Objective To clone and express the valuable Clonorchis sinensis antigen molecules which can be applied to the diagnosis of clonorchiasis. Methods Based on the sequences (Genbank) No. AF271091 (CysA) and No.AF093242 (CysB), primers were designed to amplify the two C. sinensis cysteine proteinase genes and expressed in E.cloi. The expressed proteins were purified by affinity chromatography and then tested for their immunological characters.Results The two genes were successfully cloned and expressed. Western blot showed that CysB had strong reaction with clonorchiasis sera and very weak reaction with schistosomiasis sera, while CysA showed no reactivity with the probed sera. Immunohistochemistry showed that both proteins were mainly located in adult worm intestines and the intrauterine eggs.Conclusions The results suggested that, of the two expressed C. sinensis proteins, CysB had good antigenic reactivity against sera from patients. It is a potential candidate of diagnostic antigens for clonorchiasis.

Objectives The ferredoxins are iron-sulfur proteins, which function in electron transfer reactions in a variety of systems and participate in the activation of the antimicrobial agent metronidazole. The aim of this study is to clone and characterize ferredoxin genes of Trichomonas vaginalis. Methods A cDNA expression library was constructed with T. Vaginalis total RNA. Hundreds of cDNA clones were isolated and sequenced. Sequence analysis was performed using BLAST programs, ClustalW program, etc. Results One of the cDNA clones, which has homology with T.vaginalis ferredoxin, was further analyzed. This cDNA clone has an open reading frame of 312 base pairs. The deduced precursor protein contains 103 amino acid residues with a hydrogenosome targeting sequence (MLSQCSPLRF) at the N-terminal end. The primary sequence analysis revealed that this new ferredoxin (TvFd2) has a high homology (69% identity) to the previous reported T.vaginalis ferredoxin(TvFd). Interestingly, TvFd2 is homologous to both the two subclasses of (2Fe-2S) ferredoxins, the oxidase ferredoxins and the photosynthetic ferredoxins,but with low similarity. The conserved four-cysteine residues, which are predicted to form the iron-sulfur cluster,are arranged in a typical pattern of (2Fe-2S)ferredoxins(-C-X5-C-X2-C-Xn-C-). Conclusion These data show that TvFd2 is a putative new (2Fe-2S) ferredoxin of T.vaginalis. Its biological function remains to be studied.

Objective To explore immunological mark and the outcome of disease after pharmacological treatment in human NCC with praziquantel. Methods 35 patients were treated with praziquantel for 6 months. Levels of serum IL-2,IFN-γ, IL-12, IL-5, IL-10 and TNF-α were measured before and after treatment. Results Th1 type cytokines IL-2, IFN-γand IL-12 were up regulated after treatment( P ＜ 0. 01 ) . Levels of TNF-α and Th2 type cytokines IL-5 and IL-10 were remarkable decreased after treatment( P ＜ 0. 01 ). The levels of IL-2, IFN-γ and IL-12 from the patients with good response to the treatment is higher than those with no response to the treatment. Conclusion Th1 type cytokines were up regulated while Th2 type cytokines were down regulated in the NCC patients treated with praziquantel. The protective immunity may be related to the Thl cell activation.

Objective To study the effect of of human cytomegalovirus (HCMV) infection on the expressions of Hox genes.Methods Forty eight kunming mice were randomly divided into infection group (n=32) injected with HCMVAD169 and control group (n=16) injected with saline into their brain.After 7,15,30,and 60 days,the cerebral lesions were observed by pathological method.HCMV antigen was detected by immunohistochemical method and HCMV DNA was detected by polymerase chain reaction (PCR). On the basis of developing HCMV mouse model. reverse trancriptase-polymerase chain reaction (RT-PCR) was applied to determine the expressions of Hox gene in the brains of infected mice.The expression of Hox genes were also analysed with Northern-blot by isotope labelled Hox genes oligonucleotide probes. Results A HCMV infection model was developed and extensive pathological damages in brain tissue of infected mice were observed.Meanwhile.the HCMV-LA and HCMV-DNA were also found in brain tissues of HCMV infected mice.The expression level of Hox genes in control and infected mouse brain were determined by RT-PCR and Northern-blot.RT-PCR and Northern-blot showed that mouse brain expressed Hox-A9,Hox-A10,Hox-A11,Hox-A12,and Hox-A13,but they did not express Hox-B13.After HCMV infection,murine brain was induced to express Hox-B13 gene(P＜0.01),and reached the peak at 30 d after infection.Comparing with the control group,the expression of Hox-A9 and Hox-A11 were down-regulated in infected group (P＜0.05);the expression of Hox-A10 and Hox-A13 were significantly higher in infected groups (P＜0.05).Conclusion The results suggest that HCMVAD169 is able to cause mouse CNS infection and induced the abnormal expressions of Hox genes. which provides more information for understanding the mechanism of congenital abnormal due to HCMV infection and a valuable method of clinical prevention and treatment of HCMV infection.

