Diagnostic efficacy of joint detection of Xpert MTBRIF, TB-RNA and TB-DNA in bronchoalveolar lavage fluid for smear-negative pulmonary tuberculosis / 中国热带医学

@#Abstract: Objective　To investigate the diagnostic value of joint detection of Mycobacterium tuberculosis rifampicin resistance gene (Xpert MTB/RIF), Mycobacterium tuberculosis ribonucleic acid (TB-RNA) and Mycobacterium tuberculosis deoxyribonucleic acid (TB-DNA) in bronchoalveolar lavage fluid for smear-negative pulmonary tuberculosis. Methods A total of 806 patients with suspected smear-negative pulmonary tuberculosis admitted to our hospital from May 2020 to July 2022 were selected, 506 patients diagnosed as bacterial negative pulmonary tuberculosis by clinical, X-ray and sputum samples were classified as bacterial negative pulmonary tuberculosis group, and the other 300 patients with non-tuberculous pulmonary disease were classified as non-tuberculous pulmonary disease group. XpertMTB/RIF, TB-RNA and TB-DNA in bronchoalveolar lavage fluid of all patients were detected. With clinical, X-ray and sputum specimen examination of mycobacterium tuberculosis as the gold standard, the diagnostic efficacy of alveolar lavage solution Xpert MTB/RIF, TB-RNA and TB-DNA alone and in combination was analyzed. Results The positive detection rates of Xpert MTB/RIF, TB-RNA and TB-DNA in bronchoalveolar lavage fluid of the smear-negative pulmonary tuberculosis group and the non-tuberculosis pulmonary disease group were 69.96% (354/506) and 2.67% (8/300), 61.46% (311/506) and 5.00% (15/300), and 63.64% (322/506) and 8.00% (24/300), respectively. The rates in the smear-negative pulmonary tuberculosis group were higher than those in the non-tuberculosis lung disease group, and the differences were statistically significant (χ2=342.005, 246.930, 235.687, P<0.01). Compared with the gold standard, the sensitivity, specificity, accuracy, positive predictive value and negative predictive value of Xpert MTB/RIF in the diagnosis of smear-negative pulmonary tuberculosis were 69.96%, 97.33%, 80.15%, 97.79% and 65.77%, respectively; those values of TB-RNA were 61.46%, 95.00%, 73.95%, 95.40% and 59.38%, respectively; those values of TB-DNA were 63.64%, 92.00%, 74.19%, 93.06% and 60.00%, respectively; those values of combined diagnosis with Xpert MTB/RIF, TB-RNA and TB-DNA were 61.26%, 100.00%, 75.68%, 100.00% and 60.48%, respectively; the specificity and positive predictive value of combined detection were higher than those of single detection (P<0.05). Conclusions The joint detection of Xpert MTB/RIF, TB-RNA and TB-DNA in bronchoalveolar lavage fluid can improve the diagnostic efficacy of smear-negative pulmonary tuberculosis and is worthy of clinical promotion and application.

Research Progress of DNA-Based Technologies for Postmortem Interval Estimation / 法医学杂志

In criminal investigations, postmortem interval (PMI) is important information to be inferred in homicide investigations, as well as the focus and the difficulty in forensic pathology research. Because the DNA content in different tissues is relatively constant and shows changes regularly with the extension of PMI, it has become a research hotspot of PMI estimation. This paper reviews the recent progress of PMI estimation technologies including DNA-based single cell gel electrophoresis, image analysis, flow cytometry, real-time fluorescence quantitative PCR and high-throughput sequencing, hoping to provide references for forensic medicine practice and scientific research.

Practical tips to using formalin-fixed paraffin-embedded tissue archives for molecular diagnostics in a South African setting

Background: Formalin-fixed paraffin-embedded (FFPE) tissue archives in hospitals, biobanks, and others offer a vast collection of extensive, readily available specimens for molecular testing. Unfortunately, the use of tissue samples for molecular diagnostic applications is challenging; thus, the forensic pathology FFPE tissue archives in Africa have been a largely unexploited genetic resource, with the usability of DNA obtainable from these samples being unknown.Intervention: The study, conducted from January 2015 to August 2016, determined the usefulness of FFPE tissue as a reliable source of genetic material for successful post-mortem molecular applications and diagnostics. Formalin-fixed paraffin-embedded tissue samples were collected and archived from autopsies conducted over 13 years in the forensic medicine department of the University of Pretoria (Pretoria, South Africa). Deoxyribonucleic acid from FFPE tissue samples and control blood samples was amplified by high-resolution melt real-time polymerase chain reaction before sequencing. The procurement parameters and fixation times were compared with the quantity and quality of the extracted DNA and the efficiency of its subsequent molecular applications.Lessons learnt: This study has shown that FFPE samples are still usable in molecular forensics, despite inadequate sample preparation, and offer immense value to forensic molecular diagnostics.Recommendations: FFPE samples fixed in formalin for more than 24 h should still be used in molecular diagnostics or research, as long as the primer design targets amplicons not exceeding 300 base pairs.

