Carlos J. Orihuela, United States of America

University of Alabama at Birmingham Microbiology

Author Of 3 Presentations

STREPTOCOCCUS PNEUMONIAE ANATOMICAL SITE GENE EXPRESSION ATLAS: DISSECTION OF HOST-PATHOGEN INTERACTIONS USING LARGE-SCALE DUAL TRANSCRIPTOMICS OF THREE STREPTOCOCCUS PNEUMONIAE ISOLATES IN VIVO (ID 571)

Abstract

Background

Streptococcus pneumoniae (Spn) colonizes the nasopharynx asymptomatically but is also a leading cause of community-acquired pneumonia, otitis media, bacteremia, sepsis, and meningitis. Spn pathogenesis studies have largely focused on the middle ear, airway, bloodstream, and central nervous system. Yet, increasing evidence indicates that during severe infections, Spn gains access to other vital organs.

Methods

Using dual species RNA-seq transcriptomics and mouse models of asymptomatic colonization and invasive disease, we characterized the pneumococcal (3 strains) and host transcriptomes during colonization and disease. This included the nasopharynx, lungs, and blood. Bacterial and host regulated pathways were validated using a panel of knock-out Spn strains and/or treatment of mice with relevant molecules.

Results

Spn differentially regulates genes in vivo in a site-specific manner, including upregulation of diverse scavenger pathways within the nasopharynx. Core genes highly expressed at all disease sites, and therefore targets for intervention, were identified, including pspA. Novel insights include that pneumococcal infection induces a Type I interferon response during pneumonia and invasive disease, but not colonization.

Conclusions

The pneumococcal transcriptome varies in anatomical site-specific manner and is vastly different during asymptomatic colonization versus disease. Host transcriptomes during invasive disease provide novel insights about potential therapeutic targets.

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ORAL STREPTOCOCCI: A SOURCE OF NEW AND VIRULENT CAPSULE FOR PNEUMOCOCCI (ID 244)

Abstract

Background

Pneumococcal conjugate vaccines have been very successful, but their use has increased infections by non-vaccine serotypes. Oral streptococci often harbor capsular polysaccharide (PS) synthesis loci (cps). If pneumococcus can replace its cps with oral streptococcal cps, it may increase its serotype repertoire.

Methods

Chemical structures of capsular PS from an oral streptococcus strain, SK95, and a pneumococcal strain, D39, were determined by 2-dimensional NMR. An acapsular pneumococcus strain was transformed with cps from SK95 and D39, which were compared for their virulence by measuring non-specific phagocytic killing and their ability to kill mice.

Results

SK95 and D39 both produce structurally identical type 2 capsules. Baby rabbit serum (BRS) at 10% non-specifically killed >50% of SK95; however, 12·5% BRS killed <20% of D39. Non-encapsulated pneumococci became resistant to BRS as D39 following transformation with SK95 cps. Similarly, SK95 is avirulent in well-established in vivo mouse model. When the acapsular pneumococcus was transformed with SK95 cps, the transformant became virulent and killed all mice.

Conclusions

Oral streptococcus cps can make acapsular pneumococcus virulent and inter-species cps transfer should be considered a potential mechanism for serotype replacement. Our findings highlight potential limitations of current WHO criterion for studying serotypes of pneumococci carried without first isolating bacteria.

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STREPTOCOCCUS PNEUMONIAE GENERATES HYDROXYL RADICALS TO RAPIDLY INTOXICATE AND KILL STAPHYLOCOCCUS AUREUS STRAINS (ID 256)

Abstract

Background

Streptococcus pneumoniae (Spn) strains rapidly kills Staphylococcus aureus (Sau) by producing membrane-permeable hydrogen peroxide (H2O2). The exact mechanism by which the H2O2-mediated killing occurs is not well understood.

Methods

An in vitro model that mimicked Spn-Sau contact during colonization of the upper airways and whole genome sequencing was conducted. Different Spn H2O2 mutants were constructed to confirm the Sau killing mechanism.

Results

Sau killing required outcompeting densities of Spn. A collection of MRSA/MSSA strains showed a linear sensitivity (R2=0.95) for Spn killing but the same strains had different susceptibilities when challenged against pure H2O2. WGS of these MRSA/MSSA strains revealed no association between clonal complex and susceptibility, or resistance, to Spn, or H2O2,respectively. A sublethal dose (~1 mM) of pure H2O2 when incubated with TIGR4DspxB eradicated cultures of Sau strains suggesting that Spn converts H2O2 to the hydroxyl radical (·OH). Accordingly, Sau killing was completely blocked by incubating with scavengers of ·OH radicals, DMSO, or thiourea.

Conclusions

Spn produces H2O2 which is rapidly converted to a more potent oxidant, the ·OH radical. Hydroxyl radicals does not affect Spn viability but rapidly intoxicate Sau strains. The target(s) of the ·OH radicals represents an exciting new alternative for the development of therapeutics against Sau infections.

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