Project description:Alterations in genes for penicillin-binding proteins (pbp) are well-known determinants for the resistance of Streptococcus pneumoniae to B-lactam antibiotics. Surprisingly, some mutations in non-pbp genes were also found to contribute to B-lactam resistance. Two of them discovered in the piperacillin resistant mutants P106 and P104, affect the expression of cpoA (encoding a glycosyltransferase) and of the rgtABCDHR cluster (encoding two small membrane proteins, an ABC transporter and a regulatory two-component system), respectively. cpoA and rgtABCDHR are involved in maintaining the synthesis and the proper ratio of the two major membrane glycolipids, and deletions in these genes led to complex phenotypes. In attempts to identify genetic determinants for these phenotypes, the global trancription patterns of the deletion mutants R6 delta cpoA, R6 delta rgtA and R6 delta rgtD were compared to that of the parent strain R6.
Project description:<p><em>Streptococcus pneumoniae</em> is the primary cause of community-acquired bacterial pneumonia with rates of penicillin and multi-drug resistance exceeding 80% and 40%, respectively. The innate immune response generates a variety of antimicrobial agents to control infection including zinc stress. Here, we characterized the impact of zinc intoxication on <em>S. pneumoniae</em>, revealing disruptions in central carbon metabolism, lipid biogenesis and peptidoglycan biosynthesis. Characterization of the pivotal peptidoglycan biosynthetic enzyme GlmU revealed an exquisite sensitivity to zinc inhibition. Disruption of the sole zinc efflux pathway, czcD, rendered <em>S. pneumonia</em>e highly susceptible to β-lactam antibiotics. To dysregulate zinc homeostasis in the wild-type strain, we investigated the safe-for-human use ionophore PBT2. PBT2 rendered wild-type <em>S. pneumoniae</em> strains sensitive to a range of antibiotics. Using an invasive ampicillin-resistant strain, we demonstrate in a murine pneumonia infection model the efficacy of PBT2+ampicillin treatment. These findings present a therapeutic modality to break resistance of drug-resistant <em>S. pneumoniae</em>.</p>
Project description:Shotgun metagenomic sequencing data for nasopharyngeal colonization dynamics with Streptococcus pneumoniae and associated antimicrobial-resistance in a South African birth cohort.
Project description:Pneumonia caused by Streptococcus pneumoniae is a leading cause of death worldwide, and bacterial resistance to antimicrobial drugs has become a major issue. A growing body of evidence indicates that the successful treatment of bacterial infections results from synergy between antibiotic-mediated direct antibacterial activity and the host’s immune defenses. However, the mechanisms underlying the protective immune responses induced by amoxicillin (a β-lactam antibiotic used as the first-line treatment of S. pneumoniae infections) have not been characterized. A better understanding of amoxicillin’s effects on host-pathogen interactions might facilitate the development of other treatment options. Given the crucial role of neutrophils in the control of S. pneumoniae infections, we decided to investigate amoxicillin’s impact on neutrophil development in a mouse model of pneumococcal superinfection. Although a single therapeutic dose of amoxicillin prevented local and systemic inflammatory responses, it did not impair the emergency granulopoiesis triggered in the bone marrow by S. pneumoniae. Importantly, treatment of pneumonia with amoxicillin was associated with a greater mature neutrophil count in the bone marrow; these neutrophils had specific transcriptomic and proteomic profiles. Furthermore, amoxicillin-conditioned, mature neutrophils in the bone marrow had a less activated phenotype and might be rapidly mobilized in peripheral tissues in response to systemic inflammation. Thus, by revealing a novel effect of amoxicillin on the development and functions of bone marrow neutrophils during Streptococcus pneumoniae pneumonia, our findings provide new insights into the impact of amoxicillin treatment on host immune responses.
Project description:Comparison of Streptococcus pneumoniae D39 argR1 mutant compared to D39 wild type in CDM with 10 mM arginine to define the regulon of the ArgR1 regulator under this condition. Details described in Kloosterman TG and Kuipers OP. ArgR1 and AhrC Mediate Arginine-Dependent Regulation of Arginine Acquisition- and Virulence Genes in the Human Pathogen Streptococcus pneumoniae. JBC 2011
Project description:Comparison of Streptococcus pneumoniae D39 ahrC mutant compared to D39 wild type in CDM with 10 mM arginine to define the regulon of the AhrC regulator under this condition. Details described in Kloosterman TG and Kuipers OP. ArgR1 and AhrC Mediate Arginine-Dependent Regulation of Arginine Acquisition- and Virulence Genes in the Human Pathogen Streptococcus pneumoniae. JBC 2011
Project description:Comparison of Streptococcus pneumoniae D39 wild-type grown in CDM+10 mM arginine compared to D39 wild type grown in CDM + 0.05 mM arginine to define the genome-wide transcriptional response to arginine. Details described in Kloosterman TG and Kuipers OP. ArgR1 and AhrC Mediate Arginine-Dependent Regulation of Arginine Acquisition- and Virulence Genes in the Human Pathogen Streptococcus pneumoniae. JBC 2011
Project description:Comparison of Streptococcus pneumoniae D39 argR1-ahrC mutant compared to D39 wild type in CDM with 10 mM arginine to define the regulons of the ArgR1 and AhrC regulators under this condition. Details described in Kloosterman TG and Kuipers OP. ArgR1 and AhrC Mediate Arginine-Dependent Regulation of Arginine Acquisition- and Virulence Genes in the Human Pathogen Streptococcus pneumoniae. JBC 2011
Project description:Background: Antimicrobial resistance is generally studied using a combination of growth inhibition measurements, sometimes in combination with DNA detection methods. However, the actual proteins that cause resistance such as enzymes, efflux pumps and a lack of porins cannot be detected by these methods. Improvements in liquid chromatography (LC) and mass spectrometry (MS) enabled easier and more comprehensive proteome analysis. In the current study, these three methods are combined into a multi-omics approach to analyze resistance against frequently used antibiotics within the beta-lactam, aminoglycoside and fluoroquinolone group in E. coli and K. pneumoniae. Objectives: We aimed to analyze which currently known antimicrobial resistance genes are detected at the protein level using liquid chromatography-mass spectrometry (LC-MS/MS) and to assess whether these could explain beta-lactam, aminoglycoside, and fluoroquinolone resistance in the studied isolates. Furthermore, we aimed to identify significant protein to resistance correlations which have not yet been described and to correlate the abundance of different porins to resistance. Methods: Whole genome sequencing, high-resolution LC-MS/MS and antimicrobial susceptibility testing by broth microdilution were performed for 187 clinical E. coli and K. pneumoniae isolates. Resistance genes and proteins were identified using the Comprehensive Antibiotic Resistance Database (CARD). All proteins were annotated using the NCBI RefSeq database and Prokka. Results & Conclusion: Proteins of small spectrum beta-lactamases, extended spectrum beta-lactamases, AmpC beta-lactamases, carbapenemases, and proteins of 16S ribosomal RNA methyltransferases and aminoglycoside acetyltransferases can be detected in E. coli and K. pneumoniae by LC-MS/MS. The detected mechanisms could explain phenotypic resistance in most of the studied isolates. Differences in the abundance and the primary structure of other proteins such as porins also correlated with resistance. LC-MS/MS is a different and complementary method which can be used to characterize antimicrobial resistance in detail as not only the primary resistance causing mechanisms are detected, but also secondary enhancing resistance mechanisms.