Establishment of real-time fluorescent quantitative PCR for detection of torque teno virus types 7, 8 and 10 / 中华微生物学和免疫学杂志

Objective:To establish a real-time fluorescent quantitative PCR for the detection of torque teno virus types 7 (TTV7), 8 (TTV8) and 10 (TTV10) and analyze its performance in clinical sample detection.Methods:Specific primers were designed based on the gene sequences of TTV7, TTV8 and TTV10 in GenBank. Recombinant plasmids of pMD19-T-TTV7, pMD19-T-TTV8 and pMD19-T-TTV10 were constructed and used as positive standard control to establish a real-time fluorescent quantitative PCR based on FAM-Eclipse probe method. The specificity and sensitivity of the established method were evaluated. Moreover, it was validated in terms of clinical sample detection.Results:The standard curve equations of the real-time fluorescent quantitative PCR for detecting TTV7, TTV8 and TTV10 were y=-0.340 2 x+ 114.780 0 ( R2=0.998 8), y=-0.351 1 x+ 114.940 0 ( R2=0.995 3) and y=-0.348 9 x+ 115.020 0 ( R2=0.991 7), respectively, and there was no cross-reaction with other viruses. The detection sensitivity of the established method for TTV7, TTV8 and TTV10 were 108 copies/μl, 84 copies/μl and 98 copies/μl, and the positive detection rates in clinical pediatric serum samples were 10.9%, 2.1% and 4.3%, respectively. Conclusions:The established real-time fluorescent quantitative PCR for detection of TTV7, TTV8 and TTV10 was featured by strong specificity and high sensitivity, which could be used for rapid TTV detection in clinical serum samples.

Establishment and verification of real-time fluorescent quantitative PCR detection system for ring virus 6 / 中华实验和临床病毒学杂志

Objective@#To establish a real-time quantitative PCR detection system for Torque teno virus (TTV) and verify the sensitivity and specificity of the detection system.@*Methods@#Primers and FAM-Eclipse probes were designed based on the TTV6 gene sequence registered in GenBank, and were to establish a real-time fluorescent quantitative PCR detecting way based on the FAM-Eclipse probe, the standard curve was constructed and sensitivity and specificity were analyzed.@*Results@#A quantitative PCR method for the specific detection of TTV6 were established that the standard curve equation was y=-3.0921x + 28.36, and the amplification efficiency and R2 were 99.6% and 1.000, respectively. The sensitivity of TTV6 was 1.0×10 copies/μl, and there was no cross-reactivity with other viruses. There was 1 case positive for TTV6 out of 56 throat swab samples from the patients with clinical respiratory infection.@*Conclusions@#The real-time fluorescent quantitative PCR for detecting TTV6 established by FAM-Eclipse probe had the advantages of high sensitivity and specificity. It provides an effective way for detection and quantification of viral content of TTV6 in clinical specimens.

Multiple serotypes of Rhinovirus infection occurred in inpatients of an emergency department, Bengbu, Anhui province, in winter of 2018 / 中华实验和临床病毒学杂志

Objective@#To investigate the incidence of human rhinovirus (HRV) infection in hospitalized patients in emergency department.@*Methods@#A total of 94 emergency patients admitted to the emergency department from November to December of 2018 were enrolled in this study. The rhinovirus infection and related risk factors were analyzed.@*Results@#HRV infection occurred in 17 out of 94 hospitalized patients in emergency department, the infection rate was 18.09%; Multiple HRV serotypes were prevalent from November to December of 2018, which were A9, A10, A16, A31, A73, B42 and C3. Elderly patients are at high risk of rhinovirus infection; Rhinovirus infection may increase the course of inpatients.@*Conclusions@#From November to December of 2018, multiple serotypes of HRV infections in the emergency department of Bengbu, Anhui, and HRV easily infected elderly inpatients.

