Pneumococcal Polysaccharides - Advancing Research and Development

By Cara N. Wilder, Ph.D.,1 and Susan E. Witko, B.S.2
1ATCC, Manassas,VA
2Pfizer, New York City, NY

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Carbohydrates in the form of capsular polysaccharides and lipopolysaccharides constitute a major component of the cellular surface for many bacterial species.1 In Streptococcus pneumoniae, capsular polysaccharides function as major virulence factors that protect the bacterium from phagocytosis by host immune cells. Within this species, capsular polysaccharides exhibit enormous structural diversity, resulting in significant differences in immunogenicity and antigenicity. In turn, this has presented numerous challenges toward the prevention and serological analysis of pneumococcal infections.2 In the present article, we will discuss the importance of capsular polysaccharides in S. pneumoniae infection, and will describe the use of purified serotype-specific polysaccharides in the development and evaluation of pneumococcal vaccines, the verification of novel immunoassays, and tracking bacterial disease epidemiology.

S. pneumoniae is a major human pathogen known to cause both invasive and non-invasive infections such as pneumonia, meningitis, bacteremia, otitis media, and sinusitis.2,3 More importantly, this bacterium is considered to be the leading cause of vaccine-preventable morbidity and mortality worldwide. A recent report estimated that pneumococcal disease was responsible for 445,000 hospitalizations and 22,000 deaths within the United States in 2004, corresponding to approximately $3.5 billion in direct medical costs.4 Moreover, the World Health Organization (WHO) estimates that S. pneumoniae annually contributes to the death of more than one million children under the age of five years, globally, which is more than malaria, tuberculosis, and AIDS combined.5

This pneumococcal pathogen causes disease through the production of numerous virulence factors, including pneumolysin, pneumococcal surface proteins, and capsular polysaccharides.2 Of these, the capsular polysaccharides—which are polymeric, surface-exposed carbohydrate molecules that encapsulate the bacteria—are considered to be the most important virulence factors as they shield the bacteria from neutrophil clearance. Currently, more than 90 different capsular serotypes have been identified, each distinguishable by serological response, variations in chemical structure, and related genetic mutations.2,6-11 These unique differences can associate with distinct epidemiological properties, including variations in carriage or prevalence of disease. For example, capsular types with higher ratios of charge to carbon may be physically larger, making them more resistant to neutrophil clearance.12,13

Because of their importance in pneumococcal pathogenicity, capsular polysaccharides have been components in a number of serotype-specific vaccines. In 1983, a 23-valent pneumococcal polysaccharide vaccine (PPV23, Pneumovax 23; Merck) was developed, which contained the pooled capsular polysaccharides purified from 23 different serotypes.14 Due to poor immunogenicity in infants, PPV23 was not approved for use in children younger than 2 years of age. In 2000, after nearly two decades without a suitable vaccine for juveniles, a 7-valent pneumococcal conjugate vaccine (PCV7, Prevnar; Wyeth) was developed and licensed for use in infants and young children in the United States. This vaccine was created through the covalent coupling of a protein carrier to the purified polysaccharides from serotypes 4, 6B, 9V, 14, 18C, 19F, and 23F.15 However, to extend vaccine coverage to additional serotypes, a 13-valent pneumococcal conjugate vaccine (PCV13, Prevnar 13®; Pfizer) was later licensed for pediatric and adult use. PCV13 is composed of the same serotype-conjugates included in PCV7, as well as polysaccharide conjugates for serotypes 1, 3, 5, 6A, 7F, and 19A.16

In addition to their role in pneumococcal vaccine development, purified polysaccharides are also necessary for evaluating how effectively a vaccine can induce an immune response. Currently, immunogenicity is measured using a standardized enzyme-linked immunosorbent assay (ELISA) protocol designed for the quantitation of human IgG antibodies specific for S. pneumoniae capsular polysaccharides. The protocol for this assay, which was developed in 2000 by representatives from academia, government, industry, and the WHO, is provided as a training manual containing the standard operating procedures for the preparation, execution, and analysis of the ELISA.17 In this assay, ELISA plates are first coated with purified, serotype-specific capsular pneumococcal polysaccharides (ATCC, Manassas, VA). Following adsorption of the individual polysaccharides, dilutions of absorbed human sera are then added to the ELISA plates. The serotype specific antibodies that remain bound to the purified pneumococcal polysaccharides are then detected and quantitated using an anti-human IgG antibody conjugated with alkaline phosphatase.17

