Dataset Information

In vivo dual RNA-seq reveals that neutrophil recruitment underlies differential tissue tropism of Streptococcus pneumoniae.

ABSTRACT: Streptococcus pneumoniae is a genetically diverse human-adapted pathogen commonly carried asymptomatically in the nasopharynx. We have recently shown that a single nucleotide polymorphism (SNP) in the raffinose pathway regulatory gene rafR accounts for a difference in the capacity of clonally-related strains to cause localised versus systemic infection. Using dual RNA-seq, we show that this SNP affects expression of bacterial genes encoding multiple sugar transporters, and fine-tunes carbohydrate metabolism, along with extensive rewiring of host transcriptional responses to infection, particularly expression of genes encoding cytokine and chemokine ligands and receptors. The data predict a crucial role for differential neutrophil recruitment (confirmed by in vivo neutrophil depletion and IL-17 neutralization) indicating that early detection of bacteria by the host in the lung environment is crucial for effective clearance. Thus, dual RNA-seq provides a powerful tool for understanding complex host-pathogen interactions and reveals how a single bacterial SNP can drive differential disease outcomes.

SUBMITTER: Minhas V

PROVIDER: S-EPMC7275033 | biostudies-literature | 2020 Jun

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

In vivo dual RNA-seq reveals that neutrophil recruitment underlies differential tissue tropism of Streptococcus pneumoniae.

Minhas Vikrant V Aprianto Rieza R McAllister Lauren J LJ Wang Hui H David Shannon C SC McLean Kimberley T KT Comerford Iain I McColl Shaun R SR Paton James C JC Veening Jan-Willem JW Trappetti Claudia C

Communications biology 20200605 1

Streptococcus pneumoniae is a genetically diverse human-adapted pathogen commonly carried asymptomatically in the nasopharynx. We have recently shown that a single nucleotide polymorphism (SNP) in the raffinose pathway regulatory gene rafR accounts for a difference in the capacity of clonally-related strains to cause localised versus systemic infection. Using dual RNA-seq, we show that this SNP affects expression of bacterial genes encoding multiple sugar transporters, and fine-tunes carbohydrat ...[more]

PMID: 32504007

Similar Datasets

Project description:Streptococcus pneumoniae is the dominant cause of community-acquired pneumonia worldwide. Invasion of the pleural space is common and results in increased mortality. We set out to determine the bacterial and host factors that influence invasion of the pleural space. In a murine model of pneumococcal infection, we isolated neutrophil-dominated samples of bronchoalveolar and pleural fluid containing bacteria 48 hours after infection. Using dual RNA sequencing (RNA-seq), we characterized bacterial and host transcripts that were differentially regulated between these compartments and bacteria in broth and resting neutrophils, respectively. Pleural and lung samples showed upregulation of genes involved in the positive regulation of neutrophil extravasation but downregulation of genes mediating bacterial killing. Compared to the lung samples, cells within the pleural space showed marked upregulation of many genes induced by type I interferons, which are cytokines implicated in preventing bacterial transmigration across epithelial barriers. Differences in the bacterial transcripts between the infected samples and bacteria grown in broth showed the upregulation of genes in the bacteriocin locus, the pneumococcal surface adhesin PsaA, and the glycopeptide resistance gene vanZ; the gene encoding the ClpP protease was downregulated in infection. One hundred sixty-nine intergenic putative small bacterial RNAs were also identified, of which 43 (25.4%) small RNAs had been previously described. Forty-two of the small RNAs were upregulated in pleura compared to broth, including many previously identified as being important in virulence. Our results have identified key host and bacterial responses to invasion of the pleural space that can be potentially exploited to develop alternative antimicrobial strategies for the prevention and treatment of pneumococcal pleural disease.IMPORTANCE The factors that regulate the passage of bacteria between different anatomical compartments are unclear. We have used an experimental model of infection with Streptococcus pneumoniae to examine the host and bacterial factors involved in the passage of bacteria from the lung to the pleural space. The transcriptional profile of host and bacterial cells within the pleural space and lung was analyzed using deep sequencing of the entire transcriptome using the technique of dual RNA-seq. We found significant differences in the host and bacterial RNA profiles in infection, which shed light on the key factors that allow passage of this bacterium into the pleural space.

