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ABSTRACT The gut microbiome has a pivotal function in human immunodeficiency virus (HIV). However, the associated alterations in the gut microbiome-host interaction are unknown. Herein, we aimed to investigate the gut microbiota and fecal metabolites in people living with HIV (PLWH). We collected stool samples from 70 PLWH and 34 healthy controls (HCs) and carried out 16S rRNA gene sequencing and analyzed the metabolites using liquid chromatography-mass spectrometry. Firmicutes, Proteobacteria, Actinobacteriota, and Bacteroidota were the most abundant phyla in both groups. Among genera, the level of Escherichia-Shigella was upregulated significantly in the PLWH group, whereas in the HC group, Bacteroides spp. were upregulated. Prediction of microbial function indicated significant reductions in alanine, aspartate, glutamate, and histidine metabolism. Furthermore, a comparison of the fecal metabolites between the HC and PLWH groups identified 38 differentially abundant metabolites in four differentially enriched human metabolic pathways. According to Spearman correlation analysis, there are close relationships between four differentially abundant microbiota members and five differentially abundant fecal metabolites, which might influence particular human metabolic pathways. Our findings provide a basis for further experimental investigation of the contribution of the gut microbiota and its associated metabolites to HIV/AIDS, providing a novel perspective for the further study of HIV/AIDS.IMPORTANCEGrowing evidence demonstrates that the gut microbiota is associated with HIV. This study investigated changes in the gut microbiota and fecal metabolites in PLWH. We identified 38 differentially abundant metabolites in four differentially enriched human metabolic pathways. Moreover, close relationships were noted between the four differentially abundant microbiota members and five differentially abundant fecal metabolites, which might influence particular human metabolic pathways. Thus, to benefit PLWH, potential pathobionts could be reduced (e.g., g_Enterococcus); probiotics could be increased (e.g., g_Faecalibacterium and g_Agathobacter); or certain metabolites (e.g., N-acetyl-L-phenylalanine and trehalose) could be reduced by changes in diet or the use of nutritional supplements. Our results provide insights into the interaction between the gut microbiota and the host, identifying possible targets that might be beneficial for PLWH.

As an essential nutrient element for biological growth and metabolism, sulfur is closely interlinked with the carbon and nitrogen cycles, and it is one of the limiting elements for grassland productivity. Here we investigated the spatial distribution of sulfur contents and 34S stable isotope along the North China Transect (NCT), with the aim to explore the shaping role of the aridity index (AI) gradient on sulfur cycling dynamic in arid and semi-arid grasslands. In the area with AI < 0.12, soil sulfur contents and sulfur isotopic compositions (δ34S) showed no correlation with AI, indicating that abiotic processes predominantly govern the sulfur cycle in this area. In the area where 0.12 ≤ AI < 0.32, both sulfur contents and δ34S values increased with rising AI, with microbial-mediated reduction being the primary sulfur cycling process. In the area with 0.32 ≤ AI < 0.60, soil sulfur contents continued to increase with higher AI, but δ34S significantly decreased as AI increased, suggesting plant uptake as the dominant sulfur cycling process in this area. This study demonstrated the significant impact of AI on sulfur dynamics, providing insights into the different drivers of sulfur cycling along the aridity gradient, and offering guidance for developing targeted strategies under global climate change.

Objective: To monitor post-vaccination antibody production, neutralizing activity, and their dynamics over time in people living with HIV (PLWH). Methods: We collected sera from 147 PLWH and 94 healthy controls after vaccination at different time points and examined changes in antibody levels and neutralizing activity using enzyme-linked immunosorbent assay (ELISA) and pseudovirus neutralization assay. Results: IgG levels were substantially increased in both PLWH and healthy controls after the booster injection. Antibody levels decreased significantly in both PLWH and controls five months after the booster injection. However, the rate of decrease was not significantly different between the two groups. The generated antibodies demonstrated protective efficacy against the wild-type SARS-CoV-2 strain, but very low protection against the mutant strains. Furthermore, the protection decreased over time. The vaccine was less effective in PLWH with

Abstract Effective drugs with broad spectrum safety profile to all people are highly expected to combat COVID-19 caused by SARS-CoV-2. Here we report that nelfinavir, an FDA approved drug for the treatment of HIV infection, is effective against SARS-CoV-2 and COVID-19. Preincubation of nelfinavir could inhibit the activity of the main protease of the SARS-CoV-2 (IC50 = 8.26 μM), while its antiviral activity in Vero E6 cells against a clinical isolate of SARS-CoV-2 was determined to be 2.93 μM (EC50). In comparison with vehicle-treated animals, rhesus macaque prophylactically treated with nelfinavir had significantly lower temperature and significantly reduced virus loads in the nasal and anal swabs of the animals. At necropsy, nelfinavir-treated animals had a significant reduction of the viral replication in the lungs by nearly three orders of magnitude. A prospective clinic study with 37 enrolled treatment-naive patients at Shanghai Public Health Clinical Center, which were randomized (1:1) to nelfinavir and control groups, showed that the nelfinavir treatment could shorten the duration of viral shedding by 5.5 days (9.0 vs. 14.5 days, P = 0.055) and the duration of fever time by 3.8 days (2.8 vs. 6.6 days, P = 0.014) in mild/moderate COVID-19 patients. The antiviral efficiency and clinical benefits in rhesus macaque model and in COVID-19 patients, together with its well-established good safety profile in almost all ages and during pregnancy, indicated that nelfinavir is a highly promising medication with the potential of preventative effect for the treatment of COVID-19.

Summary: Here, we present a protocol to generate a murine model of liver metastasis by directly injecting tumor cells into the portal vein under ultrasound guidance. We describe steps for animal and cell preparation and two techniques for injecting tumor cells. One technique is freehand, while the other technique is device-assisted using a 3D-printed prototype device. Finally, we describe tumor surveillance with bioluminescent imaging. : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.

