Biology subjects

Aprianto, R.

Publications and source records attributed to Aprianto, R..

7 recordsLinked to original sources

bioRxiv2024

The Streptococcus pneumoniae transcriptome in patient cerebrospinal fluid identifies novel virulence factors required for meningitis

To better understand Streptococcus pneumoniae pathogenesis we performed RNA sequencing on cerebrospinal fluid (CSF) from meningitis patients to identify bacterial genes expressed during invasion of the central nervous system. Comparison to transcriptome data for serotype 1 S. pneumoniae cultured in ex vivo human CSF defined a subset of 57 genes with high expression during human meningitis. Deletion of two of the most highly expressed genetic loci, bgaA (encodes for a {beta}-galactosidase) or the SP_1801-5 putative stress response operon, resulted in S. pneumoniae strains still able to transmigrate the blood brain barrier but which were more susceptible to complement opsonisation and unable to maintain brain infection in a murine meningitis model. In 1144 meningitis patients, infection with bgaA containing S. pneumoniae strains was associated with a higher mortality (22% versus 14% p=0.02). These data demonstrate that direct bacterial RNAseq from CSF can identify previously undescribed S. pneumoniae virulence factors required for meningitis pathogenesis.

microbiology↗

bioRxiv2023

Klebsiella pneumoniae OmpR facilitates lung infection through transcriptional regulation of key virulence factors

Bacteria must adapt to the stresses of specific environmental conditions to survive. This adaptation is often achieved by altering gene expression through two-component regulatory systems (TCSs). In Gram-negative bacteria, the response to environmental changes in osmolarity and pH are primarily mediated by the EnvZ/OmpR TCS. Although the functioning of EnvZ/OmpR has been well characterized in Escherichia coli, Salmonella enterica, and the Yersinia genus, the importance of EnvZ/OmpR TCS in the opportunistic human pathogen Klebsiella pneumoniae has been limitedly studied. Here, we investigated the importance of EnvZ/OmpR in K. pneumoniae for fitness, gene regulation, virulence, and infection. Through the generation of a markerless ompR-deletion mutant, we show that overall fitness of K. pneumoniae is not impacted in vitro. Using dual RNA-seq of K. pneumoniae co-incubated with human lung epithelial cells we demonstrate that the K. pneumoniae OmpR regulon includes important virulence factors, but shows otherwise limited overlap with the regulons of other Gram-negative bacteria. In addition, we show that deletion of ompR in K. pneumoniae leads to a stronger antibacterial transcriptional response in human lung epithelial cells. Lastly, we show that OmpR is crucial for K. pneumoniae virulence and infection through a murine lung infection model. As the adaptation of commensal bacteria to specific niches is mediated by TCSs, we show that EnvZ/OmpR plays a crucial role in successful lung infection, as well as in virulence. These results suggest that OmpR is an interesting target for anti-virulence drug discovery programs. ImportanceBacteria use two-component regulatory systems (TCSs) to adapt to changes in their environment by changing their gene expression. In this study, we show that the EnvZ/OmpR TCS of Klebsiella pneumoniae plays an important role in successfully establishing lung infection, and virulence. In addition, we discern the transcriptional response that OmpR facilitates within this clinically relevant opportunistic pathogen, and within the host. This work suggests that K. pneumoniae OmpR might be a promising target for innovative anti-infectives.

microbiology↗

bioRxiv2022

Pneumolysin promotes host cell necroptosis and bacterial competence during pneumococcal meningitis as shown by whole animal dual RNA-seq

Pneumolysin is a major virulence factor of Streptococcus pneumoniae that plays a key role in interaction with the host during invasive disease. How pneumolysin influences these dynamics between host and pathogen interaction during early phase of central nervous system infection in pneumococcal meningitis remains unclear. Using a whole animal in vivo dual RNA-seq approach, we identified pneumolysin-specific transcriptional responses in both S. pneumoniae and zebrafish (Danio rerio) during early pneumococcal meningitis. By functional enrichment analysis we identified host pathways known to be activated by pneumolysin, and discovered the importance of necroptosis for host survival. Inhibition of this pathway using the drugs necrostatin-5 or GSK872 increased host mortality during pneumococcal meningitis. On the pathogens side, we find that pneumolysin-dependent competence activation is crucial for intra-host replication and virulence and that not all bacteria activate competence at the same time. Altogether, this study provides new insights into pneumolysin-specific transcriptional responses and identifies key pathways involved in pneumococcal meningitis. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=200 SRC="FIGDIR/small/479878v1_ufig1.gif" ALT="Figure 1"> View larger version (30K): org.highwire.dtl.DTLVardef@1214d1aorg.highwire.dtl.DTLVardef@14d0e8corg.highwire.dtl.DTLVardef@1f6133aorg.highwire.dtl.DTLVardef@685955_HPS_FORMAT_FIGEXP M_FIG C_FIG HIGHLIGHTSO_LIPneumolysin-specific host and bacterial responses as identified by whole animal dual RNA-seq, available at https://veeninglab.com/dual-danio C_LIO_LIDiscovery of a functional necroptosis or necroptosis-like pathway in zebrafish C_LIO_LIHeterogeneity in competence development during infection C_LIO_LICompetence development is an important virulence determinant C_LI

microbiology↗

bioRxiv2019

In Vivo Dual RNA-Seq Analysis Reveals the Basis for Differential Tissue Tropism of Clinical Isolates of Streptococcus pneumoniae

