Prediction of prognostic and immune therapy response in lung adenocarcinoma based on MHC-I-related genes.

OBJECTIVES: The study investigated the prognostic and immune predictive potential of major histocompatibility complex class I (MHC-I) in lung adenocarcinoma (LUAD). MATERIALS AND METHODS: With The Cancer Genome Atlas (TCGA)-LUAD and Gene Expression Omnibus datasets (GSE26939, GSE72094) as the training and validation sets, respectively, we used Cox regression analysis to construct a prognostic model, and verified independence of riskscore. The predictive capacity of the model was assessed in both sets using the receiver operating characteristic curve and Kaplan-Meier survival curves. Immune analysis was performed by using ssGSEA. Additionally, immune checkpoint blockade therapy was assessed by using immunophenoscore, Tumor Immune Dysfunction and Exclusion score. Based on the cMAP database, effective small molecule compounds were predicted. RESULTS: A prognostic model was established based on 8 MHC-I-related genes, and the predictive capacity of the model was accurate. Immune analysis results revealed that patients classified as high-risk had lower levels of immune cell infiltration and impaired immune function. The low-risk group possessed a better response to immune checkpoint blockade therapy. Theobromine and pravastatin were identified as having great potential in improving the prognosis of LUAD. CONCLUSION: Overall, the study revealed MHC-I-related molecular prognostic biomarkers as robust indicators for LUAD prognosis and immune therapy response.

Differences in cytokine and chemokine profiles in cerebrospinal fluid caused by the etiology of cryptococcal meningitis and tuberculous meningitis in HIV patients.

The roles of cytokines and chemokines in HIV-associated cryptococcal meningitis (HCM) and HIV-associated tuberculous meningitis (HTBM) are debatable. In sum, 34 HIV-infected patients without meningitis, 44 HCM patients and 27 HTBM patients were enrolled for study. The concentrations of 22 cytokines/chemokines in cerebrospinal fluid (CSF) were assayed at admission. Principal component analysis (PCA), Pearson's and logistic regression analyses were used to assess the role of cytokines/chemokines in HCM and HTBM. We found the levels of T helper (Th)17, Th1 [interleukin (IL)-12p40, interferon (IFN)-γ, tumor necrosis factor (TNF)-α and TNF-ß and Th2 (IL-2/4/5/6/10)] cytokines were elevated in patients with meningitis compared with those in HIV-infected patients without central nervous system (CNS) infection. Furthermore, the IL-1Ra, IL-12p40, IL-17α and monocyte chemotactic protein-1 (MCP-1) levels were higher in HCM patients, while the IFN-γ, regulated upon activation, normal T cell expressed and secreted (RANTES) and interferon-inducible protein-10 (IP)-10 levels were higher in HTBM patients. Elevated CSF concentrations of IL-17a, TNF-ß, IL-5, IL-12p40 and IL-1Rα were closely related to meningitis, but elevated IP-10, MCP-1, RANTES and IFN-γ levels and CSF white blood cells (WBCs) were protective factors against HCM. Our study suggested that HIV-infected patients with low CSF WBCs have a high risk of HCM. Th1, Th2 and Th17 cytokines/chemokines mediate differences in the pathogenesis of HCM and TBM. Overexpressed proinflammatory MCP-1, RANTES, IFN-γ and IP-10 in CSF are protective factors against HCM but not HTBM.

Differential expression of innate immunity regulation genes in chronic HIV-1 infected adults.

