Objective To compare and analyze the differences and characteristics of three strain mouse models in-fected by influenza virus aerosol inhalation, and provide the reference for choosing the appropriate infection model in the re-search of pathogenesis of influenza and the development of vaccines and drugs.Method C57BL/6, BALB/c and ICR mice were infected with A/Puerto Rico/8/34 (H1N1) virus strain by aerosol inhalation.The symptoms and body weight of mice were observed every day.At 3, 7, 14 days after infection, the mice were sacrificed.The lungs of mice were weighed, then virus assay and pathological observation were carried out.Results The three strains of mice were infected.The sur-vival rate in the C57BL/6 mice was lower than those in the BALB/c and ICR mice.The lung index and viral load of C57BL/6 mice were significantly higher than those of ICR mice ( P<0.05) at 3 days after infection.The pathological changes of C57BL/6 mice were also more obvious than other two strains.Compared with other two mouse strains, the weight recovery of BALB/c mice was the slowest.The survival rate in BALB/c mice was higher than that of C57BL/6 mice and lower than that of ICR mice.The lung index and viral load were not significantly different among the three strains of in-fected mice.The pathological changes among the three strains of infected mice were similar, but the degrees of pathological changes in the BALB/c mice were milder than in the C57BL/6 mice and worse than in the ICR mice.Compared with other two mouse strains, the process of disease is similar, but the body weight, mortality, lung index, viral load, and the micro-scopic pathological changes were lighter in the ICR mice than in the other two strain mice.Conclusions The three strain mouse models can be established by influenza virus aerosol inhalation, but showing different characteristics.Appropriate strain mice can be chosen to build model according to different research purpose in the experiment.

Objective To develop mycobacterial inducible expression vectors which permit to overexpress Mycobacterium tuberculosis (Mtb) immunodominant antigen, and to analyze its immunogenicity after purification by affinity chromatography. Methods The regulatory region of M. smegmatis (Ms) acet-amidase(pACE) was obtained by PCR amplification, and was used as promoter to construct the mycobacteri-al inducible expression vectors, pMF series. The coding gene of Mtb chimeric antigen Ag856A2 which is a recombinant Ag85A with 2 copies of ESAT-6 inserted in its Acc Ⅰ site and showed excellent immunogenicity in the animal experiments we described previously, was cloned into the pMF vector series, and was induced to express by the addition of acetamide. The recombinant protein expressed in the Ms was purified by the Ni~(2+)-NTA affinity chromatography, the resulted homologous recombinant antigen was added into the spleen cells separated from BCG vaccinated mice, and the immunogenicity was analyzed by the IFN-γ ELISPOT as-say. Results The mycobacterial inducible expression vectors, pMF series was constructed successfully, target antigen could be. induced to express in the Ms by the addition of 0.02% acetamide, and could be puri-fied by the Ni~(2+) -NTA affinity chromatography due to the addition of 6×His tag in the vector pMF406. Fur-thermore, the mycobactefial homologous antigen could induce more IFN-γ secretion than the heterogonous one. Conclusion The mycobacterial inducible expression system based on the regulatory region of Ms acet-amidase as promoter could permit the Mtb target antigen of interest overexpression and purification, and the immunogenicity of the homologous antigen from Ms is better than that of be expressed from E. coli, which may be more potential for immunological detection of tuberculosis.

Objective To evaluate the value of enzyme-linked immunospot assay (TB ELISPOT) combined with serum latex agglutination test (LA) for diagnosis of pulmonary tuberculosis plus pulmonary cryptococcosis.Methods Serum and biopsy specimens of 76 patients, who were suspected of pulmonary tuberculosis and/or pulmonary cryptococcosis based on clinical and imaging features, were collected from March 2006 to September 2008 in Shanghai Public Health Clinical Center. TB ELISPOT assay, LA and histopathological examination were performed in all the patients. Results Histopathological and pathogenic examination confirmed pulmonary cryptococcosis in 15 cases and pulmonary tuberculosis in 22 cases, pulmonary tuberculosis plus pulmonary cryptococcosis in 8 cases. The sensitivity and specificity of TB ELISPOT were 91% and 94.4%. The sensitivity and specificity of LA were both 100%. TB ELISPOT assay and LA test were both positive in the 8 cases of pulmonary tuberculosis plus pulmonary cryptococcosis.Conclusions The value of enzyme-linked immunospot assay combined with serum latex agglutination test is high for diagnosis of pulmonary tuberculosis plus pulmonary cryptococcosis.