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 significant difference in the capacity of clonally-related strains to cause localised versus systemic infection. Here we have used dual RNA-seq to show that this SNP extensively impacts both bacterial and host transcriptomes in infected lungs. It 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 dual RNA-seq data predicted a crucial role for differential neutrophil recruitment in the distinct virulence profiles of the infecting strains and single cell analysis revealed that while reduced expression of the RafR regulon driven by a single rafR SNP provides a clear advantage for pneumococci to colonize the ear, in the lung it leads to massive recruitment of neutrophils and bacterial clearance. Importantly, the observed disease outcomes were confirmed by in vivo neutrophil depletion showing 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 revealed how a single bacterial SNP can drive differential disease outcomes.

microbiology↗

bioRxiv2018

Refining the pneumococcal competence regulon by RNA-sequencing

Competence for genetic transformation allows the opportunistic human pathogen Streptococcus pneumoniae to take up exogenous DNA for incorporation into its own genome. This ability may account for the extraordinary genomic plasticity of this bacterium, leading to antigenic variation, vaccine escape, and the spread of antibiotic resistance markers. The competence system has been thoroughly studied and its regulation is well-understood. Additionally, over the last decade, several stress factors have been shown to trigger the competent state, leading to the activation of several stress response regulons. The arrival of next-generation sequencing techniques allowed us to update the competence regulon, the latest report of which still depended on DNA microarray technology. Enabled by the availability of an up-to-date genome annotation, including transcript boundaries, we assayed time-dependent expression of all annotated features in response to competence induction, were able to identify the affected promoters and produced a more complete overview of the various regulons activated during competence. We show that 4% of all annotated genes are under direct control of competence regulators ComE and ComX, while the expression of a total of up to 17% of all genes is, either directly or indirectly, affected. Among the affected genes are various small RNAs with an as-of-yet unknown function. Besides the ComE and ComX regulons, we were also able to refine the CiaR, VraR (LiaR) and BlpR regulons, underlining the strength of combining RNA-seq with a well-annotated genome.

microbiology↗

bioRxiv2018

Deep genome annotation of the opportunistic human pathogen Streptococcus pneumoniae D39

A precise understanding of the genomic organization into transcriptional units and their regulation is essential for our comprehension of opportunistic human pathogens and how they cause disease. Using single-molecule real-time (PacBio) sequencing we unambiguously determined the genome sequence of Streptococcus pneumoniae strain D39 and revealed several inversions previously undetected by short-read sequencing. Significantly, a chromosomal inversion results in antigenic variation of PhtD, an important surface-exposed virulence factor. We generated a new genome annotation using automated tools, followed by manual curation, reflecting the current knowledge in the field. By combining sequence-driven terminator prediction, deep paired-end transcriptome sequencing and enrichment of primary transcripts by Cappable-Seq, we mapped 1,015 transcriptional start sites and 748 termination sites. Using this new genomic map, we identified several new small RNAs (sRNAs), riboswitches (including twelve previously misidentified as sRNAs), and antisense RNAs. In total, we annotated 92 new protein-encoding genes, 39 sRNAs and 165 pseudogenes, bringing the S. pneumoniae D39 repertoire to 2,151 genetic elements. We report operon structures and observed that 9% of operons lack a 5-UTR. The genome data is accessible in an online resource called PneumoBrowse (https://veeninglab.com/pneumobrowse) providing one of the most complete inventories of a bacterial genome to date. PneumoBrowse will accelerate pneumococcal research and the development of new prevention and treatment strategies.

microbiology↗

bioRxiv2018

High-resolution analysis of the pneumococcal transcriptome under a wide range of infection-relevant conditions

Streptococcus pneumoniae is an opportunistic human pathogen that typically colonizes the nasopharyngeal passage and causes lethal disease in other host niches such as the lung or the meninges. How pneumococcal genes are expressed and regulated at the different stages of its life cycle, as commensal or as pathogen, has not been entirely described. To chart the transcriptional responses of S. pneumoniae, we quantified the transcriptome under 22 different infection-relevant conditions. The transcriptomic compendium exposed a high level of dynamic expression and, strikingly, all annotated pneumococcal genomic features were expressed in at least one of the studied conditions. By computing the correlation of gene expression of every two genes across all studied conditions, we created a co-expression matrix that provides valuable information on both operon structure and regulatory processes. The co-expression data is highly consistent with well-characterized operons and regulons, such as the PyrR, ComE and ComX regulons, and has allowed us to identify a new member of the competence regulon. Finally, we created an interactive data center named PneumoExpress (www.veeninglab.com/pneumoexpress) that enables users to access the expression data as well as the co-expression matrix in an intuitive and efficient manner, providing a valuable resource to the pneumococcal research community.

microbiology↗