Project description:An important determinant of disease following Streptococcus pneumoniae (pneumococcus) lung infection is pulmonary inflammation mediated by polymorphonuclear leukocytes (PMNs). We found that upon intratracheal challenge of mice, recruitment of PMNs into the lungs within the first 3 hours coincided with decreased pulmonary pneumococci, whereas large numbers of pulmonary PMNs beyond 12 hours correlated with a greater bacterial burden. Indeed, mice that survived infection largely resolved inflammation by 72 hours, and PMN depletion at peak infiltration, i.e. 18 hours post-infection, lowered bacterial numbers and enhanced survival. We investigated host signaling pathways that influence both pneumococcus clearance and pulmonary inflammation. Pharmacologic inhibition and/or genetic ablation of enzymes that generate extracellular adenosine (EAD) (e.g. the ectoenzyme CD73) or degrade EAD (e.g. adenosine deaminase) revealed that EAD dramatically increases murine resistance to S. pneumoniae lung infection. Moreover, adenosine diminished PMN movement across endothelial monolayers in vitro, and although inhibition or deficiency of CD73 had no discernible impact on PMN recruitment within the first 6 hours after intratracheal inoculation of mice, these measures enhanced PMN numbers in the pulmonary interstitium after 18 hours of infection, culminating in dramatically elevated numbers of pulmonary PMNs at three days post-infection. When assessed at this time point, CD73-/- mice displayed increased levels of cellular factors that promote leukocyte migration, such as CXCL2 chemokine in the murine lung, as well as CXCR2 and β-2 integrin on the surface of pulmonary PMNs. The enhanced pneumococcal susceptibility of CD73-/- mice was significantly reversed by PMN depletion following infection, suggesting that EAD-mediated resistance is largely mediated by its effects on PMNs. Finally, CD73-inhibition diminished the ability of PMNs to kill pneumococci in vitro, suggesting that EAD alters both the recruitment and bacteriocidal function of PMNs. The EAD-pathway may provide a therapeutic target for regulating potentially harmful inflammatory host responses during Gram-positive bacterial pneumonia.

Project description:Enhanced reactive oxygen species production in allergic airways is well described and correlates with increased airway contractions, inflammatory cell infiltration, goblet cell metaplasia, and mucus hypersecretion. There is also an abundance of interleukin-4/interleukin-13 (IL-4/IL-13)- or interleukin-5-secreting cells that are thought to be central to the pathogenesis of allergic asthma. We postulated that the dual oxidases (DUOX1 and DUOX2), members of the nicotinamide adenine dinucleotide phosphate oxidase family that release hydrogen peroxide (H2O2) in the respiratory tract, are critical proteins in the pathogenesis of allergic airways. DUOX activity is regulated by cytokines, including IL-4 and IL-13, and DUOX-mediated H2O2 influences several important features of allergic asthma: mucin production, IL-8 secretion, and wound healing. The objective of this study was to establish the contribution of DUOXs to the development of allergic asthma in a murine model. To accomplish this goal, we utilized a DUOXA-deficient mouse model (Duoxa(-/-)) that lacked maturation factors for both DUOX1 and DUOX2. Our results are the first to demonstrate evidence of DUOX protein and DUOX functional activity in murine airway epithelium. We also demonstrate that DUOXA maturation factors are required for airway-specific H2O2 production and localization of DUOX to cilia of fully differentiated airway epithelial cells. We compared wild-type and Duoxa(-/-) mice in an ovalbumin exposure model to determine the role of DUOX in allergic asthma. In comparison to DUOX-intact mice, Duoxa(-/-) mice had reduced mucous cell metaplasia and lower levels of TH2 cytokine levels in bronchoalveolar fluid. In addition, increased airway resistance in response to methacholine was observed in Duoxa(+/+) mice, as expected, but was absent in Duoxa(-/-) mice. Surprisingly, Duoxa(-/-) mice had decreased influx of neutrophils in bronchoalveolar fluid and lung tissue sections associated with a lower level of the chemotactic cytokine IL-6. These findings suggest that DUOX-derived H2O2 has an important role in signaling neutrophils into allergic airways.

Project description:Streptococcus pneumoniae undergoes spontaneous phase variation resulting in opaque and transparent colony forms. Differences in colony opacity correlate with differences in virulence: the transparent variants are more capable of colonizing the nasopharynx, whereas the opaque variants show increased virulence during systemic infections. To gain insight into the pathogenesis of pneumococcal disease at the molecular level, protein expression patterns of the phenotypic variants of two pneumococcal strains were compared by high-resolution two-dimensional protein electrophoresis. In comparison with transparent variants, the opaque variants reduced the expression of two proteins and overexpressed one protein. The proteins were identified by mass spectrometric analysis. The protein overexpressed in the opaque phenotype revealed significant homology to elongation factor Ts of Helicobacter pylori. One of the two proteins that were underexpressed in the opaque variants revealed significant homology to the proteinase maturation protein PrtM of Lactocobacillus paracasei, a member of the family of peptidyl-prolyl cis/trans isomerases. A consensus lipoprotein signal sequence suggests that the putative proteinase maturation protein A, designated PpmA, is located at the surface of the pneumococcus and may play a role in the maturation of surface or secreted proteins. The second underexpressed protein was identified as pyruvate oxidase, SpxB. The lower SpxB expression in opaque variants most probably explains the reduced production of hydrogen peroxide, a reaction product of SpxB, in this variant. Since a spxB-defective pneumococcal mutant has decreased ability to colonize the nasopharynx (B. Spellerberg, D. R. Cundell, J. Sandros, B. J. Pearce, I. Idanpaan-Heikkila, C. Rosenow, and H. R. Masure, 1996. Mol. Microbiol. 19:803-813, 1996), our data suggest that SpxB plays an important role in enhancing the ability of transparent variants to efficiently colonize the nasopharynx.

Dataset Information

In vivo dual RNA-seq reveals that neutrophil recruitment underlies differential tissue tropism of Streptococcus pneumoniae.

Publications

In vivo dual RNA-seq reveals that neutrophil recruitment underlies differential tissue tropism of Streptococcus pneumoniae.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets