ABSTRACT Herpes simplex virus type 1 (HSV-1) mutants lacking the ICP34.5 gene are severely attenuated in mouse models and have a significant growth defect in confluent mouse embryo fibroblasts. Previously, ICP34.5 was demonstrated to have a crucial role in evading the innate immune response to infection by mediating the dephosphorylation of eIF2α, a translation initiation factor phosphorylated by PKR during the antiviral response. To further understand the role of ICP34.5 in evasion of the antiviral response, we used transcriptional profiling to examine host cell gene expression in both wild-type and ICP34.5-null virus-infected mouse embryo fibroblasts over a time course of infection. Our study revealed that cells responded to infection within 3 h through PKR-dependent eIF2α phosphorylation and that the majority of up-regulated genes at 3 h postinfection were involved in the antiviral response. HSV-1 counters this response through early expression of ICP34.5 and dephosphorylation of eIF2α. By 12 h postinfection, the differences between the number and functional classification of genes differentially up- and down-regulated between wild-type and ICP34.5-null virus-infected cells were maximal. Specifically, in wild-type virus-infected cells, the majority of changed genes were involved in metabolic and biosynthetic processes, while in ICP34.5-null virus-infected cells, mostly antiviral genes were up-regulated. Further, ICP34.5-null virus-infected cells produced greater amounts of beta interferon than wild-type virus-infected cells. These results indicate that ICP34.5 expression and function at early times postinfection have a pivotal role in the ability of HSV-1 to gain control of the host cell and maintain an environment for successful viral replication.
Incident bacterial vaginosis (BV) is associated with a shift in the vaginal microbiota from lactobacillus predominant to a diverse community of anaerobic bacteria The kinetics of this shift are poorly described. We sought to identify the dynamics of incident BV.
Methods
Twenty women with frequent BV (>3 episodes per year) self-collected vaginal swabs every 8 hours for 60 days. Swabs were analysed with quantitative PCR targeting Lactobacillus iners, Lactobacillus jensenii, Lactobacillus crispatus, Gardenerella vaginalis, Atopobium vaginae, bacterial vaginosis associated bacterium-2 (BVAB2), and Megasphaera spp. We defined bacteria as absent, low (4 DNA copies), moderate (104–108 DNA copies) or high-level (108–1011 DNA copies). Participants kept detailed diaries regarding sexual behaviour, menstruation, antibiotic use and vulvovaginal symptoms.
Results
We noted three states of the vaginal microbiota: three women had high-level lactobacilli with intermittent, transient low-level G. vaginalis throughout the sampling period (State 1); five women had high-level lactobacilli with persistent, fluctuating low to moderate-level G. vaginalis and other BV associated anaerobic species throughout the sampling period (State 2); two women had polymicrobial colonisation with high-level G. vaginalis and other BV associated species, and intermittent, transient low-level, or persistent moderate-level Lactobacillus jensenii and crispatus throughout the sampling period (State 3). Ten women shifted between States 2 and 3 on at least one occasion. Extremely rapid transition from State 2 to 3 over.
Conclusions
The vaginal microbiota is extremely dynamic and BV develops over narrow time intervals. Low-levels of BV associated species in the vagina may be a risk factor for rapid, incident BV. Future studies will identify drivers of shifts in the vaginal bacterial biota.
Objectives To compare gastrointestinal (GI) tolerability and patient preference for the new 625 mg formulation of nelfinavir (NFV) and the marketed 250 mg tablets (Viracept ® ) in HIV‐1‐infected patients. Methods Virologically controlled patients ( n =126) treated with a nelfinavir (NFV) 250 mg‐containing regimen for ≥8 weeks completed a stool diary for 14 days to assess baseline bowel function. After switching to the NFV 625 mg formulation [1250 mg twice a day (bid)] for 28 days, patients continued their stool diaries and at study completion answered a questionnaire regarding formulation preferences. Results The incidence and mean weekly duration of GI upset over a 2‐week period were lower with NFV 625 mg than with NFV 250 mg (79.8% vs. 84.9% of patients and 2.1 vs. 3.0 days, respectively). Fewer patients experienced moderate or severe diarrhoea with NFV 625 mg (6.5% vs. 11.1%), and the incidence of investigator‐assessed diarrhoea also decreased with NFV 625 mg. Importantly, there was a significant improvement overall in the incidence of diarrhoea (any grade) when patients switched to NFV 625 mg [38 of 124 (31%) improving, 69 of 124 (56%) stable and 17 of 124 (14%) worsening on NFV 625 mg; P <0.01]. At study completion, most patients expressed a preference to continue treatment with NFV 625 mg [112 of 122 (91.8%); P <0.0001], with only one patient (0.8%) preferring to resume treatment with NFV 250 mg. The new formulation was well tolerated with no new safety concerns. Conclusions The new NFV 625 mg formulation is better tolerated and preferred by patients switching from NFV 250 mg tablets. By reducing the daily pill count and improving GI tolerability, the NFV 625 mg formulation may enhance patient adherence to NFV‐containing antiretroviral regimens and thus potentially improve virological outcomes.