Las formulaciones anti-COVID-19 con ácido ribonucleico mensajero en la clasificación general de las vacunas antivirales

RESUMEN Las vacunas representan un hito fundamental para la prevención y el control de las enfermedades infectocontagiosas, con repercusión excepcional en la salud mundial. Su valor es incuestionable para evitar la aparición de numerosos padecimientos y muertes cada año. Existen numerosas clasificaciones de las vacunas, según se atienda a diferentes aspectos de su composición, síntesis o naturaleza. En este artículo se presenta una clasificación de los diseños actuales en que se sustentan las diversas plataformas tecnológicas de las vacunas antivirales. Se hace especial énfasis en las basadas en genes, entre ellas las vacunas de ácido ribonucleico mensajero que han recibido un impulso especial en su desarrollo desde el comienzo de la pandemia de la COVID-19. Las implicaciones de la respuesta satisfactoria de las vacunas de ácido ribonucleico mensajero podrían ir más allá de la actual pandemia de la COVID-19. Su éxito podría allanar el camino para el uso generalizado de esta plataforma tecnológica tanto para los patógenos emergentes como para los ya establecidos.

ABSTRACT Although the use of vaccines for disease prevention and control is a relatively recent social and health event, it has no doubt become one of the main tools of modern medicine to fight infectious diseases. The paper presents a classification of current designs substantiating the various technological platforms of antiviral vaccines, with special emphasis on those based on genes, among them messenger ribonucleic acid vaccines, which have experienced considerable development since the start of the COVID-19 pandemic. The implications of the successful response of messenger ribonucleic acid vaccines could go beyond the current COVID-19 pandemic. Its success could pave the way for the widespread use of this technology platform for both emerging and established pathogens.

Relationship of mitochondrial DNA copy number with clinicopathological characteristics and its influence on the prognosis of hepatocellular carcinoma patients / 中华消化外科杂志

Objective:To investigate the relationship of mitochondrial DNA (mtDNA) copy number with clinicopathologic characteristics and its influence on the prognosis of hepatocellular carcinoma (HCC) patients.Methods:The retrospective case-control study was conducted. The clinicopathological data of 71 HCC patients undergoing surgical treatment in the Eastern Hepatobiliary Surgery Hospital of Naval Medical University from March to June 2011 were collected. There were 61 males and 10 females, aged from 26 to 80 years, with a median age of 55 years. The mtDNA copy number of tumor tissues and adjacent normal tissues were measured for all patients. Observation indicators: (1) the mtDNA copy number of tumor tissues and adjacent normal tissues and relationship between the mtDNA copy number and clinicopathological characteristics of HCC patients; (2) follow-up; (3) related factors for the prognosis of HCC patients. Follow-up using outpatient examination or telephone interview was conducted to detect postoperative survival of patients up to September 2019. Measurement data with normal distribution were described as Mean± SD, and comparison between groups was analyzed using independent samples t test or the matched samples t test. Measurement data with skewed distribution were described as M(range). Count data were represented as absolute numbers, and comparison between groups was analyzed using the chi-square test or Fisher exact probability. Univariate and multivariate analyses were conducted using the COX regressional model. Variables with P<0.10 in the univariate analysis were included for the multivariate analysis. Survival rates were calculated using the Kaplan-Meier method, and Log-rank test was used for survival analysis. Results:(1) The mtDNA copy number of tumor tissues and adjacent normal tissues and relationship between the mtDNA copy number and clinicopathological characteristics of HCC patients: of 71 HCC patients, the mtDNA copy number was 0.85±0.08 in tumor tissues, versus 1.16±0.08 in adjacent normal tissues, showing a significant difference between them ( t=2.96, P<0.05). Of 71 HCC patients, 48 cases were mtDNA-low and 23 cases were mtDNA-high. Cases with tumor capsule as integrity or not-integrity, cases with or without microvascular (MVI) in mtDNA-low and mtDNA-high patients were 20, 28, 21, 27 and 16, 7, 4, 19, respectively, showing significant differences ( χ2=4.84, 4.74, P<0.05). (2) Follow-up: 71 patients were followed up for 2.1 to 85.3 months, with a median follow-up time of 47.8 months. The 1-, 3-, 5-year overall survival rates of 71 HCC patients were 87.3%, 64.7, 37.4%, respectively. Moreover, the 1-, 3-, 5-year overall survival rates were 81.2%, 50.0%, 29.2% of the mtDNA-low patients, versus 95.7%, 86.5%, 54.7% of the mtDNA-high patients, showing a significant difference between the two groups ( χ2=5.86, P<0.05). (3) Related factors for the prognosis of HCC patients. Results of univariate analysis showed that the number of tumor, portal vein tumor thrombus, MVI, Barcelona Clinic Liver Cancer stage, mtDNA copy number were related factors for the prognosis of HCC patients ( hazard ratios=2.211, 2.911, 3.899, 3.587, 0.440, 95% confidence intervals as 1.024?4.777, 1.485?5.704, 2.115?7.186, 1.615?7.966, 0.223?0.871, P<0.05). Results of multivariate analysis showed that MVI and mtDNA copy number were independent influencing factors for the prognosis of HCC patients ( hazard ratios=2.754, 0.437, 95% confidence intervals as 1.374?5.521, 0.205?0.932, P<0.05). Conclusions:The mtDNA copy number of HCC patients is related with tumor capsule and MVI. The mtDNA copy number and MVI are independent influencing factors for the prognosis of HCC patients.