OBJECTIVE: Chronic activation of the innate immune system plays a central role in HIV-1 disease progression. Negative regulation of innate immunity is critical in preventing the effects of this excessive activation; however, the molecules involved in this process remain to be identified. In this study, we compared the expression of immune regulation genes between HIV-1 infected individuals and healthy control participants to identify genes involved in the regulation of innate immunity in HIV-1 infection. METHODS: We conducted gene expression analysis of a series of immune regulatory genes in viremic treatment-naïve HIV-positive donors, patients receiving highly active antiretroviral therapy (HAART) and HIV-negative healthy control participants. Reverse transcription-quantitative PCR (RT-qPCR) was conducted to determine the expression levels of genes in peripheral blood mononuclear cells isolated from all participants. The spearman correlation test and linear regression analysis were performed to evaluate the correlation between gene expression level and viral load. RESULTS: The following differentially expressed genes were identified: A20, CYLD, DDX24, MARCH5, MKRN2, PTP1B, RNF125, S1PR1, SOCS1, IFI35, RBCK1, TTLL12 and USP18. The three most differentially expressed genes were A20, S1PR1, and USP18. USP18 correlated positively with viral load. CONCLUSION: Thirteen immune regulation genes were identified in comparisons of viremic treatment-naïve HIV-positive donors, HAART-treated patients and healthy control participants, indicating the potential of these genes as therapeutic targets.

Molecular characterization of H10 subtype avian influenza viruses isolated from poultry in Eastern China.

In recent years, avian-origin H10 influenza viruses have proved capable of infecting human beings, and they pose a potential public health threat. Seven H10 avian influenza viruses (AIVs), H10N3 (n = 2), H10N7 (n = 1), and H10N8 (n = 4), were isolated from chickens in Zhejiang Province, Eastern China, during surveillance of AIVs in live poultry markets in 2016 and 2017. Phylogenetic analysis indicated that Zhejiang H10 strains received gene segments from H10, H3, and H7 viruses from birds in East Asia. Animal inoculation tests showed that these isolates have low pathogenicity in mice and can replicate in this species. Our findings suggest these H10 AIVs have the ability to adapt to chicken or other poultry, and highlight the need of long-term surveillance.

Chemokine and Cytokine Cascade Caused by Skewing of the Th1-Th2 Balance Is Associated with High Intracranial Pressure in HIV-Associated Cryptococcal Meningitis.

PURPOSE: Serum cytokines/chemokines play important roles in cryptococcal meningitis, but it is unclear whether cytokines/chemokines in cerebrospinal fluid (CSF) contribute to high intracranial pressure (HICP) in HIV-associated cryptococcal meningitis (HCM). METHODS: CSF cytokines/chemokines were assayed in 17 HIV-uninfected patients, 26 HIV-infected patients without CNS infection, and 39 HCM patients at admission. Principal component analysis and correlation and logistic regression analyses were used to assess the relationships between these parameters. RESULTS: The CSF Th1, Th2, and macrophage cytokines showed an obvious increase in HCM patients as compared to the HIV-uninfected patients and HIV-infected patients without CNS infection. CSF IL-6, GM-CSF, and IL-8 were positively correlated with CSF fungal burden. Serum CD4 count, CSF Th1 cytokines (TNF-α, TNF-ß, IL-12, IL-1ß, IL-12, IL-1α, TNF-α, TNF-ß, IL-12, IL-1γ, and IL-12) and Th2 cytokines (IL-4 and IL-10) contribute to HICP. CONCLUSION: Overall, the present findings indicated that both pro- and anti-inflammatory cytokines of Th1, Th2, and macrophage origin contributed to the development of HCM. Specifically, the chemokine and cytokine cascade caused by skewing of the Th1-Th2 balance and reduced CD4 count were found to be important contributors to HICP. Summary. Our research suggested that chemokine and cytokine cascade caused by skewing of the Th1-Th2 balance in HIV-infected patients played more important role than Cryptococcus numbers and size in CSF on the development of high intracranial pressure in HIV-associated cryptococcal meningitis, providing a new understanding of mechanisms of HCM.

Isolation and characterization of novel reassortant H6N1 avian influenza viruses from chickens in Eastern China.

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.

Establishment of a multiplex real-time RT-PCR assay for rapid identification of H6 subtype avian influenza viruses.