Objective To evaluate the identification and counting effeciency of nucleated red blood cells by Sysmex XE-2100 Hematology Analyzer. Methods Accurancy: nucleated red blood cells were counted from 38 specimens by Sysmex XE-2100 Hematology Analyzer and compared with the number counted under microscopy. Precision: 3 specimens with different values were counted for the nucleated red blood cells 10 times by Sysmex XE-2100 Hematology Analyzer and compared with the number counted under microscopy. The CV(% ) value was estimated.Results There was insignificant difference between the results obtained from Sysmex XE-2100 Hematology Analyzer and those under microscopy. In the T-test, P >0.05, r=0.9893(P< 0.01).CV(% ) were 8.1% and 15.8% . It means that the Sysmex XE-2100 is more precise in analyzing the nucleated red blood cells than that under microscopy. Conclusion The nucleated red blood cells count by Sysmex XE-2100 is accurate and fast to obtain clinical data.

Abstract：Objective To develop a colloidal gold immunochromatographic strip ⁃ based method for the rapid detection of Zika virus（ZIKV）NS1 antigen. Methods The gold nanoparticles modified with the anti⁃ZIKV NS1 monoclonal antibody as the detection probe were coated on the glass ⁃fiber pad. The anti ⁃ZIKV NS1 monoclonal antibody and the goat anti ⁃mouse polyclonal antibody were immobilized on a nitrocellulose membrane as the test line and the control line，respectively. In order to achieve critical results，the ratio of the optical density （OD）of the test line to that of the control line was compared. Serial diluted ZIKV NS1 standard antigen was applied to evaluate sensitivity of the immunoassay. The culture supernatant and serum samples for arboviruses（ZIKV，Dengue virus, Japanese encephalitis virus and Chikungunya virus） were utilized to demonstrate the specificity of the method. Results The detection result could read by naked eyes within 20 minutes. The visual cut ⁃off level for the test strip was achieved at 100 ng/mL of the Zika virus NS1 standard antigen. No cross⁃reactions with Dengue virus，Japanese encephalitis virus and Chikungunya virus were observed. The strip could remain good stability within 36 weeks whether stored in 4 ℃ or room temperature（22-25 ℃）. Conclusion Apart from stability， the method was convenient，rapid and specific for ZIKV NS1 antigen，which showed a promising potential in the point of care test and the screening test.

Abstract：Objective To establish a sensitive，rapid and convenient method for the detection of dengue antigen and assist clinical diagnosis of dengue. Methods In this paper，we developed a rapid detection method for dengue antigen based on microfluidic immune magnetic beads. Solidwork software was used to design microfluidic chip，which was prepared by mechanical processing and chemical sealing. Immunomagnetic beads of dengue antibody were prepared by chemical coupling reaction. Using HRP ⁃TMB ⁃H2O2 as color system，dengue NS1 antigen was detected on microfluidic chip carrier by double antibody sandwich method. Finally，57 clinical samples were tested by the novel method and traditional ELISA kit，and the accuracy of the method was analyzed，and the advantages and disadvantages of the two methods were compared. Results 20 minutes was needed to detect dengue NS1 antigen by using the novel ELISA method，and the reaction system only needed 10 μg beads and 10 μL samples. In the verification experiment，the method could distinguish the negative from the positive obviously. The positive sample had color rendering，while the negative and blank samples had no color rendering. In terms of detection performance，the coincidence rate between the new ELISA method and the traditional ELISA method reached 100%. Conclusion The novel ELISA detection platform had the advantages of simple，rapid，reagent and sample saving，high sensitivity，good stability and high accuracy，and could be used for the detection of dengue antigen.