Prevalence and distribution of various human papillomavirus genotypes in women with high risk for cervical carcinoma

Background: In India, cervical cancer is second most common cancer in women and is the second most common cause of death due to cancers in women. Human papillomavirus (HPV), mainly genotype 16 and 18, are responsible for most of the precancerous lesions of cervix and for cervical cancer. Therefore, it is necessary to prevent the spread of HPV infection and its early treatment to decrease cervical cancer associated morbidity and mortality. Aims of this study was to find out the prevalence and distribution of various HPV genotypes in women high risk for cervical carcinoma. To find correlation between HPV DNA test and cytology report.Methods: An observational study was done on a total of 384 women who were at a high risk for cervical cancer.  Ectocervix and endocervix samples for conventional cytology along with cervical swab collection for HPV DNA isolation were obtained from women aged 21-65 years. Multiplex real time PCR used for HPV DNA isolation and genotyping and Bethseda classification 2014 was used for reporting cytology. Statistical analysis was done using SPSS version 22 and Microsoft excel 2010 software. Data assessment was done using independent t-test, Chi-square test or Fisher’s exact test.Results: Prevalence of HPV in women high risk for cervical cancer in our study was 14.58%. Most common high-risk genotypes were HPV 18 (7.03%) followed by HPV 16 (6.51%). All genotypes except genotype 31, were more frequently seen as a single infection rather than a multiple infection. In 323 subjects which were negative for Pap smear, 31 were positive for HPV DNA showing a statistically significant result with a p value of <0.0001.Conclusions: It was analysed that infection with HPV is very common (9.60%) even in women who had a negative Pap smear test showing a positive correlation between the two tests. HPV DNA testing can improve the detection rate of cervical intraepithelial lesions.

COⅠ and 16S rDNA Sequence Identification of Common Necrophagous Flies in Fujian Province / 法医学杂志

Objective To identify the species of common necrophagous flies in Fujian Province by gene fragment sequences of mitochondrial cytochrome c oxidase subunit Ⅰ （COⅠ） and 16S ribosomal deoxyribonucleic acid （16S rDNA）, and to explore the identification efficacy of these two molecular markers. Methods In total 22 common necrophagous flies were collected from the death scenes in 9 different regions in Fujian Province and DNA was extracted from the flies after morphological identification. The gene fragments of COⅠ and 16S rDNA were amplified and sequenced. All the sequences were uploaded to GeneBank and BLAST and MEGA 10.0 software were used to perform sequence alignment, homology analysis and intraspecific and interspecific genetic distance analysis. The phylogenetic trees of DNA fragment sequences of COⅠ and 16S rDNA of common necrophagous flies in Fujian Province were established by unweighted pair-group method with arithmetic means （UPGMA）, respectively. Results The flies were classified into 6 species, 5 genera and 3 families by morphological identification. The results of gene sequence analysis showed that the average number of interspecific and intraspecific genetic distance of 16S rDNA ranged from 1.8% to 8.9% and 0.0% to 2.4%, respectively. The average number of interspecific and intraspecific genetic distance of COⅠ ranged from 7.2% to 13.6% and 0.0% to 6.3%, respectively. Conclusion The gene sequences of COⅠ and 16S rDNA can accurately identify the species of different necrophagous flies, and 16S rDNA showed higher value in species identification of common calliphoridae necrophagous flies in Fujian Province.

