterol,aminotransferase (ALT) in LCB with MS patients were significantly higher than those in LCB without MS patients.There were no differences in the levels of HBeAg and HBV DNA between LCB with MS patients and LCB without MS patients.The levels of BMI,hypertension,triglyceride in LCC with MS patients were significantly higher than those in LCC without MS patients.There were no differences in fasting blood glucose,cholesterol and ALT between LCC with MS patients and LCC without MS patients.Logistic regression revealed that BMI was the independent factor in LCB and LCC with MS.Conclusions The prevalence of hyperglycemia and obesity are higher in LCB and LCC.The incidence rate of MS in LCB is less than that in the general population,while the incidence rate of MS in LCC is significantly higher than that in the general population,and it's nothing to do with the viral replication.BMI is an important factor affected in posthepatitic cirrhosis with MS.

Objective To verify the interaction between asialoglycoprotein receptor (ASGPR)and hepatitis B virus (HBV)preS1 protein in vivo and in vitro ,and identify ASGPR as a cell-surface receptor for HBV,which could elucidate the molecular mechanism of HBV infection.Methods The preS1-ASGPR interaction was examined in mammalian two-hybrid and coimmunoprecipitation system by strictly following the manufacturer’s instructions.Results ASGPR interacted specifically and directly with the preS1 domain of HBV in vivo and in vitro .Conclusion ASGPR may be a candidate receptor for HBV that mediates further step of HBV entry.

A HPLC method was established for determining the content of bai calin in Qinxiazhixiao Instant Granules. The specification of the YWG-C18 chrom atograp hic column was 4.6 mm×250 mm, 10 μm; the mobile phase was MeOH∶H2O∶THF∶H 3P O4 (200∶300∶50∶0.16); the retention time of baicalin was about 9 minutes; t he average recovery rate of baicalin was 100.65%; and the RSD 2.03%.

Objective:To investigate high mobility group protein 1(HMGB1), tumor necrosis factor-α(TNF-α)and interleukin-6(IL-6)levels and their clinical significance in elderly patients with viral pneumonia.Methods:One hundred and sixty elderly patients with viral pneumonia admitted to the Sixth Hospital Affiliated to Anhui Medical University were enrolled as research subjects.In addition, 40 elderly people who underwent regular physical examination were considered as the control group.Patients with viral pneumonia were divided into the low-risk group, middle-risk group and high-risk group according to CURB-65 scores and pneumonia severity index(PSI)scores.HMGB1, TNF-α and IL-6 levels were compared between different groups.The correlations of CURB-65 scores and PSI scores with HMGB1, TNF-α, IL-6 levels were analyzed.Multivariate Logistic regression analysis was used to examine influencing factors for the severity of viral pneumonia in elderly patients.Results:HMGB1, TNF-α and IL-6 levels were higher in research subjects than in the control group.As the severity of viral pneumonia increased, so did HMGB1, TNF-α and IL-6 levels(all P<0.05). HMGB1, TNF-α and IL-6 levels in the severe viral pneumonia group were significantly higher than those in the non-severe viral pneumonia group( P<0.05). HMGB1, TNF-α and IL-6 levels were positively correlated with CURB-65 scores and PSI scores(CURB-65 score: r=0.463, 0.392 and 0.497, P=0.015, 0.003 and 0.025; PSI score: r=0.596, 0.515 and 0.381, P=0.007, 0.011 and 0.009). HMGB1, TNF-α and IL-6 levels were influencing factors for the severity of viral pneumonia in elderly patients( OR=1.344, 1.422 and 1.351, P=0.006, 0.015 and 0.009). Conclusions:HMGB1, TNF-α and IL-6 levels are closely correlated with the severity of viral pneumonia and are helpful for early assessment of viral pneumonia.