The H6 subtype avian influenza viruses (AIVs) possess the capacity for zoonotic transmission from avian species to humans. Establishment of a specific, rapid and sensitive method to screen H6 AIVs is necessary. Based on the conserved domain of the matrix and H6 AIV hemagglutinin genes, two TaqMan minor-groove-binder probes and multiplex real-time RT-PCR primers were designed in this study. The multiplex real-time RT-PCR assay developed in this study had high specificity and repeatability and a detection limit of 30 copies per reaction. This rapid diagnostic method will be useful for clinical detection and surveillance of H6 AIVs in China.

Isolation and molecular characterization of an H5N1 swine influenza virus in China in 2015.

In 2015, an H5N1 influenza virus was isolated from a pig in Zhejiang Province, Eastern China. This strain was characterized by whole-genome sequencing with subsequent phylogenetic analysis. Phylogenetic analysis showed that all segments from this strain belonged to clade 2.3.2 and that it had received its genes from poultry influenza viruses in China. A Glu627Lys mutation associated with pathogenicity was observed in the PB2 protein. This strain was moderately pathogenic in mice and was able to replicate without prior adaptation. These results suggest that active surveillance of swine influenza should be used as an early warning system for influenza outbreaks in mammals.

Development of a TaqMan MGB RT-PCR assay for the detection of type A and subtype H10 avian influenza viruses.

H10 subtype avian influenza viruses have caused several epidemics in poultry and mammals, and specific, rapid and sensitive methods for detection are urgently needed. Herein, TaqMan minor groove binder (MGB) probes and multiplex real-time RT-PCR primers were designed based on gene regions encoding conserved domains of the nucleoprotein and H10 hemagglutinin. The developed multiplex real-time RT-PCR assay displayed high specificity, repeatability, and a detection limit of 10 copies per reaction. This diagnostic method could prove valuable for the rapid detection of H10 subtype AIVs in China.

Characterization of HIV-1 subtypes and transmitted drug resistance among treatment-naive HIV-infected individuals in Zhejiang, China, 2014-2017.

In recent years, transmitted drug resistance (TDR) has adversely impacted upon first-line therapy for HIV-infected individuals. To understand the current subtype distribution and TDR level in Zhejiang, China we performed phylogenetic analysis and genotypic drug resistance testing of treatment-naive HIV-infected individuals. A total of 153 HIV-1 Pol genes were successfully amplified. The distribution of HIV-1 genotypes was as follows: CRF01_AE (43.8%); CRF07_BC (37.9%); subtype B/B' (7.2%); CRF08_BC (5.2%); and others (5.9%). Drug resistance analysis demonstrated that 11.1% of isolates contained at least one NRTI or NNRTI resistance-associated mutations while 2.0% were identified to be resistant to PIs. These findings enhance our understanding of the genetic diversity of HIV-1 strains circulating in Zhejiang and provide some guidelines for HIV initial treatment therapy.

Characterization of reassortant H1-subtype avian influenza viruses isolated from poultry in Zhejiang Province in China from 2013 to 2015.

From 2013 to 2015, 32 H1-subtype avian influenza viruses (AIVs), H1N2 (n = 12), H1N3 (n = 14), H1N4 (n = 4) and H1N9 (n = 2), were isolated from poultry in Zhejiang Province in eastern China. These strains were characterized by whole-genome sequencing with subsequent phylogenetic analysis and genetic comparison. Phylogenetic analysis of all eight viral genes showed that these strains clustered in the AIV Eurasian lineage. These strains were found to be minimally pathogenic in mice and were able to replicate in mice without prior adaptation. Continued surveillance is needed, considering the important role of poultry in AIV reassortment.

Molecular characterization of a novel reassortant H7N6 subtype avian influenza virus from poultry in Eastern China, in 2016.

