Streptococcus pneumoniae pathogenesis

Sam KingCMP and BCMM meeting

Projects

• Does pneumococcal genomic variation contribute to development of different disease states

• Structure function analysis of pneumococcal transporters

Pneumococcal diversity

Pneumococcal bacteremia

Hemolytic uremic syndrome (HUS)

Is there something different about these isolates?Can we use that knowledge to develop tests to identify patients at risk or therapeutics?

To Identify pneumococcal sequences that contribute to HUS

Aims

1. Identify pneumococcal loci that potentially correlate with development of HUS

2. Identify pneumococcal HUS enriched sequences

3. Determine the biological contribution of these sequences to pathogenesis

Identify pneumococcal loci that potentially correlate with development of HUS

• Sequence six pneumococcal HUS isolates– Two methodologies

• Next generation (Biomedical Genomics Core)– 30 million reads for each strain– Short (~100bp), high accuracy– Has to be aligned to a sequenced genome

• Third generation sequencing (EA sequencing)– Longer reads (~3kb)– Accuracy low– Will allow generation of de novo sequence

Analysis of sequence data• Biomedical Genomics Core (Post doctoral scientist)

– Will generate accurate de novo sequence using info from both techniques

– Compare sequence with 16 sequenced genomes• King lab

– Prioritize sequence variants for analysis• Conserved in the majority of HUS isolates• Absent in the majority of non-HUS isolate• Introduce or remove open reading frames• Change promoter or coding sequence of predicted

extracellular proteins

Identify pneumococcal HUS enriched sequences

• Screen presence of up to 50 variants• In up to:

– 50 HUS isolates– 50 non-HUS blood isolates

• Determine if there is a significant correlation with HUS

Identify pneumococcal HUS enriched sequences

• Will 50 strains be enough?• If pneumococcal HUS isolates are more closely

related genetically how will that affect our data? Can we take account of that?

Biological consequences of these HUS enriched sequences

• Correlation but not causation• Contribution to pathogenesis

– Bioinformatics– Biochemical assays– Genetic approaches

Structure function analysis of pneumococcal transporters

• ATP binding cassette transporters – Conserved protein family– Five components

Substrate binding protein

Permease proteins

ATPase (nuclear binding domains)

ATP

ATP

ADP + Pi

ADP + Pi