This research amplified the phyA gene with the designed and synthesized primers specific for the phyA gene full-length coding sequence. The phyA gene was from Aspergillus niger F246 by the polymerase chain reaction(PCR), which is selected and identified in our laboratory. After sequncing the coding sequence, it was confirmed that the construction of cloning vector was succeeded. The phyA gene fragment was recovered from the pMD18T-phyA and ligated with prokaryotic expression vector pET30a+ to construct the recombinant expression plasmid pET30a+ -phyA. It was expressed with IPTG induction in E. coli for high efficiency. A new protein band with apparent molecular weight 50 kDa was detected in the lysate of the transformed cell by using SDS-PAGE. The amount of the soluble fusion protein was about 40% of large intestine bacillus soluble protein of transformed cells, estimated by absorbance scanning of SDS-PAGE and protein quantitation. It's phytase activity was eight times over the natural phyase. So this research provides the basis of the study on obtaining large and high active phytase and developmant of the new microbial ecologicalagent.
6-Phytase ; biosynthesis ; genetics ; Aspergillus niger ; enzymology ; genetics ; Base Sequence ; Cloning, Molecular ; Escherichia coli ; genetics ; metabolism ; Genes, Fungal ; Genetic Vectors ; Molecular Sequence Data ; Recombinant Fusion Proteins ; biosynthesis ; genetics

Objective:To investigate the effects of zinc oxide nanoparticles (ZnO-NPs) on neutrophil hypoxia and pyroptosis through nucleotide binding of oligomeric domain-like receptor protein 3 (NLRP3) inflammasome, and to analyze the role of pyroptosis on respiratory tract inflammotion induced by ZnO-NPs.Methods:In October 2022, primary cultured neutrophils were obtained from the abdominal aortic blood of SPF adult healthy SD rats. The neutrophils were treated with ZnO-NPs solution (0, 5, 10, 20 μg/ml) at different concentrations, and hypoxia group (5% O 2) was set up. Hypoxia and reactive oxygen species (ROS) levels were detected by flow cytometry, and the expression levels of NLRP3, apoptosis-associated speck-like protein containing a CARD (ASC), cleaved Caspase-1 were measured by Western blot. The activity of lactic dehydrogenase (LDH) in the cell supernatant was measured by coloration, and the content of interleukin-1 beta (IL-1β) in cell culture supernatant was detected by enzyme-linked immunosorbent assay (ELISA) . Results:Compared with the control group, hypoxia and ROS levels of neutrophils in hypoxia group and ZnO-NPs groups were significantly increased ( P<0.05), and NLRP3, ASC, cleaved Caspase-1 protein expression levels, LDH activity and IL-1β content were significantly increased ( P<0.05). Compared with hypoxia group, hypoxia and ROS levels of neutrophils in 5 μg/ml and 10 μg/ml ZnO-NPs groups were significantly decreased ( P<0.05), NLRP3, ASC, cleaved Caspase-1 protein expression levels, LDH activity, and IL-1β content were decreased significantly ( P<0.05). There were no significant differences in hypoxia, ROS levels, ASC, cleaved Caspase-1 protein expression levels, LDH activity, and IL-1β content between the 20 μg/ml ZnO-NPs group and the hypoxia group ( P>0.05) . Conclusion:ZnO-NPs treatment may activate the NLRP3 inflammasome to induce pyroptosis of neutrophils which may be related to ROS and hypoxia.

Objective To investigate the effects of glucocorticoid combined with spleen aminopeptide on immune function and pulmonary function in children with combined allergic rhinitis and asthma syndrome (CARAS).Methods A total of 166 cases of CARAS were divided into observation group (84 cases) and control group (82 cases),the patients in the control group were treated by nasal inhalation of Budesonide Aerosol,in addition to the treatment of control group the observation group was given spleen aminopeptide oral lyophilized powder for treatment,and two groups were treated continuously for 3 months.The effect of 2 groups of children,and the changes of immune and lung function before and after treatment were compared.Results Rhinitis and asthma were significantly reduced in two groups of children after treatment,and the reductions of the score of rhinitis symptoms and the asthma score in the observation group were more significant than those in the control group(P＜0.05).The humoral immunity index (IgG,IgM and IgA) of the 2 groups increased significantly after treatment.CD3+,CD4+ and the ratio of CD4+/CD8+ in the cellular immune indexes increased significantly,and CD8+ decreased significantly.The immune indexes in the observation group were significantly improved compared with those in the control group (P＜0.05).After treatment,the forced expiratory volume in one second(FEV1) and maximal expiratory flow rate (PEFR) of the 2 groups increased significantly compared with those before treatment,and the degree of increase in the observation group was significantly higher than that in the control group(P＜0.01).Conclusion Glucocorticoid combined with spleen aminopeptide could not only improve the symptoms and signs of children with CARAS,but also significantly enhance cellular immunity,humoral immunity and pulmonary function.