Avian influenza viruses (AIVs) have caused a large number of epidemics in domestic and wild birds, and even posed a health challenge to humans. Highly pathogenic AIVs have attracted the most public attention. However, low pathogenic AIVs, including H4, H6, and H10 subtype AIVs, have spread covertly in domestic poultry, without obvious clinical signs. The emergence of human infections with H6 and H10 AIVs and the evidence of seropositivity of H4 AIV in poultry-exposed individuals indicated that these AIVs sporadically infect humans and could cause a potential pandemic. Therefore, a rapid and sensitive diagnostic method to simultaneously detect Eurasian lineage H4, H6, and H10 subtype AIVs is urgently required. Four singleplex real-time RT-PCR (RRT-PCR) assays were established based on carefully designed primers and probes of the conserved regions of the matrix, H4, H6, and H10 genes and combined into a multiplex RRT-PCR method to simultaneously detect H4, H6, and H10 AIVs in one reaction. The detection limit of the multiplex RRT-PCR method was 1–10 copies per reaction when detecting standard plasmids, and showed no cross-reaction against other subtype AIVs and other common avian viruses. Additionally, this method was suitable to detect the AIVs in samples from different sources, the results of which showed high consistency with virus isolation and a commercial influenza detection kit. In summary, this rapid, convenient, and practical multiplex RRT-PCR method could be applied in laboratory testing and clinical screening to detect AIVs.

Vaccines are known to function as the most effective interventional therapeutics for controlling infectious diseases, including polio, smallpox, rabies, tuberculosis, influenza and SARS-CoV-2. Smallpox has been eliminated completely and polio is almost extinct because of vaccines. Rabies vaccines and Bacille Calmette-Guérin (BCG) vaccines could effectively protect humans against respective infections. However, both influenza vaccines and COVID-19 vaccines are unable to eliminate these two infectious diseases of their highly variable antigenic sites in viral proteins. Vaccine effectiveness (VE) could be negatively influenced (i.e., interfered with) by immune imprinting of previous infections or vaccinations, and repeated vaccinations could interfere with VE against infections due to mismatch between vaccine strains and endemic viral strains. Moreover, VE could also be interfered with when more than one kind of vaccine is administrated concomitantly (i.e., co-administrated), suggesting that the VE could be modulated by the vaccine-induced immunity. In this review, we revisit the evidence that support the interfered VE result from immune imprinting or repeated vaccinations in influenza and COVID-19 vaccine, and the interference in co-administration of these two types of vaccines is also discussed. Regarding the development of next-generation COVID-19 vaccines, the researchers should focus on the induction of cross-reactive T-cell responses and naive B-cell responses to overcome negative effects from the immune system itself. The strategy of co-administrating influenza and COVID-19 vaccine needs to be considered more carefully and more clinical data is needed to verify this strategy to be safe and immunogenic.

Abstract Background Rare earth elements (REE) are a group of trace elements that behave geochemically coherently. REE fractionation patterns normalized to reference materials provide a powerful tool for documenting pedogenesis. In-soil processes are particularly difficult to illustrate with respect to contemporary and past climate conditions. In this study, we characterize the rare earth element (REE) contents in bulk soils and respective geochemical fractions (e.g., exchangeable, carbonate-bound, reducible, and oxidizable fractions) and to decipher the relationships between REE geochemistry components and climatic factors across a large-scale northern China transect (NCT). Results Across the NCT, bulk REE concentrations ranged from 55.2 to 241.1 μg g−1 with a main portion in the residual fraction (49–79%), followed by oxidizable fraction (2–40%), reducible fraction (3–22%), carbonate-bound fraction (0.1–16%), and negligible exchangeable fraction. The REE contents of geochemical components (carbonate-bound, reducible, and oxidizable) in topsoils correlated to climate factors (mean annual precipitation, mean annual temperature, potential evaporation, and aridity index (AI)). The normalized abundances to the upper continental crust (UCC) composition show that the middle REE was generally enriched than the light REE and heavy REE in topsoils along the transect. The overall UCC-normalized bulk REE patterns in topsoils and subsoils were similar, characterized by weak negative Ce anomalies and positive Eu anomalies. Conclusions Our data in topsoils and depth profiles collectively suggest that cycling of REE was primarily regulated by abiotic processes in area with AI 0.3. The similar UCC normalized patterns in topsoils suggest that the REE was originated from a common source with limited influences from other sources (e.g., atmospheric dusts and anthropogenic contributions). Our results to some extent provide evidence for climatic influence REE distribution patterns both in topsoils and subsoils across the continental-scale transect. Our investigation gives insights into future studies on vertical REE mobility and its associated biogeochemical pathways.

BackgroundSince the first HIV/AIDS case appeared in 1980s, HIV/AIDS has been the focus of international attention. As a major public health problem, there are epidemiological uncertainties about the future of HIV/AIDS. It is important to monitor the global statistics of HIV/AIDS prevalence, deaths, disability adjusted life years (DALYs), and risk factors for adequate prevention and control.MethodsThe Global Burden of Disease Study 2019 database was used to analyze the burden of HIV/AIDS in 1990–2019. By extracting global, regional, and national data on HIV/AIDS prevalence, deaths, and DALYs, we described the distribution by age and sex, explored the risk factors, and analyzed the trends in HIV/AIDS.ResultsIn 2019, there were 36.85 million HIV/AIDS cases (95% UI: 35.15–38.86 million), 863.84 thousand deaths (95% UI: 78.61–99.60 thousand), and 47.63 million (95% UI: 42.63–55.65 million) DALYs. The global age-standardized HIV/AIDS prevalence, death, and DALY rates were 454.32 (95% UI: 433.76–478.59), 10.72 (95% UI: 9.70–12.39), and 601.49 (95% UI: 536.16–703.92) per 100,000 cases, respectively. In 2019, the global age-standardized HIV/AIDS prevalence, death, and DALY rates increased by 307.26 (95% UI: 304.45–312.63), 4.34 (95% UI: 3.78–4.90), and 221.91 (95% UI: 204.36–239.47) per 100,000 cases, respectively, compared to 1990. Age-standardized prevalence, death, and DALY rates decreased in high sociodemographic index (SDI) areas. High age-standardized rates were observed in low sociodemographic index areas, while low age-standardized rates were observed in high sociodemographic index areas. In 2019, the high age-standardized prevalence, death, and DALY rates were predominant in Southern Sub-Saharan Africa, and global DALYs peaked in 2004 and subsequently decreased. The highest global HIV/AIDS DALYs were in the 40–44 age group. The main risk factors affecting HIV/AIDS DALY rates included behavioral risks, drug use, partner violence, and unsafe sex.ConclusionsHIV/AIDS disease burden and risk factors vary by region, sex, and age. As access to health care increases across countries and treatment for HIV/AIDS infection improves, the HIV/AIDS disease burden is concentrated in areas with low SDIs, particularly in South Africa. Regional differences should be fully considered to target optimal prevention strategies and treatment options based on risk factors.