Isolation and identification of Rhinovirus from throat swab samples of patients with acute respiratory infection in Bengbu, Anhui province / 中华实验和临床病毒学杂志

Objective@#To isolate and identify human rhinovirus (HRV) from throat swab samples from patients with acute respiratory infection in Anhui Bengbu.@*Methods@#The throat swab specimens from 108 patients with acute upper respiratory tract infection diagnosed in the Anhui Bengbu were selected as samples. RNA was extracted and detected by HRV universal primers; H1-Hela cells were infected with the positive samples for virus isolation, and HRV VP1 gene amplification was performed and gene phylogenetic tree analysis of the successfully isolated HRV strains was done.@*Results@#Only 5 samples were positive for HRV by the real-time PCR, and only one sample showed significant cytopathic effects after three passages of H1-Hela cells were infected, and the HRV VP1 gene was amplified by RT-PCR in the sample that was named TYZQ201901. The sequence analysis showed that the VP1 nucleotide homology of the strain with other representative HRV A strains was over 95%. The gene phylogenetic tree analysis showed that the strain had the closest genetic distance to the RMH127/2013 strain, and both were on a branch and was confirmed to be HRV type A virus.@*Conclusions@#An HRV type A strain virus was isolated from throat swab samples from patients with acute respiratory infection in Anhui Province by the H1-Hela cells, and the HRV virus separation technology system were initially established.

Changes of endogenous small interfering RNA in H1-HeLa cells induced by human rhinovirus 16 infection / 中华实验和临床病毒学杂志

Objective@#To observe the changes of endogenous small interfering RNA (siRNA) in H1-Hela cells infected with human rhinovirus 16 (HRV 16).@*Methods@#To determine whether HRV16 infection induced the changes of siRNA, H1-HeLa cells were infected with HRV16 for 12 h, 24 h and 36 h, siRNAs were detected by high-throughput sequencing, second-generation sequencing) and qRT-PCR.@*Results@#The result showed that siRNA was generated differently at different time points post-infection, among which novel_sir907 and novel_sir1950 decreased at three time points. Further validation by qRT-PCR showed that novel_sir907 decreased at 12 h, 24 h and 36 h post-infection compared with the cell control, but novel_sir1950 increased at 12 h then decreased at 24 h and 36 h.@*Conclusions@#HRV16 infection induces changes endogenous siRNAs.

A preliminary study on thapsigargin promoting coxsackievirus B3 replication through activating transcription factor 6 pathway / 中华实验和临床病毒学杂志

Objective@#To investigate the effect of thapsigargin (TG) which can induce endoplasmic reticulum stress (ERS) on the replication of coxsackievirus B 3 (CV-B3).@*Methods@#After 10 MOI CV-B3 infected HeLa cells were exposed 0.25 μmol/L TG for 3 h, 6 h and 9 h, virus RNA of HeLa cells were extracted and viral replication was evaluated by real time PCR. After 0.25 μmol/L、0.08 μmol/L and 0.025 μmol/L TG exposed, the plaque of CV-B3 was used to confirm further replication of CV-B3. To verify TG induced ERS through three signal pathway, one of among PERK, ATF6 and IRE1 inhibitors GSK2656157, AEBSF and STF-083010, and 0.25 μmol/L TG were used in HeLa cells infected with 10 MOI CV-B3, replication of CV-B3 was evaluated by qRT-PCR.@*Results@#The stimulation of TG did not induce increase of virus replication after post-infection 3 h. However, TG induced replication of virus to increase 2.5 times after post-infection 6 h and 158.6 times after post-infection 9 h. And, the area of viral plaque was significantly increased. ATF6 inhibitors AEBSF significantly inhibited promotion of virus replication from TG.@*Conclusions@#TG can promote the replication of CV-B3 through ATF6 signal pathway.

A autophagy induced by non-stuctural protein 2B of human rhinovirus / 中华实验和临床病毒学杂志

Objective@#To study the intracellular location and autophagosome production of rhinovirus 16 2B protein using miniSOG labeling technique.@*Methods@#2B was fused with miniSOG and flag tags to construct pcDNA3.1-2B-miniSOG-flag plasmid, which was used to transfect HEK293 cells, LC3 protein was detected by western blot. The transfected cells were fixed, stained with DAB through the photooxidation activity of miniSOG, and used to prepare ultrathin sections. Localization of 2B-miniSOG protein in cells and ultrastructural changes of cells were observed under electron microscope.@*Results@#2B-miniSOG protein glows green under a fluorescence microscopy. Green flourescence coold be observed in the cells expressing 2B-miniSOG protein.LC-II protein increased in the cells transfected with pcDNA3.1-2B-miniSOG-flag. Under electron microscopy it was observed that 2B-miniSOG protein was located in the mitochondria, and a large number of vesicular structures appeared in the cytoplasm. Both autophagosomes and autophagic lysosomes can be observed.@*Conclusions@#Non-structural protein 2B of HRV16 can induce autophagy.