Objective This study explored the effect of the multidisciplinary-team collaborative nursing model in physical examinations of people with critical conditions (based on test results).Methods We selected 962 patients with critical conditions based on test-result values found from February to April 2018 as the general process group treated through the routine nursing model,and we also selected 1009 patients with critical conditions based on test-result values found from May to July 2018 as the multidisciplinary collaborating group using a team nursing model.The multidisciplinary collaborative nursing team members included health management center nurses,outpatient nurses,resident nurses,and ward nurses.We compared visiting rates,hospitalization rates,average visiting times,and overall satisfaction after the patients received notification of their abnormal results.There were 488 male patients (50.7％) and 474 female patients (49.3％) in the general process group,with an average age of 51.9 ± 14.9 years;there were 537 male patients (53.2％) and 472 female patients (46.8％) in the multidisciplinary collaboration group,with an average age of 51.0 ± 13.0.Results For the multidisciplinary collaboration group and the general process group,respectively,the visiting rate was 53.0％ and 44.7％ (x2=13.65);the hospitalization rate was 26.7％ and 20.9％ (x2=4.38);overall satisfaction was 97.9％ and 95.9％ (x2=6.49);and the average visiting time was 4 days and 6 days (Z=5.04).The differences were statistically significant (P＜0.05).By category,the visiting rate for radiology and ultrasound patients among the multidisciplinary collaboration group was significantly higher than in the general process group (64.4％ vs.50.8％ for radiology,45.9％ vs.37.3％ for ultrasound,x2=7.65,7.11,P＜0.05).The hospitalization rate for ultrasound patients in the multidisciplinary collaboration group was significantly higher than in the general process group (12.5％ vs.6.4％,x2=10.17,P＜0.05).The average visiting time of ultrasound,abnormal blood pressure,and laboratory testing patients was significantly lower in the general process group (4 days vs.6 days,4 d vs.7 days,4 days vs.5 days,Z=3.37,1.97,2.62,P＜0.05).The overall satisfaction of radiology patients was significantly higher than in the general process group (98.6％ vs.94.3％,x2=5.39,P＜0.05).Conclusion The multidisciplinary team collaborative nursing model improves the rate of visiting and hospitalization of patients with critical conditions after physical examination,shortening their stays,helping patients get timely diagnoses and treatment,and improving patient satisfaction,making the model worth popularizing and applying more broadly.

Objective: To analyze the drug resistance of clinical isolates of Candida tropicalis in patients with infectious diseases, and preliminarily study their molecular characteristics. Methods: 95 strains of Candida tropicalis were isolated from the fungal culture specimens of 87 patients with infectious diseases in Shanghai Public Health Clinical Center from 2012 to 2015. Meanwhile, basic clinical data of patients were collected. The drug resistance of the strains to fungal drugs was analyzed by ATB FUNGUS 3 drug sensitivity test strips. All strains were classified by Multilocus sequence typing(MLST). Then, homology analysis was conducted by MEGA 5.2 software, and the evolutionary tree was mapped by using UPGMA method. Results: Patients distribution of strains was rendered as following: 31 strains from TB patients, 21 strains from HIV/AIDS patients, 19 strains from patients with liver disease, and 24 strains from rare cause infection or fever patients. The drug resistance rate to five antifungal drugs commonly used in clinical (amphotericin B, 5-fluorine cytosine, fluconazole, itraconazole, voriconazole) were 2.11% (2 strains), 0, 26.32% (25 strains), 26.32% (25 strains), and 26.32% (25 strains) respectively. Among the 25 azole-resistant strains: 14 strains were from rare cause infection or fever patients, 8 strains were from HIV/AIDS patients, and 3 strains were from tuberculosis patients. In MLST, 72 sequence types (ST types) were produced, 70 of which were new types. Evolutionary tree analysis showed that 95 strains of clinical strains distribute as three large clusters. 24 azole resistant strains (96.0%) were located in CLUSER Ⅰ. Conclusion The isolated Candida tropicalis were mainly resistant to azole drugs. MLST typing indicates that they was closely related to their genetic background.