Electrochemical, spectroscopic, and molecular docking studies of the interaction between the anti-retroviral drug indinavir and dsDNA / 药物分析学报

In this study, an electrochemical DNA biosensor was developed using a straightforward methodology to investigate the interaction of indinavir with calf thymus double-stranded deoxyribonucleic acid (ct-dsDNA) for the first time. The decrease in the oxidation signals of deoxyguanosine (dGuo) and deoxy-adenosine (dAdo), measured by differential pulse voltammetry, upon incubation with different con-centrations of indinavir can be attributed to the binding mode of indinavir to ct-dsDNA. The currents of the dGuo and dAdo peaks decreased linearly with the concentration of indinavir in the range of 1.0-10.0μg/mL. The limit of detection and limit of quantification for indinavir were 0.29 and 0.98μg/mL, respectively, based on the dGuo signal, and 0.23 and 0.78μg/mL, respectively, based on the dAdo signal. To gain further insights into the interaction mechanism between indinavir and ct-dsDNA, spectroscopic measurements and molecular docking simulations were performed. The binding constant (Kb) between indinavir and ct-dsDNA was calculated to be 1.64 × 108 M-1, based on spectrofluorometric measure-ments. The obtained results can offer insights into the inhibitory activity of indinavir, which could help to broaden its applications. That is, indinavir can be used to inhibit other mechanisms and/or hallmarks of viral diseases.

Epigenetic therapies in patients with solid tumors: Focus on monotherapy with deoxyribonucleic acid methyltransferase inhibitors and histone deacetylase inhibitors

Epigenomics is the study of the gene expression changes due to epigenetic processes and not due to the deoxyribonucleic acid (DNA) base sequence alterations. The key mechanisms of epigenetic regulation include DNA methylation, histone modifications, and noncoding RNAs. Epigenetic alterations in cancer are predominantly linked with hypermethylation of promoters of the tumor suppressor genes, global DNA hypomethylation, and increased expression of histone deacetylases (HDAC). There is a growing need to investigate epigenetic patterns and to provide safe and effective, innovative therapeutic strategies for oncology patients, who did not improve on traditional anticancer regimens. The epi-drugs (e.g., DNA methyltransferase inhibitors, e.g., azacitidine and decitabine and HDAC inhibitors, e.g., vorinostat and romidepsin) have been approved for the clinical use. In this paper, we provide a brief overview of the mechanisms of action and targets for novel epi-drugs, focusing on their potential clinical applications in patients with solid tumors, resistant to standard oncology treatments

Sample-A Raw Data For Future Information Of Life

Gene amplification requires a biological sample which is collected prior to the demands. The sample for such analysis plays a vital role as they serve as a resource for the core material-Deoxyribonucleic acid. A sample for gene amplification or any analysis would be collected prior to the demands and type of analysis. Human blood has been an essential resource of DNA from the commencement of DNA extraction in the 19th century. After then several protocols developed according to the requirement of both samples of different forms and their retrieval as various forms and methods. The sample has various characteristics and prerequisites when collected for gene analysis. Some important characteristics of sample collection methods are significant, which are not widely taken. This work analyses various general sample collection protocols and highlights some of the major characteristics and prerequisites for the sample. This work initiates and delivers to attain the core of genomics by bringing out the basic raw materials importance and consequences on amplification which is highly sensitive.

Enhancement of pigment production potential of Serratia marcescens (GBB151) through mutation and random amplified polymorphic deoxyribonucleic acid analysis of its mutants

Serratia marcescens (GBB151) was isolated and genetically modified for high-yielding pigment production capacitythat could be employed for industrial purposes. Ethidium bromide-induced mutagenesis of GBB151 resulted inthe generation of eight mutant isolates (GBB151Ea-GBB151Eh). The chemical mutants of S. marcescens obtainedproduced 5-fold more pigment than the wild-type organism. The wild-type GBB151 produced 413.9 unit/cell,while the mutant strains produced pigments with yields ranging from 841.7 to 2008.5 unit/cell. Random amplifiedpolymorphic deoxyribonucleic acid-polymerase chain reaction analysis showed different amplicons patterns of nativeas well as mutant derivatives. The factorial analysis diagram and the dendrogram showed a degree of dissimilarityamong the wild-type bacterial isolate GBB151 and its mutants. Mutant strains GBB151Ec and GBB151Ef wereclosest to the wild type as they appeared in the same quadrant. GBB151Ed which had lost its ability to producepigment was farthest and in the different quadrant to the wild type. These study provided insight into improvement inpigment production by manipulating genetic make-up of S. marcescens, thus meeting industrial demand.

