Project description:We utilized host-pathogen dual RNA-sequencing to elucidate the transcriptomes of both Chlamydia trachomatis and the infected HeLa cell during nutritional conditions that induce persistence.
Project description:Chlamydiae are obligate intracellular bacteria comprising well-known human pathogens and ubiquitous symbionts of protists, which are characterized by a unique developmental cycle. Here we comprehensively analyzed gene expression dynamics of Protochlamydia amoebophila during infection of its Acanthamoeba host by RNA sequencing. This revealed a highly dynamic transcriptional landscape, where major transcriptional shifts are conserved among chlamydial symbionts and pathogens. Our data served to propose a time-resolved model for type III protein secretion during the developmental cycle, and we provide evidence for a biphasic metabolism of P. amoebophila during infection, which involves energy parasitism and amino acids as carbon source during initial stages and a post-replicative switch to endogenous glucose-based ATP production. This fits well with major transcriptional changes in the amoeba host, where upregulation of complex sugar breakdown precedes the P. amoebophila metabolic switch. The biphasic chlamydial metabolism represents a unique adaptation to exploit eukaryotic host cells, which likely contributed to the evolutionary success of this group of microbes.
Project description:NK cells and pulmonary macrophages both are important components of innate immunity. The interaction between NK cells and pulmonary macrophages during Chlamydia muridarum(C. muridarum)respiratory infections is poorly understood. In this study, we explored the effect of NK cells on regulation of pulmonary macrophage function during chlamydial lung infection. We found that NK depletion led to polarization of pulmonary macrophages from M1 to M2 phenotype, and this related to significantly reduced miR-155 expression in pulmonary macrophage. Using adoptive transfer approach, we found that the recipient mice receiving lung macrophages isolated from C. muridarum-infected NK-cell-depleted mice exhibited an increased bacterial load and severe inflammation in the lung upon chlamydial challenge when compared with the recipients of lung macrophages from infected IgG -treated mice. Herein, the effects of NK cells on macrophage polarization were examined in vitro. We found that NK cells from chlamydial-infected mice (iNK) significantly induced M1 polarization compared to that from sham-infected mice (uNK). Inhibition of miR-155 expression in macrophages attenuated M1 polarization induced by iNK, while miR-155 over-expression enhanced it. Furthermore, neutralization of IFN-γ in the coculture system decreased the expression of miR-155 by macrophages, and resulted in diminished M1 polarization induced by iNK cells. The data indicates that NK cells direct M1 polarization through up-regulation of miR-155 by IFN-γ production, and NK-regulated macrophage polarization is functionally relevant to host defense against chlamydial infection.
Project description:Characterization of host-pathogen interactions is critical for the development of next-generation therapies and vaccines. Classical approaches involve the use of transformed cell lines and/or animal models which may not reflect the complexity and response of the human host. We reconstituted the ciliated human bronchial epithelium in vitro using primary bronchial epithelial cells to simultaneously monitor the infection-linked global changes in nontypeable Haemophilus influenzae (NTHi) and infected host epithelia gene expression by dual RNA-seq. Acquisition of a total of nearly 2,5 billion sequences allowed construction of high-resolution strand-specific transcriptome maps of NTHi during infection of host mucosal surface and monitoring of metabolic as well as stress-induced host-adaptation strategies of this pathogen. As a part of our screening, we identified a global profile of noncoding transcripts that are candidate small RNAs regulated during human host infection in Haemophilus species. Temporal analysis of host mRNA signatures revealed significant dysregulation of target cell cytoskeleton elicited by bacterial infection, with a profound effect on intermediate filament network of bronchial epithelium. Our data provide a robust and comprehensive catalogue of regulatory responses that drive NTHi pathogenesis and gives novel insights into complex crosstalk between the host and the invading pathogen. Primary human bronchial epithelium was infected with NTHi at a multiplicity of 100:1. Total RNA was isolated at 1, 6, 24 and 72 h post-infection in three biologically-independent experiments and cDNA libraries were prepared and sequenced with Illumina HiSeq 2500 sequencer. At each time point, between 60 and 180 million total reads per sample were obtained of which approximately one-third could be aligned to non-rRNA regions of the bacterial and human genomes
Project description:Characterization of host-pathogen interactions is critical for the development of next-generation therapies and vaccines. Classical approaches involve the use of transformed cell lines and/or animal models which may not reflect the complexity and response of the human host. We reconstituted the ciliated human bronchial epithelium in vitro using primary bronchial epithelial cells to simultaneously monitor the infection-linked global changes in nontypeable Haemophilus influenzae (NTHi) and infected host epithelia gene expression by dual RNA-seq. Acquisition of a total of nearly 2,5 billion sequences allowed construction of high-resolution strand-specific transcriptome maps of NTHi during infection of host mucosal surface and monitoring of metabolic as well as stress-induced host-adaptation strategies of this pathogen. As a part of our screening, we identified a global profile of noncoding transcripts that are candidate small RNAs regulated during human host infection in Haemophilus species. Temporal analysis of host mRNA signatures revealed significant dysregulation of target cell cytoskeleton elicited by bacterial infection, with a profound effect on intermediate filament network of bronchial epithelium. Our data provide a robust and comprehensive catalogue of regulatory responses that drive NTHi pathogenesis and gives novel insights into complex crosstalk between the host and the invading pathogen.