The intestinal microbiota is an “invisible organ” in the human body, with diverse components and complex interactions. Homeostasis of the intestinal microbiota plays a pivotal role in maintaining the normal physiological process and regulating immune homeostasis. By reviewing more than one hundred related studies concerning HIV infection and intestinal microbiota from 2011 to 2023, we found that human immunodeficiency virus (HIV) infection can induce intestinal microbiota dysbiosis, which not only worsens clinical symptoms but also promotes the occurrence of post-sequelae symptoms and comorbidities. In the early stage of HIV infection, the intestinal mucosal barrier is damaged and a persistent inflammatory response is induced. Mucosal barrier damage and immune injury play a pivotal role in promoting the post-sequelae symptoms caused by HIV infection. This review summarizes the relationship between dysbiosis of the intestinal microbiota and mucosal barrier damage during HIV infection and discusses the potential mechanisms of intestinal barrier damage induced by intestinal microbiota dysbiosis and inflammation. Exploring these molecular mechanisms might provide new ideas to improve the efficacy of HIV treatment and reduce the incidence of post-sequelae symptoms.

Abnormal oxidative stress caused by human immunodeficiency virus (HIV) infection affects viral replication and causes non-acquired immune deficiency syndrome-related complications in infected individuals. The transcription factor NFE2-related factor 2 (NRF2), a key regulator of oxidative stress, responds to abnormal oxidative stress by regulating the expression of NRF2-dependent cytoprotective genes. The present study aimed to determine whether inhibition of oxidative stress could control HIV replication and improve cell survival. In this study, the NRF2 activator, methyl bardoxolone, was used to treat cells for HIV infection. The effects on HIV replication and apoptosis pathways were confirmed by NRF2 activation or knockdown. The results showed that NRF2 activation could block HIV replication in macrophages before the integration phase and inhibited the expression of apoptotic pathways in virus-exposed macrophages. The study presents an unconventional anti-viral strategy of activation antioxidant response for HIV infection blocking.

Aims: To explore the contagiousness and new SARS-CoV-2 mutations in pediatric COVID-19.Methods: This cohort study enrolled all pediatric patients admitted to 8 hospitals in Zhejiang Province of China between 21 January and 29 February 2020, their family members and close-contact classmates. Epidemiological, demographic, clinical and laboratory data were collected. Bioinformatics was used to analyze the features of SARS-CoV-2. Individuals were divided into 3 groups by the first-generation case: Groups 1 (unclear), 2 (adult), and 3 (child). The secondary attack rate (SAR) and R0 were compared among the groups.Results: The infection rate among 211 individuals was 64% (135/211). The SAR in Groups 2 and 3 was 71% (73/103) and 3% (1/30), respectively; the median R0 in Groups 2 and 3 was 2 (range: 1-8) and 0 (range: 0-1), respectively. Compared with adult cases, the SAR and R0 of pediatric cases were significantly lower (p

Abstract Background The H9N2 subtype of avian influenza virus (AIV) has become the most widespread subtype of AIV among birds in Asia, which threatens the poultry industry and human health. Therefore, it is important to establish methods for the rapid diagnosis and continuous surveillance of H9N2 subtype AIV. Methods In this study, an antigen-capture enzyme-linked immunosorbent assay (AC-ELISA) and a colloidal gold immunochromatographic test (ICT) strip using monoclonal antibodies (MAbs) 3G4 and 2G7 were established to detect H9N2 subtype AIV. Results The AC-ELISA method and ICT strip can detect H9N2 subtype AIV quickly, and do not cross-react with other subtype AIVs or other viruses. The detection limit of AC-ELISA was a hemagglutinin (HA) titer of 4 for H9N2 subtype AIV per 100 μl sample, and the limit of detection of the HA protein of AIV H9N2 was 31.5 ng/ml. The ICT strip detection limit was an HA titer of 4 for H9N2 subtype AIV per 100 μl sample. Moreover, both detection methods exhibited good reproducibility and repeatability, with coefficients of variation

Objectives: The continuous evolution of highly pathogenic H5N6 avian influenza viruses (AIVs) causes outbreaks in wildfowl and poultry, and occasional human infections. The aim of this study was to better understand the genetic relationships between these H5N6 AIVs from eastern China and other AIVs. Methods: In 2016, 1623 cloacal swabs were sampled from poultry in 18 LPMs in eastern China, and subsequently characterized systematically using gene sequencing, phylogenetic studies, and antigenic analysis. In addition, their pathogenicity in mammals was studied in BALB/c mice, which were inoculated with viruses, with survival rate and body weight recorded daily for 14 days. Results: In total, 56 H5N6 AIVs were isolated in eastern China and five representative isolates were selected for further study. In our study, the H5N6 AIVs clustered into clade 2.3.4.4, Group C, and their six internal segments were derived from H6N6 and H9N2 viruses, or both, suggesting extensive reassortant among H5N6 AIVs with other subtypes. These H5N6 viruses could replicate in the lungs without prior adaptation, and exhibited slight-to-moderate virulence in mice. Conclusions: The continuous circulation of these novel H5N6 viruses suggests the importance of persistent surveillance of H5N6 AIVs in poultry.