During the surveillance for avian influenza viruses (AIVs) in live poultry markets in Eastern China, in 2016, a novel reassortant H7N6 AIV was isolated from a chicken. Phylogenetic analysis showed that this strain received its genes from H9N2, H7N9 and H5N6 AIVs infecting poultry in China. This strain showed moderate pathogenicity in mice and was able to replicate in mice without prior adaptation. Considering that this novel reassorted H7N6 virus was isolated from poultry in this study, it is possible that chickens play an important role in the generation of novel reassorted H7N6 AIVs.

High-throughput sequencing identifies HIV-1-replication- and latency-related miRNAs in CD4+ T cell lines.

MicroRNAs are potent gene expression regulators involved in regulating various biological processes, including host-pathogen interactions. In this study, we used high-throughput sequencing to investigate cellular miRNA signatures related to HIV-1 replication and latent infection in CD4+ T cell lines, which included HIV-1-replicating H9/HTLV-IIIB, HIV-1-latently-infected CEM-Bru cells, and their parental uninfected H9 and CEM-SS cells. Relatively few miRNAs were found to be modulated by HIV-1 replication or latent infection, while the cell-lineage-specific miRNA difference was more pronounced, irrespective of HIV-1 infection. In silico analysis showed that some of our HIV-1 infection-regulated miRNA profiles echoed previous studies, while others were novel. In addition, some of the miRNAs that were differentially expressed between the productively and latently infected cells seemed to participate in shaping the differential infection state. Thus, the newly identified miRNA profiles related to HIV-1 replication and latency provide information about the interplay between HIV-1 and its host.

Amino acid substitutions involved in the adaptation of a novel highly pathogenic H5N2 avian influenza virus in mice.

BACKGROUND: H5N2 avian influenza viruses (AIVs) can infect individuals that are in frequent contact with infected birds. In 2013, we isolated a novel reassortant highly pathogenic H5N2 AIV strain [A/duck/Zhejiang/6DK19/2013(H5N2) (6DK19)] from a duck in Eastern China. This study was undertaken to understand the adaptive processes that led enhanced replication and increased virulence of 6DK19 in mammals. 6DK19 was adapted to mice using serial lung-to-lung passages (10 passages total). The virulence of the wild-type virus (WT-6DK19) and mouse-adapted virus (MA-6DK19) was determined in mice. The whole-genome sequences of MA-6DK19 and WT-6DK19 were compared to determine amino acid differences. FINDINGS: Amino acid changes were identified in the MA-DK19 PB2 (E627K), PB1 (I181T), HA (A150S), NS1 (seven amino acid extension "WRNKVAD" at the C-terminal), and NS2 (E69G) proteins. Survival and histology analyses demonstrated that MA-6DK19 was more virulent in mice than WT-6DK19. CONCLUSION: Our results suggest that these substitutions are involved in the enhanced replication efficiency and virulence of H5N2 AIVs in mammals. Continuing surveillance for H5N2 viruses in poultry that are carrying these mutations is required.

Amino acid substitutions occurring during adaptation of an emergent H5N6 avian influenza virus to mammals.

Avian influenza viruses (AIVs) are known to cross species barriers, and emergent highly pathogenic H5N6 AIVs pose a serious threat to human health and the poultry industry. Here, we serially passaged an H5N6 virus 10 times in BALB/c mice. The pathogenicity of the wild-type 6D2 (WT-6D2) and mammal-adapted 6D2 strain (MA-6D2) were compared. The viral titer in multiple organs and the death rate for MA-6D2 were significantly higher than for WT-6D2. We provide evidence that the mutations HA A150V, NA R143K and G147E, PB2 E627K, and PA A343T may be important for adaptation of H5N6 AIVs to mammals.

Multiple amino acid substitutions involved in the adaptation of avian-origin influenza A (H10N7) virus in mice.