Project description:Helicobacter pylori (H. pylori) is a highly successful pathogen that constitutes a serious threat to human health. However, the dynamic interaction between H. pylori and human gastric epithelium has not been fully documented. Here, by leveraging the advantage of dual RNA sequencing technology, we characterized a cytotoxin-associated genes A (CagA)-modulated bacterial adoption strategy by reinforcing the expression of ATP-binding cassette (ABC) transporter-related genes, metQ and HP_0888 upon co-culturing with human gastric epithelial cells (GES-1), thus, to benefit intracellular H. pylori survival through facilitating the uptake of host-provided nutrients. We further detected a generally repressed impact on electron transportation-associated genes by CagA, leading to the activation of oxidative phosphorylation. Temporal profiling of host mRNA signatures revealed down-regulation of multiple splicing regulators elicited by bacterial infection. Consequently, aberrant pre-mRNA splicing of functional genes that were involved in cell cycle process in response to H. pylori infection were identified. Moreover, we verified a protective effect for gastric H. pylori colonization against chronic dextran sulfate sodium (DSS)-induced colitis. Mechanistically, we profiled a cluster of short chain fatty acids (SCFA)-producing bacteria, Muribaculaceae that was selectively enriched in H. pylori-pre-colonized mice colon, contributing to the restoration of intestinal barrier function damaged by DSS treatment. Taken together, this study represents the first dual-transcriptome analysis of H. pylori during dynamic interaction with gastric epithelium and provide new insights into H. pylori pathogenesis.
Project description:Genital C. trachomatis (CT) infection may cause pelvic inflammatory disease (PID) that can lead to tubal factor infertility (TFI). Understanding the pathogenesis of chlamydial complications including the pathophysiological processes within the female host genital tract is of immense importance in preventing adverse pathology. In this study, we tested the hypothesis that the miRNA profile of a acute primary chlamydial infection characterized by temporary inflammation versus the profile associated with chronic genital chlamydial infections that might precipitate PID or TFI will be different. Thus, we analyzed and compared the differentially expressed miRNAs that regulate CT pathogenesis after a single genital infection and those involved in the development of PID and TFI after repeat infections. Mice (Mus musculus) were infected with Chlamydia muridarum once or twice with a month interval between infections, and then sacrificed and genital tract tissues were collected at 1, 2, 4, and 8 weeks after infection. miRNAs were differentially expressed in both first infection and the re-infection; however, the miRNA expression profile was different for both groups. Pathway analysis showed that, amongst other functions, the differentially regulated miRNA might be regulating several pathways involved in cellular and tissue development, disease conditions and toxicity. Grant number: 1SC2HD086066-03 Funding source: Eunice Kennedy Shriver National Institute of Child Health & Human Development Title: Discovering Novel Biomarkers Predictive of Tubal Infertility Caused by Chlamydia. Principal investigator: Yusuf Omosun Date: 05/01/2015-04/30/2019