El ácido desoxirribonucleico mitocondrial (ADNm) y "Las siete hijas de Eva" / Mitochondrial deoxyribonucleic acid (mRNA) and "The seven daughters of Eve"

En el año 2001, el profesor Bryan Sykes (Oxford), publicó un libro intitulado "Las Siete Hijas de Eva", tras analizar el ADN mitocondrial (ADNm) de numerosas europeas. El ADNm pasa exclusivamente a las mitocondrias de mujeres y a su vez solo las hijas lo transmiten a la suya. El estudio fue posible gracias a la Reacción en Cadena de la Polimerasa. De ese modo Sykes precisó que en Europa hay siete clanes femeninos principales, cuyas fundadoras vivieron desde hace unos 45 000 hasta hace unos 8 500 años. Palabras clave: Ácido Desoxirribonucleico Mitocondrial (ADNm). Clanes ancestros maternos y paternos. Migraciones y Genética.(AU)

In 2001, professor Bryan Sykes (Oxford), published a book in Spanish untitled "Las siete hijas de Eva", analyzing the mitochondrial DNA from numerous European women. The mDNA has the peculiarity to pass exclusively to the mitochondrias of women, and only their daughters are able to transmit it. The study was possible thanks to the availability of the Polymerase Chain Reaction. By means of the study of the mtDNA he was able to detect in Europe seven principle maternal clans from 45 000 to 8 500 years.(AU)

Relationship between Maternal PBMC HBV cccDNA and HBV Serological Markers and its Effect on HBV Intrauterine Transmission / 生物医学与环境科学（英文）

OBJECTIVE@#To investigate the relationship between maternal peripheral blood mononuclear cells (PBMC) hepatitis B virus (HBV) covalenty closed circular deoxyribonucleic acid (cccDNA) and other HBV serological markers and its effects on HBV intrauterine transmission.@*METHODS@#We enrolled 290 newborns and their hepatitis B surface antigen (HBsAg) positive mothers. HBV cccDNA in PBMC and HBV DNA in serum were detected by a real-time PCR-TaqMan probe while HBV serological markers were detected with an electrochemiluminescence immunoassay.@*RESULTS@#There was a positive correlation between the levels of PBMC HBV cccDNA and serum HBV DNA and HBeAg (r = 0.436 and 0.403, P < 0.001). The detection rate of pattern A ['HBsAg (+), HBeAg (+), and anti-HBc (+)'] was significantly higher in the PBMC HBV cccDNA positive group than in the control group (χ2 = 48.48, P < 0.001). There was a significant association between HBV intrauterine transmission and PBMC HBV cccDNA (χ2 = 9.28, P = 0.002). In the presence of serum HBV DNA, HBeAg, and PBMC HBV cccDNA, the risk of HBV intrauterine transmission was three times higher (OR = 3.69, 95% CI: 1.30-10.42) than that observed in their absence. The risk of HBV intrauterine transmission was the greatest (OR = 5.89, 95% CI: 2.35-14.72) when both PBMC HBV cccDNA and pattern A were present. A Bayesian network model showed that maternal PBMC HBV cccDNA was directly related to HBV intrauterine transmission.@*CONCLUSION@#PBMC HBV cccDNA may be a direct risk factor for HBV intrauterine transmission. Our study suggests that serological markers could be combined with PBMC-related markers in prenatal testing.

Expression of PD-1hiCXCR5-CD4+T cells in patients with systemic lupus erythematosus / 中华风湿病学杂志

Objective To investigate the expression of peripheral programmed death (PD)-1hiCXCR5-CD4+T cells and its clinical significance in systemic lupus erythematosus (SLE). Methods Peripheral blood PD-1hiCXCR5-CD4+ T cells from 21 SLE patients and 16 healthy controls were examined by flow cytometry. The levels of serum anti-double-stranded deoxyribonucleic acid (dsDNA) antibodies were determined using immunoradiometric as-say. Data were analyzed with t test and Pearson's correlation test. Results The per-centages of PD-1hiCXCR5- cells within CD4+ T cell were significantly higher in SLE patients [(2.1 ±2.0)％] compared to normal controls [(0.3±0.3)％] (t=2.959, P<0.01). The percentages of PD-1hiCXCR5-cells within CD4+T cells in moderate to severe active SLE patients (3.0 ±2.0)％ was significantly increased compared to patients with mild or inactive (1.0±1.4)％(t=2.574, P<0.05) and normal controls (0.3±0.3)％ (t=5.149, P<0.01). The percentages of PD-1hiCXCR5- cells within CD4+ T cells from SLE patients were positively related with systemic lupus erythematosus disease activity index (SLEDAI) (r=0.475, P=0.0297). SLE patients in serum anti-dsDNA antibodies positive group (2.7±2.1)％displayed a higher percentage of PD-1hiCXCR5-cells within CD4+T cells than patients in serum anti-dsDNA antibodies negative group (0.6 ±0.5)％ (t=2.303, P<0.05). The percentages of PD-1hiCXCR5-cells within CD4+T cells from SLE patients were positively correlated with anti-dsDNA antibody titers. Conclusion The percentages of PD-1hiCXCR5- cells within CD4+ T cells from SLE patients are increased and are positively correlated with SLEDAI and anti-dsDNA antibody levels. Increased percentage of PD-1hiCXCR5-cells within CD4+T cells might play an important role in the pathogenesis of SLE.