Analysis on global Rhinovirus research advances / 中华实验和临床病毒学杂志

Objective@#To explore the research status of rhinovirus (RV) through analysis of rhinovirus literature using GoPubMed.@*Methods@#"Rhinovirus" was used as the major subject word and the rhinovirus literature was collected at PubMed database (from Jan 1, 1970 to April 16, 2018). The high frequency subject words of rhinovirus related literature and the distribution of countries, cities, and journals were analyzed through a bibliometrical analysis method .@*Results@#A total of 5 367 reports were retrieved from PubMed. The quantity of rhinovirus papers increased overall year by year. The highest number of papers were mainly published in developed countries. The highest number of papers on RV were mainly published in J Virol among all journals related with rhinovirus and Tyrrell D published the highest number of papers in all authors contributed to articles on rhinovirus. The rhinovirus, human, virus, respiratory tract infection were the high frequency subject words in the rhinovirus research.@*Conclusions@#Rhinovirus research is becoming one of research hotspots according to the statistical analysis of the research literature on rhinoviruses by GoPubMed.

Knowledge of voluntary blood donation and factors influencing the willingness among rural residents in Anhui / 中国输血杂志

Objective To get a general idea of the knowing level of knowledge and policies related to voluntary blood donation among rural residents,and to explore factors influencing their willingness of voluntary blood donation.Methods Three natural villages were randomly chosen in the north,middle and south of Anhui province as the investigation sites in Jan.2008.Data were collected by interviewing the target people,as well as using a self-designed questionnaire.A total of 724 people were interviewed and 620 valid questionnaires were obtained,with an effective retrieval rate of 85.6%.Results The score of voluntary blood donation knowledge among rural residents was 19.26?3.987,and the score of relevant policy knowledge was 5.85?2.154.There were significant differences in knowledge of both voluntary blood donation and relevant policies among different age groups.Residents with different education levels also differed significantly in knowledge of relevant policies.Logistic analysis indicated that age,education and knowing level of relevant policies were the factors that had influence people's willingness of voluntary blood donation.Conclusion Knowing level of voluntary blood donation knowledge is high among rural residents while the policy knowledge level is low.Willingness of blood donation among the group of age 25 to 34 is obviously higher than that of the group of age 18 to 24;willingness of people with middle school education background is lower than illiterate people;the higher the knowledge level,the higher of people's willingness.

Abnormal DNA-binding abilities of NF-?B and glucocorticoid receptor in cortex of kindey from rats with Adriamycin-induced nephrosis / 中国医师杂志

Objective To explore the role of transcription factors such as nuclear factor-kappa B(NF-?B) and glucocorticoid receptor(GR) in the pathogenesis of Adriamycin(ADM)-induced nephrosis in rats and the therapeutic effects of dexamethasone(Dex) and cyclosporin(CsA) on these animals.Methods The DNA-binding abilities of NF-?B and GR in cortex of kindey were examined with electrophoretic mobility shift assay(EMSA) and isotopic radioautography on the 7th,14th,21th and 28th day after a single intravenous injection of ADM,and the therapeutic effects of Dex and CsA were estimated.The biochemistry parameters from blood and urine of rats and the urine protein excretion were also measured.Results The NF-?B DNA-binding ability was significantly increased after 7 days and achieved maximum after 28 days(P0.05).Conclusion The DNA-binding ability of NF-?B is abnormally increased and that of GR is decreased in cells from cortex of kindey in Adriamycin-induced nephrotic rats.The NF-?B DNA-binding ability can be inhibited and the urine protein excretion is decreased by the treatment of CsA.

Thinking of the fairness about family planning services in urban and rural / 中国医学伦理学

The paper discusses the problems and inequity in town and country family-planning service by comparing the following aspects: work thinking in the service,accessibility of family-planning service,service capacity of the organizations,resource allocation.Besides,in order to satisfy the demand of rural family-planning service market and improve the equity,some suggestions,such as integrating resource and complementing advantages to implement regional family-planning program,identifying the functions of government and changing the service thinking,increasing the investment in primary health family-planning organizations and reinforcing the co-operation between health and family-planning organizations.