OBJECTIVETo determine the function of twin-arginine translocation system (Tat) and gene cluster in Vibrio strains and to analyze the homology of tat gene cluster among different Vibrio spp. strains based on N16961 and tatABC mutant strains N169-dtat.

METHODSDifferent serotypes of biotype strains of Vibrio spp. were selected to detect the transcription of 4 genes of Tat transport system and upstream ubi aarF gene and downstream cyt551 gene by the total RNA reverse transcription and homologicity of the gene cluster by sequencing analysis.

RESULTSOur results showed that the 4 genes of tat cluster (tatA, tatB, tatC, and tatE) were intragenic and co-transcribed. We found that ubi aarF gene could be co-transcribed with tatA, tatB, but not with tatC. The electron transport chain and energy metabolism-related genes, cytochrome C551 peroxidase gene, and 4 genes located at upstream of tatABC operon were not transcribed with tatABC. Although the co-transcription between ubi aarF and tatAB was blocked in N169-dtat strain, they were still transcribed separately. Homologous analysis of genes of tat cluster in different types of Vibrio cholerae showed that tat gene cluster was a very conservative.

CONCLUSIONThe ubi and aarF gene might be co-transcribed with genes of tat cluster in Vibrio cholerae, which and the close relationship showed that they might play a key function in Vibrio cholerae.

Arginine ; Bacterial Proteins ; Cytochrome c Group ; Membrane Transport Proteins ; Vibrio cholerae

Objective To investigate the species distribution characteristics of nontuberculous mycobacteria(NTM)in acquired immunodeficiency syndrome(AIDS)patients in Shanghai,and to provide evidences for clinical treatment.Methods A total of 775 Mycobacteria strains were isolated from patients(including 129 isolates from AIDS patients and 646 isolates from HIV-negative patients)admitted to Shanghai Public Health Clinical Center during 2015.All the species were identified by the sequence analysis of 16S rRNA gene and hsp65 gene.Differences in the species distribution were compared between patients with and without HIV infection.CD4+T lymphocyte count was detected by flow cytometry and its relation with mycobacteria infection was also analyzed.Chi-square test was used for comparison between groups.Results The ratio of NTM isolation from HIV-negative patients was 15.79%(102/646),while that was 46.51% in AIDS patients(60/129),and the difference was statistically significant(x2=61.38,P <0.01).Among the 60 NTM strains isolated from AIDS patients,mycobacterium avium-intracelluare complex(MAC)and Mycobacterium gordonae were the predominant species(43.33% and 20.00%,respectively).Moreover,five Mycobacterium colombiense strains,which were relatively rare,were also obtained.A total of 102 NTM were identified from HIV-negative patients,of which Mycobacterium gordonae(32 isolates,31.37%)and MAC(30 isolates,29.41%)were the most frequently isolated.In addition,the positive rates of NTM and MAC were significantly higher in patients with CD4+T lymphocyte counts ≤50 cells/μL(58.33% and 76.92%,respectively)than those with CD4+T lymphocyte counts>50 cells/μL(20.00% and 15.38%,respectively)(x2=4.048 and 6.524,respectively,both P<0.05).Four out of five Mycobacterium colombiense infected patients died of disseminated infections,except the remaining one whose CD4+T lymphocyte counts>50 cells/μL.Conclusions The prevalence of NTM isolation is significant higher in AIDS patients than HIV-negative patients in Shanghai,and the most prevalent NTM species is MAC.The NTM infection in AIDS patients is related with low CD4+T lymphocyte counts.

OBJECTIVETo analyze virulence genes and molecular characteristics of Vibrio parahaemolyticus isolated from sporadic cases with diarrhea in tow sentinel hospitals of Shanghai, 2010-2012.

METHODSA total of 2 729 stool samples were collected from two surveillance sentinel hospitals in Shanghai 2010-2012. Vibrio parahaemolyticus strains were isolated and identified from diarrhea out patients using TCBS agar plates and biochemical reactions. Thermostable direct hemolysingene (tdh), thermostable-related hemolysin gene (trh), hemolysin gene (tlh) were detected by multiplex PCR method. Isolates were analyzed by PFGE and MLST. The PFGE profiles were analyzed using BioNumerics software.