Correlation of HBV genotyping and drug resistance mutation in Hohhot,Inner Mongolia / 中国基层医药

Objective To investigate the relationship between HBV -DNA load and serum markers in chronic hepatitis B( CHB) patients in Hohhot,Inner Mongolia,and to explore the mutation of HBV genotype and nucleoside analogue.Methods From January 2015 to December 2017,one hundred and ninety-three CHB patients hospitalized in the People＇s Hospital of Inner Mongolia were selected randomly.The clinical diagnostic criteria for all admitted patients were based on the " Guidelines for the Prevention and Treatment of Chronic Hepatitis B" jointly formulated by the Infectious Diseases Society of 2010. The HBV -DNA load of HBV was detected by real -time quantitative PCR,and the correlation between HBV -DNA load and serum markers was analyzed. Seventy -nine patients were selected from 193 hospitalized patients,PCR-reverse dot blot hybridization was used to analyze HBV genotyping and the drug resistance mutations of different genotypes.Results The differences of HBeAb level and HBV-DNA load between HBeAg positive patients and negative patients were statistically significant(all P<0.001). Of 79 serum specimens of HBV infected people,9 cases(11.4% ) were B genotypes,and 70 cases of C genotype (88.6% ).Of them,25 cases had different loci variation,the rate of variation was 31.6% (25/79),with the unit point rtS213T mutation dominated,accounting for about 24.0% (6/25).Conclusion In Hohhot Inner Mongolia patients with CHB,HBV-DNA load with HBeAg and HBe Ab level are correlated;genotype in patients including B type and C type,which is mainly genotype C;patients with CHB mainly had drug resistance to lamivudine and adefovir dipivoxil, mutations including rtS213T,and hybrid mutation.

Specification of Bacteriophage Isolated Against Clinical Methicillin-Resistant Staphylococcus Aureus

OBJECTIVES: The emergence of resistant bacteria is being increasingly reported around the world, potentially threatening millions of lives. Amongst resistant bacteria, methicillin-resistant Staphylococcus aureus (MRSA) is the most challenging to treat. This is due to emergent MRSA strains and less effective traditional antibiotic therapies to Staphylococcal infections. The use of bacteriophages (phages) against MRSA is a new, potential alternate therapy. In this study, morphology, genetic and protein structure of lytic phages against MRSA have been analysed. METHODS: Isolation of livestock and sewage bacteriophages were performed using 0.4 μm membrane filters. Plaque assays were used to determine phage quantification by double layer agar method. Pure plaques were then amplified for further characterization. Sulfate-polyacrylamide gel electrophoresis and random amplification of polymorphic DNA were run for protein evaluation, and genotyping respectively. Transmission electron microscope was also used to detect the structure and taxonomic classification of phage visually. RESULTS: Head and tail morphology of bacteriophages against MRSA were identified by transmission electron microscopy and assigned to the Siphoviridae family and the Caudovirales order. CONCLUSION: Bacteriophages are the most abundant microorganism on Earth and coexist with the bacterial population. They can destroy bacterial cells successfully and effectively. They cannot enter mammalian cells which saves the eukaryotic cells from lytic phage activity. In conclusion, phage therapy may have many potential applications in microbiology and human medicine with no side effect on eukaryotic cells.

Research of live cell membranes modification based on cholesterol-DNA / 上海交通大学学报（医学版）

Objective: To investigate the modification of live cell membranes with cholesterol linked deoxyribonucleic acid (cholesterol-DNA). Methods: The suspension L1210 cells and adherent PC-12 cells were included. L1210 cells and PC-12 cells were divided into the experimental group (incubated with cholesterol-DNA) and the control group (treated with phosphate buffer saline), respectively. The fluorescence intensity of the cells in the two groups was obtained and the experimental group cells were three-dimensional reconstructed by confocal microscopy. The morphology of the cells in the experimental group was observed by scanning electron microscopy (SEM). The effect of cholesterol-DNA modification on cell membrane fluidity was detected by fluorescence recovery after photobleaching. Results: The results of suspension cells and adherent cells were consistent. Compared with the control group, the fluorescence intensity of cell surface in the experimental group was increased (both P=0.000). Three-dimensional reconstruction of confocal microscopy showed that the fluorescence of the cells in the experimental group was distributed across the surface of the global cell. SEM showed that the morphology of the cells in the experimental group did not change with cholesterol-DNA modification. After fluorescence photobleaching, the relative fluorescence intensity of the L1210 cells in the experimental group was decreased to 0.090, and then recovered to 0.860 within 110 s. Conclusion: Cholesterol-DNA can modify the whole live cell membranes, and the modified cell membranes still have fluidity. This method can modify not only the suspension cells, but also the adherent cells.