In recent years, capsular polysaccharides have been similarly used in the development and evaluation of novel immunoassays. Though the WHO ELISA protocol provides a standardized, accurate method for the evaluation of vaccine efficacy, it has recently come under scrutiny as it is laborious and consumes large volumes of serum. To this end, the development of novel multiplex immunoassays that allow for the simultaneous measurement of specific antibodies against various pneumococcal polysaccharides have been explored.18-20 For example, a study by Klein et al. described the development and characterization of a multiplex bead-based antibody quantification assay (MBIA) based on Luminex® xMAP® technology (Luminex, Austin, TX) that required the conjugation of purified serotype-specific capsular pneumococcal polysaccharides (ATCC, Manassas, VA) to fluorescent beads. In this assay the polysaccharide-bead conjugate mixture was incubated with various dilutions of pre-adsorbed serum. Following incubation, bound antibodies were quantified through the use of phycoerythrin-labeled goat anti-human reporter antibodies.18 As compared to the traditional ELISA protocol, the MBIA procedure was able to provide a more efficient, cost-effective means of analyzing small volumes of serum; though, careful optimization, stringent validation, and continuous monitoring are still required.

Along with their function in detecting and enumerating anti-pneumococcal antibodies, the combinatorial use of capsular polysaccharides in immunoassays is ideal for tracking the epidemiology of serotype-specific pneumococcal infections following vaccination. The gold-standard method for tracking invasive S. pneumoniae infection has occurred through traditional culture-based assays. However, good-quality samples are not always available, culturing and serological analyses can be fairly time-consuming and laborious, and the detection of viable pneumococci can be hampered by use of antibiotics.21,22 Furthermore, serotype-specific assays to diagnose community-acquired pneumonia caused by S. pneumoniae only recently became available.23,24 To assess the positive impact of the 13-valent pneumococcal conjugate vaccine (PCV13, Prevnar 13®; Pfizer), serotype-specific diagnostics can provide valuable data on how effectively the vaccine reduces disease in the population, as well as assist in monitoring for the potential emergence of rare serotypes.

Pfizer, a global leader in the biopharmaceutical industry committed to producing quality products that make an impact on health worldwide, is collaborating with ATCC, a leading provider of standards for the global scientific community, to ensure that researchers worldwide have access to purified, high-quality polysaccharides. The supply of pneumococcal polysaccharides is essential in continuing to drive pneumococcal research, predicting future epidemiological shifts, and combating this deadly pathogen.

Overall, capsular polysaccharides are bacterial antigens that play a significant role in pneumococcal pathogenicity. By targeting these polymeric carbohydrates through the use of polysaccharide or polysaccharide conjugate vaccines, the disease burden of S. pneumoniae serotypes covered by these vaccines has been reduced significantly. Availability of purified pneumococcal polysaccharides has also facilitated the development of a number of novel technological advances in the development of serological assays and diagnostics, and the understanding of disease epidemiology.