RESULTSA total of 30 clinical Vibrio parahaemolyticus strains isolated from 2 729 stool samples. The anually Vibrio parahaemolyticus isolation rate during 2010 to 2012 were 1.1%(11/973), 1.0%(11/1 120) and 1.3%(8/636) respectively. The PCR positive rates of virulence genes tlh, tdh and trh were 100%, 97% and 0 respectively. The Vibrio parahaemolyticus strains were divided into 13 PFGE types (P1-P13)and 3 ST types (ST-189, ST-799, ST-3). Among 13 PFGE types, P4 was the main PFGE type, accounting for 30%(9/30). P9, P10 were accounting for 12% (4/30) respectively, P1, P2, P12, P13 were accounting for 7%(2/30) respectively, the others types were 3%(1/30) respectively. MLST analysis results showed there are three ST types, ST3 was 84%(25/30), ST189 and ST799 were accounting for 13% (4/30) and 3% (1/30) respectively.

CONCLUSIONThe infection rate of Vibrio parahaemolyticus was not very high from 2010-2012 in Shanghai, all strains were positive for tlh and negative for trh. ST3 was the major type of Vibrio parahaemolyticus.

China ; Diarrhea ; Genotype ; Hemolysin Proteins ; Hospitals ; Humans ; Multilocus Sequence Typing ; Outpatients ; Polymerase Chain Reaction ; Sentinel Surveillance ; Vibrio Infections ; Vibrio parahaemolyticus ; Virulence

Objective: To analyze the changes in the expression of hypoxia inducible factor-1α (HIF-1α) and inflammatory cytokines and to investigate the role of HIF-1α in regulating the production of inflammatory cytokines during influenza A (H1N1) virus infection. Methods: BALB/c mice were injected with H1N1 virus to establish the mouse model of H1N1 virus infection. Fifteen BALB/c mice were randomly divided into three groups: control group, H1N1 virus group and H1N1 virus+ HIF-1α inhibitor group. Inflammatory cytokines (IL-6, TNF-α, IL-1β and IL-10) in samples of serum and lung tissues were detected by Luminex and ELISA. Levels of HIF-1α in serum and lung tissue samples were detected by Western blot and ELISA, respectively. Results: Compared with the control group, the levels of inflammatory cytokines in serum (IL-6, TNF-α, IL-1β and IL-10) and lung tissues (IL-6 and TNF-α) and the expression of HIF-1α in serum and lung tissues in the H1N1 virus group were significantly increased. The levels of HIF-1α, IL-6, TNF-α IL-1β and IL-10 in lung tissues in H1N1 virus+ HIF-1α inhibitor group were significantly lower than those of the H1N1 virus group. Conclusion During H1N1 virus infection, the levels of inflammatory cytokines and HIF-1α were significantly increased. The production of inflammatory cytokines was significantly reduced after inhibiting HIF-1α expression, suggesting that HIF-1α might promote the production of inflammatory cytokines.

The ongoing pandemic of Coronavirus disease 19 (COVID-19) is caused by a newly discovered β Coronavirus named severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). How long the adaptive immunity triggered by SARS-CoV-2 can last is of critical clinical relevance in assessing the probability of second infection and efficacy of vaccination. Here we examined, using ELISA, the IgG antibodies in serum specimens collected from 17 COVID-19 patients at 6-7 months after diagnosis and the results were compared to those from cases investigated 2 weeks to 2 months post-infection. All samples were positive for IgGs against the S- and N-proteins of SARS-CoV-2. Notably, 14 samples available at 6-7 months post-infection all showed significant neutralizing activities in a pseudovirus assay, with no difference in blocking the cell-entry of the 614D and 614G variants of SARS-CoV-2. Furthermore, in 10 blood samples from cases at 6-7 months post-infection used for memory T-cell tests, we found that interferon γ-producing CD4
Adaptive Immunity/physiology* ; Adult ; Aged ; Antibodies, Neutralizing/blood* ; COVID-19/immunology* ; Cohort Studies ; Female ; Humans ; Immunoglobulin G/blood* ; Male ; Middle Aged ; SARS-CoV-2/immunology* ; T-Lymphocytes/physiology* ; Time Factors ; Viral Proteins/immunology*