Pneumococcal Adaptive Responses to Changing ...

EDITORIAL COMMENTARY

Pneumococcal Adaptive Responses to Changing Host Environments Anders P. Hakansson1,2,3 1

Department of Microbiology and Immunology, University at Buffalo, State University of New York, Buffalo; 2The Witebsky Center for Microbial Pathogenesis and Immunology, University at Buffalo, State University of New York, Buffalo; and 3New York State Center of Excellence in Bioinformatics and Life Sciences, Buffalo, New York

Keywords. Streptococcus pneumoniae; colonization; invasive disease; sepsis; hydrogen peroxidase; pyruvate oxidase; macrophages; phagocytosis. Despite being an exclusive human pathogen, Streptococcus pneumoniae (the pneumococcus) is highly adaptable to the various host niches it encounters. It is foremost an exceptional colonizer of the human nasopharynx, where it has developed an array of strategies to adjust to both host factors and other microbial flora that reside there. Upon changes in this environment, often associated with virus infection, pneumococci may leave this environment and enter otherwise sterile sites such as the sinuses, middle ears, and lungs, from where they can disseminate further to reach the bloodstream and the meninges to cause septicemia and meningitis, respectively. The mechanisms and molecules involved in the transition and dissemination to different host environments and the adaptation required to persist in these very different host niches is not entirely clear. In a study in this issue of The Journal of Infectious Diseases [1], a group from

Received 31 January 2014; accepted 3 February 2014; electronically published 6 February 2014. Correspondence: Anders P. Hakansson, PhD, University at Buffalo, State University of New York, 138 Farber Hall, 3435 Main St, Buffalo, NY 14214-3000 ([email protected]). The Journal of Infectious Diseases 2014;210:1–3 © The Author 2014. Published by Oxford University Press on behalf of the Infectious Diseases Society of America. All rights reserved. For Permissions, please e-mail: journals. [email protected]. DOI: 10.1093/infdis/jiu084

the Karolinska Institute, led by Dr Henriques-Normark, elegantly presents us with a novel mechanism explaining how pneumococci adapt to improve their survival during invasive pneumococcal disease (IPD). The authors observed that when serotype 1 strains of 2 clonal complexes were used to infect animals, large colony variants were observed from blood cultures. Similar large colony variants could be cultured from blood of patients with IPD with pneumococcal serotype 1 strains. After sequencing the strains obtained, all large colony variants contained diverse mutations in the spxB gene, encoding pyruvate oxidase that converts pyruvate to acetyl-phosphate while using oxygen to produce hydrogen peroxide. The authors further showed that the large colony variants lacking SpxB activity were more virulent when injected intraperitoneally into mice than wild-type organisms, and that this was related to a decreased early clearance of the bacteria from the bloodstream. In contrast, the lack of SpxB made these strains less able to cause nasopharyngeal colonization. The role of SpxB deficiency was not as clear-cut when the authors used a serotype 4 strain (TIGR4). Infection with wild-type and SpxB-negative TIGR4 resulted in decreased clearance of the SpxB-mutant, but no difference in virulence was observed for this strain, which

suggests that the adaptation to the bloodstream may vary between pneumococcal serotypes or clonal complexes. Either way, the results of the study highlight SpxB as a central molecule in the adaptation of pneumococci to the various environments they may encounter in the human host. SpxB has been studied for some time as a virulence factor and except for the earliest report by Spellerberg et al [2] that indicated that the Avery strain D39 (serotype 2) lacking SpxB was attenuated for both colonization and invasive disease, which is contrary to the results in this study, reports rather consistently indicate that pneumococcal strains of various serotypes (including D39) require SpxB for optimal nasopharyngeal colonization, but that spxB is dramatically down-regulated in the lung and bloodstream [3–5]. The role of SpxB during colonization is likely multifold. SpxB activity in the presence of oxygen results in secretion of hydrogen peroxide that has been shown to protect pneumococci against other species in the nasopharyngeal microbiome [6, 7]. However, SpxB may also act to enhance colonization through improved biofilm formation in several ways. Pneumococci have recently been shown to organize as biofilms during nasopharyngeal colonization [8–10], and optimal biofilm formation requires constant bacterial


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(See the major article by Syk et al on pages 4–13.)

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unclear how various strains and serotypes accomplish this repression. Another interesting avenue for future research will be to characterize the mechanism of the decreased clearance observed in SpxB-negative bacteria. One possible explanation is that SpxBnegative mutants are known to upregulate their capsule production [20], which would certainly increase resistance to phagocytosis. However, the mechanisms involved are likely going to be more complex and multifactorial than that. SpxB is regulated by SpxR, which also regulates other genes involved in colonization [21], suggesting that SpxB is part of at least one genetic regulatory network. A better understanding of the complex genetic networks affected by repression or mutation of spxB is a fascinating topic for future studies. Additionally, how spxB expression interplays with other factors required for persistence in the bloodstream, such as pneumococcal surface protein A and pneumolysin [22, 23], would help us better understand the adaptation of pneumococci to the bloodstream environment. Finally, this study and others indicate that the serotype 1 strains that have emerged in the wake of the pneumococcal conjugate vaccine to cause an increased incidence of IPD worldwide may use separate mechanisms to adapt to their host environment than most other pneumococcal strains. Besides the mutations of spxB observed in this study, another study has identified pneumolysin alleles with lower or no cytotoxicity in invasive serotype 1 strains that appear to be beneficial for pathogenicity of these strains. Yet when these alleles were transferred to a different serotype, the strain became attenuated, despite the fact that a slightly increased growth in the blood was observed [24]. Combined, this study provides new insight into the mechanism of pneumococcal adaptation to avoid clearance and increase their survival in the bloodstream, an important aspect of pneumococcal pathogenesis.