Purine nucleoside phosphorylase (PNP) enables the breakdown and recycling of guanine nucleosides. PNP insufficiency in humans is paradoxically associated with both immunodeficiency and autoimmunity, but the mechanistic basis for these outcomes is incompletely understood. Here, we identify two immune lineage-dependent consequences of PNP inactivation dictated by distinct gene interactions. During T cell development, PNP inactivation is synthetically lethal with downregulation of the dNTP triphosphohydrolase SAMHD1. This interaction requires deoxycytidine kinase activity and is antagonized by microenvironmental deoxycytidine. In B lymphocytes and macrophages, PNP regulates Toll-like receptor 7 signaling by controlling the levels of its (deoxy)guanosine nucleoside ligands. Overriding this regulatory mechanism promotes germinal center formation in the absence of exogenous antigen and accelerates disease in a mouse model of autoimmunity. This work reveals that one purine metabolism gene protects against immunodeficiency and autoimmunity via independent mechanisms operating in distinct immune lineages and identifies PNP as a potentially novel metabolic immune checkpoint.

Abstract Background In 2020, a new coronavirus, SARS-CoV-2, quickly spread worldwide within a few months. Although coronaviruses typically infect the upper or lower respiratory tract, the virus RNA can be detected in plasma. The risk of transmitting coronavirus via transfusion of blood products remains. As more asymptomatic infections are identified in COVID-19 cases, blood safety has become particularly important. Methylene blue (MB) photochemical technology has been proven to inactivate lipid-enveloped viruses with high efficiency and safety. The present study aimed to investigate the SARS-CoV-2 inactivation effects of MB in plasma. Methods The SARS-CoV-2 virus strain was isolated from Zhejiang University. The live virus was harvested from cultured VERO-E6 cells, and mixed with MB in plasma. The MB final concentrations were 0, 1, 2, and 4 μM. The “BX-1 AIDS treatment instrument” was used at room temperature, the illumination adjusted to 55,000 ± 0.5 million Lux, and the plasma was irradiated for 0, 2, 5, 10, 20, and 40 mins using light at a single wavelength of 630 nm. Virus load changes were measured using quantitative reverse transcription- PCR. Results BX-1 could effectively eliminate SARS-CoV-2 within 2 mins in plasma, and the virus titer declined to 4.5 log10 TCID50 (median tissue culture infectious dose)/mL. Conclusion BX-1 is based on MB photochemical technology, which was designed to inactivate HIV-1 virus in plasma. It was proven to be safe and reliable in clinical trials of HIV treatment. In this study, we showed that BX-1 could also be applied to inactivate SARS-CoV-2. During the current outbreak, this technique it has great potential for ensuring the safety of blood transfusions, for plasma transfusion therapy in recovering patients, and for preparing inactivated vaccines.

Abstract Background Highly pathogenic influenza A (H5N8) viruses have caused several worldwide outbreaks in birds and are of potential risk to humans. Thus, a specific, rapid and sensitive method for detection is urgently needed. Methods In the present study, TaqMan minor groove binder probes and multiplex real-time RT-PCR primers were designed to target the H5 hemagglutinin and N8 neuraminidase genes. A total of 38 strains of avian influenza viruses and other viruses were selected to test the performance of the assay. Results The results showed that only H5 and N8 avian influenza viruses yielded a positive signal, while all other subtypes avian influenza viruses and other viruses were negative. High specificity, repeatability, and sensitivity were achieved, with a detection limit of 10 copies per reaction. Conclusions The developed assay could be a powerful tool for rapid detection of H5N8 influenza viruses in the future.

ABSTRACTThe mutations in the SARS-CoV-2 virus genome during COVID-19 dissemination are unclear. In 788 COVID-19 patients from Zhejiang province, we observed decreased rate of severe/critical cases compared with patients in Wuhan. For mechanisms exploration, we isolated one strain of SARS-CoV-2 (ZJ01) from a mild COVID-19 patient. Thirty-five specific gene mutations were identified. Phylogenetic and relative synonymous codon usage analysis suggested that ZJ01 may be a potential evolutionary branch of SARS-CoV-2. We classified 54 global virus strains based on the base (C or T) at positions 8824 and 28247 while ZJ01 has T at both sites. The prediction of the Furin cleavage site (FCS) and sequence alignment indicated that the FCS may be an important site of coronavirus evolution. ZJ01 mutations identified near the FCS (F1-2) caused changes in the structure and electrostatic distribution of the S surface protein, further affecting the binding capacity of Furin. Single-cell sequencing and ACE2-Furin co-expression results confirmed that the Furin expression was especially higher in glands, liver, kidneys, and colon. The evolutionary pattern of SARS-CoV-2 towards FCS formation may result in its clinical symptom becoming closer to HKU-1 and OC43 caused mild flu-like symptoms, further showing its potential in differentiating into mild COVID-19 subtypes.

ABSTRACTThe H7N9 viruses have been circulating for six years. The insertion of a polybasic cleavage site in the haemagglutinin (HA) protein of H7N9 has resulted in the emergence of a highly pathogenic (HP) avian influenza virus. Currently, there are limited studies on neutralizing monoclonal antibodies(mAbs) against HP H7N9 AIVs. In this study, mice were immunized with inactivated H7N9 vaccine of A/ZJU01/PR8/2013 to produce murine mAbs. Finally, two murine mAbs against the HA of low pathogenic (LP) virus were produced and characterized. Characterization included determining mAbs binding breadth and affinity, in vitro neutralization capacity, and potential in vivo protection. Two of these mAbs, 1H10 and 2D1, have been identified to have therapeutic and prophylactic efficacy against the HP strain in mouse passive transfer-viral challenge experiments. The mAb 1H10 was most efficacious, even if the treatment-time was as late as 72 h post-infection, or the therapeutic dose was as low as 1 mg/kg; and it was confirmed to have haemagglutination inhibition and neutralizing activity on both LP-and HP-H7N9 strains. Further study indicated that the protection provided by 2D1 was mediated by antibody-dependent cellular cytotoxicity. The mAbs described here provide promising results and merit further development into potential antiviral therapeutics for H7N9 infection.