  1. Weintraub A. Immunology of bacterial polysaccharide antigens. Carbohydrate research 338: 2539-2547, 2003. 
  2. Song JY, Nahm MH, Moseley MA. Clinical implications of pneumococcal serotypes: invasive disease potential, clinical presentations, and antibiotic resistance. J Korean Med Sci 28: 4-15, 2013. 
  3. Kaltoft MS, Skov Sorensen UB, Slotved HC, Konradsen HB. An easy method for detection of nasopharyngeal carriage of multiple Streptococcus pneumoniae serotypes. J Microbiol Methods 75: 540-544, 2008. 
  4. Huang SS, et al. Healthcare utilization and cost of pneumococcal disease in the United States. Vaccine 29: 3398-3412, 2011. 
  5. WHO. Pneumonia Fact Sheet, 2013. 
  6. Park IH, et al. Discovery of a new capsular serotype (6C) within serogroup 6 of Streptococcus pneumoniae. J Clin Microbiol 45: 1225-1233, 2007. 
  7. Zartler ER, et al. Structure of the capsular polysaccharide of pneumococcal serotype 11A reveals a novel acetylglycerol that is the structural basis for 11A subtypes. J Biol Chem 284: 7318-7329, 2009. 
  8. Jin P, et al. First report of putative Streptococcus pneumoniae serotype 6D among nasopharyngeal isolates from Fijian children. J Infect Dis 200: 1375-1380, 2009. 
  9. Calix JJ, Dagan R, Pelton SI, Porat N, Nahm MH. Differential occurrence of Streptococcus pneumoniae serotype 11E between asymptomatic carriage and invasive pneumococcal disease isolates reflects a unique model of pathogen microevolution. Clinical infectious diseases: an official publication of the Infectious Diseases Society of America 54: 794-799, 2012. 
  10. Calix JJ, et al. Biochemical, genetic, and serological characterization of two capsule subtypes among Streptococcus pneumoniae Serotype 20 strains: discovery of a new pneumococcal serotype. J Biol Chem 287: 27885-27894, 2012. 
  11. Henrichsen J. Six newly recognized types of Streptococcus pneumoniae. J Clin Microbiol 33: 2759-2762, 1995. 
  12. Hausdorff WP, Feikin DR, Klugman KP. Epidemiological differences among pneumococcal serotypes. Lancet Infect Dise 5: 83-93, 2005. 
  13. Weinberger DM, et al. Pneumococcal capsular polysaccharide structure predicts serotype prevalence. PLoS pathogens 5, e1000476, 2009. 
  14. Robbins JB, et al. Considerations for formulating the second-generation pneumococcal capsular polysaccharide vaccine with emphasis on the cross-reactive types within groups. J Infect Dise 148: 1136-1159, 1983. 
  15. ACIP. Preventing pneumococcal disease among infants and young children: recommendation of the Advisory Committee on Immunization Practices (ACIP). MMWR Recomm Rep 49: 1-35, 2000. 
  16. FDA. Product approval information---licensing action, package insert: Prevnar 13 (pneumococcal 13-valent conjugate vaccine), Pfizer. Rockville, MD: US Department of Health and Human Services, Food and Drug Administration, 2010. 
  17. World Health Organization Pneumococcal Serology Reference Laboratories at the Institute of Child Health, U. C. L., London, England, and the Department of Pathology at the University of Alabama at Birmingham, Birmingham Alabama, USA. Training manual for Enzyme linked immunosorbent assay for the quantification of Streptococcus pneumoniae serotype specific IgG (Pn PS ELISA), 2000. 
  18. Klein DL, et al. Development and characterization of a multiplex bead-based immunoassay to quantify pneumococcal capsular polysaccharide-specific antibodies. Clin Vaccine Immunol: CVI 19: 1276-1282, 2012. 
  19. Goldblatt D, Ashton L, Zhang Y, Antonello J, Marchese RD. Comparison of a new multiplex binding assay versus the enzyme-linked immunosorbent assay for measurement of serotype-specific pneumococcal capsular polysaccharide IgG. Clin Vaccine Immunol: CVI 18: 1744-1751, 2011. 
  20. Elberse KE, Tcherniaeva I, Berbers GA, Schouls LM. Optimization and application of a multiplex bead-based assay to quantify serotypespecific IgG against Streptococcus pneumoniae polysaccharides: response to the booster vaccine after immunization with the pneumococcal 7-valent conjugate vaccine. Clin Vaccine Immunol: CVI 17: 674-682, 2010. 
  21. Ewig S, et al. Factors associated with unknown aetiology in patients with community-acquired pneumonia. Eur Resp J 20: 1254-1262, 2002. 
  22. Musher DM, Montoya R, Wanahita A. Diagnostic value of microscopic examination of Gram-stained sputum and sputum cultures in patients with bacteremic pneumococcal pneumonia. Clinical infectious diseases : an official publication of the Infectious Diseases Society of America 39: 165-169, 2004. 
  23. Pride MW HS, Wu K, et al. Validation of an immunodiagnostic assay for detection of 13 Streptococcus pneumoniae serotype-specific polysaccharides in human urine. Clin Vaccine Immunol 19: 1131-1141, 2012. 
  24. Sheppard CL HT, Smith MD, George RC. Development of a sensitive, multiplexed immunoassay using xMAP beads for detection of serotype-specific Streptococcus pneumoniae antigen in urine samples. J Med Microbiol 60: 49-55, 2011.


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