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1. Deletion of genes involved in the ketogluconate metabolism, Entner-Doudoroff pathway, and glucose dehydrogenase increase local and invasive virulence phenotypes in Streptococcus pneumoniae.

Author

Fen Z Hu, Jarosław E Król, Chen Hsuan Sherry Tsai, Rory A Eutsey, Luisa N Hiller, Bhaswati Sen, Azad Ahmed, Todd Hillman, Farrel J Buchinsky, Laura Nistico, Bethany Dice, Mark Longwell, Edward Horsey, and Garth D Ehrlich

Subjects
Medicine, Science
Abstract

Streptococcus pneumoniae displays increased resistance to antibiotic therapy following biofilm formation. A genome-wide search revealed that SP 0320 and SP 0675 (respectively annotated as 5-keto-D-gluconate-5-reductase and glucose dehydrogenase) contain the highest degree of homology to CsgA of Myxococcus xanthus, a signaling factor that promotes cell aggregation and biofilm formation. Single and double SP 0320 and SP 0675 knockout mutants were created in strain BS72; however, no differences were observed in the biofilm-forming phenotypes of mutants compared to the wild type strain. Using the chinchilla model of otitis media and invasive disease, all three mutants exhibited greatly increased virulence compared to the wild type strain (increased pus formation, tympanic membrane rupture, mortality rates). The SP 0320 gene is located in an operon with SP 0317, SP 0318 and SP 0319, which we bioinformatically annotated as being part of the Entner-Doudoroff pathway. Deletion of SP 0317 also resulted in increased mortality in chinchillas; however, mutations in SP 0318 and SP 0319 did not alter the virulence of bacteria compared to the wild type strain. Complementing the SP 0317, SP 0320 and SP 0675 mutant strains reversed the virulence phenotype. We prepared recombinant SP 0317, SP 0318, SP 0320 and SP 0675 proteins and confirmed their functions. These data reveal that disruption of genes involved in the degradation of ketogluconate, the Entner-Doudoroff pathway, and glucose dehydrogenase significantly increase the virulence of bacteria in vivo; two hypothetical models involving virulence triggered by reduced in carbon-flux through the glycolytic pathways are presented.

Published
2019
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2. Deletion of genes involved in the ketogluconate metabolism, Entner-Doudoroff pathway, and glucose dehydrogenase increase local and invasive virulence phenotypes in Streptococcus pneumoniae

Author

Edward Horsey, Fen Z. Hu, Luisa Hiller, Farrel J. Buchinsky, Jarosław E. Król, Bethany Dice, Laura Nistico, Garth D. Ehrlich, Bhaswati Sen, Chen Hsuan Sherry Tsai, Rory Eutsey, Azad Ahmed, Todd Hillman, and Mark Longwell

Subjects
0301 basic medicine, Operon, Mutant, Artificial Gene Amplification and Extension, Otology, medicine.disease_cause, Pathology and Laboratory Medicine, Polymerase Chain Reaction, Biochemistry, Glucose dehydrogenase, Chinchilla, Medicine and Health Sciences, Myxococcus xanthus, Sequence Deletion, Mammals, Multidisciplinary, biology, Virulence, Organic Compounds, Monosaccharides, Eukaryota, Pneumococcus, Cell aggregation, 3. Good health, Bacterial Pathogens, Enzymes, Mutant Strains, Chemistry, Phenotype, Streptococcus pneumoniae, Medical Microbiology, Physical Sciences, Vertebrates, Medicine, Carbohydrate Metabolism, Pathogens, Oxidoreductases, Glycolysis, Research Article, Science, 030106 microbiology, Carbohydrates, Research and Analysis Methods, Microbiology, Rodents, Pneumococcal Infections, 03 medical and health sciences, Bacterial Proteins, medicine, Genetics, Animals, Molecular Biology Techniques, Entner–Doudoroff pathway, Microbial Pathogens, Molecular Biology, Dehydrogenases, Bacteria, Organic Chemistry, Organisms, Chemical Compounds, Biology and Life Sciences, Streptococcus, Proteins, Glucose 1-Dehydrogenase, biology.organism_classification, Otitis Media, 030104 developmental biology, Glucose, Otorhinolaryngology, Biofilms, Mutation, Amniotes, Chinchillas, Enzymology
Abstract

Streptococcus pneumoniae displays increased resistance to antibiotic therapy following biofilm formation. A genome-wide search revealed that SP 0320 and SP 0675 (respectively annotated as 5-keto-D-gluconate-5-reductase and glucose dehydrogenase) contain the highest degree of homology to CsgA of Myxococcus xanthus, a signaling factor that promotes cell aggregation and biofilm formation. Single and double SP 0320 and SP 0675 knockout mutants were created in strain BS72; however, no differences were observed in the biofilm-forming phenotypes of mutants compared to the wild type strain. Using the chinchilla model of otitis media and invasive disease, all three mutants exhibited greatly increased virulence compared to the wild type strain (increased pus formation, tympanic membrane rupture, mortality rates). The SP 0320 gene is located in an operon with SP 0317, SP 0318 and SP 0319, which we bioinformatically annotated as being part of the Entner-Doudoroff pathway. Deletion of SP 0317 also resulted in increased mortality in chinchillas; however, mutations in SP 0318 and SP 0319 did not alter the virulence of bacteria compared to the wild type strain. Complementing the SP 0317, SP 0320 and SP 0675 mutant strains reversed the virulence phenotype. We prepared recombinant SP 0317, SP 0318, SP 0320 and SP 0675 proteins and confirmed their functions. These data reveal that disruption of genes involved in the degradation of ketogluconate, the Entner-Doudoroff pathway, and glucose dehydrogenase significantly increase the virulence of bacteria in vivo; two hypothetical models involving virulence triggered by reduced in carbon-flux through the glycolytic pathways are presented.

Published
2019

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