There is a worldwide pandemic of Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection; yet our understanding remains limited on the characteristic of antibodies, especially for dynamic long-term tracking. Sequential serum samples were collected up to 416 days post onset of symptoms (POS) from 102 patients who were hospitalized with coronavirus disease 2019 (COVID-19). Immunoglobulin (Ig)G, IgM, and IgA levels targeting SARS-CoV-2 spike 1 receptor-binding domain (S1-RBD), spike 2 extracellular domain (S2-ECD), and nucleocapsid protein (N) were quantified as well as neutralizing activity. We were pleasantly surprised to find that the antibody remained detective and effective for more than a year POS. We also found the varied reactions of different antibodies as time passed: N-IgA rose most rapidly in the early stage of infection, while S2-IgG was present at a high level in the long time of observation. This study described the long traceable antibody response of the COVID-19 and offered hints about targets to screen for postinfectious immunity and for vaccination development of SARS-CoV-2.

Abstract Background The major infectious diseases of hepatitis B has constituted an acute public health challenge in China. An effective and affordable HBV control model is urgently needed. A national project of Community-based Collaborative Innovation HBV (CCI-HBV) demonstration areas has optimized the existing community healthcare resources and obtained initial results in HBV control. Methods Based on the existing community healthcare network, CCI-HBV project combined the community health management and health contract signing service for long-staying residents in hepatitis B screening. Moreover, HBV field research strategy was popularized in CCI-HBV areas. After screening, patients with seropositive results were enrolled in corresponding cohorts and received treatment at an early stage. And the uninfected people received medical supports including health education through new media, behavior intervention and HBV vaccinations. In this process, a cloud-based National Information Platform (NIP) was established to collect and store residents’ epidemiological data. In addition, a special quality control team was set up for CCI project. Results After two rounds of screening, HBsAg positive rate dropped from 5.05% (with 5,173,003 people screened) to 4.57% (with 3,819,675 people screened), while the rate of new HBV infections was 0.28 per 100 person-years in the fixed cohorts of 2,584,322 people. The quality control team completed PPS sampling simultaneously and established the serum sample database with 2,800,000 serum samples for unified testing. Conclusions CCI-HBV project has established a large-scale field research to conduct whole-population screening and intervention. We analyzed the HBsAg prevalence and new infection rate of HBV in the fixed population for the epidemic trend and intervention effect. The purpose of CCI-HBV project is to establish and evaluate a practical model of grid management and field strategy, to realize the new goal to control hepatitis B in China. To provide policymakers with a feasible model, our results are directly applicable. Trial registration The project was funded by the Major Projects of Science Research for the 11th and 12th five-year plans of China, entitled “The prevention and control of AIDS, viral hepatitis and other major infectious diseases”, Grant Nos. 2009ZX10004901, 2011ZX10004901, 2013ZX10004904, 2014ZX10004007 and 2014ZX10004008.

Viral DNA integrated in host cells is a major barrier to completely curing HIV-1. However, genome editing using the recently developed technique of clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 has the potential to eradicate HIV-1. The present study aimed to use a lentiviral vector-based CRISPR/Cas9 system combined with dual-small/single guide RNAs (sgRNAs) to attack HIV-1 DNA in the latency reactivation model J-Lat 10.6 cell line and to assess off-target effects using whole-genome sequencing (WGS). We designed 12 sgRNAs targeting HIV-1 DNA, and selected high-efficiency sgRNAs for further pairwise combinations after a preliminary evaluation of the editing efficiency. Three combinations of dual-sgRNAs/Cas9 with high editing efficiency were screened successfully from multiple combinations. Among these combinations, the incidences of insertions and deletions in the sgRNA-targeted regions reached 76% and above, and no credible off-target sites were detected using WGS. The results provided comprehensive basic experimental evidence and methodological recommendations for future personalized HIV-1 treatment using CRISPR/Cas9 genome editing technology.

Background Elite controller refers to a patient with human immunodeficiency virus infection with an undetected viral load in the absence of highly active antiretroviral therapy. Studies on gene expression and regulation in these individuals are limited but significant, and have helped researchers and clinicians to understand the interrelationships between HIV and its host. Methods We collected CD4 T-cell samples from two elite controllers (ECs), two HIV-positive infected patients (HPs), and two healthy controls (HCs) to perform second-generation transcriptome sequencing. Using the Cufflinks software, we calculated the Fragments Per Kilobase of transcript per Million fragments mapped (FPKM) and identified differentially expressed (DE) mRNAs and long non-coding RNAs (lncRNAs), with corrected P value < 0.05 (based on a false discovery rate (FDR) < 0.05). We then constructed a protein-protein interaction network using cytoHubba and a long non-coding RNA-mRNA co-expression network based on the Pearson correlation coefficient. Results In total, 1109 linear correlations of DE lncRNAs targeting DE mRNAs were found and several interesting interactions were identified as being associated with viral infections and immune responses within the networks based on these correlations. Among these lncRNA-mRNA relationships, hub mRNAs including HDAC6, MAPK8, MAPK9, ATM and their corresponding annotated co-expressed lncRNAs presented strong correlations with the MAPK-NF-kappa B pathway, which plays a role in the reactivation and replication of the virus. Conclusions Using RNA-sequencing, we systematically analyzed the expression profiles of lncRNAs and mRNAs from CD4+ T cells from ECs, HPs, and HCs, and constructed a co-expression network based on the relationships among DE transcripts and database annotations. This was the first study to examine gene transcription in elite controllers and to study their functional relationships. Our results provide a reference for subsequent functional verification at the molecular or cellular level.