Respiratory tract infection；Multiplex PCR；Children Abstract:Objective The aim of the study was to evaluate a multiplex molecular real ⁃ time assay for the diagnosis of respiratory viruses of childhood acute respiratory virus infection，and analyze the symptoms associated with the pathogens which were detected from those samples. Methods From August 2018 to August 2019，a sample of 275 swallow swabs from children with acute respiratory infections in Chengdu Children′s Specialist Hospital was collected and tested using the RespiFinder®SMART 22 test kit. The kit is based on multiplex PCR technology and can detect and distinguish 22 pathogens related to respiratory infections at one time. Results Of the 275 samples，245（89.1%）detected positive viral infections， of which 160 were single virus infections，59 were multiple viral infections，17 were positive for Mycoplasma pneumoniae， and 9 were positive for Chlamydia pneumoniae. The most common pathogens were rhinovirus 77 cases（28.0%），adenovirus 45 cases（16.4%）and respiratory syncytial virus 34 cases（12.4%）. Influenza virus was detected in 28（10.2%）samples， parainfluenza in 25 cases（9.1%），coronavirus in 15 cases（5.5%），Boca virus in 12 cases（4.4%），and human metapneumovirus in 9 cases （3.3%）. Rhinovirus （28.0%） was the most common pathogen in upper respiratory tract infections，while respiratory syncytial virus（12.4%）was the most common pathogen in lower respiratory tract infections. It has been found that multiple viral infections in children were associated with severe respiratory symptoms in clinical manifestations. Conclusion Compared with ordinary PCR detection，the multiplex PCR could detect multiple pathogens at one time in experiment，which greatly shortens the detection time and had important clinical significance for rapid diagnosis of multiple respiratory pathogens. Thus，it could be useful for detecting several viruses causing respiratory tract disorders.

Objective To analyze the status of knowledge，attitude and practice of the coronavirus disease（COVID ⁃19）among the adults of 18⁃59 years old in China，and to provide scientific basis for corresponding health education strategies. Methods In the rapid development phrase of COVID ⁃ 19，subjects from all provinces or municipalities of China were invited to participate in a quick questionnaire online survey on January 29th，2020. Results The effective response rate of completing questionnaire was 97.41%（3 083/3 165）. 98.54% of the subjects reported that they were very terrified. The main reasons included the high contagion（64.71%）and lack of effective treatments（19.92%）；94.45% of the subjects were concerned that they and their family members would be infected by the novel coronavirus. 99.42% knew that the virus could be transmitted from person to person；97.89% and 93.87% knew that it could spread through respiratory tract and contact respectively，97.73% knew that patients without symptom could also be contagious，96.37% knew that persons in close contact to COVID⁃19 patient were required to be quarantined for at least 14 days of medical observation. 99.09% knew that the pathogen of this disease was novel coronavirus. 65.46% knew that both medical protective masks and surgical masks could prevent COVID⁃19 effectively. 99.68% had confidence in defeating COVID⁃19，and 85.86% believed that COVID⁃19 would be controlled within the next 3 months. Study subjects mainly obtained health information through WeChat（88.97%）or websites（82.06%）. The proportions of the subjects who can cover mouth and nose when coughing or sneezing，avoid hand contacting with eyes，mouth or nose，practice hand hygiene，wear masks outside，avoid exposure to respiratory patients，and avoid the crowded were 89.85%，85.44%，95.13%，96.89%，92.18% and 96.27%，respectively. Multivariate Logistic analysis showed that gender（OR=0.544，95%CI：0.440⁃0.673，P<0.001），age（OR=1.844，95%CI：1.466⁃2.320，P<0.001），recognition （OR=2.200，95% CI：1.780 ⁃ 2.718，P<0.001） were associated with those good behaviors. Conclusion After the happened，the government and society′s vigorous publicity to the public achieved good results. The public are highly concerned and have a high awareness of the knowledge of COVID ⁃ 19. They adopt protective measures proactively. Females，middle⁃aged，and individuals with insufficient recognition are likely under⁃protected. In the different epidemic stages of the emerging infectious disease，health education should be carried out to the public based on scientific evidences.

Objective To screen cell growth and senescence-related genes of the parasitic pmtist Trichomonas vaginalis,we launched an EST program and isolated two cDNA clones from a T.vaginalis cDNA library,which showed high homology in deduced amino acid sequences to yeast Sir2 and designated as TvSir2 and TvSir2-like.Method The cDNA sequence of TvSIR2 had a length of 1034 base pairs (bp) with an open reading frame of 915 bp,and TvSIR2-like,1214 bp with an open reading frame of 1116 bp.Result The two deduced amino acid sequences shared all the three conserved cole domains with yeast Sir2 and its homologues,suggesting that the two clones were Sir2 homologues. A cDNA fragment from each cDNA clone was subvloned into the expression vector pET-41a.The expression of the fusion proteins in E.coli BL21 stains was induced by isopropylthio-β-D-galactoside (IPTG).Two anti-sera were prepared by immunizing two guinea pigs with the purified fusion proteins, Western-blot analysis demonstrated that each anti-serum reacted with the corresponding recombinant protein and detected a clear band (TvSir2,34 000 Mr;TvSir2-like,42 000 Mr)in protein extracts of the protist.Immunofluolescence techniques showed that TvSir2 and TvSir2-like proteins were both localized in the legions of perinuelear (ER) and Golgi complex.Conclusion Our data suggest that TvSir2 and TvSir2-like were two members of Sir2 family.Their biological functions in the protist would be further studied.