Vesicular stomatitis virus expressing Zaire Ebola virus (EBOV) glycoprotein (VSVΔG/EBOVgp) could be used as a vaccine to meet the 2014 Ebola virus outbreak. To characterize the host response to this vaccine, we used mRNA sequencing to analyze peripheral blood mononuclear cells (PBMCs) from cynomolgus macaques after VSVΔG/EBOVgp immunization and subsequent EBOV challenge. We found a controlled transcriptional response that transitioned to immune regulation as the EBOV was cleared. This observation supports the safety of the vaccine.
Abstract Using a mouse mutagenesis screen, we have identified CD83 as being critical for the development of CD4+ T cells and for their function postactivation. CD11c+ dendritic cells develop and function normally in mice with a mutated CD83 gene but CD4+ T cell development is substantially reduced. Additionally, we now show that those CD4+ cells that develop in a CD83 mutant animal fail to respond normally following allogeneic stimulation. This is at least in part due to an altered cytokine expression pattern characterized by an increased production of IL-4 and IL-10 and diminished IL-2 production. Thus, in addition to its role in selection of CD4+ T cells, absence of CD83 results in the generation of cells with an altered activation and cytokine profile.
Investigating the human fallopian tube (FT) microbiota has significant implications for understanding the pathogenesis of ovarian cancer (OC). In this large prospective study, we collected swabs intraoperatively from the FT and other surgical sites as controls to profile the microbiota in the FT and to assess its relationship with OC. Eighty-one OC and 106 non-cancer patients were enrolled and 1001 swabs were processed for 16S rRNA gene PCR and sequencing. We identified 84 bacterial species that may represent the FT microbiota and found a clear shift in the microbiota of the OC patients when compared to the non-cancer patients. Of the top 20 species that were most prevalent in the FT of OC patients, 60% were bacteria that predominantly reside in the gastrointestinal tract, while 30% normally reside in the mouth. Serous carcinoma had higher prevalence of almost all 84 FT bacterial species compared to the other OC subtypes. The clear shift in the FT microbiota in OC patients establishes the scientific foundation for future investigation into the role of these bacteria in the pathogenesis of OC.
ABSTRACT We previously reported widespread differential expression of long non-protein-coding RNAs (ncRNAs) in response to virus infection. Here, we expanded the study through small RNA transcriptome sequencing analysis of the host response to both severe acute respiratory syndrome coronavirus (SARS-CoV) and influenza virus infections across four founder mouse strains of the Collaborative Cross, a recombinant inbred mouse resource for mapping complex traits. We observed differential expression of over 200 small RNAs of diverse classes during infection. A majority of identified microRNAs (miRNAs) showed divergent changes in expression across mouse strains with respect to SARS-CoV and influenza virus infections and responded differently to a highly pathogenic reconstructed 1918 virus compared to a minimally pathogenic seasonal influenza virus isolate. Novel insights into miRNA expression changes, including the association with pathogenic outcomes and large differences between in vivo and in vitro experimental systems, were further elucidated by a survey of selected miRNAs across diverse virus infections. The small RNAs identified also included many non-miRNA small RNAs, such as small nucleolar RNAs (snoRNAs), in addition to nonannotated small RNAs. An integrative sequencing analysis of both small RNAs and long transcripts from the same samples showed that the results revealing differential expression of miRNAs during infection were largely due to transcriptional regulation and that the predicted miRNA-mRNA network could modulate global host responses to virus infection in a combinatorial fashion. These findings represent the first integrated sequencing analysis of the response of host small RNAs to virus infection and show that small RNAs are an integrated component of complex networks involved in regulating the host response to infection. IMPORTANCE Most studies examining the host transcriptional response to infection focus only on protein-coding genes. However, mammalian genomes transcribe many short and long non-protein-coding RNAs (ncRNAs). With the advent of deep-sequencing technologies, systematic transcriptome analysis of the host response, including analysis of ncRNAs of different sizes, is now possible. Using this approach, we recently discovered widespread differential expression of host long (>200 nucleotide [nt]) ncRNAs in response to virus infection. Here, the samples described in the previous report were again used, but we sequenced another fraction of the transcriptome to study very short (about 20 to 30 nt) ncRNAs. We demonstrated that virus infection also altered expression of many short ncRNAs of diverse classes. Putting the results of the two studies together, we show that small RNAs may also play an important role in regulating the host response to virus infection.
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