Western diet induces endogen oxidative deoxyribonucleic acid damage and infl ammation in Wistar rats / Dieta ocidental induz endógeno dano oxidativo do ácido desoxirribonucleico e infl amação em ratos Wistar

ABSTRACT Objective Nutritional diseases such as metabolic syndrome, cardiovascular disorder, chronic inflammation or even cancer are observed in people who sustain their lifestyle by Western diet due to high calorie intake. The origin of these diseases are the degraded deoxyribonucleic acid structure. In this study, we investigated whether Western diet produced endogenous oxidative deoxyribonucleic acid damage, apoptosis or inflammation. Methods Twenty-eight male Wistar rats, aged 10-12 weeks, were divided into four groups. The rats in control group received the standard diet and the remaining rats were given one of the following three diets for four weeks: a high-fat diet containing 35% fat, a high-sucrose diet containing 69% sucrose and Western diet comprising both two types of diets. After treatment the serum 8-hydroxy-2-deoxyguanosine, poly (adenosine diphosphate ribose) polymerase-1, chitinase-3-like protein 1, soluble urokinase-type plasminogen activator receptor, Fas ligand and cytochrome c levels were measured. Results It was observed no changes in the serum soluble urokinase-type plasminogen activator receptor, Fas ligand and cytochrome c levels whereas a statistically significant increase in the serum 8-hydroxy-2-deoxyguanosine, poly (adenosine diphosphate ribose) polymerase-1 and chitinase-3-like protein 1 levels were found only in rats that were given Western diet. Conclusion The findings show that Western diet produced endogenous oxidative deoxyribonucleic acid damage, which then increased serum poly (adenosine diphosphate ribose) polymerase-1 levels, eventually leading to inflammation.

RESUMO Objetivo Doenças nutricionais, como síndrome metabólica, distúrbios cardiovasculares, inflamação crônica ou mesmo câncer, são observadas em pessoas que sustentam seu estilo de vida na dieta ocidental, caracterizada pela alta ingestão de calorias. Dado que a origem dessas doenças é a estrutura degradada do ácido desoxirribonucleico, o presente estudo investigou se a dieta ocidental produzia dano oxidativo endógeno ao ácido desoxirribonucleico, apoptose ou inflamação. Métodos Foram utilizados 28 ratos Wistar machos, com idade entre 10-12 semanas, divididos em quatro grupos. Os ratos do grupo controle receberam a dieta padrão, ao passo que os ratos restantes receberam uma das três dietas seguintes por quatro semanas: uma dieta rica em gordura contendo 35% de gordura; uma dieta rica em sacarose contendo 69% de sacarose; e dieta ocidental compreendendo os dois tipos de dietas. Após o tratamento soro 8-hidroxi-2-desoxiguanosina, poli (adenosina difosfato ribose) polimerase-1, quitinase-3-like proteína 1, uroquinase solúvel tipo de receptor ativador de plasminogênio, os níveis do ligante Fas e do citocromo c foram medidos. Resultados Não foram observadas alterações nos níveis séricos de uroquinase solúvel tipo de receptor ativador de plasminogênio, ligante Fas e citocromo c, enquanto um aumento estatisticamente significativo nos níveis séricos de 8-hidroxi-2-desoxiguanosina, poli (adenosina difosfato ribose) polimerase-1 e quitinase-3-like proteína 1 foi encontrado apenas em ratos que receberam dieta ocidental. Conclusão Os resultados mostram que a dieta ocidental produziu danos no ácido desoxirribonucleico oxidativo endógeno, o que aumentou os níveis séricos de poli (adenosina difosfato ribose) polimerase-1, levando à inflamação.