Abstract Background The H6N1 subtype of avian influenza viruses (AIVs) can infect people with an influenza-like illness; the H6N1 viruses possess the ability for zoonotic transmission from avians into mammals, and possibly pose a threat to human health. Methods In 2017, live poultry markets (LPMs) in Zhejiang Province were surveyed for AIVs. To better understand the genetic relationships between these strains from Eastern China and other AIVs, all gene segments of these strains were sequenced and compared with sequences available in GenBank. In this study, we analyzed the receptor-binding specificity, antigenic characteristics, and pathogenicity of these two H6N1 viruses. Results In 2017, two H6N1 AIVs were isolated from chickens during surveillance for AIVs in LPMs in Eastern China. Phylogenetic analysis showed that these strains shared genetic characteristics from H6, H10, H1, and H4 AIVs found in ducks and wild birds in East Asia. These AIV strains were able to replicate in mice without prior adaptation. Conclusions In this study, we report the discovery of new strains of H6N1 viruses from chickens with novel gene reassortments. Our results suggest that these chickens play an important role generating novel reassortments in AIVs, and emphasize the need for continued surveillance of AIV strains circulating in poultry.

Background: To determine the safety and immunogenicity of a novel recombinant adenovirus type 5 vector based Ebola virus disease vaccine (Ad5-EBOV) in Africans in China. Methods: A phase 1, dose-escalation, open-label trial was conducted. 61 healthy Africans were sequentially enrolled, with 31 participants receiving one shot intramuscular injection and 30 participants receiving a double-shot regimen. Primary and secondary end points related to safety and immunogenicity were assessed within 28 d after vaccination. This study was registered with ClinicalTrials.gov (NCT02401373). Results: Ad5-EBOV is well tolerated and no adverse reaction of grade 3 or above was observed. 53 (86.89%) participants reported at least one adverse reaction within 28 d of vaccination. The most common reaction was fever and the mild pain at injection site, and there were no significant difference between these 2 groups. Ebola glycoprotein-specific antibodies appeared in all 61 participants and antibodies titers peaked after 28 d of vaccination. The geometric mean titres (GMTs) were similar between these 2 groups (1919.01 vs 1684.70 P = 0.5562). The glycoprotein-specific T-cell responses rapidly peaked after 14 d of vaccination and then decreased, however, the percentage of subjects with responses were much higher in the high-dose group (60.00% vs 9.68%, P = 0.0014). Pre-existing Ad5 neutralizing antibodies could significantly dampen the specific humoral immune response and cellular response to the vaccine. Conclusion: The application of Ad5-EBOV demonstrated safe in Africans in China and a specific GP antibody and T-cell response could occur 14 d after the first immunization. This acceptable safety profile provides a reliable basis to proceed with trials in Africa.

Abstract Objective To identify factors associated with recent HIV infection among men who have sex with men (MSM) in Huzhou City, Eastern China, who seek sexual partners via the internet. Methods Adult members of Tencent QQ instant messaging software, who completed a face-to-face interview questionnaire and serological HIV testing at baseline, were enrolled into this prospective cohort study. Questionnaires and serological testing were also completed at 6 and 12 months. Cox proportional hazards was used for initial bivariate analyses of differences between participants with or without recent HIV infection, and for subsequent forward stepwise multiple regression of statistically significant variables. Results Out of 415 baseline participants, 283 completed the 12-month follow-up (25 with recent HIV infection and 258 without; recent HIV infection rate, 8.20 per 100 person-years). Recent HIV infection was shown to be significantly associated with ≤ high school education (versus college/university), registered residence outside Huzhou City (versus within Huzhou City), having ≥ two male sexual partners, and syphilis infection. Conclusions Improving education levels, treating syphilis promptly, and reducing the number of male sexual partners may reduce HIV transmission among MSM in Eastern China.

H5N6 avian influenza viruses (AIVs) can cause severe pneumonia and death in humans. However, the molecular determinants of H5N6 influenza virus mammalian adaption are still unclear. Three amino acid substitutions (HA A150V, PA A343T, PB2 E627K) are observed in H5N6 virus A/duck/Zhejiang/6D2/2013 (6D2) in lung-to-lung passage in mice. These substitutions are crucial to the pathogenicity of mouse-adapted virus. In this study, we investigated the contribution of each amino acid substitution in the virus by reverse genetics. The results demonstrate that HA A150V greatly altered the receptor binding preference of 6D2. Virus bearing this substitution acquired increased mortality than mice infected with wild-type 6D2. The PA A343T substitution mildly enhanced viral polymerase activity but the reduced survival rate in mice indicates this substitution may change the immunoreaction of the host. The well-known PB2 E627K substitution increased eight folds the relative polymerase activity compared to PA A343T and resulted in 100% death rate in mice. In addition, we show that PA A343T dramatically exacerbates the effect of PB2 E627K on viral polymerase activity; when combined, these two substitutions work synergistically. However, HA A150V and PA A343T seemed to attenuate PB2 E627K in vivo, which implies the difference between mixed viral populations under natural condition and single population under experiment, specialization and cooperation in quasispecies is important in the process of adaption. This study suggests that HA A150V, PA A343T, and PB2 E627K are crucial in the adaption and increased pathogenicity of H5N6 in mammalian hosts.