Note Potential conflict of interest. Author certifies no potential conflicts of interest. The author has submitted the ICMJE Form for Disclosure of Potential Conflicts of Interest. Conflicts that the editors consider relevant to the content of the manuscript have been disclosed.

References 1. Syk A, Norman M, Fernebro J, et al. Emergence of hyper-virulent mutants resistant to early clearance during systemic serotype 1 pneumococcal infection in mouse and man. J Infect Dis 2014. 2. Spellerberg B, Cundell DR, Sandros J, et al. Pyruvate oxidase, as a determinant of virulence in Streptococcus pneumoniae. Mol Microbiol 1996; 19:803–13. 3. Orihuela CJ, Gao G, Francis KP, Yu J, Tuomanen EI. Tissue-specific contributions of pneumococcal virulence factors to pathogenesis. J Infect Dis 2004; 190:1661–9. 4. LeMessurier KS, Ogunniyi AD, Paton JC. Differential expression of key pneumococcal virulence genes in vivo. Microbiology 2006; 152:305–11. 5. Mahdi LK, Ogunniyi AD, LeMessurier KS, Paton JC. Pneumococcal virulence gene expression and host cytokine profiles during pathogenesis of invasive disease. Infect Immun 2008; 76:646–57. 6. Pericone CD, Overweg K, Hermans PW, Weiser JN. Inhibitory and bactericidal effects of hydrogen peroxide production by Streptococcus pneumoniae on other inhabitants of the upper respiratory tract. Infect Immun 2000; 68:3990–7. 7. Regev-Yochay G, Trzcinski K, Thompson CM, Malley R, Lipsitch M. Interference between Streptococcus pneumoniae and Staphylococcus aureus: in vitro hydrogen peroxide-mediated killing by Streptococcus pneumoniae. J Bacteriol 2006; 188:4996–5001. 8. Marks LR, Parameswaran GI, Hakansson AP. Pneumococcal interactions with epithelial cells are crucial for optimal biofilm formation and colonization in vitro and in vivo. Infect Immun 2012; 80:2744–60. 9. Shak JR, Ludewick HP, Howery KE, et al. Novel role for the Streptococcus pneumoniae toxin pneumolysin in the assembly of biofilms. MBio 2013; 4:e00655–13. 10. Blanchette-Cain K, Hinojosa CA, Akula Suresh Babu R, et al. Streptococcus pneumoniae biofilm formation is strain dependent, multifactorial, and associated with reduced invasiveness and immunoreactivity during colonization. MBio 2013; 4:e00745-13. 11. Wei H, Havarstein LS. Fratricide is essential for efficient gene transfer between pneumococci in biofilms. Appl Environ Microbiol 2012; 78:5897–905. 12. Marks LR, Reddinger RM, Hakansson AP. High levels of genetic recombination during nasopharyngeal carriage and biofilm


growth and death, which is intricately intertwined with competence or genetic exchange [11, 12]. SpxB appears to be involved in both of these processes, first by producing hydrogen peroxide, which, when accumulated, causes pneumococcal suicide [13], and second because SpxB is also required for induction of competence through an as yet unknown mechanism [14]. Although SpxB is required for optimal colonization, the current study [1] indicates that SpxB expression is detrimental for survival in the bloodstream to the degree that at least serotype 1 strains inactivate their spxB gene through genomic mutation. The increased bacterial numbers of SpxB-negative pneumococci observed early during infection were not due to an increased growth rate but to a decreased clearance, specifically associated with SIGN-R1-positive macrophages in the marginal zone of the spleen. Neutrophils were shown to have no role in this early clearance. This finding concurs with other reports that have long observed that splenectomized animals and human individuals are highly susceptible to IPD, and that clearance is due to splenic macrophages [15–17], whereas neutrophils and T cells appear to be more important for clearance in the lung and inhibition of dissemination to the bloodstream [18, 19]. Besides highlighting SpxB as an important switch for persistence in the nasopharynx and in the bloodstream, the Syk et al study also proposes that some pneumococcal strains or serotypes go to great lengths to inactivate SpxB to avoid elimination from the blood. In serotypes, such as types 2, 4, and 6A, repression of spxB is observed [5], but whether these strains and other strains also adapt to the bloodstream environment through mutations in the spxB gene and whether such mutant strains are more virulent in these strain backgrounds are interesting questions for future research. It is therefore clear that repression of spxB during invasive disease is important for pneumococcal survival, although it is still

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spxB ( pyruvate oxidase) virulence factor gene by a CBS-HotDog domain protein (SpxR) in serotype 2 Streptococcus pneumoniae. Mol Microbiol 2008; 67:729–46. 22. Ogunniyi AD, LeMessurier KS, Graham RM, et al. Contributions of pneumolysin, pneumococcal surface protein A (PspA), and PspC to pathogenicity of Streptococcus pneumoniae D39 in a mouse model. Infect Immun 2007; 75:1843–51. 23. Benton KA, VanCott JL, Briles DE. Role of tumor necrosis factor alpha in the host response of mice to bacteremia caused by pneumolysin-deficient Streptococcus pneumoniae. Infect Immun 1998; 66:839–42. 24. Harvey RM, Ogunniyi AD, Chen AY, Paton JC. Pneumolysin with low hemolytic activity confers an early growth advantage to Streptococcus pneumoniae in the blood. Infect Immun 2011; 79:4122–30.


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