Association of Cytokines with Alanine Aminotransferase, Hepatitis B Virus Surface Antigen and Hepatitis B Envelope Antigen Levels in Chronic Hepatitis B / 中华医学杂志(英文版)

Background@#Cytokines play an important role in occurrence and recovery of hepatitis B virus (HBV) infection. The aim of this study was to investigate the changes of cytokines concentration and its correlation to alanine aminotransferase (ALT), HBV deoxyribonucleic acid (HBV-DNA), hepatitis B envelope antigen (HBeAg), and HBV surface antigen (HBsAg) in the development of chronic hepatitis B (CHB).@*Methods@#Thirteen healthy individuals (HI), 30 chronic HBV-infected patients in immune tolerant (IT) phase, and 55 CHB patients were enrolled between August 2015 and May 2017. The peripheral blood samples were collected from all individuals. The levels of interferon (IFN)-α2, interleukin (IL)-10, transforming growth factor (TGF)-β1, HBV-DNA, HBsAg, and HBeAg and liver function were measured. The quantitative determinations of cytokines levels, including IFN-α2, IL-10, and TGF-β1 were performed using Luminex multiplex technology. The correlation of cytokines to ALT, HBV-DNA, HBsAg, and HBeAg was analyzed by linear regression analysis.@*Results@#IFN-α2 levels were similar between HI and IT groups (15.35 [5.70, 67.65] pg/ml vs. 15.24 [4.07, 30.73] pg/ml, Z = -0.610, P = 0.542), while it elevated significantly in CHB group (35.29 [15.94, 70.15] pg/ml vs. 15.24 [4.07, 30.73] pg/ml; Z = -2.522, P = 0.012). Compared with HI group (3.73 [2.98, 11.92] pg/ml), IL-10 concentrations in IT group (5.02 [2.98, 10.11] pg/ml), and CHB group (7.48 [3.10, 18.00] pg/ml) slightly increased (χ = 2.015, P = 0.365), and there was no significant difference between IT and CHB group (Z = -1.419, P = 0.156). The TGF-β1 levels among HI (3.59 ± 0.20 pg/ml), IT (3.62 ± 0.55 pg/ml), and CHB groups (3.64 ± 0.30 pg/ml) were similar (χ = 2.739, P = 0.254). In all chronic HBV-infected patients (including patients in IT and CHB groups), the elevation of IFN-α2 level was significantly associated with ALT level (β= 0.389, t = 2.423, P = 0.018), and was also negatively correlated to HBV-DNA load (β = -0.358, t = -2.308, P = 0.024), HBsAg (β = -0.359, t = -2.288, P = 0.025), and HBeAg contents (β = -0.355, t = -2.258, P = 0.027). However, when both ALT level and cytokines were included as independent variable, HBV-DNA load, HBsAg, and HBeAg contents were only correlated to ALT level (β = -0.459, t = -4.225, P = 0.000; β = -0.616, t = -6.334, P = 0.000; and β = -0.290, t = -2.433, P = 0.018; respectively).@*Conclusions@#IFN-α2 elevation was associated with ALT level in patients with chronic HBV infection. However, in CHB patients, only ALT level was correlated to HBV-DNA, HBsAg and HBeAg contents.

Study on the effects of hemolysis, fatty blood and various storage conditions on HBV DNA,HIV RNA nucleic acid screening / 天津医药

Objective To study the influences of hemolysis, fatty blood, storage temperature and storage time on blood HIV RNA and HBV DNA. Methods The HBV DNA and HIV RNA samples (the concentration was 12-30 times of limit of detection of reagent), which were stored for 4 h, 1 d, 3 d,1 w and 4 w under the conditions of 4℃, 25℃, 37℃and-30℃, were detected using Roche MPX V2.0 kit. The HBV DNA and HIV RNA samples (the concentration was 2-5 times of limit of detection of reagent) were detected by the 6 groups of hemolytic samples (the concentrations of hemoglobin were 97 g/L, 34 g/L,17 g/L, 8 g/L, 5 g/L and 3 g/L, respectively). NAT test was performed at 5-group lipid samples (the concentrations of triglyceride were 7.93 mmol/L, 3.80 mmol/L, 2.63 mmol/L, 1.83 mmol/L and 1.49 mmol/L, respectively) and the control group samples (the concentrations of hemoglobin and triglyceride were 0 g/L and 0.95 mmol/L respectively). Results There were no significant differences in Ct values of HIV RNA or HBV DNA between 4 h to 4 w at 25℃(P>0.05). There were significant differences in Ct values of HBV DNA after preservation for 3 d and 1 w under 37 ℃ compared with those of preservation for 4 h,1 d and 2 d (P<0.005). When Hb concentration was reached to 97 g/L, the results of HBV DNA and HIV RNA were negative. When the concentration of Hb was less than 34 g/L, compared with the control group, there were no significant differences in Ct values of HIV RNA and HBV DNA (P>0.05). When the TG concentration was≤7.93 mmol/L, there were no significant differences in Ct values between the control group and the TG group (P>0.05). Conclusion The samples of HIV RNA and HBV DNA detected by Roche MPX V2.0 kit can be stored at room temperature (25℃) for four weeks. When the concentrations of TG and Hb are less than 7.93 mmol/L and 34 g/L respectively, there are no effects on HIV RNA or HBV DNA samples detected by Roche MPX V2.0 kit.