Background. Butyrylcholinesterase (BChE) is synthesized mainly in the liver and an important marker in many infectious/inflammatory diseases, but its role in acquired immunodeficiency syndrome (AIDS) patients is not clear. We wished to ascertain if BChE level is associated with the progression/prognosis of AIDS patients. Methods. BChE levels (in U/L) were measured in 505 patients;

Background/Aims: The main obstacle to cure HIV infection is the existence of long-lasting latent reservoirs. Many efforts have been made to understand basal mechanisms of HIV-1 latency, in which miRNAs play an important role. However, integrated analysis of miRNA and mRNA expression in HIV-1 latency is lacking. Methods and Results: Global miRNA and mRNA expression was determined by microarrays and quantitative reverse transcription PCR in well-characterized HIV-1 latently and actively infected cells, respectively. Interactions of miRNA-mRNA, mRNA-mRNA, and transcription factor-miRNA pairs were assembled into the function network. Our results show that transcription regulation related genes were mostly enriched in HIV-1 latently infected cells. Gene set enrichment analysis revealed nuclear transport related pathways were up-regulated in the latency group. Network dynamic analysis highlighted many gene-pairs sharing the largest changes in different HIV-1 infection state. 83.33% miRNA-target pairs were validated against database, and RHOB related genes constitute the interface between HIV-1 latency and replication state. Conclusion: We show for the first time a joint miRNA and mRNA expression profile related to a HIV-1 latency phenotype, outline a dynamic network of potential regulators involving in HIV-1 latency or replication state, and gain new insights into the source messages for affecting HIV-1 latency.

Background: Co-infection of hepatitis virus is common in human immunodeficiency virus (HIV) infected adults in China. But little is known about hepatitis virus co-infection in pediatric HIV-infected subjects. The study aimed to investigate the impact of hepatitis B virus (HBV) and/or hepatitis C virus (HCV) co-infection and highly active antiretroviral therapy (HAART) on liver function of pediatric HIV-infected subjects. Materials and Methods: A cohort study including 101 pediatric HIV-infected subjects with HBV/HCV co-infection and 44 pediatric comparators with HIV mono-infection was carried out in Henan Province of China from September 2011 to September 2012. All patients received HAART for 1-year. HBV and HCV infection was determined by antibody tests. HIV RNA load, CD4 + T-cell counts and liver function were determined before and after HAART. The Student′s t-test or a one-way ANOVA was used for normally distributed values and A Mann-Whitney U-test was performed for values without normal distribution using SPSS statistical package 18.0 (SPSS Inc.). Results: After HAART for 1-year, the median levels of viral load were decreased to lower limit of detection in 90.34% pediatric HIV-infected subjects with/without HBV/HCV co-infection (P < 0.001), and CD4 + T-cell counts increased significantly (P < 0.001). Compared with the pre-HAART, mean level of alanine aminotransferase (ALT) in each group had a significant increase after HAART (P < 0.01). The mean levels of ALT and aspartate aminotransferase (AST) in nevirapine (NVP) based HAART group increased significantly after HAART (P < 0.01). Mean change values of ALT and AST were significantly higher in the NVP based regimen group than in the efavirenz (EFV) based regimen group (P < 0.01). For HIV/HBV/HCV co-infected patients, mean change values of ALT and AST in NVP-based HAART group was significantly higher than that in EFV-based HAART group (P < 0.01). Conclusion: Highly active antiretroviral therapy can damage liver function in pediatric HIV-infected subjects, especially in those with HBV/HCV co-infection. NVP was more harmful to liver function of pediatric HIV-infected subjects than EFV.

Currently, there are three algorithms for screening of syphilis: traditional algorithm, reverse algorithm and European Centre for Disease Prevention and Control (ECDC) algorithm. To date, there is not a generally recognized diagnostic algorithm. When syphilis meets HIV, the situation is even more complex. To evaluate their screening performance and impact on the seroprevalence of syphilis in HIV-infected individuals, we conducted a cross-sectional study included 865 serum samples from HIV-infected patients in a tertiary hospital. Every sample (one per patient) was tested with toluidine red unheated serum test (TRUST), T. pallidum particle agglutination assay (TPPA), and Treponema pallidum enzyme immunoassay (TP-EIA) according to the manufacturer's instructions. The results of syphilis serological testing were interpreted following different algorithms respectively. We directly compared the traditional syphilis screening algorithm with the reverse syphilis screening algorithm in this unique population. The reverse algorithm achieved remarkable higher seroprevalence of syphilis than the traditional algorithm (24.9% vs. 14.2%, p < 0.0001). Compared to the reverse algorithm, the traditional algorithm also had a missed serodiagnosis rate of 42.8%. The total percentages of agreement and corresponding kappa values of tradition and ECDC algorithm compared with those of reverse algorithm were as follows: 89.4%,0.668; 99.8%, 0.994. There was a very good strength of agreement between the reverse and the ECDC algorithm. Our results supported the reverse (or ECDC) algorithm in screening of syphilis in HIV-infected populations. In addition, our study demonstrated that screening of HIV-populations using different algorithms may result in a statistically different seroprevalence of syphilis.

Recently, novel variants of H5 highly pathogenic avian influenza viruses (AIVs) have been frequently isolated from poultry and wild birds in Asia, Europe and North America. Live poultry markets (LPMs) play an important role in the dissemination of influenza viruses. Four H5N2 AIVs were isolated from poultry during surveillance of AIVs in LPMs in Eastern China, in 2015. Whole-genome sequencing, combined with phylogenetic and antigenic analyses were performed to characterize these viruses. These H5N2 viruses had undergone extensive reassortment resulting in two genetic groups of viruses in poultry. These viruses exhibited slightly pathogenicity in mice, and replicated without prior adaptation. The continued circulation of these novel H5N2 viruses may represent a threat to human health.

Domestic ducks are natural reservoirs of avian influenza viruses and serve as reassortant hosts for new virus subtypes. We isolated 2 novel influenza A(H5N8) viruses from domestic ducks in eastern China, sequenced their genomes, and tested their pathogenicity in chickens and mice. Circulation of these viruses may pose health risks for humans.

