Environmental and Nutritional Factors That Affect Growth and Metabolism of the Pneumococcal Serotype 2 Strain D39 and Its Nonencapsulated Derivative Strain

Links between carbohydrate metabolism and virulence in Streptococcus pneumoniae have been recurrently established. To investigate these links further we developed a chemically defined medium (CDM) and standardized growth conditions that allowed for high growth yields of the related pneumococcal strains D39 and R6. The utilization of the defined medium enabled the evaluation of different environmental and nutritional factors on growth and fermentation patterns under controlled conditions of pH, temperature and gas atmosphere. The same growth conditions impacted differently on the nonencapsulated R6, and its encapsulated progenitor D39. A semi-aerobic atmosphere and a raised concentration of uracil, a fundamental component of the D39 capsule, improved considerably D39 growth rate and biomass. In contrast, in strain R6, the growth rate was enhanced by strictly anaerobic conditions and uracil had no effect on biomass. In the presence of oxygen, the difference in the growth rates was mainly attributed to a lower activity of pyruvate oxidase in strain D39. Our data indicate an intricate connection between capsule production in strain D39 and uracil availability. In this study, we have also successfully applied the in vivo NMR technique to study sugar metabolism in S. pneumoniae R6. Glucose consumption, end-products formation and evolution of intracellular metabolite pools were monitored online by C-NMR. Additionally, the pools of NTP and inorganic phosphate were followed by P-NMR after a pulse of glucose. These results represent the first metabolic profiling data obtained non-invasively for S. pneumoniae, and pave the way to a better understanding of regulation of central metabolism. Citation: Carvalho SM, Kuipers OP, Neves AR (2013) Environmental and Nutritional Factors That Affect Growth and Metabolism of the Pneumococcal Serotype 2 Strain D39 and Its Nonencapsulated Derivative Strain R6. PLoS ONE 8(3): e58492. doi:10.1371/journal.pone.0058492 Editor: Malcolm James Horsburgh, University of Liverpool, United Kingdom Received October 5, 2012; Accepted February 5, 2013; Published March 7, 2013 Copyright: 2013 Carvalho et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Funding: S. M. Carvalho was supported by a Fundação para a Ciência e a Tecnologia (FCT) Ph. D. grant (SFRH/BD/35947/2007). This work was supported by Fundação para a Ciência e a Tecnologia, Portugal (FCT) and FEDER, projects PDCT/BIA-MIC/56235/2004, PTDC/BIA-MIC/099963/2008 and PTDC/SAU-MII/100964/ 2008 and grant PEst-OE/EQB/LA0004/2011. The NMR spectrometers are part of The National NMR Network (REDE/1517/RMN/2005), supported by ‘‘Programa Operacional Ciência e Inovação (POCTI) 2010’’ and FCT. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: The authors have declared that no competing interests exist. * E-mail: [email protected] ¤ Current address: Novo Nordisk Foundation Center for Biosustainability, Technical University of Denmark, Hørsholm, Denmark