To identify substitutions that are possibly associated with the adaptation of avian-origin H10N7 virus to mammals, adaptation of the H10N7 virus in mouse lung was carried out by serial lung-to-lung passage. Genomic analysis of the mouse-adapted virus revealed amino acid changes in the PB2 (E627K), PA (T97I), and HA (G409E) proteins, and this virus was more virulent in mice than the wild-type virus. Our results suggest that these substitutions are involved in the enhancement of the replication efficiency of avian-origin H10N7 virus, resulting in severe disease in mice. Continued poultry surveillance of these substitutions in H10N7 viruses is required.

Isolation and genetic characterization of novel reassortant H6N6 subtype avian influenza viruses isolated from chickens in eastern China.

H6 subtype avian influenza viruses (AIVs) possess the ability to cross the species barrier to infect mammals and pose a threat to human health. From June 2014 to July 2015, 12 H6N6 AIVs were isolated from chickens in live-poultry markets in Zhejiang Province, Eastern China. Phylogenetic analysis showed that these isolates received their genes from H6 and H9N2 subtype AIVs of poultry in China. These novel reassortant viruses showed moderate pathogenicity in mice and were able to replicate in mice without prior adaptation. Considering that novel reassorted H6N6 viruses were isolated from chickens in this study, it is possible that these chickens play an important role in the generation of novel reassorted H6N6 AIVs, and these results emphasize the need for continued surveillance of the H6N6 AIVs circulating in poultry.

Genetic and molecular characterization of a novel reassortant H2N8 subtype avian influenza virus isolated from a domestic duck in Zhejiang Province in China.

The circulation of the H2 subtype influenza viruses in domestic animals increases the risk of human exposure to these viruses. An H2N8 avian influenza virus (AIV) was isolated from a domestic duck during AIV surveillance of poultry in live poultry markets (LPMs) in Zhejiang Province, Eastern China, in 2013. The phylogenetic trees suggested that this strain is a novel reassortant virus derived from multiple AIV subtypes from aquatic birds and poultry in Eastern Asia. Although this reassortant strain exhibited low pathogenicity in mice, it was able to replicate in the lungs of the mice without prior adaptation. Continued surveillance of domestic ducks in LPMs is required for early detection of AIV outbreaks in poultry and humans.

Isolation and molecular characterization of reassortant H11N3 subtype avian influenza viruses isolated from domestic ducks in Zhejiang Province in China.

In July 2013, six H11N3 subtype avian influenza viruses (AIVs) were isolated from domestic ducks in Zhejiang Province in Eastern China. These strains were characterized by whole genome sequencing with subsequent phylogenetic analysis and genetic comparison. Phylogenetic analysis of all eight viral genes showed that these strains clustered in the AIV Eurasian lineage, and these strains received their genes from H11, H7, and H1 AIVs in Eastern China. These strains were found to be minimally pathogenic in mice, and were able to replicate in mice without prior adaptation. Continued surveillance is needed considering the important role of domestic ducks in AIV reassortment.

Molecular characterization of a novel reassortant H1N2 influenza virus containing genes from the 2009 pandemic human H1N1 virus in swine from eastern China.

Pandemic outbreaks of H1N1 swine influenza virus have been reported since 2009. Reassortant H1N2 viruses that contain genes from the pandemic H1N1 virus have been isolated in Italy and the United States. However, there is limited information regarding the molecular characteristics of reassortant H1N2 swine influenza viruses in eastern China. Active influenza surveillance programs in Zhejiang Province identified a novel H1N2 influenza virus isolated from pigs displaying clinical signs of influenza virus infection. Whole-genome sequencing was performed and this strain was compared with other influenza viruses available in GenBank. Phylogenetic analysis suggested that the novel strain contained genes from the 2009 pandemic human H1N1 and swine H3N2 viruses. BALB/c mice were infected with the isolated virus to assess its virulence in mice. While the novel H1N2 isolate replicated well in mice, it was found to be less virulent. These results provide additional evidence that swine serve as intermediate hosts or 'mixing vessels' for novel influenza viruses. They also emphasize the importance of surveillance in the swine population for use as an early warning system for influenza outbreaks in swine and human populations.