The HIV epidemic in men who have sex with men (MSM) continues to grow in most countries. However, the phylodynamic and virological differences among HIV-1 strains circulating in MSM and other populations are not well characterized.Nearly full-length genomes (NFLGs) of the HIV-1 CRF01_AE were obtained from the Los Alamos HIV database. Phylogenetic analyses were conducted using the NFLG, gag, pol and env genes, using the maximum likelihood method. Selection pressure analyses at the codon level were performed for each gene in the phylogenetic clusters using PAML.Sequences isolated from MSM in China clustered in Clusters 1 (92.5%) and 2 (85.71%). The major risk factor for Cluster 3 was heterosexual transmission (62.16%). The ratio of non-synonymous to synonymous substitutions in the env gene (0.7-0.75) was higher than the gag (0.26-0.34) or pol (0.21-0.26) genes. In env gene, Cluster 1 (4.56×10(-3) subs/site/year) and 2 (6.01×10(-3) subs/site/year) had higher evolutionary rates than Cluster 3 (1.14×10(-3) subs/site/year). Positive selection affected 4.2-6.58% of the amino acid sites in the env gene. Two sites (HXB2:136 and 316) evolved similarly in Clusters 1 and 2, but not Cluster 3.The HIV-1 CRF01_AE in MSM is evolving differently than in other populations.

BACKGROUND: Female sex workers (FSWs) play an important role in transmitting HIV and syphilis from high-risk groups to the general population. However, the trends in HIV and syphilis epidemics in Chinese FSWs in the period after 2000 are unclear to date. METHODS: The Preferred Reporting Items for Systematic Reviews and Meta-Analyses Statement was followed. Seven databases were searched for published peer-reviewed articles. The incidence of HIV and syphilis in FSWs in different time periods, provinces and workplaces in China were separately pooled by meta-analysis. Correlation analysis was conducted between HIV and syphilis incidence and study time, respectively. RESULTS: After 1,662 articles were screened, 190 published papers were included in the final analysis. Estimated HIV prevalence was 0.284% (95% CI: 0.080-0.488%) in the period 2000-2002, 0.211% (95% CI: 0.149-0.273%) in 2003-2005, 0.242% (95% CI: 0.190-0.294%) in 2006-2008 and 0.041% (95% CI: 0.024-0.058%) in 2009-2011. The corresponding syphilis prevalence was 9.669% (95% CI: 7.810-11.529%), 4.970% (95% CI: 4.384-5.556%), 4.404% (95% CI: 4.032-4.775%) and 3.169% (95% CI: 2.738-3.600%), respectively. Spearman rank correlation coefficients were -0.165 (p = 0.002) between HIV prevalence and study time, and -0.209 (p = 0.000) between syphilis prevalence and study time. The combined HIV prevalence was 0.318% (95% CI: 0.156-0.479%) in medium and high-tier workplaces and 0.393% (95% CI: 0.176-0.610%) in low-tier workplaces. The corresponding syphilis prevalence was 3.216% (95% CI: 2.192-4.240%) and 13.817% (95% CI: 10.589-17.044%), respectively. CONCLUSIONS: Our data suggested a decline in HIV and syphilis epidemics in FSWs in China on a national level during the study period (2000-2011). FSWs in low-tier workplaces should be given more attention in the future to ensure they are included in prevention programs for HIV and sexually transmitted diseases.

INTRODUCTION: Pancreatic ductal adenocarcinoma (PDAC) is projected to rise to the second leading cause of U.S. cancer-related deaths by 2020. Novel therapeutic targets are desperately needed. MicroRNAs (miRs) are small noncoding RNAs that function by suppressing gene expression and are dysregulated in cancer. miR-21 is overexpressed in PDAC tumor cells (TC) and is associated with decreased survival, chemoresistance and invasion. Dysregulation of miR regulatory networks in PDAC tumor-associated fibroblasts (TAFs) have not been previously described. In this study, we show that miR-21 expression in TAFs promotes TC invasion. METHODS: In-situ hybridization for miR-21 was performed on the 153 PDAC patient UCLA tissue microarray and 23 patient-matched lymph node metastases. Stromal and TC histoscores were correlated with clinicopathologic parameters by univariate and multivariate Cox regression. miR-21 positive cells were further characterized by immunofluorescence for mesenchymal/epithelial markers. For in vitro studies, TAFs were isolated from freshly resected human PDAC tumors by the outgrowth method. miR-21 was overexpressed/inhibited in fibroblasts and then co-cultured with GFP-MiaPaCa TCs to assess TC invasion in modified Boyden chambers. RESULTS: miR-21 was upregulated in TAFs of 78% of tumors, and high miR-21 significantly correlated with decreased overall survival (P = 0.04). Stromal miR-21 expression was also significantly associated with lymph node invasion (P = 0.004), suggesting that it is driving TC spread. Co-immunofluorescence revealed that miR-21 colocalized with peritumoral fibroblasts expressing α-smooth muscle actin. Moreover, expression of miR-21 in primary TAFs correlated with miR-21 in TAFs from patient-matched LN metastases; evidence that PDAC tumor cells induce TAFs to express miR-21. miR-21 expression in TAFs and TCs promotes invasion of TCs and is inhibited with anti-miR-21. CONCLUSIONS: miR-21 expression in PDAC TAFs is associated with decreased overall survival and promotes TC invasion. Anti-miR-21 may represent a novel therapeutic strategy for dual targeting of both tumor and stroma in PDAC.

Development of an effective HIV management is enticed by the fact that long-term non-progressors (LTNP) restrict viral replication spontaneously, but is hindered by HIV-1 latency. Given that the most overlapping characteristics found between HIV-1 LTNP and latency, detailed analysis of the difference would disclose the essentials of latency. In this study, microarray data from our previous study was combined with HIV-1 latency and LTNP data obtained from NCBI GEO database. Principal variance component analysis and hierarchical clustering verified the removal of batch effect across platform. The analysis revealed a total of 456 differential expressed genes with >2-fold change and B-statistic >0. Bayesian inference was used to reconstitute the transcriptional network of HIV-1 latency or LTNP, respectively. Gene regulation was reprogrammed under different disease condition. By network interference, KPNA2 and ATP5G3 were identified as the hubs in latency network which mediate nuclear export and RNA processing. These data offer comparative insights into HIV-1 latency, which will facilitate the understanding of the genetic basis of HIV-1 latency in vivo and serve as a clue for future treatment dealing with key targets in HIV-1 latency.